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Huang CW, Zhang WZ, Liao Y, Hu T, Li JM, Wang CL. A targeted approach: Gene and RNA editing for neurodegenerative disease treatment. Life Sci 2025; 376:123756. [PMID: 40412606 DOI: 10.1016/j.lfs.2025.123756] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/24/2025] [Revised: 05/15/2025] [Accepted: 05/21/2025] [Indexed: 05/27/2025]
Abstract
With the global aging trend, neurodegenerative diseases (NDs) have emerged as a significant public health concern in the 21st century, imposing substantial economic burdens on families and society. NDs are characterized by cognitive and motor decline, resulting from a combination of genetic and environmental factors. Currently, there is no cure for NDs. Gene and RNA editing therapies offer new possibilities for addressing NDs. Gene editing involves modifying mutant genes associated with NDs, while RNA editing can directly modify RNA molecules to regulate the protein translation process, potentially influencing the expression of genes related to NDs. In this review, we examined the historical evolution, mechanisms of action, applications in NDs, advantages and disadvantages, as well as ethical and safety considerations of gene and RNA editing. While gene and RNA editing technologies hold promise for treating NDs, further research and development are needed to address safety, efficacy, and treatment timing issues, ultimately offering improved treatment options for ND patients. Our review provides valuable insights for future gene and RNA editing applications in ND treatment.
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Affiliation(s)
- Chen-Wei Huang
- Department of Stress Medicine, Faculty of Psychology, Naval Medical University, Shanghai, 200433, China
| | - Wang-Zheqi Zhang
- Department of Anesthesiology, Changhai Hospital, Naval Medical University, Shanghai 200433, China; School of Anesthesiology, Naval Medical University, Shanghai 200433, China
| | - Yan Liao
- Department of Anesthesiology, Changhai Hospital, Naval Medical University, Shanghai 200433, China; School of Anesthesiology, Naval Medical University, Shanghai 200433, China
| | - Ting Hu
- Department of Stress Medicine, Faculty of Psychology, Naval Medical University, Shanghai, 200433, China
| | - Jia-Mei Li
- Department of Neurology, The 971st Hospital of Navy, Qingdao 266071, China.
| | - Chang-Li Wang
- Department of Anesthesiology, Changhai Hospital, Naval Medical University, Shanghai 200433, China.
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Sheng H, Gao P, Yang C, Quilichini TD, Kochian LV, Datla R, Xiang D. Advances in Genome Editing Through Haploid Induction Systems. Int J Mol Sci 2025; 26:4779. [PMID: 40429922 PMCID: PMC12112045 DOI: 10.3390/ijms26104779] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/12/2025] [Revised: 05/05/2025] [Accepted: 05/09/2025] [Indexed: 05/29/2025] Open
Abstract
Groundbreaking advances in gene editing technologies are transforming modern plant breeding by enabling precise genetic modifications that dramatically accelerate crop improvement. Haploid and diploid induction systems have emerged as particularly powerful tools in this landscape, offering both efficient gene editing capabilities and rapid production of homozygous lines while seamlessly integrating with the advanced genome-editing platforms such as CRISPR-Cas systems. This review synthesizes the current state of knowledge regarding the mechanisms, applications, and recent progress in haploid and diploid induction systems for gene editing. We examine their transformative potential for enhancing genetic gains and compressing breeding timelines, with significant implications for global food security. Additionally, we provide a critical analysis of emerging challenges of genome editing in crops and outline promising future directions for research and development.
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Affiliation(s)
- Huajin Sheng
- Department of Biology, College of Arts and Science, University of Saskatchewan, Saskatoon, SK S7N 5E2, Canada; (H.S.); (T.D.Q.)
| | - Peng Gao
- Agriculture and Agri-Food Canada, Saskatoon Research and Development Centre, 107 Science Place, Saskatoon, SK S7N 0X2, Canada;
| | - Changye Yang
- Aquatic and Crop Resource Development, National Research Council Canada, Saskatoon, SK S7N 0W9, Canada;
| | - Teagen D. Quilichini
- Department of Biology, College of Arts and Science, University of Saskatchewan, Saskatoon, SK S7N 5E2, Canada; (H.S.); (T.D.Q.)
| | - Leon V. Kochian
- Department of Plant Sciences, University of Saskatchewan, Saskatoon, SK S7N 5A8, Canada;
| | - Raju Datla
- Global Institute for Food Security, University of Saskatchewan, Saskatoon, SK S7N 4J8, Canada;
| | - Daoquan Xiang
- Aquatic and Crop Resource Development, National Research Council Canada, Saskatoon, SK S7N 0W9, Canada;
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Boff MO, Xavier FAC, Diz FM, Gonçalves JB, Ferreira LM, Zambeli J, Pazzin DB, Previato TTR, Erwig HS, Gonçalves JIB, Bruzzo FTK, Marinowic D, da Costa JC, Zanirati G. mTORopathies in Epilepsy and Neurodevelopmental Disorders: The Future of Therapeutics and the Role of Gene Editing. Cells 2025; 14:662. [PMID: 40358185 PMCID: PMC12071303 DOI: 10.3390/cells14090662] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2025] [Revised: 02/06/2025] [Accepted: 02/06/2025] [Indexed: 05/15/2025] Open
Abstract
mTORopathies represent a group of neurodevelopmental disorders linked to dysregulated mTOR signaling, resulting in conditions such as tuberous sclerosis complex, focal cortical dysplasia, hemimegalencephaly, and Smith-Kingsmore Syndrome. These disorders often manifest with epilepsy, cognitive impairments, and, in some cases, structural brain anomalies. The mTOR pathway, a central regulator of cell growth and metabolism, plays a crucial role in brain development, where its hyperactivation leads to abnormal neuroplasticity, tumor formation, and heightened neuronal excitability. Current treatments primarily rely on mTOR inhibitors, such as rapamycin, which reduce seizure frequency and tumor size but fail to address underlying genetic causes. Advances in gene editing, particularly via CRISPR/Cas9, offer promising avenues for precision therapies targeting the genetic mutations driving mTORopathies. New delivery systems, including viral and non-viral vectors, aim to enhance the specificity and efficacy of these therapies, potentially transforming the management of these disorders. While gene editing holds curative potential, challenges remain concerning delivery, long-term safety, and ethical considerations. Continued research into mTOR mechanisms and innovative gene therapies may pave the way for transformative, personalized treatments for patients affected by these complex neurodevelopmental conditions.
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Affiliation(s)
- Marina Ottmann Boff
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
- School of Medicine, Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90619-900, RS, Brazil
| | - Fernando Antônio Costa Xavier
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
- Graduate Program in Medicine and Health Sciences, School of Medicine, Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90619-900, RS, Brazil
| | - Fernando Mendonça Diz
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
| | - Júlia Budelon Gonçalves
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
| | - Laura Meireles Ferreira
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
- School of Medicine, Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90619-900, RS, Brazil
| | - Jean Zambeli
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
- School of Medicine, University of the Valley of the Rio dos Sinos (UNISINOS), São Leopoldo 93022-750, RS, Brazil
| | - Douglas Bottega Pazzin
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
- Graduate Program in Pediatrics and Child Health, School of Medicine, Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90619-900, RS, Brazil
| | - Thales Thor Ramos Previato
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
- Graduate Program in Biomedical Gerontology, School of Medicine, Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90619-900, RS, Brazil
| | - Helena Scartassini Erwig
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
- School of Health and Life, Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90619-900, RS, Brazil
| | - João Ismael Budelon Gonçalves
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
| | - Fernanda Thays Konat Bruzzo
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
| | - Daniel Marinowic
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
- School of Health and Life, Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90619-900, RS, Brazil
| | - Jaderson Costa da Costa
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
| | - Gabriele Zanirati
- Brain Institute of Rio Grande do Sul (BraIns), Pontifical Catholic University of Rio Grande do Sul (PUCRS), Porto Alegre 90610-000, RS, Brazil; (M.O.B.); (F.A.C.X.); (F.M.D.); (J.B.G.); (L.M.F.); (J.Z.); (D.B.P.); (T.T.R.P.); (H.S.E.); (J.I.B.G.); (F.T.K.B.); (D.M.); (J.C.d.C.)
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4
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Halim CE, Deng S, Crasta KC, Yap CT. Interplay Between the Cytoskeleton and DNA Damage Response in Cancer Progression. Cancers (Basel) 2025; 17:1378. [PMID: 40282554 PMCID: PMC12025774 DOI: 10.3390/cancers17081378] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/05/2025] [Revised: 04/14/2025] [Accepted: 04/16/2025] [Indexed: 04/29/2025] Open
Abstract
DNA damage has emerged as a critical factor in fuelling the development and progression of cancer. DNA damage response (DDR) pathways lie at the crux of cell fate decisions following DNA damage induction, which can either trigger the repair of detrimental DNA lesions to protect cancer cells or induce the cell death machinery to eliminate damaged cells. Cytoskeletal dynamics have a critical role to play and influence the proper function of DDR pathways. Microfilaments, intermediate filaments, microtubules, and their associated proteins are well involved in the DDR. For instance, they are not only implicated in the recruitment of specific DDR molecules to the sites of DNA damage but also in the regulation of the mobility of the damaged DNA to repair sites in the periphery of the nucleus. The exquisite roles that these cytoskeletal proteins play in different DDR pathways, such as non-homologous end joining (NHEJ), homologous recombination (HR), base excision repair (BER), and nucleotide excision repair (NER), in cancer cells are extensively discussed in this review. Many cancer treatments are reliant upon inducing DNA damage in cancer cells to eliminate them; thus, it is important to shed light on factors that could affect their efficacy. Although the cytoskeleton is intricately involved in the DDR process, this has often been overlooked in cancer research and has not been exploited in developing DDR-targeting cancer therapy. Understanding the interplay between the cytoskeleton and the DDR in cancer will then provide insights into improving the development of cancer therapies that can leverage the synergistic action of DDR inhibitors and cytoskeleton-targeting agents.
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Affiliation(s)
- Clarissa Esmeralda Halim
- Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117593, Singapore; (C.E.H.); (S.D.); (K.C.C.)
- NUS Centre for Cancer Research (N2CR), Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117599, Singapore
| | - Shuo Deng
- Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117593, Singapore; (C.E.H.); (S.D.); (K.C.C.)
- NUS Centre for Cancer Research (N2CR), Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117599, Singapore
| | - Karen Carmelina Crasta
- Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117593, Singapore; (C.E.H.); (S.D.); (K.C.C.)
- Healthy Longevity Translational Research Programme, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117593, Singapore
| | - Celestial T. Yap
- Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117593, Singapore; (C.E.H.); (S.D.); (K.C.C.)
- NUS Centre for Cancer Research (N2CR), Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117599, Singapore
- National University Cancer Institute, National University Health System, Singapore 119074, Singapore
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Cao ML, Han RY, Chen SD, Zhao DY, Shi MY, Zou JH, Li L, Jiang HK. Gene Editing: An Effective Tool for the Future Treatment of Kidney Disease. J Inflamm Res 2025; 18:4001-4018. [PMID: 40125088 PMCID: PMC11927957 DOI: 10.2147/jir.s506760] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2024] [Accepted: 02/18/2025] [Indexed: 03/25/2025] Open
Abstract
Gene editing technology involves modifying target genes to alter genetic traits and generate new phenotypes. Beginning with zinc-finger nucleases (ZFN) and transcription activator-like effector nucleases (TALEN), the field has evolved through the advent of clustered regularly interspaced short palindromic repeats and CRISPR-associated protein (CRISPR-Cas) systems, and more recently to base editors (BE) and prime editors (PE). These innovations have provided deep insights into the molecular mechanisms of complex biological processes and have paved the way for novel therapeutic strategies for a range of diseases. Gene editing is now being applied in the treatment of both genetic and acquired kidney diseases, as well as in kidney transplantation and the correction of genetic mutations. This review explores the current applications of mainstream gene editing technologies in biology, with a particular emphasis on their roles in kidney disease research and treatment of. It also addresses the limitations and challenges associated with these technologies, while offering perspectives on their future potential in this field.
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Affiliation(s)
- Mei-Ling Cao
- Department of Neonatology, The First Hospital of China Medical University, Shenyang, Liaoning, 110001, People’s Republic of China
| | - Rui-Yi Han
- Department of Pediatrics, The First Hospital of China Medical University, Shenyang, Liaoning, 110001, People’s Republic of China
| | - Si-Da Chen
- Department of Orthopaedic Surgery, Shengjing Hospital of China Medical University, Shenyang, Liaoning, 110004, People’s Republic of China
| | - Dan-Yang Zhao
- Department of Pediatrics, The First Hospital of China Medical University, Shenyang, Liaoning, 110001, People’s Republic of China
| | - Ming-Yue Shi
- Department of Pediatrics, The First Hospital of China Medical University, Shenyang, Liaoning, 110001, People’s Republic of China
| | - Jia-Hui Zou
- Department of Pediatrics, The First Hospital of China Medical University, Shenyang, Liaoning, 110001, People’s Republic of China
| | - Lei Li
- Department of Orthopaedic Surgery, Shengjing Hospital of China Medical University, Shenyang, Liaoning, 110004, People’s Republic of China
| | - Hong-Kun Jiang
- Department of Pediatrics, The First Hospital of China Medical University, Shenyang, Liaoning, 110001, People’s Republic of China
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Balamurugan S, Li DW, Wang X, Li HY. Unleashing the potential of biotechnological strategies for the sustainable production of microalgal polysaccharides. Crit Rev Food Sci Nutr 2025:1-18. [PMID: 40084718 DOI: 10.1080/10408398.2025.2475240] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 03/16/2025]
Abstract
The prevailing trend toward the increased application of natural polysaccharides in the food, cosmetics, and pharmaceutical sectors has provided the impetus for exploring sustainable biological feedstocks. Amongst them, photoautotrophic microalgae have garnered huge research and commercial interests for polysaccharide production by photosynthesis, thereby concurrently attaining carbon sequestration and green production of valuable metabolites. However, conventional approaches for enhancing polysaccharide accumulation warrant adverse conditions, which in turn hinder cellular growth and productivity. Hence, there exists a pressing demand to harness biotechnological approaches for empowering photosynthetic algae as a sustainable feedstock for polysaccharide production. Meanwhile, it remains an untapped tool for the commercial production of microalgal products, despite the recent advancements in synthetic biology. In this review, we discuss the existing intricacies in polysaccharide biosynthetic circuits and propose crucial strategies to circumvent those techno-biological complexities. We also highlight the possible approaches to circumventing such limitations to successfully employ metabolic engineering for the large-scale production of microalgal polysaccharides. The technologically feasible directions for unleashing the biotechnological potential of microalgae as green cell factories are projected toward the sustainable biosynthesis of polysaccharides.
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Affiliation(s)
- Srinivasan Balamurugan
- Department of Biotechnology, School of Bioengineering, SRM Institute of Science and Technology, Kattankulathur, India
| | - Da-Wei Li
- Key Laboratory of Eutrophication and Red Tide Prevention of Guangdong Higher Education Institutes, Jinan University, Guangzhou, China
| | - Xiang Wang
- Key Laboratory of Eutrophication and Red Tide Prevention of Guangdong Higher Education Institutes, Jinan University, Guangzhou, China
| | - Hong-Ye Li
- Key Laboratory of Eutrophication and Red Tide Prevention of Guangdong Higher Education Institutes, Jinan University, Guangzhou, China
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Li P, Li JY, Ma YJ, Wang XW, Chen JP, Li YY. DNA Damaging Agents Induce RNA Structural and Transcriptional Changes for Genes Associated with Redox Homeostasis in Arabidopsis thaliana. PLANTS (BASEL, SWITZERLAND) 2025; 14:780. [PMID: 40094761 PMCID: PMC11901513 DOI: 10.3390/plants14050780] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/15/2025] [Revised: 02/25/2025] [Accepted: 02/26/2025] [Indexed: 03/19/2025]
Abstract
Living organisms are constantly exposed to various DNA damaging agents. While the mechanisms of DNA damage and DNA repair are well understood, the impact of these agents on RNA secondary structure and subsequent function remains elusive. In this study, we explore the effects of DNA damaging reagent methyl methanesulfonate (MMS) on arabidopsis gene expression and RNA secondary structure using the dimethyl sulfate (DMS) mutational profiling with sequencing (DMS-MaPseq) method. Our analyses reveal that changes in transcriptional levels and mRNA structure are key factors in response to DNA damaging agents. MMS treatment leads to the up-regulation of arabidopsis RBOHs (respiratory burst oxidase homologues) and alteration in the RNA secondary structure of GSTF9 and GSTF10, thereby enhancing mRNA translation efficiency. Redox homeostasis manipulated by RBOHs and GSTFs plays a crucial role in MMS-induced primary root growth inhibition. In conclusion, our findings shed light on the effects of DNA damaging agents on RNA structure and potential mRNA translation, which provide a new insight to understand the mechanism of DNA damage.
