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Grant-Bier J, Ruppert K, Hayward B, Usdin K, Kumari D. MSH2 is not required for either maintenance of DNA methylation or repeat contraction at the FMR1 locus in fragile X syndrome or the FXN locus in Friedreich's ataxia. Epigenetics Chromatin 2025; 18:24. [PMID: 40296143 PMCID: PMC12036138 DOI: 10.1186/s13072-025-00588-4] [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: 12/20/2024] [Accepted: 04/11/2025] [Indexed: 04/30/2025] Open
Abstract
BACKGROUND Repeat-induced epigenetic changes are observed in many repeat expansion disorders (REDs). These changes result in transcriptional deficits and/or silencing of the associated gene. MSH2, a mismatch repair protein that is required for repeat expansion in the REDs, has been implicated in the maintenance of DNA methylation seen in the region upstream of the expanded CTG repeats at the DMPK locus in myotonic dystrophy type 1 (DM1). Here, we investigated the role of MSH2 in aberrant DNA methylation in two additional REDs, fragile X syndrome (FXS) that is caused by a CGG repeat expansion in the 5' untranslated region (UTR) of the Fragile X Messenger Ribonucleoprotein 1 (FMR1) gene, and Friedreich's ataxia (FRDA) that is caused by a GAA repeat expansion in intron 1 of the frataxin (FXN) gene. RESULTS In contrast to what is seen at the DMPK locus in DM1, loss of MSH2 did not decrease DNA methylation at the FMR1 promoter in FXS embryonic stem cells (ESCs) or increase FMR1 transcription. This difference was not due to the differences in the CpG density of the two loci as a decrease in DNA methylation was also not observed in a less CpG dense region upstream of the expanded GAA repeats in the FXN gene in MSH2 null induced pluripotent stem cells (iPSCs) derived from FRDA patient fibroblasts. Surprisingly, given previous reports, we found that FMR1 reactivation was associated with a high frequency of MSH2-independent CGG-repeat contractions that resulted a permanent loss of DNA methylation. MSH2-independent GAA-repeat contractions were also seen in FRDA cells. CONCLUSIONS Our results suggest that there are mechanistic differences in the way that DNA methylation is maintained in the region upstream of expanded repeats among different REDs even though they share a similar mechanism of repeat expansion. The high frequency of transcription-induced MSH2-dependent and MSH2-independent contractions we have observed may contribute to the mosaicism that is frequently seen in carriers of FMR1 alleles with expanded CGG-repeat tracts. These contractions may reflect the underlying problems associated with transcription through the repeat. Given the recent interest in the therapeutic use of transcription-driven repeat contractions, our data may have interesting mechanistic, prognostic, and therapeutic implications.
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Affiliation(s)
- Jessalyn Grant-Bier
- Section on Gene Structure and Disease, Laboratory of Cell and Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, 20892, USA
- Present address: Cellular and Molecular Biology Program, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, 19104, USA
| | - Kathryn Ruppert
- Section on Gene Structure and Disease, Laboratory of Cell and Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, 20892, USA
| | - Bruce Hayward
- Section on Gene Structure and Disease, Laboratory of Cell and Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, 20892, USA
| | - Karen Usdin
- Section on Gene Structure and Disease, Laboratory of Cell and Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, 20892, USA
| | - Daman Kumari
- Section on Gene Structure and Disease, Laboratory of Cell and Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, 20892, USA.
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Tithi TI, Mo J, Borcherding N, Jo S, Kates HR, Cho E, Cash KE, Honda M, Wang L, Ahmed KK, Shirlekar K, Chen L, Gibson-Corley K, Weigel R, Spies M, Kolb R, Zhang W. The distinct roles of MSH2 and MLH1 in basal-like breast cancer and immune modulation. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2025:2023.07.20.549745. [PMID: 37745359 PMCID: PMC10515760 DOI: 10.1101/2023.07.20.549745] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 09/26/2023]
Abstract
The mismatch repair (MMR) pathway is known as a tumor suppressive pathway and genes involved in MMR are commonly mutated in hereditary colorectal or other cancer types. However, the function of MMR genes/proteins in breast cancer progression and metastasis are largely undefined. We found that MSH2, but not MLH1, is highly enriched in basal-like breast cancer (BLBC) and that its protein expression is inversely correlated with overall survival time (OS). MSH2 expression is frequently elevated due to genomic amplification or gain-of-expression in BLBC, which results in increased MSH2 protein to pair with MSH6 (collectively referred to as MutSα). Genetic deletion of MSH2 or MLH1 results in a contrasting phenotype in metastasis, with MSH2 -deletion leading to reduced metastasis and MLH1 -deletion to enhanced liver or lung metastasis. Mechanistically, MSH2 - but not MLH1 - binds to the promoter region of interferon α receptor 1 ( IFNAR1 ) and suppresses its expression in BLBC. Deletion of MSH2 initiates a chain of immune reactions via the upregulation of IFNAR1 expression and the activation of type 1 interferon signaling, which explains a highly immune active tumor microenvironment in tumors with MSH2-deficiency. Our study supports the contrasting functions of MSH2 and MLH1 in BLBC progression and metastasis due to the differential regulation of IFNAR1 expression, which challenges the paradigm of the MMR pathway as a universal tumor suppressive mechanism.
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Mentani A, Maresca M, Shiriaeva A. Prime Editing: Mechanistic Insights and DNA Repair Modulation. Cells 2025; 14:277. [PMID: 39996750 PMCID: PMC11853414 DOI: 10.3390/cells14040277] [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: 12/12/2024] [Revised: 01/15/2025] [Accepted: 01/24/2025] [Indexed: 02/26/2025] Open
Abstract
Prime editing is a genome editing technique that allows precise modifications of cellular DNA without relying on donor DNA templates. Recently, several different prime editor proteins have been published in the literature, relying on single- or double-strand breaks. When prime editing occurs, the DNA undergoes one of several DNA repair pathways, and these processes can be modulated with the use of inhibitors. Firstly, this review provides an overview of several DNA repair mechanisms and their modulation by known inhibitors. In addition, we summarize different published prime editors and provide a comprehensive overview of associated DNA repair mechanisms. Finally, we discuss the delivery and safety aspects of prime editing.
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Affiliation(s)
- Astrid Mentani
- Genome Engineering, Discovery Science, BioPharmaceuticals R&D, AstraZeneca, 43183 Mölndal, Sweden;
| | | | - Anna Shiriaeva
- Genome Engineering, Discovery Science, BioPharmaceuticals R&D, AstraZeneca, 43183 Mölndal, Sweden;
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Marks SA, Zhou ZX, Lujan SA, Burkholder AB, Kunkel TA. Evidence that DNA polymerase δ proofreads errors made by DNA polymerase α across the Saccharomyces cerevisiae nuclear genome. DNA Repair (Amst) 2024; 143:103768. [PMID: 39332392 DOI: 10.1016/j.dnarep.2024.103768] [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: 07/10/2024] [Revised: 09/09/2024] [Accepted: 09/17/2024] [Indexed: 09/29/2024]
Abstract
We show that the rates of single base substitutions, additions, and deletions across the nuclear genome are strongly increased in a strain harboring a mutator variant of DNA polymerase α combined with a mutation that inactivates the 3´-5´ exonuclease activity of DNA polymerase δ. Moreover, tetrad dissections attempting to produce a haploid triple mutant lacking Msh6, which is essential for DNA mismatch repair (MMR) of base•base mismatches made during replication, result in tiny colonies that grow very slowly and appear to be aneuploid and/or defective in oxidative metabolism. These observations are consistent with the hypothesis that during initiation of nuclear DNA replication, single-base mismatches made by naturally exonuclease-deficient DNA polymerase α are extrinsically proofread by DNA polymerase δ, such that in the absence of this proofreading, the mutation rate is strongly elevated. Several implications of these data are discussed, including that the mutational signature of defective extrinsic proofreading in yeast could appear in human tumors.
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Affiliation(s)
- Sarah A Marks
- Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, DHHS, Research Triangle Park, NC 27709, USA
| | - Zhi-Xiong Zhou
- Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, DHHS, Research Triangle Park, NC 27709, USA
| | - Scott A Lujan
- Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, DHHS, Research Triangle Park, NC 27709, USA
| | - Adam B Burkholder
- Office of Environmental Science Cyberinfrastructure, National Institute of Environmental Health Sciences, National Institutes of Health, DHHS, Research Triangle Park, NC 27709, USA
| | - Thomas A Kunkel
- Genome Integrity and Structural Biology Laboratory, National Institute of Environmental Health Sciences, National Institutes of Health, DHHS, Research Triangle Park, NC 27709, USA.
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Zhang Z, Ma S, Li S, Chen Z, Song R, Wang Z. MSH6 germline mutations leading to Lynch syndrome-associated cholangiocarcinoma: a case report. Front Oncol 2024; 14:1414665. [PMID: 39161380 PMCID: PMC11330888 DOI: 10.3389/fonc.2024.1414665] [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: 04/09/2024] [Accepted: 07/22/2024] [Indexed: 08/21/2024] Open
Abstract
Lynch syndrome, a hereditary cancer susceptibility syndrome, arises from pathogenic mutations in mismatch repair genes. This syndrome is strongly linked to colorectal and endometrial cancers, as well as an elevated risk for other cancers such as gastric, ovarian, renal pelvis/ureter, and prostate. Notably, Lynch syndrome is rarely associated with cholangiocarcinoma (CCA). In this case study, we present a unique instance of Lynch syndrome-related CCA resulting from a singular MSH6 mutation. Notably, our findings reveal discrepancies between immunohistochemistry (IHC) and microsatellite stability results compared to genetic testing outcomes. This discrepancy underscores the limitations of solely relying on IHC analysis and microsatellite stability testing for the detection of hereditary tumors, emphasizing the crucial role of genetic testing in such cases. This insight enhances our comprehension of the mechanisms involved in cancer development and underscores the significance of thorough analysis integrating immunohistochemistry and genetic testing for diagnosing Lynch syndrome-related cancers.
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Affiliation(s)
| | | | | | | | | | - Zhanpeng Wang
- Department of hepatobiliary and pancreatic surgery, China-Japan Union Hospital, Jilin University, Changchun, China
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Kadyrova LY, Mieczkowski PA, Kadyrov FA. MutLα suppresses error-prone DNA mismatch repair and preferentially protects noncoding DNA from mutations. J Biol Chem 2024; 300:107406. [PMID: 38782208 PMCID: PMC11231602 DOI: 10.1016/j.jbc.2024.107406] [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: 04/04/2024] [Revised: 05/15/2024] [Accepted: 05/16/2024] [Indexed: 05/25/2024] Open
Abstract
The DNA mismatch repair (MMR) system promotes genome stability and protects humans from certain types of cancer. Its primary function is the correction of DNA polymerase errors. MutLα is an important eukaryotic MMR factor. We have examined the contributions of MutLα to maintaining genome stability. We show here that loss of MutLα in yeast increases the genome-wide mutation rate by ∼130-fold and generates a genome-wide mutation spectrum that consists of small indels and base substitutions. We also show that loss of yeast MutLα leads to error-prone MMR that produces T > C base substitutions in 5'-ATA-3' sequences. In agreement with this finding, our examination of human whole-genome DNA sequencing data has revealed that loss of MutLα in induced pluripotent stem cells triggers error-prone MMR that leads to the formation of T > C mutations in 5'-NTN-3' sequences. Our further analysis has shown that MutLα-independent MMR plays a role in suppressing base substitutions in N3 homopolymeric runs. In addition, we describe that MutLα preferentially protects noncoding DNA from mutations. Our study defines the contributions of MutLα-dependent and independent mechanisms to genome-wide MMR.
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Affiliation(s)
- Lyudmila Y Kadyrova
- Division of Biochemistry and Molecular Biology, Department of Biomedical Sciences, Southern Illinois University School of Medicine, Carbondale, Illinois, USA
| | - Piotr A Mieczkowski
- Department of Genetics, Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, North Carolina, USA
| | - Farid A Kadyrov
- Division of Biochemistry and Molecular Biology, Department of Biomedical Sciences, Southern Illinois University School of Medicine, Carbondale, Illinois, USA.
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7
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Kadyrova LY, Mieczkowski PA, Kadyrov FA. MutLα suppresses error-prone DNA mismatch repair and preferentially protects noncoding DNA from mutations. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.04.01.587563. [PMID: 38617288 PMCID: PMC11014525 DOI: 10.1101/2024.04.01.587563] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 04/16/2024]
Abstract
The DNA mismatch repair (MMR) system promotes genome stability and protects humans from certain types of cancer. Its primary function is the correction of DNA polymerase errors. MutLα is an important eukaryotic MMR factor. We have examined the contributions of MutLα to maintaining genome stability. We show here that loss of MutLα in yeast increases the genome-wide mutation rate by ~130-fold and generates a genome-wide mutation spectrum that consists of small indels and base substitutions. We also show that loss of yeast MutLα leads to error-prone MMR that produces T>C base substitutions in 5'-ATA-3' sequences. In agreement with this finding, our examination of human whole genome DNA sequencing data has revealed that loss of MutLα in induced pluripotent stem cells triggers error-prone MMR that leads to the formation of T>C mutations in 5'-NTN-3' sequences. Our further analysis has shown that MutLα-independent MMR plays a role in suppressing base substitutions in N3 homopolymeric runs. In addition, we describe that MutLα preferentially defends noncoding DNA from mutations. Our study defines the contributions of MutLα-dependent and independent mechanisms to genome-wide MMR.
