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Ikeda M, Kobayashi K, Nakayama-Sadakiyo Y, Sato Y, Tobita A, Saito M, Yamasu K. Transcriptome Analysis Suggested Striking Transition Around the End of Epiboly in the Gene Regulatory Network Downstream of the Oct4-Type POU Gene in Zebrafish Embryos. Dev Growth Differ 2025. [PMID: 40490365 DOI: 10.1111/dgd.70012] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/13/2025] [Revised: 05/21/2025] [Accepted: 05/23/2025] [Indexed: 06/11/2025]
Abstract
Zebrafish pou5f3 encodes a Class V POU transcription factor, Pou5f3, which regulates various developmental processes, including neurogenesis and brain formation. In the current study, we attempted to comprehensively identify the Pou5f3 downstream genes around the end of epiboly, when the competence of the mid-hindbrain region to Pou5f3 suppression changes drastically, by the microarray method and a heat-inducible dominant-interference pou5f3 gene (en-pou5f3) that functionally suppresses pou5f3. At late epiboly and early somitogenesis stages, we identified genes whose expression was altered in en-pou5f3-induced embryos, revealing numerous genes regulated differently by Pou5f3 at the two stages. The validity of the microarray data was confirmed by whole mount in situ hybridization and quantitative RT-PCR. Many of the downstream genes were implicated by the Gene ontology (GO) analyses in transcriptional regulation and neural development and were enriched with sox genes and bHLH genes such as her genes. Interestingly, we noticed a tendency that Notch-dependent her genes were activated, whereas Notch-independent her genes were downregulated by Pou5f3 suppression. Among the Notch-independent her genes, her3, which is orthologous to mammalian Hes3, was suggested to be strongly activated endogenously by Pou5f3. In the upstream DNA of this gene, we found two noncoding conserved sequences (NCRs), which harbored consensus binding sites for Pou5f3, Sox, and Nanog. We further showed in reporter assays that the transcriptional regulatory activity of the her3 upstream DNA was strongly enhanced by SoxB1, and this SoxB1-mediated activation was weakened by Pou5f3. Deletion experiments showed that both upstream NCRs were involved in transcriptional repression.
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Affiliation(s)
- Masaaki Ikeda
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Shimo-Okubo, Sakura-ku, Saitama City, Saitama, Japan
| | - Kana Kobayashi
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Shimo-Okubo, Sakura-ku, Saitama City, Saitama, Japan
| | - Yukiko Nakayama-Sadakiyo
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Shimo-Okubo, Sakura-ku, Saitama City, Saitama, Japan
| | - Yuto Sato
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Shimo-Okubo, Sakura-ku, Saitama City, Saitama, Japan
| | - Ayano Tobita
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Shimo-Okubo, Sakura-ku, Saitama City, Saitama, Japan
| | - Mika Saito
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Shimo-Okubo, Sakura-ku, Saitama City, Saitama, Japan
| | - Kyo Yamasu
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Shimo-Okubo, Sakura-ku, Saitama City, Saitama, Japan
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Prista von Bonhorst F, Gandrillon O, Herbach U, Robert C, Chazaud C, De Decker Y, Gonze D, Dupont G. Uncovering candidate Nanog-Helper genes in early mouse embryo differentiation using differential entropy and network inference. Sci Rep 2025; 15:19975. [PMID: 40481039 PMCID: PMC12144205 DOI: 10.1038/s41598-025-03956-y] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/25/2025] [Accepted: 05/23/2025] [Indexed: 06/11/2025] Open
Abstract
In the preimplantation mammalian embryo, stochastic cell-to-cell expression heterogeneity is followed by signal reinforcement to initiate the specification of Inner Cell Mass (ICM) cells into Epiblast (Epi). The expression of NANOG, the key transcription factor for the Epi fate, is necessary but not sufficient: coincident expression of other factors is required. To identify possible Nanog-helper genes, we analyzed gene expression variability in five time-stamped single-cell transcriptomic datasets using differential entropy, a quantitative measure of cell-to-cell heterogeneity. The entropy of Nanog displays a peak-shaped temporal pattern from the 16-cell to the 64-cell stage, consistent with its key role in Epi specification. By estimating the entropy profiles of the 21 genes common to all five datasets, we identified three genes - Pecam1, Sox2, and Hnf4a - whose variability in expression patterns mirrors that of Nanog. We further performed gene regulatory network inference using CARDAMOM, an algorithm that exploits temporal dynamics and transcriptional bursting. The results revealed that these three genes exhibit reciprocal activation with Nanog at the 32-cell stage. This regulatory motif reinforces fate-switching decisions and co-expression states. Our innovative analysis of single-cell transcriptomic data thus uncovers a likely role for Pecam1, Sox2, and Hnf4a as key genes that, when coincidentally expressed with Nanog, initiate ICM differentiation.
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Affiliation(s)
| | - Olivier Gandrillon
- Univ Lyon, CNRS, ENS de Lyon, INSERM, UMR5239, LBMC, U1210, 69364, Lyon, France
| | - Ulysse Herbach
- CNRS, Inria, IECL, Université de Lorraine, F-54000, Nancy, France
| | - Corentin Robert
- Unit of Theoretical Chronobiology, Université libre de Bruxelles (ULB), 1050, Brussels, Belgium
- Non-linear Physical Chemistry Unit, Université libre de Bruxelles (ULB), 1050, Brussels, Belgium
| | - Claire Chazaud
- CNRS, INSERM, GReD Institute, Université Clermont Auvergne, Clermont-Ferrand, France
| | - Yannick De Decker
- Non-linear Physical Chemistry Unit, Université libre de Bruxelles (ULB), 1050, Brussels, Belgium
| | - Didier Gonze
- Unit of Theoretical Chronobiology, Université libre de Bruxelles (ULB), 1050, Brussels, Belgium
| | - Geneviève Dupont
- Unit of Theoretical Chronobiology, Université libre de Bruxelles (ULB), 1050, Brussels, Belgium
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Kongtanawanich K, Prasopporn S, Jamnongsong S, Thongsin N, Payungwong T, Okada S, Hokland M, Wattanapanitch M, Jirawatnotai S. A live single-cell reporter system reveals drug-induced plasticity of a cancer stem cell-like population in cholangiocarcinoma. Sci Rep 2024; 14:22619. [PMID: 39349745 PMCID: PMC11442615 DOI: 10.1038/s41598-024-73581-8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/04/2024] [Accepted: 09/18/2024] [Indexed: 10/04/2024] Open
Abstract
Cancer stem cells (CSC) play an important role in carcinogenesis and are acknowledged to be responsible for chemoresistance in cholangiocarcinoma (CCA). Studying CCA CSC has been challenging, due to lack of consensus CSC markers, and to their plastic nature. Since dual expression of the core pluripotent factors SOX2/OCT4 has been shown to correlate with poor outcome in CCA patients, we selected the SOX2/OCT4 activating short half-life GFP-based live reporter (SORE6-dsCopGFP) to study CSC dynamics at the single-cell level. Transduction of five human CCA cell lines resulted in the expression of 1.8-13.1% GFP-positive (SORE6POS) cells. By live imaging, we found that SORE6POS CCA cells possess self-renewal capacity and that they can be induced to differentiate. Significantly, the SORE6POS cells were highly tumorigenic, both in vitro and in vivo, thus implicating the characteristics of primary CSCs. When we then analyzed for selected CSC-related markers, we found that the majority of both CD133+/CD44+, and CD133+/LGR5+ CCA cells were SORE6POS cells. Exposing transduced cells to standard CCA chemotherapy revealed higher growth rate inhibition at 50% (GR50s) for SORE6POS cells compared to GFP-negative (SORE6NEG) ones indicating that these CSC-like cells were more resistant to the treatment. Moreover, the chemotherapy induced SORE6POS from SORE6NEG cells, while retaining the existing SORE6POS population. Finally, treatment of transduced cells with CDK4/6 inhibitors in vitro for 3 days resulted in a lowered CSC number in the culture. Thus, applying a live reporter system allowed us to elucidate the stem cell diversity and drug-induced plasticity of CCA CSCs. These findings have clear implications for future management of such patients.
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Affiliation(s)
| | - Sunisa Prasopporn
- Department of Pharmacology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
- Siriraj Center of Research Excellence for Precision Medicine and Systems Pharmacology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Supawan Jamnongsong
- Department of Pharmacology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
- Siriraj Center of Research Excellence for Precision Medicine and Systems Pharmacology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Nontaphat Thongsin
- Siriraj Center for Regenerative Medicine, Research Department, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Tongchai Payungwong
- Department of Pharmacology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
- Siriraj Center of Research Excellence for Precision Medicine and Systems Pharmacology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Seiji Okada
- Division of Hematopoiesis, Joint Research Center for Human Retrovirus Infection, Kumamoto University, Kumamoto, Japan
| | | | - Methichit Wattanapanitch
- Siriraj Center for Regenerative Medicine, Research Department, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand
| | - Siwanon Jirawatnotai
- Department of Pharmacology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand.
- Siriraj Center of Research Excellence for Precision Medicine and Systems Pharmacology, Faculty of Medicine Siriraj Hospital, Mahidol University, Bangkok, Thailand.
- Division of Hematopoiesis, Joint Research Center for Human Retrovirus Infection, Kumamoto University, Kumamoto, Japan.
- Faculty of Pharmacy, Silpakorn University, Nakhon Pathom, Thailand.
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Waite JB, Boytz R, Traeger AR, Lind TM, Lumbao-Conradson K, Torigoe SE. A suboptimal OCT4-SOX2 binding site facilitates the naïve-state specific function of a Klf4 enhancer. PLoS One 2024; 19:e0311120. [PMID: 39348365 PMCID: PMC11441684 DOI: 10.1371/journal.pone.0311120] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/05/2024] [Accepted: 09/12/2024] [Indexed: 10/02/2024] Open
Abstract
Enhancers have critical functions in the precise, spatiotemporal control of transcription during development. It is thought that enhancer grammar, or the characteristics and arrangements of transcription factor binding sites, underlie the specific functions of developmental enhancers. In this study, we sought to identify grammatical constraints that direct enhancer activity in the naïve state of pluripotency, focusing on the enhancers for the naïve-state specific gene, Klf4. Using a combination of biochemical tests, reporter assays, and endogenous mutations in mouse embryonic stem cells, we have studied the binding sites for the transcription factors OCT4 and SOX2. We have found that the three Klf4 enhancers contain suboptimal OCT4-SOX2 composite binding sites. Substitution with a high-affinity OCT4-SOX2 binding site in Klf4 enhancer E2 rescued enhancer function and Klf4 expression upon loss of the ESRRB and STAT3 binding sites. We also observed that the low-affinity of the OCT4-SOX2 binding site is crucial to drive the naïve-state specific activities of Klf4 enhancer E2. Altogether, our work suggests that the affinity of OCT4-SOX2 binding sites could facilitate enhancer functions in specific states of pluripotency.
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Affiliation(s)
- Jack B Waite
- Biochemistry & Molecular Biology Program, Lewis & Clark College, Portland, Oregon, United States of America
| | - RuthMabel Boytz
- Biology Department, Lewis & Clark College, Portland, Oregon, United States of America
| | - Alexis R Traeger
- Biochemistry & Molecular Biology Program, Lewis & Clark College, Portland, Oregon, United States of America
| | - Torrey M Lind
- Biochemistry & Molecular Biology Program, Lewis & Clark College, Portland, Oregon, United States of America
| | - Koya Lumbao-Conradson
- Biochemistry & Molecular Biology Program, Lewis & Clark College, Portland, Oregon, United States of America
| | - Sharon E Torigoe
- Biochemistry & Molecular Biology Program, Lewis & Clark College, Portland, Oregon, United States of America
- Biology Department, Lewis & Clark College, Portland, Oregon, United States of America
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5
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Yu M, Wang F, Gang H, Liu C. Research progress of nanog gene in fish. Mol Genet Genomics 2024; 299:88. [PMID: 39313603 DOI: 10.1007/s00438-024-02182-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/24/2024] [Accepted: 09/07/2024] [Indexed: 09/25/2024]
Abstract
Nanog is a crucial regulatory factor in maintaining the self-renewal and pluripotency of embryonic stem cells. It is involved in various biological processes, such as early embryonic development, cell reprogramming, cell cycle regulation, the proliferation and migration of primordial germ cells. While research on this gene has primarily focused on mammals, there has been a growing interest in studying nanog in fish. However, there is a notable lack of comprehensive reviews regarding this gene in fish, which is essential for guiding future research. This review aims to provide a thorough summary of the gene's structure, expression patterns, functions and regulatory mechanisms in fish. The findings suggest that nanog probably has both conserved and divergent functions in regulating cell pluripotency, early embryonic development, and germ cell development in teleosts compared to other species, including mammals. These insights lay the foundation for future research and applications of the nanog gene, providing a new perspective for understanding the evolution and conserved charactristics of teleost nanog.
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Affiliation(s)
- Miao Yu
- Engineering Technology Research Center of Henan Province for Aquatic Animal Cultivation, Engineering Laboratory of Henan Province for Aquatic Animal Disease Control, Observation and Research Station On Water Ecosystem in Danjiangkou Reservoir of Henan Province, College of Fisheries, Henan Normal University, Xinxiang, 453007, China.
| | - Fangyuan Wang
- Engineering Technology Research Center of Henan Province for Aquatic Animal Cultivation, Engineering Laboratory of Henan Province for Aquatic Animal Disease Control, Observation and Research Station On Water Ecosystem in Danjiangkou Reservoir of Henan Province, College of Fisheries, Henan Normal University, Xinxiang, 453007, China
| | - Huihui Gang
- Engineering Technology Research Center of Henan Province for Aquatic Animal Cultivation, Engineering Laboratory of Henan Province for Aquatic Animal Disease Control, Observation and Research Station On Water Ecosystem in Danjiangkou Reservoir of Henan Province, College of Fisheries, Henan Normal University, Xinxiang, 453007, China
| | - Chuanhu Liu
- School of 3D Printing, Xinxiang University, Xinxiang, 453003, China.
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Yuikawa T, Sato T, Ikeda M, Tsuruoka M, Yasuda K, Sato Y, Nasu K, Yamasu K. Elongation of the developing spinal cord is driven by Oct4-type transcription factor-mediated regulation of retinoic acid signaling in zebrafish embryos. Dev Dyn 2024; 253:404-422. [PMID: 37850839 DOI: 10.1002/dvdy.666] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/23/2023] [Revised: 10/04/2023] [Accepted: 10/05/2023] [Indexed: 10/19/2023] Open
Abstract
BACKGROUND Elongation of the spinal cord is dependent on neural development from neuromesodermal progenitors in the tail bud. We previously showed the involvement of the Oct4-type gene, pou5f3, in this process in zebrafish mainly by dominant-interference gene induction, but, to compensate for the limitation of this transgene approach, mutant analysis was indispensable. pou5f3 involvement in the signaling pathways was another unsolved question. RESULTS We examined the phenotypes of pou5f3 mutants and the effects of Pou5f3 activation by the tamoxifen-ERT2 system in the posterior neural tube, together confirming the involvement of pou5f3. The reporter assays using P19 cells implicated tail bud-related transcription factors in pou5f3 expression. Regulation of tail bud development by retinoic acid (RA) signaling was confirmed by treatment of embryos with RA and the synthesis inhibitor, and in vitro reporter assays further showed that RA signaling regulated pou5f3 expression. Importantly, the expression of the RA degradation enzyme gene, cyp26a1, was down-regulated in embryos with disrupted pou5f3 activity. CONCLUSIONS The involvement of pou5f3 in spinal cord extension was supported by using mutants and the gain-of-function approach. Our findings further suggest that pou5f3 regulates the RA level, contributing to neurogenesis in the posterior neural tube.
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Affiliation(s)
- Tatsuya Yuikawa
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Saitama City, Saitama, Japan
| | - Takehisa Sato
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Saitama City, Saitama, Japan
| | - Masaaki Ikeda
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Saitama City, Saitama, Japan
| | - Momo Tsuruoka
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Saitama City, Saitama, Japan
| | - Kaede Yasuda
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Saitama City, Saitama, Japan
| | - Yuto Sato
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Saitama City, Saitama, Japan
| | - Kouhei Nasu
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Saitama City, Saitama, Japan
| | - Kyo Yamasu
- Division of Life Science, Graduate School of Science and Engineering, Saitama University, Saitama City, Saitama, Japan
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Dehghanian F, Bovio PP, Gather F, Probst S, Naghsh-Nilchi A, Vogel T. ZFP982 confers mouse embryonic stem cell characteristics by regulating expression of Nanog, Zfp42, and Dppa3. BIOCHIMICA ET BIOPHYSICA ACTA. MOLECULAR CELL RESEARCH 2024; 1871:119686. [PMID: 38342310 DOI: 10.1016/j.bbamcr.2024.119686] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 08/01/2023] [Revised: 01/27/2024] [Accepted: 02/01/2024] [Indexed: 02/13/2024]
Abstract
BACKGROUND Understanding the genetic underpinnings of protein networks conferring stemness is of broad interest for basic and translational research. METHODS We used multi-omics analyses to identify and characterize stemness genes, and focused on the zinc finger protein 982 (Zfp982) that regulates stemness through the expression of Nanog, Zfp42, and Dppa3 in mouse embryonic stem cells (mESC). RESULTS Zfp982 was expressed in stem cells, and bound to chromatin through a GCAGAGKC motif, for example near the stemness genes Nanog, Zfp42, and Dppa3. Nanog and Zfp42 were direct targets of ZFP982 that decreased in expression upon knockdown and increased upon overexpression of Zfp982. We show that ZFP982 expression strongly correlated with stem cell characteristics, both on the transcriptional and morphological levels. Zfp982 expression decreased with progressive differentiation into ecto-, endo- and mesodermal cell lineages, and knockdown of Zfp982 correlated with morphological and transcriptional features of differentiated cells. Zfp982 showed transcriptional overlap with members of the Hippo signaling pathway, one of which was Yap1, the major co-activator of Hippo signaling. Despite the observation that ZFP982 and YAP1 interacted and localized predominantly to the cytoplasm upon differentiation, the localization of YAP1 was not influenced by ZFP982 localization. CONCLUSIONS Together, our study identified ZFP982 as a transcriptional regulator of early stemness genes, and since ZFP982 is under the control of the Hippo pathway, underscored the importance of the context-dependent Hippo signals for stem cell characteristics.