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Affiliation(s)
- Ping Li
- State Key Laboratory for Quality and Safety of Agro-Products, Key Laboratory of Biotechnology in Plant Protection of MARA, Zhejiang Key Laboratory of Green Plant Protection, Institute of Plant Virology, Ningbo University, Ningbo 315211, China
| | - Jiong-Yi Li
- State Key Laboratory for Quality and Safety of Agro-Products, Key Laboratory of Biotechnology in Plant Protection of MARA, Zhejiang Key Laboratory of Green Plant Protection, Institute of Plant Virology, Ningbo University, Ningbo 315211, China
| | - Yu-Jiao Ma
- State Key Laboratory for Quality and Safety of Agro-Products, Key Laboratory of Biotechnology in Plant Protection of MARA, Zhejiang Key Laboratory of Green Plant Protection, Institute of Plant Virology, Ningbo University, Ningbo 315211, China
| | - Xiao-Wei Wang
- State Key Laboratory for Quality and Safety of Agro-Products, Key Laboratory of Biotechnology in Plant Protection of MARA, Zhejiang Key Laboratory of Green Plant Protection, Institute of Plant Virology, Ningbo University, Ningbo 315211, China
- Ministry of Agriculture Key Lab of Molecular Biology of Crop Pathogens and Insects, Institute of Insect Sciences, Zhejiang University, Hangzhou 310058, China
| | - Jian-Ping Chen
- State Key Laboratory for Quality and Safety of Agro-Products, Key Laboratory of Biotechnology in Plant Protection of MARA, Zhejiang Key Laboratory of Green Plant Protection, Institute of Plant Virology, Ningbo University, Ningbo 315211, China
| | - Yi-Yuan Li
- State Key Laboratory for Quality and Safety of Agro-Products, Key Laboratory of Biotechnology in Plant Protection of MARA, Zhejiang Key Laboratory of Green Plant Protection, Institute of Plant Virology, Ningbo University, Ningbo 315211, China
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Sobral AF, Dinis-Oliveira RJ, Barbosa DJ. CRISPR-Cas technology in forensic investigations: Principles, applications, and ethical considerations. Forensic Sci Int Genet 2025; 74:103163. [PMID: 39437497 DOI: 10.1016/j.fsigen.2024.103163] [Citation(s) in RCA: 3] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/20/2024] [Revised: 10/08/2024] [Accepted: 10/09/2024] [Indexed: 10/25/2024]
Abstract
CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats and CRISPR-associated proteins) systems are adaptive immune systems originally present in bacteria, where they are essential to protect against external genetic elements, including viruses and plasmids. Taking advantage of this system, CRISPR-Cas-based technologies have emerged as incredible tools for precise genome editing, thus significantly advancing several research fields. Forensic sciences represent a multidisciplinary field that explores scientific methods to investigate and resolve legal issues, particularly criminal investigations and subject identification. Consequently, it plays a critical role in the justice system, providing scientific evidence to support judicial investigations. Although less explored, CRISPR-Cas-based methodologies demonstrate strong potential in the field of forensic sciences due to their high accuracy and sensitivity, including DNA profiling and identification, interpretation of crime scene investigations, detection of food contamination or fraud, and other aspects related to environmental forensics. However, using CRISPR-Cas-based methodologies in human samples raises several ethical issues and concerns regarding the potential misuse of individual genetic information. In this manuscript, we provide an overview of potential applications of CRISPR-Cas-based methodologies in several areas of forensic sciences and discuss the legal implications that challenge their routine implementation in this research field.
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Affiliation(s)
- Ana Filipa Sobral
- Associate Laboratory i4HB - Institute for Health and Bioeconomy, University Institute of Health Sciences - CESPU, Gandra 4585-116, Portugal; UCIBIO - Applied Molecular Biosciences Unit, Toxicologic Pathology Research Laboratory, University Institute of Health Sciences (1H-TOXRUN, IUCS-CESPU), Gandra 4585-116, Portugal.
| | - Ricardo Jorge Dinis-Oliveira
- Associate Laboratory i4HB - Institute for Health and Bioeconomy, University Institute of Health Sciences - CESPU, Gandra 4585-116, Portugal; UCIBIO - Applied Molecular Biosciences Unit, Translational Toxicology Research Laboratory, University Institute of Health Sciences (1H-TOXRUN, IUCS-CESPU), Gandra 4585-116, Portugal; Department of Public Health and Forensic Sciences and Medical Education, Faculty of Medicine, University of Porto, Porto 4200-319, Portugal; FOREN - Forensic Science Experts, Dr. Mário Moutinho Avenue, No. 33-A, Lisbon 1400-136, Portugal.
| | - Daniel José Barbosa
- Associate Laboratory i4HB - Institute for Health and Bioeconomy, University Institute of Health Sciences - CESPU, Gandra 4585-116, Portugal; UCIBIO - Applied Molecular Biosciences Unit, Translational Toxicology Research Laboratory, University Institute of Health Sciences (1H-TOXRUN, IUCS-CESPU), Gandra 4585-116, Portugal.
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9
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Tong R, Jing F, Li Y, Pan L, Yu X, Zhang N, Liao Q. Mechanisms of intestinal DNA damage and inflammation induced by ammonia nitrogen exposure in Litopenaeus vannamei. Comp Biochem Physiol C Toxicol Pharmacol 2025; 287:110070. [PMID: 39522856 DOI: 10.1016/j.cbpc.2024.110070] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/27/2024] [Revised: 10/11/2024] [Accepted: 11/07/2024] [Indexed: 11/16/2024]
Abstract
Ammonia nitrogen, a common aquaculture pollutant, harms crustaceans by causing intestinal inflammation, though its exact mechanisms are unclear. Thus, we exposed shrimp to 0, 2, 10 and 20 mg/L NH4Cl exposure for 0, 3, 6, 12, 24, 48, 72 h, and explored the intestinal stress, apoptosis, proliferation, inflammation and its histopathological changes. This research indicated that ammonia nitrogen exposure heightens plasma dopamine (DA), 5-hydroxytryptamine (5-HT), norepinephrine (NE), and acetylcholine (ACh) levels, alters gene expression of neurotransmitter receptors in the intestine, triggering the PLCCa2+ pathway and induces endoplasmic reticulum stress. Additionally, mitochondrial fission-related genes (Drp1, FIS1) significantly increase, the level of reactive oxygen species (ROS) was significantly elevated in the intestine, which induced DNA damage effects and initiated the DNA repair function, mainly through the base excision repair pathway, but with a low repair efficiency. By determining the expression of key genes of caspase-dependent and non-caspase-dependent apoptotic pathways, it was found that ammonia nitrogen exposure induced apoptosis in intestinal cells, proliferation key signaling pathways such as Wnt, EGFR and FOXO signaling showed an overall decrease after ammonia nitrogen exposure, combined with the gene expression of cell cycle proteins and proliferation markers, indicated that the proliferation of intestinal cells was inhibited. Performing pearson correlation analysis of intestinal cell damage, proliferation, and inflammatory factors, we hypothesized that ammonia nitrogen exposure induces intestinal endoplasmic reticulum stress and mitochondrial fission, induces elevated ROS, leads to DNA damage, and causes inflammation and damage in intestinal tissues by the underlying mechanism of promoting apoptosis and inhibiting proliferation.
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Affiliation(s)
- Ruixue Tong
- The Key Laboratory of Mariculture, Ministry of Education, Ocean University of China, Qingdao 266003, PR China
| | - Futao Jing
- Shandong Fisheries Development and Resources Conservation Center, Jinan 250013, China
| | - Yaobing Li
- The Key Laboratory of Mariculture, Ministry of Education, Ocean University of China, Qingdao 266003, PR China
| | - Luqing Pan
- The Key Laboratory of Mariculture, Ministry of Education, Ocean University of China, Qingdao 266003, PR China.
| | - Xin Yu
- The Key Laboratory of Mariculture, Ministry of Education, Ocean University of China, Qingdao 266003, PR China
| | - Ning Zhang
- The Key Laboratory of Mariculture, Ministry of Education, Ocean University of China, Qingdao 266003, PR China
| | - Qilong Liao
- The Key Laboratory of Mariculture, Ministry of Education, Ocean University of China, Qingdao 266003, PR China
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10
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Yu JZ, Kiss Z, Ma W, Liang R, Li T. Preclinical Models for Functional Precision Lung Cancer Research. Cancers (Basel) 2024; 17:22. [PMID: 39796653 PMCID: PMC11718887 DOI: 10.3390/cancers17010022] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2024] [Revised: 12/20/2024] [Accepted: 12/23/2024] [Indexed: 01/13/2025] Open
Abstract
Patient-centered precision oncology strives to deliver individualized cancer care. In lung cancer, preclinical models and technological innovations have become critical in advancing this approach. Preclinical models enable deeper insights into tumor biology and enhance the selection of appropriate systemic therapies across chemotherapy, targeted therapies, immunotherapies, antibody-drug conjugates, and emerging investigational treatments. While traditional human lung cancer cell lines offer a basic framework for cancer research, they often lack the tumor heterogeneity and intricate tumor-stromal interactions necessary to accurately predict patient-specific clinical outcomes. Patient-derived xenografts (PDXs), however, retain the original tumor's histopathology and genetic features, providing a more reliable model for predicting responses to systemic therapeutics, especially molecularly targeted therapies. For studying immunotherapies and antibody-drug conjugates, humanized PDX mouse models, syngeneic mouse models, and genetically engineered mouse models (GEMMs) are increasingly utilized. Despite their value, these in vivo models are costly, labor-intensive, and time-consuming. Recently, patient-derived lung cancer organoids (LCOs) have emerged as a promising in vitro tool for functional precision oncology studies. These LCOs demonstrate high success rates in growth and maintenance, accurately represent the histology and genomics of the original tumors and exhibit strong correlations with clinical treatment responses. Further supported by advancements in imaging, spatial and single-cell transcriptomics, proteomics, and artificial intelligence, these preclinical models are reshaping the landscape of drug development and functional precision lung cancer research. This integrated approach holds the potential to deliver increasingly accurate, personalized treatment strategies, ultimately enhancing patient outcomes in lung cancer.
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Affiliation(s)
- Jie-Zeng Yu
- Division of Hematology/Oncology, Department of Internal Medicine, University of California Davis School of Medicine, University of California Davis Comprehensive Cancer Center, Sacramento, CA 95817, USA; (J.-Z.Y.); (Z.K.); (W.M.); (R.L.)
| | - Zsofia Kiss
- Division of Hematology/Oncology, Department of Internal Medicine, University of California Davis School of Medicine, University of California Davis Comprehensive Cancer Center, Sacramento, CA 95817, USA; (J.-Z.Y.); (Z.K.); (W.M.); (R.L.)
| | - Weijie Ma
- Division of Hematology/Oncology, Department of Internal Medicine, University of California Davis School of Medicine, University of California Davis Comprehensive Cancer Center, Sacramento, CA 95817, USA; (J.-Z.Y.); (Z.K.); (W.M.); (R.L.)
- Department of Pathology and Laboratory Medicine, Dartmouth Hitchcock Medical Center, Geisel School of Medicine at Dartmouth, Lebanon, NH 03756, USA
| | - Ruqiang Liang
- Division of Hematology/Oncology, Department of Internal Medicine, University of California Davis School of Medicine, University of California Davis Comprehensive Cancer Center, Sacramento, CA 95817, USA; (J.-Z.Y.); (Z.K.); (W.M.); (R.L.)
| | - Tianhong Li
- Division of Hematology/Oncology, Department of Internal Medicine, University of California Davis School of Medicine, University of California Davis Comprehensive Cancer Center, Sacramento, CA 95817, USA; (J.-Z.Y.); (Z.K.); (W.M.); (R.L.)
- Medical Service, Hematology/Oncology, Veterans Affairs Northern California Health Care System, Mather, CA 10535, USA
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11
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Flores-Vega JJ, Puente-Rivera J, Sosa-Mondragón SI, Camacho-Nuez M, Alvarez-Sánchez ME. RAD51 recombinase and its paralogs: Orchestrating homologous recombination and unforeseen functions in protozoan parasites. Exp Parasitol 2024; 267:108847. [PMID: 39414114 DOI: 10.1016/j.exppara.2024.108847] [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: 05/20/2024] [Revised: 09/30/2024] [Accepted: 10/14/2024] [Indexed: 10/18/2024]
Abstract
The DNA of protozoan parasites is highly susceptible to damage, either induced by environmental agents or spontaneously generated during cellular metabolism through reactive oxygen species (ROS). Certain phases of the cell cycle, such as meiotic recombination, and external factors like ionizing radiation (IR), ultraviolet light (UV), or chemical genotoxic agents further increase this susceptibility. Among the various types of DNA damage, double-stranded breaks (DSBs) are the most critical, as they are challenging to repair and can result in genetic instability or cell death. DSBs caused by environmental stressors are primarily repaired via one of two major pathways: non-homologous end joining (NHEJ) or homologous recombination (HR). In multicellular eukaryotes, NHEJ predominates, but in unicellular eukaryotes such as protozoan parasites, HR seems to be the principal mechanism for DSB repair. The HR pathway is orchestrated by proteins from the RAD52 epistasis group, including RAD51, RAD52, RAD54, RAD55, and the MRN complex. This review focuses on elucidating the diverse roles and significance of RAD51 recombinase and its paralogs in protozoan parasites, such as Acanthamoeba castellanii, Entamoeba histolytica (Amoebozoa), apicomplexan parasites (Chromalveolata), Naegleria fowleri, Giardia spp., Trichomonas vaginalis, and trypanosomatids (Excavata), where they primarily function in HR. Additionally, we analyze the diversity of proteins involved in HR, both upstream and downstream of RAD51, and discuss the implications of these processes in parasitic protozoa.
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Affiliation(s)
- Jose Jesús Flores-Vega
- Posgrado en Ciencias Genómicas, Universidad Autónoma de la Ciudad de México (UACM), San Lorenzo #290, Col. Del Valle, CP 03100, Mexico City, Mexico
| | - Jonathan Puente-Rivera
- Posgrado en Ciencias Genómicas, Universidad Autónoma de la Ciudad de México (UACM), San Lorenzo #290, Col. Del Valle, CP 03100, Mexico City, Mexico; División de Investigación. Hospital Juárez de México, Ciudad de México, 07760, Mexico.
| | - Sharon Itzel Sosa-Mondragón
- Posgrado en Ciencias Genómicas, Universidad Autónoma de la Ciudad de México (UACM), San Lorenzo #290, Col. Del Valle, CP 03100, Mexico City, Mexico
| | - Minerva Camacho-Nuez
- Posgrado en Ciencias Genómicas, Universidad Autónoma de la Ciudad de México (UACM), San Lorenzo #290, Col. Del Valle, CP 03100, Mexico City, Mexico
| | - María Elizbeth Alvarez-Sánchez
- Posgrado en Ciencias Genómicas, Universidad Autónoma de la Ciudad de México (UACM), San Lorenzo #290, Col. Del Valle, CP 03100, Mexico City, Mexico.
