|
1
|
Lorca V, Rueda D, Martín-Morales L, Fernández-Aceñero MJ, Grolleman J, Poves C, Llovet P, Tapial S, García-Barberán V, Sanz J, Pérez-Segura P, de Voer RM, Díaz-Rubio E, de la Hoya M, Caldés T, Garre P. Contribution of New Adenomatous Polyposis Predisposition Genes in an Unexplained Attenuated Spanish Cohort by Multigene Panel Testing. Sci Rep 2019; 9:9814. [PMID: 31285513 PMCID: PMC6614360 DOI: 10.1038/s41598-019-46403-5] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/15/2018] [Accepted: 06/28/2019] [Indexed: 12/25/2022] Open
Abstract
Attenuated adenomatous polyposis (AAP) is a heterogeneous syndrome in terms of clinical manifestations, heritability and etiology of the disease. Genetic heterogeneity and low penetrance alleles are probably the best explanation for this variability. Certainly, it is known that APC and MUTYH are high penetrance predisposition genes for adenomatous polyposis, but they only account for 5-10% of AAP. Other new predisposition genes, such as POLE, POLD1, NTHL1, AXIN2 or MSH3, have been recently described and have been associated with AAP, but their relative contribution is still not well defined. In order to evaluate the genetic predisposition to AAP in a hospital based population, germline DNAs from 158 AAP subjects were screened for genetic variants in the coding regions and intron-exon boundaries of seven associated genes through a next-generation sequencing (NGS) custom gene panel. Splicing, segregation studies, somatic mutational screening and RNA quantitative expression assays were conducted for selected variants. In four of the probands the adenoma susceptibility could be explained by actionable mutations in APC or MUTYH, and one other patient was a double carrier of two truncating variants in both POLE and NTHL1. Furthermore, 16 additional patients harbored uncertain significance variants in the remaining tested genes. This report gives information about the contribution of the newly described adenomatous polyposis predisposition genes in a Spanish attenuated polyposis cohort. Our results highly support the convenience of NGS multigene panels for attenuated polyposis genetic screening and reveals POLE frameshift variants as a plausible susceptibility mechanism for AAP.
Collapse
Affiliation(s)
- Víctor Lorca
- Laboratorio de Oncología Molecular, Hospital Clínico San Carlos, IdISSC, CIBERONC, Madrid, Spain
| | - Daniel Rueda
- Laboratorio de Cáncer Hereditario, Servicio de Bioquímica, i + 12, Hospital 12 de Octubre, Madrid, Spain
| | - Lorena Martín-Morales
- Laboratorio de Oncología Molecular, Hospital Clínico San Carlos, IdISSC, CIBERONC, Madrid, Spain
| | | | - Judith Grolleman
- Department of Human Genetics, Radboud University Medical Center, Nijmegen, The Netherlands
| | - Carmen Poves
- Servicio de Aparato Digestivo, Hospital Clínico San Carlos, Madrid, Spain
| | - Patricia Llovet
- Laboratorio de Oncología Molecular, Hospital Clínico San Carlos, IdISSC, CIBERONC, Madrid, Spain
| | - Sandra Tapial
- Laboratorio de Cáncer Hereditario, Servicio de Bioquímica, i + 12, Hospital 12 de Octubre, Madrid, Spain
| | - Vanesa García-Barberán
- Laboratorio de Oncología Molecular, Hospital Clínico San Carlos, IdISSC, CIBERONC, Madrid, Spain
| | - Julián Sanz
- Servicio de Anatomía Patológica, Hospital Clínico San Carlos, Madrid, Spain
| | - Pedro Pérez-Segura
- Servicio de Oncología Médica, Hospital Clínico San Carlos, CIBERONC, Madrid, Spain
| | - Richarda M de Voer
- Department of Human Genetics, Radboud University Medical Center, Nijmegen, The Netherlands
| | - Eduardo Díaz-Rubio
- Servicio de Oncología Médica, Hospital Clínico San Carlos, CIBERONC, Madrid, Spain
| | - Miguel de la Hoya
- Laboratorio de Oncología Molecular, Hospital Clínico San Carlos, IdISSC, CIBERONC, Madrid, Spain
| | - Trinidad Caldés
- Laboratorio de Oncología Molecular, Hospital Clínico San Carlos, IdISSC, CIBERONC, Madrid, Spain
| | - Pilar Garre
- Laboratorio de Oncología Molecular, Hospital Clínico San Carlos, IdISSC, CIBERONC, Madrid, Spain.
| |
Collapse
|
|
2
|
Raetz AG, David SS. When you're strange: Unusual features of the MUTYH glycosylase and implications in cancer. DNA Repair (Amst) 2019; 80:16-25. [PMID: 31203172 DOI: 10.1016/j.dnarep.2019.05.005] [Citation(s) in RCA: 27] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/23/2019] [Revised: 05/23/2019] [Accepted: 05/29/2019] [Indexed: 02/06/2023]
Abstract
MUTYH is a base-excision repair glycosylase that removes adenine opposite 8-oxoguanine (OG). Variants of MUTYH defective in functional activity lead to MUTYH-associated polyposis (MAP), which progresses to cancer with very high penetrance. Whole genome and whole exome sequencing studies have found MUTYH deficiencies in an increasing number of cancer types. While the canonical OG:A repair activity of MUTYH is well characterized and similar to bacterial MutY, here we review more recent evidence that MUTYH has activities independent of OG:A repair and appear centered on the interdomain connector (IDC) region of MUTYH. We summarize evidence that MUTYH is involved in rapid DNA damage response (DDR) signaling, including PARP activation, 9-1-1 and ATR signaling, and SIRT6 activity. MUTYH alters survival and DDR to a wide variety of DNA damaging agents in a time course that is not consistent with the formation of OG:A mispairs. Studies that suggest MUTYH inhibits the repair of alkyl-DNA damage and cyclopyrimidine dimers (CPDs) is reviewed, and evidence of a synthetic lethal interaction with mismatch repair (MMR) is summarized. Based on these studies we suggest that MUTYH has evolved from an OG:A mispair glycosylase to a multifunctional scaffold for DNA damage response signaling.
Collapse
Affiliation(s)
- Alan G Raetz
- Department of Chemistry, University of California, Davis, Davis, CA, USA.
| | - Sheila S David
- Department of Chemistry, University of California, Davis, Davis, CA, USA.
| |
Collapse
|
|
3
|
McDonnell KJ, Chemler JA, Bartels PL, O'Brien E, Marvin ML, Ortega J, Stern RH, Raskin L, Li GM, Sherman DH, Barton JK, Gruber SB. A human MUTYH variant linking colonic polyposis to redox degradation of the [4Fe4S] 2+ cluster. Nat Chem 2018; 10:873-880. [PMID: 29915346 PMCID: PMC6060025 DOI: 10.1038/s41557-018-0068-x] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2017] [Accepted: 04/20/2018] [Indexed: 12/26/2022]
Abstract
The human DNA repair enzyme MUTYH excises mispaired adenine residues in oxidized DNA. Homozygous MUTYH mutations underlie the autosomal, recessive cancer syndrome MUTYH-associated polyposis. We report a MUTYH variant, p.C306W (c.918C>G), with a tryptophan residue in place of native cysteine, that ligates the [4Fe4S] cluster in a patient with colonic polyposis and family history of early age colon cancer. In bacterial MutY, the [4Fe4S] cluster is redox active, allowing rapid localization to target lesions by long-range, DNA-mediated signalling. In the current study, using DNA electrochemistry, we determine that wild-type MUTYH is similarly redox-active, but MUTYH C306W undergoes rapid oxidative degradation of its cluster to [3Fe4S]+, with loss of redox signalling. In MUTYH C306W, oxidative cluster degradation leads to decreased DNA binding and enzyme function. This study confirms redox activity in eukaryotic DNA repair proteins and establishes MUTYH C306W as a pathogenic variant, highlighting the essential role of redox signalling by the [4Fe4S] cluster.
Collapse
Affiliation(s)
- Kevin J McDonnell
- University of Southern California Norris Comprehensive Cancer Center, Los Angeles, CA, USA
| | - Joseph A Chemler
- Life Sciences Institute, University of Michigan, Ann Arbor, MI, USA
| | - Phillip L Bartels
- Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA, USA
| | - Elizabeth O'Brien
- Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA, USA
| | - Monica L Marvin
- Department of Human Genetics, University of Michigan, Ann Arbor, MI, USA
| | - Janice Ortega
- Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, TX, USA
| | - Ralph H Stern
- Division of Molecular Medicine and Genetics, Department of Internal Medicine, University of Michigan, Ann Arbor, MI, USA
| | | | - Guo-Min Li
- Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, TX, USA
| | - David H Sherman
- Life Sciences Institute, University of Michigan, Ann Arbor, MI, USA.
- Departments of Medicinal Chemistry, Chemistry and Microbiology & Immunology, University of Michigan, Ann Arbor, MI, USA.
| | - Jacqueline K Barton
- Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA, USA.
| | - Stephen B Gruber
- University of Southern California Norris Comprehensive Cancer Center, Los Angeles, CA, USA.
| |
Collapse
|
|
4
|
Nuñez NN, Majumdar C, Lay KT, David SS. Fe-S Clusters and MutY Base Excision Repair Glycosylases: Purification, Kinetics, and DNA Affinity Measurements. Methods Enzymol 2018; 599:21-68. [PMID: 29746241 DOI: 10.1016/bs.mie.2017.11.035] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/09/2023]
Abstract
A growing number of iron-sulfur (Fe-S) cluster cofactors have been identified in DNA repair proteins. MutY and its homologs are base excision repair (BER) glycosylases that prevent mutations associated with the common oxidation product of guanine (G), 8-oxo-7,8-dihydroguanine (OG) by catalyzing adenine (A) base excision from inappropriately formed OG:A mispairs. The finding of an [4Fe-4S]2+ cluster cofactor in MutY, Endonuclease III, and structurally similar BER enzymes was surprising and initially thought to represent an example of a purely structural role for the cofactor. However, in the two decades subsequent to the initial discovery, purification and in vitro analysis of bacterial MutYs and mammalian homologs, such as human MUTYH and mouse Mutyh, have demonstrated that proper Fe-S cluster coordination is required for OG:A substrate recognition and adenine excision. In addition, the Fe-S cluster in MutY has been shown to be capable of redox chemistry in the presence of DNA. The work in our laboratory aimed at addressing the importance of the MutY Fe-S cluster has involved a battery of approaches, with the overarching hypothesis that understanding the role(s) of the Fe-S cluster is intimately associated with understanding the biological and chemical properties of MutY and its unique damaged DNA substrate as a whole. In this chapter, we focus on methods of enzyme expression and purification, detailed enzyme kinetics, and DNA affinity assays. The methods described herein have not only been leveraged to provide insight into the roles of the MutY Fe-S cluster but have also been provided crucial information needed to delineate the impact of inherited variants of the human homolog MUTYH associated with a colorectal cancer syndrome known as MUTYH-associated polyposis or MAP. Notably, many MAP-associated variants have been found adjacent to the Fe-S cluster further underscoring the intimate relationship between the cofactor, MUTYH-mediated DNA repair, and disease.