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Affiliation(s)
- Fariba Dehghanian
- Department of Cell and Molecular Biology & Microbiology, Faculty of Biological Science and Technology, University of Isfahan, HezarJarib Street, Isfahan 81746-73441, Iran; Department of Molecular Embryology, Institute of Anatomy and Cell Biology, Faculty of Medicine, University of Freiburg, 79104 Freiburg, Germany.
| | - Patrick Piero Bovio
- Department of Molecular Embryology, Institute of Anatomy and Cell Biology, Faculty of Medicine, University of Freiburg, 79104 Freiburg, Germany; Faculty of Biology, University of Freiburg, Freiburg, Germany
| | - Fabian Gather
- Department of Molecular Embryology, Institute of Anatomy and Cell Biology, Faculty of Medicine, University of Freiburg, 79104 Freiburg, Germany
| | - Simone Probst
- Institute of Experimental and Clinical Pharmacology and Toxicology, Faculty of Medicine, University of Freiburg, Freiburg, Germany
| | - Amirhosein Naghsh-Nilchi
- Department of Cell and Molecular Biology & Microbiology, Faculty of Biological Science and Technology, University of Isfahan, HezarJarib Street, Isfahan 81746-73441, Iran
| | - Tanja Vogel
- Department of Molecular Embryology, Institute of Anatomy and Cell Biology, Faculty of Medicine, University of Freiburg, 79104 Freiburg, Germany; Centre for Basics in Neuromodulation (Neuromodul Basics), Freiburg, Germany.
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8
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Kwak S, Song CL, Cho YS, Choi I, Byun JE, Jung H, Lee J. Txnip regulates the Oct4-mediated pluripotency circuitry via metabolic changes upon differentiation. Cell Mol Life Sci 2024; 81:142. [PMID: 38485770 PMCID: PMC10940461 DOI: 10.1007/s00018-024-05161-y] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/14/2023] [Revised: 01/16/2024] [Accepted: 02/06/2024] [Indexed: 03/18/2024]
Abstract
Thioredoxin interacting protein (Txnip) is a stress-responsive factor regulating Trx1 for redox balance and involved in diverse cellular processes including proliferation, differentiation, apoptosis, inflammation, and metabolism. However, the biological role of Txnip function in stem cell pluripotency has yet to be investigated. Here, we reveal the novel functions of mouse Txnip in cellular reprogramming and differentiation onset by involving in glucose-mediated histone acetylation and the regulation of Oct4, which is a fundamental component of the molecular circuitry underlying pluripotency. During reprogramming or PSC differentiation process, cellular metabolic and chromatin remodeling occur in order to change its cellular fate. Txnip knockout promotes induced pluripotency but hinders initial differentiation by activating pluripotency factors and promoting glycolysis. This alteration affects the intracellular levels of acetyl-coA, a final product of enhanced glycolysis, resulting in sustained histone acetylation on active PSC gene regions. Moreover, Txnip directly interacts with Oct4, thereby repressing its activity and consequently deregulating Oct4 target gene transcriptions. Our work suggests that control of Txnip expression is crucial for cell fate transitions by modulating the entry and exit of pluripotency.
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Affiliation(s)
- Sojung Kwak
- Developmental Biology Laboratory, Environmental Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 34141, Republic of Korea
| | - Cho Lok Song
- Developmental Biology Laboratory, Environmental Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 34141, Republic of Korea
| | - Yee Sook Cho
- Stem Cell Research Laboratory, Immunotherapy Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 34141, Republic of Korea
- Department of Bioscience, KRIBB School, University of Science and Technology, Daejeon, 34141, Republic of Korea
| | - Inpyo Choi
- Immunotherapy Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 34141, Republic of Korea
| | - Jae-Eun Byun
- Aging Convergence Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 34141, Republic of Korea
- Department of Biochemistry, School of Life Sciences, Chungbuk National University, Cheongju, 28644, Republic of Korea
| | - Haiyoung Jung
- Department of Bioscience, KRIBB School, University of Science and Technology, Daejeon, 34141, Republic of Korea.
- Immunotherapy Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 34141, Republic of Korea.
- Aging Convergence Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 34141, Republic of Korea.
| | - Jungwoon Lee
- Developmental Biology Laboratory, Environmental Disease Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, 34141, Republic of Korea.
- Department of Bioscience, KRIBB School, University of Science and Technology, Daejeon, 34141, Republic of Korea.
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9
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Francia MG, Verneri P, Oses C, Vazquez Echegaray C, Garcia MR, Toro A, Levi V, Guberman AS. AKT1 induces Nanog promoter in a SUMOylation-dependent manner in different pluripotent contexts. BMC Res Notes 2023; 16:309. [PMID: 37919788 PMCID: PMC10623886 DOI: 10.1186/s13104-023-06598-3] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/31/2023] [Accepted: 10/26/2023] [Indexed: 11/04/2023] Open
Abstract
AKT/PKB is a kinase crucial for pluripotency maintenance in pluripotent stem cells. Multiple post-translational modifications modulate its activity. We have previously demonstrated that AKT1 induces the expression of the pluripotency transcription factor Nanog in a SUMOylation-dependent manner in mouse embryonic stem cells. Here, we studied different cellular contexts and main candidates that could mediate this induction. Our results strongly suggest the pluripotency transcription factors OCT4 and SOX2 are not essential mediators. Additionally, we concluded that this induction takes place in different pluripotent contexts but not in terminally differentiated cells. Finally, the cross-matching analysis of ESCs, iPSCs and MEFs transcriptomes and AKT1 phosphorylation targets provided new clues about possible factors that could be involved in the SUMOylation-dependent Nanog induction by AKT.
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Affiliation(s)
- Marcos Gabriel Francia
- Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales (IQUIBICEN), Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, CONICET, Universidad de Buenos Aires, Buenos Aires, Argentina
| | - Paula Verneri
- Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales (IQUIBICEN), Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, CONICET, Universidad de Buenos Aires, Buenos Aires, Argentina
| | - Camila Oses
- Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales (IQUIBICEN), Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, CONICET, Universidad de Buenos Aires, Buenos Aires, Argentina
| | - Camila Vazquez Echegaray
- Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales (IQUIBICEN), Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, CONICET, Universidad de Buenos Aires, Buenos Aires, Argentina
- Lund Stem Cell Center, Department of Molecular Medicine and Gene Therapy, Lund University, Lund, Sweden
| | - Mora Reneé Garcia
- Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales (IQUIBICEN), Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, CONICET, Universidad de Buenos Aires, Buenos Aires, Argentina
| | - Ayelen Toro
- Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales (IQUIBICEN), Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, CONICET, Universidad de Buenos Aires, Buenos Aires, Argentina
| | - Valeria Levi
- Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales (IQUIBICEN), Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, CONICET, Universidad de Buenos Aires, Buenos Aires, Argentina
| | - Alejandra Sonia Guberman
- Instituto de Química Biológica de la Facultad de Ciencias Exactas y Naturales (IQUIBICEN), Departamento de Química Biológica, Facultad de Ciencias Exactas y Naturales, CONICET, Universidad de Buenos Aires, Buenos Aires, Argentina.
- Departamento de Fisiología y Biología Molecular y Celular, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, Buenos Aires, Argentina.
- Laboratorio de Regulación Génica en Células Madre (CONICET-UBA), Intendente Guiraldes 2160 Pab. 2, 4to Piso, QB-71, C1428EGA, Buenos Aires, Argentina.
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10
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Kar S, Niharika, Roy A, Patra SK. Overexpression of SOX2 Gene by Histone Modifications: SOX2 Enhances Human Prostate and Breast Cancer Progression by Prevention of Apoptosis and Enhancing Cell Proliferation. Oncology 2023; 101:591-608. [PMID: 37549026 DOI: 10.1159/000531195] [Citation(s) in RCA: 8] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/22/2022] [Accepted: 05/02/2023] [Indexed: 08/09/2023]
Abstract
INTRODUCTION SOX2 plays a crucial role in tumor development, cancer stem cell maintenance, and cancer progression. Mechanisms of SOX2 gene regulation in human breast and prostate cancers are not established yet. METHODS SOX2 expression in prostate and breast cancer tissues and cell lines was determined by qRT-PCR, Western blot, and immunochemistry, followed by the investigation of pro-tumorigenic properties like cell proliferation, migration, and apoptosis by gene knockdown and treatment with epigenetic modulators and ChIP. RESULTS Prostate and breast cancer tissues showed very high expression of SOX2. All cancer cell lines DU145 and PC3 (prostate) and MCF7 and MDA-MB-231 (breast) exhibited high expression of SOX2. Inhibition of SOX2 drastically decreased cell proliferation and migration. Epigenetic modulators enhanced SOX2 gene expression in both cancer types. DNA methylation pattern in SOX2 promoter could not be appreciably counted for SOX2 overexpression. Activation of SOX2 gene promoter was due to very high deposition of H3K4me3 and H3K9acS10p and drastic decrease of H3K9me3 and H3K27me3. CONCLUSION Histone modification is crucial for the overexpression of SOX2 during tumor development and cancer progression. These findings show the avenue of co-targeting SOX2 and its active epigenetic modifier enzymes to effectively treat aggressive prostate and breast cancers.
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Affiliation(s)
- Swayamsiddha Kar
- Epigenetics and Cancer Research Laboratory, Biochemistry and Molecular Biology Group, Department of Life Science, National Institute of Technology, Rourkela, India
| | - Niharika
- Epigenetics and Cancer Research Laboratory, Biochemistry and Molecular Biology Group, Department of Life Science, National Institute of Technology, Rourkela, India
| | - Ankan Roy
- Epigenetics and Cancer Research Laboratory, Biochemistry and Molecular Biology Group, Department of Life Science, National Institute of Technology, Rourkela, India
| | - Samir Kumar Patra
- Epigenetics and Cancer Research Laboratory, Biochemistry and Molecular Biology Group, Department of Life Science, National Institute of Technology, Rourkela, India
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11
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Yu W, Peng Y, Peng X, Li Z, Liu C, Yang L, Gao Y, Liang S, Yuan B, Chen C, Kim NH, Jiang H, Zhang J. 6-Gingerol Improves In Vitro Porcine Embryo Development by Reducing Oxidative Stress. Animals (Basel) 2023; 13:ani13081315. [PMID: 37106877 PMCID: PMC10135256 DOI: 10.3390/ani13081315] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/25/2023] [Revised: 04/01/2023] [Accepted: 04/10/2023] [Indexed: 04/29/2023] Open
Abstract
6-Gingerol, the main active ingredient in ginger, exhibits a variety of biological activities, such as antioxidant, anti-inflammatory, and anticancer activities, and can affect cell development. However, the effects of 6-gingerol on mammalian reproductive processes, especially early embryonic development, are unclear. This study explored whether 6-gingerol can be used to improve the quality of in vitro-cultured porcine embryos. The results showed that 5 μM 6-gingerol significantly increased the blastocyst formation rates of porcine early embryos. 6-Gingerol attenuated intracellular reactive oxygen species accumulation and autophagy, increased intracellular glutathione levels, and increased mitochondrial activity. In addition, 6-gingerol upregulated NANOG, SRY-box transcription factor 2, cytochrome c oxidase subunit II, mechanistic target of rapamycin kinase, and RPTOR independent companion of MTOR complex 2 while downregulating Caspase 3, baculoviral IAP repeat containing 5, autophagy related 12, and Beclin 1. Most importantly, 6-gingerol significantly increased the levels of p-extracellular regulated protein kinase 1/2 while reducing the levels of p-c-Jun N-terminal kinase 1/2/3 and p-p38. These results indicate that 6-gingerol can promote the development of porcine early embryos in vitro.
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Affiliation(s)
- Wenjie Yu
- Department of Laboratory Animals, Jilin Provincial Key Laboratory of Animal Model, Jilin University, Changchun 130062, China
| | - Yanxia Peng
- Department of Laboratory Animals, Jilin Provincial Key Laboratory of Animal Model, Jilin University, Changchun 130062, China
| | - Xinyue Peng
- Department of Laboratory Animals, Jilin Provincial Key Laboratory of Animal Model, Jilin University, Changchun 130062, China
| | - Ze Li
- Department of Laboratory Animals, Jilin Provincial Key Laboratory of Animal Model, Jilin University, Changchun 130062, China
| | - Chang Liu
- School of Grains, Jilin Business and Technology College, Changchun 130507, China
| | - Liu Yang
- Tongyu Grassland Management Station, Changchun 137200, China
| | - Yan Gao
- Department of Laboratory Animals, Jilin Provincial Key Laboratory of Animal Model, Jilin University, Changchun 130062, China
| | - Shuang Liang
- Department of Laboratory Animals, Jilin Provincial Key Laboratory of Animal Model, Jilin University, Changchun 130062, China
| | - Bao Yuan
- Department of Laboratory Animals, Jilin Provincial Key Laboratory of Animal Model, Jilin University, Changchun 130062, China
| | - Chengzhen Chen
- Department of Laboratory Animals, Jilin Provincial Key Laboratory of Animal Model, Jilin University, Changchun 130062, China
| | - Nam-Hyung Kim
- Department of Laboratory Animals, Jilin Provincial Key Laboratory of Animal Model, Jilin University, Changchun 130062, China
- Department of Animal Science, Chungbuk National University, Cheongju 361-763, Chungbuk, Republic of Korea
| | - Hao Jiang
- Department of Laboratory Animals, Jilin Provincial Key Laboratory of Animal Model, Jilin University, Changchun 130062, China
- Department of Animal Science, Chungbuk National University, Cheongju 361-763, Chungbuk, Republic of Korea
| | - Jiabao Zhang
- Department of Laboratory Animals, Jilin Provincial Key Laboratory of Animal Model, Jilin University, Changchun 130062, China
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12
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Xu Y, Xu M, Li X, Weng X, Su Z, Zhang M, Tan J, Zeng H, Li X, Nie L, Gong J, Chen N, Chen X, Zhou Q. SOX9 and HMGB3 co-operatively transactivate NANOG and promote prostate cancer progression. Prostate 2023; 83:440-453. [PMID: 36541373 DOI: 10.1002/pros.24476] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/06/2021] [Revised: 06/03/2021] [Accepted: 06/29/2021] [Indexed: 12/24/2022]
Abstract
BACKGROUND The homeodomain-containing transcription factor NANOG is overexpressed in prostate adenocarcinoma (PCa) and predicts poor prognosis. The SOX family transcription factor SOX9, as well as the transcription co-activator HMGB3 of the HMGB family, are also overexpressed and may play pivotal roles in PCa. However, it is unknown whether SOX9 and HMGB3 interact with each other, or if they regulate NANOG gene transcription. METHODS We identified potential SOX9 responsive elements in NANOG promoter, and investigated if SOX9 regulated NANOG transcription in co-operation with HMGB3 by experimental analysis of potential SOX9 binding sites in NANOG promoter, reporter gene transcription assays with or without interference or artificial overexpression of SOX9 and/or HMGB3, and protein-binding assays of SOX9-HMGB3 interaction. Clinicopathologic and prognostic significance of SOX9-HMGB3 overexpression in PCa was analyzed. RESULTS SOX9 activated NANOG gene transcription by preferentially binding to a highly conserved consensus cis-regulatory element (-573 to -568) in NANOG promoter, and promoted the expression of NANOG downstream oncogenic genes. Importantly, HMGB3 functioned as a partner of SOX9 to co-operatively enhance transactivation of NANOG by interacting with SOX9, predominantly via the HMG Box A domain of HMGB3. Overexpression of SOX9 and/or HMGB3 enhanced PCa cell survival and cell migration and were significantly associated with PCa progression. Notably, Cox proportional regression analysis showed that co-overexpression of both SOX9 and HMGB3 was an independent unfavorable prognosticator for both CRPC-free survival (relative risk [RR] = 3.779，95% confidence interval [CI]: 1.159-12.322, p = 0.028) and overall survival (RR = 3.615，95% CI: 1.101-11.876, p = 0.034). CONCLUSIONS These findings showed a novel SOX9/HMGB3/NANOG regulatory mechanism, deregulation of which played important roles in PCa progression.