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12
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Zhu W, Pan S, Zhang J, Xu J, Zhang R, Zhang Y, Fu Z, Wang Y, Hu C, Xu Z. The role of hyperthermia in the treatment of tumor. Crit Rev Oncol Hematol 2024; 204:104541. [PMID: 39461607 DOI: 10.1016/j.critrevonc.2024.104541] [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: 06/02/2024] [Revised: 09/19/2024] [Accepted: 10/17/2024] [Indexed: 10/29/2024] Open
Abstract
Despite recent advancements in the diagnosis and treatment options for cancer, it remains one of the most serious threats to health. Hyperthermia (HT) has emerged as a highly promising area of research due to its safety and cost-effectiveness. Currently, based on temperature, HT can be categorized into thermal ablation and mild hyperthermia. Thermal ablation involves raising the temperature within the tumor to over 60°C, resulting in direct necrosis in the central region of the tumor. In contrast, mild hyperthermia operates at relatively lower temperatures, typically in the range of 41-45°C, to induce damage to tumor cells. Furthermore, HT also serves as an immune adjuvant strategy in radiotherapy, chemotherapy, and immunotherapy, enhancing the effectiveness of radiotherapy, increasing the uptake of chemotherapy drugs, and reprogramming the tumor microenvironment through the induction of immunogenic cell death, thereby promoting the recruitment of endogenous immune cells. This article reviews the current status and development of hyperthermia, outlines potential mechanisms by which hyperthermia inhibits tumors, describes clinical trial attempts combining hyperthermia with radiotherapy, chemotherapy, and immunotherapy, and discusses the relationship between nanoparticles and hyperthermia.
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Affiliation(s)
- Weiwei Zhu
- Second Clinical Medical College, Zhejiang Chinese Medical University, Hangzhou 310053, China; Department of Gastric surgery, The Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital), Institutes of Basic Medicine and Cancer (IBMC), Chinese Academy of Sciences, Hangzhou 310022, China
| | - Siwei Pan
- Department of Gastric surgery, The Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital), Institutes of Basic Medicine and Cancer (IBMC), Chinese Academy of Sciences, Hangzhou 310022, China; Key Laboratory of Prevention, Diagnosis and Therapy of Upper Gastrointestinal Cancer of Zhejiang Province, Hangzhou 310022, China; Zhejiang Provincial Research Center for Upper Gastrointestinal Tract Cancer, Zhejiang Cancer Hospital, Hangzhou 310022, China
| | - Jiaqing Zhang
- Second Clinical Medical College, Zhejiang Chinese Medical University, Hangzhou 310053, China; Department of Gastric surgery, The Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital), Institutes of Basic Medicine and Cancer (IBMC), Chinese Academy of Sciences, Hangzhou 310022, China
| | - Jingli Xu
- Second Clinical Medical College, Zhejiang Chinese Medical University, Hangzhou 310053, China; Department of Gastric surgery, The Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital), Institutes of Basic Medicine and Cancer (IBMC), Chinese Academy of Sciences, Hangzhou 310022, China
| | - Ruolan Zhang
- Second Clinical Medical College, Zhejiang Chinese Medical University, Hangzhou 310053, China; Department of Gastric surgery, The Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital), Institutes of Basic Medicine and Cancer (IBMC), Chinese Academy of Sciences, Hangzhou 310022, China
| | - Yanqiang Zhang
- Department of Gastric surgery, The Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital), Institutes of Basic Medicine and Cancer (IBMC), Chinese Academy of Sciences, Hangzhou 310022, China; Key Laboratory of Prevention, Diagnosis and Therapy of Upper Gastrointestinal Cancer of Zhejiang Province, Hangzhou 310022, China; Zhejiang Provincial Research Center for Upper Gastrointestinal Tract Cancer, Zhejiang Cancer Hospital, Hangzhou 310022, China
| | - Zhenjie Fu
- Department of Gastric surgery, The Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital), Institutes of Basic Medicine and Cancer (IBMC), Chinese Academy of Sciences, Hangzhou 310022, China; Key Laboratory of Prevention, Diagnosis and Therapy of Upper Gastrointestinal Cancer of Zhejiang Province, Hangzhou 310022, China; Zhejiang Provincial Research Center for Upper Gastrointestinal Tract Cancer, Zhejiang Cancer Hospital, Hangzhou 310022, China
| | - Yuqi Wang
- Department of Gastric surgery, The Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital), Institutes of Basic Medicine and Cancer (IBMC), Chinese Academy of Sciences, Hangzhou 310022, China; Key Laboratory of Prevention, Diagnosis and Therapy of Upper Gastrointestinal Cancer of Zhejiang Province, Hangzhou 310022, China; Zhejiang Provincial Research Center for Upper Gastrointestinal Tract Cancer, Zhejiang Cancer Hospital, Hangzhou 310022, China
| | - Can Hu
- Department of Gastric surgery, The Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital), Institutes of Basic Medicine and Cancer (IBMC), Chinese Academy of Sciences, Hangzhou 310022, China; Key Laboratory of Prevention, Diagnosis and Therapy of Upper Gastrointestinal Cancer of Zhejiang Province, Hangzhou 310022, China; Zhejiang Provincial Research Center for Upper Gastrointestinal Tract Cancer, Zhejiang Cancer Hospital, Hangzhou 310022, China.
| | - Zhiyuan Xu
- Department of Gastric surgery, The Cancer Hospital of the University of Chinese Academy of Sciences (Zhejiang Cancer Hospital), Institutes of Basic Medicine and Cancer (IBMC), Chinese Academy of Sciences, Hangzhou 310022, China; Key Laboratory of Prevention, Diagnosis and Therapy of Upper Gastrointestinal Cancer of Zhejiang Province, Hangzhou 310022, China; Zhejiang Provincial Research Center for Upper Gastrointestinal Tract Cancer, Zhejiang Cancer Hospital, Hangzhou 310022, China.
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13
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Kumar S, Singh A, Bist CMS, Sharma M. Advancements in genetic techniques and functional genomics for enhancing crop traits and agricultural sustainability. Brief Funct Genomics 2024; 23:607-623. [PMID: 38679487 DOI: 10.1093/bfgp/elae017] [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: 02/08/2024] [Revised: 04/03/2024] [Accepted: 04/16/2024] [Indexed: 05/01/2024] Open
Abstract
Genetic variability is essential for the development of new crop varieties with economically beneficial traits. The traits can be inherited from wild relatives or induced through mutagenesis. Novel genetic elements can then be identified and new gene functions can be predicted. In this study, forward and reverse genetics approaches were described, in addition to their applications in modern crop improvement programs and functional genomics. By using heritable phenotypes and linked genetic markers, forward genetics searches for genes by using traditional genetic mapping and allele frequency estimation. Despite recent advances in sequencing technology, omics and computation, genetic redundancy remains a major challenge in forward genetics. By analyzing close-related genes, we will be able to dissect their functional redundancy and predict possible traits and gene activity patterns. In addition to these predictions, sophisticated reverse gene editing tools can be used to verify them, including TILLING, targeted insertional mutagenesis, gene silencing, gene targeting and genome editing. By using gene knock-down, knock-up and knock-out strategies, these tools are able to detect genetic changes in cells. In addition, epigenome analysis and editing enable the development of novel traits in existing crop cultivars without affecting their genetic makeup by increasing epiallelic variants. Our understanding of gene functions and molecular dynamics of various biological phenomena has been revised by all of these findings. The study also identifies novel genetic targets in crop species to improve yields and stress tolerances through conventional and non-conventional methods. In this article, genetic techniques and functional genomics are specifically discussed and assessed for their potential in crop improvement.
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Affiliation(s)
- Surender Kumar
- Department of Biotechnology, College of Horticulture, Dr. Y. S. Parmar University of Horticulture and Forestry, Nauni, Solan-173230, Himachal Pradesh, India
| | - Anupama Singh
- Department of Biotechnology, College of Horticulture, Dr. Y. S. Parmar University of Horticulture and Forestry, Nauni, Solan-173230, Himachal Pradesh, India
| | - Chander Mohan Singh Bist
- Indian Council of Agricultural Research (ICAR)-Central Potato Research Institute, Shimla-171001, Himachal Pradesh, India
| | - Munish Sharma
- Department of Plant Sciences, Central University of Himachal Pradesh, Dharamshala-176215, Himachal Pradesh, India
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14
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van de Kamp G, Heemskerk T, Kanaar R, Essers J. Synergistic Roles of Non-Homologous End Joining and Homologous Recombination in Repair of Ionizing Radiation-Induced DNA Double Strand Breaks in Mouse Embryonic Stem Cells. Cells 2024; 13:1462. [PMID: 39273031 PMCID: PMC11393957 DOI: 10.3390/cells13171462] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2024] [Revised: 08/23/2024] [Accepted: 08/27/2024] [Indexed: 09/15/2024] Open
Abstract
DNA double strand breaks (DSBs) are critical for the efficacy of radiotherapy as they lead to cell death if not repaired. DSBs caused by ionizing radiation (IR) initiate histone modifications and accumulate DNA repair proteins, including 53BP1, which forms distinct foci at damage sites and serves as a marker for DSBs. DSB repair primarily occurs through Non-Homologous End Joining (NHEJ) and Homologous Recombination (HR). NHEJ directly ligates DNA ends, employing proteins such as DNA-PKcs, while HR, involving proteins such as Rad54, uses a sister chromatid template for accurate repair and functions in the S and G2 phases of the cell cycle. Both pathways are crucial, as illustrated by the IR sensitivity in cells lacking DNA-PKcs or Rad54. We generated mouse embryonic stem (mES) cells which are knockout (KO) for DNA-PKcs and Rad54 to explore the combined role of HR and NHEJ in DSB repair. We found that cells lacking both DNA-PKcs and Rad54 are hypersensitive to X-ray radiation, coinciding with impaired 53BP1 focus resolution and a more persistent G2 phase cell cycle block. Additionally, mES cells deficient in DNA-PKcs or both DNA-PKcs and Rad54 exhibit an increased nuclear size approximately 18-24 h post-irradiation. To further explore the role of Rad54 in the absence of DNA-PKcs, we generated DNA-PKcs KO mES cells expressing GFP-tagged wild-type (WT) or ATPase-defective Rad54 to track the Rad54 foci over time post-irradiation. Cells lacking DNA-PKcs and expressing ATPase-defective Rad54 exhibited a similar phenotypic response to IR as those lacking both DNA-PKcs and Rad54. Despite a strong G2 phase arrest, live-cell imaging showed these cells eventually progress through mitosis, forming micronuclei. Additionally, mES cells lacking DNA-PKcs showed increased Rad54 foci over time post-irradiation, indicating an enhanced reliance on HR for DSB repair without DNA-PKcs. Our findings underscore the essential roles of HR and NHEJ in maintaining genomic stability post-IR in mES cells. The interplay between these pathways is crucial for effective DSB repair and cell cycle progression, highlighting potential targets for enhancing radiotherapy outcomes.
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Affiliation(s)
- Gerarda van de Kamp
- Department of Molecular Genetics, Erasmus MC Cancer Institute, Erasmus University Medical Center, 3015 GD Rotterdam, The Netherlands
- Oncode Institute, Erasmus University Medical Center, 3015 GD Rotterdam, The Netherlands
| | - Tim Heemskerk
- Department of Molecular Genetics, Erasmus MC Cancer Institute, Erasmus University Medical Center, 3015 GD Rotterdam, The Netherlands
- Oncode Institute, Erasmus University Medical Center, 3015 GD Rotterdam, The Netherlands
| | - Roland Kanaar
- Department of Molecular Genetics, Erasmus MC Cancer Institute, Erasmus University Medical Center, 3015 GD Rotterdam, The Netherlands
- Oncode Institute, Erasmus University Medical Center, 3015 GD Rotterdam, The Netherlands
| | - Jeroen Essers
- Oncode Institute, Erasmus University Medical Center, 3015 GD Rotterdam, The Netherlands
- Department of Vascular Surgery, Erasmus University Medical Center, 3015 GD Rotterdam, The Netherlands
- Department of Radiotherapy, Erasmus MC Cancer Institute, Erasmus University Medical Center, 3015 GD Rotterdam, The Netherlands
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15
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Adegbaju MS, Ajose T, Adegbaju IE, Omosebi T, Ajenifujah-Solebo SO, Falana OY, Shittu OB, Adetunji CO, Akinbo O. Genetic engineering and genome editing technologies as catalyst for Africa's food security: the case of plant biotechnology in Nigeria. Front Genome Ed 2024; 6:1398813. [PMID: 39045572 PMCID: PMC11263695 DOI: 10.3389/fgeed.2024.1398813] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/10/2024] [Accepted: 05/15/2024] [Indexed: 07/25/2024] Open
Abstract
Many African countries are unable to meet the food demands of their growing population and the situation is worsened by climate change and disease outbreaks. This issue of food insecurity may lead to a crisis of epic proportion if effective measures are not in place to make more food available. Thus, deploying biotechnology towards the improvement of existing crop varieties for tolerance or resistance to both biotic and abiotic stresses is crucial to increasing crop production. In order to optimize crop production, several African countries have implemented strategies to make the most of this innovative technology. For example, Nigerian government has implemented the National Biotechnology Policy to facilitate capacity building, research, bioresource development and commercialization of biotechnology products for over two decades. Several government ministries, research centers, universities, and agencies have worked together to implement the policy, resulting in the release of some genetically modified crops to farmers for cultivation and Commercialization, which is a significant accomplishment. However, the transgenic crops were only brought to Nigeria for confined field trials; the manufacturing of the transgenic crops took place outside the country. This may have contributed to the suspicion of pressure groups and embolden proponents of biotechnology as an alien technology. Likewise, this may also be the underlying issue preventing the adoption of biotechnology products in other African countries. It is therefore necessary that African universities develop capacity in various aspects of biotechnology, to continuously train indigenous scientists who can generate innovative ideas tailored towards solving problems that are peculiar to respective country. Therefore, this study intends to establish the role of genetic engineering and genome editing towards the achievement of food security in Africa while using Nigeria as a case study. In our opinion, biotechnology approaches will not only complement conventional breeding methods in the pursuit of crop improvements, but it remains a viable and sustainable means of tackling specific issues hindering optimal crop production. Furthermore, we suggest that financial institutions should offer low-interest loans to new businesses. In order to promote the growth of biotechnology products, especially through the creation of jobs and revenues through molecular farming.
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Affiliation(s)
- Muyiwa Seyi Adegbaju
- Department of Crop, Soil and Pest Management, Federal University of Technology Akure, Akure, Ondo, Nigeria
| | - Titilayo Ajose
- Fruits and Spices Department, National Horticultural Institute, Ibadan, Oyo, Nigeria
| | | | - Temitayo Omosebi
- Department of Agricultural Technology, Federal College of Forestry, Jos, Nigeria
| | | | - Olaitan Yetunde Falana
- Department of Genetics, Genomic and Bioinformatics, National Biotechnology Research and Development Agency, Abuja, Nigeria
| | - Olufunke Bolatito Shittu
- Department of Microbiology, College of Biosciences, Federal University of Agriculture, Abeokuta, Nigeria
| | | | - Olalekan Akinbo
- African Union Development Agency-NEPAD, Office of Science, Technology and Innovation, Midrand, South Africa
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16
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Yao H, Zhang M, Wang D. The next decade of SET: from an oncoprotein to beyond. J Mol Cell Biol 2024; 16:mjad082. [PMID: 38157418 PMCID: PMC11267991 DOI: 10.1093/jmcb/mjad082] [Citation(s) in RCA: 3] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/20/2023] [Revised: 11/22/2023] [Accepted: 12/24/2023] [Indexed: 01/03/2024] Open
Abstract
This year marks the fourth decade of research into the protein SET, which was discovered in 1992. SET was initially identified as an oncoprotein but later shown to be a multifaceted protein involved in regulating numerous biological processes under both physiological and pathophysiological conditions. SET dysfunction is closely associated with diseases, such as cancer and Alzheimer's disease. With the increasing understanding of how SET works and how it is regulated in cells, targeting aberrant SET has emerged as a potential strategy for disease intervention. In this review, we present a comprehensive overview of the advancements in SET studies, encompassing its biological functions, regulatory networks, clinical implications, and pharmacological inhibitors. Furthermore, we provide insights into the future prospects of SET research, with a particular emphasis on its promising potential in the realm of immune modulation.
