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Jiang YZ, Hu LY, Chen MS, Wang XJ, Tan CN, Xue PP, Yu T, He XY, Xiang LX, Xiao YN, Li XL, Ran Q, Li ZJ, Chen L. GATA binding protein 2 mediated ankyrin repeat domain containing 26 high expression in myeloid-derived cell lines. World J Stem Cells 2024; 16:538-550. [PMID: 38817334 PMCID: PMC11135246 DOI: 10.4252/wjsc.v16.i5.538] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/26/2023] [Revised: 03/12/2024] [Accepted: 04/12/2024] [Indexed: 05/24/2024] Open
Abstract
BACKGROUND Thrombocytopenia 2, an autosomal dominant inherited disease characterized by moderate thrombocytopenia, predisposition to myeloid malignancies and normal platelet size and function, can be caused by 5'-untranslated region (UTR) point mutations in ankyrin repeat domain containing 26 (ANKRD26). Runt related transcription factor 1 (RUNX1) and friend leukemia integration 1 (FLI1) have been identified as negative regulators of ANKRD26. However, the positive regulators of ANKRD26 are still unknown. AIM To prove the positive regulatory effect of GATA binding protein 2 (GATA2) on ANKRD26 transcription. METHODS Human induced pluripotent stem cells derived from bone marrow (hiPSC-BM) and urothelium (hiPSC-U) were used to examine the ANKRD26 expression pattern in the early stage of differentiation. Then, transcriptome sequencing of these iPSCs and three public transcription factor (TF) databases (Cistrome DB, animal TFDB and ENCODE) were used to identify potential TF candidates for ANKRD26. Furthermore, overexpression and dual-luciferase reporter experiments were used to verify the regulatory effect of the candidate TFs on ANKRD26. Moreover, using the GENT2 platform, we analyzed the relationship between ANKRD26 expression and overall survival in cancer patients. RESULTS In hiPSC-BMs and hiPSC-Us, we found that the transcription levels of ANKRD26 varied in the absence of RUNX1 and FLI1. We sequenced hiPSC-BM and hiPSC-U and identified 68 candidate TFs for ANKRD26. Together with three public TF databases, we found that GATA2 was the only candidate gene that could positively regulate ANKRD26. Using dual-luciferase reporter experiments, we showed that GATA2 directly binds to the 5'-UTR of ANKRD26 and promotes its transcription. There are two identified binding sites of GATA2 that are located 2 kb upstream of the TSS of ANKRD26. In addition, we discovered that high ANKRD26 expression is always related to a more favorable prognosis in breast and lung cancer patients. CONCLUSION We first discovered that the transcription factor GATA2 plays a positive role in ANKRD26 transcription and identified its precise binding sites at the promoter region, and we revealed the importance of ANKRD26 in many tissue-derived cancers.
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Affiliation(s)
- Yang-Zhou Jiang
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Lan-Yue Hu
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Mao-Shan Chen
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Xiao-Jie Wang
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Cheng-Ning Tan
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Pei-Pei Xue
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Teng Yu
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Xiao-Yan He
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Li-Xin Xiang
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Yan-Ni Xiao
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Xiao-Liang Li
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Qian Ran
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Zhong-Jun Li
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China
| | - Li Chen
- Laboratory of Radiation Biology, Department of Blood Transfusion, Laboratory Medicine Center, The Second Affiliated Hospital, Third Military Medical University, Chongqing 400037, China
- Hematopoietic Acute Radiation Syndrome Medical and Pharmaceutical Basic Research Innovation Center, Ministry of Education of the People's Republic of China, Chongqing 400037, China.
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Pang C, Wu X, Nikuze L, Wei H. Analysis of clinical characteristics and treatment efficacy in two pediatric cases of ANKRD26-related thrombocytopenia. Platelets 2023; 34:2262607. [PMID: 37852929 DOI: 10.1080/09537104.2023.2262607] [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: 08/11/2022] [Accepted: 09/19/2023] [Indexed: 10/20/2023]
Abstract
ANKRD26-related thrombocytopenia (ANKRD26-RT or THC2, MIM 188 000), an autosomal dominant thrombocytopenia, is unresponsive to immunosuppressive therapy and susceptible to hematological malignancies. A large number of pediatric patients are diagnosed with immune thrombocytopenia (ITP) every year; however, thrombocytopenia of genetic origin is often missed. Extensive characterization of ANKRD26-RT will help prevent missed diagnosis and misdiagnosis. Furthermore, identification of ANKRD26-RT will help in the formulation of an accurate diagnosis and a treatment plan. In our study, we report cases of two Chinese pediatric patients with ANKRD26-RT and analyze their clinical characteristics, gene mutations, and treatment modalities. Both patients were 1-year-old and presented with mild bleeding (World Health Organization(WHO) score grade 1), different degrees of platelet reduction, normal mean platelet volume, and megakaryocyte maturation impairment not obvious. Genetic tests revealed that both patients had ANKRD26 gene mutations.Patient 1 had a mutation c.-140C>G of the 5' untranslated region (UTR), and patient 2 had a mutation of c.-127A>T of 5'UTR. Both patients were treated with eltrombopag, and the treatment was no response, with no adverse reactions.
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Affiliation(s)
- Congfei Pang
- Department of Pediatrics, The Sixth Affiliated Hospital of Guangxi Medical University: The First People's Hospital of Yulin, Yulin, Guangxi, P.R. China
| | - Xiaomei Wu
- Department of Pediatrics, Red Cross Hospital of Yulin city, Yulin, Guangxi, P.R. China
| | | | - Hongying Wei
- Department of Pediatrics, The Second Affiliated Hospital of Guangxi Medical University, Nanning, Guangxi, P.R. China
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Kast DJ, Jansen S. Purification of modified mammalian actin isoforms for in vitro reconstitution assays. Eur J Cell Biol 2023; 102:151363. [PMID: 37778219 PMCID: PMC10872616 DOI: 10.1016/j.ejcb.2023.151363] [Citation(s) in RCA: 5] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/19/2023] [Revised: 08/19/2023] [Accepted: 09/26/2023] [Indexed: 10/03/2023] Open
Abstract
In vitro reconstitution assays using purified actin have greatly improved our understanding of cytoskeletal dynamics and their regulation by actin-binding proteins. However, early purification methods consisted of harsh conditions to obtain pure actin and often did not include correct maturation and obligate modification of the isolated actin monomers. Novel insights into the folding requirements and N-terminal processing of actin as well as a better understanding of the interaction of actin with monomer sequestering proteins such as DNaseI, profilin and gelsolin, led to the development of more gentle approaches to obtain pure recombinant actin isoforms with known obligate modifications. This review summarizes the approaches that can be employed to isolate natively folded endogenous and recombinant actin from tissues and cells. We further emphasize the use and limitations of each method and describe how these methods can be implemented to study actin PTMs, disease-related actin mutations and novel actin-like proteins.
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Affiliation(s)
- David J Kast
- Department of Cell Biology and Physiology, Washington University in St. Louis, Saint Louis, MO, 63110, United States.
| | - Silvia Jansen
- Department of Cell Biology and Physiology, Washington University in St. Louis, Saint Louis, MO, 63110, United States.
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Vyas H, Alcheikh A, Lowe G, Stevenson WS, Morgan NV, Rabbolini DJ. Prevalence and natural history of variants in the ANKRD26 gene: a short review and update of reported cases. Platelets 2022; 33:1107-1112. [PMID: 35587581 PMCID: PMC9555274 DOI: 10.1080/09537104.2022.2071853] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022]
Abstract
ANKRD26 is a highly conserved gene located on chromosome 10p12.1 which has shown to play a role in normal megakaryocyte differentiation. ANKRD26-related thrombocytopenia, or thrombocytopenia 2, is an inherited thrombocytopenia with mild bleeding diathesis resulting from point mutations the 5ʹUTR of the ANKRD26 gene. Point mutations in the 5ʹUTR region have been shown to prevent transcription factor-mediated downregulation of ANKRD26 in normal megakaryocyte differentiation. Patients with ANKRD26-related thrombocytopenia have a predisposition to developing hematological malignancies, with acute myeloid leukemia and myelodysplastic syndrome most commonly described in the literature. We review the clinical features and biological mechanisms of ANKRD26-related thrombocytopenia and summarize known cases in the literature.