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Affiliation(s)
- Yunyi Xu
- Department of Pathology and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China
| | - Miao Xu
- Department of Pathology and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China
| | - Xinglan Li
- Department of Pathology and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China
| | - Xin Weng
- Department of Pathology and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China
| | - Zhengzheng Su
- Department of Pathology and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China
| | - Mengni Zhang
- Department of Pathology and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China
| | - Junya Tan
- Department of Pathology and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China
| | - Hao Zeng
- Department of Urology and Institute of Urology, West China Hospital, Sichuan University, Chengdu, China
| | - Xun Li
- Department of Ophthalmology and Research Laboratory of Ophthalmology and Vision Sciences, West China Hospital, Sichuan University, Chengdu, China
| | - Ling Nie
- Department of Pathology and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China
| | - Jing Gong
- Department of Pathology and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China
| | - Ni Chen
- Department of Pathology and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China
| | - Xueqin Chen
- Department of Pathology and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China
| | - Qiao Zhou
- Department of Pathology and State Key Laboratory of Biotherapy, West China Hospital, Sichuan University, Chengdu, China
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13
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Hernandez JC, Chen CL, Machida T, Uthaya Kumar DB, Tahara SM, Montana J, Sher L, Liang J, Jung JU, Tsukamoto H, Machida K. LIN28 and histone H3K4 methylase induce TLR4 to generate tumor-initiating stem-like cells. iScience 2023; 26:106254. [PMID: 36949755 PMCID: PMC10025994 DOI: 10.1016/j.isci.2023.106254] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/03/2020] [Revised: 01/09/2022] [Accepted: 02/16/2023] [Indexed: 02/24/2023] Open
Abstract
Chemoresistance and plasticity of tumor-initiating stem-like cells (TICs) promote tumor recurrence and metastasis. The gut-originating endotoxin-TLR4-NANOG oncogenic axis is responsible for the genesis of TICs. This study investigated mechanisms as to how TICs arise through transcriptional, epigenetic, and post-transcriptional activation of oncogenic TLR4 pathways. Here, we expressed constitutively active TLR4 (caTLR4) in mice carrying pLAP-tTA or pAlb-tTA, under a tetracycline withdrawal-inducible system. Liver progenitor cell induction accelerated liver tumor development in caTLR4-expressing mice. Lentiviral shRNA library screening identified histone H3K4 methylase SETD7 as central to activation of TLR4. SETD7 combined with hypoxia induced TLR4 through HIF2 and NOTCH. LIN28 post-transcriptionally stabilized TLR4 mRNA via de-repression of let-7 microRNA. These results supported a LIN28-TLR4 pathway for the development of HCCs in a hypoxic microenvironment. These findings not only advance our understanding of molecular mechanisms responsible for TIC generation in HCC, but also represent new therapeutic targets for the treatment of HCC.
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Affiliation(s)
- Juan Carlos Hernandez
- Departments of Molecular Microbiology and Immunology, University of Southern California, Los Angeles, CA 90033, USA
- MS Biotechnology Program, California State University Channel Islands, Camarillo, CA 93012, USA
| | - Chia-Lin Chen
- Departments of Molecular Microbiology and Immunology, University of Southern California, Los Angeles, CA 90033, USA
- Department of Life Sciences & Institute of Genome Sciences, National Yang Ming Chiao Tung University, Taipei 110, Taiwan
| | - Tatsuya Machida
- Departments of Molecular Microbiology and Immunology, University of Southern California, Los Angeles, CA 90033, USA
| | - Dinesh Babu Uthaya Kumar
- Departments of Molecular Microbiology and Immunology, University of Southern California, Los Angeles, CA 90033, USA
| | - Stanley M. Tahara
- Departments of Molecular Microbiology and Immunology, University of Southern California, Los Angeles, CA 90033, USA
| | - Jared Montana
- Departments of Molecular Microbiology and Immunology, University of Southern California, Los Angeles, CA 90033, USA
| | - Linda Sher
- Department of Surgery, University of Southern California, Los Angeles, CA 90033, USA
| | | | - Jae U. Jung
- Departments of Molecular Microbiology and Immunology, University of Southern California, Los Angeles, CA 90033, USA
| | - Hidekazu Tsukamoto
- Department of Pathology, University of Southern California, Los Angeles, CA 90033, USA
- Southern California Research Center for ALPD and Cirrhosis, Los Angeles, CA, USA
- Department of Veterans Affairs Greater Los Angeles Healthcare System, Los Angeles, CA, USA
| | - Keigo Machida
- Departments of Molecular Microbiology and Immunology, University of Southern California, Los Angeles, CA 90033, USA
- Southern California Research Center for ALPD and Cirrhosis, Los Angeles, CA, USA
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14
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Karami Fath M, Garousi S, Mottahedi M, Ghasemzadeh N, Salmani K, Olfati F, Beit Saeed M, Sotoudeh S, Barati G. The role of hypoxia-inducible factors in breast cancer stem cell specification. Pathol Res Pract 2023; 243:154349. [PMID: 36791562 DOI: 10.1016/j.prp.2023.154349] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/24/2022] [Revised: 01/24/2023] [Accepted: 01/27/2023] [Indexed: 01/31/2023]
Abstract
Breast tumor is heterogeneous cancer with high morbidity and mortality rates, particularly in developing countries. Despite new efforts to reduce the breast cancer implications, the number of newly diagnosed cases is increasing worldwide. It is believed that cancer stem cells (CSCs) are responsible for the implication of cancers including breast cancer. Although CSCs compose a small population in tumor bulks, they play a crucial role in tumor initiation, progression, metastasis, and chemotherapeutic resistance. These events are mediated by the hypoxia-inducible factor (HIF) pathway which regulates the transcription of genes involved in CSC maintenance and tumorigenesis. In this review, we discussed the mechanisms by which hypoxia- or chemotherapy-induced HIFs promote breast CSC specification.
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Affiliation(s)
- Mohsen Karami Fath
- Department of Cellular and Molecular Biology, Faculty of Biological Sciences, Kharazmi University, Tehran, Iran
| | - Setareh Garousi
- Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Mehran Mottahedi
- Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran
| | | | - Kiana Salmani
- Department of Genetics, Faculty of Advanced Science and Technology, Tehran Medical Sciences, Islamic Azad University, Tehran, Iran
| | - Fatemeh Olfati
- Department of Reproductive Health, Faculty of Nursing and Midwifery, Tehran University of Medical Sciences, Tehran, Iran
| | - Miad Beit Saeed
- Faculty of Nursing and Midwifery, Abadan Islamic Azad University, Abadan, Iran
| | - Sina Sotoudeh
- Faculty of Nursing and Midwifery, Guilan University of Medical Sciences, Guilan, Iran
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15
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von Eyben FE, Kristiansen K, Kapp DS, Hu R, Preda O, Nogales FF. Epigenetic Regulation of Driver Genes in Testicular Tumorigenesis. Int J Mol Sci 2023; 24:ijms24044148. [PMID: 36835562 PMCID: PMC9966837 DOI: 10.3390/ijms24044148] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/19/2022] [Revised: 02/15/2023] [Accepted: 02/15/2023] [Indexed: 02/22/2023] Open
Abstract
In testicular germ cell tumor type II (TGCT), a seminoma subtype expresses an induced pluripotent stem cell (iPSC) panel with four upregulated genes, OCT4/POU5F1, SOX17, KLF4, and MYC, and embryonal carcinoma (EC) has four upregulated genes, OCT4/POU5F1, SOX2, LIN28, and NANOG. The EC panel can reprogram cells into iPSC, and both iPSC and EC can differentiate into teratoma. This review summarizes the literature on epigenetic regulation of the genes. Epigenetic mechanisms, such as methylations of cytosines on the DNA string and methylations and acetylations of histone 3 lysines, regulate expression of these driver genes between the TGCT subtypes. In TGCT, the driver genes contribute to well-known clinical characteristics and the driver genes are also important for aggressive subtypes of many other malignancies. In conclusion, epigenetic regulation of the driver genes are important for TGCT and for oncology in general.
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Affiliation(s)
- Finn E. von Eyben
- Center for Tobacco Control Research, Birkevej 17, 5230 Odense, Denmark
- Correspondence: ; Tel.: +45-66145862
| | - Karsten Kristiansen
- Laboratory of Genomics and Molecular Biomedicine, August Krogh Building Department of Biology, University of Copenhagen, Universitetsparken 13, 2100 Copenhagen, Denmark
- BGI-Research, BGI-Shenzhen, Shenzhen 518120, China
- Institute of Metagenomics, Qingdao-Europe Advanced Institute for Life Sciences, BGI-Qingdao, Qingdao 166555, China
| | - Daniel S. Kapp
- Department of Radiation Oncology, Stanford University, Stanford, CA 94305, USA
| | - Rong Hu
- Department of Pathology, Laboratory Medicine, University of Wisconsin Hospital and Clinics, Madison, WI 53792, USA
| | - Ovidiu Preda
- Department of Pathology, San Cecilio University Hospital, 18071 Granada, CP, Spain
| | - Francisco F. Nogales
- Department of Pathology, School of Medicine, University Granada, 18071 Granada, CP, Spain
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16
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Choi EB, Vodnala M, Saini P, Anugula S, Zerbato M, Ho JJ, Wang J, Ho Sui SJ, Yoon J, Roels M, Inouye C, Fong YW. Transcription factor SOX15 regulates stem cell pluripotency and promotes neural fate during differentiation by activating the neurogenic gene Hes5. J Biol Chem 2023; 299:102996. [PMID: 36764520 PMCID: PMC10023989 DOI: 10.1016/j.jbc.2023.102996] [Citation(s) in RCA: 7] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/05/2022] [Revised: 02/01/2023] [Accepted: 02/02/2023] [Indexed: 02/11/2023] Open
Abstract
SOX2 and SOX15 are Sox family transcription factors enriched in embryonic stem cells (ESCs). The role of SOX2 in activating gene expression programs essential for stem cell self-renewal and acquisition of pluripotency during somatic cell reprogramming is well-documented. However, the contribution of SOX15 to these processes is unclear and often presumed redundant with SOX2 largely because overexpression of SOX15 can partially restore self-renewal in SOX2-deficient ESCs. Here, we show that SOX15 contributes to stem cell maintenance by cooperating with ESC-enriched transcriptional coactivators to ensure optimal expression of pluripotency-associated genes. We demonstrate that SOX15 depletion compromises reprogramming of fibroblasts to pluripotency which cannot be compensated by SOX2. Ectopic expression of SOX15 promotes the reversion of a postimplantation, epiblast stem cell state back to a preimplantation, ESC-like identity even though SOX2 is expressed in both cell states. We also uncover a role of SOX15 in lineage specification, by showing that loss of SOX15 leads to defects in commitment of ESCs to neural fates. SOX15 promotes neural differentiation by binding to and activating a previously uncharacterized distal enhancer of a key neurogenic regulator, Hes5. Together, these findings identify a multifaceted role of SOX15 in induction and maintenance of pluripotency and neural differentiation.
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Affiliation(s)
- Eun-Bee Choi
- Brigham Regenerative Medicine Center, Brigham and Women's Hospital, Boston, Massachusetts, USA; Department of Medicine, Cardiovascular Medicine Division, Harvard Medical School, Boston, Massachusetts, USA; Harvard Stem Cell Institute, Cambridge, Massachusetts, USA
| | - Munender Vodnala
- Brigham Regenerative Medicine Center, Brigham and Women's Hospital, Boston, Massachusetts, USA; Department of Medicine, Cardiovascular Medicine Division, Harvard Medical School, Boston, Massachusetts, USA; Harvard Stem Cell Institute, Cambridge, Massachusetts, USA
| | - Prince Saini
- Brigham Regenerative Medicine Center, Brigham and Women's Hospital, Boston, Massachusetts, USA; Department of Medicine, Cardiovascular Medicine Division, Harvard Medical School, Boston, Massachusetts, USA; Harvard Stem Cell Institute, Cambridge, Massachusetts, USA
| | - Sharath Anugula
- Brigham Regenerative Medicine Center, Brigham and Women's Hospital, Boston, Massachusetts, USA; Department of Medicine, Cardiovascular Medicine Division, Harvard Medical School, Boston, Massachusetts, USA; Harvard Stem Cell Institute, Cambridge, Massachusetts, USA
| | - Madeleine Zerbato
- Brigham Regenerative Medicine Center, Brigham and Women's Hospital, Boston, Massachusetts, USA; Department of Medicine, Cardiovascular Medicine Division, Harvard Medical School, Boston, Massachusetts, USA; Harvard Stem Cell Institute, Cambridge, Massachusetts, USA
| | - Jaclyn J Ho
- Department of Molecular and Cell Biology, Li Ka Shing Center for Biomedical and Health Sciences, California Institute for Regenerative Medicine Center of Excellence, University of California at Berkeley, Berkeley, California, USA; Howard Hughes Medical Institute, Berkeley, California, USA
| | - Jianing Wang
- Brigham Regenerative Medicine Center, Brigham and Women's Hospital, Boston, Massachusetts, USA; Department of Medicine, Cardiovascular Medicine Division, Harvard Medical School, Boston, Massachusetts, USA; Harvard Stem Cell Institute, Cambridge, Massachusetts, USA
| | - Shannan J Ho Sui
- Harvard Chan Bioinformatics Core, Harvard T.H. Chan School of Public Health, Boston, Massachusetts, USA
| | - Joon Yoon
- Harvard Chan Bioinformatics Core, Harvard T.H. Chan School of Public Health, Boston, Massachusetts, USA
| | - Marielle Roels
- Brigham Regenerative Medicine Center, Brigham and Women's Hospital, Boston, Massachusetts, USA; Department of Medicine, Cardiovascular Medicine Division, Harvard Medical School, Boston, Massachusetts, USA
| | - Carla Inouye
- Department of Molecular and Cell Biology, Li Ka Shing Center for Biomedical and Health Sciences, California Institute for Regenerative Medicine Center of Excellence, University of California at Berkeley, Berkeley, California, USA; Howard Hughes Medical Institute, Berkeley, California, USA
| | - Yick W Fong
- Brigham Regenerative Medicine Center, Brigham and Women's Hospital, Boston, Massachusetts, USA; Department of Medicine, Cardiovascular Medicine Division, Harvard Medical School, Boston, Massachusetts, USA; Harvard Stem Cell Institute, Cambridge, Massachusetts, USA.
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17
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Park J, Park S, Lee JS. Role of the Paf1 complex in the maintenance of stem cell pluripotency and development. FEBS J 2023; 290:951-961. [PMID: 35869661 DOI: 10.1111/febs.16582] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/05/2022] [Revised: 06/26/2022] [Accepted: 07/21/2022] [Indexed: 11/30/2022]
Abstract
Cell identity is determined by the transcriptional regulation of a cell-type-specific gene group. The Paf1 complex (Paf1C), an RNA polymerase II-associating factor, is an important transcriptional regulator that not only participates in transcription elongation and termination but also affects transcription-coupled histone modifications and chromatin organisation. Recent studies have shown that Paf1C is involved in the expression of genes required for self-renewal and pluripotency in stem cells and tumorigenesis. In this review, we focused on the role of Paf1C as a critical transcriptional regulator in cell fate decisions. Paf1C affects the pluripotency of stem cells by regulating the expression of core transcription factors such as Oct4 and Nanog. In addition, Paf1C directly binds to the promoters or distant elements of target genes, thereby maintaining the pluripotency in embryonic stem cells derived from an early stage of the mammalian embryo. Paf1C is upregulated in cancer stem cells, as compared with that in cancer cells, suggesting that Paf1C may be a target for cancer therapy. Interestingly, Paf1C is involved in multiple developmental stages in Drosophila, zebrafish, mice and even humans, thereby displaying a trend for the correlation between Paf1C and cell fate. Thus, we propose that Paf1C is a critical contributor to cell differentiation, cell specification and its characteristics and could be employed as a therapeutic target in developmental diseases.
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Affiliation(s)
- Jiyeon Park
- Department of Molecular Bioscience, College of Biomedical Science, Kangwon National University, Chuncheon-si, Korea
| | - Shinae Park
- Department of Molecular Bioscience, College of Biomedical Science, Kangwon National University, Chuncheon-si, Korea
| | - Jung-Shin Lee
- Department of Molecular Bioscience, College of Biomedical Science, Kangwon National University, Chuncheon-si, Korea
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18
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Patel I, Parchem RJ. Regulation of Oct4 in stem cells and neural crest cells. Birth Defects Res 2022; 114:983-1002. [PMID: 35365980 PMCID: PMC9525453 DOI: 10.1002/bdr2.2007] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/03/2022] [Revised: 02/25/2022] [Accepted: 03/14/2022] [Indexed: 12/30/2022]
Abstract
During embryonic development, cells gradually restrict their developmental potential as they exit pluripotency and differentiate into various cell types. The POU transcription factor Oct4 (encoded by Pou5f1) lies at the center of the pluripotency machinery that regulates stemness and differentiation in stem cells, and is required for reprogramming of somatic cells into induced pluripotent stem cells (iPSCs). Several studies have revealed that Oct4 and other stemness genes are also expressed in multipotent cell populations such as neural crest cells (NCCs), and are required to expand the NCC developmental potential. Transcriptional regulation of Oct4 has been studied extensively in stem cells during early embryonic development and reprogramming, but not in NCCs. Here, we review how Oct4 is regulated in pluripotent stem cells, and address some of the gaps in knowledge about regulation of the pluripotency network in NCCs.