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Affiliation(s)
- Han Yao
- State Key Laboratory of Common Mechanism Research for Major Diseases & Department of Medical Genetics, Institute of Basic Medical Sciences & School of Basic Medicine, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100005, China
| | - Meng Zhang
- State Key Laboratory of Common Mechanism Research for Major Diseases & Department of Medical Genetics, Institute of Basic Medical Sciences & School of Basic Medicine, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100005, China
| | - Donglai Wang
- State Key Laboratory of Common Mechanism Research for Major Diseases & Department of Medical Genetics, Institute of Basic Medical Sciences & School of Basic Medicine, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100005, China
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Ng YB, Akincilar SC. Shaping DNA damage responses: Therapeutic potential of targeting telomeric proteins and DNA repair factors in cancer. Curr Opin Pharmacol 2024; 76:102460. [PMID: 38776747 DOI: 10.1016/j.coph.2024.102460] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/10/2022] [Revised: 10/06/2023] [Accepted: 10/11/2023] [Indexed: 05/25/2024]
Abstract
Shelterin proteins regulate genomic stability by preventing inappropriate DNA damage responses (DDRs) at telomeres. Unprotected telomeres lead to persistent DDR causing cell cycle inhibition, growth arrest, and apoptosis. Cancer cells rely on DDR to protect themselves from DNA lesions and exogenous DNA-damaging agents such as chemotherapy and radiotherapy. Therefore, targeting DDR machinery is a promising strategy to increase the sensitivity of cancer cells to existing cancer therapies. However, the success of these DDR inhibitors depends on other mutations, and over time, patients develop resistance to these therapies. This suggests the need for alternative approaches. One promising strategy is co-inhibiting shelterin proteins with DDR molecules, which would offset cellular fitness in DNA repair in a mutation-independent manner. This review highlights the associations and dependencies of the shelterin complex with the DDR proteins and discusses potential co-inhibition strategies that might improve the therapeutic potential of current inhibitors.
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Affiliation(s)
- Yu Bin Ng
- Laboratory of NFκB Signalling, Institute of Molecular and Cell Biology (IMCB), Agency for Science, Technology and Research (A∗STAR), 61 Biopolis Drive, Proteos, Singapore 138673, Republic of Singapore
| | - Semih Can Akincilar
- Laboratory of NFκB Signalling, Institute of Molecular and Cell Biology (IMCB), Agency for Science, Technology and Research (A∗STAR), 61 Biopolis Drive, Proteos, Singapore 138673, Republic of Singapore.
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Naik B, Kumar V, Rizwanuddin S, Mishra S, Kumar V, Saris PEJ, Khanduri N, Kumar A, Pandey P, Gupta AK, Khan JM, Rustagi S. Biofortification as a solution for addressing nutrient deficiencies and malnutrition. Heliyon 2024; 10:e30595. [PMID: 38726166 PMCID: PMC11079288 DOI: 10.1016/j.heliyon.2024.e30595] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/16/2023] [Revised: 04/30/2024] [Accepted: 04/30/2024] [Indexed: 05/12/2024] Open
Abstract
Malnutrition, defined as both undernutrition and overnutrition, is a major global health concern affecting millions of people. One possible way to address nutrient deficiency and combat malnutrition is through biofortification. A comprehensive review of the literature was conducted to explore the current state of biofortification research, including techniques, applications, effectiveness and challenges. Biofortification is a promising strategy for enhancing the nutritional condition of at-risk populations. Biofortified varieties of basic crops, including rice, wheat, maize and beans, with elevated amounts of vital micronutrients, such as iron, zinc, vitamin A and vitamin C, have been successfully developed using conventional and advanced technologies. Additionally, the ability to specifically modify crop genomes to improve their nutritional profiles has been made possible by recent developments in genetic engineering, such as CRISPR-Cas9 technology. The health conditions of people have been shown to improve and nutrient deficiencies were reduced when biofortified crops were grown. Particularly in environments with limited resources, biofortification showed considerable promise as a long-term and economical solution to nutrient shortages and malnutrition. To fully exploit the potential of biofortified crops to enhance public health and global nutrition, issues such as consumer acceptance, regulatory permitting and production and distribution scaling up need to be resolved. Collaboration among governments, researchers, non-governmental organizations and the private sector is essential to overcome these challenges and promote the widespread adoption of biofortification as a key part of global food security and nutrition strategies.
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Affiliation(s)
- Bindu Naik
- Department of Food Science and Technology, Graphic Era (Deemed to Be) University, Bell Road, Clement Town, Dehradun, 248002, Uttarakhand, India
- School of Agriculture, Graphic Hill University, Clement Town, Dehradun, Uttarakhand, India
| | - Vijay Kumar
- Himalayan School of Biosciences, Swami Rama Himalayan University, Swami Rama Nagar, Jolly Grant, Dehradun, 248016, Uttarakhand, India
| | - Sheikh Rizwanuddin
- Department of Food Science and Technology, Graphic Era (Deemed to Be) University, Bell Road, Clement Town, Dehradun, 248002, Uttarakhand, India
| | - Sadhna Mishra
- Faculty of Agricultural Sciences, GLA University, Mathura, India
| | - Vivek Kumar
- Himalayan School of Biosciences, Swami Rama Himalayan University, Swami Rama Nagar, Jolly Grant, Dehradun, 248016, Uttarakhand, India
| | - Per Erik Joakim Saris
- Department of Microbiology, Faculty of Agriculture and Forestry, University of Helsinki, 00100, Helsinki, Finland
| | - Naresh Khanduri
- Himalayan School of Biosciences, Swami Rama Himalayan University, Swami Rama Nagar, Jolly Grant, Dehradun, 248016, Uttarakhand, India
| | - Akhilesh Kumar
- Himalayan School of Biosciences, Swami Rama Himalayan University, Swami Rama Nagar, Jolly Grant, Dehradun, 248016, Uttarakhand, India
| | - Piyush Pandey
- Soil and Environment Microbiology Laboratory, Department of Microbiology, Assam University, Silchur, 788011, Assam, India
| | - Arun Kumar Gupta
- Department of Food Science and Technology, Graphic Era (Deemed to Be) University, Bell Road, Clement Town, Dehradun, 248002, Uttarakhand, India
| | - Javed Masood Khan
- Department of Food Science and Nutrition, Faculty of Food and Agricultural Sciences, King Saud University, 2460, Riyadh, 11451, Saudi Arabia
| | - Sarvesh Rustagi
- Department of Food Technology, Uttaranchal University, Dehradun, 248007, Uttarakhand, India
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Negroni YL, Doro I, Tamborrino A, Luzzi I, Fortunato S, Hensel G, Khosravi S, Maretto L, Stevanato P, Lo Schiavo F, de Pinto MC, Krupinska K, Zottini M. The Arabidopsis Mitochondrial Nucleoid-Associated Protein WHIRLY2 Is Required for a Proper Response to Salt Stress. PLANT & CELL PHYSIOLOGY 2024; 65:576-589. [PMID: 38591870 PMCID: PMC11094760 DOI: 10.1093/pcp/pcae025] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/29/2023] [Revised: 03/04/2024] [Accepted: 03/07/2024] [Indexed: 04/10/2024]
Abstract
In the last years, plant organelles have emerged as central coordinators of responses to internal and external stimuli, which can induce stress. Mitochondria play a fundamental role as stress sensors being part of a complex communication network between the organelles and the nucleus. Among the different environmental stresses, salt stress poses a significant challenge and requires efficient signaling and protective mechanisms. By using the why2 T-DNA insertion mutant and a novel knock-out mutant prepared by CRISPR/Cas9-mediated genome editing, this study revealed that WHIRLY2 is crucial for protecting mitochondrial DNA (mtDNA) integrity during salt stress. Loss-of-function mutants show an enhanced sensitivity to salt stress. The disruption of WHIRLY2 causes the impairment of mtDNA repair that results in the accumulation of aberrant recombination products, coinciding with severe alterations in nucleoid integrity and overall mitochondria morphology besides a compromised redox-dependent response and misregulation of antioxidant enzymes. The results of this study revealed that WHIRLY2-mediated structural features in mitochondria (nucleoid compactness and cristae) are important for an effective response to salt stress.
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Affiliation(s)
- Yuri L Negroni
- Department of Biology, University of Padova, Via U. Bassi 58/b, Padova 35131, Italy
| | - Irene Doro
- Department of Biology, University of Padova, Via U. Bassi 58/b, Padova 35131, Italy
| | - Alberto Tamborrino
- Department of Biology, University of Padova, Via U. Bassi 58/b, Padova 35131, Italy
| | - Irene Luzzi
- Department of Biology, University of Padova, Via U. Bassi 58/b, Padova 35131, Italy
| | - Stefania Fortunato
- Department of Biosciences, Biotechnology and Environment, University of Bari, Campus Universitario, Via Orabona, 4, Bari 70125, Italy
| | - Götz Hensel
- Plant Reproductive Biology, Department of Physiology and Cell Biology, IPK, Corrensstraße 3, Seeland, Gatersleben D-06466, Germany
| | - Solmaz Khosravi
- Plant Reproductive Biology, Department of Physiology and Cell Biology, IPK, Corrensstraße 3, Seeland, Gatersleben D-06466, Germany
| | - Laura Maretto
- Department of Agronomy, Food, Natural Resources, Animal and Environment, University of Padova, Viale Università 16, Legnaro, Padova 35020, Italy
| | - Piergiorgio Stevanato
- Department of Agronomy, Food, Natural Resources, Animal and Environment, University of Padova, Viale Università 16, Legnaro, Padova 35020, Italy
| | - Fiorella Lo Schiavo
- Department of Biology, University of Padova, Via U. Bassi 58/b, Padova 35131, Italy
| | - Maria Concetta de Pinto
- Department of Biosciences, Biotechnology and Environment, University of Bari, Campus Universitario, Via Orabona, 4, Bari 70125, Italy
| | - Karin Krupinska
- Botanisches Institut, Christian-Albrechts-Universität zu Kiel, Am Botanischen Garten 1-9, Kiel D-24098, Germany
| | - Michela Zottini
- Department of Biology, University of Padova, Via U. Bassi 58/b, Padova 35131, Italy
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20
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Fallah J, Xu J, Weinstock C, Gao X, Heiss BL, Maguire WF, Chang E, Agrawal S, Tang S, Amiri-Kordestani L, Pazdur R, Kluetz PG, Suzman DL. Efficacy of Poly(ADP-ribose) Polymerase Inhibitors by Individual Genes in Homologous Recombination Repair Gene-Mutated Metastatic Castration-Resistant Prostate Cancer: A US Food and Drug Administration Pooled Analysis. J Clin Oncol 2024; 42:1687-1698. [PMID: 38484203 PMCID: PMC11095872 DOI: 10.1200/jco.23.02105] [Citation(s) in RCA: 39] [Impact Index Per Article: 39.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2023] [Revised: 11/29/2023] [Accepted: 12/20/2023] [Indexed: 05/09/2024] Open
Abstract
PURPOSE We performed a pooled analysis of multiple trials of poly(ADP-ribose) polymerase inhibitors (PARPi) in metastatic castration-resistant prostate cancer (mCRPC) to investigate the efficacy of PARPi in each individual homologous recombination repair (HRR) mutated (m) gene. PATIENTS AND METHODS We pooled patient-level data from trials of PARPi in mCRPC that reported mutation status in individual HRR genes. Any HRR gene with available data across all the randomized trials of PARPi in first-line mCRPC was selected. The hazard ratios (HRs; 95% CI) for radiographic progression-free survival (rPFS; by blinded independent review) and overall survival (OS) of a PARPi plus an androgen receptor pathway inhibitor (ARPI) relative to placebo plus an ARPI in the pool of three randomized trials in first-line mCRPC were calculated using Kaplan-Meier estimates and a Cox proportional hazards model. RESULTS In ATMm (N = 268), rPFS HR was 1.05 (0.74 to 1.49) and OS HR was 1.18 (0.82 to 1.71). In BRCA1m (N = 64), rPFS HR was 0.51 (0.23 to 1.1) and OS HR was 0.74 (0.34 to 1.61). In BRCA2m (N = 422), rPFS HR was 0.31 (0.23 to 0.42) and OS HR was 0.66 (0.49 to 0.89). In CDK12m (N = 164), rPFS HR was 0.50 (0.32 to 0.80) and OS HR was 0.63 (0.39 to 0.99). In CHEK2m (N = 172), rPFS HR was 1.06 (0.67 to 1.66) and OS HR was 1.53 (0.95 to 2.46). In PALB2m (N = 41) rPFS HR was 0.52 (0.23 to 1.17) and OS HR was 0.78 (0.34 to 1.8). CONCLUSION In this pooled analysis, benefit from PARPi appeared greatest for patients with BRCA1m, BRCA2m, CDK12m, and PALB2m. Given limitations of this exploratory analysis, the apparent lack of benefit from PARPi in patients with CHEK2m or ATMm should be further explored in future clinical trials.
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Affiliation(s)
- Jaleh Fallah
- Center for Drug Evaluation and Research (CDER), U.S. Food and Drug Administration, Silver Spring, MD
| | - Jianjin Xu
- Center for Drug Evaluation and Research (CDER), U.S. Food and Drug Administration, Silver Spring, MD
| | - Chana Weinstock
- Center for Drug Evaluation and Research (CDER), U.S. Food and Drug Administration, Silver Spring, MD
| | - Xin Gao
- Center for Drug Evaluation and Research (CDER), U.S. Food and Drug Administration, Silver Spring, MD
| | - Brian L. Heiss
- Center for Drug Evaluation and Research (CDER), U.S. Food and Drug Administration, Silver Spring, MD
| | - William F. Maguire
- Center for Drug Evaluation and Research (CDER), U.S. Food and Drug Administration, Silver Spring, MD
| | - Elaine Chang
- Center for Drug Evaluation and Research (CDER), U.S. Food and Drug Administration, Silver Spring, MD
| | - Sundeep Agrawal
- Center for Drug Evaluation and Research (CDER), U.S. Food and Drug Administration, Silver Spring, MD
| | - Shenghui Tang
- Center for Drug Evaluation and Research (CDER), U.S. Food and Drug Administration, Silver Spring, MD
| | - Laleh Amiri-Kordestani
- Center for Drug Evaluation and Research (CDER), U.S. Food and Drug Administration, Silver Spring, MD
- Oncology Center of Excellence (OCE), U.S. Food and Drug Administration, Silver Spring, MD
| | - Richard Pazdur
- Center for Drug Evaluation and Research (CDER), U.S. Food and Drug Administration, Silver Spring, MD
- Oncology Center of Excellence (OCE), U.S. Food and Drug Administration, Silver Spring, MD
| | - Paul G. Kluetz
- Center for Drug Evaluation and Research (CDER), U.S. Food and Drug Administration, Silver Spring, MD
- Oncology Center of Excellence (OCE), U.S. Food and Drug Administration, Silver Spring, MD
| | - Daniel L. Suzman
- Center for Drug Evaluation and Research (CDER), U.S. Food and Drug Administration, Silver Spring, MD
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21
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Mohamad Zamberi NN, Abuhamad AY, Low TY, Mohtar MA, Syafruddin SE. dCas9 Tells Tales: Probing Gene Function and Transcription Regulation in Cancer. CRISPR J 2024; 7:73-87. [PMID: 38635328 DOI: 10.1089/crispr.2023.0078] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 04/20/2024] Open
Abstract
Clustered regularly interspaced short palindromic repeats (CRISPR)-based genome editing is evolving into an essential tool in the field of biological and medical research. Notably, the development of catalytically deactivated Cas9 (dCas9) enzyme has substantially broadened its traditional boundaries in gene editing or perturbation. The conjugation of dCas9 with various molecular effectors allows precise control over transcriptional processes, epigenetic modifications, visualization of chromosomal dynamics, and several other applications. This expanded repertoire of CRISPR-Cas9 applications has emerged as an invaluable molecular tool kit that empowers researchers to comprehensively interrogate and gain insights into health and diseases. This review delves into the advancements in Cas9 protein engineering, specifically on the generation of various dCas9 tools that have significantly enhanced the CRISPR-based technology capability and versatility. We subsequently discuss the multifaceted applications of dCas9, especially in interrogating the regulation and function of genes that involve in supporting cancer pathogenesis. In addition, we also delineate the designing and utilization of dCas9-based tools as well as highlighting its current constraints and transformative potentials in cancer research.