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Affiliation(s)
- Lyudmila Y. Kadyrova
- Division of Biochemistry and Molecular Biology, Department of Biomedical Sciences, Southern Illinois University School of Medicine, Carbondale, IL 62901, USA
| | - Piotr A. Mieczkowski
- Department of Genetics, Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, North Carolina 27599, USA
| | - Farid A. Kadyrov
- Division of Biochemistry and Molecular Biology, Department of Biomedical Sciences, Southern Illinois University School of Medicine, Carbondale, IL 62901, USA
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Li Y, Zhang Y, Shah SB, Chang CY, Wang H, Wu X. MutSβ protects common fragile sites by facilitating homology-directed repair at DNA double-strand breaks with secondary structures. Nucleic Acids Res 2024; 52:1120-1135. [PMID: 38038265 PMCID: PMC10853791 DOI: 10.1093/nar/gkad1112] [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: 08/10/2023] [Revised: 10/14/2023] [Accepted: 11/07/2023] [Indexed: 12/02/2023] Open
Abstract
Common fragile sites (CFSs) are regions prone to chromosomal rearrangements, thereby contributing to tumorigenesis. Under replication stress (RS), CFSs often harbor under-replicated DNA regions at the onset of mitosis, triggering homology-directed repair known as mitotic DNA synthesis (MiDAS) to complete DNA replication. In this study, we identified an important role of DNA mismatch repair protein MutSβ (MSH2/MSH3) in facilitating MiDAS and maintaining CFS stability. Specifically, we demonstrated that MutSβ is required for the increased mitotic recombination induced by RS or FANCM loss at CFS-derived AT-rich and structure-prone sequences (CFS-ATs). We also found that MSH3 exhibits synthetic lethality with FANCM. Mechanistically, MutSβ is required for homologous recombination (HR) especially when DNA double-strand break (DSB) ends contain secondary structures. We also showed that upon RS, MutSβ is recruited to Flex1, a specific CFS-AT, in a PCNA-dependent but MUS81-independent manner. Furthermore, MutSβ interacts with RAD52 and promotes RAD52 recruitment to Flex1 following MUS81-dependent fork cleavage. RAD52, in turn, recruits XPF/ERCC1 to remove DNA secondary structures at DSB ends, enabling HR/break-induced replication (BIR) at CFS-ATs. We propose that the specific requirement of MutSβ in processing DNA secondary structures at CFS-ATs underlies its crucial role in promoting MiDAS and maintaining CFS integrity.
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Affiliation(s)
- Youhang Li
- Beijing Key Laboratory of DNA Damage Response and College of Life Sciences, Capital Normal University, Beijing 100048, China
- Department of Molecular Medicine, The Scripps Research Institute, La Jolla, CA 92037, USA
| | - Yunkun Zhang
- Beijing Key Laboratory of DNA Damage Response and College of Life Sciences, Capital Normal University, Beijing 100048, China
| | - Sameer Bikram Shah
- Department of Molecular Medicine, The Scripps Research Institute, La Jolla, CA 92037, USA
| | - Chia-Yu Chang
- Department of Molecular Medicine, The Scripps Research Institute, La Jolla, CA 92037, USA
| | - Hailong Wang
- Beijing Key Laboratory of DNA Damage Response and College of Life Sciences, Capital Normal University, Beijing 100048, China
| | - Xiaohua Wu
- Department of Molecular Medicine, The Scripps Research Institute, La Jolla, CA 92037, USA
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Pretta A, Ziranu P, Giampieri R, Pinna G, Randon G, Donisi C, Ravarino A, Loi F, Deias G, Palmas E, Pretta G, Morano F, Semonella F, Mariani S, Deidda MA, Pusceddu V, Puzzoni M, Lai E, Solinas C, Restivo A, Zorcolo L, Barbara R, Berardi R, Faa G, Pietrantonio F, Scartozzi M. Mismatch Repair system protein deficiency as a resistance factor for locally advanced rectal adenocarcinoma patients receiving neoadjuvant chemo-radiotherapy. Br J Cancer 2023; 129:1619-1624. [PMID: 37749283 PMCID: PMC10646038 DOI: 10.1038/s41416-023-02444-2] [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: 03/21/2023] [Accepted: 09/14/2023] [Indexed: 09/27/2023] Open
Abstract
BACKGROUND Available data on Mismatch Repair system (MMR) deficiency are conflicting and derived from small studies. Our study aimed to evaluate the therapeutic implications of MMR status in patients with locally advanced rectal cancer (LARC). METHODS We retrospectively collected data from 318 patients affected by LARC treated in Italy at the Medical Oncology Units of the University Hospital of Cagliari, Istituto Nazionale dei Tumori Milan, and AOU Ospedali Riuniti Ancona. All patients underwent neoadjuvant chemoradiotherapy. The primary objective was major TRG while secondary objectives were pathological complete response, disease-free survival (DFS) and overall survival (OS). RESULTS One hundred sixty patients (148 pMMR and 12 dMMR) were included in the exploratory cohort and 158 (146 pMMR and 12 dMMR) were included in the validation cohort. A major TRG has been shown in 42.6% and 43.1% patients with pMMR in exploratory and validation cohort, respectively; while no major TRG have been shown in dMMR patients in both cohorts. Exploratory and validation cohorts showed a statistically significant higher mDFS in pMMR patients compared to dMMR: NR vs. 14 months and NR vs. 17 months, respectively. CONCLUSION Our results indicated an association between dMMR and poor response to preoperative chemoradiotherapy and they represent a hypothesis-generating data for new neoadjuvant strategies.
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Affiliation(s)
- Andrea Pretta
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy.
| | - Pina Ziranu
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy
| | - Riccardo Giampieri
- Medical Oncology Unit, Azienda Ospedaliera Universitaria delle Marche, Ancona, Italy
| | - Giovanna Pinna
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy
| | - Giovanni Randon
- Medical Oncology Department, Fondazione IRCCS Istituto Nazionale dei Tumori, Milano, Italy
| | - Clelia Donisi
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy
| | - Alberto Ravarino
- UOC Anatomia Patologica, AOU Cagliari, University of Cagliari, Cagliari, Italy
| | - Francesco Loi
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy
| | - Giulia Deias
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy
| | - Enrico Palmas
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy
| | - Gianluca Pretta
- Science Department, King's School Hove, Hangleton Way, Hangleton, East Sussex, BN3 8BN, UK
| | - Federica Morano
- Medical Oncology Department, Fondazione IRCCS Istituto Nazionale dei Tumori, Milano, Italy
| | - Francesca Semonella
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy
| | - Stefano Mariani
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy
| | | | - Valeria Pusceddu
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy
| | - Marco Puzzoni
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy
| | - Eleonora Lai
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy
| | - Cinzia Solinas
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy
| | - Angelo Restivo
- Department of General Surgery, University Hospital and University of Cagliari, Cagliari, Italy
| | - Luigi Zorcolo
- Department of General Surgery, University Hospital and University of Cagliari, Cagliari, Italy
| | - Raffaele Barbara
- UOC Radioterapia Oncologica, Azienda Ospedaliera "Brotzu", Cagliari, Italy
| | - Rossana Berardi
- Medical Oncology Unit, Azienda Ospedaliera Universitaria delle Marche, Ancona, Italy
| | - Gavino Faa
- UOC Anatomia Patologica, AOU Cagliari, University of Cagliari, Cagliari, Italy
| | - Filippo Pietrantonio
- Medical Oncology Department, Fondazione IRCCS Istituto Nazionale dei Tumori, Milano, Italy
| | - Mario Scartozzi
- Medical Oncology Unit, University Hospital and University of Cagliari, Cagliari, Italy
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Oh JM, Kang Y, Park J, Sung Y, Kim D, Seo Y, Lee E, Ra J, Amarsanaa E, Park YU, Lee S, Hwang J, Kim H, Schärer O, Cho S, Lee C, Takata KI, Lee J, Myung K. MSH2-MSH3 promotes DNA end resection during homologous recombination and blocks polymerase theta-mediated end-joining through interaction with SMARCAD1 and EXO1. Nucleic Acids Res 2023; 51:5584-5602. [PMID: 37140056 PMCID: PMC10287916 DOI: 10.1093/nar/gkad308] [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: 08/16/2022] [Revised: 04/04/2023] [Accepted: 04/27/2023] [Indexed: 05/05/2023] Open
Abstract
DNA double-strand break (DSB) repair via homologous recombination is initiated by end resection. The extent of DNA end resection determines the choice of the DSB repair pathway. Nucleases for end resection have been extensively studied. However, it is still unclear how the potential DNA structures generated by the initial short resection by MRE11-RAD50-NBS1 are recognized and recruit proteins, such as EXO1, to DSB sites to facilitate long-range resection. We found that the MSH2-MSH3 mismatch repair complex is recruited to DSB sites through interaction with the chromatin remodeling protein SMARCAD1. MSH2-MSH3 facilitates the recruitment of EXO1 for long-range resection and enhances its enzymatic activity. MSH2-MSH3 also inhibits access of POLθ, which promotes polymerase theta-mediated end-joining (TMEJ). Collectively, we present a direct role of MSH2-MSH3 in the initial stages of DSB repair by promoting end resection and influencing the DSB repair pathway by favoring homologous recombination over TMEJ.
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Affiliation(s)
- Jung-Min Oh
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
- Department of Oral Biochemistry, Dental and Life Science Institute, School of Dentistry, Pusan National University, Yangsan 50612, Republic of Korea
| | - Yujin Kang
- Department of Biological Sciences, Ulsan National Institute of Science and Technology, Ulsan44919, Republic of Korea
| | - Jumi Park
- Department of Biological Sciences, Ulsan National Institute of Science and Technology, Ulsan44919, Republic of Korea
| | - Yubin Sung
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
| | - Dayoung Kim
- Department of Biomedical Engineering, Ulsan National Institute of Science and Technology, Ulsan44919, Republic of Korea
| | - Yuri Seo
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
| | - Eun A Lee
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
| | - Jae Sun Ra
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
| | - Enkhzul Amarsanaa
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
- Department of Biological Sciences, Ulsan National Institute of Science and Technology, Ulsan44919, Republic of Korea
| | - Young-Un Park
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
| | - Seon Young Lee
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
| | - Jung Me Hwang
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
| | - Hongtae Kim
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
- Department of Biological Sciences, Ulsan National Institute of Science and Technology, Ulsan44919, Republic of Korea
| | - Orlando Schärer
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
- Department of Biological Sciences, Ulsan National Institute of Science and Technology, Ulsan44919, Republic of Korea
| | - Seung Woo Cho
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
- Department of Biomedical Engineering, Ulsan National Institute of Science and Technology, Ulsan44919, Republic of Korea
| | - Changwook Lee
- Department of Biological Sciences, Ulsan National Institute of Science and Technology, Ulsan44919, Republic of Korea
| | - Kei-ichi Takata
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
- Department of Biological Sciences, Ulsan National Institute of Science and Technology, Ulsan44919, Republic of Korea
| | - Ja Yil Lee
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
- Department of Biological Sciences, Ulsan National Institute of Science and Technology, Ulsan44919, Republic of Korea
| | - Kyungjae Myung
- Center for Genomic Integrity, Institute for Basic Science (IBS), Ulsan 44919, Republic of Korea
- Department of Biomedical Engineering, Ulsan National Institute of Science and Technology, Ulsan44919, Republic of Korea
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Park JC, Kim YJ, Han JH, Kim D, Park MJ, Kim J, Jang HK, Bae S, Cha HJ. MutSα and MutSβ as size-dependent cellular determinants for prime editing in human embryonic stem cells. MOLECULAR THERAPY. NUCLEIC ACIDS 2023; 32:914-922. [PMCID: PMC10280094 DOI: 10.1016/j.omtn.2023.05.015] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 10/04/2022] [Accepted: 05/10/2023] [Indexed: 06/22/2023]
Abstract
Precise genome editing in human pluripotent stem cells (hPSCs) has potential applications in isogenic disease modeling and ex vivo stem cell therapy, necessitating diverse genome editing tools. However, unlike differentiated somatic cells, hPSCs have unique cellular properties that maintain genome integrity, which largely determine the overall efficiency of an editing tool. Considering the high demand for prime editors (PEs), it is imperative to characterize the key molecular determinants of PE outcomes in hPSCs. Through homozygous knockout (KO) of MMR pathway key proteins MSH2, MSH3, and MSH6, we reveal that MutSα and MutSβ determine PE efficiency in an editing size-dependent manner. Notably, MSH2 perturbation disrupted both MutSα and MutSβ complexes, dramatically escalating PE efficiency from base mispair to 10 bases, up to 50 folds. Similarly, impaired MutSα by MSH6 KO improved editing efficiency from single to three base pairs, while defective MutSβ by MSH3 KO heightened efficiency from three to 10 base pairs. Thus, the size-dependent effect of MutSα and MutSβ on prime editing implies that MMR is a vital PE efficiency determinant in hPSCs and highlights the distinct roles of MutSα and MutSβ in its outcome.