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Affiliation(s)
- Hrushikesh Vyas
- Institute of Cardiovascular Sciences, College of Medical and Dental Sciences, University of Birmingham, Birmingham, UK
| | - Ahmad Alcheikh
- Northern Blood Research Centre, Kolling Institute, University of Sydney, Sydney, Australia
| | - Gillian Lowe
- Comprehensive Care Haemophilia Centre, University Hospital Birmingham NHS Foundation Trust, Birmingham, UK
| | - William S Stevenson
- Northern Blood Research Centre, Kolling Institute, University of Sydney, Sydney, Australia.,Department of Haematology and Transfusion Medicine, Royal North Shore Hospital, Sydney, Australia
| | - Neil V Morgan
- Institute of Cardiovascular Sciences, College of Medical and Dental Sciences, University of Birmingham, Birmingham, UK
| | - David J Rabbolini
- Northern Blood Research Centre, Kolling Institute, University of Sydney, Sydney, Australia
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Kasahara Y, Osuka S, Takasaki N, Bayasula, Koya Y, Nakanishi N, Murase T, Nakamura T, Goto M, Iwase A, Kajiyama H. Primate-specific POTE-actin gene could play a role in human folliculogenesis by controlling the proliferation of granulosa cells. Cell Death Discov 2021; 7:186. [PMID: 34285194 PMCID: PMC8292509 DOI: 10.1038/s41420-021-00566-1] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/23/2021] [Revised: 06/06/2021] [Accepted: 06/25/2021] [Indexed: 11/16/2022] Open
Abstract
Patients with primary ovarian insufficiency (POI) often have a high prevalence of autoimmune disorders. To identify antigenic molecules associated with ovarian autoimmunity, we performed immunoprecipitation (IP) screening using serum from patients with POI and the established human granulosa cell line (HGrC1). POTE ankyrin domain family member E (POTEE) and POTE ankyrin domain family member F (POTEF), proteins specific to primates, were identified as candidate antigens. Using immunohistochemistry (IHC) with human ovarian tissue, POTEE or POTEF was weakly seen in the granulosa cells (GCs) of primordial follicles and primary follicles, and strongly in large antral follicles and luteal cells. Interestingly, no signals were detected in growing GCs in secondary, preantral, and small antral follicles. Thus, to explore the function of POTEE and POTEF in human folliculogenesis, we established HGrC1 cell lines with drug-inducible expression of POTEF. Expression of POTEF significantly suppressed cell proliferation in HGrC1 cells. Furthermore, chaperonin containing TCP-1 complex (CCT) components, which affect folding proteins required for cell proliferation, was bound to the actin domain of POTEF protein. Although CCT is normally localized only around the Golgi apparatus, TCP-1α, a component of CCT, co-migrated closer to the cell membrane when POTEF expression was induced. These data suggest that the interaction between POTEF and CCT components impairs the usual function of CCT during cell growth. In addition, over-accumulation of POTEF in HGrC1 cells leads to autophagic failure. It was recently reported that knockout of an autophagic gene in mice leads to a phenotype similar to human POI. These results suggested that a proper amount of POTEF is required for the maintenance of GCs in follicle pools, whereas POTEF overaccumulation might be involved in follicle atresia and the development of POI. We also showed the possibility that POTEF could be an antigen involved in ovarian autoimmunity.
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Affiliation(s)
- Yukiyo Kasahara
- Department of Obstetrics and Gynecology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya, 466-8550, Japan
| | - Satoko Osuka
- Department of Obstetrics and Gynecology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya, 466-8550, Japan. .,Department of Maternal and Perinatal Medicine, Nagoya University Hospital, 65 Tsurumai-cho, Showa-ku, Nagoya, 466-8550, Japan.
| | - Nobuyoshi Takasaki
- Bell Research Center for Reproductive Health and Cancer, Nagoya University Graduate School of Medicine, Nagoya, 466-8550, Japan
| | - Bayasula
- Bell Research Center for Reproductive Health and Cancer, Nagoya University Graduate School of Medicine, Nagoya, 466-8550, Japan
| | - Yoshihiro Koya
- Bell Research Center for Reproductive Health and Cancer, Nagoya University Graduate School of Medicine, Nagoya, 466-8550, Japan
| | - Natsuki Nakanishi
- Department of Obstetrics and Gynecology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya, 466-8550, Japan
| | - Tomohiko Murase
- Department of Obstetrics and Gynecology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya, 466-8550, Japan
| | - Tomoko Nakamura
- Department of Obstetrics and Gynecology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya, 466-8550, Japan
| | - Maki Goto
- Department of Obstetrics and Gynecology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya, 466-8550, Japan
| | - Akira Iwase
- Department of Obstetrics and Gynecology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi, 371-8511, Japan
| | - Hiroaki Kajiyama
- Department of Obstetrics and Gynecology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya, 466-8550, Japan
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Novel genetic features of human and mouse Purkinje cell differentiation defined by comparative transcriptomics. Proc Natl Acad Sci U S A 2020; 117:15085-15095. [PMID: 32546527 PMCID: PMC7334519 DOI: 10.1073/pnas.2000102117] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Comparative transcriptomics between differentiating human pluripotent stem cells (hPSCs) and developing mouse neurons offers a powerful approach to compare genetic and epigenetic pathways in human and mouse neurons. To analyze human Purkinje cell (PC) differentiation, we optimized a protocol to generate human pluripotent stem cell-derived Purkinje cells (hPSC-PCs) that formed synapses when cultured with mouse cerebellar glia and granule cells and fired large calcium currents, measured with the genetically encoded calcium indicator jRGECO1a. To directly compare global gene expression of hPSC-PCs with developing mouse PCs, we used translating ribosomal affinity purification (TRAP). As a first step, we used Tg(Pcp2-L10a-Egfp) TRAP mice to profile actively transcribed genes in developing postnatal mouse PCs and used metagene projection to identify the most salient patterns of PC gene expression over time. We then created a transgenic Pcp2-L10a-Egfp TRAP hPSC line to profile gene expression in differentiating hPSC-PCs, finding that the key gene expression pathways of differentiated hPSC-PCs most closely matched those of late juvenile mouse PCs (P21). Comparative bioinformatics identified classical PC gene signatures as well as novel mitochondrial and autophagy gene pathways during the differentiation of both mouse and human PCs. In addition, we identified genes expressed in hPSC-PCs but not mouse PCs and confirmed protein expression of a novel human PC gene, CD40LG, expressed in both hPSC-PCs and native human cerebellar tissue. This study therefore provides a direct comparison of hPSC-PC and mouse PC gene expression and a robust method for generating differentiated hPSC-PCs with human-specific gene expression for modeling developmental and degenerative cerebellar disorders.
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Evolutionary Dynamics of the POTE Gene Family in Human and Nonhuman Primates. Genes (Basel) 2020; 11:genes11020213. [PMID: 32085667 PMCID: PMC7073761 DOI: 10.3390/genes11020213] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/17/2019] [Revised: 02/06/2020] [Accepted: 02/13/2020] [Indexed: 12/20/2022] Open
Abstract
POTE (prostate, ovary, testis, and placenta expressed) genes belong to a primate-specific gene family expressed in prostate, ovary, and testis as well as in several cancers including breast, prostate, and lung cancers. Due to their tumor-specific expression, POTEs are potential oncogenes, therapeutic targets, and biomarkers for these malignancies. This gene family maps within human and primate segmental duplications with a copy number ranging from two to 14 in different species. Due to the high sequence identity among the gene copies, specific efforts are needed to assemble these loci in order to correctly define the organization and evolution of the gene family. Using single-molecule, real-time (SMRT) sequencing, in silico analyses, and molecular cytogenetics, we characterized the structure, copy number, and chromosomal distribution of the POTE genes, as well as their expression in normal and disease tissues, and provided a comparative analysis of the POTE organization and gene structure in primate genomes. We were able, for the first time, to de novo sequence and assemble a POTE tandem duplication in marmoset that is misassembled and collapsed in the reference genome, thus revealing the presence of a second POTE copy. Taken together, our findings provide comprehensive insights into the evolutionary dynamics of the primate-specific POTE gene family, involving gene duplications, deletions, and long interspersed nuclear element (LINE) transpositions to explain the actual repertoire of these genes in human and primate genomes.
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Galera P, Dulau-Florea A, Calvo KR. Inherited thrombocytopenia and platelet disorders with germline predisposition to myeloid neoplasia. Int J Lab Hematol 2019; 41 Suppl 1:131-141. [PMID: 31069978 DOI: 10.1111/ijlh.12999] [Citation(s) in RCA: 41] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/02/2019] [Revised: 02/07/2019] [Accepted: 02/10/2019] [Indexed: 12/21/2022]
Abstract
Advances in molecular genetic sequencing techniques have contributed to the elucidation of previously unknown germline mutations responsible for inherited thrombocytopenia (IT). Regardless of age of presentation and severity of symptoms related to thrombocytopenia and/or platelet dysfunction, a subset of patients with IT are at increased risk of developing myeloid neoplasms during their life time, particularly those with germline autosomal dominant mutations in RUNX1, ANKRD26, and ETV6. Patients may present with isolated thrombocytopenia and megakaryocytic dysmorphia or atypia on baseline bone marrow evaluation, without constituting myelodysplasia (MDS). Bone marrow features may overlap with idiopathic thrombocytopenic purpura (ITP) or sporadic MDS leading to misdiagnosis. Progression to myelodysplastic syndrome/ acute myeloid leukemia (MDS/AML) may be accompanied by progressive bi- or pancytopenia, multilineage dysplasia, increased blasts, cytogenetic abnormalities, acquisition of bi-allelic mutations in the underlying gene with germline mutation, or additional somatic mutations in genes associated with myeloid malignancy. A subset of patients may present with MDS/AML at a young age, underscoring the growing concern for evaluating young patients with MDS/AML for germline mutations predisposing to myeloid neoplasm. Early recognition of germline mutation and predisposition to myeloid malignancy permits appropriate treatment, adequate monitoring for disease progression, proper donor selection for hematopoietic stem cell transplantation, as well as genetic counseling of the affected patients and their family members. Herein, we describe the clinical and diagnostic features of IT with germline mutations predisposing to myeloid neoplasms focusing on mutations involving RUNX1, ANKRD26, and ETV6.