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Affiliation(s)
- Nurul Nadia Mohamad Zamberi
- UKM Medical Molecular Biology Institute, Universiti Kebangsaan Malaysia, Cheras, Malaysia, Department of Biochemistry, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, Serdang, Malaysia
| | - Asmaa Y Abuhamad
- Bionanotechnology Research Group, Department of Biochemistry, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, Serdang, Malaysia
| | - Teck Yew Low
- UKM Medical Molecular Biology Institute, Universiti Kebangsaan Malaysia, Cheras, Malaysia, Department of Biochemistry, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, Serdang, Malaysia
| | - M Aiman Mohtar
- UKM Medical Molecular Biology Institute, Universiti Kebangsaan Malaysia, Cheras, Malaysia, Department of Biochemistry, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, Serdang, Malaysia
| | - Saiful Effendi Syafruddin
- UKM Medical Molecular Biology Institute, Universiti Kebangsaan Malaysia, Cheras, Malaysia, Department of Biochemistry, Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, Serdang, Malaysia
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22
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Gim GM, Jang G. Outlook on genome editing application to cattle. J Vet Sci 2024; 25:e10. [PMID: 38311323 PMCID: PMC10839183 DOI: 10.4142/jvs.23133] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/15/2023] [Revised: 08/04/2023] [Accepted: 08/20/2023] [Indexed: 02/07/2024] Open
Abstract
In livestock industry, there is growing interest in methods to increase the production efficiency of livestock to address food shortages, given the increasing global population. With the advancements in gene engineering technology, it is a valuable tool and has been intensively utilized in research specifically focused on human disease. In historically, this technology has been used with livestock to create human disease models or to produce recombinant proteins from their byproducts. However, in recent years, utilizing gene editing technology, cattle with identified genes related to productivity can be edited, thereby enhancing productivity in response to climate change or specific disease instead of producing recombinant proteins. Furthermore, with the advancement in the efficiency of gene editing, it has become possible to edit multiple genes simultaneously. This cattle breed improvement has been achieved by discovering the genes through the comprehensive analysis of the entire genome of cattle. The cattle industry has been able to address gene bottlenecks that were previously impossible through conventional breeding systems. This review concludes that gene editing is necessary to expand the cattle industry, improving productivity in the future. Additionally, the enhancement of cattle through gene editing is expected to contribute to addressing environmental challenges associated with the cattle industry. Further research and development in gene editing, coupled with genomic analysis technologies, will significantly contribute to solving issues that conventional breeding systems have not been able to address.
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Affiliation(s)
| | - Goo Jang
- LARTBio Inco, Seoul 06221, Korea
- Laboratory of Theriogenology and Biotechnology, Department of Veterinary Clinical Science, College of Veterinary Medicine and the Research Institute of Veterinary Science, Seoul National University, Seoul 08826, Korea
- Comparative medicine Disease Research Center, Seoul National University, Seoul 08826, Korea
- Faculty of Veterinary Medicine, Universitas Airlangga, Surabaya 60115, Indonesia.
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23
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Tatwavedi D, Pellagatti A, Boultwood J. Recent advances in the application of induced pluripotent stem cell technology to the study of myeloid malignancies. Adv Biol Regul 2024; 91:100993. [PMID: 37827894 DOI: 10.1016/j.jbior.2023.100993] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/22/2023] [Accepted: 09/25/2023] [Indexed: 10/14/2023]
Abstract
Acquired myeloid malignancies are a spectrum of clonal disorders known to be caused by sequential acquisition of genetic lesions in hematopoietic stem and progenitor cells, leading to their aberrant self-renewal and differentiation. The increasing use of induced pluripotent stem cell (iPSC) technology to study myeloid malignancies has helped usher a paradigm shift in approaches to disease modeling and drug discovery, especially when combined with gene-editing technology. The process of reprogramming allows for the capture of the diversity of genetic lesions and mutational burden found in primary patient samples into individual stable iPSC lines. Patient-derived iPSC lines, owing to their self-renewal and differentiation capacity, can thus be a homogenous source of disease relevant material that allow for the study of disease pathogenesis using various functional read-outs. Furthermore, genome editing technologies like CRISPR/Cas9 enable the study of the stepwise progression from normal to malignant hematopoiesis through the introduction of specific driver mutations, individually or in combination, to create isogenic lines for comparison. In this review, we survey the current use of iPSCs to model acquired myeloid malignancies including myelodysplastic syndromes (MDS), myeloproliferative neoplasms (MPN), acute myeloid leukemia and MDS/MPN overlap syndromes. The use of iPSCs has enabled the interrogation of the underlying mechanism of initiation and progression driving these diseases. It has also made drug testing, repurposing, and the discovery of novel therapies for these diseases possible in a high throughput setting.
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Affiliation(s)
- Dharamveer Tatwavedi
- Blood Cancer UK Molecular Haematology Unit, Nuffield Division of Clinical Laboratory Sciences, Radcliffe Department of Medicine, University of Oxford, Oxford, UK.
| | - Andrea Pellagatti
- Blood Cancer UK Molecular Haematology Unit, Nuffield Division of Clinical Laboratory Sciences, Radcliffe Department of Medicine, University of Oxford, Oxford, UK
| | - Jacqueline Boultwood
- Blood Cancer UK Molecular Haematology Unit, Nuffield Division of Clinical Laboratory Sciences, Radcliffe Department of Medicine, University of Oxford, Oxford, UK.
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24
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Au TY, Arudkumar J, Assavarittirong C, Benjamin S. Killing two birds with one stone: CRISPR/Cas9 CCR5 knockout hematopoietic stem cells transplantation to treat patients with HIV infection and hematological malignancies concurrently. Clin Exp Med 2023; 23:4163-4175. [PMID: 37500934 DOI: 10.1007/s10238-023-01129-7] [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: 02/07/2023] [Accepted: 06/23/2023] [Indexed: 07/29/2023]
Abstract
Human immunodeficiency virus (HIV) is known to cause hematological malignancy. Hematopoietic stem cell transplantation (HPSCT) is an advanced treatment for that. Currently, there are three successful HIV-eliminated cases, and two received HPSCT from CCR5-absent donors. It is well established that the CCR5 protein on the cell surface assists human immunodeficiency virus entry. Preliminary studies have revealed that knocking out CCR5 and/or CXCR4 may inhibit the viral entry of HIV, which may prove promising in the further development of HIV treatment options. Herein, we suggest performing autologous or allogeneic HSCT with CCR5 KO hematopoietic stem cells in patients who suffer from complicated HIV conditions, particularly drug-resistant HIV or a concurrent diagnosis of HIV with lymphoma/leukemia, to achieve complete HIV remission. Nevertheless, at the clinical forefront of CRISPR-HIV technology, more efforts should be directed to advance nonhuman primate (NHP) models for studies of HIV pathogenesis and off-target assessments within this system. CRISPR-Cas9 knock out of host HSCT-expressing CCR5 or CXCR4 may confer HIV-resistance, which when applied to bedside therapeutics in an allogeneic or autologous manner can warrant a permanent and effective treatment outcome.
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Affiliation(s)
- Tsz Yuen Au
- Center for Medical Education in English, Poznan University of Medical Sciences, Poznan, Poland
| | - Jayshen Arudkumar
- South Australian Health and Medical Research Institute, Adelaide, SA, Australia.
- The University of Adelaide, Adelaide, SA, Australia.
| | - Chanika Assavarittirong
- Center for Medical Education in English, Poznan University of Medical Sciences, Poznan, Poland
| | - Shamiram Benjamin
- Center for Medical Education in English, Poznan University of Medical Sciences, Poznan, Poland
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25
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Hu J, Ferlez B, Dau J, Crickard JB. Rad53 regulates the lifetime of Rdh54 at homologous recombination intermediates. Nucleic Acids Res 2023; 51:11688-11705. [PMID: 37850655 PMCID: PMC10681728 DOI: 10.1093/nar/gkad848] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/22/2023] [Revised: 09/12/2023] [Accepted: 09/21/2023] [Indexed: 10/19/2023] Open
Abstract
Rdh54 is a conserved DNA translocase that participates in homologous recombination (HR), DNA checkpoint adaptation, and chromosome segregation. Saccharomyces cerevisiae Rdh54 is a known target of the Mec1/Rad53 signaling axis, which globally protects genome integrity during DNA metabolism. While phosphorylation of DNA repair proteins by Mec1/Rad53 is critical for HR progression little is known about how specific post translational modifications alter HR reactions. Phosphorylation of Rdh54 is linked to protection of genomic integrity but the consequences of modification remain poorly understood. Here, we demonstrate that phosphorylation of the Rdh54 C-terminus by the effector kinase Rad53 regulates Rdh54 clustering activity as revealed by single molecule imaging. This stems from phosphorylation dependent and independent interactions between Rdh54 and Rad53. Genetic assays reveal that loss of phosphorylation leads to phenotypic changes resulting in loss-of-heterozygosity (LOH) outcomes. Our data highlight Rad53 as a key regulator of HR intermediates through activation and attenuation of Rdh54 motor function.
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Affiliation(s)
- Jingyi Hu
- Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853, USA
| | - Bryan Ferlez
- Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853, USA
| | - Jennifer Dau
- Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853, USA
| | - J Brooks Crickard
- Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY 14853, USA
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26
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Wang R, Sun Y, Li C, Xue Y, Ba X. Targeting the DNA Damage Response for Cancer Therapy. Int J Mol Sci 2023; 24:15907. [PMID: 37958890 PMCID: PMC10648182 DOI: 10.3390/ijms242115907] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/21/2023] [Revised: 10/30/2023] [Accepted: 11/01/2023] [Indexed: 11/15/2023] Open
Abstract
Over the course of long-term evolution, cells have developed intricate defense mechanisms in response to DNA damage; these mechanisms play a pivotal role in maintaining genomic stability. Defects in the DNA damage response pathways can give rise to various diseases, including cancer. The DNA damage response (DDR) system is instrumental in safeguarding genomic stability. The accumulation of DNA damage and the weakening of DDR function both promote the initiation and progression of tumors. Simultaneously, they offer opportunities and targets for cancer therapeutics. This article primarily elucidates the DNA damage repair pathways and the progress made in targeting key proteins within these pathways for cancer treatment. Among them, poly (ADP-ribose) polymerase 1 (PARP1) plays a crucial role in DDR, and inhibitors targeting PARP1 have garnered extensive attention in anticancer research. By delving into the realms of DNA damage and repair, we aspire to explore more precise and effective strategies for cancer therapy and to seek novel avenues for intervention.
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Affiliation(s)
- Ruoxi Wang
- Center for Cell Structure and Function, Key Laboratory of Animal Resistance Biology of Shandong Province, College of Life Sciences, Shandong Normal University, Jinan 250014, China; (R.W.); (Y.S.)
| | - Yating Sun
- Center for Cell Structure and Function, Key Laboratory of Animal Resistance Biology of Shandong Province, College of Life Sciences, Shandong Normal University, Jinan 250014, China; (R.W.); (Y.S.)
| | - Chunshuang Li
- Key Laboratory of Molecular Epigenetics of Ministry of Education, College of Life Sciences, Northeast Normal University, Changchun 130024, China; (C.L.); (Y.X.)
| | - Yaoyao Xue
- Key Laboratory of Molecular Epigenetics of Ministry of Education, College of Life Sciences, Northeast Normal University, Changchun 130024, China; (C.L.); (Y.X.)
| | - Xueqing Ba
- Key Laboratory of Molecular Epigenetics of Ministry of Education, College of Life Sciences, Northeast Normal University, Changchun 130024, China; (C.L.); (Y.X.)
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27
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Baghini SS, Razeghian E, Malayer SK, Pecho RDC, Obaid M, Awfi ZS, Zainab HA, Shamsara M. Recent advances in the application of genetic and epigenetic modalities in the improvement of antibody-producing cell lines. Int Immunopharmacol 2023; 123:110724. [PMID: 37582312 DOI: 10.1016/j.intimp.2023.110724] [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: 01/31/2023] [Revised: 07/25/2023] [Accepted: 07/26/2023] [Indexed: 08/17/2023]
Abstract
There are numerous applications for recombinant antibodies (rAbs) in biological and toxicological research. Monoclonal antibodies are synthesized using genetic engineering and other related processes involved in the generation of rAbs. Because they can identify specific antigenic sites on practically any molecule, including medicines, hormones, microbial antigens, and cell receptors, rAbs are particularly useful in scientific research. The key benefits of rAbs are improved repeatability, control, and consistency, shorter manufacturing times than with hybridoma technology, an easier transition from one format of antibody to another, and an animal-free process. The engineering of the host cell has recently been developed method for enhancing the production efficiency and improving the quality of antibodies from mammalian cell lines. In this light, genetic engineering is mostly utilized to manage cellular chaperones, decrease cell death, increase cell viability, change the microRNAs (miRNAs) pattern in mammalian cells, and glycoengineered cell lines. Here, we shed light on how genetic engineering can be used therapeutically to produce antibodies at higher levels with greater potency and effectiveness.
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Affiliation(s)
- Sadegh Shojaei Baghini
- Plant Biotechnology Department, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.
| | - Ehsan Razeghian
- Human Genetics Division, Medical Biotechnology Department, National Institute of Genetics Engineering and Biotechnology (NIGEB), Tehran, Iran
| | - Setare Kakavand Malayer
- Department of Biology, Faculty of Biological Science, Tehran North Branch, Islamic Azad University, Tehran, Iran
| | | | | | - Zinah Salem Awfi
- Department of Dental Industry Techniques, Al-Noor University College, Nineveh, Iraq.
| | - H A Zainab
- Department of Pharmacy, Al-Zahrawi University College, Karbala, Iraq.
| | - Mehdi Shamsara
- Department of Animal Biotechnology, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran.