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Affiliation(s)
- Ju-Chan Park
- College of Pharmacy, Seoul National University, Seoul, Republic of Korea
| | - Yun-Jeong Kim
- College of Pharmacy, Seoul National University, Seoul, Republic of Korea
| | - Jun Hee Han
- Department of Chemistry, Hanyang University, Seoul, Republic of Korea
| | - Dayeon Kim
- College of Pharmacy, Seoul National University, Seoul, Republic of Korea
| | - Mihn Jeong Park
- College of Pharmacy, Seoul National University, Seoul, Republic of Korea
| | - Jumee Kim
- College of Pharmacy, Seoul National University, Seoul, Republic of Korea
| | - Hyeon-Ki Jang
- Division of Chemical Engineering and Bioengineering, College of Art Culture and Engineering, Kangwon National University, Chuncheon, South Korea
| | - Sangsu Bae
- College of Medicine, Seoul National University, Seoul, Republic of Korea
| | - Hyuk-Jin Cha
- College of Pharmacy, Seoul National University, Seoul, Republic of Korea
- Research Institute of Pharmaceutical Sciences, Seoul National University, Seoul, Republic of Korea
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12
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Kadyrova LY, Mieczkowski PA, Kadyrov FA. Genome-wide contributions of the MutSα- and MutSβ-dependent DNA mismatch repair pathways to the maintenance of genetic stability in S. cerevisiae. J Biol Chem 2023; 299:104705. [PMID: 37059180 DOI: 10.1016/j.jbc.2023.104705] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/24/2023] [Revised: 04/07/2023] [Accepted: 04/08/2023] [Indexed: 04/16/2023] Open
Abstract
The DNA mismatch repair (MMR) system is a major DNA repair system that suppresses inherited and sporadic cancers in humans. In eukaryotes the MutSα-dependent and MutSβ-dependent MMR pathways correct DNA polymerase errors. Here, we investigated these two pathways on a whole-genome level in S. cerevisiae. We found that inactivation of MutSα-dependent MMR by deletion of the MSH6 gene increases the genome-wide mutation rate by ∼17-fold, and loss of MutSβ-dependent MMR via deletion of MSH3 elevates the genome-wide mutation rate by ∼4-fold. We also found that MutSα-dependent MMR does not show a preference for protecting coding or noncoding DNA from mutations, whereas MutSβ-dependent MMR preferentially protects noncoding DNA from mutations. The most frequent mutations in the msh6Δ strain are C>T transitions, whereas 1-6-bp deletions are the most common genetic alterations in the msh3Δ strain. Strikingly, MutSα-dependent MMR is more important than MutSβ-dependent MMR for protection from 1-bp insertions, while MutSβ-dependent MMR has a more critical role in the defense against 1-bp deletions and 2-6-bp indels. We also determined that a mutational signature of yeast MSH6 loss is similar to mutational signatures of human MMR deficiency. Furthermore, our analysis showed that compared to other 5'-NCN-3' trinucleotides, 5'-GCA-3' trinucleotides are at the highest risk of accumulating C>T transitions at the central position in the msh6Δ cells and that the presence of a G/A base at the -1 position is important for the efficient MutSα-dependent suppression of C>T transitions. Our results highlight key differences between the roles of the MutSα-dependent and MutSβ-dependent MMR pathways.
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Affiliation(s)
- Lyudmila Y Kadyrova
- Department of Biochemistry and Molecular Biology, Southern Illinois University School of Medicine, Carbondale, IL 62901, USA
| | - Piotr A Mieczkowski
- Department of Genetics, Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, North Carolina 27599, USA
| | - Farid A Kadyrov
- Department of Biochemistry and Molecular Biology, Southern Illinois University School of Medicine, Carbondale, IL 62901, USA.
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13
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Ovejero-Sánchez M, González-Sarmiento R, Herrero AB. DNA Damage Response Alterations in Ovarian Cancer: From Molecular Mechanisms to Therapeutic Opportunities. Cancers (Basel) 2023; 15:448. [PMID: 36672401 PMCID: PMC9856346 DOI: 10.3390/cancers15020448] [Citation(s) in RCA: 9] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/14/2022] [Revised: 01/03/2023] [Accepted: 01/04/2023] [Indexed: 01/12/2023] Open
Abstract
The DNA damage response (DDR), a set of signaling pathways for DNA damage detection and repair, maintains genomic stability when cells are exposed to endogenous or exogenous DNA-damaging agents. Alterations in these pathways are strongly associated with cancer development, including ovarian cancer (OC), the most lethal gynecologic malignancy. In OC, failures in the DDR have been related not only to the onset but also to progression and chemoresistance. It is known that approximately half of the most frequent subtype, high-grade serous carcinoma (HGSC), exhibit defects in DNA double-strand break (DSB) repair by homologous recombination (HR), and current evidence indicates that probably all HGSCs harbor a defect in at least one DDR pathway. These defects are not restricted to HGSCs; mutations in ARID1A, which are present in 30% of endometrioid OCs and 50% of clear cell (CC) carcinomas, have also been found to confer deficiencies in DNA repair. Moreover, DDR alterations have been described in a variable percentage of the different OC subtypes. Here, we overview the main DNA repair pathways involved in the maintenance of genome stability and their deregulation in OC. We also recapitulate the preclinical and clinical data supporting the potential of targeting the DDR to fight the disease.
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Affiliation(s)
- María Ovejero-Sánchez
- Institute of Biomedical Research of Salamanca (IBSAL), 37007 Salamanca, Spain
- Molecular Medicine Unit, Department of Medicine, University of Salamanca, 37007 Salamanca, Spain
- Institute of Molecular and Cellular Biology of Cancer (IBMCC), University of Salamanca-Spanish National Research Council, 37007 Salamanca, Spain
| | - Rogelio González-Sarmiento
- Institute of Biomedical Research of Salamanca (IBSAL), 37007 Salamanca, Spain
- Molecular Medicine Unit, Department of Medicine, University of Salamanca, 37007 Salamanca, Spain
- Institute of Molecular and Cellular Biology of Cancer (IBMCC), University of Salamanca-Spanish National Research Council, 37007 Salamanca, Spain
| | - Ana Belén Herrero
- Institute of Biomedical Research of Salamanca (IBSAL), 37007 Salamanca, Spain
- Molecular Medicine Unit, Department of Medicine, University of Salamanca, 37007 Salamanca, Spain
- Institute of Molecular and Cellular Biology of Cancer (IBMCC), University of Salamanca-Spanish National Research Council, 37007 Salamanca, Spain
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14
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Britton BM, London JA, Martin-Lopez J, Jones ND, Liu J, Lee JB, Fishel R. Exploiting the distinctive properties of the bacterial and human MutS homolog sliding clamps on mismatched DNA. J Biol Chem 2022; 298:102505. [PMID: 36126773 PMCID: PMC9597889 DOI: 10.1016/j.jbc.2022.102505] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/22/2021] [Revised: 09/13/2022] [Accepted: 09/15/2022] [Indexed: 12/30/2022] Open
Abstract
MutS homologs (MSHs) are highly conserved core components of DNA mismatch repair. Mismatch recognition provokes ATP-binding by MSH proteins that drives a conformational transition from a short-lived lesion-searching clamp to an extremely stable sliding clamp on the DNA. Here, we have expanded on previous bulk biochemical studies to examine the stability, lifetime, and kinetics of bacterial and human MSH sliding clamps on mismatched DNA using surface plasmon resonance and single-molecule analysis of fluorescently labeled proteins. We found that ATP-bound MSH complexes bound to blocked-end or very long mismatched DNAs were extremely stable over a range of ionic conditions. These observations underpinned the development of a high-throughput Förster resonance energy transfer system that specifically detects the formation of MSH sliding clamps on mismatched DNA. The Förster resonance energy transfer system is capable of distinguishing between HsMSH2-HsMSH3 and HsMSH2-HsMSH6 and appears suitable for chemical inhibitor screens. Taken together, our results provide additional insight into MSH sliding clamps as well as methods to distinguish their functions in mismatch repair.
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Affiliation(s)
- Brooke M Britton
- Department of Cancer Biology and Genetics, The Ohio State University Wexner Medical Center, Columbus, Ohio, USA
| | - James A London
- Department of Cancer Biology and Genetics, The Ohio State University Wexner Medical Center, Columbus, Ohio, USA
| | - Juana Martin-Lopez
- Department of Cancer Biology and Genetics, The Ohio State University Wexner Medical Center, Columbus, Ohio, USA
| | - Nathan D Jones
- Department of Cancer Biology and Genetics, The Ohio State University Wexner Medical Center, Columbus, Ohio, USA
| | - Jiaquan Liu
- Department of Cancer Biology and Genetics, The Ohio State University Wexner Medical Center, Columbus, Ohio, USA
| | - Jong-Bong Lee
- Department of Physics, Pohang University of Science and Technology (POSTECH), Pohang, Korea; Interdisciplinary Bioscience and Bioengineering, POSTECH, Pohang, Korea
| | - Richard Fishel
- Department of Cancer Biology and Genetics, The Ohio State University Wexner Medical Center, Columbus, Ohio, USA.
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15
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Lund Johansen E, Fribert Thusgaard C, Thomassen M, Eriksen Boonen S, Marie Jochumsen K. Germline Pathogenic Variants Associated with Ovarian Cancer: A Historical Overview. Gynecol Oncol Rep 2022; 44:101105. [DOI: 10.1016/j.gore.2022.101105] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/20/2022] [Revised: 11/02/2022] [Accepted: 11/06/2022] [Indexed: 11/09/2022] Open
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16
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Abstract
Roughly 3% of the human genome consists of microsatellites or tracts of short tandem repeats (STRs). These STRs are often unstable, undergoing high-frequency expansions (increases) or contractions (decreases) in the number of repeat units. Some microsatellite instability (MSI) is seen at multiple STRs within a single cell and is associated with certain types of cancer. A second form of MSI is characterised by expansion of a single gene-specific STR and such expansions are responsible for a group of 40+ human genetic disorders known as the repeat expansion diseases (REDs). While the mismatch repair (MMR) pathway prevents genome-wide MSI, emerging evidence suggests that some MMR factors are directly involved in generating expansions in the REDs. Thus, MMR suppresses some forms of expansion while some MMR factors promote expansion in other contexts. This review will cover what is known about the paradoxical effect of MMR on microsatellite expansion in mammalian cells.
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17
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Vali-Pour M, Park S, Espinosa-Carrasco J, Ortiz-Martínez D, Lehner B, Supek F. The impact of rare germline variants on human somatic mutation processes. Nat Commun 2022; 13:3724. [PMID: 35764656 PMCID: PMC9240060 DOI: 10.1038/s41467-022-31483-1] [Citation(s) in RCA: 17] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/05/2021] [Accepted: 06/17/2022] [Indexed: 02/07/2023] Open
Abstract
Somatic mutations are an inevitable component of ageing and the most important cause of cancer. The rates and types of somatic mutation vary across individuals, but relatively few inherited influences on mutation processes are known. We perform a gene-based rare variant association study with diverse mutational processes, using human cancer genomes from over 11,000 individuals of European ancestry. By combining burden and variance tests, we identify 207 associations involving 15 somatic mutational phenotypes and 42 genes that replicated in an independent data set at a false discovery rate of 1%. We associate rare inherited deleterious variants in genes such as MSH3, EXO1, SETD2, and MTOR with two phenotypically different forms of DNA mismatch repair deficiency, and variants in genes such as EXO1, PAXIP1, RIF1, and WRN with deficiency in homologous recombination repair. In addition, we identify associations with other mutational processes, such as APEX1 with APOBEC-signature mutagenesis. Many of the genes interact with each other and with known mutator genes within cellular sub-networks. Considered collectively, damaging variants in the identified genes are prevalent in the population. We suggest that rare germline variation in diverse genes commonly impacts mutational processes in somatic cells.
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Affiliation(s)
- Mischan Vali-Pour
- Centre for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology (BIST), Barcelona, Spain
- Universitat Pompeu Fabra (UPF), Barcelona, Spain
| | - Solip Park
- Centro Nacional de Investigaciones Oncológicas (CNIO), Madrid, Spain
| | - Jose Espinosa-Carrasco
- Institute for Research in Biomedicine (IRB Barcelona), The Barcelona Institute of Science and Technology (BIST), Barcelona, Spain
| | - Daniel Ortiz-Martínez
- Institute for Research in Biomedicine (IRB Barcelona), The Barcelona Institute of Science and Technology (BIST), Barcelona, Spain
| | - Ben Lehner
- Centre for Genomic Regulation (CRG), The Barcelona Institute of Science and Technology (BIST), Barcelona, Spain.
- Universitat Pompeu Fabra (UPF), Barcelona, Spain.
- Catalan Institution for Research and Advanced Studies (ICREA), Barcelona, Spain.
| | - Fran Supek
- Institute for Research in Biomedicine (IRB Barcelona), The Barcelona Institute of Science and Technology (BIST), Barcelona, Spain.
- Catalan Institution for Research and Advanced Studies (ICREA), Barcelona, Spain.
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18
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Genome Integrity and Neurological Disease. Int J Mol Sci 2022; 23:ijms23084142. [PMID: 35456958 PMCID: PMC9025063 DOI: 10.3390/ijms23084142] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/03/2022] [Revised: 04/02/2022] [Accepted: 04/05/2022] [Indexed: 02/06/2023] Open
Abstract
Neurological complications directly impact the lives of hundreds of millions of people worldwide. While the precise molecular mechanisms that underlie neuronal cell loss remain under debate, evidence indicates that the accumulation of genomic DNA damage and consequent cellular responses can promote apoptosis and neurodegenerative disease. This idea is supported by the fact that individuals who harbor pathogenic mutations in DNA damage response genes experience profound neuropathological manifestations. The review article here provides a general overview of the nervous system, the threats to DNA stability, and the mechanisms that protect genomic integrity while highlighting the connections of DNA repair defects to neurological disease. The information presented should serve as a prelude to the Special Issue “Genome Stability and Neurological Disease”, where experts discuss the role of DNA repair in preserving central nervous system function in greater depth.
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19
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The nuclease activity of DNA2 promotes exonuclease 1-independent mismatch repair. J Biol Chem 2022; 298:101831. [PMID: 35300981 PMCID: PMC9036127 DOI: 10.1016/j.jbc.2022.101831] [Citation(s) in RCA: 7] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2021] [Revised: 03/09/2022] [Accepted: 03/10/2022] [Indexed: 12/03/2022] Open
Abstract
The DNA mismatch repair (MMR) system is a major DNA repair system that corrects DNA replication errors. In eukaryotes, the MMR system functions via mechanisms both dependent on and independent of exonuclease 1 (EXO1), an enzyme that has multiple roles in DNA metabolism. Although the mechanism of EXO1-dependent MMR is well understood, less is known about EXO1-independent MMR. Here, we provide genetic and biochemical evidence that the DNA2 nuclease/helicase has a role in EXO1-independent MMR. Biochemical reactions reconstituted with purified human proteins demonstrated that the nuclease activity of DNA2 promotes an EXO1-independent MMR reaction via a mismatch excision-independent mechanism that involves DNA polymerase δ. We show that DNA polymerase ε is not able to replace DNA polymerase δ in the DNA2-promoted MMR reaction. Unlike its nuclease activity, the helicase activity of DNA2 is dispensable for the ability of the protein to enhance the MMR reaction. Further examination established that DNA2 acts in the EXO1-independent MMR reaction by increasing the strand-displacement activity of DNA polymerase δ. These data reveal a mechanism for EXO1-independent mismatch repair.