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Affiliation(s)
- Pallavi Galera
- Department of Laboratory Medicine, Hematology Section, Clinical Center, National Institutes of Health (NIH), Bethesda, Maryland
| | - Alina Dulau-Florea
- Department of Laboratory Medicine, Hematology Section, Clinical Center, National Institutes of Health (NIH), Bethesda, Maryland
| | - Katherine R Calvo
- Department of Laboratory Medicine, Hematology Section, Clinical Center, National Institutes of Health (NIH), Bethesda, Maryland
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Sharma A, Albahrani M, Zhang W, Kufel CN, James SR, Odunsi K, Klinkebiel D, Karpf AR. Epigenetic activation of POTE genes in ovarian cancer. Epigenetics 2019; 14:185-197. [PMID: 30764732 DOI: 10.1080/15592294.2019.1581590] [Citation(s) in RCA: 20] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/27/2022] Open
Abstract
The POTE gene family consists of 14 homologous genes localized to autosomal pericentromeres, and a sub-set of POTEs are cancer-testis antigen (CTA) genes. POTEs are over-expressed in epithelial ovarian cancer (EOC), including the high-grade serous subtype (HGSC), and expression of individual POTEs correlates with chemoresistance and reduced survival in HGSC. The mechanisms driving POTE overexpression in EOC and other cancers is unknown. Here, we investigated the role of epigenetics in regulating POTE expression, with a focus on DNA hypomethylation. Consistent with their pericentromeric localization, Pan-POTE expression in EOC correlated with expression of the pericentromeric repeat NBL2, which was not the case for non-pericentromeric CTAs. POTE genomic regions contain LINE-1 (L1) sequences, and Pan-POTE expression correlated with both global and POTE-specific L1 hypomethylation in EOC. Analysis of individual POTEs using RNA-seq and DNA methylome data from fallopian tube epithelia (FTE) and HGSC revealed that POTEs C, E, and F have increased expression in HGSC in conjunction with DNA hypomethylation at 5' promoter or enhancer regions. Moreover, POTEs C/E/F showed additional increased expression in recurrent HGSC in conjunction with 5' hypomethylation, using patient-matched samples. Experiments using decitabine treatment and DNMT knockout cell lines verified a functional contribution of DNA methylation to POTE repression, and epigenetic drug combinations targeting histone deacetylases (HDACs) and histone methyltransferases (HMTs) in combination with decitabine further increased POTE expression. In summary, several alterations of the cancer epigenome, including pericentromeric activation, global and locus-specific L1 hypomethylation, and locus-specific 5' CpG hypomethylation, converge to promote POTE expression in ovarian cancer.
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Affiliation(s)
- Ashok Sharma
- a Eppley Institute , University of Nebraska Medical Center , Omaha , NE , USA.,b Fred & Pamela Buffett Cancer Center , University of Nebraska Medical Center , Omaha , NE , USA
| | - Mustafa Albahrani
- a Eppley Institute , University of Nebraska Medical Center , Omaha , NE , USA.,b Fred & Pamela Buffett Cancer Center , University of Nebraska Medical Center , Omaha , NE , USA
| | - Wa Zhang
- a Eppley Institute , University of Nebraska Medical Center , Omaha , NE , USA.,b Fred & Pamela Buffett Cancer Center , University of Nebraska Medical Center , Omaha , NE , USA
| | - Christina N Kufel
- c Department of Pharmacology and Therapeutics , Roswell Park Comprehensive Cancer Center , Buffalo , NY , USA
| | - Smitha R James
- c Department of Pharmacology and Therapeutics , Roswell Park Comprehensive Cancer Center , Buffalo , NY , USA
| | - Kunle Odunsi
- d Department of Immunology , Roswell Park Comprehensive Cancer Center , Buffalo , NY , USA.,e Department of Gynecologic Oncology , Roswell Park Comprehensive Cancer Center , Buffalo , NY , USA.,f Center for Immunotherapy , Roswell Park Comprehensive Cancer Center , Buffalo , NY , USA
| | - David Klinkebiel
- b Fred & Pamela Buffett Cancer Center , University of Nebraska Medical Center , Omaha , NE , USA.,g Department of Biochemistry and Molecular Biology , University of Nebraska Medical Center , Omaha , NE , USA
| | - Adam R Karpf
- a Eppley Institute , University of Nebraska Medical Center , Omaha , NE , USA.,b Fred & Pamela Buffett Cancer Center , University of Nebraska Medical Center , Omaha , NE , USA.,c Department of Pharmacology and Therapeutics , Roswell Park Comprehensive Cancer Center , Buffalo , NY , USA
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Barger CJ, Zhang W, Sharma A, Chee L, James SR, Kufel CN, Miller A, Meza J, Drapkin R, Odunsi K, Klinkebiel D, Karpf AR. Expression of the POTE gene family in human ovarian cancer. Sci Rep 2018; 8:17136. [PMID: 30459449 PMCID: PMC6244393 DOI: 10.1038/s41598-018-35567-1] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/05/2018] [Accepted: 11/07/2018] [Indexed: 12/23/2022] Open
Abstract
The POTE family includes 14 genes in three phylogenetic groups. We determined POTE mRNA expression in normal tissues, epithelial ovarian and high-grade serous ovarian cancer (EOC, HGSC), and pan-cancer, and determined the relationship of POTE expression to ovarian cancer clinicopathology. Groups 1 & 2 POTEs showed testis-specific expression in normal tissues, consistent with assignment as cancer-testis antigens (CTAs), while Group 3 POTEs were expressed in several normal tissues, indicating they are not CTAs. Pan-POTE and individual POTEs showed significantly elevated expression in EOC and HGSC compared to normal controls. Pan-POTE correlated with increased stage, grade, and the HGSC subtype. Select individual POTEs showed increased expression in recurrent HGSC, and POTEE specifically associated with reduced HGSC OS. Consistent with tumors, EOC cell lines had significantly elevated Pan-POTE compared to OSE and FTE cells. Notably, Group 1 & 2 POTEs (POTEs A/B/B2/C/D), Group 3 POTE-actin genes (POTEs E/F/I/J/KP), and other Group 3 POTEs (POTEs G/H/M) show within-group correlated expression, and pan-cancer analyses of tumors and cell lines confirmed this relationship. Based on their restricted expression in normal tissues and increased expression and association with poor prognosis in ovarian cancer, POTEs are potential oncogenes and therapeutic targets in this malignancy.
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Affiliation(s)
- Carter J Barger
- Eppley Institute for Cancer Research, University of Nebraska Medical Center, Omaha, NE, 68198-6805, USA
- Fred & Pamela Buffett Cancer Center, University of Nebraska Medical Center, Omaha, NE, 68198-6805, USA
| | - Wa Zhang
- Eppley Institute for Cancer Research, University of Nebraska Medical Center, Omaha, NE, 68198-6805, USA
- Fred & Pamela Buffett Cancer Center, University of Nebraska Medical Center, Omaha, NE, 68198-6805, USA
| | - Ashok Sharma
- Eppley Institute for Cancer Research, University of Nebraska Medical Center, Omaha, NE, 68198-6805, USA
- Fred & Pamela Buffett Cancer Center, University of Nebraska Medical Center, Omaha, NE, 68198-6805, USA
| | - Linda Chee
- Eppley Institute for Cancer Research, University of Nebraska Medical Center, Omaha, NE, 68198-6805, USA
- Fred & Pamela Buffett Cancer Center, University of Nebraska Medical Center, Omaha, NE, 68198-6805, USA
| | - Smitha R James
- Department of Pharmacology and Therapeutics, Roswell Park Comprehensive Cancer Center, Buffalo, NY, 14263, USA
| | - Christina N Kufel
- Department of Pharmacology and Therapeutics, Roswell Park Comprehensive Cancer Center, Buffalo, NY, 14263, USA
| | - Austin Miller
- Department of Biostatistics, Roswell Park Comprehensive Cancer Center, Buffalo, NY, 14263, USA
| | - Jane Meza
- Department of Biostatistics, University of Nebraska Medical Center, Omaha, NE, 68198-4375, USA
| | - Ronny Drapkin
- Penn Ovarian Cancer Research Center, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA, 19104, USA
| | - Kunle Odunsi
- Department of Immunology, Roswell Park Comprehensive Cancer Center, Buffalo, NY, 14263, USA
- Department of Gynecologic Oncology, Roswell Park Comprehensive Cancer Center, Buffalo, NY, 14263, USA
- Center for Immunotherapy, Roswell Park Comprehensive Cancer Center, Buffalo, NY, 14263, USA
| | - David Klinkebiel
- Fred & Pamela Buffett Cancer Center, University of Nebraska Medical Center, Omaha, NE, 68198-6805, USA
- Department of Biochemistry, University of Nebraska Medical Center, Omaha, NE, 68198, USA
| | - Adam R Karpf
- Eppley Institute for Cancer Research, University of Nebraska Medical Center, Omaha, NE, 68198-6805, USA.