Collapse
Affiliation(s)
| | | | - Kori T Lay
- University of California, Davis, CA, United States
| | | |
Collapse
|
|
5
|
Kunrath-Lima M, Repolês BM, Alves CL, Furtado C, Rajão MA, Macedo AM, Franco GR, Pena SDJ, Valenzuela L, Wisnovsky S, Kelley SO, Galanti N, Cabrera G, Machado CR. Characterization of Trypanosoma cruzi MutY DNA glycosylase ortholog and its role in oxidative stress response. INFECTION GENETICS AND EVOLUTION 2017; 55:332-342. [PMID: 28970112 DOI: 10.1016/j.meegid.2017.09.030] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/11/2017] [Revised: 09/25/2017] [Accepted: 09/26/2017] [Indexed: 12/11/2022]
Abstract
Trypanosoma cruzi is a protozoan parasite and the causative agent of Chagas disease. Like most living organisms, it is susceptible to oxidative stress, and must adapt to distinct environments. Hence, DNA repair is essential for its survival and the persistence of infection. Therefore, we studied whether T. cruzi has a homolog counterpart of the MutY enzyme (TcMYH), important in the DNA Base Excision Repair (BER) mechanism. Analysis of T. cruzi genome database showed that this parasite has a putative MutY DNA glycosylase sequence. We performed heterologous complementation assays using this genomic sequence. TcMYH complemented the Escherichia coli MutY- strain, reducing the mutation rate to a level similar to wild type. In in vitro assays, TcMYH was able to remove an adenine that was opposite to 8-oxoguanine. We have also constructed a T. cruzi lineage that overexpresses MYH. Although in standard conditions this lineage has similar growth to control cells, the overexpressor is more sensitive to hydrogen peroxide and glucose oxidase than the control, probably due to accumulation of AP sites in its DNA. Localization experiments with GFP-fused TcMYH showed this enzyme is present in both nucleus and mitochondrion. QPCR and MtOX results reinforce the presence and function of TcMYH in these two organelles. Our data suggest T. cruzi has a functional MYH DNA glycosylase, which participates in nuclear and mitochondrial DNA Base Excision Repair.
Collapse
Affiliation(s)
- Marianna Kunrath-Lima
- Departamento de Bioquímica e Imunologia, ICB, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627, Caixa Postal 486, Belo Horizonte 30161-970, MG, Brazil
| | - Bruno Marçal Repolês
- Departamento de Bioquímica e Imunologia, ICB, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627, Caixa Postal 486, Belo Horizonte 30161-970, MG, Brazil
| | - Ceres Luciana Alves
- Departamento de Bioquímica e Imunologia, ICB, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627, Caixa Postal 486, Belo Horizonte 30161-970, MG, Brazil
| | - Carolina Furtado
- Departamento de Bioquímica e Imunologia, ICB, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627, Caixa Postal 486, Belo Horizonte 30161-970, MG, Brazil
| | - Matheus Andrade Rajão
- Departamento de Bioquímica e Imunologia, ICB, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627, Caixa Postal 486, Belo Horizonte 30161-970, MG, Brazil
| | - Andrea Mara Macedo
- Departamento de Bioquímica e Imunologia, ICB, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627, Caixa Postal 486, Belo Horizonte 30161-970, MG, Brazil
| | - Glória Regina Franco
- Departamento de Bioquímica e Imunologia, ICB, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627, Caixa Postal 486, Belo Horizonte 30161-970, MG, Brazil.
| | - Sérgio Danilo Junho Pena
- Departamento de Bioquímica e Imunologia, ICB, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627, Caixa Postal 486, Belo Horizonte 30161-970, MG, Brazil.
| | - Lucía Valenzuela
- Programa de Biología Celular y Molecular, Instituto de Ciencias Biomédicas, Facultad de Medicina, Universidad de Chile, Santiago, Chile
| | - Simon Wisnovsky
- Department of Biochemistry, Faculty of Medicine, University of Toronto, Toronto, Ontario, Canada
| | - Shana O Kelley
- Department of Biochemistry, Faculty of Medicine, University of Toronto, Toronto, Ontario, Canada.
| | - Norbel Galanti
- Programa de Biología Celular y Molecular, Instituto de Ciencias Biomédicas, Facultad de Medicina, Universidad de Chile, Santiago, Chile.
| | - Gonzalo Cabrera
- Programa de Biología Celular y Molecular, Instituto de Ciencias Biomédicas, Facultad de Medicina, Universidad de Chile, Santiago, Chile.
| | - Carlos Renato Machado
- Departamento de Bioquímica e Imunologia, ICB, Universidade Federal de Minas Gerais, Av. Antônio Carlos, 6627, Caixa Postal 486, Belo Horizonte 30161-970, MG, Brazil.
| |
Collapse
|
|
6
|
Banda DM, Nuñez NN, Burnside MA, Bradshaw KM, David SS. Repair of 8-oxoG:A mismatches by the MUTYH glycosylase: Mechanism, metals and medicine. Free Radic Biol Med 2017; 107:202-215. [PMID: 28087410 PMCID: PMC5457711 DOI: 10.1016/j.freeradbiomed.2017.01.008] [Citation(s) in RCA: 71] [Impact Index Per Article: 8.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/17/2016] [Revised: 01/01/2017] [Accepted: 01/04/2017] [Indexed: 12/12/2022]
Abstract
Reactive oxygen and nitrogen species (RONS) may infringe on the passing of pristine genetic information by inducing DNA inter- and intra-strand crosslinks, protein-DNA crosslinks, and chemical alterations to the sugar or base moieties of DNA. 8-Oxo-7,8-dihydroguanine (8-oxoG) is one of the most prevalent DNA lesions formed by RONS and is repaired through the base excision repair (BER) pathway involving the DNA repair glycosylases OGG1 and MUTYH in eukaryotes. MUTYH removes adenine (A) from 8-oxoG:A mispairs, thus mitigating the potential of G:C to T:A transversion mutations from occurring in the genome. The paramount role of MUTYH in guarding the genome is well established in the etiology of a colorectal cancer predisposition syndrome involving variants of MUTYH, referred to as MUTYH-associated polyposis (MAP). In this review, we highlight recent advances in understanding how MUTYH structure and related function participate in the manifestation of human disease such as MAP. Here we focus on the importance of MUTYH's metal cofactor sites, including a recently discovered "Zinc linchpin" motif, as well as updates to the catalytic mechanism. Finally, we touch on the insight gleaned from studies with MAP-associated MUTYH variants and recent advances in understanding the multifaceted roles of MUTYH in the cell, both in the prevention of mutagenesis and tumorigenesis.
Collapse
Affiliation(s)
- Douglas M Banda
- Department of Chemistry, University of California, Davis, One Shields Avenue, Davis, CA 95616, United States
| | - Nicole N Nuñez
- Department of Chemistry, University of California, Davis, One Shields Avenue, Davis, CA 95616, United States
| | - Michael A Burnside
- Department of Chemistry, University of California, Davis, One Shields Avenue, Davis, CA 95616, United States
| | - Katie M Bradshaw
- Department of Chemistry, University of California, Davis, One Shields Avenue, Davis, CA 95616, United States
| | - Sheila S David
- Department of Chemistry, University of California, Davis, One Shields Avenue, Davis, CA 95616, United States.