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20
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Prime editing efficiency and fidelity are enhanced in the absence of mismatch repair. Nat Commun 2022; 13:760. [PMID: 35140211 PMCID: PMC8828784 DOI: 10.1038/s41467-022-28442-1] [Citation(s) in RCA: 95] [Impact Index Per Article: 31.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/06/2021] [Accepted: 01/26/2022] [Indexed: 02/07/2023] Open
Abstract
Prime editing (PE) is a powerful genome engineering approach that enables the introduction of base substitutions, insertions and deletions into any given genomic locus. However, the efficiency of PE varies widely and depends not only on the genomic region targeted, but also on the genetic background of the edited cell. Here, to determine which cellular factors affect PE efficiency, we carry out a focused genetic screen targeting 32 DNA repair factors, spanning all reported repair pathways. We show that, depending on cell line and type of edit, ablation of mismatch repair (MMR) affords a 2–17 fold increase in PE efficiency, across several human cell lines, types of edits and genomic loci. The accumulation of the key MMR factors MLH1 and MSH2 at PE sites argues for direct involvement of MMR in PE control. Our results shed new light on the mechanism of PE and suggest how its efficiency might be optimised. Prime Editing is a versatile genome engineering tool. Here, the authors identify the DNA repair pathway known as mismatch repair as inhibitory for Prime Editing, thus, loss of mismatch repair enhances the efficiency of Prime Editing.
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21
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Barroso-González J, García-Expósito L, Galaviz P, Lynskey ML, Allen JAM, Hoang S, Watkins SC, Pickett HA, O'Sullivan RJ. Anti-recombination function of MutSα restricts telomere extension by ALT-associated homology-directed repair. Cell Rep 2021; 37:110088. [PMID: 34879271 PMCID: PMC8724847 DOI: 10.1016/j.celrep.2021.110088] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/20/2021] [Revised: 09/13/2021] [Accepted: 11/10/2021] [Indexed: 01/02/2023] Open
Abstract
Alternative lengthening of telomeres (ALT) is a telomere-elongation mechanism observed in ~15% of cancer subtypes. Current models indicate that ALT is mediated by homology-directed repair mechanisms. By disrupting MSH6 gene expression, we show that the deficiency of MutSα (MSH2/MSH6) DNA mismatch repair complex causes striking telomere hyperextension. Mechanistically, we show MutSα is specifically recruited to telomeres in ALT cells by associating with the proliferating-cell nuclear antigen (PCNA) subunit of the ALT telomere replisome. We also provide evidence that MutSα counteracts Bloom (BLM) helicase, which adopts a crucial role in stabilizing hyper-extended telomeres and maintaining the survival of MutSα-deficient ALT cancer cells. Lastly, we propose a model in which MutSα deficiency impairs heteroduplex rejection, leading to premature initiation of telomere DNA synthesis that coincides with an accumulation of telomere variant repeats (TVRs). These findings provide evidence that the MutSα DNA mismatch repair complex acts to restrain unwarranted ALT. Barroso-Gonzalez et al. show that the mismatch repair complex MutSα restricts the alternative lengthening of telomeres (ALT) pathway in cancer cells. MutSα has an anti-recombination function and limits recombination between heteroduplex sequences at telomeres, in part by counteracting the Bloom helicase (BLM).
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Affiliation(s)
- Jonathan Barroso-González
- Department of Pharmacology and Chemical Biology, School of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
| | - Laura García-Expósito
- Department of Pharmacology and Chemical Biology, School of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
| | - Pablo Galaviz
- Bioinformatics Unit, Children's Medical Research Institute, Faculty of Medicine and Health, University of Sydney, Westmead, NSW 2145, Australia
| | - Michelle Lee Lynskey
- Department of Pharmacology and Chemical Biology, School of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
| | - Joshua A M Allen
- Telomere Length Regulation Unit, Children's Medical Research Institute, Faculty of Medicine and Health, University of Sydney, Westmead, NSW 2145, Australia
| | - SongMy Hoang
- Department of Pharmacology and Chemical Biology, School of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
| | - Simon C Watkins
- Department of Cell Biology, School of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA
| | - Hilda A Pickett
- Telomere Length Regulation Unit, Children's Medical Research Institute, Faculty of Medicine and Health, University of Sydney, Westmead, NSW 2145, Australia
| | - Roderick J O'Sullivan
- Department of Pharmacology and Chemical Biology, School of Medicine, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA.
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22
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Olkinuora AP, Peltomäki PT, Aaltonen LA, Rajamäki K. From APC to the genetics of hereditary and familial colon cancer syndromes. Hum Mol Genet 2021; 30:R206-R224. [PMID: 34329396 PMCID: PMC8490010 DOI: 10.1093/hmg/ddab208] [Citation(s) in RCA: 19] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/06/2021] [Revised: 07/01/2021] [Accepted: 07/05/2021] [Indexed: 11/12/2022] Open
Abstract
Hereditary colorectal cancer (CRC) syndromes attributable to high penetrance mutations represent 9-26% of young-onset CRC cases. The clinical significance of many of these mutations is understood well enough to be used in diagnostics and as an aid in patient care. However, despite the advances made in the field, a significant proportion of familial and early-onset cases remains molecularly uncharacterized and extensive work is still needed to fully understand the genetic nature of CRC susceptibility. With the emergence of next-generation sequencing and associated methods, several predisposition loci have been unraveled, but validation is incomplete. Individuals with cancer-predisposing mutations are currently enrolled in life-long surveillance, but with the development of new treatments, such as cancer vaccinations, this might change in the not so distant future for at least some individuals. For individuals without a known cause for their disease susceptibility, prevention and therapy options are less precise. Herein, we review the progress achieved in the last three decades with a focus on how CRC predisposition genes were discovered. Furthermore, we discuss the clinical implications of these discoveries and anticipate what to expect in the next decade.
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Affiliation(s)
- Alisa P Olkinuora
- Department of Medical and Clinical Genetics, Medicum, University of Helsinki, 00014 Helsinki, Finland
- iCAN Digital Precision Cancer Medicine Flagship, University of Helsinki, 00014 Helsinki, Finland
| | - Päivi T Peltomäki
- Department of Medical and Clinical Genetics, Medicum, University of Helsinki, 00014 Helsinki, Finland
- iCAN Digital Precision Cancer Medicine Flagship, University of Helsinki, 00014 Helsinki, Finland
| | - Lauri A Aaltonen
- Department of Medical and Clinical Genetics, Medicum, University of Helsinki, 00014 Helsinki, Finland
- iCAN Digital Precision Cancer Medicine Flagship, University of Helsinki, 00014 Helsinki, Finland
- Applied Tumor Genomics Research Program, Research Programs Unit, University of Helsinki, 00014 Helsinki, Finland
| | - Kristiina Rajamäki
- Department of Medical and Clinical Genetics, Medicum, University of Helsinki, 00014 Helsinki, Finland
- Applied Tumor Genomics Research Program, Research Programs Unit, University of Helsinki, 00014 Helsinki, Finland
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23
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Shioi S, Shimamoto A, Song Y, Hidaka K, Nakamura M, Take A, Hayashi N, Takiguchi S, Fujikane R, Hidaka M, Oda S, Nakatsu Y. DNA polymerase delta Exo domain stabilizes mononucleotide microsatellites in human cells. DNA Repair (Amst) 2021; 108:103216. [PMID: 34530183 DOI: 10.1016/j.dnarep.2021.103216] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2021] [Revised: 08/19/2021] [Accepted: 08/26/2021] [Indexed: 01/16/2023]
Abstract
In prokaryotes and yeasts, DNA polymerase proofreading (PPR) and DNA mismatch repair (MMR) cooperatively counteracts replication errors leading to repeat sequence destabilization (i.e. insertions/deletions of repeat units). However, PPR has not thus far been regarded as a mechanism stabilizing repeat sequences in higher eukaryotic cells. In a human cancer cell line, DLD-1, which carries mutations in both MSH6 and the Exo domain of POLD1, we previously observed that mononucleotide microsatellites were markedly destabilized whereas being stable in the simple MMR-defective backgrounds. In this study, we introduced the Exo domain mutation found in DLD-1 cells into MSH2-null HeLa cell clones, using CRISPR/Cas9 system. In the established Exo-/MMR-mutated HeLa clones, mononucleotide repeat sequences were remarkably destabilized as in DLD-1 cells. In contrast, dinucleotide microsatellites were readily destabilized in the parental MMR-deficient backgrounds, and the instability was not notably increased in the genome-edited HeLa clones. Here, we show an involvement of the Exo domain functions of DNA polymerase delta in mononucleotide repeat stabilization in human cells, which also suggests a possible role division between DNA polymerase and MMR in repeat maintenance in the human genome.
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Affiliation(s)
- Seijiro Shioi
- Cancer Genetics Laboratory, Clinical Research Institute, National Hospital Organization Kyushu Cancer Center, Fukuoka, Japan
| | - Akiyoshi Shimamoto
- Cancer Genetics Laboratory, Clinical Research Institute, National Hospital Organization Kyushu Cancer Center, Fukuoka, Japan
| | - Yingxia Song
- Department of Medical Biophysics and Radiation Biology, Faculty of Medical Sciences, Kyushu University, Japan
| | - Kyoko Hidaka
- Centre for Fundamental Education, University of Kitakyushu, Kitakyushu, Japan
| | - Maki Nakamura
- Cancer Genetics Laboratory, Clinical Research Institute, National Hospital Organization Kyushu Cancer Center, Fukuoka, Japan
| | - Ayumi Take
- Cancer Genetics Laboratory, Clinical Research Institute, National Hospital Organization Kyushu Cancer Center, Fukuoka, Japan
| | - Namiko Hayashi
- Cancer Genetics Laboratory, Clinical Research Institute, National Hospital Organization Kyushu Cancer Center, Fukuoka, Japan
| | - Soichi Takiguchi
- Cancer Genetics Laboratory, Clinical Research Institute, National Hospital Organization Kyushu Cancer Center, Fukuoka, Japan
| | - Ryosuke Fujikane
- Department of Physiological Science and Molecular Biology, Fukuoka Dental College, Fukuoka, Japan
| | - Masumi Hidaka
- Department of Physiological Science and Molecular Biology, Fukuoka Dental College, Fukuoka, Japan
| | - Shinya Oda
- Cancer Genetics Laboratory, Clinical Research Institute, National Hospital Organization Kyushu Cancer Center, Fukuoka, Japan.
| | - Yoshimichi Nakatsu
- Department of Medical Biophysics and Radiation Biology, Faculty of Medical Sciences, Kyushu University, Japan.
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24
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Kratz K, Artola-Borán M, Kobayashi-Era S, Koh G, Oliveira G, Kobayashi S, Oliveira A, Zou X, Richter J, Tsuda M, Sasanuma H, Takeda S, Loizou JI, Sartori AA, Nik-Zainal S, Jiricny J. FANCD2-Associated Nuclease 1 Partially Compensates for the Lack of Exonuclease 1 in Mismatch Repair. Mol Cell Biol 2021; 41:e0030321. [PMID: 34228493 PMCID: PMC8384067 DOI: 10.1128/mcb.00303-21] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/23/2021] [Accepted: 06/28/2021] [Indexed: 11/20/2022] Open
Abstract
Germline mutations in the mismatch repair (MMR) genes MSH2, MSH6, MLH1, and PMS2 are linked to cancer of the colon and other organs, characterized by microsatellite instability and a large increase in mutation frequency. Unexpectedly, mutations in EXO1, encoding the only exonuclease genetically implicated in MMR, are not linked to familial cancer and cause a substantially weaker mutator phenotype. This difference could be explained if eukaryotic cells possessed additional exonucleases redundant with EXO1. Analysis of the MLH1 interactome identified FANCD2-associated nuclease 1 (FAN1), a novel enzyme with biochemical properties resembling EXO1. We now show that FAN1 efficiently substitutes for EXO1 in MMR assays and that this functional complementation is modulated by its interaction with MLH1. FAN1 also contributes to MMR in vivo; cells lacking both EXO1 and FAN1 have an MMR defect and display resistance to N-methyl-N-nitrosourea (MNU) and 6-thioguanine (TG). Moreover, FAN1 loss amplifies the mutational profile of EXO1-deficient cells, suggesting that the two nucleases act redundantly in the same antimutagenic pathway. However, the increased drug resistance and mutator phenotype of FAN1/EXO1-deficient cells are less prominent than those seen in cells lacking MSH6 or MLH1. Eukaryotic cells thus apparently possess additional mechanisms that compensate for the loss of EXO1.