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Affiliation(s)
- Ivanshi Patel
- Department of Molecular and Cellular BiologyBaylor College of MedicineHoustonTexasUSA,Stem Cells and Regenerative Medicine Center, Center for Cell and Gene TherapyBaylor College of MedicineHoustonTexasUSA,Dan L Duncan Comprehensive Cancer CenterBaylor College of MedicineHoustonTexasUSA
| | - Ronald J. Parchem
- Department of Molecular and Cellular BiologyBaylor College of MedicineHoustonTexasUSA,Stem Cells and Regenerative Medicine Center, Center for Cell and Gene TherapyBaylor College of MedicineHoustonTexasUSA,Dan L Duncan Comprehensive Cancer CenterBaylor College of MedicineHoustonTexasUSA
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19
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Glover HJ, Shparberg RA, Morris MB. L-Proline Supplementation Drives Self-Renewing Mouse Embryonic Stem Cells to a Partially Primed Pluripotent State: The Early Primitive Ectoderm-Like Cell. METHODS IN MOLECULAR BIOLOGY (CLIFTON, N.J.) 2022; 2490:11-24. [PMID: 35486235 DOI: 10.1007/978-1-0716-2281-0_2] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
Abstract
Mouse embryonic stem cells (mESCs) can be grown under a variety of culture conditions as discrete cell states along the pluripotency continuum, ranging from the least mature "ground state" to being "primed" to differentiate. Cells along this continuum are demarcated by differences in gene expression, X chromosome inactivation, ability to form chimeras and epigenetic marks. We have developed a protocol to differentiate "naïve" mESCs to a "partially primed" state by adding the amino acid L-proline to self-renewal medium. These cells express the primitive ectoderm markers Dnmt3b and Fgf5, and are thus called early primitive ectoderm-like (EPL) cells. In addition to changes in gene expression, these cells undergo a morphological change to flattened, dispersed colonies, have an increased proliferation rate, and a predisposition to neural fate. EPL cells can be used to study the cell states along the pluripotency continuum, peri-implantation embryogenesis, and as a starting point for efficient neuronal differentiation.
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Affiliation(s)
- Hannah J Glover
- Bosch Institute and Discipline of Physiology, School of Medical Sciences, University of Sydney, Camperdown, NSW, Australia.
| | - Rachel A Shparberg
- Bosch Institute and Discipline of Physiology, School of Medical Sciences, University of Sydney, Camperdown, NSW, Australia
| | - Michael B Morris
- Bosch Institute and Discipline of Physiology, School of Medical Sciences, University of Sydney, Camperdown, NSW, Australia.
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Zhong Z, Xu Y, Feng Y, Ao L, Jiang Y. Characterization of the Nanog gene involved in the gonadal development in pearlscale angelfish (Centropyge vrolikii). FISH PHYSIOLOGY AND BIOCHEMISTRY 2022; 48:303-319. [PMID: 35138521 DOI: 10.1007/s10695-022-01054-8] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/05/2021] [Accepted: 01/30/2022] [Indexed: 06/14/2023]
Abstract
The homeodomain transcription factor Nanog plays a crucial role in the embryonic and gonadal development and the maintenance of embryonic stem cells (ESCs), interacting with transcription factors such as Oct4 and Sox2 in mammals. Nevertheless, its pathways to molecular mechanisms remain unclear as to teleosts. This study investigates the role of the Nanog gene in gonadal development and sex reversal of pearlscale angelfish (Centropyge vrolikii). To understand the expression pattern of gonadal development, we identified the Nanog gene of C. vrolikii, which we named Cv-Nanog. The full-length cDNA sequence of Cv-Nanog was 2,136 bp in length and encoded a homeodomain protein of 436 amino acid residues. The gene structure and western blot prove results that Cv-Nanog was homologous to the Nanog gene of mammalians. The protein sequence comparison demonstrates that the Cv-Nanog shared a high degree of similarity with orthologs from other vertebrates in the conserved homeodomain. The Cv-Nanog gene was substantially expressed in gonads, and the expression was significantly higher in the ovaries than in the testis, according to quantitative real-time PCR (qRT-PCR) and western blot analyses. In situ hybridization reveals that the transcripts were located in the cytoplasm and membrane of the oocytes in the ovaries and testes. The expression of Cv-Nanog mRNA was weak in Sertoli cells but strong in germ cells. After overexpression of Cv-Nanog, the expression levels of pluripotent factors Sox2 and Oct4 increased significantly with 21.5-fold and 12.2-fold, respectively. Simultaneously, the TGF-beta signaling pathway was activated, and the gonadal cell growth was promoted. The expression of ovary-bias genes Cyp19a and Foxl2 was upregulated, and the expression of testis-bias genes Sox9 and Dmrt1 was downregulated to promote ovarian development. These results imply that the Nanog gene might play a crucial role in the process of gonadal development and sexual reversion in C. vrolikii. This study provides new insight to understand the molecular regulatory mechanism of the Nanog gene further and important clues for the future studies in gonadal development.
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Affiliation(s)
- Zhaowei Zhong
- Key Laboratory of Healthy Mariculture for the East China Sea, Ministry of Agriculture and Rural Affairs, Fisheries College, Jimei University, Xiamen, 361021, China
- National Demonstration Center for Experimental Aquatic Science and Technology Education, Jimei University, Xiamen, 361021, China
| | - Yan Xu
- Key Laboratory of Healthy Mariculture for the East China Sea, Ministry of Agriculture and Rural Affairs, Fisheries College, Jimei University, Xiamen, 361021, China
- National Demonstration Center for Experimental Aquatic Science and Technology Education, Jimei University, Xiamen, 361021, China
| | - Yan Feng
- Key Laboratory of Healthy Mariculture for the East China Sea, Ministry of Agriculture and Rural Affairs, Fisheries College, Jimei University, Xiamen, 361021, China
- National Demonstration Center for Experimental Aquatic Science and Technology Education, Jimei University, Xiamen, 361021, China
| | - Lulu Ao
- Key Laboratory of Healthy Mariculture for the East China Sea, Ministry of Agriculture and Rural Affairs, Fisheries College, Jimei University, Xiamen, 361021, China
- National Demonstration Center for Experimental Aquatic Science and Technology Education, Jimei University, Xiamen, 361021, China
| | - Yonghua Jiang
- Key Laboratory of Healthy Mariculture for the East China Sea, Ministry of Agriculture and Rural Affairs, Fisheries College, Jimei University, Xiamen, 361021, China.
- National Demonstration Center for Experimental Aquatic Science and Technology Education, Jimei University, Xiamen, 361021, China.
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21
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Zaveri L, Dhawan J. Inducible expression of Oct-3/4 reveals synergy with Klf4 in targeting Cyclin A2 to enhance proliferation during early reprogramming. Biochem Biophys Res Commun 2022; 587:29-35. [PMID: 34864392 DOI: 10.1016/j.bbrc.2021.11.058] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2021] [Revised: 11/02/2021] [Accepted: 11/15/2021] [Indexed: 11/18/2022]
Abstract
During reprogramming of somatic cells, heightened proliferation is one of the earliest changes observed. While other early events such as mesenchymal-to-epithelial transition have been well studied, the mechanisms by which the cell cycle switches from a slow cycling state to a faster cycling state are still incompletely understood. To investigate the role of Oct-3/4 in this early transition, we created a 4-Hydroxytamoxifen (OHT) dependent Oct-3/4 Estrogen Receptor fusion (OctER). We confirmed that OctER can substitute for Oct-3/4 to reprogram mouse embryonic fibroblasts to a pluripotent state. During the early stages of reprograming, Oct-3/4 and Klf4 individually did not affect cell proliferation but in combination hastened the cell cycle. Using OctER + Klf4, we found that proliferative enhancement is OHT dose-dependent, suggesting that OctER is the driver of this transition. We identified Cyclin A2 as a likely target of Oct-3/4 + Klf4. In mESC, Klf4 and Oct-3/4 bind ∼100bp upstream of Cyclin A2 CCRE, suggesting a potential regulatory role. Using inducible OctER, we show a dose-dependent induction of Cyclin A2 promoter-reporter activity. Taken together, our results suggest that Cyclin A2 is a key early target during reprogramming, and support the view that a rapid cell cycle assists the transition to pluripotency.
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Affiliation(s)
- Lamuk Zaveri
- Institute for Stem Cell Science and Regenerative Medicine, Bengaluru, 560068, India; CSIR-Centre for Cellular and Molecular Biology, Hyderabad, 500007, India; Manipal Academy of Higher Education, Manipal, Karnataka, India
| | - Jyotsna Dhawan
- Institute for Stem Cell Science and Regenerative Medicine, Bengaluru, 560068, India; CSIR-Centre for Cellular and Molecular Biology, Hyderabad, 500007, India.
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22
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Zhang T, Zhou H, Wang K, Wang X, Wang M, Zhao W, Xi X, Li Y, Cai M, Zhao W, Xu Y, Shao R. Role, molecular mechanism and the potential target of breast cancer stem cells in breast cancer development. Biomed Pharmacother 2022; 147:112616. [PMID: 35008001 DOI: 10.1016/j.biopha.2022.112616] [Citation(s) in RCA: 23] [Impact Index Per Article: 7.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/24/2021] [Revised: 01/01/2022] [Accepted: 01/02/2022] [Indexed: 02/06/2023] Open
Abstract
Breast cancer (BC) is one of the most common malignant tumors in women globally, and its occurrence has surpassed lung cancer and become the biggest threat for women. At present, breast cancer treatment includes surgical resection or postoperative chemotherapy and radiotherapy. However, tumor relapse and metastasis usually lead to current therapy failure thanks to breast cancer stem cells (BCSCs)-mediated tumorigenicity and drug resistance. Drug resistance is mainly due to the long-term quiescent G0 phase, strong DNA repairability, and high expression of ABC transporter, and the tumorigenicity is reflected in the activation of various proliferation pathways related to BCSCs. Therefore, understanding the characteristics of BCSCs and their intracellular and extracellular molecular mechanisms is crucial for the development of targeted drugs for BCSCs. To this end, we discussed the latest developments in BCSCs research, focusing on the analysis of specific markers, critical signaling pathways that maintain the stemness of BCSCs，such as NOTCH, Wnt/β-catenin, STAT3, Hedgehog, and Hippo-YAP signaling, immunomicroenviroment and summarizes targeting therapy strategies for stemness maintenance and differentiation, which provides a theoretical basis for further exploration of treating breast cancer and preventing relapse derived from BCSCs.
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Affiliation(s)
- Tianshu Zhang
- Key Laboratory of Antibiotic Bioengineering, Ministry of Health, Laboratory of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
| | - Huimin Zhou
- Key Laboratory of Antibiotic Bioengineering, Ministry of Health, Laboratory of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
| | - Kexin Wang
- Key Laboratory of Antibiotic Bioengineering, Ministry of Health, Laboratory of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
| | - Xiaowei Wang
- Key Laboratory of Antibiotic Bioengineering, Ministry of Health, Laboratory of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
| | - Mengyan Wang
- Key Laboratory of Antibiotic Bioengineering, Ministry of Health, Laboratory of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
| | - Wenxia Zhao
- Key Laboratory of Antibiotic Bioengineering, Ministry of Health, Laboratory of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
| | - Xiaoming Xi
- Key Laboratory of Antibiotic Bioengineering, Ministry of Health, Laboratory of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
| | - Yang Li
- Key Laboratory of Antibiotic Bioengineering, Ministry of Health, Laboratory of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
| | - Meilian Cai
- Key Laboratory of Antibiotic Bioengineering, Ministry of Health, Laboratory of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
| | - Wuli Zhao
- Key Laboratory of Antibiotic Bioengineering, Ministry of Health, Laboratory of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China.
| | - Yanni Xu
- Key Laboratory of Antibiotic Bioengineering, Ministry of Health, Laboratory of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China.
| | - Rongguang Shao
- Key Laboratory of Antibiotic Bioengineering, Ministry of Health, Laboratory of Oncology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China
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23
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German OL, Vallese-Maurizi H, Soto TB, Rotstein NP, Politi LE. Retina stem cells, hopes and obstacles. World J Stem Cells 2021; 13:1446-1479. [PMID: 34786153 PMCID: PMC8567457 DOI: 10.4252/wjsc.v13.i10.1446] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/28/2021] [Revised: 07/14/2021] [Accepted: 09/17/2021] [Indexed: 02/07/2023] Open
Abstract
Retinal degeneration is a major contributor to visual dysfunction worldwide. Although it comprises several eye diseases, loss of retinal pigment epithelial (RPE) and photoreceptor cells are the major contributors to their pathogenesis. Early therapies included diverse treatments, such as provision of anti-vascular endothelial growth factor and many survival and trophic factors that, in some cases, slow down the progression of the degeneration, but do not effectively prevent it. The finding of stem cells (SC) in the eye has led to the proposal of cell replacement strategies for retina degeneration. Therapies using different types of SC, such as retinal progenitor cells (RPCs), embryonic SC, pluripotent SCs (PSCs), induced PSCs (iPSCs), and mesenchymal stromal cells, capable of self-renewal and of differentiating into multiple cell types, have gained ample support. Numerous preclinical studies have assessed transplantation of SC in animal models, with encouraging results. The aim of this work is to revise the different preclinical and clinical approaches, analyzing the SC type used, their efficacy, safety, cell attachment and integration, absence of tumor formation and immunorejection, in order to establish which were the most relevant and successful. In addition, we examine the questions and concerns still open in the field. The data demonstrate the existence of two main approaches, aimed at replacing either RPE cells or photoreceptors. Emerging evidence suggests that RPCs and iPSC are the best candidates, presenting no ethical concerns and a low risk of immunorejection. Clinical trials have already supported the safety and efficacy of SC treatments. Serious concerns are pending, such as the risk of tumor formation, lack of attachment or integration of transplanted cells into host retinas, immunorejection, cell death, and also ethical. However, the amazing progress in the field in the last few years makes it possible to envisage safe and effective treatments to restore vision loss in a near future.
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Affiliation(s)
- Olga L German
- Department of Biology, Biochemistry and Pharmacy, Universidad Nacional del Sur, Bahia blanca 8000, Buenos Aires, Argentina
- Department of Biology, Biochemistry and Pharmacy, Universidad Nacional del Sur, and Neurobiology Department, Instituto de Investigaciones Bioquímicas de Bahía Blanca (INIBIBB) Conicet, Bahía Blanca 8000, Buenos Aires, Argentina
| | - Harmonie Vallese-Maurizi
- Department of Biology, Biochemistry and Pharmacy, Universidad Nacional del Sur, Bahia blanca 8000, Buenos Aires, Argentina
- Department of Biology, Biochemistry and Pharmacy, Universidad Nacional del Sur, and Neurobiology Department, Instituto de Investigaciones Bioquímicas de Bahía Blanca (INIBIBB) Conicet, Bahía Blanca 8000, Buenos Aires, Argentina
| | - Tamara B Soto
- Department of Biology, Biochemistry and Pharmacy, Universidad Nacional del Sur, and Neurobiology Department, Instituto de Investigaciones Bioquímicas de Bahía Blanca (INIBIBB) Conicet, Bahía Blanca 8000, Buenos Aires, Argentina
| | - Nora P Rotstein
- Department of Biology, Biochemistry and Pharmacy, Universidad Nacional del Sur, Bahia blanca 8000, Buenos Aires, Argentina
- Department of Biology, Biochemistry and Pharmacy, Universidad Nacional del Sur, and Neurobiology Department, Instituto de Investigaciones Bioquímicas de Bahía Blanca (INIBIBB) Conicet, Bahía Blanca 8000, Buenos Aires, Argentina
| | - Luis Enrique Politi
- Department of Biology, Biochemistry and Pharmacy, Universidad Nacional del Sur, and Neurobiology Department, Instituto de Investigaciones Bioquímicas de Bahía Blanca (INIBIBB) Conicet, Bahía Blanca 8000, Buenos Aires, Argentina
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24
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Veraguas-Davila D, Cordero MF, Saez S, Saez-Ruiz D, Gonzalez A, Saravia F, Castro FO, Rodriguez-Alvarez L. Domestic cat embryos generated without zona pellucida are capable of developing in vitro but exhibit abnormal gene expression and a decreased implantation rate. Theriogenology 2021; 174:36-46. [PMID: 34416562 DOI: 10.1016/j.theriogenology.2021.08.013] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/07/2021] [Revised: 08/10/2021] [Accepted: 08/11/2021] [Indexed: 11/25/2022]
Abstract
The removal of the zona pellucida has been used to improve the in vitro development of domestic cat embryos generated by IVF and SCNT. However, the in vivo development of domestic cat embryos generated without the zona pellucida has not been evaluated. The objective of this study was to evaluate the effects of zona pellucida removal on the in vitro and in vivo development of domestic cat embryos generated by IVF. For this purpose, two experimental groups were created: 1) domestic cat embryos cultured in vitro (Zona-intact group, ZI) and 2) domestic cat embryos cultured in vitro without the zona pellucida (Zona-free group, ZF). Domestic cat embryos were generated by IVF and cultured in vitro for 8 days. In the ZF group, the zona pellucida was removed after IVF, and embryos were cultured using the well of the well system (WOW). Cleavage, morula and blastocyst rates were evaluated in both groups. The diameter and total cell number of blastocysts were assessed. Relative expression of pluripotency (OCT4, SOX2 and NANOG), differentiation (CDX2 and GATA6) and apoptotic markers (BAX and BCL2) was evaluated in blastocysts. Finally, to evaluate in vivo development, embryos at days 5, 6 and 7 of development were transferred into recipient domestic cats, and ultrasonography was performed to evaluate implantation. No differences were observed in the cleavage, morula or blastocyst rates between embryos from the ZI and ZF groups. The diameter (mean ± SD) of blastocysts from the ZF group was greater (253.4 ± 83.3 μm) than that from the ZI group (210.5 ± 78.5 μm). No differences were observed in the relative expression of OCT4, CDX2 or GATA6. However, the relative expression of SOX2 and NANOG was significantly reduced in ZF blastocysts compared to ZI blastocysts. Furthermore, the relative expression of BAX was higher in ZF blastocysts than in ZI blastocysts. Finally, four pregnancies were confirmed after the transfer of ZI embryos (n = 110). However, no pregnancies were observed after the transfer of ZF embryos at the morula or blastocyst stage (n = 56). In conclusion, domestic cat embryos cultured without the zona pellucida were able to develop in vitro until the blastocyst stage. However, the removal of the zona pellucida negatively affected the gene expression of pluripotency and apoptosis markers, and ZF embryos were unable to implant. This might indicate that the removal of the zona pellucida is detrimental for the implantation and in vivo development of domestic cat embryos.