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28
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Zhou Y, Qiu C, Fu Q, Li T, Zhang X, Zhu C, Qin X, Wu B. Pan-Cancer Analysis of Oncogenic Role of RAD54L and Experimental Validation in Hepatocellular Carcinoma. J Inflamm Res 2023; 16:3997-4017. [PMID: 37719938 PMCID: PMC10503553 DOI: 10.2147/jir.s426558] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/08/2023] [Accepted: 09/01/2023] [Indexed: 09/19/2023] Open
Abstract
Background RAD54L is a prominent member of the SWI2/SNF2 protein family, primarily involved in the homologous recombination repair (HRR) process, thereby playing a pivotal role in the repair of DNA double-strand breaks (DSBs). RAD54L has been implicated in the development of numerous tumors. Consequently, we aimed to investigate the potential contribution of RAD54L in pan-cancer. Methods Various databases and analytical tools were employed for bioinformatics analysis. Moreover, in vitro experiments were conducted to corroborate the findings from the bioinformatics analysis and delve deeper into the role of RAD54L in hepatocellular carcinoma (HCC). Results RAD54L expression demonstrated a significant elevation in the majority of tumors, and its overexpression was strongly associated with unfavorable survival outcomes. RAD54L displayed robust correlations with the infiltration levels of various immune cells, including cancer associated fibroblasts (CAFs), endothelial cells, and myeloid-derived suppressor cells (MDSCs). Additionally, associations were observed between RAD54L and key factors such as tumor mutation burden (TMB), microsatellite instability (MSI), multiple immune checkpoints, and immune cell infiltration. Moreover, a close relationship was observed between RAD54L expression levels in HCC and clinicopathological characteristics, as well as immune cell infiltration. Experimental techniques including qRT-PCR, Western blotting, colony-forming, Cell Counting Kit-8 (CCK-8), wound-healing, and transwell assays were employed, which collectively demonstrated that RAD54L promoted the proliferation and migration of HCC cells. Conclusion RAD54L exhibits robust expression in both pan-cancer and HCC, exerting a significant influence on the proliferation and migration of HCC cells. These findings highlight its potential as a promising biomarker for pan-cancer and a prospective target for immunotherapy.
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Affiliation(s)
- Yongzhen Zhou
- Department of General Surgery, the Affiliated Changzhou No. 2 People’s Hospital of Nanjing Medical University, Changzhou, 213000, People’s Republic of China
- Graduate School, Bengbu Medical College, Bengbu, 233003, People’s Republic of China
| | - Chenjie Qiu
- Department of General Surgery, Changzhou Hospital of Traditional Chinese Medicine, Changzhou, 213000, People’s Republic of China
| | - Qingsheng Fu
- Graduate School, Bengbu Medical College, Bengbu, 233003, People’s Republic of China
| | - Tao Li
- Department of General Surgery, the Affiliated Changzhou No. 2 People’s Hospital of Nanjing Medical University, Changzhou, 213000, People’s Republic of China
| | - Xudong Zhang
- Department of General Surgery, the Affiliated Changzhou No. 2 People’s Hospital of Nanjing Medical University, Changzhou, 213000, People’s Republic of China
| | - Chunfu Zhu
- Department of General Surgery, the Affiliated Changzhou No. 2 People’s Hospital of Nanjing Medical University, Changzhou, 213000, People’s Republic of China
| | - Xihu Qin
- Department of General Surgery, the Affiliated Changzhou No. 2 People’s Hospital of Nanjing Medical University, Changzhou, 213000, People’s Republic of China
| | - Baoqiang Wu
- Department of General Surgery, the Affiliated Changzhou No. 2 People’s Hospital of Nanjing Medical University, Changzhou, 213000, People’s Republic of China
- Graduate School, Bengbu Medical College, Bengbu, 233003, People’s Republic of China
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Guo Y, Zhao G, Gao X, Zhang L, Zhang Y, Cai X, Yuan X, Guo X. CRISPR/Cas9 gene editing technology: a precise and efficient tool for crop quality improvement. PLANTA 2023; 258:36. [PMID: 37395789 DOI: 10.1007/s00425-023-04187-z] [Citation(s) in RCA: 7] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Subscribe] [Scholar Register] [Received: 01/18/2023] [Accepted: 06/18/2023] [Indexed: 07/04/2023]
Abstract
MAIN CONCLUSION This review provides a direction for crop quality improvement and ideas for further research on the application of CRISPR/Cas9 gene editing technology for crop improvement. Various important crops, such as wheat, rice, soybean and tomato, are among the main sources of food and energy for humans. Breeders have long attempted to improve crop yield and quality through traditional breeding methods such as crossbreeding. However, crop breeding progress has been slow due to the limitations of traditional breeding methods. In recent years, clustered regularly spaced short palindromic repeat (CRISPR)/Cas9 gene editing technology has been continuously developed. And with the refinement of crop genome data, CRISPR/Cas9 technology has enabled significant breakthroughs in editing specific genes of crops due to its accuracy and efficiency. Precise editing of certain key genes in crops by means of CRISPR/Cas9 technology has improved crop quality and yield and has become a popular strategy for many breeders to focus on and adopt. In this paper, the present status and achievements of CRISPR/Cas9 gene technology as applied to the improvement of quality in several crops are reviewed. In addition, the shortcomings, challenges and development prospects of CRISPR/Cas9 gene editing technology are discussed.
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Affiliation(s)
- Yingxin Guo
- College of Biological and Chemical Engineering, Qilu Institute of Technology, Jinan, 250200, Shandong, People's Republic of China
| | - Guangdong Zhao
- College of Life Sciences, Linyi University, Linyi, 276000, Shandong, People's Republic of China
| | - Xing Gao
- College of Biological and Chemical Engineering, Qilu Institute of Technology, Jinan, 250200, Shandong, People's Republic of China
| | - Lin Zhang
- College of Biological and Chemical Engineering, Qilu Institute of Technology, Jinan, 250200, Shandong, People's Republic of China
| | - Yanan Zhang
- College of Biological and Chemical Engineering, Qilu Institute of Technology, Jinan, 250200, Shandong, People's Republic of China
| | - Xiaoming Cai
- College of Biological and Chemical Engineering, Qilu Institute of Technology, Jinan, 250200, Shandong, People's Republic of China
| | - Xuejiao Yuan
- College of Biological and Chemical Engineering, Qilu Institute of Technology, Jinan, 250200, Shandong, People's Republic of China.
| | - Xingqi Guo
- State Key Laboratory of Crop Biology, College of Life Sciences, Shandong Agricultural University, Taian, 271018, Shandong, People's Republic of China.
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Li M, Niu X, Li S, Fu S, Li Q, Xu M, Wang C, Wu S. CRISPR/Cas9 Based Cell-Type Specific Gene Knock-Out in Arabidopsis Roots. PLANTS (BASEL, SWITZERLAND) 2023; 12:2365. [PMID: 37375990 DOI: 10.3390/plants12122365] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/19/2023] [Revised: 06/14/2023] [Accepted: 06/15/2023] [Indexed: 06/29/2023]
Abstract
CRISPR/Cas9 (hereafter Cas9)-mediated gene knockout is one of the most important tools for studying gene function. However, many genes in plants play distinct roles in different cell types. Engineering the currently used Cas9 system to achieve cell-type-specific knockout of functional genes is useful for addressing the cell-specific functions of genes. Here we employed the cell-specific promoters of the WUSCHEL RELATED HOMEOBOX 5 (WOX5), CYCLIND6;1 (CYCD6;1), and ENDODERMIS7 (EN7) genes to drive the Cas9 element, allowing tissue-specific targeting of the genes of interest. We designed the reporters to verify the tissue-specific gene knockout in vivo. Our observation of the developmental phenotypes provides strong evidence for the involvement of SCARECROW (SCR) and GIBBERELLIC ACID INSENSITIVE (GAI) in the development of quiescent center (QC) and endodermal cells. This system overcomes the limitations of traditional plant mutagenesis techniques, which often result in embryonic lethality or pleiotropic phenotypes. By allowing cell-type-specific manipulation, this system has great potential to help us better understand the spatiotemporal functions of genes during plant development.
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Affiliation(s)
- Meng Li
- College of Life Sciences and Horticultural Plant Biology Metabolomics Center, Fujian Agriculture and Forestry University, Fuzhou 350002, China
| | - Xufang Niu
- College of Life Sciences and Horticultural Plant Biology Metabolomics Center, Fujian Agriculture and Forestry University, Fuzhou 350002, China
| | - Shuang Li
- College of Life Sciences and Horticultural Plant Biology Metabolomics Center, Fujian Agriculture and Forestry University, Fuzhou 350002, China
| | - Shasha Fu
- College of Life Sciences and Horticultural Plant Biology Metabolomics Center, Fujian Agriculture and Forestry University, Fuzhou 350002, China
| | - Qianfang Li
- College of Life Sciences and Horticultural Plant Biology Metabolomics Center, Fujian Agriculture and Forestry University, Fuzhou 350002, China
| | - Meizhi Xu
- College of Life Sciences and Horticultural Plant Biology Metabolomics Center, Fujian Agriculture and Forestry University, Fuzhou 350002, China
| | - Chunhua Wang
- College of Life Sciences and Horticultural Plant Biology Metabolomics Center, Fujian Agriculture and Forestry University, Fuzhou 350002, China
| | - Shuang Wu
- College of Life Sciences and Horticultural Plant Biology Metabolomics Center, Fujian Agriculture and Forestry University, Fuzhou 350002, China
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Patel L, Pritchard CC. Molecular testing of DNA damage response pathways in prostate cancer patients. Curr Opin Oncol 2023; 35:224-230. [PMID: 36966502 DOI: 10.1097/cco.0000000000000934] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 03/27/2023]
Abstract
PURPOSE OF REVIEW Personalizing prostate cancer therapy requires germline and tumor molecular tests that predict who will respond to specific treatments and who may not. The review covers molecular testing of DNA damage response pathways, the first biomarker-driven precision target with clinical utility for treatment selection in patients with castration resistant prostate cancer (CRPC). RECENT FINDINGS Recurrent somatic and germline variants cause deficiency of the mismatch repair (MMR) or homologous recombination (HR) pathways in about a quarter of CRPC patients. In prospective clinical trials, patients with deleterious variants in the MMR pathway more frequently experience a therapeutic response to immune checkpoint inhibitors (ICI). Similarly, somatic and germline events affecting HR predict response to poly(ADP) ribose polymerase inhibitor (PARPi) therapy. Molecular testing of these pathways currently involves assaying for loss of function variants in individual genes and for the genome-wide consequences of repair deficiency. SUMMARY DNA damage response pathways are the first major area of molecular genetic testing in CRPC settings and offer insights into this new paradigm. Our hope is that eventually an arsenal of molecularly-guided therapies will be developed across many pathways to enable precision medicine options for most men with prostate cancer.
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Affiliation(s)
- Lalit Patel
- Department of Genetics, The University of Texas MD Anderson Cancer Center, Houston, Texas
| | - Colin C Pritchard
- Department of Laboratory Medicine and Pathology, University of Washington
- Brotman Baty Institute for Precision Medicine, Seattle, Washington, USA
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Garaycoechea JI, Quinlan C, Luijsterburg MS. Pathological consequences of DNA damage in the kidney. Nat Rev Nephrol 2023; 19:229-243. [PMID: 36702905 DOI: 10.1038/s41581-022-00671-z] [Citation(s) in RCA: 15] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 12/09/2022] [Indexed: 01/27/2023]
Abstract
DNA lesions that evade repair can lead to mutations that drive the development of cancer, and cellular responses to DNA damage can trigger senescence and cell death, which are associated with ageing. In the kidney, DNA damage has been implicated in both acute and chronic kidney injury, and in renal cell carcinoma. The susceptibility of the kidney to chemotherapeutic agents that damage DNA is well established, but an unexpected link between kidney ciliopathies and the DNA damage response has also been reported. In addition, human genetic deficiencies in DNA repair have highlighted DNA crosslinks, DNA breaks and transcription-blocking damage as lesions that are particularly toxic to the kidney. Genetic tools in mice, as well as advances in kidney organoid and single-cell RNA sequencing technologies, have provided important insights into how specific kidney cell types respond to DNA damage. The emerging view is that in the kidney, DNA damage affects the local microenvironment by triggering a damage response and cell proliferation to replenish injured cells, as well as inducing systemic responses aimed at reducing exposure to genotoxic stress. The pathological consequences of DNA damage are therefore key to the nephrotoxicity of DNA-damaging agents and the kidney phenotypes observed in human DNA repair-deficiency disorders.
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Affiliation(s)
- Juan I Garaycoechea
- Hubrecht Institute-KNAW, University Medical Center Utrecht, Utrecht, The Netherlands.
| | - Catherine Quinlan
- Department of Paediatrics, University of Melbourne, Parkville, Australia
- Department of Nephrology, Royal Children's Hospital, Melbourne, Australia
- Department of Kidney Regeneration, Murdoch Children's Research Institute, Melbourne, Australia
| | - Martijn S Luijsterburg
- Department of Human Genetics, Leiden University Medical Center (LUMC), Leiden, The Netherlands.
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Ghantous L, Volman Y, Hefez R, Wald O, Stern E, Friehmann T, Chajut A, Bremer E, Elhalel MD, Rachmilewitz J. The DNA damage response pathway regulates the expression of the immune checkpoint CD47. Commun Biol 2023; 6:245. [PMID: 36882648 PMCID: PMC9992352 DOI: 10.1038/s42003-023-04615-6] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/29/2022] [Accepted: 02/21/2023] [Indexed: 03/09/2023] Open
Abstract
CD47 is a cell surface ligand expressed on all nucleated cells. It is a unique immune checkpoint protein acting as "don't eat me" signal to prevent phagocytosis and is constitutively overexpressed in many tumors. However, the underlying mechanism(s) for CD47 overexpression is not clear. Here, we show that irradiation (IR) as well as various other genotoxic agents induce elevated expression of CD47. This upregulation correlates with the extent of residual double-strand breaks (DSBs) as determined by γH2AX staining. Interestingly, cells lacking mre-11, a component of the MRE11-RAD50-NBS1 (MRN) complex that plays a central role in DSB repair, or cells treated with the mre-11 inhibitor, mirin, fail to elevate the expression of CD47 upon DNA damage. On the other hand, both p53 and NF-κB pathways or cell-cycle arrest do not play a role in CD47 upregualtion upon DNA damage. We further show that CD47 expression is upregulated in livers harvested from mice treated with the DNA-damage inducing agent Diethylnitrosamine (DEN) and in cisplatin-treated mesothelioma tumors. Hence, our results indicate that CD47 is upregulated following DNA damage in a mre-11-dependent manner. Chronic DNA damage response in cancer cells might contribute to constitutive elevated expression of CD47 and promote immune evasion.
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Affiliation(s)
- Lucy Ghantous
- Goldyne Savad Institute of Gene Therapy, Hadassah Medical Center, Faculty of Medicine, Hebrew University of Jerusalem, Jerusalem, Israel
- Department of Nephrology and Hypertension, Hadassah Medical Center, Faculty of Medicine, Hebrew University of Jerusalem, Jerusalem, Israel
| | - Yael Volman
- Goldyne Savad Institute of Gene Therapy, Hadassah Medical Center, Faculty of Medicine, Hebrew University of Jerusalem, Jerusalem, Israel
| | - Ruth Hefez
- Goldyne Savad Institute of Gene Therapy, Hadassah Medical Center, Faculty of Medicine, Hebrew University of Jerusalem, Jerusalem, Israel
| | - Ori Wald
- Goldyne Savad Institute of Gene Therapy, Hadassah Medical Center, Faculty of Medicine, Hebrew University of Jerusalem, Jerusalem, Israel
- Department of Cardiothoracic Surgery, Hadassah Medical Center, Faculty of Medicine, Hebrew University of Jerusalem, Jerusalem, Israel
| | - Esther Stern
- Goldyne Savad Institute of Gene Therapy, Hadassah Medical Center, Faculty of Medicine, Hebrew University of Jerusalem, Jerusalem, Israel
| | - Tomer Friehmann
- Goldyne Savad Institute of Gene Therapy, Hadassah Medical Center, Faculty of Medicine, Hebrew University of Jerusalem, Jerusalem, Israel
| | | | - Edwin Bremer
- Department of Hematology, University Medical Center Groningen, Groningen, The Netherlands
| | - Michal Dranitzki Elhalel
- Department of Nephrology and Hypertension, Hadassah Medical Center, Faculty of Medicine, Hebrew University of Jerusalem, Jerusalem, Israel.
| | - Jacob Rachmilewitz
- Goldyne Savad Institute of Gene Therapy, Hadassah Medical Center, Faculty of Medicine, Hebrew University of Jerusalem, Jerusalem, Israel.