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Affiliation(s)
- Katja Kratz
- Institute of Molecular Cancer Research, University of Zurich, Zurich, Switzerland
| | - Mariela Artola-Borán
- Institute of Molecular Cancer Research, University of Zurich, Zurich, Switzerland
| | - Saho Kobayashi-Era
- Institute of Molecular Cancer Research, University of Zurich, Zurich, Switzerland
- Institute of Biochemistry of the ETH Zurich, Zurich, Switzerland
| | - Gene Koh
- Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, United Kingdom
- Academic Department of Medical Genetics, The Clinical School, University of Cambridge, Cambridge, United Kingdom
- MRC Cancer Unit, The Clinical School, University of Cambridge, Cambridge, United Kingdom
| | - Goncalo Oliveira
- Institute of Cancer Research, Department of Medicine I, Comprehensive Cancer Centre, Medical University of Vienna, Vienna, Austria
| | - Shunsuke Kobayashi
- Institute of Molecular Cancer Research, University of Zurich, Zurich, Switzerland
- Institute of Biochemistry of the ETH Zurich, Zurich, Switzerland
| | - Andreia Oliveira
- Institute of Molecular Cancer Research, University of Zurich, Zurich, Switzerland
- Institute of Biochemistry of the ETH Zurich, Zurich, Switzerland
| | - Xueqing Zou
- Wellcome Sanger Institute, Wellcome Genome Campus, Hinxton, United Kingdom
- Academic Department of Medical Genetics, The Clinical School, University of Cambridge, Cambridge, United Kingdom
- MRC Cancer Unit, The Clinical School, University of Cambridge, Cambridge, United Kingdom
| | - Julia Richter
- Institute of Biochemistry of the ETH Zurich, Zurich, Switzerland
| | - Masataka Tsuda
- Department of Radiation Genetics, Graduate School of Medicine, Kyoto University, Kyoto, Japan
| | - Hiroyuki Sasanuma
- Department of Radiation Genetics, Graduate School of Medicine, Kyoto University, Kyoto, Japan
| | - Shunichi Takeda
- Department of Radiation Genetics, Graduate School of Medicine, Kyoto University, Kyoto, Japan
| | - Joanna I. Loizou
- Institute of Cancer Research, Department of Medicine I, Comprehensive Cancer Centre, Medical University of Vienna, Vienna, Austria
| | | | - Serena Nik-Zainal
- Academic Department of Medical Genetics, The Clinical School, University of Cambridge, Cambridge, United Kingdom
- MRC Cancer Unit, The Clinical School, University of Cambridge, Cambridge, United Kingdom
| | - Josef Jiricny
- Institute of Molecular Cancer Research, University of Zurich, Zurich, Switzerland
- Institute of Biochemistry of the ETH Zurich, Zurich, Switzerland
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25
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Zou X, Koh GCC, Nanda AS, Degasperi A, Urgo K, Roumeliotis TI, Agu CA, Badja C, Momen S, Young J, Amarante TD, Side L, Brice G, Perez-Alonso V, Rueda D, Gomez C, Bushell W, Harris R, Choudhary JS, Jiricny J, Skarnes WC, Nik-Zainal S. A systematic CRISPR screen defines mutational mechanisms underpinning signatures caused by replication errors and endogenous DNA damage. NATURE CANCER 2021; 2:643-657. [PMID: 34164627 PMCID: PMC7611045 DOI: 10.1038/s43018-021-00200-0] [Citation(s) in RCA: 91] [Impact Index Per Article: 22.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 08/04/2020] [Accepted: 03/17/2021] [Indexed: 02/02/2023]
Abstract
Mutational signatures are imprints of pathophysiological processes arising through tumorigenesis. We generated isogenic CRISPR-Cas9 knockouts (Δ) of 43 genes in human induced pluripotent stem cells, cultured them in the absence of added DNA damage, and performed whole-genome sequencing of 173 subclones. ΔOGG1, ΔUNG, ΔEXO1, ΔRNF168, ΔMLH1, ΔMSH2, ΔMSH6, ΔPMS1, and ΔPMS2 produced marked mutational signatures indicative of being critical mitigators of endogenous DNA modifications. Detailed analyses revealed mutational mechanistic insights, including how 8-oxo-dG elimination is sequence-context-specific while uracil clearance is sequence-context-independent. Mismatch repair (MMR) deficiency signatures are engendered by oxidative damage (C>A transversions), differential misincorporation by replicative polymerases (T>C and C>T transitions), and we propose a 'reverse template slippage' model for T>A transversions. ΔMLH1, ΔMSH6, and ΔMSH2 signatures were similar to each other but distinct from ΔPMS2. Finally, we developed a classifier, MMRDetect, where application to 7,695 WGS cancers showed enhanced detection of MMR-deficient tumors, with implications for responsiveness to immunotherapies.
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Affiliation(s)
- Xueqing Zou
- Academic Department of Medical Genetics, School of Clinical Medicine, University of Cambridge, Cambridge, UK
- MRC Cancer Unit, University of Cambridge, Cambridge, UK
- Wellcome Sanger Institute, Hinxton, UK
| | - Gene Ching Chiek Koh
- Academic Department of Medical Genetics, School of Clinical Medicine, University of Cambridge, Cambridge, UK
- MRC Cancer Unit, University of Cambridge, Cambridge, UK
- Wellcome Sanger Institute, Hinxton, UK
| | - Arjun Scott Nanda
- Academic Department of Medical Genetics, School of Clinical Medicine, University of Cambridge, Cambridge, UK
- MRC Cancer Unit, University of Cambridge, Cambridge, UK
| | - Andrea Degasperi
- Academic Department of Medical Genetics, School of Clinical Medicine, University of Cambridge, Cambridge, UK
- MRC Cancer Unit, University of Cambridge, Cambridge, UK
- Wellcome Sanger Institute, Hinxton, UK
| | | | | | | | - Cherif Badja
- Academic Department of Medical Genetics, School of Clinical Medicine, University of Cambridge, Cambridge, UK
- MRC Cancer Unit, University of Cambridge, Cambridge, UK
- Wellcome Sanger Institute, Hinxton, UK
| | - Sophie Momen
- Academic Department of Medical Genetics, School of Clinical Medicine, University of Cambridge, Cambridge, UK
- MRC Cancer Unit, University of Cambridge, Cambridge, UK
| | - Jamie Young
- Academic Department of Medical Genetics, School of Clinical Medicine, University of Cambridge, Cambridge, UK
| | - Tauanne Dias Amarante
- Academic Department of Medical Genetics, School of Clinical Medicine, University of Cambridge, Cambridge, UK
- MRC Cancer Unit, University of Cambridge, Cambridge, UK
| | - Lucy Side
- UCL Institute for Women's Health, Great Ormond Street Hospital, London, UK
- Wessex Clinical Genetics Service, Princess Anne Hospital, Southampton, UK
| | - Glen Brice
- Southwest Thames Regional Genetics Service, St George's University of London, London, UK
| | - Vanesa Perez-Alonso
- Pediatrics Department, Doce de Octubre University Hospital, i+12 Research Institute, Madrid, Spain
| | - Daniel Rueda
- Hereditary Cancer Laboratory, Doce de Octubre University Hospital, i+12 Research Institute, Madrid, Spain
| | | | | | - Rebecca Harris
- Academic Department of Medical Genetics, School of Clinical Medicine, University of Cambridge, Cambridge, UK
- Wellcome Sanger Institute, Hinxton, UK
| | - Jyoti S Choudhary
- The Institute of Cancer Research, Chester Beatty Laboratories, London, UK
| | - Josef Jiricny
- Institute of Molecular Life Sciences, University of Zurich, Zurich, Switzerland
- Institute of Biochemistry, ETH Zurich, Zurich, Switzerland
| | - William C Skarnes
- Wellcome Sanger Institute, Hinxton, UK
- William Harvey Research Institute, Queen Mary University of London, London, UK
| | - Serena Nik-Zainal
- Academic Department of Medical Genetics, School of Clinical Medicine, University of Cambridge, Cambridge, UK.
- MRC Cancer Unit, University of Cambridge, Cambridge, UK.
- Wellcome Sanger Institute, Hinxton, UK.
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26
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Ortega J, Lee GS, Gu L, Yang W, Li GM. Mispair-bound human MutS-MutL complex triggers DNA incisions and activates mismatch repair. Cell Res 2021; 31:542-553. [PMID: 33510387 PMCID: PMC8089094 DOI: 10.1038/s41422-021-00468-y] [Citation(s) in RCA: 31] [Impact Index Per Article: 7.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2020] [Accepted: 12/17/2020] [Indexed: 01/30/2023] Open
Abstract
DNA mismatch repair (MMR) relies on MutS and MutL ATPases for mismatch recognition and strand-specific nuclease recruitment to remove mispaired bases in daughter strands. However, whether the MutS-MutL complex coordinates MMR by ATP-dependent sliding on DNA or protein-protein interactions between the mismatch and strand discrimination signal is ambiguous. Using functional MMR assays and systems preventing proteins from sliding, we show that sliding of human MutSα is required not for MMR initiation, but for final mismatch removal. MutSα recruits MutLα to form a mismatch-bound complex, which initiates MMR by nicking the daughter strand 5' to the mismatch. Exonuclease 1 (Exo1) is then recruited to the nick and conducts 5' → 3' excision. ATP-dependent MutSα dissociation from the mismatch is necessary for Exo1 to remove the mispaired base when the excision reaches the mismatch. Therefore, our study has resolved a long-standing puzzle, and provided new insights into the mechanism of MMR initiation and mispair removal.
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Affiliation(s)
- Janice Ortega
- Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, TX USA
| | - Grace Sanghee Lee
- Department of Toxicology and Cancer Biology, University of Kentucky College of Medicine, Lexington, KY USA ,Present Address: Division of Viral Hepatitis, National Center for HIV/AIDS, Viral Hepatitis, STD and TB Prevention, Centers for Disease Control and Prevention, Atlanta, GA USA
| | - Liya Gu
- Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, TX USA
| | - Wei Yang
- Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD USA
| | - Guo-Min Li
- Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, TX USA ,Department of Toxicology and Cancer Biology, University of Kentucky College of Medicine, Lexington, KY USA
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27
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Young SJ, West SC. Coordinated roles of SLX4 and MutSβ in DNA repair and the maintenance of genome stability. Crit Rev Biochem Mol Biol 2021; 56:157-177. [PMID: 33596761 PMCID: PMC7610648 DOI: 10.1080/10409238.2021.1881433] [Citation(s) in RCA: 13] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2020] [Revised: 01/06/2021] [Accepted: 01/22/2021] [Indexed: 12/14/2022]
Abstract
SLX4 provides a molecular scaffold for the assembly of multiple protein complexes required for the maintenance of genome stability. It is involved in the repair of DNA crosslinks, the resolution of recombination intermediates, the response to replication stress and the maintenance of telomere length. To carry out these diverse functions, SLX4 interacts with three structure-selective endonucleases, MUS81-EME1, SLX1 and XPF-ERCC1, as well as the telomere binding proteins TRF2, RTEL1 and SLX4IP. Recently, SLX4 was shown to interact with MutSβ, a heterodimeric protein involved in DNA mismatch repair, trinucleotide repeat instability, crosslink repair and recombination. Importantly, MutSβ promotes the pathogenic expansion of CAG/CTG trinucleotide repeats, which is causative of myotonic dystrophy and Huntington's disease. The colocalization and specific interaction of MutSβ with SLX4, together with their apparently overlapping functions, are suggestive of a common role in reactions that promote DNA maintenance and genome stability. This review will focus on the role of SLX4 in DNA repair, the interplay between MutSβ and SLX4, and detail how they cooperate to promote recombinational repair and DNA crosslink repair. Furthermore, we speculate that MutSβ and SLX4 may provide an alternative cellular mechanism that modulates trinucleotide instability.
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Affiliation(s)
- Sarah J Young
- DNA Recombination and Repair Laboratory, The Francis Crick Institute, London, UK
| | - Stephen C West
- DNA Recombination and Repair Laboratory, The Francis Crick Institute, London, UK
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28
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Gartner A, Engebrecht J. DNA repair, recombination, and damage signaling. Genetics 2021; 220:6522877. [PMID: 35137093 PMCID: PMC9097270 DOI: 10.1093/genetics/iyab178] [Citation(s) in RCA: 35] [Impact Index Per Article: 8.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/10/2021] [Accepted: 10/10/2021] [Indexed: 01/09/2023] Open
Abstract
DNA must be accurately copied and propagated from one cell division to the next, and from one generation to the next. To ensure the faithful transmission of the genome, a plethora of distinct as well as overlapping DNA repair and recombination pathways have evolved. These pathways repair a large variety of lesions, including alterations to single nucleotides and DNA single and double-strand breaks, that are generated as a consequence of normal cellular function or by external DNA damaging agents. In addition to the proteins that mediate DNA repair, checkpoint pathways have also evolved to monitor the genome and coordinate the action of various repair pathways. Checkpoints facilitate repair by mediating a transient cell cycle arrest, or through initiation of cell suicide if DNA damage has overwhelmed repair capacity. In this chapter, we describe the attributes of Caenorhabditis elegans that facilitate analyses of DNA repair, recombination, and checkpoint signaling in the context of a whole animal. We review the current knowledge of C. elegans DNA repair, recombination, and DNA damage response pathways, and their role during development, growth, and in the germ line. We also discuss how the analysis of mutational signatures in C. elegans is helping to inform cancer mutational signatures in humans.
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Affiliation(s)
- Anton Gartner
- Department for Biological Sciences, IBS Center for Genomic Integrity, Ulsan National Institute of Science and Technology, Ulsan 689-798, Republic of Korea,Corresponding author: (A.G.); (J.E.)
| | - JoAnne Engebrecht
- Department of Molecular and Cellular Biology, University of California Davis, Davis, CA 95616, USA,Corresponding author: (A.G.); (J.E.)
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29
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Monakhova MV, Milakina MA, Trikin RM, Oretskaya TS, Kubareva EA. Functional Specifics of the MutL Protein of the DNA Mismatch Repair System in Different Organisms. RUSSIAN JOURNAL OF BIOORGANIC CHEMISTRY 2020. [DOI: 10.1134/s1068162020060217] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/23/2022]
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30
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Li S, Shi B, Liu X, An HX. Acetylation and Deacetylation of DNA Repair Proteins in Cancers. Front Oncol 2020; 10:573502. [PMID: 33194676 PMCID: PMC7642810 DOI: 10.3389/fonc.2020.573502] [Citation(s) in RCA: 17] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2020] [Accepted: 09/17/2020] [Indexed: 12/12/2022] Open
Abstract
Hundreds of DNA repair proteins coordinate together to remove the diverse damages for ensuring the genomic integrity and stability. The repair system is an extensive network mainly encompassing cell cycle arrest, chromatin remodeling, various repair pathways, and new DNA fragment synthesis. Acetylation on DNA repair proteins is a dynamic epigenetic modification orchestrated by lysine acetyltransferases (HATs) and lysine deacetylases (HDACs), which dramatically affects the protein functions through multiple mechanisms, such as regulation of DNA binding ability, protein activity, post-translational modification (PTM) crosstalk, and protein–protein interaction. Accumulating evidence has indicated that the aberrant acetylation of DNA repair proteins contributes to the dysfunction of DNA repair ability, the pathogenesis and progress of cancer, as well as the chemosensitivity of cancer cells. In the present scenario, targeting epigenetic therapy is being considered as a promising method at par with the conventional cancer therapeutic strategies. This present article provides an overview of the recent progress in the functions and mechanisms of acetylation on DNA repair proteins involved in five major repair pathways, which warrants the possibility of regulating acetylation on repair proteins as a therapeutic target in cancers.