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Affiliation(s)
- Daniel Veraguas-Davila
- Department of Animal Science, Faculty of Veterinary Sciences, Universidad de Concepcion, Chillán, Chile.
| | - Maria Francisca Cordero
- Department of Animal Science, Faculty of Veterinary Sciences, Universidad de Concepcion, Chillán, Chile
| | - Soledad Saez
- Department of Animal Science, Faculty of Veterinary Sciences, Universidad de Concepcion, Chillán, Chile
| | - Darling Saez-Ruiz
- Department of Animal Science, Faculty of Veterinary Sciences, Universidad de Concepcion, Chillán, Chile
| | - Alejandro Gonzalez
- Department of Clinical Science, Faculty of Veterinary Sciences, Universidad de Concepcion, Chillán, Chile
| | - Fernando Saravia
- Department of Animal Science, Faculty of Veterinary Sciences, Universidad de Concepcion, Chillán, Chile
| | - Fidel Ovidio Castro
- Department of Animal Science, Faculty of Veterinary Sciences, Universidad de Concepcion, Chillán, Chile
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25
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Ghafouri-Fard S, Moghadam MHB, Shoorei H, Bahroudi Z, Taheri M, Taheriazam A. The impact of non-coding RNAs on normal stem cells. Biomed Pharmacother 2021; 142:112050. [PMID: 34426251 DOI: 10.1016/j.biopha.2021.112050] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2021] [Revised: 08/02/2021] [Accepted: 08/12/2021] [Indexed: 12/11/2022] Open
Abstract
Self-renewal and differentiation into diverse cells are two main characteristics of stem cells. These cells have important roles in development and homeostasis of different tissues and are supposed to facilitate tissue regeneration. Function of stem cells is regulated by dynamic interactions between external signaling, epigenetic factors, and molecules that regulate expression of genes. Among the highly appreciated regulators of function of stem cells are long non-coding RNAs (lncRNAs) and microRNAs (miRNAs). Impact of miR-342-5p, miR-145, miR-1297, miR-204-5p, miR-132, miR-128-3p, hsa-miR-302, miR-26b-5p and miR-10a are among miRNAs that regulate function of stem cells. Among lncRNAs, AK141205, ANCR, MEG3, Pnky, H19, TINCR, HULC, EPB41L4A-AS1 and SNHG7 have important roles in the regulation of stem cells. In the current paper, we aimed at reviewing the importance of miRNAs and lncRNAs in differentiation of stem cells both in normal and diseased conditions. For this purpose, we searched PubMed/Medline and google scholar databases using "stem cell" AND "lncRNA", or "long non-coding RNA", or "microRNA" or "miRNA".
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Affiliation(s)
- Soudeh Ghafouri-Fard
- Department of Medical Genetics, School of Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran
| | | | - Hamed Shoorei
- Department of Anatomical Sciences, Faculty of Medicine, Birjand University of Medical Sciences, Birjand, Iran
| | - Zahra Bahroudi
- Department of Anatomical Sciences, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran
| | - Mohammad Taheri
- Skull Base Research Center, Loghman Hakim Hospital, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
| | - Afshin Taheriazam
- Department of Orthopedics, Tehran Medical Sciences Branch, Islamic Azad University, Tehran, Iran.
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26
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Auxin-degron system identifies immediate mechanisms of OCT4. Stem Cell Reports 2021; 16:1818-1831. [PMID: 34143975 PMCID: PMC8282470 DOI: 10.1016/j.stemcr.2021.05.016] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/29/2020] [Revised: 05/19/2021] [Accepted: 05/20/2021] [Indexed: 12/11/2022] Open
Abstract
The pluripotency factor OCT4 is essential for the maintenance of naive pluripotent stem cells in vitro and in vivo. However, the specific role of OCT4 in this process remains unknown. Here, we developed a rapid protein-level OCT4 depletion system that demonstrates that the immediate downstream response to loss of OCT4 is reduced expression of key pluripotency factors. Our data show a requirement for OCT4 for the efficient transcription of several key pluripotency factors and suggest that expression of trophectoderm markers is a subsequent event. In addition, we find that NANOG is able to bind to the genome in the absence of OCT4, and this binding is in fact enhanced. Globally, however, the active enhancer-associated histone mark H3K27ac is depleted. Our work establishes that, while OCT4 is required for the maintenance of the naive transcription factor network, at a normal embryonic stem cell levels it antagonizes this network through inhibition of NANOG binding.
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27
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Tan A, Prasad R, Jho EH. TFEB regulates pluripotency transcriptional network in mouse embryonic stem cells independent of autophagy-lysosomal biogenesis. Cell Death Dis 2021; 12:343. [PMID: 33795648 PMCID: PMC8016867 DOI: 10.1038/s41419-021-03632-9] [Citation(s) in RCA: 16] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2020] [Revised: 03/17/2021] [Accepted: 03/17/2021] [Indexed: 12/20/2022]
Abstract
Transcription factor EB (TFEB), a well-known master regulator of autophagy and lysosomal biogenesis, is a member of the microphthalmia family of transcription factors (MiT family). Over the years, TFEB has been shown to have diverse roles in various physiological processes such as clearance for intracellular pathogenic factors and having developmental functions such as dendritic maturation, as well as osteoclast, and endoderm differentiation. However, in the present study, we propose a novel mechanism for TFEB governing pluripotency of mouse ESCs (mESCs) by regulating the pluripotency transcriptional network (PTN) in these cells. We observed high levels of TFEB mRNA and protein levels in undifferentiated mESCs. Interestingly, we found a reduction of Nanog and Sox2 levels in TFEB knockout (KO) mESCs while pluripotency was maintained as there was an upregulation of TFE3, a potent stem cell maintenance factor. In consistent, double knockout of TFEB/TFE3 (TFEB/3 DKO) reduced mESC pluripotency, as indicated by the loss of ESC morphology, reduction of ESC markers, and the emergence of differentiation markers. We further discovered that Nanog was a TFEB target gene in undifferentiated mESCs. TFEB also promoted sex-determining region Y-box2 (Sox2) transcription by forming a heterodimer with Sox2 in mESCs. Notably, Sox2, Oct4, and Nanog were also binding to the TFEB promoter and thus generating a feed-forward loop in relation to TFEB. Although high levels of nuclear TFEB are expected to enhance autophagy-lysosomal activity, undifferentiated mESC remarkably displayed low basal autophagy-lysosomal activity. Overexpression or knockout of TFEB did not affect the expression of TFEB lysosomal-autophagy target genes and TFEB also had a lesser binding affinity to its own lysosomal promoter-target genes in mESCs compared to differentiated cells. Collectively, these findings define a newly incorporative, moonlighting function for TFEB in regulating PTN, independent of its autophagy-lysosomal biogenesis roles.
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Affiliation(s)
- Anderson Tan
- Department of Life Science, University of Seoul, Seoul, 130-743, Republic of Korea
| | - Renuka Prasad
- Department of Life Science, University of Seoul, Seoul, 130-743, Republic of Korea
| | - Eek-Hoon Jho
- Department of Life Science, University of Seoul, Seoul, 130-743, Republic of Korea.
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28
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Jindal GA, Farley EK. Enhancer grammar in development, evolution, and disease: dependencies and interplay. Dev Cell 2021; 56:575-587. [PMID: 33689769 PMCID: PMC8462829 DOI: 10.1016/j.devcel.2021.02.016] [Citation(s) in RCA: 72] [Impact Index Per Article: 18.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2020] [Revised: 02/15/2021] [Accepted: 02/16/2021] [Indexed: 12/19/2022]
Abstract
Each language has standard books describing that language's grammatical rules. Biologists have searched for similar, albeit more complex, principles relating enhancer sequence to gene expression. Here, we review the literature on enhancer grammar. We introduce dependency grammar, a model where enhancers encode information based on dependencies between enhancer features shaped by mechanistic, evolutionary, and biological constraints. Classifying enhancers based on the types of dependencies may identify unifying principles relating enhancer sequence to gene expression. Such rules would allow us to read the instructions for development within genomes and pinpoint causal enhancer variants underlying disease and evolutionary changes.
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Affiliation(s)
- Granton A Jindal
- Division of Cardiology, Department of Medicine, University of California San Diego, La Jolla, CA 92093, USA; Division of Biological Sciences, Section of Molecular Biology, University of California San Diego, La Jolla, CA 92093, USA
| | - Emma K Farley
- Division of Cardiology, Department of Medicine, University of California San Diego, La Jolla, CA 92093, USA; Division of Biological Sciences, Section of Molecular Biology, University of California San Diego, La Jolla, CA 92093, USA.
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29
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Choi HJ, Jin SD, Rengaraj D, Kim JH, Pain B, Han JY. Differential transcriptional regulation of the NANOG gene in chicken primordial germ cells and embryonic stem cells. J Anim Sci Biotechnol 2021; 12:40. [PMID: 33658075 PMCID: PMC7931612 DOI: 10.1186/s40104-021-00563-5] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/21/2020] [Accepted: 01/26/2021] [Indexed: 01/06/2023] Open
Abstract
Background NANOG is a core transcription factor (TF) in embryonic stem cells (ESCs) and primordial germ cells (PGCs). Regulation of the NANOG gene by TFs, epigenetic factors, and autoregulatory factors is well characterized in ESCs, and transcriptional regulation of NANOG is well established in these cells. Although NANOG plays a key role in germ cells, the molecular mechanism underlying its transcriptional regulation in PGCs has not been studied. Therefore, we investigated the mechanism that regulates transcription of the chicken NANOG (cNANOG) gene in PGCs and ESCs. Results We first identified the transcription start site of cNANOG by 5′-rapid amplification of cDNA ends PCR analysis. Then, we measured the promoter activity of various 5′ flanking regions of cNANOG in chicken PGCs and ESCs using the luciferase reporter assay. cNANOG expression required transcriptional regulatory elements, which were positively regulated by POU5F3 (OCT4) and SOX2 and negatively regulated by TP53 in PGCs. The proximal region of the cNANOG promoter contains a positive transcriptional regulatory element (CCAAT/enhancer-binding protein (CEBP)-binding site) in ESCs. Furthermore, small interfering RNA-mediated knockdown demonstrated that POU5F3, SOX2, and CEBP played a role in cell type-specific transcription of cNANOG. Conclusions We show for the first time that different trans-regulatory elements control transcription of cNANOG in a cell type-specific manner. This finding might help to elucidate the mechanism that regulates cNANOG expression in PGCs and ESCs.
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Affiliation(s)
- Hee Jung Choi
- Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Sciences, Seoul National University, Seoul, 08826, South Korea
| | - So Dam Jin
- Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Sciences, Seoul National University, Seoul, 08826, South Korea
| | - Deivendran Rengaraj
- Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Sciences, Seoul National University, Seoul, 08826, South Korea
| | - Jin Hwa Kim
- Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Sciences, Seoul National University, Seoul, 08826, South Korea
| | - Bertrand Pain
- Univ Lyon, Universite ́Lyon 1, INSERM, INRAE, Stem Cell and Brain Research Institute, U1208, USC1361, 69500, Bron, France
| | - Jae Yong Han
- Department of Agricultural Biotechnology and Research Institute of Agriculture and Life Sciences, Seoul National University, Seoul, 08826, South Korea. .,Institute for Biomedical Sciences, Shinshu University, Minamiminowa, Nagano, 399-4598, Japan.
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30
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Stirparo GG, Kurowski A, Yanagida A, Bates LE, Strawbridge SE, Hladkou S, Stuart HT, Boroviak TE, Silva JCR, Nichols J. OCT4 induces embryonic pluripotency via STAT3 signaling and metabolic mechanisms. Proc Natl Acad Sci U S A 2021; 118:e2008890118. [PMID: 33452132 PMCID: PMC7826362 DOI: 10.1073/pnas.2008890118] [Citation(s) in RCA: 38] [Impact Index Per Article: 9.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/28/2022] Open
Abstract
OCT4 is a fundamental component of the molecular circuitry governing pluripotency in vivo and in vitro. To determine how OCT4 establishes and protects the pluripotent lineage in the embryo, we used comparative single-cell transcriptomics and quantitative immunofluorescence on control and OCT4 null blastocyst inner cell masses at two developmental stages. Surprisingly, activation of most pluripotency-associated transcription factors in the early mouse embryo occurs independently of OCT4, with the exception of the JAK/STAT signaling machinery. Concurrently, OCT4 null inner cell masses ectopically activate a subset of trophectoderm-associated genes. Inspection of metabolic pathways implicates the regulation of rate-limiting glycolytic enzymes by OCT4, consistent with a role in sustaining glycolysis. Furthermore, up-regulation of the lysosomal pathway was specifically detected in OCT4 null embryos. This finding implicates a requirement for OCT4 in the production of normal trophectoderm. Collectively, our findings uncover regulation of cellular metabolism and biophysical properties as mechanisms by which OCT4 instructs pluripotency.