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Genome editing in cancer: Challenges and potential opportunities. Bioact Mater 2023; 21:394-402. [PMID: 36185740 PMCID: PMC9483578 DOI: 10.1016/j.bioactmat.2022.08.013] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2022] [Revised: 08/21/2022] [Accepted: 08/24/2022] [Indexed: 11/29/2022] Open
Abstract
Ever since its mechanism was discovered back in 2012, the CRISPR/Cas9 system have revolutionized the field of genome editing. While at first it was seen as a therapeutic tool mostly relevant for curing genetic diseases, it has been recently shown to also hold the potential to become a clinically relevant therapy for cancer. However, there are multiple challenges that must be addressed prior to clinical testing. Predominantly, the safety of the system when used for in-vivo therapies, including off-target activity and the effects of the double strand break induction on genomic stability. Here, we will focus on the inherent challenges in the CRISPR/Cas9 system and discuss various opportunities to overcoming these challenges.
In recent years, several works have shown that knocking down key genes by CRISPR/Cas9 based could potentially be a new type of cancer therapy. This has been made possible due to advances in the fields of In-vivo delivery, such as lentiviral vectors and lipid nanoparticles. Limiting CRISPR/Cas9 activity to the tumor and minimizing off-target activity are challenges that must be overcome before proceeding to the clinic. We review approaches arising from multiple disciplines that could overcome these challenges. The combination of these multi-disciplinary approaches should be able to overcome the different challenges and open the way to the clinic.
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Kong Q, Li J, Wang S, Feng X, Shou H. Combination of Hairy Root and Whole-Plant Transformation Protocols to Achieve Efficient CRISPR/Cas9 Genome Editing in Soybean. PLANTS (BASEL, SWITZERLAND) 2023; 12:1017. [PMID: 36903878 PMCID: PMC10005656 DOI: 10.3390/plants12051017] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 01/16/2023] [Revised: 02/17/2023] [Accepted: 02/20/2023] [Indexed: 06/18/2023]
Abstract
The new gene-editing technology CRISPR/Cas system has been widely used for genome engineering in various organisms. Since the CRISPR/Cas gene-editing system has a certain possibility of low efficiency and the whole plant transformation of soybean is time-consuming and laborious, it is important to evaluate the editing efficiency of designed CRISPR constructs before the stable whole plant transformation process starts. Here, we provide a modified protocol for generating transgenic hairy soybean roots to assess the efficiency of guide RNA (gRNA) sequences of the CRISPR/Cas constructs within 14 days. The cost- and space-effective protocol was first tested in transgenic soybean harboring the GUS reporter gene for the efficiency of different gRNA sequences. Targeted DNA mutations were detected in 71.43-97.62% of the transgenic hairy roots analyzed as evident by GUS staining and DNA sequencing of the target region. Among the four designed gene-editing sites, the highest editing efficiency occurred at the 3' terminal of the GUS gene. In addition to the reporter gene, the protocol was tested for the gene-editing of 26 soybean genes. Among the gRNAs selected for stable transformation, the editing efficiency of hairy root transformation and stable transformation ranged from 5% to 88.8% and 2.7% to 80%, respectively. The editing efficiencies of stable transformation were positively correlated with those of hairy root transformation with a Pearson correlation coefficient (r) of 0.83. Our results demonstrated that soybean hairy root transformation could rapidly assess the efficiency of designed gRNA sequences on genome editing. This method can not only be directly applied to the functional study of root-specific genes, but more importantly, it can be applied to the pre-screening of gRNA in CRISPR/Cas gene editing.
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Affiliation(s)
- Qihui Kong
- State Key Laboratory of Plant Physiology and Biochemistry, College of Life Sciences, Zhejiang University, Hangzhou 310058, China
- Zhejiang Lab, Hangzhou 310012, China
| | - Jie Li
- State Key Laboratory of Plant Physiology and Biochemistry, College of Life Sciences, Zhejiang University, Hangzhou 310058, China
| | - Shoudong Wang
- Zhejiang Lab, Hangzhou 310012, China
- Key Laboratory of Soybean Molecular Design Breeding, Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences, Changchun 130102, China
| | - Xianzhong Feng
- Zhejiang Lab, Hangzhou 310012, China
- Key Laboratory of Soybean Molecular Design Breeding, Northeast Institute of Geography and Agroecology, Chinese Academy of Sciences, Changchun 130102, China
| | - Huixia Shou
- State Key Laboratory of Plant Physiology and Biochemistry, College of Life Sciences, Zhejiang University, Hangzhou 310058, China
- Zhejiang Lab, Hangzhou 310012, China
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36
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Qi Q, Hu B, Jiang W, Wang Y, Yan J, Ma F, Guan Q, Xu J. Advances in Plant Epigenome Editing Research and Its Application in Plants. Int J Mol Sci 2023; 24:ijms24043442. [PMID: 36834852 PMCID: PMC9961165 DOI: 10.3390/ijms24043442] [Citation(s) in RCA: 9] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/31/2022] [Revised: 01/29/2023] [Accepted: 02/02/2023] [Indexed: 02/11/2023] Open
Abstract
Plant epistatic regulation is the DNA methylation, non-coding RNA regulation, and histone modification of gene sequences without altering the genome sequence, thus regulating gene expression patterns and the growth process of plants to produce heritable changes. Epistatic regulation in plants can regulate plant responses to different environmental stresses, regulate fruit growth and development, etc. Genome editing can effectively improve plant genetic efficiency by targeting the design and efficient editing of genome-specific loci with specific nucleases, such as zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALEN), and clustered regularly interspaced short palindromic repeats/CRISPR-associated 9 (CRISPR/Cas9). As research progresses, the CRISPR/Cas9 system has been widely used in crop breeding, gene expression, and epistatic modification due to its high editing efficiency and rapid translation of results. In this review, we summarize the recent progress of CRISPR/Cas9 in epigenome editing and look forward to the future development direction of this system in plant epigenetic modification to provide a reference for the application of CRISPR/Cas9 in genome editing.
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Affiliation(s)
- Qiaoyun Qi
- State Key Laboratory of Crop Stress Biology for Arid Areas/Shaanxi Key Laboratory of Apple, College of Horticulture, Northwest A&F University, Xianyang 712100, China
| | - Bichun Hu
- State Key Laboratory of Crop Stress Biology for Arid Areas/Shaanxi Key Laboratory of Apple, College of Horticulture, Northwest A&F University, Xianyang 712100, China
| | - Weiyu Jiang
- State Key Laboratory of Crop Stress Biology for Arid Areas/Shaanxi Key Laboratory of Apple, College of Horticulture, Northwest A&F University, Xianyang 712100, China
| | - Yixiong Wang
- State Key Laboratory of Crop Stress Biology for Arid Areas/Shaanxi Key Laboratory of Apple, College of Horticulture, Northwest A&F University, Xianyang 712100, China
| | - Jinjiao Yan
- College of Forestry, Northwest A&F University, Xianyang 712100, China
| | - Fengwang Ma
- State Key Laboratory of Crop Stress Biology for Arid Areas/Shaanxi Key Laboratory of Apple, College of Horticulture, Northwest A&F University, Xianyang 712100, China
| | - Qingmei Guan
- State Key Laboratory of Crop Stress Biology for Arid Areas/Shaanxi Key Laboratory of Apple, College of Horticulture, Northwest A&F University, Xianyang 712100, China
| | - Jidi Xu
- State Key Laboratory of Crop Stress Biology for Arid Areas/Shaanxi Key Laboratory of Apple, College of Horticulture, Northwest A&F University, Xianyang 712100, China
- Correspondence:
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Li C, Zhu H, Jin S, Maksoud LM, Jain N, Sun J, Gao Y. Structural basis of DNA polymerase θ mediated DNA end joining. Nucleic Acids Res 2023; 51:463-474. [PMID: 36583344 PMCID: PMC9841435 DOI: 10.1093/nar/gkac1201] [Citation(s) in RCA: 9] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/22/2022] [Revised: 11/30/2022] [Accepted: 12/05/2022] [Indexed: 12/31/2022] Open
Abstract
DNA polymerase θ (Pol θ) plays an essential role in the microhomology-mediated end joining (MMEJ) pathway for repairing DNA double-strand breaks. However, the mechanisms by which Pol θ recognizes microhomologous DNA ends and performs low-fidelity DNA synthesis remain unclear. Here, we present cryo-electron microscope structures of the polymerase domain of Lates calcarifer Pol θ with long and short duplex DNA at up to 2.4 Å resolution. Interestingly, Pol θ binds to long and short DNA substrates similarly, with extensive interactions around the active site. Moreover, Pol θ shares a similar active site as high-fidelity A-family polymerases with its finger domain well-closed but differs in having hydrophilic residues surrounding the nascent base pair. Computational simulations and mutagenesis studies suggest that the unique insertion loops of Pol θ help to stabilize short DNA binding and assemble the active site for MMEJ repair. Taken together, our results illustrate the structural basis of Pol θ-mediated MMEJ.
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Affiliation(s)
- Chuxuan Li
- Department of Biosciences, Rice University, Houston, TX 77005, USA
| | - Hanwen Zhu
- Department of Structural Biology, St. Jude Children’s Research Hospital, Memphis, TN 38105, USA
| | - Shikai Jin
- Department of Biosciences, Rice University, Houston, TX 77005, USA
- Center for Theoretical Biological Physics, Rice University, Houston, TX 77005, USA
| | - Leora M Maksoud
- Department of Biosciences, Rice University, Houston, TX 77005, USA
| | - Nikhil Jain
- Department of Structural Biology, St. Jude Children’s Research Hospital, Memphis, TN 38105, USA
| | - Ji Sun
- Department of Structural Biology, St. Jude Children’s Research Hospital, Memphis, TN 38105, USA
| | - Yang Gao
- Department of Biosciences, Rice University, Houston, TX 77005, USA
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Cheon H, Wang Y, Wightman SM, Jackson MW, Stark GR. How cancer cells make and respond to interferon-I. Trends Cancer 2023; 9:83-92. [PMID: 36216730 PMCID: PMC9797472 DOI: 10.1016/j.trecan.2022.09.003] [Citation(s) in RCA: 83] [Impact Index Per Article: 41.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2022] [Revised: 09/06/2022] [Accepted: 09/12/2022] [Indexed: 11/07/2022]
Abstract
Acute exposure of cancer cells to high concentrations of type I interferon (IFN-I) drives growth arrest and apoptosis, whereas chronic exposure to low concentrations provides important prosurvival advantages. Tyrosine-phosphorylated IFN-stimulated gene (ISG) factor 3 (ISGF3) drives acute deleterious responses to IFN-I, whereas unphosphorylated (U-)ISGF3, lacking tyrosine phosphorylation, drives essential constitutive prosurvival mechanisms. Surprisingly, programmed cell death-ligand 1 (PD-L1), often expressed on the surfaces of tumor cells and well recognized for its importance in inactivating cytotoxic T cells, also has important cell-intrinsic protumor activities, including dampening acute responses to cytotoxic high levels of IFN-I and sustaining the expression of the low levels that benefit tumors. More thorough understanding of the newly recognized complex roles of IFN-I in cancer may lead to the identification of novel therapeutic strategies.
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Affiliation(s)
- HyeonJoo Cheon
- Department of Oncology, Wayne State University School of Medicine, Karmanos Cancer Institute, Detroit, MI, USA
| | - Yuxin Wang
- Department of Cancer Biology, Lerner Research Institute, Cleveland Clinic, Cleveland, OH, USA
| | - Samantha M. Wightman
- Department of Cancer Biology, Lerner Research Institute, Cleveland Clinic, Cleveland, OH, USA
| | - Mark W. Jackson
- Department of Pathology, Case Western Reserve University School of Medicine, Cleveland, OH, USA
| | - George R. Stark
- Department of Cancer Biology, Lerner Research Institute, Cleveland Clinic, Cleveland, OH, USA,Correspondence: (G.R. Stark)
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Lee C, Leem J, Oh JS. Selective utilization of non-homologous end-joining and homologous recombination for DNA repair during meiotic maturation in mouse oocytes. Cell Prolif 2022; 56:e13384. [PMID: 36564861 PMCID: PMC10068936 DOI: 10.1111/cpr.13384] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/09/2022] [Revised: 11/29/2022] [Accepted: 12/08/2022] [Indexed: 12/25/2022] Open
Abstract
DNA double-strand breaks (DSBs) are highly toxic lesions that can cause genomic instability and can be repaired by non-homologous end-joining (NHEJ) and homologous recombination (HR) pathways. Despite extensive studies about DSB repair pathways, the roles of each pathway during meiotic maturation in oocytes are not well understood. Here we show that oocytes selectively utilize NHEJ and HR to repair DSBs during meiotic maturation. Inhibition of NHEJ impaired the meiotic maturation of oocytes with DNA damage by activating the spindle assembly checkpoint (SAC) with a concomitant increase in metaphase I (MI) arrest and DNA damage levels. In contrast, oocytes with DNA damage bypassed SAC-mediated MI arrest despite the presence of fragmented DNA when HR was inhibited. Notably, this bypass of SAC arrest by HR inhibition was associated with a loss of centromere integrity and subsequent impairment of chromosome architecture. Our results demonstrate that, while NHEJ is critical for the meiotic maturation of oocytes with DNA damage, HR is essential to maintain centromere integrity against DNA damage during meiotic maturation, revealing distinct roles of NHEJ and HR during meiotic maturation in mouse oocytes.
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Affiliation(s)
- Crystal Lee
- Department of Integrative Biotechnology, Sungkyunkwan University, Suwon, South Korea
| | - Jiyeon Leem
- Department of Integrative Biotechnology, Sungkyunkwan University, Suwon, South Korea
| | - Jeong Su Oh
- Department of Integrative Biotechnology, Sungkyunkwan University, Suwon, South Korea.,Biomedical Institute for Convergence at SKKU (BICS), Sungkyunkwan University, Suwon, South Korea
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40
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Factors to Consider for the Correct Use of γH2AX in the Evaluation of DNA Double-Strand Breaks Damage Caused by Ionizing Radiation. Cancers (Basel) 2022; 14:cancers14246204. [PMID: 36551689 PMCID: PMC9776434 DOI: 10.3390/cancers14246204] [Citation(s) in RCA: 21] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/22/2022] [Revised: 12/07/2022] [Accepted: 12/13/2022] [Indexed: 12/23/2022] Open
Abstract
People exposed to ionizing radiation (IR) both for diagnostic and therapeutic purposes is constantly increasing. Since the use of IR involves a risk of harmful effects, such as the DNA DSB induction, an accurate determination of this induced DNA damage and a correct evaluation of the risk-benefit ratio in the clinical field are of key relevance. γH2AX (the phosphorylated form of the histone variant H2AX) is a very early marker of DSBs that can be induced both in physiological conditions, such as in the absence of specific external agents, and by external factors such as smoking, heat, background environmental radiation, and drugs. All these internal and external conditions result in a basal level of γH2AX which must be considered for the correct assessment of the DSBs after IR exposure. In this review we analyze the most common conditions that induce H2AX phosphorylation, including specific exogenous stimuli, cellular states, basic environmental factors, and lifestyles. Moreover, we discuss the most widely used methods for γH2AX determination and describe the principal applications of γH2AX scoring, paying particular attention to clinical studies. This knowledge will help us optimize the use of available methods in order to discern the specific γH2AX following IR-induced DSBs from the basal level of γH2AX in the cells.