- Fred & Pamela Buffett Cancer Center, University of Nebraska Medical Center, Omaha, NE, 68198-6805, USA.
- Department of Pharmacology and Therapeutics, Roswell Park Comprehensive Cancer Center, Buffalo, NY, 14263, USA.
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11
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Wang L, Li M, Zhan Y, Ban X, Zeng T, Zhu Y, Yun J, Guan XY, Li Y. Down-regulation of POTEG predicts poor prognosis in esophageal squamous cell carcinoma patients. Mol Carcinog 2018; 57:886-895. [PMID: 29566278 PMCID: PMC6001627 DOI: 10.1002/mc.22809] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/29/2017] [Revised: 03/08/2018] [Accepted: 03/20/2018] [Indexed: 12/18/2022]
Abstract
POTE ankyrin domain family, member G (poteg) belongs to POTE family. The POTE family is composed of many proteins which are very closely related and expressed in prostate, ovary, testis, and placenta. Some POTE paralogs are related with some cancers. Here we showed that down‐regulation of POTEG was detected in about 60% primary esophageal squamous cell carcinoma (ESCC) tumor tissues. Clinical association studies determined that POTEG down‐regulation was significantly correlated with tumor differentiation, lymph nodes metastasis and TNM staging. Kaplan‐Meier analysis determined that POTEG down‐regulation was associated with poorer clinical outcomes of ESCC patients (P = 0.026). Functional studies showed that POTEG overexpression could suppress tumor cell growth and metastasis capacity in vitro and in vivo. Molecular analyses revealed that POTEG downregulated CDKs, leading to subsequent inhibition of Rb phosphorylation, and consequently arrested Cell Cycle at G1/S Checkpoint. POTEG overexpression induced apoptosis by activating caspases and PARP, and regulating canonical mitochondrial apoptotic pathways. On the other side, POTEG inhibited epithelial‐mesenchymal transition and suppressed tumor cell metastasis. In conclusion, our study reveals a functionally important control mechanism of POTEG in esophageal cancer pathogenesis, suggesting potential use in the ESCC intervention and therapeutic strategies.
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Affiliation(s)
- Ling Wang
- State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou, P.R. China.,Guangdong Esophageal Cancer Institute, Guangzhou, P.R. China
| | - Mengqing Li
- State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou, P.R. China
| | - Yuting Zhan
- State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou, P.R. China
| | - Xiaojiao Ban
- State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou, P.R. China
| | - Tingting Zeng
- State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou, P.R. China
| | - Yinghui Zhu
- State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou, P.R. China
| | - Jingping Yun
- State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou, P.R. China
| | - Xin-Yuan Guan
- State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou, P.R. China.,Department of Clinical Oncology, The University of Hong Kong, Hong Kong, P.R. China
| | - Yan Li
- State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou, P.R. China
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12
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Balduini A, Raslova H, Di Buduo CA, Donada A, Ballmaier M, Germeshausen M, Balduini CL. Clinic, pathogenic mechanisms and drug testing of two inherited thrombocytopenias, ANKRD26-related Thrombocytopenia and MYH9-related diseases. Eur J Med Genet 2018; 61:715-722. [PMID: 29545013 DOI: 10.1016/j.ejmg.2018.01.014] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/25/2017] [Revised: 01/08/2018] [Accepted: 01/27/2018] [Indexed: 12/21/2022]
Abstract
Inherited thrombocytopenias (ITs) are a heterogeneous group of disorders characterized by low platelet count resulting in impaired hemostasis. Patients can have spontaneous hemorrhages and/or excessive bleedings provoked by hemostatic challenges as trauma or surgery. To date, ITs encompass 32 different rare monogenic disorders caused by mutations of 30 genes. This review will focus on the major discoveries that have been made in the last years on the diagnosis, treatment and molecular mechanisms of ANKRD26-Related Thrombocytopenia and MYH9-Related Diseases. Furthermore, we will discuss the use a Thrombopoietin mimetic as a novel approach to treat the thrombocytopenia in these patients. We will propose the use of a new 3D bone marrow model to study the mechanisms of action of these drugs and to test their efficacy and safety in patients. The overall purpose of this review is to point out that important progresses have been made in understanding the pathogenesis of ANKRD26-Related Thrombocytopenia and MYH9-Related Diseases and new therapeutic approaches have been proposed and tested. Future advancement in this research will rely in the development of more physiological models to study the regulation of human platelet biogenesis, disease mechanisms and specific pharmacologic targets.
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Affiliation(s)
- Alessandra Balduini
- University of Pavia, Pavia, Italy; IRCCS Policlinico San Matteo Foundation, Pavia, Italy.
| | - Hana Raslova
- INSERM UMR 1170, Gustave Roussy Cancer Campus, Université Paris-Saclay, Equipe Labellisée par la Ligue Nationale Contre le Cancer, Villejuif, France
| | - Christian A Di Buduo
- University of Pavia, Pavia, Italy; IRCCS Policlinico San Matteo Foundation, Pavia, Italy
| | - Alessandro Donada
- INSERM UMR 1170, Gustave Roussy Cancer Campus, Université Paris-Saclay, Equipe Labellisée par la Ligue Nationale Contre le Cancer, Villejuif, France
| | | | | | - Carlo L Balduini
- University of Pavia, Pavia, Italy; IRCCS Policlinico San Matteo Foundation, Pavia, Italy.
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13
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Baptista RLR, Dos Santos ACE, Gutiyama LM, Solza C, Zalcberg IR. Familial Myelodysplastic/Acute Leukemia Syndromes-Myeloid Neoplasms with Germline Predisposition. Front Oncol 2017; 7:206. [PMID: 28955657 PMCID: PMC5600909 DOI: 10.3389/fonc.2017.00206] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/09/2017] [Accepted: 08/23/2017] [Indexed: 12/16/2022] Open
Abstract
Although most cases of myeloid neoplasms are sporadic, a small subset has been associated with germline mutations. The 2016 revision of the World Health Organization classification included these cases in a myeloid neoplasm group with a predisposing germline mutational background. These patients must have a different management and their families should get genetic counseling. Cases identification and outline of the major known syndromes characteristics will be discussed in this text.
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Affiliation(s)
| | | | - Luciana Mayumi Gutiyama
- Divisão de Laboratórios do Centro de Transplantes de Medula Óssea (CEMO), Instituto Nacional do Câncer, Rio de Janeiro, Brazil
| | - Cristiana Solza
- Departamento de Medicina Interna/Hematologia, Hospital Universitário Pedro Ernesto, Rio de Janeiro, Brazil
| | - Ilana Renault Zalcberg
- Divisão de Laboratórios do Centro de Transplantes de Medula Óssea (CEMO), Instituto Nacional do Câncer, Rio de Janeiro, Brazil
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14
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Parry ML, Blanck G. Flat cells come full sphere: Are mutant cytoskeletal-related proteins oncoprotein-monsters or useful immunogens? Hum Vaccin Immunother 2015. [PMID: 26225584 DOI: 10.1080/21645515.2015.1073428] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/06/2023] Open
Abstract
Osteogenesis imperfecta is inherited as a dominant disease because if one allele is mutated, it contributes a mutant, destructive subunit polypeptide to collagen, which requires many subunits to form normal, polymeric, collagenous structures. Recent cancer genome atlas (TCGA) data indicate that cytoskeletal-related proteins are among the most commonly mutated proteins in human cancers, in distinct mutation frequency groups, i.e., including low mutation frequency groups. Part of the explanation for this observation is likely to be the fact that many of the coding regions for these proteins are very large, and indeed, it is likely these coding regions are mutated in many cells that never become cancerous. However, it would not be surprising if mutations in cytoskeletal proteins, when combined with oncoprotein or tumor suppressor protein mutations, had significant impacts on cancer development, for a number of reasons, including results obtained almost 5 decades ago indicating that well-spread cells in tissue culture, with well-formed cytoskeletons, were less tumorigenic than spherical cells with disrupted cytoskeletons. This raises the question, are mutant cytoskeletal proteins, which would likely interfere with polymer formation, a new class of oncoproteins, in particular, dominant negative oncoproteins? If these proteins are so commonly mutant, could they be the bases for common cancer vaccines?