| |
Collapse
|
|
7
|
Pilati C, Shinde J, Alexandrov LB, Assié G, André T, Hélias‐Rodzewicz Z, Ducoudray R, Le Corre D, Zucman‐Rossi J, Emile J, Bertherat J, Letouzé E, Laurent‐Puig P. Mutational signature analysis identifies
MUTYH
deficiency in colorectal cancers and adrenocortical carcinomas. J Pathol 2017; 242:10-15. [DOI: 10.1002/path.4880] [Citation(s) in RCA: 99] [Impact Index Per Article: 12.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/10/2016] [Revised: 01/09/2017] [Accepted: 01/16/2017] [Indexed: 12/12/2022]
Affiliation(s)
- Camilla Pilati
- INSERM UMR‐S1147, Personalized Medicine, Pharmacogenomics, Therapeutic Optimization Université Paris Descartes Paris France
| | - Jayendra Shinde
- INSERM Unité Mixte de Recherche (UMR) 1162 Génomique Fonctionnelle des Tumeurs Solides, Equipe Labellisée Ligue contre le Cancer Paris France
- Université Paris Descartes Labex Immuno‐Oncology Sorbonne Paris Cité Paris France
- Université Paris 13 Sorbonne Paris Cité, Unité de Formation et de Recherche (UFR) Santé, Médecine, Biologie Humaine (SMBH) Bobigny France
- Université Paris Diderot Institut Universitaire d'Hématologie Paris France
| | - Ludmil B Alexandrov
- Theoretical Biology and Biophysics (T‐6) Los Alamos National Laboratory Los Alamos NM USA
- Center for Nonlinear Studies Los Alamos National Laboratory Los Alamos NM USA
| | - Guillaume Assié
- INSERM U1016, CNRS UMR 8104 Paris Descartes University Institut Cochin, Paris France
- Center for Rare Adrenal Diseases, Department of Endocrinology, Assistance Publique‐Hôpitaux de Paris Hôpital Cochin, Paris France
| | - Thierry André
- Department of Medical Oncology AP‐HP, Hospital Saint‐Antoine Paris France
- Université Pierre et Marie Curie (UMPC) Paris VI Paris France
| | - Zofia Hélias‐Rodzewicz
- Department of Pathology AP‐HP, Hôpital Ambroise Paré Paris France
- EA 4340, Université de Versailles Versailles France
| | - Romain Ducoudray
- Department of Pathology AP‐HP, Hôpital Ambroise Paré Paris France
- EA 4340, Université de Versailles Versailles France
| | - Delphine Le Corre
- INSERM UMR‐S1147, Personalized Medicine, Pharmacogenomics, Therapeutic Optimization Université Paris Descartes Paris France
| | - Jessica Zucman‐Rossi
- INSERM Unité Mixte de Recherche (UMR) 1162 Génomique Fonctionnelle des Tumeurs Solides, Equipe Labellisée Ligue contre le Cancer Paris France
- Université Paris Descartes Labex Immuno‐Oncology Sorbonne Paris Cité Paris France
- Université Paris 13 Sorbonne Paris Cité, Unité de Formation et de Recherche (UFR) Santé, Médecine, Biologie Humaine (SMBH) Bobigny France
- Université Paris Diderot Institut Universitaire d'Hématologie Paris France
| | - Jean‐François Emile
- Department of Pathology AP‐HP, Hôpital Ambroise Paré Paris France
- EA 4340, Université de Versailles Versailles France
| | - Jérôme Bertherat
- INSERM U1016, CNRS UMR 8104 Paris Descartes University Institut Cochin, Paris France
- Center for Rare Adrenal Diseases, Department of Endocrinology, Assistance Publique‐Hôpitaux de Paris Hôpital Cochin, Paris France
| | - Eric Letouzé
- INSERM Unité Mixte de Recherche (UMR) 1162 Génomique Fonctionnelle des Tumeurs Solides, Equipe Labellisée Ligue contre le Cancer Paris France
- Université Paris Descartes Labex Immuno‐Oncology Sorbonne Paris Cité Paris France
- Université Paris 13 Sorbonne Paris Cité, Unité de Formation et de Recherche (UFR) Santé, Médecine, Biologie Humaine (SMBH) Bobigny France
- Université Paris Diderot Institut Universitaire d'Hématologie Paris France
| | - Pierre Laurent‐Puig
- INSERM UMR‐S1147, Personalized Medicine, Pharmacogenomics, Therapeutic Optimization Université Paris Descartes Paris France
| |
Collapse
|
|
8
|
Komine K, Shimodaira H, Takao M, Soeda H, Zhang X, Takahashi M, Ishioka C. Functional Complementation Assay for 47 MUTYH Variants in a MutY-Disrupted Escherichia coli Strain. Hum Mutat 2016; 36:704-11. [PMID: 25820570 PMCID: PMC4682456 DOI: 10.1002/humu.22794] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/27/2014] [Accepted: 03/25/2015] [Indexed: 12/27/2022]
Abstract
MUTYH-associated polyposis (MAP) is an adenomatous polyposis transmitted in an autosomal-recessive pattern, involving biallelic inactivation of the MUTYH gene. Loss of a functional MUTYH protein will result in the accumulation of G:T mismatched DNA caused by oxidative damage. Although p.Y179C and p.G396D are the two most prevalent MUTYH variants, more than 200 missense variants have been detected. It is difficult to determine whether these variants are disease-causing mutations or single-nucleotide polymorphisms. To understand the functional consequences of these variants, we generated 47 MUTYH gene variants via site-directed mutagenesis, expressed the encoded proteins in MutY-disrupted Escherichia coli, and assessed their abilities to complement the functional deficiency in the E. coli by monitoring spontaneous mutation rates. Although the majority of variants exhibited intermediate complementation relative to the wild type, some variants severely interfered with this complementation. However, some variants retained functioning similar to the wild type. In silico predictions of functional effects demonstrated a good correlation. Structural prediction of MUTYH based on the MutY protein structure allowed us to interpret effects on the protein stability or catalytic activity. These data will be useful for evaluating the functional consequences of missense MUTYH variants detected in patients with suspected MAP.
Collapse
Affiliation(s)
- Keigo Komine
- Department of Clinical Oncology, Institute of Development, Aging and Cancer, Tohoku University, Aoba-ku, Sendai, Japan.,Department of Clinical Oncology, Tohoku University Hospital, Aoba-ku, Sendai, Japan
| | - Hideki Shimodaira
- Department of Clinical Oncology, Institute of Development, Aging and Cancer, Tohoku University, Aoba-ku, Sendai, Japan.,Department of Clinical Oncology, Tohoku University Hospital, Aoba-ku, Sendai, Japan
| | - Masashi Takao
- Department of Molecular Genetics, Institute of Development, Aging and Cancer, Tohoku University, Aoba-ku, Sendai, Japan
| | - Hiroshi Soeda
- Department of Clinical Oncology, Institute of Development, Aging and Cancer, Tohoku University, Aoba-ku, Sendai, Japan.,Department of Clinical Oncology, Tohoku University Hospital, Aoba-ku, Sendai, Japan
| | - Xiaofei Zhang
- Department of Clinical Oncology, Institute of Development, Aging and Cancer, Tohoku University, Aoba-ku, Sendai, Japan
| | - Masanobu Takahashi
- Department of Clinical Oncology, Institute of Development, Aging and Cancer, Tohoku University, Aoba-ku, Sendai, Japan.,Department of Clinical Oncology, Tohoku University Hospital, Aoba-ku, Sendai, Japan
| | - Chikashi Ishioka
- Department of Clinical Oncology, Institute of Development, Aging and Cancer, Tohoku University, Aoba-ku, Sendai, Japan.,Department of Clinical Oncology, Tohoku University Hospital, Aoba-ku, Sendai, Japan
| |
Collapse
|
|
9
|
Shinmura K, Kato H, Goto M, Yamada H, Tao H, Nakamura S, Sugimura H. Functional Evaluation of Nine Missense-Type Variants of the Human DNA Glycosylase Enzyme MUTYH in the Japanese Population. Hum Mutat 2016; 37:350-3. [PMID: 26694661 DOI: 10.1002/humu.22949] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/10/2015] [Accepted: 12/15/2015] [Indexed: 11/08/2022]
Abstract
Biallelic germline mutations of MUTYH, the gene encoding DNA glycosylase, cause MUTYH-associated polyposis (MAP), characterized by multiple colorectal adenomas and carcinoma(s). However, a considerable number of MUTYH variants are still functionally uncharacterized. Herein, we report the results of functional evaluation of nine missense-type MUTYH variant proteins in the Japanese population. The DNA glycosylase activity and ability to suppress mutations caused by 8-hydroxyguanine, an oxidized form of guanine, were examined for the nine variants of type 2 MUTYH, a nuclear form of the enzyme, by DNA cleavage activity assay and supF forward mutation assay, respectively. Both activities were severely defective in the p.N210S MUTYH type 2 variant corresponding to p.N238S in the reference MUTYH form and partially defective in p.R219G variant corresponding to p.R247G, but nearly fully retained in seven other variants examined. Our results suggest that p.N238S and p.R247G are likely to be pathogenic alleles for MAP.
Collapse
Affiliation(s)
- Kazuya Shinmura
- Department of Tumor Pathology, Hamamatsu University School of Medicine, Hamamatsu, Japan
| | - Hisami Kato
- Department of Tumor Pathology, Hamamatsu University School of Medicine, Hamamatsu, Japan
| | - Masanori Goto
- Division of Carcinogenesis and Prevention, National Cancer Center Research Institute, Tokyo, Japan
| | - Hidetaka Yamada
- Department of Tumor Pathology, Hamamatsu University School of Medicine, Hamamatsu, Japan
| | - Hong Tao
- Department of Tumor Pathology, Hamamatsu University School of Medicine, Hamamatsu, Japan
| | - Satoki Nakamura
- Department of Tumor Pathology, Hamamatsu University School of Medicine, Hamamatsu, Japan
| | - Haruhiko Sugimura
- Department of Tumor Pathology, Hamamatsu University School of Medicine, Hamamatsu, Japan
| |
Collapse
|
|
10
|
Friedhoff P, Li P, Gotthardt J. Protein-protein interactions in DNA mismatch repair. DNA Repair (Amst) 2015; 38:50-57. [PMID: 26725162 DOI: 10.1016/j.dnarep.2015.11.013] [Citation(s) in RCA: 35] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/02/2015] [Revised: 11/11/2015] [Accepted: 11/30/2015] [Indexed: 11/25/2022]
Abstract
The principal DNA mismatch repair proteins MutS and MutL are versatile enzymes that couple DNA mismatch or damage recognition to other cellular processes. Besides interaction with their DNA substrates this involves transient interactions with other proteins which is triggered by the DNA mismatch or damage and controlled by conformational changes. Both MutS and MutL proteins have ATPase activity, which adds another level to control their activity and interactions with DNA substrates and other proteins. Here we focus on the protein-protein interactions, protein interaction sites and the different levels of structural knowledge about the protein complexes formed with MutS and MutL during the mismatch repair reaction.