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Affiliation(s)
- Giuliano G Stirparo
- Wellcome Trust-Medical Research Council Stem Cell Institute, Jeffrey Cheah Biomedical Centre, University of Cambridge, CB2 0AW Cambridge, United Kingdom;
- Living Systems Institute, University of Exeter, EX4 4QD Exeter, United Kingdom
| | - Agata Kurowski
- Wellcome Trust-Medical Research Council Stem Cell Institute, Jeffrey Cheah Biomedical Centre, University of Cambridge, CB2 0AW Cambridge, United Kingdom
- Department of Pharmacological Sciences, Icahn School of Medicine at Mount Sinai, New York, NY 10029
| | - Ayaka Yanagida
- Wellcome Trust-Medical Research Council Stem Cell Institute, Jeffrey Cheah Biomedical Centre, University of Cambridge, CB2 0AW Cambridge, United Kingdom
- Living Systems Institute, University of Exeter, EX4 4QD Exeter, United Kingdom
| | - Lawrence E Bates
- Wellcome Trust-Medical Research Council Stem Cell Institute, Jeffrey Cheah Biomedical Centre, University of Cambridge, CB2 0AW Cambridge, United Kingdom
- Department of Biochemistry, University of Cambridge, CB2 1GA Cambridge, United Kingdom
| | - Stanley E Strawbridge
- Wellcome Trust-Medical Research Council Stem Cell Institute, Jeffrey Cheah Biomedical Centre, University of Cambridge, CB2 0AW Cambridge, United Kingdom
| | - Siarhei Hladkou
- Wellcome Trust-Medical Research Council Stem Cell Institute, Jeffrey Cheah Biomedical Centre, University of Cambridge, CB2 0AW Cambridge, United Kingdom
- Department of Biochemistry, University of Cambridge, CB2 1GA Cambridge, United Kingdom
| | - Hannah T Stuart
- Wellcome Trust-Medical Research Council Stem Cell Institute, Jeffrey Cheah Biomedical Centre, University of Cambridge, CB2 0AW Cambridge, United Kingdom
| | - Thorsten E Boroviak
- Department of Physiology, Development and Neuroscience, University of Cambridge, CB2 3EG Cambridge, United Kingdom
- Centre for Trophoblast Research, University of Cambridge, CB2 3EG Cambridge, United Kingdom
| | - Jose C R Silva
- Wellcome Trust-Medical Research Council Stem Cell Institute, Jeffrey Cheah Biomedical Centre, University of Cambridge, CB2 0AW Cambridge, United Kingdom
- Department of Biochemistry, University of Cambridge, CB2 1GA Cambridge, United Kingdom
| | - Jennifer Nichols
- Wellcome Trust-Medical Research Council Stem Cell Institute, Jeffrey Cheah Biomedical Centre, University of Cambridge, CB2 0AW Cambridge, United Kingdom;
- Department of Physiology, Development and Neuroscience, University of Cambridge, CB2 3EG Cambridge, United Kingdom
- Centre for Trophoblast Research, University of Cambridge, CB2 3EG Cambridge, United Kingdom
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31
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Veraguas D, Aguilera C, Echeverry D, Saez-Ruiz D, Castro FO, Rodriguez-Alvarez L. Embryo aggregation allows the production of kodkod (Leopardus guigna) blastocysts after interspecific SCNT. Theriogenology 2020; 158:148-157. [PMID: 32961350 DOI: 10.1016/j.theriogenology.2020.09.006] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/05/2020] [Revised: 07/30/2020] [Accepted: 09/06/2020] [Indexed: 12/22/2022]
Abstract
The kodkod (Leopardus guigna) is a small felid endemic of Chile and is considered a vulnerable species. Domestic cat oocytes have been successfully used as recipient cytoplast to reprogram somatic cells from different felids by interspecific somatic cell nuclear transfer (iSCNT). The developmental competence of felid embryos generated by iSCNT can be improved by the aggregation method using a zona-free culture system. The objective of this research was to evaluate the developmental competence of kodkod embryos generated by iSCNT using domestic cat oocytes and the aggregation method. For this purpose, five experimental group were done: (1) cat embryos generated by IVF, (2) cat embryos generated by SCNT (Ca1x), (3) aggregated cat embryos generated by SCNT (Ca2x), (4) kodkod embryos generated by iSCNT (K1x) and (5) aggregated kodkod embryos generated by iSCNT (K2x). Cleavage, morulae and blastocyst rates were estimated. The blastocyst diameter was evaluated. The gene expression level of pluripotency (OCT4, SOX2 and NANOG) and differentiation markers (CDX2 and GATA6) was analyzed in blastocysts. Morulae rate was higher in the IVF group and when cloned embryos were cultured in aggregates (IVF: 68.2%, Ca2x: 58.0% and K2x: 62.4%) compared to individually cultured kodkod embryos (K1x: 37.0%) (P < 0.05). Embryo aggregation increased blastocysts formation in the Ca2x group (30.9%) to a similar rate compared to the IVF group (44.5%) (P > 0.05). No blastocysts were generated in the K1x group, whereas blastocysts formation was obtained in K2x group (5.9%). The diameter of blastocysts from the K2x group (172.8 μm) was significantly lower than blastocysts from the Ca2x group (P < 0.05). The relative expression of OCT4 was lower in blastocysts from Ca1x than in blastocysts from IVF (P < 0.05). Furthermore, CDX2 expression was lower in blastocysts from Ca2x than in blastocysts from Ca1x and IVF groups (P < 0.05). In kodkod embryos, only one blastocyst from the K2x group expressed OCT4. No expression of SOX2, NANOG, CDX2 and GATA6 was detected in kodkod blastocysts. In conclusion, after iSCNT, domestic cat oocytes support the development of kodkod embryos until the morula stage. The aggregation method increases the morulae rate of kodkod cloned embryos and allows blastocysts formation. However, kodkod blastocysts have a poor morphological quality and a lacking expression of pluripotency and differentiation markers, probably caused by an incomplete nuclear reprogramming.
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Affiliation(s)
- Daniel Veraguas
- Department of Animal Science, Faculty of Veterinary Sciences, Universidad de Concepcion, Chillán, Chile
| | - Constanza Aguilera
- Department of Animal Science, Faculty of Veterinary Sciences, Universidad de Concepcion, Chillán, Chile
| | - Diana Echeverry
- Department of Animal Science, Faculty of Veterinary Sciences, Universidad de Concepcion, Chillán, Chile
| | - Darling Saez-Ruiz
- Department of Animal Science, Faculty of Veterinary Sciences, Universidad de Concepcion, Chillán, Chile
| | - Fidel Ovidio Castro
- Department of Animal Science, Faculty of Veterinary Sciences, Universidad de Concepcion, Chillán, Chile
| | - Lleretny Rodriguez-Alvarez
- Department of Animal Science, Faculty of Veterinary Sciences, Universidad de Concepcion, Chillán, Chile.
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32
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Cui W, Marcho C, Wang Y, Degani R, Golan M, Tremblay KD, Rivera-Pérez JA, Mager J. MED20 is essential for early embryogenesis and regulates NANOG expression. Reproduction 2020; 157:215-222. [PMID: 30571656 DOI: 10.1530/rep-18-0508] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/25/2018] [Accepted: 12/17/2018] [Indexed: 12/15/2022]
Abstract
Mediator is an evolutionarily conserved multi-subunit complex, bridging transcriptional activators and repressors to the general RNA polymerase II (Pol II) initiation machinery. Though the Mediator complex is crucial for the transcription of almost all Pol II promoters in eukaryotic organisms, the phenotypes of individual Mediator subunit mutants are each distinct. Here, we report for the first time, the essential role of subunit MED20 in early mammalian embryo development. Although Med20 mutant mouse embryos exhibit normal morphology at E3.5 blastocyst stage, they cannot be recovered at early post-gastrulation stages. Outgrowth assays show that mutant blastocysts cannot hatch from the zona pellucida, indicating impaired blastocyst function. Assessments of cell death and cell lineage specification reveal that apoptosis, inner cell mass, trophectoderm and primitive endoderm markers are normal in mutant blastocysts. However, the epiblast marker NANOG is ectopically expressed in the trophectoderm of Med20 mutants, indicative of defects in trophoblast specification. These results suggest that MED20 specifically, and the Mediator complex in general, are essential for the earliest steps of mammalian development and cell lineage specification.
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Affiliation(s)
- Wei Cui
- Department of Veterinary and Animal Sciences, University of Massachusetts, Amherst, Massachusetts, USA.,Animal Models Core Facility, Institute for Applied Life Sciences (IALS), University of Massachusetts, Amherst, Massachusetts, USA
| | - Chelsea Marcho
- Department of Veterinary and Animal Sciences, University of Massachusetts, Amherst, Massachusetts, USA
| | - Yongsheng Wang
- Key Laboratory of Animal Biotechnology of the Ministry of Agriculture, College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, PR China
| | - Rinat Degani
- Department of Veterinary and Animal Sciences, University of Massachusetts, Amherst, Massachusetts, USA
| | - Morgane Golan
- Department of Veterinary and Animal Sciences, University of Massachusetts, Amherst, Massachusetts, USA
| | - Kimberly D Tremblay
- Department of Veterinary and Animal Sciences, University of Massachusetts, Amherst, Massachusetts, USA
| | - Jaime A Rivera-Pérez
- Division of Genes and Development, Department of Pediatrics, University of Massachusetts Medical School, Worcester, Massachusetts, USA
| | - Jesse Mager
- Department of Veterinary and Animal Sciences, University of Massachusetts, Amherst, Massachusetts, USA
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33
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Wuputra K, Ku CC, Wu DC, Lin YC, Saito S, Yokoyama KK. Prevention of tumor risk associated with the reprogramming of human pluripotent stem cells. J Exp Clin Cancer Res 2020; 39:100. [PMID: 32493501 PMCID: PMC7268627 DOI: 10.1186/s13046-020-01584-0] [Citation(s) in RCA: 46] [Impact Index Per Article: 9.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2020] [Accepted: 04/22/2020] [Indexed: 02/07/2023] Open
Abstract
Human pluripotent embryonic stem cells have two special features: self-renewal and pluripotency. It is important to understand the properties of pluripotent stem cells and reprogrammed stem cells. One of the major problems is the risk of reprogrammed stem cells developing into tumors. To understand the process of differentiation through which stem cells develop into cancer cells, investigators have attempted to identify the key factors that generate tumors in humans. The most effective method for the prevention of tumorigenesis is the exclusion of cancer cells during cell reprogramming. The risk of cancer formation is dependent on mutations of oncogenes and tumor suppressor genes during the conversion of stem cells to cancer cells and on the environmental effects of pluripotent stem cells. Dissecting the processes of epigenetic regulation and chromatin regulation may be helpful for achieving correct cell reprogramming without inducing tumor formation and for developing new drugs for cancer treatment. This review focuses on the risk of tumor formation by human pluripotent stem cells, and on the possible treatment options if it occurs. Potential new techniques that target epigenetic processes and chromatin regulation provide opportunities for human cancer modeling and clinical applications of regenerative medicine.
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Affiliation(s)
- Kenly Wuputra
- Graduate Institute of Medicine, Kaohsiung Medical University, 100 Shih-Chuan 1st Rd., San-Ming District, Kaohsiung, 807, Taiwan
- Regenerative Medicine and Cell Therapy Research Center, Kaohsiung Medical University Hospital, Kaohsiung, 807, Taiwan
| | - Chia-Chen Ku
- Graduate Institute of Medicine, Kaohsiung Medical University, 100 Shih-Chuan 1st Rd., San-Ming District, Kaohsiung, 807, Taiwan
- Regenerative Medicine and Cell Therapy Research Center, Kaohsiung Medical University Hospital, Kaohsiung, 807, Taiwan
| | - Deng-Chyang Wu
- Regenerative Medicine and Cell Therapy Research Center, Kaohsiung Medical University Hospital, Kaohsiung, 807, Taiwan
- Division of Gastroenterology, Department of Internal Medicine, Kaohsiung Medical University Hospital, Kaohsiung, 807, Taiwan
| | - Ying-Chu Lin
- School of Dentistry, School of Medicine, Kaohsiung Medical University, Kaohsiung, 807, Taiwan
| | - Shigeo Saito
- Waseda University Research Institute for Science and Engineering, Shinjuku, Tokyo, 162-8480, Japan.
- Saito Laboratory of Cell Technology Institute, Yaita, Tochigi, 329-1571, Japan.
| | - Kazunari K Yokoyama
- Graduate Institute of Medicine, Kaohsiung Medical University, 100 Shih-Chuan 1st Rd., San-Ming District, Kaohsiung, 807, Taiwan.
- Regenerative Medicine and Cell Therapy Research Center, Kaohsiung Medical University Hospital, Kaohsiung, 807, Taiwan.
- Waseda University Research Institute for Science and Engineering, Shinjuku, Tokyo, 162-8480, Japan.
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34
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Emura N, Saito Y, Miura R, Sawai K. Effect of Downregulating the Hippo Pathway Members YAP1 and LATS2 Transcripts on Early Development and Gene Expression Involved in Differentiation in Porcine Embryos. Cell Reprogram 2020; 22:62-70. [PMID: 32150685 DOI: 10.1089/cell.2019.0082] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/03/2023] Open
Abstract
In mouse development, differentiation of the inner cell mass (ICM) and trophectoderm (TE) during the transition from the morula to blastocyst stage is regulated by the Hippo pathway; however, the functions of the Hippo pathway in porcine embryogenesis have not been investigated. In the present study, we examined the gene expression patterns of the Hippo pathway members yes-associated protein 1 (YAP1) and large tumor suppressor 2 (LATS2) and the functions of these genes during porcine preimplantation development using RNA interference. Both YAP1 and LATS2 mRNA levels were shown high in the in vitro matured oocytes and 1-cell stage embryos and fell progressively with development. YAP1 nuclear localization was detected at the morula and blastocyst stages. Downregulation of either YAP1 or LATS2 inhibited porcine preimplantation development and affected the expression levels of POU class 5 homeobox 1 (OCT-4) and SRY-related HMG-box gene 2 (SOX2), transcription factors necessary for the ICM/TE differentiation. Taken together, YAP1 and LATS2 are essential for porcine preimplantation development, and it is possible that the Hippo pathway has important roles in porcine ICM/TE segregation.
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Affiliation(s)
- Natsuko Emura
- The United Graduate School of Agricultural Sciences, Iwate University, Morioka, Japan
| | - Yuriko Saito
- Faculty of Agriculture, Iwate University, Morioka, Japan
| | - Ruri Miura
- Faculty of Agriculture, Iwate University, Morioka, Japan
| | - Ken Sawai
- The United Graduate School of Agricultural Sciences, Iwate University, Morioka, Japan.,Faculty of Agriculture, Iwate University, Morioka, Japan
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35
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Inomata C, Yuikawa T, Nakayama-Sadakiyo Y, Kobayashi K, Ikeda M, Chiba M, Konishi C, Ishioka A, Tsuda S, Yamasu K. Involvement of an Oct4-related PouV gene, pou5f3/pou2, in neurogenesis in the early neural plate of zebrafish embryos. Dev Biol 2020; 457:30-42. [DOI: 10.1016/j.ydbio.2019.09.002] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/07/2019] [Revised: 09/09/2019] [Accepted: 09/10/2019] [Indexed: 01/03/2023]
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36
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Park SW, Do HJ, Choi W, Kim JH. Fli-1 promotes proliferation and upregulates NANOGP8 expression in T-lymphocyte leukemia cells. Biochimie 2019; 168:1-9. [PMID: 31626853 DOI: 10.1016/j.biochi.2019.10.005] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/03/2019] [Accepted: 10/10/2019] [Indexed: 11/27/2022]
Abstract
Friend leukemia integration 1 (Fli-1) is a member of the E26 transformation-specific (ETS) transcription factor family. Fli-1 regulates normal hematopoiesis and vasculogenesis, and its aberrant expression underlies virus-induced leukemias and various types of human cancers. NANOGP8, a retro-pseudogene of stem cell mediator NANOG, is expressed predominantly in cancer cells and plays a role in tumorigenesis. In this study, we demonstrate that Fli-1 expression enhances human acute T-cell leukemia Jurkat cell proliferation and that Fli-1 acts as a transcriptional activator of NANOGP8 expression in these cells. NANOGP8 and Fli-1 are highly expressed in Jurkat cells, whereas NANOG was undetectable at both the RNA and protein levels. Moreover, the expression of endogenous NANOGP8 was significantly influenced by gain of function and loss of function of Fli-1. Promoter-reporter assays showed that NANOGP8 transcription was significantly upregulated by dose-dependent Fli-1 overexpression. A series of deletion mutagenesis of NANOGP8 promoter sequence revealed that NANOGP8 promoter activity was tightly regulated and found the minimal promoter region sufficient to activate NANOGP8 transcription mediated by Fli-1. Moreover, site-directed mutagenesis of the putative binding site abolished both NANOGP8 full-length and minimal promoter activities. Binding assays revealed that Fli-1 directly interacts with the potent binding site in NANOG promoter region. Taken together, our data demonstrate that Fli-1 is a novel upstream transcriptional activator of NANOGP8 and provide the molecular details of Fli-1-mediated NANOGP8 gene expression. Ultimately, these findings may contribute to understanding the expanded regulatory mechanisms of oncogenic NANOGP8 and ETS family transcription factors in leukemogenesis.
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Affiliation(s)
- Sung-Won Park
- Department of Biomedical Science, College of Life Science, CHA University, Seongnam-Si, Gyeonggi-Do, 13488, South Korea
| | - Hyun-Jin Do
- Department of Biomedical Science, College of Life Science, CHA University, Seongnam-Si, Gyeonggi-Do, 13488, South Korea
| | - Wonbin Choi
- Department of Biomedical Science, College of Life Science, CHA University, Seongnam-Si, Gyeonggi-Do, 13488, South Korea
| | - Jae-Hwan Kim
- Department of Biomedical Science, College of Life Science, CHA University, Seongnam-Si, Gyeonggi-Do, 13488, South Korea.
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37
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Terada M, Kawamata M, Kimura R, Sekiya S, Nagamatsu G, Hayashi K, Horisawa K, Suzuki A. Generation of
Nanog
reporter mice that distinguish pluripotent stem cells from unipotent primordial germ cells. Genesis 2019; 57:e23334. [DOI: 10.1002/dvg.23334] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/17/2019] [Revised: 08/26/2019] [Accepted: 08/27/2019] [Indexed: 12/25/2022]
Affiliation(s)
- Maiko Terada
- Division of Organogenesis and Regeneration Medical Institute of Bioregulation, Kyushu University Fukuoka Japan
| | - Masaki Kawamata
- Division of Organogenesis and Regeneration Medical Institute of Bioregulation, Kyushu University Fukuoka Japan
| | - Ryota Kimura
- Division of Organogenesis and Regeneration Medical Institute of Bioregulation, Kyushu University Fukuoka Japan
| | - Sayaka Sekiya
- Division of Organogenesis and Regeneration Medical Institute of Bioregulation, Kyushu University Fukuoka Japan
| | - Go Nagamatsu
- Department of Stem Cell Biology and Medicine Graduate School of Medical Sciences, Kyushu University Fukuoka Japan
| | - Katsuhiko Hayashi
- Department of Stem Cell Biology and Medicine Graduate School of Medical Sciences, Kyushu University Fukuoka Japan
| | - Kenichi Horisawa
- Division of Organogenesis and Regeneration Medical Institute of Bioregulation, Kyushu University Fukuoka Japan
| | - Atsushi Suzuki
- Division of Organogenesis and Regeneration Medical Institute of Bioregulation, Kyushu University Fukuoka Japan
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38
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Chen B, Zhu Z, Li L, Ye W, Zeng J, Gao J, Wang S, Zhang L, Huang Z. Effect of overexpression of Oct4 and Sox2 genes on the biological and oncological characteristics of gastric cancer cells. Onco Targets Ther 2019; 12:4667-4682. [PMID: 31417271 PMCID: PMC6592062 DOI: 10.2147/ott.s209734] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/23/2019] [Accepted: 05/07/2019] [Indexed: 11/23/2022] Open
Abstract
Objective: Using the gastric cancer cell line SGC7901, we constructed a cell line that overexpressed octamer-binding protein 4 (Oct4) and SRY-box 2 (Sox2) to explore the stem cell oncological and biological characteristics of these cells and to elucidate the mechanisms of Oct4 and Sox2 in cancer. Methods: A lentiviral vector containing the Sox2 gene was constructed and transfected into a gastric cancer cell line overexpressing Oct4 (SGC7901-Oct4) to obtain a stably transfected cell line (SGC7901-Oct4-Sox2). Oct4 and Sox2 expression was detected by RT-PCR and Western blotting. The proliferation, drug resistance, migration, and invasion abilities of the cells were assessed using in vitro (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium), drug resistance, scratch-wound migration, transwell migration, transwell invasion, and spherical clone formation assays, and their tumorigenic ability was assessed using a tumor formation experiment in mice. Results: Compared with the control group, the expression of Oct4, Sox2, CD44, and E-cadherin was significantly higher in the group that overexpressed Oct4 and Sox2, while the expression of c-Myc and Klf4 did not significantly change. The proliferation, drug resistance, migration, and invasion abilities were significantly enhanced in the overexpression group, and the tumorigenic ability in mice was also significantly enhanced, with significantly increased tumor size and weight. Conclusion: The proliferation, drug resistance, migration, invasion, and tumorigenic abilities of SGC7901 cells overexpressing Oct4 and Sox2 were significantly improved. Oct4 and Sox2 play important roles in the proliferation, migration, invasion, and tumorigenicity of gastric cancer cells, and the two genes may be synergistic to a certain degree.