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Affiliation(s)
- Michele L Parry
- a Department of Molecular Medicine ; Morsani College of Medicine; University of South Florida ; Tampa , FL USA
| | - George Blanck
- a Department of Molecular Medicine ; Morsani College of Medicine; University of South Florida ; Tampa , FL USA.,b Immunology Program; Moffitt Cancer Center and Research Institute ; Tampa , FL USA
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15
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Wang Q, Li X, Ren S, Cheng N, Zhao M, Zhang Y, Li J, Cai W, Zhao C, Cao W, Zhou C. Serum levels of the cancer-testis antigen POTEE and its clinical significance in non-small-cell lung cancer. PLoS One 2015; 10:e0122792. [PMID: 25860145 PMCID: PMC4393100 DOI: 10.1371/journal.pone.0122792] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/16/2014] [Accepted: 02/15/2015] [Indexed: 02/01/2023] Open
Abstract
Background POTEE (POTE ankyrin domain family, member E) is a newly identified cancer-testis antigen that has been found to be expressed in a wide variety of human cancers including cancers of the colon, prostate, lung, breast, ovary, and pancreas. Aim To measure the serum levels of POTEE in patients with non-small-cell lung cancer (NSCLC) and to explore the clinical significance of POTEE in NSCLC. Patients and Methods 104 NSCLC patients, 66 benign lung disease patients and 80 healthy volunteers were enrolled in this study from May 2013 to February 2014. Serum POTEE levels were measured using enzyme-linked immunosorbent assay (ELISA). Numerical variables were recorded as means ± standard deviation (SD) and analyzed by independent t tests. Categorical variables were calculated as rates and were analyzed using a χ2 test or Fisher’s exact test. Survival curves were estimated and compared using the Kaplan-Meier method and log-rank tests. Results Serum POTEE levels were significantly higher in NSCLC patients than in benign lung disease patients and healthy controls (mean ± SD [pg/ml], 324.38± 13.84 vs. 156.93 ± 17.38 and 139.09 ± 15.80, P<0.001) and were significantly correlated with TNM stage. Survival analysis revealed that patients with low serum POTEE had longer progression-free survival (PFS) than those with high serum POTEE (P=0.021). Cox multivariate analysis indicated that POTEE was an independent prognostic factor of progression-free survival (P =0.009, hazard ratio, 2.440). Conclusions Serum POTEE level in NSCLC patients is associated with TNM stage and is a potential prognostic factor.
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Affiliation(s)
- Qi Wang
- Department of Lung Cancer and Immunology, Shanghai Pulmonary Hospital, Tongji University, Tongji University Medical School Cancer Institute, Shanghai, People’s Republic of China
| | - Xuefei Li
- Department of Lung Cancer and Immunology, Shanghai Pulmonary Hospital, Tongji University, Tongji University Medical School Cancer Institute, Shanghai, People’s Republic of China
| | - Shengxiang Ren
- Department of Lung Cancer and Immunology, Shanghai Pulmonary Hospital, Tongji University, Tongji University Medical School Cancer Institute, Shanghai, People’s Republic of China
| | - Ningning Cheng
- Department of Lung Cancer and Immunology, Shanghai Pulmonary Hospital, Tongji University, Tongji University Medical School Cancer Institute, Shanghai, People’s Republic of China
| | - Mingchuan Zhao
- Department of Lung Cancer and Immunology, Shanghai Pulmonary Hospital, Tongji University, Tongji University Medical School Cancer Institute, Shanghai, People’s Republic of China
| | - Yishi Zhang
- Department of Lung Cancer and Immunology, Shanghai Pulmonary Hospital, Tongji University, Tongji University Medical School Cancer Institute, Shanghai, People’s Republic of China
| | - Jiayu Li
- Department of Lung Cancer and Immunology, Shanghai Pulmonary Hospital, Tongji University, Tongji University Medical School Cancer Institute, Shanghai, People’s Republic of China
| | - Weijing Cai
- Department of Lung Cancer and Immunology, Shanghai Pulmonary Hospital, Tongji University, Tongji University Medical School Cancer Institute, Shanghai, People’s Republic of China
| | - Chao Zhao
- Department of Lung Cancer and Immunology, Shanghai Pulmonary Hospital, Tongji University, Tongji University Medical School Cancer Institute, Shanghai, People’s Republic of China
| | - Wa Cao
- Department of Lung Cancer and Immunology, Shanghai Pulmonary Hospital, Tongji University, Tongji University Medical School Cancer Institute, Shanghai, People’s Republic of China
| | - Caicun Zhou
- Department of Lung Cancer and Immunology, Shanghai Pulmonary Hospital, Tongji University, Tongji University Medical School Cancer Institute, Shanghai, People’s Republic of China
- * E-mail:
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16
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Du X, Servin B, Womack JE, Cao J, Yu M, Dong Y, Wang W, Zhao S. An update of the goat genome assembly using dense radiation hybrid maps allows detailed analysis of evolutionary rearrangements in Bovidae. BMC Genomics 2014; 15:625. [PMID: 25052253 PMCID: PMC4141111 DOI: 10.1186/1471-2164-15-625] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/24/2013] [Accepted: 07/10/2014] [Indexed: 01/02/2023] Open
Abstract
Background The domestic goat (Capra hircus), an important livestock species, belongs to a clade of Ruminantia, Bovidae, together with cattle, buffalo and sheep. The history of genome evolution and chromosomal rearrangements on a small scale in ruminants remain speculative. Recently completed goat genome sequence was released but is still in a draft stage. The draft sequence used a variety of assembly packages, as well as a radiation hybrid (RH) map of chromosome 1 as part of its validation. Results Using an improved RH mapping pipeline, whole-genome dense maps of 45,953 SNP markers were constructed with statistical confidence measures and the saturated maps provided a fine map resolution of approximate 65 kb. Linking RH maps to the goat sequences showed that the assemblies of scaffolds/super-scaffolds were globally accurate. However, we observed certain flaws linked to the process of anchoring chromosome using conserved synteny with cattle. Chromosome assignments, long-range order, and orientation of the scaffolds were reassessed in an updated genome sequence version. We also present new results exploiting the updated goat genome sequence to understand genomic rearrangements and chromosome evolution between mammals during species radiations. The sequence architecture of rearrangement sites between the goat and cattle genomes presented abundant segmental duplication on regions of goat chromosome 9 and 14, as well as new insertions in homologous cattle genome regions. This complex interplay between duplicated sequences and Robertsonian translocations highlights the rearrangement mechanism of centromeric nonallelic homologous recombination (NAHR) in mammals. We observed that species-specific shifts in ANKRD26 gene duplication are coincident with breakpoint reuse in divergent lineages and this gene family may play a role in chromosome stabilization in chromosome evolution. Conclusions We generated dense maps of the complete whole goat genome. The chromosomal maps allowed us to anchor and orientate assembled genome scaffolds along the chromosomes, annotate chromosome rearrangements and thereby get a better understanding of the genome evolution of ruminants and other mammals. Electronic supplementary material The online version of this article (doi:10.1186/1471-2164-15-625) contains supplementary material, which is available to authorized users.
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Affiliation(s)
| | | | | | | | | | | | - Wen Wang
- Key lab of animal genetics, breeding and reproduction of ministry education, College of Animal Science and Technology, Huazhong Agricultural University, Wuhan 430070, People's Republic of China.
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17
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Bluteau D, Balduini A, Balayn N, Currao M, Nurden P, Deswarte C, Leverger G, Noris P, Perrotta S, Solary E, Vainchenker W, Debili N, Favier R, Raslova H. Thrombocytopenia-associated mutations in the ANKRD26 regulatory region induce MAPK hyperactivation. J Clin Invest 2014; 124:580-91. [PMID: 24430186 DOI: 10.1172/jci71861] [Citation(s) in RCA: 127] [Impact Index Per Article: 11.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/28/2013] [Accepted: 10/31/2013] [Indexed: 11/17/2022] Open
Abstract
Point mutations in the 5' UTR of ankyrin repeat domain 26 (ANKRD26) are associated with familial thrombocytopenia 2 (THC2) and a predisposition to leukemia. Here, we identified underlying mechanisms of ANKRD26-associated thrombocytopenia. Using megakaryocytes (MK) isolated from THC2 patients and healthy subjects, we demonstrated that THC2-associated mutations in the 5' UTR of ANKRD26 resulted in loss of runt-related transcription factor 1 (RUNX1) and friend leukemia integration 1 transcription factor (FLI1) binding. RUNX1 and FLI1 binding at the 5' UTR from healthy subjects led to ANKRD26 silencing during the late stages of megakaryopoiesis and blood platelet development. We showed that persistent ANKRD26 expression in isolated MKs increased signaling via the thrombopoietin/myeloproliferative leukemia virus oncogene (MPL) pathway and impaired proplatelet formation by MKs. Importantly, we demonstrated that ERK inhibition completely rescued the in vitro proplatelet formation defect. Our data identify a mechanism for development of the familial thrombocytopenia THC2 that is related to abnormal MAPK signaling.