Collapse
Affiliation(s)
- Peter Friedhoff
- Institute for Biochemistry FB 08, Justus Liebig University, Heinrich-Buff-Ring 17, D-35392 Giessen, Germany.
| | - Pingping Li
- Institute for Biochemistry FB 08, Justus Liebig University, Heinrich-Buff-Ring 17, D-35392 Giessen, Germany
| | - Julia Gotthardt
- Institute for Biochemistry FB 08, Justus Liebig University, Heinrich-Buff-Ring 17, D-35392 Giessen, Germany
| |
Collapse
|
|
11
|
Oka S, Leon J, Tsuchimoto D, Sakumi K, Nakabeppu Y. MUTYH, an adenine DNA glycosylase, mediates p53 tumor suppression via PARP-dependent cell death. Oncogenesis 2014; 3:e121. [PMID: 25310643 PMCID: PMC4216901 DOI: 10.1038/oncsis.2014.35] [Citation(s) in RCA: 35] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/28/2014] [Revised: 07/15/2014] [Accepted: 08/25/2014] [Indexed: 12/12/2022] Open
Abstract
p53-regulated caspase-independent cell death has been implicated in suppression of tumorigenesis, however, the regulating mechanisms are poorly understood. We previously reported that 8-oxoguanine (8-oxoG) accumulation in nuclear DNA (nDNA) and mitochondrial DNA triggers two distinct caspase-independent cell death through buildup of single-strand DNA breaks by MutY homolog (MUTYH), an adenine DNA glycosylase. One pathway depends on poly-ADP-ribose polymerase (PARP) and the other depends on calpains. Deficiency of MUTYH causes MUTYH-associated familial adenomatous polyposis. MUTYH thereby suppresses tumorigenesis not only by avoiding mutagenesis, but also by inducing cell death. Here, we identified the functional p53-binding site in the human MUTYH gene and demonstrated that MUTYH is transcriptionally regulated by p53, especially in the p53/DNA mismatch repair enzyme, MLH1-proficient colorectal cancer-derived HCT116+Chr3 cells. MUTYH-small interfering RNA, an inhibitor for p53 or PARP suppressed cell death without an additive effect, thus revealing that MUTYH is a potential mediator of p53 tumor suppression, which is known to be upregulated by MLH1. Moreover, we found that the p53-proficient, mismatch repair protein, MLH1-proficient colorectal cancer cell line express substantial levels of MUTYH in nuclei but not in mitochondria, suggesting that 8-oxoG accumulation in nDNA triggers MLH1/PARP-dependent cell death. These results provide new insights on the molecular mechanism of tumorigenesis and potential new strategies for cancer therapies.
Collapse
Affiliation(s)
- S Oka
- 1] Division of Neurofunctional Genomics, Department of Immunobiology and Neuroscience, Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan [2] Research Center for Nucleotide Pool, Kyushu University, Fukuoka, Japan
| | - J Leon
- Division of Neurofunctional Genomics, Department of Immunobiology and Neuroscience, Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan
| | - D Tsuchimoto
- 1] Division of Neurofunctional Genomics, Department of Immunobiology and Neuroscience, Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan [2] Research Center for Nucleotide Pool, Kyushu University, Fukuoka, Japan
| | - K Sakumi
- 1] Division of Neurofunctional Genomics, Department of Immunobiology and Neuroscience, Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan [2] Research Center for Nucleotide Pool, Kyushu University, Fukuoka, Japan
| | - Y Nakabeppu
- 1] Division of Neurofunctional Genomics, Department of Immunobiology and Neuroscience, Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan [2] Research Center for Nucleotide Pool, Kyushu University, Fukuoka, Japan
| |
Collapse
|
|
12
|
Kurian AW, Hare EE, Mills MA, Kingham KE, McPherson L, Whittemore AS, McGuire V, Ladabaum U, Kobayashi Y, Lincoln SE, Cargill M, Ford JM. Clinical evaluation of a multiple-gene sequencing panel for hereditary cancer risk assessment. J Clin Oncol 2014; 32:2001-9. [PMID: 24733792 PMCID: PMC4067941 DOI: 10.1200/jco.2013.53.6607] [Citation(s) in RCA: 382] [Impact Index Per Article: 34.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022] Open
Abstract
PURPOSE Multiple-gene sequencing is entering practice, but its clinical value is unknown. We evaluated the performance of a customized germline-DNA sequencing panel for cancer-risk assessment in a representative clinical sample. METHODS Patients referred for clinical BRCA1/2 testing from 2002 to 2012 were invited to donate a research blood sample. Samples were frozen at -80° C, and DNA was extracted from them after 1 to 10 years. The entire coding region, exon-intron boundaries, and all known pathogenic variants in other regions were sequenced for 42 genes that had cancer risk associations. Potentially actionable results were disclosed to participants. RESULTS In total, 198 women participated in the study: 174 had breast cancer and 57 carried germline BRCA1/2 mutations. BRCA1/2 analysis was fully concordant with prior testing. Sixteen pathogenic variants were identified in ATM, BLM, CDH1, CDKN2A, MUTYH, MLH1, NBN, PRSS1, and SLX4 among 141 women without BRCA1/2 mutations. Fourteen participants carried 15 pathogenic variants, warranting a possible change in care; they were invited for targeted screening recommendations, enabling early detection and removal of a tubular adenoma by colonoscopy. Participants carried an average of 2.1 variants of uncertain significance among 42 genes. CONCLUSION Among women testing negative for BRCA1/2 mutations, multiple-gene sequencing identified 16 potentially pathogenic mutations in other genes (11.4%; 95% CI, 7.0% to 17.7%), of which 15 (10.6%; 95% CI, 6.5% to 16.9%) prompted consideration of a change in care, enabling early detection of a precancerous colon polyp. Additional studies are required to quantify the penetrance of identified mutations and determine clinical utility. However, these results suggest that multiple-gene sequencing may benefit appropriately selected patients.
Collapse
Affiliation(s)
- Allison W Kurian
- Allison W. Kurian, Meredith A. Mills, Kerry E. Kingham, Lisa McPherson, Alice S. Whittemore, Valerie McGuire, Uri Ladabaum, James M. Ford, Stanford University School of Medicine, Stanford; Emily E. Hare, Yuya Kobayashi, Stephen E. Lincoln, Michele Cargill, InVitae, San Francisco, CA
| | - Emily E Hare
- Allison W. Kurian, Meredith A. Mills, Kerry E. Kingham, Lisa McPherson, Alice S. Whittemore, Valerie McGuire, Uri Ladabaum, James M. Ford, Stanford University School of Medicine, Stanford; Emily E. Hare, Yuya Kobayashi, Stephen E. Lincoln, Michele Cargill, InVitae, San Francisco, CA
| | - Meredith A Mills
- Allison W. Kurian, Meredith A. Mills, Kerry E. Kingham, Lisa McPherson, Alice S. Whittemore, Valerie McGuire, Uri Ladabaum, James M. Ford, Stanford University School of Medicine, Stanford; Emily E. Hare, Yuya Kobayashi, Stephen E. Lincoln, Michele Cargill, InVitae, San Francisco, CA
| | - Kerry E Kingham
- Allison W. Kurian, Meredith A. Mills, Kerry E. Kingham, Lisa McPherson, Alice S. Whittemore, Valerie McGuire, Uri Ladabaum, James M. Ford, Stanford University School of Medicine, Stanford; Emily E. Hare, Yuya Kobayashi, Stephen E. Lincoln, Michele Cargill, InVitae, San Francisco, CA
| | - Lisa McPherson
- Allison W. Kurian, Meredith A. Mills, Kerry E. Kingham, Lisa McPherson, Alice S. Whittemore, Valerie McGuire, Uri Ladabaum, James M. Ford, Stanford University School of Medicine, Stanford; Emily E. Hare, Yuya Kobayashi, Stephen E. Lincoln, Michele Cargill, InVitae, San Francisco, CA
| | - Alice S Whittemore
- Allison W. Kurian, Meredith A. Mills, Kerry E. Kingham, Lisa McPherson, Alice S. Whittemore, Valerie McGuire, Uri Ladabaum, James M. Ford, Stanford University School of Medicine, Stanford; Emily E. Hare, Yuya Kobayashi, Stephen E. Lincoln, Michele Cargill, InVitae, San Francisco, CA
| | - Valerie McGuire
- Allison W. Kurian, Meredith A. Mills, Kerry E. Kingham, Lisa McPherson, Alice S. Whittemore, Valerie McGuire, Uri Ladabaum, James M. Ford, Stanford University School of Medicine, Stanford; Emily E. Hare, Yuya Kobayashi, Stephen E. Lincoln, Michele Cargill, InVitae, San Francisco, CA
| | - Uri Ladabaum
- Allison W. Kurian, Meredith A. Mills, Kerry E. Kingham, Lisa McPherson, Alice S. Whittemore, Valerie McGuire, Uri Ladabaum, James M. Ford, Stanford University School of Medicine, Stanford; Emily E. Hare, Yuya Kobayashi, Stephen E. Lincoln, Michele Cargill, InVitae, San Francisco, CA
| | - Yuya Kobayashi
- Allison W. Kurian, Meredith A. Mills, Kerry E. Kingham, Lisa McPherson, Alice S. Whittemore, Valerie McGuire, Uri Ladabaum, James M. Ford, Stanford University School of Medicine, Stanford; Emily E. Hare, Yuya Kobayashi, Stephen E. Lincoln, Michele Cargill, InVitae, San Francisco, CA
| | - Stephen E Lincoln
- Allison W. Kurian, Meredith A. Mills, Kerry E. Kingham, Lisa McPherson, Alice S. Whittemore, Valerie McGuire, Uri Ladabaum, James M. Ford, Stanford University School of Medicine, Stanford; Emily E. Hare, Yuya Kobayashi, Stephen E. Lincoln, Michele Cargill, InVitae, San Francisco, CA
| | - Michele Cargill
- Allison W. Kurian, Meredith A. Mills, Kerry E. Kingham, Lisa McPherson, Alice S. Whittemore, Valerie McGuire, Uri Ladabaum, James M. Ford, Stanford University School of Medicine, Stanford; Emily E. Hare, Yuya Kobayashi, Stephen E. Lincoln, Michele Cargill, InVitae, San Francisco, CA
| | - James M Ford
- Allison W. Kurian, Meredith A. Mills, Kerry E. Kingham, Lisa McPherson, Alice S. Whittemore, Valerie McGuire, Uri Ladabaum, James M. Ford, Stanford University School of Medicine, Stanford; Emily E. Hare, Yuya Kobayashi, Stephen E. Lincoln, Michele Cargill, InVitae, San Francisco, CA.
| |
Collapse
|
|
13
|
Impaired 8-hydroxyguanine repair activity of MUTYH variant p.Arg109Trp found in a Japanese patient with early-onset colorectal cancer. OXIDATIVE MEDICINE AND CELLULAR LONGEVITY 2014; 2014:617351. [PMID: 24799981 PMCID: PMC3988950 DOI: 10.1155/2014/617351] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 11/29/2013] [Revised: 02/05/2014] [Accepted: 02/11/2014] [Indexed: 12/13/2022]
Abstract
PURPOSE The biallelic inactivation of the 8-hydroxyguanine repair gene MUTYH leads to MUTYH-associated polyposis (MAP), which is characterized by colorectal multiple polyps and carcinoma(s). However, only limited information regarding MAP in the Japanese population is presently available. Since early-onset colorectal cancer (CRC) is a characteristic of MAP and might be caused by the inactivation of another 8-hydroxyguanine repair gene, OGG1, we investigated whether germline MUTYH and OGG1 mutations are involved in early-onset CRC in Japanese patients. METHODS Thirty-four Japanese patients with early-onset CRC were examined for germline MUTYH and OGG1 mutations using sequencing. RESULTS Biallelic pathogenic mutations were not found in any of the patients; however, a heterozygous p.Arg19∗ MUTYH variant and a heterozygous p.Arg109Trp MUTYH variant were detected in one patient each. The p.Arg19∗ and p.Arg109Trp corresponded to p.Arg5∗ and p.Arg81Trp, respectively, in the type 2 nuclear-form protein. The defective DNA repair activity of p.Arg5∗ is apparent, while that of p.Arg81Trp has been demonstrated using DNA cleavage and supF forward mutation assays. CONCLUSION These results suggest that biallelic MUTYH or OGG1 pathogenic mutations are rare in Japanese patients with early-onset CRC; however, the p.Arg19∗ and p.Arg109Trp MUTYH variants are associated with functional impairments.