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Affiliation(s)
- Borong Chen
- Department of Gastrointestinal Surgery, Xiamen Cancer Hospital, The First Affiliated Hospital of Xiamen University, Xiamen, Fujian 361003, People's Republic of China
| | - Zhipeng Zhu
- Department of Gastrointestinal Surgery, Xiamen Cancer Hospital, The First Affiliated Hospital of Xiamen University, Xiamen, Fujian 361003, People's Republic of China
| | - Lulu Li
- Department of Gastrointestinal Surgery, Xiamen Cancer Hospital, The First Affiliated Hospital of Xiamen University, Xiamen, Fujian 361003, People's Republic of China
| | - Weipeng Ye
- Department of Clinical Medicine, Fujian Medical University, Fuzhou, Fujian 350004, People's Republic of China
| | - Junjie Zeng
- Department of Gastrointestinal Surgery, Xiamen Cancer Hospital, The First Affiliated Hospital of Xiamen University, Xiamen, Fujian 361003, People's Republic of China
| | - Jin Gao
- Department of Gastrointestinal Surgery, Xiamen Cancer Hospital, The First Affiliated Hospital of Xiamen University, Xiamen, Fujian 361003, People's Republic of China
| | - Shengjie Wang
- Department of Gastrointestinal Surgery, Xiamen Cancer Hospital, The First Affiliated Hospital of Xiamen University, Xiamen, Fujian 361003, People's Republic of China
| | - Liang Zhang
- Department of Gastrointestinal Surgery, Xiamen Cancer Hospital, The First Affiliated Hospital of Xiamen University, Xiamen, Fujian 361003, People's Republic of China
| | - Zhengjie Huang
- Department of Gastrointestinal Surgery, Xiamen Cancer Hospital, The First Affiliated Hospital of Xiamen University, Xiamen, Fujian 361003, People's Republic of China.,Department of Clinical Medicine, Fujian Medical University, Fuzhou, Fujian 350004, People's Republic of China
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39
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Sybirna A, Wong FCK, Surani MA. Genetic basis for primordial germ cells specification in mouse and human: Conserved and divergent roles of PRDM and SOX transcription factors. Curr Top Dev Biol 2019; 135:35-89. [PMID: 31155363 DOI: 10.1016/bs.ctdb.2019.04.004] [Citation(s) in RCA: 25] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/06/2023]
Abstract
Primordial germ cells (PGCs) are embryonic precursors of sperm and egg that pass on genetic and epigenetic information from one generation to the next. In mammals, they are induced from a subset of cells in peri-implantation epiblast by BMP signaling from the surrounding tissues. PGCs then initiate a unique developmental program that involves comprehensive epigenetic resetting and repression of somatic genes. This is orchestrated by a set of signaling molecules and transcription factors that promote germ cell identity. Here we review significant findings on mammalian PGC biology, in particular, the genetic basis for PGC specification in mice and human, which has revealed an evolutionary divergence between the two species. We discuss the importance and potential basis for these differences and focus on several examples to illustrate the conserved and divergent roles of critical transcription factors in mouse and human germline.
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Affiliation(s)
- Anastasiya Sybirna
- Wellcome Trust Cancer Research UK Gurdon Institute, University of Cambridge, Cambridge, United Kingdom; Physiology, Development and Neuroscience Department, University of Cambridge, Cambridge, United Kingdom; Wellcome Trust/Medical Research Council Cambridge Stem Cell Institute, University of Cambridge, Cambridge, United Kingdom.
| | - Frederick C K Wong
- Wellcome Trust Cancer Research UK Gurdon Institute, University of Cambridge, Cambridge, United Kingdom; Physiology, Development and Neuroscience Department, University of Cambridge, Cambridge, United Kingdom
| | - M Azim Surani
- Wellcome Trust Cancer Research UK Gurdon Institute, University of Cambridge, Cambridge, United Kingdom; Physiology, Development and Neuroscience Department, University of Cambridge, Cambridge, United Kingdom; Wellcome Trust/Medical Research Council Cambridge Stem Cell Institute, University of Cambridge, Cambridge, United Kingdom.
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Gisler S, Gonçalves JP, Akhtar W, de Jong J, Pindyurin AV, Wessels LFA, van Lohuizen M. Multiplexed Cas9 targeting reveals genomic location effects and gRNA-based staggered breaks influencing mutation efficiency. Nat Commun 2019; 10:1598. [PMID: 30962441 PMCID: PMC6453899 DOI: 10.1038/s41467-019-09551-w] [Citation(s) in RCA: 43] [Impact Index Per Article: 7.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2018] [Accepted: 03/14/2019] [Indexed: 12/16/2022] Open
Abstract
Understanding the impact of guide RNA (gRNA) and genomic locus on CRISPR-Cas9 activity is crucial to design effective gene editing assays. However, it is challenging to profile Cas9 activity in the endogenous cellular environment. Here we leverage our TRIP technology to integrate ~ 1k barcoded reporter genes in the genomes of mouse embryonic stem cells. We target the integrated reporters (IRs) using RNA-guided Cas9 and characterize induced mutations by sequencing. We report that gRNA-sequence and IR locus explain most variation in mutation efficiency. Predominant insertions of a gRNA-specific nucleotide are consistent with template-dependent repair of staggered DNA ends with 1-bp 5' overhangs. We confirm that such staggered ends are induced by Cas9 in mouse pre-B cells. To explain observed insertions, we propose a model generating primarily blunt and occasionally staggered DNA ends. Mutation patterns indicate that gRNA-sequence controls the fraction of staggered ends, which could be used to optimize Cas9-based insertion efficiency.
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Affiliation(s)
- Santiago Gisler
- Division of Molecular Genetics, Oncode and The Netherlands Cancer Institute, Plesmanlaan 121, Amsterdam, 1066 CX, The Netherlands
| | - Joana P Gonçalves
- Department of Intelligent Systems, Delft University of Technology, Van Mourik Broekmanweg 6, Delft, 2628 XE, The Netherlands
- Division of Molecular Carcinogenesis, Oncode and The Netherlands Cancer Institute, Plesmanlaan 121, Amsterdam, 1066 CX, The Netherlands
| | - Waseem Akhtar
- Division of Molecular Genetics, Oncode and The Netherlands Cancer Institute, Plesmanlaan 121, Amsterdam, 1066 CX, The Netherlands
| | - Johann de Jong
- Division of Molecular Carcinogenesis, Oncode and The Netherlands Cancer Institute, Plesmanlaan 121, Amsterdam, 1066 CX, The Netherlands
- Data & Translational Sciences Group, UCB Biosciences GmbH, Alfred-Nobel-Straße 10, Monheim am Rhein, 40789, Germany
| | - Alexey V Pindyurin
- Institute of Molecular and Cellular Biology, Siberian Branch of Russian Academy of Sciences, Acad. Lavrentiev Ave. 8, Novosibirsk, 630090, Russia
- Division of Gene Regulation, Oncode and The Netherlands Cancer Institute, Plesmanlaan 121, Amsterdam, 1066 CX, The Netherlands
| | - Lodewyk F A Wessels
- Department of Intelligent Systems, Delft University of Technology, Van Mourik Broekmanweg 6, Delft, 2628 XE, The Netherlands.
- Division of Molecular Carcinogenesis, Oncode and The Netherlands Cancer Institute, Plesmanlaan 121, Amsterdam, 1066 CX, The Netherlands.
| | - Maarten van Lohuizen
- Division of Molecular Genetics, Oncode and The Netherlands Cancer Institute, Plesmanlaan 121, Amsterdam, 1066 CX, The Netherlands.
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Cho J, Teshigawara R, Kameda M, Yamaguchi S, Tada T. Nucleus-localized adiponectin is survival gatekeeper through miR-214-mediated AIFM2 regulation. Genes Cells 2019; 24:126-138. [PMID: 30474186 DOI: 10.1111/gtc.12658] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/22/2018] [Revised: 11/19/2018] [Accepted: 11/19/2018] [Indexed: 01/23/2023]
Abstract
Adiponectin secreted from adipocytes into plasma has anti-aging, anti-obesity and anti-inflammation effects. Here, we detected intracellular adiponectin localized in the nuclei of human and mouse pluripotent stem cells, mouse germ cells and some somatic cells. Nucleus-localized (Nu) adiponectin protein is characterized by an N-terminal truncated monomer form in a native state, compared with intact multimer forms of cytoplasm-localized (Cy) adiponectin protein. Doxycycline-induced over-expression of ADIPONECTIN caused cell death in human and mouse Nu-type pluripotent stem cells. Genome-wide gene expression analysis indicated that apoptosis by ADIPONECTIN over-expression was induced in accompany with upregulation of AIFM2 and MEG3. Upregulation of AIFM2 and MEG3 and down-regulation of miR-214-3p verified by qPCR analyses after ADIPONECTIN over-expression indicated that the MEG3/miR-214/AIFM2 pathway played a role in the apoptotic cell death of pluripotent cells. Adiponectin-induced cell death was rescued by the treatment with miR-214-3p mimic. Global data analysis shows that Nu adiponectin has a role in microRNA-mediated post-transcription regulation, cell-cell interactions and chromatin remodeling as a survival gatekeeper.
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Affiliation(s)
- Junkwon Cho
- Laboratory of Developmental Epigenome, Department of Regeneration Science and Engineering, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
| | - Rika Teshigawara
- Laboratory of Developmental Epigenome, Department of Regeneration Science and Engineering, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
| | - Masahiro Kameda
- Laboratory of Developmental Epigenome, Department of Regeneration Science and Engineering, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
| | - Shinpei Yamaguchi
- Laboratory of Stem Cell Pathology, Graduate School of Frontier Biosciences, Osaka University, Suita, Japan
| | - Takashi Tada
- Laboratory of Developmental Epigenome, Department of Regeneration Science and Engineering, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
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Yu CY, Chuang CY, Kuo HC. Trans-spliced long non-coding RNA: an emerging regulator of pluripotency. Cell Mol Life Sci 2018; 75:3339-3351. [PMID: 29961157 PMCID: PMC11105688 DOI: 10.1007/s00018-018-2862-4] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/07/2017] [Revised: 05/21/2018] [Accepted: 06/25/2018] [Indexed: 01/08/2023]
Abstract
With dual capacities for unlimited self-renewal and pluripotent differentiation, pluripotent stem cells (PSCs) give rise to many cell types in our body and PSC culture systems provide an unparalleled opportunity to study early human development and disease. Accumulating evidence indicates that the molecular mechanisms underlying pluripotency maintenance in PSCs involve many factors. Among these regulators, recent studies have shown that long non-coding RNAs (lncRNAs) can affect the pluripotency circuitry by cooperating with master pluripotency-associated factors. Additionally, trans-spliced RNAs, which are generated by combining two or more pre-mRNA transcripts to produce a chimeric RNA, have been identified as regulators of various biological processes, including human pluripotency. In this review, we summarize and discuss current knowledge about the roles of lncRNAs, including trans-spliced lncRNAs, in controlling pluripotency.
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Affiliation(s)
- Chun-Ying Yu
- Institute of Cellular and Organismic Biology, Academia Sinica, No. 128, Sec. 2, Academia Road, Nankang, Taipei, 11529, Taiwan
| | - Ching-Yu Chuang
- Genomics Research Center, Academia Sinica, Taipei, 11529, Taiwan
| | - Hung-Chih Kuo
- Institute of Cellular and Organismic Biology, Academia Sinica, No. 128, Sec. 2, Academia Road, Nankang, Taipei, 11529, Taiwan.
- Genomics Research Center, Academia Sinica, Taipei, 11529, Taiwan.
- College of Medicine, Graduate Institute of Medical Genomics and Proteomics, National Taiwan University, Taipei, Taiwan.
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Kang S, Yun J, Kim DY, Jung SY, Kim YJ, Park JH, Ji ST, Jang WB, Ha J, Kim JH, Baek SH, Kwon SM. Adequate concentration of B cell leukemia/lymphoma 3 (Bcl3) is required for pluripotency and self-renewal of mouse embryonic stem cells via downregulation of Nanog transcription. BMB Rep 2018; 51:92-97. [PMID: 29335071 PMCID: PMC5836563 DOI: 10.5483/bmbrep.2018.51.2.219] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/20/2017] [Indexed: 01/06/2023] Open
Abstract
B cell leukemia/lymphoma 3 (Bcl3) plays a pivotal role in immune homeostasis, cellular proliferation, and cell survival, as a co-activator or co-repressor of transcription of the NF-κB family. Recently, it was reported that Bcl3 positively regulates pluripotency genes, including Oct4, in mouse embryonic stem cells (mESCs). However, the role of Bcl3 in the maintenance of pluripotency and self-renewal activity is not fully established. Here, we report the dynamic regulation of the proliferation, pluripotency, and self-renewal of mESCs by Bcl3 via an influence on Nanog transcriptional activity. Bcl3 expression is predominantly observed in immature mESCs, but significantly decreased during cell differentiation by LIF depletion and in mESC-derived EBs. Importantly, the knockdown of Bcl3 resulted in the loss of self-renewal ability and decreased cell proliferation. Similarly, the ectopic expression of Bcl3 also resulted in a significant reduction of proliferation, and the self-renewal of mESCs was demonstrated by alkaline phosphatase staining and clonogenic single cell-derived colony assay. We further examined that Bcl3-mediated regulation of Nanog transcriptional activity in mESCs, which indicated that Bcl3 acts as a transcriptional repressor of Nanog expression in mESCs. In conclusion, we demonstrated that a sufficient concentration of Bcl3 in mESCs plays a critical role in the maintenance of pluripotency and the self-renewal of mESCs via the regulation of Nanog transcriptional activity.