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18
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Balduini CL, Pecci A, Noris P. Inherited thrombocytopenias: the evolving spectrum. Hamostaseologie 2012; 32:259-70. [PMID: 22972471 DOI: 10.5482/ha12050001] [Citation(s) in RCA: 39] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/15/2012] [Accepted: 08/28/2012] [Indexed: 12/23/2022] Open
Abstract
The chapter of inherited thrombocytopenias has expanded greatly over the last decade and many "new" forms deriving from mutations in "new" genes have been identified. Nevertheless, nearly half of patients remain without a definite diagnosis because their illnesses have not yet been described. The diagnostic approach to these diseases can still take advantage of the algorithm proposed by the Italian Platelet Study Group in 2003, although an update is required to include the recently described disorders. So far, transfusions of platelet concentrates have represented the main tool for preventing or treating bleedings, while haematopoietic stem cell transplantation has been reserved for patients with very severe forms. However, recent disclosure that an oral thrombopoietin mimetic is effective in increasing platelet count in patients with MYH9-related thrombocytopenia opened new therapeutic perspectives. This review summarizes the general aspects of inherited thrombocytopenias and describes in more detail MYH9-related diseases (encompassing four thrombocytopenias previously recognized as separate diseases) and the recently described ANKRD26-related thrombocytopenia, which are among the most frequent forms of inherited thrombocytopenia.
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Affiliation(s)
- C L Balduini
- Department of Internal Medicine, University of Pavia – IRCCS Policlinico San Matteo Foundation, Pavia, Italy.
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19
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Liu XF, Bera TK, Kahue C, Escobar T, Fei Z, Raciti GA, Pastan I. ANKRD26 and its interacting partners TRIO, GPS2, HMMR and DIPA regulate adipogenesis in 3T3-L1 cells. PLoS One 2012; 7:e38130. [PMID: 22666460 PMCID: PMC3364200 DOI: 10.1371/journal.pone.0038130] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2012] [Accepted: 05/04/2012] [Indexed: 01/04/2023] Open
Abstract
Partial inactivation of the Ankyrin repeat domain 26 (Ankrd26) gene causes obesity and diabetes in mice and increases spontaneous and induced adipogenesis in mouse embryonic fibroblasts. However, it is not yet known how the Ankrd26 protein carries out its biological functions. We identified by yeast two-hybrid and immunoprecipitation assays the triple functional domain protein (TRIO), the G protein pathway suppressor 2 (GPS2), the delta-interacting protein A (DIPA) and the hyaluronan-mediated motility receptor (HMMR) as ANKRD26 interacting partners. Adipogenesis of 3T3-L1 cells was increased by selective down-regulation of Ankrd26, Trio, Gps2, Hmmr and Dipa. Furthermore, GPS2 and DIPA, which are normally located in the nucleus, were translocated to the cytoplasm, when the C-terminus of ANKRD26 was introduced into these cells. These findings provide biochemical evidence that ANKRD26, TRIO, GPS2 and HMMR are novel and important regulators of adipogenesis and identify new targets for the modulation of adipogenesis.
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Affiliation(s)
- Xiu-Fen Liu
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Tapan K. Bera
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Charissa Kahue
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Thelma Escobar
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Zhaoliang Fei
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Gregory A. Raciti
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America
| | - Ira Pastan
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland, United States of America
- * E-mail:
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20
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Fei Z, Bera TK, Liu X, Xiang L, Pastan I. Ankrd26 gene disruption enhances adipogenesis of mouse embryonic fibroblasts. J Biol Chem 2011; 286:27761-8. [PMID: 21669876 DOI: 10.1074/jbc.m111.248435] [Citation(s) in RCA: 43] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
We previously reported that partial disruption of the Ankrd26 gene in mice leads to hyperphagia and leptin-resistant obesity. To determine whether the Ankrd26 mutation can affect the development of adipocytes, we studied mouse embryo fibroblasts (MEFs) from the mutant mice. We found that Ankrd26(-/-) MEFs have a higher rate of spontaneous adipogenesis than normal MEFs and that adipocyte formation is greatly increased when the cells are induced with troglitazone alone or with a mixture of troglitazone, insulin, dexamethasone, and methylisobutylxanthine. Increased adipogenesis was detected as an increase in lipid droplet formation and in the expression of several markers of adipogenesis. There was an increase in expression of early stage adipogenesis genes such as Krox20, KLF5, C/EBPβ, C/EBPδ, and late stage adipogenesis regulators KLF15, C/EBPα, PPARγ, and aP2. There was also an increase in adipocyte stem cell markers CD34 and Sca-1 and preadipocyte markers Gata2 and Pref-1, indicating an increase in both stem cells and progenitor cells in the mutant MEFs. Furthermore, ERK was found constitutively activated in Anrd26(-/-) MEFs, and the addition of MEK inhibitors to mutant cells blocked ERK activation, decreased adipogenesis induction, and significantly reduced expression of C/EBPδ, KLF15, PPARγ2, CD34, and Pref-1 genes. We conclude that Ankrd26 gene disruption promotes adipocyte differentiation at both the progenitor commitment and differentiation steps and that ERK activation plays a role in this process.
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Affiliation(s)
- Zhaoliang Fei
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892-4264, USA
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21
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Brun ME, Lana E, Rivals I, Lefranc G, Sarda P, Claustres M, Mégarbané A, De Sario A. Heterochromatic genes undergo epigenetic changes and escape silencing in immunodeficiency, centromeric instability, facial anomalies (ICF) syndrome. PLoS One 2011; 6:e19464. [PMID: 21559330 PMCID: PMC3084872 DOI: 10.1371/journal.pone.0019464] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/01/2011] [Accepted: 03/30/2011] [Indexed: 12/01/2022] Open
Abstract
Immunodeficiency, Centromeric Instability, Facial Anomalies (ICF) syndrome is a rare autosomal recessive disorder that is characterized by a marked immunodeficiency, severe hypomethylation of the classical satellites 2 and 3 associated with disruption of constitutive heterochromatin, and facial anomalies. Sixty percent of ICF patients have mutations in the DNMT3B (DNA methyltransferase 3B) gene, encoding a de novo DNA methyltransferase. In the present study, we have shown that, in ICF lymphoblasts and peripheral blood, juxtacentromeric heterochromatic genes undergo dramatic changes in DNA methylation, indicating that they are bona fide targets of the DNMT3B protein. DNA methylation in heterochromatic genes dropped from about 80% in normal cells to approximately 30% in ICF cells. Hypomethylation was observed in five ICF patients and was associated with activation of these silent genes. Although DNA hypomethylation occurred in all the analyzed heterochromatic genes and in all the ICF patients, gene expression was restricted to some genes, every patient having his own group of activated genes. Histone modifications were preserved in ICF patients. Heterochromatic genes were associated with histone modifications that are typical of inactive chromatin: they had low acetylation on H3 and H4 histones and were slightly enriched in H3K9Me(3), both in ICF and controls. This was also the case for those heterochromatic genes that escaped silencing. This finding suggests that gene activation was not generalized to all the cells, but rather was restricted to a clonal cell population that may contribute to the phenotypic variability observed in ICF syndrome. A slight increase in H3K27 monomethylation was observed both in heterochromatin and active euchromatin in ICF patients; however, no correlation between this modification and activation of heterochromatic genes was found.
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Affiliation(s)
| | - Erica Lana
- INSERM U827, Montpellier, France
- Université Montpellier 1, Montpellier, France
| | | | - Gérard Lefranc
- CNRS UPR 1142, Montpellier, France
- Université Montpellier 2, Montpellier, France
| | | | - Mireille Claustres
- INSERM U827, Montpellier, France
- Université Montpellier 1, Montpellier, France
- CHRU, Montpellier, France
| | - André Mégarbané
- Unité de Génétique Médicale and Laboratoire Associé INSERM à l’UMR S910, Faculty of Medicine, Saint Joseph University, Beirut, Lebanon
- Institut Jérôme Lejeune, Paris, France
| | - Albertina De Sario
- INSERM U827, Montpellier, France
- Université Montpellier 1, Montpellier, France
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Mutations in the 5' UTR of ANKRD26, the ankirin repeat domain 26 gene, cause an autosomal-dominant form of inherited thrombocytopenia, THC2. Am J Hum Genet 2011; 88:115-20. [PMID: 21211618 DOI: 10.1016/j.ajhg.2010.12.006] [Citation(s) in RCA: 157] [Impact Index Per Article: 11.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/05/2010] [Revised: 12/11/2010] [Accepted: 12/16/2010] [Indexed: 11/24/2022] Open
Abstract
THC2, an autosomal-dominant thrombocytopenia described so far in only two families, has been ascribed to mutations in MASTL or ACBD5. Here, we show that ANKRD26, another gene within the THC2 locus, and neither MASTL nor ACBD5, is mutated in eight unrelated families. ANKRD26 was also found to be mutated in the family previously reported to have an ACBD5 mutation. We identified six different ANKRD26 mutations, which were clustered in a highly conserved 19 bp sequence located in the 5' untranslated region. Mutations were not detected in 500 controls and are absent from the 1000 Genomes database. Available data from an animal model and Dr. Watson's genome give evidence against haploinsufficiency as the pathogenetic mechanism for ANKRD26-mediated thrombocytopenia. The luciferase reporter assay suggests that these 5' UTR mutations might enhance ANKRD26 expression. ANKRD26 is the ancestor of a family of primate-specific genes termed POTE, which have been recently identified as a family of proapoptotic proteins. Dysregulation of apoptosis might therefore be the pathogenetic mechanism, as demonstrated for another thrombocytopenia, THC4. Further investigation is needed to provide evidence supporting this hypothesis.