Collapse
|
|
14
|
Guarinos C, Juárez M, Egoavil C, Rodríguez-Soler M, Pérez-Carbonell L, Salas R, Cubiella J, Rodríguez-Moranta F, de-Castro L, Bujanda L, Serradesanferm A, Nicolás-Pérez D, Herráiz M, Fernández-Bañares F, Herreros-de-Tejada A, Aguirre E, Balmaña J, Rincón ML, Pizarro A, Polo-Ortiz F, Castillejo A, Alenda C, Payá A, Soto JL, Jover R. Prevalence and characteristics of MUTYH-associated polyposis in patients with multiple adenomatous and serrated polyps. Clin Cancer Res 2014; 20:1158-68. [PMID: 24470512 DOI: 10.1158/1078-0432.ccr-13-1490] [Citation(s) in RCA: 43] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
PURPOSE The present study aimed to determine the prevalence of MUTYH mutations in patients with multiple colonic polyps and to explore the best strategy for diagnosing MUTYH-associated polyposis (MAP) in these patients. EXPERIMENTAL DESIGN This study included 405 patients with at least 10 colonic polyps each. All cases were genetically tested for the three most frequent MUTYH mutations. Whole-gene analysis was performed in heterozygous patients and in 216 patients lacking the three most frequent mutations. Polyps from 56 patients were analyzed for the KRAS-Gly12Cys and BRAF V600E somatic mutations. RESULTS Twenty-seven (6.7%) patients were diagnosed with MAP, of which 40.8% showed serrated polyps. The sensitivity of studying only the three common variants was 74.1%. Of 216 patients without any monoallelic mutation in common variants, whole-gene analysis revealed biallelic pathogenic mutation in only one. G396D mutation was associated with serrated lesions and older age at diagnosis. There was a strong association between germinal MUTYH mutation and KRAS Gly12Cys somatic mutation in polyps. BRAF V600E mutation was found in 74% of serrated polyps in MUTYH-negative patients and in none of the polyps of MAP patients. CONCLUSIONS We observed a low frequency of MUTYH mutations among patients with multiple adenomatous and serrated polyps. The MAP phenotype frequently included patients with serrated polyps, especially when G396D mutation was involved. Our results show that somatic molecular markers of polyps can be useful in identifying MAP cases and support the need for the complete MUTYH gene analysis only in patients heterozygous for recurrent variants.
Collapse
Affiliation(s)
- Carla Guarinos
- Authors' Affiliations: Unidad de Investigación, Departments of Pathology, and Gastroenterology, Hospital General Universitario, Alicante; Department of Gastroenterology, Complexo Hospitalario Universitario de Ourense, Ourense; Department of Gastroenterology, Bellvitge University Hospital; Institut de Malaties Digestives i Metabòliques, CIBERehd, Hospital Clínic; Department of Oncology, Hospital Vall d'Hebrón, Barcelona; Gastroenterology Department, Complexo Hospitalario de Vigo, Vigo; Department of Gastroenterology, Hospital Donostia, CIBERehd, Universidad del País Vasco, San Sebastián; Department of Gastroenterology, Hospital Universitario de Canarias, La Laguna, Tenerife; Department of Gastroenterology, Clínica Universitaria de Navarra, Pamplona; Department of Gastroenterology, Hospital Mútua de Terrassa, Terrassa; Department of Gastroenterology, Hospital Puerta de Hierro, Madrid; Department of Oncology, Hospital Arnau de Vilanova, Lleida; Department of Gastroenterology, Hospital Bidasoa, Irún; Department of Gastroenterology, Hospital Universitario Virgen del Rocío, Sevilla; Department of Gastroenterology, Hospital Basurto, Bilbao; Department of Molecular Genetics, Hospital General Universitario, Elche, Spain; and Gastrointestinal Cancer Research Laboratory, Baylor Research Institute, Baylor University Medical Center, Dallas, Texas
| | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | | |
Collapse
|
|
15
|
MUTYH-associated colorectal cancer and adenomatous polyposis. Surg Today 2013; 44:593-600. [PMID: 23605219 DOI: 10.1007/s00595-013-0592-7] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2012] [Accepted: 03/04/2013] [Indexed: 12/28/2022]
Abstract
MUTYH-associated polyposis (MAP) was first described in 2002. MUTYH is a component of a base excision repair system that protects the genomic information from oxidative damage. When the MUTYH gene product is impaired by bi-allelic germline mutation, it leads to the mutation of cancer-related genes, such as the APC and/or the KRAS genes, via G to T transversion. MAP is a hereditary colorectal cancer syndrome inherited in an autosomal-recessive fashion. The clinical features of MAP include the presence of 10-100 adenomatous polyps in the colon, and early onset of colorectal cancer. Ethnic and geographical differences in the pattern of the MUTYH gene mutations have been suggested. In Caucasian patients, c.536A>G (Y179C) and c.1187G>A (G396D) mutations are frequently detected. In the Asian population, Y179C and G396D are uncommon, whereas other variants are suggested to be the major causes of MAP. We herein review the literature on MUTYH-associated colorectal cancer and adenomatous polyposis.
Collapse
|
|
16
|
Abstract
The mismatch repair (MMR) system detects non-Watson-Crick base pairs and strand misalignments arising during DNA replication and mediates their removal by catalyzing excision of the mispair-containing tract of nascent DNA and its error-free resynthesis. In this way, MMR improves the fidelity of replication by several orders of magnitude. It also addresses mispairs and strand misalignments arising during recombination and prevents synapses between nonidentical DNA sequences. Unsurprisingly, MMR malfunction brings about genomic instability that leads to cancer in mammals. But MMR proteins have recently been implicated also in other processes of DNA metabolism, such as DNA damage signaling, antibody diversification, and repair of interstrand cross-links and oxidative DNA damage, in which their functions remain to be elucidated. This article reviews the progress in our understanding of the mechanism of replication error repair made during the past decade.
Collapse
Affiliation(s)
- Josef Jiricny
- Institute of Molecular Cancer Research, University of Zurich and ETH Zurich, 8057 Zurich, Switzerland.
| |
Collapse
|
|
17
|
Turco E, Ventura I, Minoprio A, Russo MT, Torreri P, Degan P, Molatore S, Ranzani GN, Bignami M, Mazzei F. Understanding the role of the Q338H MUTYH variant in oxidative damage repair. Nucleic Acids Res 2013; 41:4093-103. [PMID: 23460202 PMCID: PMC3627602 DOI: 10.1093/nar/gkt130] [Citation(s) in RCA: 24] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022] Open
Abstract
The MUTYH DNA–glycosylase is indirectly engaged in the repair of the miscoding 7,8-dihydro-8-oxo-2′-deoxyguanine (8-oxodG) lesion by removing adenine erroneously incorporated opposite the oxidized purine. Inherited biallelic mutations in the MUTYH gene are responsible for a recessive syndrome, the MUTYH-associated polyposis (MAP), which confers an increased risk of colorectal cancer. In this study, we functionally characterized the Q338H variant using recombinant proteins, as well as cell-based assays. This is a common variant among human colorectal cancer genes, which is generally considered, unrelated to the MAP phenotype but recently indicated as a low-penetrance allele. We demonstrate that the Q338H variant retains a wild-type DNA–glycosylase activity in vitro, but it shows a reduced ability to interact with the replication sensor RAD9:RAD1:HUS1 (9–1–1) complex. In comparison with Mutyh−/− mouse embryo fibroblasts expressing a wild-type MUTYH cDNA, the expression of Q338H variant was associated with increased levels of DNA 8-oxodG, hypersensitivity to oxidant and accumulation of the population in the S phase of the cell cycle. Thus, an inefficient interaction of MUTYH with the 9–1–1 complex leads to a repair-defective phenotype, indicating that a proper communication between MUTYH enzymatic function and the S phase checkpoint is needed for effective repair of oxidative damage.
Collapse
Affiliation(s)
- Eleonora Turco
- Department of Environment and Primary Prevention, Istituto Superiore di Sanità, 00161 Roma, Italy
| | | | | | | | | | | | | | | | | | | |
Collapse
|
|
18
|
Mazzei F, Viel A, Bignami M. Role of MUTYH in human cancer. Mutat Res 2013; 743-744:33-43. [PMID: 23507534 DOI: 10.1016/j.mrfmmm.2013.03.003] [Citation(s) in RCA: 70] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/26/2012] [Revised: 03/06/2013] [Accepted: 03/09/2013] [Indexed: 06/01/2023]
Abstract
MUTYH, a human ortholog of MutY, is a post-replicative DNA glycosylase, highly conserved throughout evolution, involved in the correction of mismatches resulting from a faulty replication of the oxidized base 8-hydroxyguanine (8-oxodG). In particular removal of adenine from A:8-oxodG mispairs by MUTYH activity is followed by error-free base excision repair (BER) events, leading to the formation of C:8-oxodG base pairs. These are the substrate of another BER enzyme, the OGG1 DNA glycosylase, which then removes 8-oxodG from DNA. Thus the combined action of OGG1 and MUTYH prevents oxidative damage-induced mutations, i.e. GC->TA transversions. Germline mutations in MUTYH are associated with a recessively heritable colorectal polyposis, now referred to as MUTYH-associated polyposis (MAP). Here we will review the phenotype(s) associated with MUTYH inactivation from bacteria to mammals, the structure of the MUTYH protein, the molecular mechanisms of its enzymatic activity and the functional characterization of MUTYH variants. The relevance of these results will be discussed to define the role of specific human mutations in colorectal cancer risk together with the possible role of MUTYH inactivation in sporadic cancer.