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Affiliation(s)
- Songhwa Kang
- Laboratory for Vascular Medicine and Stem Cell Biology, Medical Research Institute, Department of Physiology, School of Medicine, Pusan National University, Yangsan 50612, Korea
| | - Jisoo Yun
- Laboratory for Vascular Medicine and Stem Cell Biology, Medical Research Institute, Department of Physiology, School of Medicine, Pusan National University, Yangsan 50612, Korea
| | - Da Yeon Kim
- Laboratory for Vascular Medicine and Stem Cell Biology, Medical Research Institute, Department of Physiology, School of Medicine, Pusan National University, Yangsan 50612, Korea
| | - Seok Yun Jung
- Laboratory for Vascular Medicine and Stem Cell Biology, Medical Research Institute, Department of Physiology, School of Medicine, Pusan National University, Yangsan 50612, Korea
| | - Yeon Ju Kim
- Laboratory for Vascular Medicine and Stem Cell Biology, Medical Research Institute, Department of Physiology, School of Medicine, Pusan National University, Yangsan 50612, Korea
| | - Ji Hye Park
- Laboratory for Vascular Medicine and Stem Cell Biology, Medical Research Institute, Department of Physiology, School of Medicine, Pusan National University, Yangsan 50612, Korea
| | - Seung Taek Ji
- Laboratory for Vascular Medicine and Stem Cell Biology, Medical Research Institute, Department of Physiology, School of Medicine, Pusan National University, Yangsan 50612, Korea
| | - Woong Bi Jang
- Laboratory for Vascular Medicine and Stem Cell Biology, Medical Research Institute, Department of Physiology, School of Medicine, Pusan National University, Yangsan 50612, Korea
| | - Jongseong Ha
- Laboratory for Vascular Medicine and Stem Cell Biology, Medical Research Institute, Department of Physiology, School of Medicine, Pusan National University, Yangsan 50612, Korea
| | - Jae Ho Kim
- Research Institute of Convergence Biomedical Science and Technology, Pusan National University Yangsan Hospital, Yangsan 50612, Korea
| | - Sang Hong Baek
- Laboratory of Cardiovascular Disease, Division of Cardiology, School of Medicine, The Catholic University, Seoul 06591, Korea
| | - Sang-Mo Kwon
- Laboratory for Vascular Medicine and Stem Cell Biology, Medical Research Institute, Department of Physiology, School of Medicine, Pusan National University, Yangsan 50612; Research Institute of Convergence Biomedical Science and Technology, Pusan National University Yangsan Hospital, Yangsan 50612, Korea
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Ali MS, Gill KS, Saglio G, Cilloni D, Soden DM, Forde PF. Expressional changes in stemness markers post electrochemotherapy in pancreatic cancer cells. Bioelectrochemistry 2018; 122:84-92. [DOI: 10.1016/j.bioelechem.2018.03.009] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/04/2017] [Revised: 03/14/2018] [Accepted: 03/14/2018] [Indexed: 12/12/2022]
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Ermakov A, Daks A, Fedorova O, Shuvalov O, Barlev NA. Ca 2+ -depended signaling pathways regulate self-renewal and pluripotency of stem cells. Cell Biol Int 2018; 42:1086-1096. [PMID: 29851182 DOI: 10.1002/cbin.10998] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/03/2017] [Accepted: 05/25/2018] [Indexed: 12/15/2022]
Abstract
Ca2+ -mediated signaling is widely spread in nature and plays critical role in the individual development of various organisms ranging from microorganisms to mammals. In vertebrates, Ca2+ is involved in important developmental events: fertilization, body plan establishment, and organogenesis. The two later events are defined by embryonic stem cells (ESCs). ESCs are capable of self-renewal and are pluripotent by nature, that is, can give rise to all types of cells that make up the body. Given the paramount importance of Ca2+ signalization in the development, it is therefore not surprising this process also plays role in the biology of stem cells. In this review, we scrutinize the published experimental data on the role of Ca2+ ions in embryonic stem cells self-renewal and pluripotency. In line with this, we also discuss possible mechanisms of p53 inhibition as a major hindrance to self-renewal of ESCs. Finally, we argue about the role of G-protein-coupled receptors (GPCRs), the largest family of heteromeric transmembrane receptors, and GPCR-mediated signalization in stem cells, and propose the role for the GPCR-G-protein-PLC-Ca2+ -downstream signaling pathway in the regulation of pluripotency of both mouse and human ESCs.
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Affiliation(s)
| | - Alexandra Daks
- Institute of Cytology RAS, Saint-Petersburg 194064, Russia
| | - Olga Fedorova
- Institute of Cytology RAS, Saint-Petersburg 194064, Russia
| | - Oleg Shuvalov
- Institute of Cytology RAS, Saint-Petersburg 194064, Russia
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Gas41 links histone acetylation to H2A.Z deposition and maintenance of embryonic stem cell identity. Cell Discov 2018; 4:28. [PMID: 29900004 PMCID: PMC5995911 DOI: 10.1038/s41421-018-0027-0] [Citation(s) in RCA: 47] [Impact Index Per Article: 6.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/19/2018] [Revised: 03/22/2018] [Accepted: 03/23/2018] [Indexed: 01/07/2023] Open
Abstract
The histone variant H2A.Z is essential for maintaining embryonic stem cell (ESC) identity in part by keeping developmental genes in a poised bivalent state. However, how H2A.Z is deposited into the bivalent domains remains unknown. In mammals, two chromatin remodeling complexes, Tip60/p400 and SRCAP, exchange the canonical histone H2A for H2A.Z in the chromatin. Here we show that Glioma Amplified Sequence 41 (Gas41), a shared subunit of the two H2A.Z-depositing complexes, functions as a reader of histone lysine acetylation and recruits Tip60/p400 and SRCAP to deposit H2A.Z into specific chromatin regions including bivalent domains. The YEATS domain of Gas41 bound to acetylated histone H3K27 and H3K14 both in vitro and in cells. The crystal structure of the Gas41 YEATS domain in complex with the H3K27ac peptide revealed that, similar to the AF9 and ENL YEATS domains, Gas41 YEATS forms a serine-lined aromatic cage for acetyllysine recognition. Consistently, mutations in the aromatic residues of the Gas41 YEATS domain abrogated the interaction. In mouse ESCs, knockdown of Gas41 led to flattened morphology of ESC colonies, as the result of derepression of differentiation genes. Importantly, the abnormal morphology was rescued by expressing wild-type Gas41, but not the YEATS domain mutated counterpart that does not recognize histone acetylation. Mechanically, we found that Gas41 depletion led to reduction of H2A.Z levels and a concomitant reduction of H3K27me3 levels on bivalent domains. Together, our study reveals an essential role of the Gas41 YEATS domain in linking histone acetylation to H2A.Z deposition and maintenance of ESC identity.
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Patra SK, Vemulawada C, Soren MM, Sundaray JK, Panda MK, Barman HK. Molecular characterization and expression patterns of Nanog gene validating its involvement in the embryonic development and maintenance of spermatogonial stem cells of farmed carp, Labeo rohita. J Anim Sci Biotechnol 2018; 9:45. [PMID: 29992021 PMCID: PMC5994655 DOI: 10.1186/s40104-018-0260-2] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/18/2017] [Accepted: 04/25/2018] [Indexed: 12/24/2022] Open
Abstract
Background The homeobox containing transcription factor Nanog plays crucial roles in embryonic development/proliferation and/or maintenance of spermatogonial stem cells (SSCs) via interacting with transcription factors such as Oct4 and Sox2 in mammals. However, knowledge of its exact mechanistic pathways remains unexploited. Very little is known about teleost Nanog. Information on the Nanog gene of farmed rohu carp (Labeo rohita) is lacking. We cloned and characterized the Nanog gene of rohu carp to understand the expression pattern in early developmental stages and also deduced the genomic organization including promoter elements. Results Rohu Nanog (LrNanog) cDNA comprised an open reading frame of 1,161 nucleotides bearing a structural homeodomain; whereas, the genomic structure contained four exons and three introns suggesting that it is homologous to mammalian counterparts. Phylogenetically, it was closely related to freshwater counterparts. Protein sequence (386 AA of 42.65 kDa) comparison revealed its low similarity with other vertebrate counterparts except that of the conserved homeodomain. Tissue distribution analysis revealed the existence of LrNanog transcripts only in adult gonads. The heightened abundances in the ovary and proliferating spermatogonia suggested its participations in maternal inheritance and male germ cell development. The potentiating abundances from fertilized egg onwards peaking at blastula stage vis- à-vis decreasing levels from gastrula stage onwards demonstrated its role in embryonic stem cell development. We also provided evidence of its presence in SSCs by western blotting analysis. Further, the promoter region was characterized, predicting a basal core promoter and other consensus elements. Conclusion The molecular characterization of LrNanog and its documented expression profiling at transcript and protein levels are indicative of its functional linkage with embryonic/spermatogonial stem cell maintenance. This is the first report of LrNanog genomic organization including its promoter sequence information with predicted regulatory elements of a large-bodied carp species. This will be useful for elucidating its mechanism expression in future. Nanog could be used as a potential biomarker for proliferating carp SSCs.
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Affiliation(s)
- Swagat K Patra
- 1Fish Genetics and Biotechnology Division, ICAR - Central Institute of Freshwater Aquaculture, Kausalyaganga, Bhubaneswar, Odisha 751002 India
| | - Chakrpani Vemulawada
- 1Fish Genetics and Biotechnology Division, ICAR - Central Institute of Freshwater Aquaculture, Kausalyaganga, Bhubaneswar, Odisha 751002 India
| | - Meenati M Soren
- 1Fish Genetics and Biotechnology Division, ICAR - Central Institute of Freshwater Aquaculture, Kausalyaganga, Bhubaneswar, Odisha 751002 India
| | - Jitendra K Sundaray
- 1Fish Genetics and Biotechnology Division, ICAR - Central Institute of Freshwater Aquaculture, Kausalyaganga, Bhubaneswar, Odisha 751002 India
| | - Manoj K Panda
- 2Center of Biotechnology, Siksha 'O' Anusandhan University, Bhubaneswar, India
| | - Hirak K Barman
- 1Fish Genetics and Biotechnology Division, ICAR - Central Institute of Freshwater Aquaculture, Kausalyaganga, Bhubaneswar, Odisha 751002 India
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Omole AE, Fakoya AOJ. Ten years of progress and promise of induced pluripotent stem cells: historical origins, characteristics, mechanisms, limitations, and potential applications. PeerJ 2018; 6:e4370. [PMID: 29770269 PMCID: PMC5951134 DOI: 10.7717/peerj.4370] [Citation(s) in RCA: 118] [Impact Index Per Article: 16.9] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2017] [Accepted: 01/24/2018] [Indexed: 12/11/2022] Open
Abstract
The discovery of induced pluripotent stem cells (iPSCs) by Shinya Yamanaka in 2006 was heralded as a major breakthrough of the decade in stem cell research. The ability to reprogram human somatic cells to a pluripotent embryonic stem cell-like state through the ectopic expression of a combination of embryonic transcription factors was greeted with great excitement by scientists and bioethicists. The reprogramming technology offers the opportunity to generate patient-specific stem cells for modeling human diseases, drug development and screening, and individualized regenerative cell therapy. However, fundamental questions have been raised regarding the molecular mechanism of iPSCs generation, a process still poorly understood by scientists. The efficiency of reprogramming of iPSCs remains low due to the effect of various barriers to reprogramming. There is also the risk of chromosomal instability and oncogenic transformation associated with the use of viral vectors, such as retrovirus and lentivirus, which deliver the reprogramming transcription factors by integration in the host cell genome. These challenges can hinder the therapeutic prospects and promise of iPSCs and their clinical applications. Consequently, extensive studies have been done to elucidate the molecular mechanism of reprogramming and novel strategies have been identified which help to improve the efficiency of reprogramming methods and overcome the safety concerns linked with iPSC generation. Distinct barriers and enhancers of reprogramming have been elucidated, and non-integrating reprogramming methods have been reported. Here, we summarize the progress and the recent advances that have been made over the last 10 years in the iPSC field, with emphasis on the molecular mechanism of reprogramming, strategies to improve the efficiency of reprogramming, characteristics and limitations of iPSCs, and the progress made in the applications of iPSCs in the field of disease modelling, drug discovery and regenerative medicine. Additionally, this study appraises the role of genomic editing technology in the generation of healthy iPSCs.
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Affiliation(s)
- Adekunle Ebenezer Omole
- Department of Basic Sciences, American University of Antigua College of Medicine, St. John's, Antigua
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Wu G, Xu R, Zhang P, Xiao T, Fu Y, Zhang Y, Du Y, Ye J, Cheng J, Jiang H. Estrogen regulates stemness and senescence of bone marrow stromal cells to prevent osteoporosis via ERβ-SATB2 pathway. J Cell Physiol 2018; 233:4194-4204. [PMID: 29030963 DOI: 10.1002/jcp.26233] [Citation(s) in RCA: 53] [Impact Index Per Article: 7.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2017] [Accepted: 09/29/2017] [Indexed: 12/22/2022]
Abstract
Decline of pluripotency in bone marrow stromal cells (BMSCs) associated with estrogen deficiency leads to a bone formation defect in osteoporosis. Special AT-rich sequence binding protein 2 (SATB2) is crucial for maintaining stemness and osteogenic differentiation of BMSCs. However, whether SATB2 is involved in estrogen-deficiency associated-osteoporosis is largely unknown. In this study, we found that estrogen mediated pluripotency and senescence of BMSCs, primarily through estrogen receptor beta (ERβ). BMSCs from the OVX rats displayed increased senescence and weaker SATB2 expression, stemness, and osteogenic differentiation, while estrogen could rescue these phenotypes. Inhibition of ERβ or ERα confirmed that SATB2 was associated with ERβ in estrogen-mediated pluripotency and senescence of BMSCs. Furthermore, estrogen mediated the upregulation of SATB2 through the induction of ERβ binding to estrogen response elements (ERE) located at -488 of the SATB2 gene. SATB2 overexpression alleviated senescence and enhanced stemness and osteogenic differentiation of OVX-BMSCs. SATB2-modified BMSCs transplantation could prevent trabecular bone loss in an ovariectomized rat model. Collectively, our study revealed the role of SATB2 in stemness, senescence, and osteogenesis of OVX-BMSCs. These results indicate that estrogen prevents osteoporosis by promoting stemness and osteogenesis, and inhibiting senescence of BMSCs through an ERβ-SATB2 pathway. Therefore, SATB2 is a novel anti-osteoporosis target gene.
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Affiliation(s)
- Geng Wu
- Department of Stomatology, The First People's Hospital of Lianyungang City, Lianyungang, China
| | - Rongyao Xu
- Jiangsu Key Laboratory of Oral Diseases, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Ping Zhang
- Jiangsu Key Laboratory of Oral Diseases, Nanjing Medical University, Nanjing, Jiangsu, China
- Department of Oral and Maxillofacial Surgery, Affiliated Hospital of Stomatology, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Tao Xiao
- Jiangsu Key Laboratory of Oral Diseases, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Yu Fu
- Jiangsu Key Laboratory of Oral Diseases, Nanjing Medical University, Nanjing, Jiangsu, China
- Department of Oral and Maxillofacial Surgery, Affiliated Hospital of Stomatology, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Yuchao Zhang
- Jiangsu Key Laboratory of Oral Diseases, Nanjing Medical University, Nanjing, Jiangsu, China
- Department of Oral and Maxillofacial Surgery, Affiliated Hospital of Stomatology, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Yifei Du
- Jiangsu Key Laboratory of Oral Diseases, Nanjing Medical University, Nanjing, Jiangsu, China
- Department of Oral and Maxillofacial Surgery, Affiliated Hospital of Stomatology, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Jinhai Ye
- Jiangsu Key Laboratory of Oral Diseases, Nanjing Medical University, Nanjing, Jiangsu, China
- Department of Oral and Maxillofacial Surgery, Affiliated Hospital of Stomatology, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Jie Cheng
- Jiangsu Key Laboratory of Oral Diseases, Nanjing Medical University, Nanjing, Jiangsu, China
- Department of Oral and Maxillofacial Surgery, Affiliated Hospital of Stomatology, Nanjing Medical University, Nanjing, Jiangsu, China
| | - Hongbing Jiang
- Jiangsu Key Laboratory of Oral Diseases, Nanjing Medical University, Nanjing, Jiangsu, China
- Department of Oral and Maxillofacial Surgery, Affiliated Hospital of Stomatology, Nanjing Medical University, Nanjing, Jiangsu, China
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Malik V, Zimmer D, Jauch R. Diversity among POU transcription factors in chromatin recognition and cell fate reprogramming. Cell Mol Life Sci 2018; 75:1587-1612. [PMID: 29335749 PMCID: PMC11105716 DOI: 10.1007/s00018-018-2748-5] [Citation(s) in RCA: 55] [Impact Index Per Article: 7.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/04/2017] [Revised: 12/23/2017] [Accepted: 01/08/2018] [Indexed: 12/28/2022]
Abstract
The POU (Pit-Oct-Unc) protein family is an evolutionary ancient group of transcription factors (TFs) that bind specific DNA sequences to direct gene expression programs. The fundamental importance of POU TFs to orchestrate embryonic development and to direct cellular fate decisions is well established, but the molecular basis for this activity is insufficiently understood. POU TFs possess a bipartite 'two-in-one' DNA binding domain consisting of two independently folding structural units connected by a poorly conserved and flexible linker. Therefore, they represent a paradigmatic example to study the molecular basis for the functional versatility of TFs. Their modular architecture endows POU TFs with the capacity to accommodate alternative composite DNA sequences by adopting different quaternary structures. Moreover, associations with partner proteins crucially influence the selection of their DNA binding sites. The plentitude of DNA binding modes confers the ability to POU TFs to regulate distinct genes in the context of different cellular environments. Likewise, different binding modes of POU proteins to DNA could trigger alternative regulatory responses in the context of different genomic locations of the same cell. Prominent POU TFs such as Oct4, Brn2, Oct6 and Brn4 are not only essential regulators of development but have also been successfully employed to reprogram somatic cells to pluripotency and neural lineages. Here we review biochemical, structural, genomic and cellular reprogramming studies to examine how the ability of POU TFs to select regulatory DNA, alone or with partner factors, is tied to their capacity to epigenetically remodel chromatin and drive specific regulatory programs that give cells their identities.
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Affiliation(s)
- Vikas Malik
- CAS Key Laboratory of Regenerative Biology, Joint School of Life Sciences, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou Medical University, Guangzhou, 511436, China
- Genome Regulation Laboratory, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China
| | - Dennis Zimmer
- CAS Key Laboratory of Regenerative Biology, Joint School of Life Sciences, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou Medical University, Guangzhou, 511436, China
- Genome Regulation Laboratory, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China
| | - Ralf Jauch
- CAS Key Laboratory of Regenerative Biology, Joint School of Life Sciences, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou Medical University, Guangzhou, 511436, China.
- Genome Regulation Laboratory, Guangdong Provincial Key Laboratory of Stem Cell and Regenerative Medicine, South China Institute for Stem Cell Biology and Regenerative Medicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, 510530, China.
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