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Sahab ZJ, Hall MD, Zhang L, Cheema AK, Byers SW. Tumor Suppressor RARRES1 Regulates DLG2, PP2A, VCP, EB1, and Ankrd26. J Cancer 2010; 1:14-22. [PMID: 20842219 PMCID: PMC2931349 DOI: 10.7150/jca.1.14] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022] Open
Abstract
Retinoic Acid Receptor Responder (RARRES1) initially identified as a novel retinoic acid receptor regulated gene in the skin is a putative tumor suppressor of unknown function. RARRES1 was knocked down in immortalized human prostatic epithelial cell line PWR-1E cells and differential protein expression was identified using differential in-gel electrophoresis (DIGE) followed by matrix-assisted laser desorption ionization (MALDI) mass spectrometry and western Blot analysis excluding highly abundant proteins routinely identified in almost all proteomics projects. Knock-down of RARRES1: 1- down-regulates PP2A, an enzyme involved in the negative regulation of the growth hormone-stimulated signal transduction pathways; 2- down-regulates Valosin-containing protein causing impaired autophagy; 3- up-regulates the tumor suppressor disks large 2; 4- up-regulates Ankrd26 that belongs to the POTE family of genes that are highly expressed in cancer patients with poor outcome; and 5- down-regulates EB1, a protein that is involved in spindle dynamics and chromosome alignment during mitosis.
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Affiliation(s)
- Ziad J Sahab
- 1. Georgetown University Medical Center, Lombardi Comprehensive Cancer Center, Department of Oncology, Washington, DC, 20007, USA
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24
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Abstract
The primate-specific gene family, POTE, is expressed in many cancers but only in a limited number of normal tissues (testis, ovary, prostate). The 13 POTE paralogs are dispersed among 8 human chromosomes. They evolved by gene duplication and remodeling from an ancestral gene, Ankrd26, recently implicated in controlling body size and obesity. In addition, several POTE paralogs are fused to an actin retrogene producing POTE-actin fusion proteins. The biological function of the POTE genes is unknown, but their high expression in primary spermatocytes, some of which are undergoing apoptosis, suggests a role in inducing programmed cell death. We have chosen Hela cells as a model to study POTE function in human cancer, and have identified POTE-2alpha-actin as the major transcript and the protein it encodes in Hela cells. Transfection experiments show that both POTE-2alpha-actin and POTE-2gammaC are localized to actin filaments close to the inner plasma membrane. Transient expression of POTE-2alpha-actin or POTE-2gammaC induces apoptosis in Hela cells. Using wild-type and mutant mouse embryo cells, we find apoptosis induced by over-expression of POTE-2gammaC is decreased in Bak ( -/- ) or Bak ( -/- ) Bax ( -/- ) cells indicating POTE is acting through a mitochondrial pathway. Endogenous POTE-actin protein levels but not RNA levels increased in a time dependent manner by stimulation of death receptors with their cognate ligands. Our data indicates that the POTE gene family encodes a new family of proapoptotic proteins.
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Affiliation(s)
- Xiu Fen Liu
- National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264, USA
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25
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Wang Y, Leung FCC. Discovery of a long inverted repeat in human POTE genes. Genomics 2009; 94:278-83. [PMID: 19463943 DOI: 10.1016/j.ygeno.2009.05.005] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/16/2009] [Revised: 05/08/2009] [Accepted: 05/13/2009] [Indexed: 01/18/2023]
Abstract
POTE gene family is tightly related to prostate, ovary, testis and placenta cancers. We recently identified an intronic long inverted repeat (LIR) in some members of the POTE gene family. Due to the capacity of inducing gene amplification, the POTE intronic LIRs may be involved in over-expression of the POTE genes. Our study aimed to understand the origin of the LIR in primates. We collected the LIR and its flanking sequences within rhesus monkey, chimpanzee and human genomes. The rhesus monkey genome only has half-sized LIRs (lack one repeat copy), whereas the human and chimpanzee genomes contain both full-sized and half-sized LIRs. Phylogenetic tree indicates that the LIR is formed after divergence of rhesus monkey and the common ancestor of human and chimpanzee. The POTE genes containing a full-sized LIR were amplified in the human genome.
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Affiliation(s)
- Yong Wang
- School of Biological Sciences and Genome Research Centre, The University of Hong Kong, Pokfulam, Hong Kong, China
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26
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Bera T, Lee B. Mining of Genome Sequence Databases to Identify New Targets for Prostate and Breast Cancer Therapy. Genomics 2008. [DOI: 10.3109/9781420067064-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022]
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27
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Ko MJ, Choi HS, Ahn JI, Kim SY, Jeong HS, Chung HJ. Gene Expression Profiling in C57BL/6 Mice Treated with the Anorectic Drugs Sibutramine and Phendimetrazine and Their Mechanistic Implications. Genomics Inform 2008. [DOI: 10.5808/gi.2008.6.3.117] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
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28
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Bera TK, Saint Fleur A, Ha D, Yamada M, Lee Y, Lee B, Hahn Y, Kaufman DS, Pera M, Pastan I. Selective POTE paralogs on chromosome 2 are expressed in human embryonic stem cells. Stem Cells Dev 2008; 17:325-32. [PMID: 18447647 PMCID: PMC7233169 DOI: 10.1089/scd.2007.0079] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022] Open
Abstract
POTE is a primate-specific gene family that encodes cancer testis antigens that contain three domains, although the proteins vary greatly in size. The amino-terminal domain is novel and has three cysteine-rich domains of 37 amino acids. The second and third domains are rich in ankyrin repeats and spectrin-like helices respectively. In humans, 13 highly homologous paralogs are dispersed among eight chromosomes. Some members of the POTE gene family have an actin insertion at the carboxyl end of the protein. The expression of the POTE gene in normal adult tissues is restricted, but several POTE paralogs are frequently expressed in many cancers including breast, prostate, and lung cancers. We show here that POTE is expressed in several human embryonic stem (ES) cell lines. We found that UC06, WA01 and ES03 cell lines express mainly a POTE-2gamma transcript but ES02 and ES04 cell lines predominantly express POTE-2alpha. The WA09 cell line expressed both POTE-2gamma and POTE-2alpha. There is no detectable POTE gene expression in fetal tissues (ages 16-36 weeks). The POTE paralogs that are expressed in ES cells may have a specific function during lineage-specific differentiation of ES cells.
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Affiliation(s)
- Tapan K Bera
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
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29
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Ise T, Das S, Nagata S, Maeda H, Lee Y, Onda M, Anver MR, Bera TK, Pastan I. Expression of POTE protein in human testis detected by novel monoclonal antibodies. Biochem Biophys Res Commun 2008; 365:603-8. [PMID: 17996727 PMCID: PMC2189737 DOI: 10.1016/j.bbrc.2007.10.195] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/17/2007] [Accepted: 10/27/2007] [Indexed: 10/22/2022]
Abstract
The POTE gene family is composed of 13 highly homologous paralogs preferentially expressed in prostate, ovary, testis, and placenta. We produced 10 monoclonal antibodies (MAbs) against three representative POTE paralogs: POTE-21, POTE-2gammaC, and POTE-22. One reacted with all three paralogs, six MAbs reacted with POTE-2gammaC and POTE-22, and three MAbs were specific to POTE-21. Epitopes of all 10 MAbs were located in the cysteine-rich repeats (CRRs) motifs located at the N-terminus of each POTE paralog. Testing the reactivity of each MAb with 12 different CRRs revealed slight differences among the antigenic determinants, which accounts for differences in cross-reactivity. Using MAbs HP8 and PG5 we were able to detect a POTE-actin fusion protein in human testis by immunoprecipitation followed by Western blotting. By immunohistochemistry we demonstrated that the POTE protein is expressed in primary spermatocytes, implying a role in spermatogenesis.