Collapse
Affiliation(s)
- Filomena Mazzei
- Department of Environment, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Roma, Italy
| | - Alessandra Viel
- Experimental Oncology 1, Centro di Riferimento Oncologico, IRCCS, Via F.Gallini 2, 33081 Aviano, PN, Italy
| | - Margherita Bignami
- Department of Environment, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Roma, Italy.
| |
Collapse
|
|
19
|
Edelbrock MA, Kaliyaperumal S, Williams KJ. Structural, molecular and cellular functions of MSH2 and MSH6 during DNA mismatch repair, damage signaling and other noncanonical activities. Mutat Res 2013; 743-744:53-66. [PMID: 23391514 DOI: 10.1016/j.mrfmmm.2012.12.008] [Citation(s) in RCA: 95] [Impact Index Per Article: 7.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/05/2012] [Revised: 12/28/2012] [Accepted: 12/31/2012] [Indexed: 11/18/2022]
Abstract
The field of DNA mismatch repair (MMR) has rapidly expanded after the discovery of the MutHLS repair system in bacteria. By the mid 1990s yeast and human homologues to bacterial MutL and MutS had been identified and their contribution to hereditary non-polyposis colorectal cancer (HNPCC; Lynch syndrome) was under intense investigation. The human MutS homologue 6 protein (hMSH6), was first reported in 1995 as a G:T binding partner (GTBP) of hMSH2, forming the hMutSα mismatch-binding complex. Signal transduction from each DNA-bound hMutSα complex is accomplished by the hMutLα heterodimer (hMLH1 and hPMS2). Molecular mechanisms and cellular regulation of individual MMR proteins are now areas of intensive research. This review will focus on molecular mechanisms associated with mismatch binding, as well as emerging evidence that MutSα, and in particular, MSH6, is a key protein in MMR-dependent DNA damage response and communication with other DNA repair pathways within the cell. MSH6 is unstable in the absence of MSH2, however it is the DNA lesion-binding partner of this heterodimer. MSH6, but not MSH2, has a conserved Phe-X-Glu motif that recognizes and binds several different DNA structural distortions, initiating different cellular responses. hMSH6 also contains the nuclear localization sequences required to shuttle hMutSα into the nucleus. For example, upon binding to O(6)meG:T, MSH6 triggers a DNA damage response that involves altered phosphorylation within the N-terminal disordered domain of this unique protein. While many investigations have focused on MMR as a post-replication DNA repair mechanism, MMR proteins are expressed and active in all phases of the cell cycle. There is much more to be discovered about regulatory cellular roles that require the presence of MutSα and, in particular, MSH6.
Collapse
Affiliation(s)
| | - Saravanan Kaliyaperumal
- Division of Comparative Medicine and Pathology, New England Primate Research Center, One Pine Hill Drive, Southborough, MA 01772, USA.
| | - Kandace J Williams
- University of Toledo College of Medicine and Life Sciences, Department of Biochemistry & Cancer Biology, 3000 Transverse Dr., Toledo, OH 43614, USA.
| |
Collapse
|
|
20
|
Shinmura K, Goto M, Tao H, Matsuura S, Matsuda T, Sugimura H. Impaired suppressive activities of human MUTYH variant proteins against oxidative mutagenesis. World J Gastroenterol 2012; 18:6935-42. [PMID: 23322991 PMCID: PMC3531677 DOI: 10.3748/wjg.v18.i47.6935] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/20/2012] [Revised: 09/19/2012] [Accepted: 09/22/2012] [Indexed: 02/06/2023] Open
Abstract
AIM: To investigate the suppressive activity of MUTYH variant proteins against mutations caused by oxidative lesion, 8-hydroxyguanine (8OHG), in human cells.
METHODS: p.R154H, p.M255V, p.L360P, and p.P377L MUTYH variants, which were previously found in patients with colorectal polyposis and cancer, were selected for use in this study. Human H1299 cancer cell lines inducibly expressing wild-type (WT) MUTYH (type 2) or one of the 4 above-mentioned MUTYH variants were established using the piggyBac transposon vector system, enabling the genomic integration of the transposon sequence for MUTYH expression. MUTYH expression was examined after cumate induction using Western blotting analysis and immunofluorescence analysis. The intracellular localization of MUTYH variants tagged with FLAG was also immunofluorescently examined. Next, the mutation frequency in the supF of the shuttle plasmid pMY189 containing a single 8OHG residue at position 159 of the supF was compared between empty vector cells and cells expressing WT MUTYH or one of the 4 MUTYH variants using a supF forward mutation assay.
RESULTS: The successful establishment of human cell lines inducibly expressing WT MUTYH or one of the 4 MUTYH variants was concluded based on the detection of MUTYH expression in these cell lines after treatment with cumate. All of the MUTYH variants and WT MUTYH were localized in the nucleus, and nuclear localization was also observed for FLAG-tagged MUTYH. The mutation frequency of supF was 2.2 × 10-2 in the 8OHG-containing pMY189 plasmid and 2.5 × 10-4 in WT pMY189 in empty vector cells, which was an 86-fold increase with the introduction of 8OHG. The mutation frequency (4.7 × 10-3) of supF in the 8OHG-containing pMY189 plasmid in cells overexpressing WT MUTYH was significantly lower than in the empty vector cells (P < 0.01). However, the mutation frequencies of the supF in the 8OHG-containing pMY189 plasmid in cells overexpressing the p.R154H, p.M255V, p.L360P, or p.P377L MUTYH variant were 1.84 × 10-2, 1.55 × 10-2, 1.91 × 10-2, and 1.96 × 10-2, respectively, meaning that no significant difference was observed in the mutation frequency between the empty vector cells and cells overexpressing MUTYH mutants.
CONCLUSION: The suppressive activities of p.R154H, p.M255V, p.L360P, and p.P377L MUTYH variants against mutations caused by 8OHG are thought to be severely impaired in human cells.
Collapse
|
|
21
|
Venesio T, Balsamo A, D'Agostino VG, Ranzani GN. MUTYH-associated polyposis (MAP), the syndrome implicating base excision repair in inherited predisposition to colorectal tumors. Front Oncol 2012; 2:83. [PMID: 22876359 PMCID: PMC3410368 DOI: 10.3389/fonc.2012.00083] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/11/2012] [Accepted: 07/16/2012] [Indexed: 12/24/2022] Open
Abstract
In 2002, Al-Tassan and co-workers described for the first time a recessive form of inherited polyposis associated with germline mutations of MUTYH, a gene encoding a base excision repair (BER) protein that counteracts the DNA damage induced by the oxidative stress. MUTYH-associated polyposis (MAP) is now a well-defined cancer susceptibility syndrome, showing peculiar molecular features that characterize disease progression. However, some aspects of MAP, including diagnostic criteria, genotype-phenotype correlations, pathogenicity of variants, as well as relationships between BER and other DNA repair pathways, are still poorly understood. A deeper knowledge of the MUTYH expression pattern is likely to refine our understanding of the protein role and, finally, to improve guidances for identifying and handling MAP patients.
Collapse
Affiliation(s)
- Tiziana Venesio
- Unit of Pathology, Institute for Cancer Research and Treatment Candiolo, Torino, Italy
| | | | | | | |
Collapse
|
|
22
|
Wallace SS, Murphy DL, Sweasy JB. Base excision repair and cancer. Cancer Lett 2012; 327:73-89. [PMID: 22252118 DOI: 10.1016/j.canlet.2011.12.038] [Citation(s) in RCA: 234] [Impact Index Per Article: 18.0] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/22/2011] [Revised: 12/20/2011] [Accepted: 12/24/2011] [Indexed: 01/13/2023]
Abstract
Base excision repair is the system used from bacteria to man to remove the tens of thousands of endogenous DNA damages produced daily in each human cell. Base excision repair is required for normal mammalian development and defects have been associated with neurological disorders and cancer. In this paper we provide an overview of short patch base excision repair in humans and summarize current knowledge of defects in base excision repair in mouse models and functional studies on short patch base excision repair germ line polymorphisms and their relationship to cancer. The biallelic germ line mutations that result in MUTYH-associated colon cancer are also discussed.
Collapse
Affiliation(s)
- Susan S Wallace
- Department of Microbiology and Molecular Genetics, The Markey Center for Molecular Genetics, University of Vermont, Burlington, 05405-0068, United States.
| | | | | |
Collapse
|
|
23
|
Poulsen MLM, Bisgaard ML. MUTYH Associated Polyposis (MAP). Curr Genomics 2011; 9:420-35. [PMID: 19506731 PMCID: PMC2691665 DOI: 10.2174/138920208785699562] [Citation(s) in RCA: 63] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/18/2008] [Revised: 04/28/2008] [Accepted: 05/04/2008] [Indexed: 01/04/2023] Open
Abstract
MUTYH Associated Polyposis (MAP), a Polyposis predisposition caused by biallelic mutations in the Base Excision Repair (BER) gene MUTYH, confers a marked risk of colorectal cancer (CRC). The MAP phenotype is difficult to distinguish from other hereditary CRC syndromes. Especially from Familial Adenomatous Polyposis (FAP) and to a lesser extend Lynch Syndrome, which are caused by germline mutations in the APC and Mismatch Repair (MMR) genes, respectively. Here we review research findings regarding MUTYH interactions, genotypic and phenotypic characteristics of MAP, as well as surveillance and treatment of the disease. The applied papers, published between 1/1 2002- 1/2 2008, were found through PubMed. The exact role of MUTYH in CRC tumorgenesis is still uncertain, although MAP tumors show distinct molecular features, including somatic G:C>T:A transversions in the APC gene. Furthermore, cooperation between the BER and the MMR systems exists, as MUTYH interacts with MMR gene-products. Possibly, monoallelic defects in both pathways are of significance to CRC development. Specific MUTYH variants are found to be characteristic in distinct ethnic populations, which could facilitate future genetic screening. Knowledge concerning functional consequences of many MUTYH germline mutations remains sparse. Most thoroughly investigated are the two most common MUTYH variants, Y179C and G396D, both generating dysfunctional gene products. Phenotypic features of MAP include: development of 10-100 colorectal adenomas, debuting at 46-47 years, often CRC at time of clinical diagnosis, and in some, development of extracolonic manifestations.