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41
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Nucleic acid-based scaffold systems and application in enzyme cascade catalysis. Appl Microbiol Biotechnol 2022; 107:9-23. [DOI: 10.1007/s00253-022-12315-0] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/30/2022] [Revised: 11/23/2022] [Accepted: 11/24/2022] [Indexed: 12/05/2022]
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42
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Wei Y, Zhao Z, Ma X. Description of CRISPR-Cas9 development and its prospects in human papillomavirus-driven cancer treatment. Front Immunol 2022; 13:1037124. [PMID: 36479105 PMCID: PMC9721393 DOI: 10.3389/fimmu.2022.1037124] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/05/2022] [Accepted: 10/17/2022] [Indexed: 11/22/2022] Open
Abstract
Human papillomaviruses (HPVs) have been recognized as the etiologic agents of various cancers and are called HPV-driven cancers. Concerning HPV-mediated carcinogenic action, gene therapy can cure cancer at the molecular level by means of the correction of specific genes or sites. CRISPR-Cas9, as a novel genetic editing technique, can correct errors in the genome and change the gene expression and function in cells efficiently, quickly, and with relative ease. Herein, we overviewed studies of CRISPR-mediated gene remedies for HPV-driven cancers and summarized the potential applications of CRISPR-Cas9 in gene therapy for cancer.
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Affiliation(s)
- Yuhao Wei
- Department of Biotherapy, Cancer Center, West China Hospital, Sichuan University, Chengdu, Sichuan, China,West China School of Medicine, West China Hospital, Sichuan University, Chengdu, Sichuan, China
| | - Zhen Zhao
- Department of Biotherapy, Cancer Center, West China Hospital, Sichuan University, Chengdu, Sichuan, China,West China School of Medicine, West China Hospital, Sichuan University, Chengdu, Sichuan, China
| | - Xuelei Ma
- Department of Biotherapy, Cancer Center, West China Hospital, Sichuan University, Chengdu, Sichuan, China,*Correspondence: Xuelei Ma,
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43
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Çerçi B, Uzay IA, Kara MK, Dinçer P. Clinical trials and promising preclinical applications of CRISPR/Cas gene editing. Life Sci 2022; 312:121204. [PMID: 36403643 DOI: 10.1016/j.lfs.2022.121204] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/12/2022] [Revised: 11/03/2022] [Accepted: 11/14/2022] [Indexed: 11/18/2022]
Abstract
Treatment of genetic disorders by genomic manipulation has been the unreachable goal of researchers for many decades. Although our understanding of the genetic basis of genetic diseases has advanced tremendously in the last few decades, the tools developed for genomic editing were not efficient and practical for their use in the clinical setting until now. The recent advancements in the research of Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) and CRISPR-associated protein (Cas) systems offered an easy and efficient way to edit the genome and accelerated the research on their potential use in the treatment of genetic disorders. In this review, we summarize the clinical trials that evaluate the CRISPR/Cas systems for treating different genetic diseases and highlight promising preclinical research on CRISPR/Cas mediated treatment of a great diversity of genetic disorders. Ultimately, we discuss the future of CRISPR/Cas mediated genome editing in genetic diseases.
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Affiliation(s)
- Barış Çerçi
- Faculty of Medicine, Hacettepe University, Ankara 06100, Turkey.
| | - Ihsan Alp Uzay
- Faculty of Medicine, Hacettepe University, Ankara 06100, Turkey
| | | | - Pervin Dinçer
- Department of Medical Biology, Faculty of Medicine, Hacettepe University, Ankara 06100, Turkey
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44
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The Molecular and Cellular Strategies of Glioblastoma and Non-Small-Cell Lung Cancer Cells Conferring Radioresistance. Int J Mol Sci 2022; 23:ijms232113577. [PMID: 36362359 PMCID: PMC9656305 DOI: 10.3390/ijms232113577] [Citation(s) in RCA: 9] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/12/2022] [Revised: 11/02/2022] [Accepted: 11/03/2022] [Indexed: 11/09/2022] Open
Abstract
Ionizing radiation (IR) has been shown to play a crucial role in the treatment of glioblastoma (GBM; grade IV) and non-small-cell lung cancer (NSCLC). Nevertheless, recent studies have indicated that radiotherapy can offer only palliation owing to the radioresistance of GBM and NSCLC. Therefore, delineating the major radioresistance mechanisms may provide novel therapeutic approaches to sensitize these diseases to IR and improve patient outcomes. This review provides insights into the molecular and cellular mechanisms underlying GBM and NSCLC radioresistance, where it sheds light on the role played by cancer stem cells (CSCs), as well as discusses comprehensively how the cellular dormancy/non-proliferating state and polyploidy impact on their survival and relapse post-IR exposure.
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45
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Rittiner J, Cumaran M, Malhotra S, Kantor B. Therapeutic modulation of gene expression in the disease state: Treatment strategies and approaches for the development of next-generation of the epigenetic drugs. Front Bioeng Biotechnol 2022; 10:1035543. [PMID: 36324900 PMCID: PMC9620476 DOI: 10.3389/fbioe.2022.1035543] [Citation(s) in RCA: 12] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/02/2022] [Accepted: 10/05/2022] [Indexed: 11/18/2022] Open
Abstract
Epigenetic dysregulation is an important determinant of many pathological conditions and diseases. Designer molecules that can specifically target endogenous DNA sequences provide a means to therapeutically modulate gene function. The prokaryote-derived CRISPR/Cas editing systems have transformed our ability to manipulate the expression program of genes through specific DNA and RNA targeting in living cells and tissues. The simplicity, utility, and robustness of this technology have revolutionized epigenome editing for research and translational medicine. Initial success has inspired efforts to discover new systems for targeting and manipulating nucleic acids on the epigenetic level. The evolution of nuclease-inactive and RNA-targeting Cas proteins fused to a plethora of effector proteins to regulate gene expression, epigenetic modifications and chromatin interactions opened up an unprecedented level of possibilities for the development of "next-generation" gene therapy therapeutics. The rational design and construction of different types of designer molecules paired with viral-mediated gene-to-cell transfers, specifically using lentiviral vectors (LVs) and adeno-associated vectors (AAVs) are reviewed in this paper. Furthermore, we explore and discuss the potential of these molecules as therapeutic modulators of endogenous gene function, focusing on modulation by stable gene modification and by regulation of gene transcription. Notwithstanding the speedy progress of CRISPR/Cas-based gene therapy products, multiple challenges outlined by undesirable off-target effects, oncogenicity and other virus-induced toxicities could derail the successful translation of these new modalities. Here, we review how CRISPR/Cas-based gene therapy is translated from research-grade technological system to therapeutic modality, paying particular attention to the therapeutic flow from engineering sophisticated genome and epigenome-editing transgenes to delivery vehicles throughout efficient and safe manufacturing and administration of the gene therapy regimens. In addition, the potential solutions to some of the obstacles facing successful CRISPR/Cas utility in the clinical research are discussed in this review. We believe, that circumventing these challenges will be essential for advancing CRISPR/Cas-based tools towards clinical use in gene and cell therapies.
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Affiliation(s)
- Joseph Rittiner
- Department of Neurobiology, Duke University Medical Center, Durham, NC, United States
- Viral Vector Core, Duke University Medical Center, Durham, NC, United States
- Duke Center for Advanced Genomic Technologies, Durham, NC, United States
| | - Mohanapriya Cumaran
- Department of Neurobiology, Duke University Medical Center, Durham, NC, United States
- Viral Vector Core, Duke University Medical Center, Durham, NC, United States
- Duke Center for Advanced Genomic Technologies, Durham, NC, United States
| | - Sahil Malhotra
- Department of Neurobiology, Duke University Medical Center, Durham, NC, United States
- Viral Vector Core, Duke University Medical Center, Durham, NC, United States
- Duke Center for Advanced Genomic Technologies, Durham, NC, United States
| | - Boris Kantor
- Department of Neurobiology, Duke University Medical Center, Durham, NC, United States
- Viral Vector Core, Duke University Medical Center, Durham, NC, United States
- Duke Center for Advanced Genomic Technologies, Durham, NC, United States
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46
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Sowa DJ, Warner MM, Tetenych A, Koechlin L, Balari P, Rascon Perez JP, Caba C, Andres SN. The Mycobacterium tuberculosis Ku C-terminus is a multi-purpose arm for binding DNA and LigD and stimulating ligation. Nucleic Acids Res 2022; 50:11040-11057. [PMID: 36250639 PMCID: PMC9638933 DOI: 10.1093/nar/gkac906] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/06/2021] [Revised: 09/20/2022] [Accepted: 10/05/2022] [Indexed: 11/13/2022] Open
Abstract
Bacterial non-homologous end joining requires the ligase, LigD and Ku. Ku finds the break site, recruits LigD, and then assists LigD to seal the phosphodiester backbone. Bacterial Ku contains a core domain conserved with eukaryotes but has a unique C-terminus that can be divided into a minimal C-terminal region that is conserved and an extended C-terminal region that varies in sequence and length between species. Here, we examine the role of Mycobacterium tuberculosis Ku C-terminal variants, where we removed either the extended or entire C-terminus to investigate the effects on Ku–DNA binding, rates of Ku-stimulated ligation, and binding affinity of a direct Ku–LigD interaction. We find that the extended C-terminus limits DNA binding and identify key amino acids that contribute to this effect through alanine-scanning mutagenesis. The minimal C-terminus is sufficient to stimulate ligation of double-stranded DNA, but the Ku core domain also contributes to stimulating ligation. We further show that wildtype Ku and the Ku core domain alone directly bind both ligase and polymerase domains of LigD. Our results suggest that Ku-stimulated ligation involves direct interactions between the Ku core domain and the LigD ligase domain, in addition to the extended Ku C-terminus and the LigD polymerase domain.
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Affiliation(s)
- Dana J Sowa
- Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada.,Michael DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario L8S 4L8, Canada
| | - Monica M Warner
- Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada.,Michael DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario L8S 4L8, Canada
| | - Andriana Tetenych
- Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada.,Michael DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario L8S 4L8, Canada
| | - Lucas Koechlin
- Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada.,Michael DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario L8S 4L8, Canada
| | - Pardis Balari
- Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada
| | - Jose Pablo Rascon Perez
- Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada
| | - Cody Caba
- Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada.,Michael DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario L8S 4L8, Canada
| | - Sara N Andres
- Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario L8S 4K1, Canada.,Michael DeGroote Institute for Infectious Disease Research, McMaster University, Hamilton, Ontario L8S 4L8, Canada
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47
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Advances in CRISPR/Cas9. BIOMED RESEARCH INTERNATIONAL 2022; 2022:9978571. [PMID: 36193328 PMCID: PMC9525763 DOI: 10.1155/2022/9978571] [Citation(s) in RCA: 9] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 06/02/2022] [Revised: 08/09/2022] [Accepted: 08/22/2022] [Indexed: 11/30/2022]
Abstract
CRISPR/Cas9 technology has become the most examined gene editing technology in recent years due to its simple design, yet low cost, high efficiency, and simple operation, which can also achieve simultaneous editing of multiple loci. It can also be carried out without using plasmids, saving lots of troubles caused by plasmids. CRISPR/Cas9 has shown great potential in the study of genes or genomic functions in microorganisms, plants, animals, and human beings. In this review, we will examine the history, structure, and basic mechanisms of the CRISPR/Cas9 system, describe its great value in precision medicine and sgRNA library screening, and dig its great potential in a new field: DNA information storage.
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48
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Tong L, Yu H, Huang X, Shen J, Xiao G, Chen L, Wang H, Xing L, Chen D. Current understanding of osteoarthritis pathogenesis and relevant new approaches. Bone Res 2022; 10:60. [PMID: 36127328 PMCID: PMC9489702 DOI: 10.1038/s41413-022-00226-9] [Citation(s) in RCA: 195] [Impact Index Per Article: 65.0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/04/2022] [Revised: 05/27/2022] [Accepted: 06/19/2022] [Indexed: 12/20/2022] Open
Abstract
Osteoarthritis (OA) is the most common degenerative joint disease that causes painful swelling and permanent damage to the joints in the body. The molecular mechanisms of OA are currently unknown. OA is a heterogeneous disease that affects the entire joint, and multiple tissues are altered during OA development. To better understand the pathological mechanisms of OA, new approaches, methods, and techniques need to be used to understand OA pathogenesis. In this review, we first focus on the epigenetic regulation of OA, with a particular focus on DNA methylation, histone modification, and microRNA regulation, followed by a summary of several key mediators in OA-associated pain. We then introduce several innovative techniques that have been and will continue to be used in the fields of OA and OA-associated pain, such as CRISPR, scRNA sequencing, and lineage tracing. Next, we discuss the timely updates concerning cell death regulation in OA pathology, including pyroptosis, ferroptosis, and autophagy, as well as their individual roles in OA and potential molecular targets in treating OA. Finally, our review highlights new directions on the role of the synovial lymphatic system in OA. An improved understanding of OA pathogenesis will aid in the development of more specific and effective therapeutic interventions for OA.
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Affiliation(s)
- Liping Tong
- Research Center for Computer-aided Drug Discovery, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518005, China
| | - Huan Yu
- Research Center for Computer-aided Drug Discovery, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518005, China
- Faculty of Pharmaceutical Sciences, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518055, China
| | - Xingyun Huang
- Research Center for Computer-aided Drug Discovery, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518005, China
- Faculty of Pharmaceutical Sciences, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518055, China
| | - Jie Shen
- Department of Orthopedic Surgery, School of Medicine, Washington University in St. Louis, St. Louis, MO, 63110, USA
| | - Guozhi Xiao
- School of Medicine, Southern University of Science and Technology, Shenzhen, 518055, China
| | - Lin Chen
- Department of Wound Repair and Rehabilitation, State Key Laboratory of Trauma, Burns and Combined Injury, Daping Hospital, Army Medical University, Chongqing, 400042, China
| | - Huaiyu Wang
- Research Center for Human Tissues and Organs Degeneration, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518055, China
| | - Lianping Xing
- Department of Pathology and Laboratory of Medicine, Center for Musculoskeletal Research, University of Rochester Medical Center, Rochester, NY, 14642, USA
| | - Di Chen
- Research Center for Computer-aided Drug Discovery, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518005, China.
- Faculty of Pharmaceutical Sciences, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen, 518055, China.
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49
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Past, Present, and Future of Genome Modification in Escherichia coli. Microorganisms 2022; 10:microorganisms10091835. [PMID: 36144436 PMCID: PMC9504249 DOI: 10.3390/microorganisms10091835] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/21/2022] [Revised: 09/05/2022] [Accepted: 09/05/2022] [Indexed: 12/04/2022] Open
Abstract
Escherichia coli K-12 is one of the most well-studied species of bacteria. This species, however, is much more difficult to modify by homologous recombination (HR) than other model microorganisms. Research on HR in E. coli has led to a better understanding of the molecular mechanisms of HR, resulting in technical improvements and rapid progress in genome research, and allowing whole-genome mutagenesis and large-scale genome modifications. Developments using λ Red (exo, bet, and gam) and CRISPR-Cas have made E. coli as amenable to genome modification as other model microorganisms, such as Saccharomyces cerevisiae and Bacillus subtilis. This review describes the history of recombination research in E. coli, as well as improvements in techniques for genome modification by HR. This review also describes the results of large-scale genome modification of E. coli using these technologies, including DNA synthesis and assembly. In addition, this article reviews recent advances in genome modification, considers future directions, and describes problems associated with the creation of cells by design.
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50
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Abstract
ABSTRACT Preoperative neoadjuvant chemoradiotherapy, combined with total mesorectal excision, has become the standard treatment for advanced localized rectal cancer (RC). However, the biological complexity and heterogeneity of tumors may contribute to cancer recurrence and metastasis in patients with radiotherapy-resistant RC. The identification of factors leading to radioresistance and markers of radiosensitivity is critical to identify responsive patients and improve radiotherapy outcomes. MicroRNAs (miRNAs) are small, endogenous, and noncoding RNAs that affect various cellular and molecular targets. miRNAs have been shown to play important roles in multiple biological processes associated with RC. In this review, we summarized the signaling pathways of miRNAs, including apoptosis, autophagy, the cell cycle, DNA damage repair, proliferation, and metastasis during radiotherapy in patients with RC. Also, we evaluated the potential role of miRNAs as radiotherapeutic biomarkers for RC.
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