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Affiliation(s)
- Shiqin Li
- Department of Medical Oncology, Xiang'an Hospital of Xiamen University, Xiamen, China
| | - Bingbing Shi
- Department of Medical Oncology, Xiang'an Hospital of Xiamen University, Xiamen, China
| | - Xinli Liu
- Department of Medical Oncology, Xiang'an Hospital of Xiamen University, Xiamen, China
| | - Han-Xiang An
- Department of Medical Oncology, Xiang'an Hospital of Xiamen University, Xiamen, China
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31
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Young SJ, Sebald M, Shah Punatar R, Larin M, Masino L, Rodrigo-Brenni MC, Liang CC, West SC. MutSβ Stimulates Holliday Junction Resolution by the SMX Complex. Cell Rep 2020; 33:108289. [PMID: 33086055 DOI: 10.1016/j.celrep.2020.108289] [Citation(s) in RCA: 20] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/08/2020] [Revised: 09/02/2020] [Accepted: 09/29/2020] [Indexed: 12/20/2022] Open
Abstract
MutSα and MutSβ play important roles in DNA mismatch repair and are linked to inheritable cancers and degenerative disorders. Here, we show that MSH2 and MSH3, the two components of MutSβ, bind SLX4 protein, a scaffold for the assembly of the SLX1-SLX4-MUS81-EME1-XPF-ERCC1 (SMX) trinuclease complex. SMX promotes the resolution of Holliday junctions (HJs), which are intermediates in homologous recombinational repair. We find that MutSβ binds HJs and stimulates their resolution by SLX1-SLX4 or SMX in reactions dependent upon direct interactions between MutSβ and SLX4. In contrast, MutSα does not stimulate HJ resolution. MSH3-depleted cells exhibit reduced sister chromatid exchanges and elevated levels of homologous recombination ultrafine bridges (HR-UFBs) at mitosis, consistent with defects in the processing of recombination intermediates. These results demonstrate a role for MutSβ in addition to its established role in the pathogenic expansion of CAG/CTG trinucleotide repeats, which is causative of myotonic dystrophy and Huntington's disease.
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Affiliation(s)
- Sarah J Young
- The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK
| | - Marie Sebald
- The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK
| | | | - Meghan Larin
- The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK
| | - Laura Masino
- The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK
| | | | - Chih-Chao Liang
- The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK
| | - Stephen C West
- The Francis Crick Institute, 1 Midland Road, London NW1 1AT, UK.
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32
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Karakaidos P, Karagiannis D, Rampias T. Resolving DNA Damage: Epigenetic Regulation of DNA Repair. Molecules 2020; 25:molecules25112496. [PMID: 32471288 PMCID: PMC7321228 DOI: 10.3390/molecules25112496] [Citation(s) in RCA: 33] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/29/2020] [Revised: 05/22/2020] [Accepted: 05/25/2020] [Indexed: 12/18/2022] Open
Abstract
Epigenetic research has rapidly evolved into a dynamic field of genome biology. Chromatin regulation has been proved to be an essential aspect for all genomic processes, including DNA repair. Chromatin structure is modified by enzymes and factors that deposit, erase, and interact with epigenetic marks such as DNA and histone modifications, as well as by complexes that remodel nucleosomes. In this review we discuss recent advances on how the chromatin state is modulated during this multi-step process of damage recognition, signaling, and repair. Moreover, we examine how chromatin is regulated when different pathways of DNA repair are utilized. Furthermore, we review additional modes of regulation of DNA repair, such as through the role of global and localized chromatin states in maintaining expression of DNA repair genes, as well as through the activity of epigenetic enzymes on non-nucleosome substrates. Finally, we discuss current and future applications of the mechanistic interplays between chromatin regulation and DNA repair in the context cancer treatment.
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Affiliation(s)
| | - Dimitris Karagiannis
- Department of Genetics and Development, Columbia University Medical Center, New York, NY 10032, USA;
| | - Theodoros Rampias
- Biomedical Research Foundation of the Academy of Athens, 11527 Athens, Greece;
- Correspondence: ; Tel.: +30-210-659-7469
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33
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Tamura K, Kaneda M, Futagawa M, Takeshita M, Kim S, Nakama M, Kawashita N, Tatsumi-Miyajima J. Genetic and genomic basis of the mismatch repair system involved in Lynch syndrome. Int J Clin Oncol 2019; 24:999-1011. [PMID: 31273487 DOI: 10.1007/s10147-019-01494-y] [Citation(s) in RCA: 45] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/11/2019] [Accepted: 06/17/2019] [Indexed: 12/11/2022]
Abstract
Lynch syndrome is a cancer-predisposing syndrome inherited in an autosomal-dominant manner, wherein colon cancer and endometrial cancer develop frequently in the family, it results from a loss-of-function mutation in one of four different genes (MLH1, MSH2, MSH6, and PMS2) encoding mismatch repair proteins. Being located immediately upstream of the MSH2 gene, EPCAM abnormalities can affect MSH2 and cause Lynch syndrome. Mismatch repair proteins are involved in repairing of incorrect pairing (point mutations and deletion/insertion of simple repetitive sequences, so-called microsatellites) that can arise during DNA replication. MSH2 forms heterodimers with MSH6 or MSH3 (MutSα, MutSβ, respectively) and is involved in mismatch-pair recognition and initiation of repair. MLH1 forms a complex with PMS2, and functions as an endonuclease. If the mismatch repair system is thoroughly working, genome integrity is maintained completely. Lynch syndrome is a state of mismatch repair deficiency due to a monoallelic abnormality of any mismatch repair genes. The phenotype indicating the mismatch repair deficiency can be frequently shown as a microsatellite instability in tumors. Children with germline biallelic mismatch repair gene abnormalities were reported to develop conditions such as gastrointestinal polyposis, colorectal cancer, brain cancer, leukemia, etc., and so on, demonstrating the need to respond with new concepts in genetic counseling. In promoting cancer genome medicine in a new era, such as by utilizing immune checkpoints, it is important to understand the genetic and genomic molecular background, including the status of mismatch repair deficiency.
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Affiliation(s)
- Kazuo Tamura
- Division of Medical Genetics, Master of Science, Graduate School of Science and Engineering Research, Kindai University, Higashiosaka, Japan.
| | - Motohide Kaneda
- Division of Medical Genetics, Master of Science, Graduate School of Science and Engineering Research, Kindai University, Higashiosaka, Japan
| | - Mashu Futagawa
- Division of Medical Genetics, Master of Science, Graduate School of Science and Engineering Research, Kindai University, Higashiosaka, Japan
| | - Miho Takeshita
- Division of Medical Genetics, Master of Science, Graduate School of Science and Engineering Research, Kindai University, Higashiosaka, Japan
| | - Sanghyuk Kim
- Division of Medical Genetics, Master of Science, Graduate School of Science and Engineering Research, Kindai University, Higashiosaka, Japan
| | - Mina Nakama
- Division of Clinical Genetics, Gifu University Hospital, Gifu, Japan
| | - Norihito Kawashita
- Division of Medical Genetics, Master of Science, Graduate School of Science and Engineering Research, Kindai University, Higashiosaka, Japan
| | - Junko Tatsumi-Miyajima
- Division of Medical Genetics, Master of Science, Graduate School of Science and Engineering Research, Kindai University, Higashiosaka, Japan
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34
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Nalawansha DA, Zhang Y, Herath K, Pflum MKH. HDAC1 Substrate Profiling Using Proteomics-Based Substrate Trapping. ACS Chem Biol 2018; 13:3315-3324. [PMID: 30421914 PMCID: PMC6563814 DOI: 10.1021/acschembio.8b00737] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Histone deacetylase (HDAC) proteins are overexpressed in multiple diseases, including cancer, and have emerged as anticancer drug targets. HDAC proteins regulate cellular processes, such as the cell cycle, apoptosis, and cell proliferation, by deacetylating histone and non-histone substrates. Although a plethora of acetylated proteins have been identified using large-scale proteomic approaches, the HDAC proteins responsible for their dynamic deacetylation have been poorly studied. For example, few substrates of HDAC1 have been identified, which is mainly due to the scarcity of substrate identification tools. We recently developed a mutant trapping strategy to identify novel substrates of HDAC1. Herein, we introduce an improved version of the trapping method that uses mass spectrometry (MS)-based proteomics to identify multiple substrates simultaneously. Among the substrate hits, CDK1, AIFM1, MSH6, and RuvB-like 1 were identified as likely HDAC1 substrates. These newly discovered HDAC1 substrates are involved in various biological processes, suggesting novel functions of HDAC1 apart from epigenetics. Substrate trapping combined with MS-based proteomics provides an efficient approach to HDAC1 substrate identification and contributes to the full characterization of HDAC function in normal and disease states.
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Affiliation(s)
| | - Yuchen Zhang
- Department of Chemistry, Wayne State University, 5101 Cass Avenue, Detroit, MI 48202
| | - Kavinda Herath
- Department of Chemistry, Wayne State University, 5101 Cass Avenue, Detroit, MI 48202
| | - Mary Kay H. Pflum
- Department of Chemistry, Wayne State University, 5101 Cass Avenue, Detroit, MI 48202
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35
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Patil V, Mahalingam K. A four-protein expression prognostic signature predicts clinical outcome of lower-grade glioma. Gene 2018; 679:57-64. [DOI: 10.1016/j.gene.2018.08.001] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/21/2018] [Accepted: 08/01/2018] [Indexed: 01/07/2023]
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36
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Duan FX, Gu GL, Yang HR, Yu PF, Zhang Z. Must Peutz-Jeghers syndrome patients have the LKB1/STK11 gene mutation? A case report and review of the literature. World J Clin Cases 2018; 6:224-232. [PMID: 30148152 PMCID: PMC6107527 DOI: 10.12998/wjcc.v6.i8.224] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/21/2018] [Revised: 04/23/2018] [Accepted: 05/11/2018] [Indexed: 02/05/2023] Open
Abstract
Peutz-Jeghers syndrome (PJS) is an autosomal dominant inherited disease, which is characterized by mucocutaneous pigmentation and multiple gastrointestinal hamartoma polyps. The germline mutation of LKB1/STK11 gene on chromosome 19p13.3 is considered to be the hereditary cause of PJS. However, must a patient with PJS have the LKB1/STK11 gene mutation? We here report a case of a male patient who had typical manifestations of PJS and a definite family history, but did not have LKB1/STK11 gene mutation. By means of high-throughput sequencing technology, only mutations in APC gene (c.6662T > C: p.Met2221Thr) and MSH6 gene (c.3488A > T: p.Glu1163Val) were detected. The missense mutations in APC and MSH6 gene may lead to abnormalities in structure and function of their expression products, and may result in the occurrence of PJS. This study suggests that some other genetic disorders may cause PJS besides LKB1/STK11 gene mutation.
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Affiliation(s)
- Fu-Xiao Duan
- Department of General Surgery, Air Force General Hospital of Chinese PLA, Beijing 100142, China
| | - Guo-Li Gu
- Department of General Surgery, Air Force General Hospital of Chinese PLA, Beijing 100142, China
| | - Hai-Rui Yang
- Department of General Surgery, Air Force General Hospital of Chinese PLA, Beijing 100142, China
| | - Peng-Fei Yu
- Department of General Surgery, Air Force General Hospital of Chinese PLA, Beijing 100142, China
| | - Zhi Zhang
- Department of General Surgery, Air Force General Hospital of Chinese PLA, Beijing 100142, China
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Meier B, Volkova NV, Hong Y, Schofield P, Campbell PJ, Gerstung M, Gartner A. Mutational signatures of DNA mismatch repair deficiency in C. elegans and human cancers. Genome Res 2018; 28:666-675. [PMID: 29636374 PMCID: PMC5932607 DOI: 10.1101/gr.226845.117] [Citation(s) in RCA: 90] [Impact Index Per Article: 12.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/27/2017] [Accepted: 01/02/2018] [Indexed: 12/18/2022]
Abstract
Throughout their lifetime, cells are subject to extrinsic and intrinsic mutational processes leaving behind characteristic signatures in the genome. DNA mismatch repair (MMR) deficiency leads to hypermutation and is found in different cancer types. Although it is possible to associate mutational signatures extracted from human cancers with possible mutational processes, the exact causation is often unknown. Here, we use C. elegans genome sequencing of pms-2 and mlh-1 knockouts to reveal the mutational patterns linked to C. elegans MMR deficiency and their dependency on endogenous replication errors and errors caused by deletion of the polymerase ε subunit pole-4 Signature extraction from 215 human colorectal and 289 gastric adenocarcinomas revealed three MMR-associated signatures, one of which closely resembles the C. elegans MMR spectrum and strongly discriminates microsatellite stable and unstable tumors (AUC = 98%). A characteristic difference between human and C. elegans MMR deficiency is the lack of elevated levels of NCG > NTG mutations in C. elegans, likely caused by the absence of cytosine (CpG) methylation in worms. The other two human MMR signatures may reflect the interaction between MMR deficiency and other mutagenic processes, but their exact cause remains unknown. In summary, combining information from genetically defined models and cancer samples allows for better aligning mutational signatures to causal mutagenic processes.