Collapse
Affiliation(s)
- M L M Poulsen
- Department of Cellular and Molecular Medicine, University of Copenhagen, 2200 Copenhagen N, Denmark
| | | |
Collapse
|
|
24
|
Gu Y, Fang TT, Chao TH. Modification of allele-specific polymerase chain reaction for hMYH mutation analysis in Taiwanese patients with colorectal cancer. J Taiwan Inst Chem Eng 2011. [DOI: 10.1016/j.jtice.2010.11.002] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/18/2022]
|
|
25
|
Romano CA, Sontz PA, Barton JK. Mutants of the base excision repair glycosylase, endonuclease III: DNA charge transport as a first step in lesion detection. Biochemistry 2011; 50:6133-45. [PMID: 21651304 DOI: 10.1021/bi2003179] [Citation(s) in RCA: 37] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
Abstract
Endonuclease III (EndoIII) is a base excision repair glycosylase that targets damaged pyrimidines and contains a [4Fe-4S] cluster. We have proposed a model where BER proteins that contain redox-active [4Fe-4S] clusters utilize DNA charge transport (CT) as a first step in the detection of DNA lesions. Here, several mutants of EndoIII were prepared to probe their efficiency of DNA/protein charge transport. Cyclic voltammetry experiments on DNA-modified electrodes show that aromatic residues F30, Y55, Y75, and Y82 help mediate charge transport between DNA and the [4Fe-4S] cluster. On the basis of circular dichroism studies to measure protein stability, mutations at residues W178 and Y185 are found to destabilize the protein; these residues may function to protect the [4Fe-4S] cluster. Atomic force microscopy studies furthermore reveal a correlation in the ability of mutants to carry out protein/DNA CT and their ability to relocalize onto DNA strands containing a single base mismatch; EndoIII mutants that are defective in carrying out DNA/protein CT do not redistribute onto mismatch-containing strands, consistent with our model. These results demonstrate a link between the ability of the repair protein to carry out DNA CT and its ability to relocalize near lesions, thus pointing to DNA CT as a key first step in the detection of base damage in the genome.
Collapse
Affiliation(s)
- Christine A Romano
- Division of Chemistry and Chemical Engineering, California Institute of Technolog, Pasadena, California 91125, USA
| | | | | |
Collapse
|
|
26
|
Morak M, Laner A, Bacher U, Keiling C, Holinski-Feder E. MUTYH-associated polyposis - variability of the clinical phenotype in patients with biallelic and monoallelic MUTYH mutations and report on novel mutations. Clin Genet 2011; 78:353-63. [PMID: 20618354 DOI: 10.1111/j.1399-0004.2010.01478.x] [Citation(s) in RCA: 34] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
To further characterize 215 APC mutation-negative patients with colorectal neoplasias classified in classical, attenuated, or atypical familial adenomatous polyposis (FAP) coli we performed mutation screening in the Mut Y homologue (MUTYH) gene. The incidence was 15% for biallelic and 3.7% for monoallelic MUTYH mutations. We describe six novel MUTYH mutations in biallelic constellation and two novel monoallelic missense mutations. Of 33 MUTYH-associated polyposis coli (MAP) patients 57% were attenuated familial adenomatous polyposis (AFAP) patients, 10% display early-onset classical FAP and 18% had only few adenomas at higher age. Biallelic cases had a high incidence of extracolonic polyposis in 32% and colorectal cancer (CRC) in 33% of the cases. The clinical picture of MAP ranged from classical FAP or synchronous CRC at age 30 years to few adenomas at age 54 years without evidence of CRC, initially suspected for hereditary non-polyposis colorectal cancer (HNPCC). The mean age of onset was 43 years, with 11 (33%) patients being younger than 40 years of age, indicating that the clinical manifestation can be earlier than so far reported. Monoallelic MUTYH mutation carriers had a positive family history in seven of eight cases allowing the hypothesis of a disease-causing synergism of MUTYH mutations with other genes.
Collapse
Affiliation(s)
- M Morak
- University Hospital of the Ludwig-Maximilians-University, Campus Innenstadt, Munich, Germany
| | | | | | | | | |
Collapse
|
|
27
|
Goto M, Shinmura K, Nakabeppu Y, Tao H, Yamada H, Tsuneyoshi T, Sugimura H. Adenine DNA glycosylase activity of 14 human MutY homolog (MUTYH) variant proteins found in patients with colorectal polyposis and cancer. Hum Mutat 2010; 31:E1861-74. [PMID: 20848659 PMCID: PMC3051265 DOI: 10.1002/humu.21363] [Citation(s) in RCA: 35] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022]
Abstract
Biallelic inactivating germline mutations in the base excision repair MUTYH (MYH) gene have been shown to predispose to MUTYH-associated polyposis (MAP), which is characterized by multiple colorectal adenomas and carcinomas. In this study, we successfully prepared highly homogeneous human MUTYH type 2 recombinant proteins and compared the DNA glycosylase activity of the wild-type protein and fourteen variant-type proteins on adenine mispaired with 8-hydroxyguanine, an oxidized form of guanine. The adenine DNA glycosylase activity of the p.I195V protein, p.G368D protein, p.M255V protein, and p.Y151C protein was 66.9%, 15.2%, 10.7%, and 4.5%, respectively, of that of the wild-type protein, and the glycosylase activity of the p.R154H, p.L360P, p.P377L, p.452delE, p.R69X, and p.Q310X proteins as well as of the p.D208N negative control form was extremely severely impaired. The glycosylase activity of the p.V47E, p.R281C, p.A345V, and p.S487F proteins, on the other hand, was almost the same as that of the wild-type protein. These results should be of great value in accurately diagnosing MAP and in fully understanding the mechanism by which MUTYH repairs DNA in which adenine is mispaired with 8-hydroxyguanine. © 2010 Wiley-Liss, Inc.
Collapse
Affiliation(s)
- Masanori Goto
- First Department of Pathology, Hamamatsu University School of Medicine, Japan
| | | | | | | | | | | | | |
Collapse
|
|
28
|
Out AA, Tops CM, Nielsen M, Weiss MM, van Minderhout IJ, Fokkema IF, Buisine MP, Claes K, Colas C, Fodde R, Fostira F, Franken PF, Gaustadnes M, Heinimann K, Hodgson SV, Hogervorst FB, Holinski-Feder E, Lagerstedt-Robinson K, Olschwang S, Ans M.W. VDO, Redeker EJ, Scott RJ, Vankeirsbilck B, Grønlund RV, Wijnen JT, Wikman FP, Aretz S, Sampson JR, Devilee P, den Dunnen JT, Hes FJ. Leiden open variation database of the MUTYH gene. Hum Mutat 2010; 31:1205-15. [DOI: 10.1002/humu.21343] [Citation(s) in RCA: 60] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
|
|
29
|
Luncsford PJ, Chang DY, Shi G, Bernstein J, Madabushi A, Patterson DN, Lu AL, Toth EA. A structural hinge in eukaryotic MutY homologues mediates catalytic activity and Rad9-Rad1-Hus1 checkpoint complex interactions. J Mol Biol 2010; 403:351-70. [PMID: 20816984 DOI: 10.1016/j.jmb.2010.08.045] [Citation(s) in RCA: 42] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/15/2010] [Revised: 08/25/2010] [Accepted: 08/25/2010] [Indexed: 02/07/2023]
Abstract
The DNA glycosylase MutY homologue (MYH or MUTYH) removes adenines misincorporated opposite 8-oxoguanine as part of the base excision repair pathway. Importantly, defects in human MYH (hMYH) activity cause the inherited colorectal cancer syndrome MYH-associated polyposis. A key feature of MYH activity is its coordination with cell cycle checkpoint via interaction with the Rad9-Rad1-Hus1 (9-1-1) complex. The 9-1-1 complex facilitates cell cycle checkpoint activity and coordinates this activity with ongoing DNA repair. The interdomain connector (IDC, residues 295-350) between the catalytic domain and the 8-oxoguanine recognition domain of hMYH is a critical element that maintains interactions with the 9-1-1 complex. We report the first crystal structure of a eukaryotic MutY protein, a fragment of hMYH (residues 65-350) that consists of the catalytic domain and the IDC. Our structure reveals that the IDC adopts a stabilized conformation projecting away from the catalytic domain to form a docking scaffold for 9-1-1. We further examined the role of the IDC using Schizosaccharomyces pombe MYH as model system. In vitro studies of S. pombe MYH identified residues I261 and E262 of the IDC (equivalent to V315 and E316 of the hMYH IDC) as critical for maintaining the MYH/9-1-1 interaction. We determined that the eukaryotic IDC is also required for DNA damage selection and robust enzymatic activity. Our studies also provide the first evidence that disruption of the MYH/9-1-1 interaction diminishes the repair of oxidative DNA damage in vivo. Thus, preserving the MYH/9-1-1 interaction contributes significantly to minimizing the mutagenic potential of oxidative DNA damage.