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Affiliation(s)
- Tomoko Ise
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264 USA
| | - Sudipto Das
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264 USA
| | - Satoshi Nagata
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264 USA
| | - Hiroshi Maeda
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264 USA
| | - Yoomi Lee
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264 USA
| | - Masanori Onda
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264 USA
| | - Miriam R. Anver
- Pathology/Histotechnology Laboratory, SAIC-Frederick, National Cancer Institute-Frederick, Frederick, MD 21702-1201 USA
| | - Tapan K. Bera
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264 USA
| | - Ira Pastan
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264 USA
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30
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Bera TK, Liu XF, Yamada M, Gavrilova O, Mezey E, Tessarollo L, Anver M, Hahn Y, Lee B, Pastan I. A model for obesity and gigantism due to disruption of the Ankrd26 gene. Proc Natl Acad Sci U S A 2008; 105:270-5. [PMID: 18162531 PMCID: PMC2224199 DOI: 10.1073/pnas.0710978105] [Citation(s) in RCA: 53] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/31/2007] [Indexed: 11/18/2022] Open
Abstract
Obesity is a major health hazard that is caused by a combination of genetic and behavioral factors. Several models of obesity have been described in mice that have defects in the production of peptide hormones, in the function of cell membrane receptors, or in a transcription factor required for neuronal cell development. We have been investigating the function of a family of genes (POTE and ANKRD26) that encode proteins that are associated with the inner aspect of the cell membrane and that contain both ankyrin repeats and spectrin helices, motifs known to interact with signaling proteins in the cell. To assess the function of ANKRD26, we prepared a mutant mouse with partial inactivation of the Ankrd26 gene. We find that the homozygous mutant mice develop extreme obesity, insulin resistance, and an increase in body size. The obesity is associated with hyperphagia with no reduction in energy expenditure and activity. The Ankrd26 protein is expressed in the arcuate and ventromedial nuclei within the hypothalamus and in the ependyma and the circumventricular organs that act as an interface between the peripheral circulation and the brain. In the enlarged hearts of the mutant mice, the levels of both phospho-Akt and mTOR were elevated. These results show that alterations in an unidentified gene can lead to obesity and identify a molecular target for the treatment of obesity.
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Affiliation(s)
- Tapan K. Bera
- *Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Xiu-Fen Liu
- *Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Masanori Yamada
- *Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Oksana Gavrilova
- Mouse Metabolism Core Laboratory, National Institute of Diabetes, Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892-0803
| | - Eva Mezey
- Craniofacial and Skeletal Diseases Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD 20892-4479
| | - Lino Tessarollo
- Mouse Cancer Genetics Program, Center for Cancer Research, and
| | - Miriam Anver
- Pathology/Histotechnology Laboratory, SAIC-Frederick, National Cancer Institute, Frederick, MD 21702
| | - Yoonsoo Hahn
- *Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Byungkook Lee
- *Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Ira Pastan
- *Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
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31
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Das S, Ise T, Nagata S, Maeda H, Bera TK, Pastan I. Palmitoylation of POTE family proteins for plasma membrane targeting. Biochem Biophys Res Commun 2007; 363:751-6. [PMID: 17904529 PMCID: PMC2170890 DOI: 10.1016/j.bbrc.2007.09.045] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/28/2007] [Accepted: 09/07/2007] [Indexed: 11/24/2022]
Abstract
The POTE gene family is composed of 13 paralogs and likely evolved by duplications and remodeling of the human genome. One common property of POTE proteins is their localization on the inner aspect of the plasma membrane. To determine the structural elements required for membrane localization, we expressed mutants of different POTEs in 293T cells as EGFP fusion proteins. We also tested their palmitoylation by a biotin-switch assay. Our data indicate that the membrane localizations of different POTEs are mediated by similar 3-4 short cysteine rich repeats (CRRs) near the amino-terminuses and that palmitoylation on paired cysteine residues in each CRR motif is responsible for the localization. Multiple palmitoylation in the small CRRs can result in the strong association of whole POTEs with plasma membrane.
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Affiliation(s)
| | | | - Satoshi Nagata
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892-4264, USA
| | | | - Tapan K. Bera
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892-4264, USA
| | - Ira Pastan
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, Maryland 20892-4264, USA
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32
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Lee Y, Ise T, Ha D, Saint Fleur A, Hahn Y, Liu XF, Nagata S, Lee B, Bera TK, Pastan I. Evolution and expression of chimeric POTE-actin genes in the human genome. Proc Natl Acad Sci U S A 2006; 103:17885-90. [PMID: 17101985 PMCID: PMC1693842 DOI: 10.1073/pnas.0608344103] [Citation(s) in RCA: 36] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
We previously described a primate-specific gene family, POTE, that is expressed in many cancers but in a limited number of normal organs. The 13 POTE genes are dispersed among eight different chromosomes and evolved by duplications and remodeling of the human genome from an ancestral gene, ANKRD26. Based on sequence similarity, the POTE gene family members can be divided into three groups. By genome database searches, we identified an actin retroposon insertion at the carboxyl terminus of one of the ancestral POTE paralogs. By Northern blot analysis, we identified the expected 7.5-kb POTE-actin chimeric transcript in a breast cancer cell line. The protein encoded by the POTE-actin transcript is predicted to be 120 kDa in size. Using anti-POTE mAbs that recognize the amino-terminal portion of the POTE protein, we detected the 120-kDa POTE-actin fusion protein in breast cancer cell lines known to express the fusion transcript. These data demonstrate that insertion of a retroposon produced an altered functional POTE gene. This example indicates that new functional human genes can evolve by insertion of retroposons.
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Affiliation(s)
- Yoomi Lee
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Tomoko Ise
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Duc Ha
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Ashley Saint Fleur
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Yoonsoo Hahn
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Xiu-Fen Liu
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Satoshi Nagata
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Byungkook Lee
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Tapan K. Bera
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
| | - Ira Pastan
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892-4264
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33
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Murad H, Collet P, Brunner E, Schohn H, Bécuwe P, Devignes MD, Dauça M, Domenjoud L. Immunoselection and characterization of a human genomic PPAR binding fragment located within POTE genes. Biochimie 2006; 89:329-36. [PMID: 17070643 DOI: 10.1016/j.biochi.2006.09.017] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2006] [Accepted: 09/14/2006] [Indexed: 11/18/2022]
Abstract
Peroxisome proliferator-activated receptors (PPARs) are ligand-inducible transcription factors and belong to the nuclear hormone receptor superfamily. They form heterodimers with retinoid X receptor (RXR) and bind to specific PPAR-response elements. To identify novel PPAR target genes, we developed an affinity method to isolate human genomic fragments containing binding sites for PPARs. For this, an antibody raised against all PPAR subtypes was used. Immunoselected fragments were amplified and sequenced. One of them, ISF1029, was mapped by BLAT and BLAST searches on different human chromosomes, downstream of several POTE genes. ISF1029 contained three hexamers strongly related to the AGGTCA motif organized according to a DR0/3 motif. The latter was found to bind to PPARalpha in gel mobility shift and supershift assays and to exhibit a downregulation potentiality in transfection experiments under clofibrate treatment. POTE genes were shown to be highly expressed in human Caco-2 colorectal adenocarcinoma cells and downregulated by fenofibrate and 9-cis-retinoic acid, as attested by RT-PCR assays. Microarray analysis confirmed and extended to the human T98-G glioblastoma cells, the downregulation of several POTE genes expression by Wy-14,643, a potent PPARalpha activator. Our data provide new insights about the pleiotropic action of PPARs.
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Affiliation(s)
- Hossam Murad
- EA3446 Proliférateurs de Peroxysomes Université Henri Poincaré-Nancy I, Faculté des Sciences, BP239, 54506 Vandoeuvre-les-Nancy, France
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Bera TK, Saint Fleur A, Lee Y, Kydd A, Hahn Y, Popescu NC, Zimonjic DB, Lee B, Pastan I. POTE paralogs are induced and differentially expressed in many cancers. Cancer Res 2006; 66:52-6. [PMID: 16397215 DOI: 10.1158/0008-5472.can-05-3014] [Citation(s) in RCA: 55] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/29/2023]
Abstract
To identify new antigens that are targets for the immunotherapy of prostate and breast cancer, we used expressed sequence tag and genomic databases and discovered POTE, a new primate-specific gene family. Each POTE gene encodes a protein that contains three domains, although the proteins vary greatly in size. The NH2-terminal domain is novel and has properties of an extracellular domain but does not contain a signal sequence. The second and third domains are rich in ankyrin repeats and spectrin-like helices, respectively. The protein encoded by POTE-21, the first family member discovered, is localized on the plasma membrane of the cell. In humans, 13 highly homologous paralogs are dispersed among eight chromosomes. The expression of POTE genes in normal tissues is restricted to prostate, ovary, testis, and placenta. A survey of several cancer samples showed that POTE was expressed in 6 of 6 prostate, 12 of 13 breast, 5 of 5 colon, 5 of 6 lung, and 4 of 5 ovarian cancers. To determine the relative expression of each POTE paralog in cancer and normal samples, we employed a PCR-based cloning and analysis method. We found that POTE-2alpha, POTE-2beta, POTE-2gamma, and POTE-22 are predominantly expressed in cancers whereas POTE expression in normal tissues is somewhat more diverse. Because POTE is primate specific and is expressed in testis and many cancers but only in a few normal tissues, we conclude POTE is a new primate-specific member of the cancer-testis antigen family. It is likely that POTE has a unique role in primate biology.
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Affiliation(s)
- Tapan K Bera
- Laboratory of Molecular Biology, Center for Cancer Research, National Cancer Institute, NIH, Bethesda, Maryland 20892-4264, USA
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