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Affiliation(s)
- Bettina Meier
- Centre for Gene Regulation and Expression, University of Dundee, Dundee DD1 5EH, United Kingdom
| | - Nadezda V Volkova
- European Molecular Biology Laboratory, European Bioinformatics Institute (EMBL-EBI), Hinxton CB10 1SD, United Kingdom
| | - Ye Hong
- Centre for Gene Regulation and Expression, University of Dundee, Dundee DD1 5EH, United Kingdom
| | - Pieta Schofield
- Centre for Gene Regulation and Expression, University of Dundee, Dundee DD1 5EH, United Kingdom
- Division of Computational Biology, University of Dundee, Dundee DD1 5EH, United Kingdom
| | - Peter J Campbell
- Cancer Genome Project, Wellcome Trust Sanger Institute, Hinxton, Cambridge CB10 1SA, United Kingdom
- Department of Haematology, University of Cambridge, Cambridge CB2 0XY, United Kingdom
- Department of Haematology, Addenbrooke's Hospital, Cambridge CB2 0XY, United Kingdom
| | - Moritz Gerstung
- European Molecular Biology Laboratory, European Bioinformatics Institute (EMBL-EBI), Hinxton CB10 1SD, United Kingdom
| | - Anton Gartner
- Centre for Gene Regulation and Expression, University of Dundee, Dundee DD1 5EH, United Kingdom
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Cama A, Genuardi M, Guanti G, Radice P, Varesco L. Molecular Genetics of Hereditary Non-Polyposis Colorectal Cancer (HNPCC). TUMORI JOURNAL 2018; 82:122-35. [PMID: 8644374 DOI: 10.1177/030089169608200206] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/29/2022]
Abstract
The story of the molecular genetics of HNPCC is one of astonishingly rapid achievements. In just 16 months, from May 1993 to September 1994, four different genes, namely hMSH2, hMLH1, hPMS1 and hPMS2 have been identified and demonstrated to be associated with the disease. Their cloning was facilitated by the finding that tumor cells in HNPCC patients display a hypermutability of DNA short tandem repeats (microsatellite instability). In fact, HNPCC associated genes are the human counterparts of genetic elements known to control the fidelity of DNA replication in lower organisms. So far, more than 50 germline mutations of hMSH2 and hMLH1 genes have been reported in HNPCC kindreds. In addition, somatic mutations have been documented in hereditary as well as sporadic cancers. Unfortunately, the molecular diagnosis of HNPCC is hampered by the lack of mutational “hot spots” and of clearly defined genotype-phenotype correlations and different screening methods are to be employed for the analysis of affected and at-risk individuals.
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Affiliation(s)
- A Cama
- Cattedra di Patologia Generale, Università Gabriele D'Annuzio, Chieti,Italy
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Dahal BK, Kadyrova LY, Delfino KR, Rogozin IB, Gujar V, Lobachev KS, Kadyrov FA. Involvement of DNA mismatch repair in the maintenance of heterochromatic DNA stability in Saccharomyces cerevisiae. PLoS Genet 2017; 13:e1007074. [PMID: 29069084 PMCID: PMC5673234 DOI: 10.1371/journal.pgen.1007074] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/18/2017] [Revised: 11/06/2017] [Accepted: 10/15/2017] [Indexed: 11/30/2022] Open
Abstract
Heterochromatin contains a significant part of nuclear DNA. Little is known about the mechanisms that govern heterochromatic DNA stability. We show here that in the yeast Saccharomyces cerevisiae (i) DNA mismatch repair (MMR) is required for the maintenance of heterochromatic DNA stability, (ii) MutLα (Mlh1-Pms1 heterodimer), MutSα (Msh2-Msh6 heterodimer), MutSβ (Msh2-Msh3 heterodimer), and Exo1 are involved in MMR at heterochromatin, (iii) Exo1-independent MMR at heterochromatin frequently leads to the formation of Pol ζ-dependent mutations, (iv) MMR cooperates with the proofreading activity of Pol ε and the histone acetyltransferase Rtt109 in the maintenance of heterochromatic DNA stability, (v) repair of base-base mismatches at heterochromatin is less efficient than repair of base-base mismatches at euchromatin, and (vi) the efficiency of repair of 1-nt insertion/deletion loops at heterochromatin is similar to the efficiency of repair of 1-nt insertion/deletion loops at euchromatin. Eukaryotic mismatch repair is an important intracellular process that defends DNA against mutations. Inactivation of mismatch repair in human cells strongly increases the risk of cancer initiation and development. Although significant progress has been made in understanding mismatch repair at euchromatin, mismatch repair at heterochromatin is not well understood. Baker’s yeast is a key model organism to study mismatch repair. We determined that in baker’s yeast (1) mismatch repair protects heterochromatic DNA from mutations, (2) the MutLα, MutSα, MutSβ, and Exo1 proteins play important roles in mismatch repair at heterochromatin, (3) Exo1-independent mismatch repair at heterochromatin is an error-prone process; (4) mismatch repair cooperates with two other intracellular processes to protect the stability of heterochromatic DNA; and (5) the efficiency of repair of base-base mismatches at heterochromatin is lower than the efficiency of repair of base-base mismatches at euchromatin, but the efficiency of 1-nt insertion/deletion loop repair at heterochromatin is similar to the efficiency of 1-nt insertion/deletion loop repair at euchromatin.
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Affiliation(s)
- Basanta K. Dahal
- Department of Biochemistry and Molecular Biology, Southern Illinois University School of Medicine, Carbondale, IL, United States of America
| | - Lyudmila Y. Kadyrova
- Department of Biochemistry and Molecular Biology, Southern Illinois University School of Medicine, Carbondale, IL, United States of America
| | - Kristin R. Delfino
- Center for Clinical Research, Southern Illinois University School of Medicine, Springfield, IL, United States of America
| | - Igor B. Rogozin
- National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, MD, United States of America
| | - Vaibhavi Gujar
- Department of Biochemistry and Molecular Biology, Southern Illinois University School of Medicine, Carbondale, IL, United States of America
| | - Kirill S. Lobachev
- School of Biological Sciences and Institute for Bioengineering and Bioscience, Georgia Institute of Technology, Atlanta, GA, United States of America
| | - Farid A. Kadyrov
- Department of Biochemistry and Molecular Biology, Southern Illinois University School of Medicine, Carbondale, IL, United States of America
- * E-mail:
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The Role of Blm Helicase in Homologous Recombination, Gene Conversion Tract Length, and Recombination Between Diverged Sequences in Drosophilamelanogaster. Genetics 2017; 207:923-933. [PMID: 28912341 DOI: 10.1534/genetics.117.300285] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/15/2017] [Accepted: 09/10/2017] [Indexed: 11/18/2022] Open
Abstract
DNA double-strand breaks (DSBs) are a particularly deleterious class of DNA damage that threatens genome integrity. DSBs are repaired by three pathways: nonhomologous-end joining (NHEJ), homologous recombination (HR), and single-strand annealing (SSA). Drosophila melanogaster Blm (DmBlm) is the ortholog of Saccharomyces cerevisiae SGS1 and human BLM, and has been shown to suppress crossovers in mitotic cells and repair mitotic DNA gaps via HR. To further elucidate the role of DmBlm in repair of a simple DSB, and in particular recombination mechanisms, we utilized the Direct Repeat of white (DR-white) and Direct Repeat of whitewith mutations (DR-white.mu) repair assays in multiple mutant allele backgrounds. DmBlm null and helicase-dead mutants both demonstrated a decrease in repair by noncrossover HR, and a concurrent increase in non-HR events, possibly including SSA, crossovers, deletions, and NHEJ, although detectable processing of the ends was not significantly impacted. Interestingly, gene conversion tract lengths of HR repair events were substantially shorter in DmBlm null but not helicase-dead mutants, compared to heterozygote controls. Using DR-white.mu, we found that, in contrast to Sgs1, DmBlm is not required for suppression of recombination between diverged sequences. Taken together, our data suggest that DmBlm helicase function plays a role in HR, and the steps that contribute to determining gene conversion tract length are helicase-independent.
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41
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Liccardo R, De Rosa M, Rossi GB, Carlomagno N, Izzo P, Duraturo F. Incomplete Segregation of MSH6 Frameshift Variants with Phenotype of Lynch Syndrome. Int J Mol Sci 2017; 18:ijms18050999. [PMID: 28481244 PMCID: PMC5454912 DOI: 10.3390/ijms18050999] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/03/2017] [Revised: 04/21/2017] [Accepted: 05/02/2017] [Indexed: 12/23/2022] Open
Abstract
Lynch syndrome (LS), the most frequent form of hereditary colorectal cancer, involves mutations in mismatch repair genes. The aim of this study was to identify mutations in MSH6 from 97 subjects negative for mutations in MLH1 and MSH2. By direct sequencing, we identified 27 MSH6 variants, of which, nine were novel. To verify the pathogenicity of these novel variants, we performed in silico and segregation analyses. Three novel variants were predicted by in silico analysis as damaging mutations and segregated with the disease phenotype; while a novel frameshift deletion variant that was predicted to yield a premature stop codon did not segregate with the LS phenotype in three of four cases in the family. Interestingly, another frame-shift variant identified in this study, already described in the literature, also did not segregate with the LS phenotype in one of two affected subjects in the family. In all affected subjects of both families, no mutation was detected in other MMR genes. Therefore, it is expected that within these families, other genetic factors contribute to the disease either alone or in combination with MSH6 variants. We conclude that caution should be exercised in counseling for MSH6-associated LS family members.
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Affiliation(s)
- Raffaella Liccardo
- Department of Molecular Medicine and Medical Biotechnology, Federico II University Medical School, 80131 Naples, Italy.
| | - Marina De Rosa
- Department of Molecular Medicine and Medical Biotechnology, Federico II University Medical School, 80131 Naples, Italy.
| | - Giovanni Battista Rossi
- Endoscopy Unit, Fondazione Pascale National Institute for Study and Care of Tumors, 80131 Naples, Italy.
| | - Nicola Carlomagno
- General Surgery Unit-Advanced Biomedical Science Department, Federico II University Medical School, 80131 Naples, Italy.
| | - Paola Izzo
- Department of Molecular Medicine and Medical Biotechnology, Federico II University Medical School, 80131 Naples, Italy.
- CEINGE-Biotecnologie Avanzate, 80145 Naples, Italy.
| | - Francesca Duraturo
- Department of Molecular Medicine and Medical Biotechnology, Federico II University Medical School, 80131 Naples, Italy.
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42
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Kadyrova LY, Dahal BK, Kadyrov FA. The Major Replicative Histone Chaperone CAF-1 Suppresses the Activity of the DNA Mismatch Repair System in the Cytotoxic Response to a DNA-methylating Agent. J Biol Chem 2016; 291:27298-27312. [PMID: 27872185 DOI: 10.1074/jbc.m116.760561] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/26/2016] [Revised: 11/15/2016] [Indexed: 11/06/2022] Open
Abstract
The DNA mismatch repair (MMR) system corrects DNA mismatches in the genome. It is also required for the cytotoxic response of O6-methylguanine-DNA methyltransferase (MGMT)-deficient mammalian cells and yeast mgt1Δ rad52Δ cells to treatment with Sn1-type methylating agents, which produce cytotoxic O6-methylguanine (O6-mG) DNA lesions. Specifically, an activity of the MMR system causes degradation of irreparable O6-mG-T mispair-containing DNA, triggering cell death; this process forms the basis of treatments of MGMT-deficient cancers with Sn1-type methylating drugs. Recent research supports the view that degradation of irreparable O6-mG-T mispair-containing DNA by the MMR system and CAF-1-dependent packaging of the newly replicated DNA into nucleosomes are two concomitant processes that interact with each other. Here, we studied whether CAF-1 modulates the activity of the MMR system in the cytotoxic response to Sn1-type methylating agents. We found that CAF-1 suppresses the activity of the MMR system in the cytotoxic response of yeast mgt1Δ rad52Δ cells to the prototypic Sn1-type methylating agent N-methyl-N'-nitro-N-nitrosoguanidine. We also report evidence that in human MGMT-deficient cell-free extracts, CAF-1-dependent packaging of irreparable O6-mG-T mispair-containing DNA into nucleosomes suppresses its degradation by the MMR system. Taken together, these findings suggest that CAF-1-dependent incorporation of irreparable O6-mG-T mispair-containing DNA into nucleosomes suppresses its degradation by the MMR system, thereby defending the cell against killing by the Sn1-type methylating agent.
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Affiliation(s)
- Lyudmila Y Kadyrova
- From the Department of Biochemistry and Molecular Biology, Southern Illinois University, School of Medicine, Carbondale, Illinois 62901
| | - Basanta K Dahal
- From the Department of Biochemistry and Molecular Biology, Southern Illinois University, School of Medicine, Carbondale, Illinois 62901
| | - Farid A Kadyrov
- From the Department of Biochemistry and Molecular Biology, Southern Illinois University, School of Medicine, Carbondale, Illinois 62901
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43
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Affiliation(s)
- Guo-Min Li
- Department of Biochemistry and Molecular Biology, Norris Comprehensive Cancer Center, University of Southern California Keck School of Medicine, University of Southern California, Los Angeles, CA 90033.
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44
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Lorge E, Moore MM, Clements J, O'Donovan M, Fellows MD, Honma M, Kohara A, Galloway S, Armstrong MJ, Thybaud V, Gollapudi B, Aardema MJ, Tanir JY. Standardized cell sources and recommendations for good cell culture practices in genotoxicity testing. MUTATION RESEARCH-GENETIC TOXICOLOGY AND ENVIRONMENTAL MUTAGENESIS 2016; 809:1-15. [PMID: 27692294 DOI: 10.1016/j.mrgentox.2016.08.001] [Citation(s) in RCA: 43] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/29/2016] [Revised: 07/29/2016] [Accepted: 08/02/2016] [Indexed: 11/16/2022]
Abstract
Good cell culture practice and characterization of the cell lines used are of critical importance in in vitro genotoxicity testing. The objective of this initiative was to make continuously available stocks of the characterized isolates of the most frequently used mammalian cell lines in genotoxicity testing anywhere in the world ('IVGT' cell lines). This project was organized under the auspices of the International Life Sciences Institute (ILSI) Health and Environmental Sciences Institute (HESI) Project Committee on the Relevance and Follow-up of Positive Results in In Vitro Genetic Toxicity (IVGT) Testing. First, cell isolates were identified that are as close as possible to the isolate described in the initial publications reporting their use in genotoxicity testing. The depositors of these cell lines managed their characterization and their expansion for preparing continuously available stocks of these cells that are stored at the European Collection of Cell Cultures (ECACC, UK) and the Japanese Collection of Research Bioresources (JCRB, Japan). This publication describes how the four 'IVGT' cell lines, i.e. L5178Y TK+/- 3.7.2C, TK6, CHO-WBL and CHL/IU, were prepared for deposit at the ECACC and JCRB cell banks. Recommendations for handling these cell lines and monitoring their characteristics are also described. The growth characteristics of these cell lines (growth rates and cell cycles), their identity (karyotypes and genetic status) and ranges of background frequencies of select endpoints are also reported to help in the routine practice of genotoxicity testing using these cell lines.
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Affiliation(s)
- E Lorge
- Servier Group, 45520, Gidy, France
| | - M M Moore
- Ramboll Environ, Little Rock, AR, 72201, USA
| | - J Clements
- Covance Laboratories Ltd, Harrogate, HG3 1PY, UK
| | - M O'Donovan
- O'Donovan GT Consulting Ltd., Epperstone, Nottingham, NG14 6AG, UK
| | - M D Fellows
- AstraZeneca, Drug Safety and Metabolism, Cambridge, CB4 0WG, UK
| | - M Honma
- National Institute of Health Sciences, Tokyo, Japan
| | - A Kohara
- JCRB Cell Bank, National Institutes of Biomedical Innovation, Health and Nutrition, Osaka, Japan
| | - S Galloway
- Merck Research Laboratories, W 45-316, West Point, PA 19486, USA
| | - M J Armstrong
- Merck Research Laboratories, W 45-316, West Point, PA 19486, USA
| | - V Thybaud
- Sanofi, 94400, Vitry sur Seine, France
| | - B Gollapudi
- Exponent, Inc., 1910 St. Andrews St., Midland, MI 48640, USA
| | - M J Aardema
- Marilyn Aardema Consulting LLC, Fairfield, OH 45014, USA
| | - J Y Tanir
- ILSI Health and Environmental Sciences Institute, Washington, DC 20005, USA.
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45
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Modrich P. Mechanismen der Fehlpaarungsreparatur in E. coliund im Menschen (Nobel-Aufsatz). Angew Chem Int Ed Engl 2016. [DOI: 10.1002/ange.201601412] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/10/2022]
Affiliation(s)
- Paul Modrich
- Howard Hughes Medical Institute and Department of Biochemistry; Duke University, Medical Center; Durham NC 27710 USA
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46
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Modrich P. Mechanisms in E. coli and Human Mismatch Repair (Nobel Lecture). Angew Chem Int Ed Engl 2016; 55:8490-501. [PMID: 27198632 PMCID: PMC5193110 DOI: 10.1002/anie.201601412] [Citation(s) in RCA: 71] [Impact Index Per Article: 7.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/09/2016] [Indexed: 11/11/2022]
Abstract
DNA molecules are not completely stable, they are subject to chemical or photochemical damage and errors that occur during DNA replication resulting in mismatched base pairs. Through mechanistic studies Paul Modrich showed how replication errors are corrected by strand-directed mismatch repair in Escherichia coli and human cells.
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Affiliation(s)
- Paul Modrich
- Howard Hughes Medical Institute and Department of Biochemistry, Duke University, Medical Center, Durham, NC, 27710, USA.
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47
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Guo J, Gu L, Leffak M, Li GM. MutSβ promotes trinucleotide repeat expansion by recruiting DNA polymerase β to nascent (CAG)n or (CTG)n hairpins for error-prone DNA synthesis. Cell Res 2016; 26:775-86. [PMID: 27255792 PMCID: PMC5129881 DOI: 10.1038/cr.2016.66] [Citation(s) in RCA: 40] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/26/2016] [Revised: 05/15/2016] [Accepted: 05/17/2016] [Indexed: 12/12/2022] Open
Abstract
Expansion of (CAG)•(CTG) repeats causes a number of familial neurodegenerative disorders. Although the underlying mechanism remains largely unknown, components involved in DNA mismatch repair, particularly mismatch recognition protein MutSβ (a MSH2-MSH3 heterodimer), are implicated in (CAG)•(CTG) repeat expansion. In addition to recognizing small insertion-deletion loop-outs, MutSβ also specifically binds DNA hairpin imperfect heteroduplexes formed within (CAG)n•(CTG)n sequences. However, whether or not and how MutSβ binding triggers expansion of (CAG)•(CTG) repeats remain unknown. We show here that purified recombinant MutSβ physically interacts with DNA polymerase β (Polβ) and stimulates Polβ-catalyzed (CAG)n or (CTG)n hairpin retention. Consistent with these in vitro observations, MutSβ and Polβ interact with each other in vivo, and colocalize at (CAG)•(CTG) repeats during DNA replication. Our data support a model for error-prone processing of (CAG)n or (CTG)n hairpins by MutSβ and Polβ during DNA replication and/or repair: MutSβ recognizes (CAG)n or (CTG)n hairpins formed in the nascent DNA strand, and recruits Polβ to the complex, which then utilizes the hairpin as a primer for extension, leading to (CAG)•(CTG) repeat expansion. This study provides a novel mechanism for trinucleotide repeat expansion in both dividing and non-dividing cells.
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Affiliation(s)
- Jinzhen Guo
- Department of Basic Medical Sciences, Tsinghua University School of Medicine, Beijing 100084, China.,Department of Biochemistry and Molecular Biology, Norris Comprehensive Cancer Center, University of Southern California Keck School of Medicine, 1450 Biggy Street, Los Angeles, CA 90033, USA
| | - Liya Gu
- Department of Biochemistry and Molecular Biology, Norris Comprehensive Cancer Center, University of Southern California Keck School of Medicine, 1450 Biggy Street, Los Angeles, CA 90033, USA
| | - Michael Leffak
- Department of Biochemistry and Molecular Biology, Boonshoft School of Medicine, Wright State University, Dayton, OH 45435, USA
| | - Guo-Min Li
- Department of Basic Medical Sciences, Tsinghua University School of Medicine, Beijing 100084, China.,Department of Biochemistry and Molecular Biology, Norris Comprehensive Cancer Center, University of Southern California Keck School of Medicine, 1450 Biggy Street, Los Angeles, CA 90033, USA
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48
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Negureanu L, Salsbury FR. Insights into protein - DNA interactions, stability and allosteric communications: a computational study of mutSα-DNA recognition complexes. J Biomol Struct Dyn 2016; 29:757-76. [PMID: 22208277 DOI: 10.1080/07391102.2012.10507412] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
DNA mismatch repair proteins (MMR) maintain genetic stability by recognizing and repairing mismatched bases and insertion/deletion loops mistakenly incorporated during DNA replication, and initiate cellular response to certain types of DNA damage. Loss of MMR in mammalian cells has been linked to resistance to certain DNA damaging chemotherapeutic agents, as well as to increase risk of cancer. Mismatch repair pathway is considered to involve the concerted action of at least 20 proteins. The most abundant MMR mismatch-binding factor in eukaryotes, MutSα, recognizes and initiates the repair of base-base mismatches and small insertion/deletion. We performed molecular dynamics simulations on mismatched and damaged MutSα-DNA complexes. A comprehensive DNA binding site analysis of relevant conformations shows that MutSα proteins recognize the mismatched and platinum cross-linked DNA substrates in significantly different modes. Distinctive conformational changes associated with MutSα binding to mismatched and damaged DNA have been identified and they provide insight into the involvement of MMR proteins in DNA-repair and DNA-damage pathways. Stability and allosteric interactions at the heterodimer interface associated with the mismatch and damage recognition step allow for prediction of key residues in MMR cancer-causing mutations. A rigorous hydrogen bonding analysis for ADP molecules at the ATPase binding sites is also presented. Due to extended number of known MMR cancer causing mutations among the residues proved to make specific contacts with ADP molecules, recommendations for further studies on similar mutagenic effects were made.
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49
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Zhang XM, Gavande N, Parajuli P, Bepler G. Implications of the USP10-HDAC6 axis in lung cancer - A path to precision medicine. JOURNAL OF CANCER BIOLOGY 2016; 2:10.46439/cancerbiology.2.015. [PMID: 34746935 PMCID: PMC8570638 DOI: 10.46439/cancerbiology.2.015] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 11/04/2022]
Abstract
Lung cancer is the leading cause of cancer death among both men and women in the United States. Because lung cancer is genetically heterogeneous, tailored therapy alone or in combination with chemotherapy would increase patient overall survival as compared with the one-size-fits-all chemotherapy. TP53-mutant lung cancer accounts for more than half of all lung cancer cases and is oftentimes more aggressive and resistant to chemotherapy. Directly targeting mutant p53 has not yet been successful, so identification of novel therapy targets and biomarkers in the TP53-mutant lung cancer is urgently needed to increase the overall survival in this subgroup. Deubiquitinating enzymes (DUBs) regulate a vast majority of proteins (DUBs' substrates) via removal of ubiquitin moieties or ubiquitin chains from these proteins, thereby altering the stability and/or functions of these substrates. In this review, we will focus on a DUB, referred to as ubiquitin-specific peptidase 10 (USP10) whose substrates include both oncogenic proteins and tumor suppressors. Therefore, targeting USP10 in cancer is highly context-dependent. Here, we will discuss USP10's functions in cancer by examining its various known substrates. In particular, we will elaborate our recent findings in the oncogenic role of USP10 in the TP53-mutant subgroup of lung cancer, focusing on USP10's function in the DNA damage response (DDR) via histone deacetylase 6 (HDAC6). Overall, these findings support the notion that targeting USP10 in the TP53-mutant subgroup of NSCLC would sensitize patients to cisplatin-based chemotherapy. Generating potent and specific clinically relevant USP10 inhibitors would benefit the TP53-mutant subgroup of NSCLC patients.
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Affiliation(s)
- Xiaohong Mary Zhang
- Department of Oncology, Wayne State University School of Medicine, Karmanos Cancer Institute, 4100 John R. Street, Detroit, Michigan, 48201, USA
| | - Navnath Gavande
- Department Pharmaceutical Sciences, Eugene Applebaum College of Pharmacy and Health Sciences, Wayne State University, 259 Mack Avenue, Detroit, Michigan, 48201, USA
| | - Prahlad Parajuli
- Department of Oncology, Wayne State University School of Medicine, Karmanos Cancer Institute, 4100 John R. Street, Detroit, Michigan, 48201, USA
| | - Gerold Bepler
- Department of Oncology, Wayne State University School of Medicine, Karmanos Cancer Institute, 4100 John R. Street, Detroit, Michigan, 48201, USA
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Zhang M, Hu C, Tong D, Xiang S, Williams K, Bai W, Li GM, Bepler G, Zhang X. Ubiquitin-specific Peptidase 10 (USP10) Deubiquitinates and Stabilizes MutS Homolog 2 (MSH2) to Regulate Cellular Sensitivity to DNA Damage. J Biol Chem 2016; 291:10783-91. [PMID: 26975374 DOI: 10.1074/jbc.m115.700047] [Citation(s) in RCA: 32] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/22/2015] [Indexed: 11/06/2022] Open
Abstract
MSH2 is a key DNA mismatch repair protein, which plays an important role in genomic stability. In addition to its DNA repair function, MSH2 serves as a sensor for DNA base analogs-provoked DNA replication errors and binds to various DNA damage-induced adducts to trigger cell cycle arrest or apoptosis. Loss or depletion of MSH2 from cells renders resistance to certain DNA-damaging agents. Therefore, the level of MSH2 determines DNA damage response. Previous studies showed that the level of MSH2 protein is modulated by the ubiquitin-proteasome pathway, and histone deacetylase 6 (HDAC6) serves as an ubiquitin E3 ligase. However, the deubiquitinating enzymes, which regulate MSH2 remain unknown. Here we report that ubiquitin-specific peptidase 10 (USP10) interacts with and stabilizes MSH2. USP10 deubiquitinates MSH2 in vitro and in vivo Moreover, the protein level of MSH2 is positively correlated with the USP10 protein level in a panel of lung cancer cell lines. Knockdown of USP10 in lung cancer cells exhibits increased cell survival and decreased apoptosis upon the treatment of DNA-methylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) and antimetabolite 6-thioguanine (6-TG). The above phenotypes can be rescued by ectopic expression of MSH2. In addition, knockdown of MSH2 decreases the cellular mismatch repair activity. Overall, our results suggest a novel USP10-MSH2 pathway regulating DNA damage response and DNA mismatch repair.
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Affiliation(s)
- Mu Zhang
- From the Department of Pathology and Cell Biology, Morsani College of Medicine, Tampa, Florida 33612
| | - Chen Hu
- From the Department of Pathology and Cell Biology, Morsani College of Medicine, Tampa, Florida 33612
| | - Dan Tong
- Department of Toxicology and Cancer Biology, University of Kentucky College of Medicine, Lexington, Kentucky 40536
| | - Shengyan Xiang
- From the Department of Pathology and Cell Biology, Morsani College of Medicine, Tampa, Florida 33612
| | - Kendra Williams
- From the Department of Pathology and Cell Biology, Morsani College of Medicine, Tampa, Florida 33612
| | - Wenlong Bai
- From the Department of Pathology and Cell Biology, Morsani College of Medicine, Tampa, Florida 33612, Cancer Biology and Evolution Program, H. Lee Moffitt Cancer Center and Research Institute, Tampa, Florida 33612, and
| | - Guo-Min Li
- Department of Toxicology and Cancer Biology, University of Kentucky College of Medicine, Lexington, Kentucky 40536
| | - Gerold Bepler
- Molecular Therapeutics Program, Karmanos Cancer Institute, Detroit, Michigan 48201
| | - Xiaohong Zhang
- From the Department of Pathology and Cell Biology, Morsani College of Medicine, Tampa, Florida 33612, Cancer Biology and Evolution Program, H. Lee Moffitt Cancer Center and Research Institute, Tampa, Florida 33612, and
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