Collapse
Affiliation(s)
- Paz J Luncsford
- Department of Biochemistry and Molecular Biology, University of Maryland School of Medicine, 108 North Greene Street, Baltimore, MD 21201, USA
| | | | | | | | | | | | | | | |
Collapse
|
|
30
|
Kundu S, Brinkmeyer MK, Eigenheer RA, David SS. Ser 524 is a phosphorylation site in MUTYH and Ser 524 mutations alter 8-oxoguanine (OG): a mismatch recognition. DNA Repair (Amst) 2010; 9:1026-37. [PMID: 20724227 DOI: 10.1016/j.dnarep.2010.07.002] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/09/2010] [Revised: 06/22/2010] [Accepted: 07/02/2010] [Indexed: 12/17/2022]
Abstract
MUTYH-associated polyposis (MAP) is a colorectal cancer predisposition syndrome that is caused by inherited biallelic mutations in the base excision repair (BER) gene, MUTYH. MUTYH is a DNA glycosylase that removes adenine (A) misinserted opposite 8-oxo-7,8-dihydro-2'-deoxyguanosine (OG). In this work, wild type (WT) MUTYH overexpressed using a baculovirus-driven insect cell expression system (BEVS) provided significantly higher levels of enzyme compared to bacterial overexpression. The isolated MUTYH enzyme was analyzed for potential post-translational modifications using mass spectrometry. An in vivo phosphorylation site was validated at Serine 524, which is located in the C-terminal OG recognition domain within the proliferating cell nuclear antigen (PCNA) binding region. Characterization of the phosphomimetic (S524D) and phosphoablating (S524A) mutants together with the observation that Ser 524 can be phosphorylated suggest that this residue may play an important regulatory role in vivo by altering stability and OG:A mismatch affinity.
Collapse
Affiliation(s)
- Sucharita Kundu
- Department of Chemistry, University of California, Davis, Davis, CA 95616, USA
| | | | | | | |
Collapse
|
|
31
|
Nielsen M, Morreau H, Vasen HFA, Hes FJ. MUTYH-associated polyposis (MAP). Crit Rev Oncol Hematol 2010; 79:1-16. [PMID: 20663686 DOI: 10.1016/j.critrevonc.2010.05.011] [Citation(s) in RCA: 111] [Impact Index Per Article: 7.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/16/2010] [Revised: 05/11/2010] [Accepted: 05/27/2010] [Indexed: 12/13/2022] Open
Abstract
The human mutY homologue (MUTYH) gene is responsible for inheritable polyposis and colorectal cancer. This review discusses the molecular genetic aspects of the MUTYH gene and protein, the clinical impact of mono- and biallelic MUTYH mutations and histological aspects of the MUTYH tumors. Furthermore, the relationship between MUTYH and the mismatch repair genes in colorectal cancer (CRC) families is examined. Finally, the role of other base excision repair genes in polyposis and CRC patients is discussed.
Collapse
Affiliation(s)
- Maartje Nielsen
- Department Clinical Genetics, Leiden University Medical Centre, Albinusdreef, Leiden, The Netherlands.
| | | | | | | |
Collapse
|
|
32
|
Functional analysis of MUTYH mutated proteins associated with familial adenomatous polyposis. DNA Repair (Amst) 2010; 9:700-7. [PMID: 20418187 DOI: 10.1016/j.dnarep.2010.03.008] [Citation(s) in RCA: 36] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2009] [Revised: 03/01/2010] [Accepted: 03/26/2010] [Indexed: 02/08/2023]
Abstract
The MUTYH DNA glycosylase specifically removes adenine misincorporated by replicative polymerases opposite the oxidized purine 8-oxo-7,8-dihydroguanine (8-oxoG). A defective protein activity results in the accumulation of G>T transversions because of unrepaired 8-oxoG:A mismatches. In humans, MUTYH germline mutations are associated with a recessive form of familial adenomatous polyposis and colorectal cancer predisposition (MUTYH-associated polyposis, MAP). Here we studied the repair capacity of the MUTYH variants R171W, E466del, 137insIW, Y165C and G382D, identified in MAP patients. Following expression and purification of human proteins from a bacterial system, we investigated MUTYH incision capacity on an 8-oxoG:A substrate by standard glycosylase assays. For the first time, we employed the surface plasmon resonance (SPR) technology for real-time recording of the association/dissociation of wild-type and MUTYH variants from an 8-oxoG:A DNA substrate. When compared to the wild-type protein, R171W, E466del and Y165C variants showed a severe reduction in the binding affinity towards the substrate, while 137insIW and G382D mutants manifested only a slight decrease mainly due to a slower rate of association. This reduced binding was always associated with impairment of glycosylase activity, with adenine removal being totally abrogated in R171W, E466del and Y165C and only partially reduced in 137insIW and G382D. Our findings demonstrate that SPR analysis is suitable to identify defective enzymatic behaviour even when mutant proteins display minor alterations in substrate recognition.
Collapse
|
|
33
|
Molatore S, Russo MT, D'Agostino VG, Barone F, Matsumoto Y, Albertini AM, Minoprio A, Degan P, Mazzei F, Bignami M, Ranzani GN. MUTYH mutations associated with familial adenomatous polyposis: functional characterization by a mammalian cell-based assay. Hum Mutat 2010; 31:159-66. [PMID: 19953527 DOI: 10.1002/humu.21158] [Citation(s) in RCA: 36] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/09/2023]
Abstract
MUTYH-associated polyposis (MAP) is a colorectal cancer syndrome, due to biallelic mutations of MUTYH. This Base Excision Repair gene encodes for a DNA glycosylase that specifically mitigates the high mutagenic potential of the 8-hydroxyguanine (8-oxodG) along the DNA. Aim of this study was to characterize the biological effects, in a mammalian cell background, of human MUTYH mutations identified in MAP patients (137insIW [c.411_416dupATGGAT; p.137insIleTrp]; R171W [c.511C>T; p.Arg171Trp]; E466del [c.1395_1397delGGA; p.Glu466del]; Y165C [c.494A>G; p.Tyr165Cys]; and G382D [c.1145G>A; p.Gly382Asp]). We set up a novel assay in which the human proteins were expressed in Mutyh(-/-) mouse defective cells. Several parameters, including accumulation of 8-oxodG in the genome and hypersensitivity to oxidative stress, were then used to evaluate the consequences of MUTYH expression. Human proteins were also obtained from Escherichia coli and their glycosylase activity was tested in vitro. The cell-based analysis demonstrated that all MUTYH variants we investigated were dysfunctional in Base Excision Repair. In vitro data complemented the in vivo observations, with the exception of the G382D mutant, which showed a glycosylase activity very similar to the wild-type protein. Our cell-based assay can provide useful information on the significance of MUTYH variants, improving molecular diagnosis and genetic counseling in families with mutations of uncertain pathogenicity.
Collapse
Affiliation(s)
- Sara Molatore
- Department of Genetics and Microbiology, University of Pavia, Pavia, Italy
| | | | | | | | | | | | | | | | | | | | | |
Collapse
|
|
34
|
Kundu S, Brinkmeyer MK, Livingston AL, David SS. Adenine removal activity and bacterial complementation with the human MutY homologue (MUTYH) and Y165C, G382D, P391L and Q324R variants associated with colorectal cancer. DNA Repair (Amst) 2010; 8:1400-10. [PMID: 19836313 DOI: 10.1016/j.dnarep.2009.09.009] [Citation(s) in RCA: 47] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2008] [Revised: 09/17/2009] [Accepted: 09/19/2009] [Indexed: 10/20/2022]
Abstract
MUTYH-associated polyposis (MAP) is the only inherited colorectal cancer syndrome that is associated with inherited biallelic mutations in a base excision repair gene. The MUTYH glycosylase plays an important role in preventing mutations associated with 8-oxoguanine (OG) by removing adenine residues that have been misincorporated opposite OG. MAP-associated mutations are present throughout MUTYH, with a large number coding for missense variations. To date the available information on the functional properties of MUTYH variants is conflicting. In this study, a kinetic analysis of the adenine glycosylase activity of MUTYH and several variants was undertaken using a correction for active fraction to control for differences due to overexpression and purification. Using these methods, the rate constants for steps involved in the adenine removal process were determined for the MAP variants Y165C, G382D, P391L and Q324R MUTYH. Under single-turnover conditions, the rate of adenine removal for these four variants was found to be 30-40% of WT MUTYH. In addition, the ability of MUTYH and the variants to suppress mutations and complement for the absence of MutY in Escherichia coli was assessed using rifampicin resistance assays. The presence of WT and Q324R MUTYH resulted in complete suppression of the mutation frequency, while G382D MUTYH showed reduced ability to suppress the mutation frequency. In contrast, the mutation frequency observed upon expression of P391L and Y165C MUTYH were similar to the controls, suggesting no activity toward preventing DNA mutations. Notably, though all variations studied herein resulted in similar reductions in adenine glycosylase activity, the effects in the bacterial complementation are quite different. This suggests that the consequences of a specific amino acid variation on overall repair in a cellular context may be magnified.
Collapse
Affiliation(s)
- Sucharita Kundu
- Department of Chemistry, University of California Davis, One Shields Avenue, Davis, CA 95616, USA
| | | | | | | |
Collapse
|
|
35
|
Gómez-Fernández N, Castellví-Bel S, Fernández-Rozadilla C, Balaguer F, Muñoz J, Madrigal I, Milà M, Graña B, Vega A, Castells A, Carracedo A, Ruiz-Ponte C. Molecular analysis of the APC and MUTYH genes in Galician and Catalonian FAP families: a different spectrum of mutations? BMC MEDICAL GENETICS 2009; 10:57. [PMID: 19531215 PMCID: PMC2702373 DOI: 10.1186/1471-2350-10-57] [Citation(s) in RCA: 38] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 12/11/2008] [Accepted: 06/16/2009] [Indexed: 12/28/2022]
Abstract
BACKGROUND Familial adenomatous polyposis (FAP) is an autosomal dominant-inherited colorectal cancer syndrome, caused by germline mutations in the APC gene. Recently, biallelic mutations in MUTYH have also been identified in patients with multiple colorectal adenomas and in APC-negative patients with FAP. The aim of this work is therefore to determine the frequency of APC and MUTYH mutations among FAP families from two Spanish populations. METHODS Eighty-two unrelated patients with classical or attenuated FAP were screened for APC germline mutations. MUTYH analysis was then conducted in those APC-negative families and in 9 additional patients from a previous study. Direct sequencing, SSCP analysis and TaqMan genotyping were used to identify point and frameshift mutations, meanwhile large rearrangements in the APC gene were screened by multiplex ligation-dependent probe amplification (MLPA). RESULTS APC germline mutations were found in 39% of the patients and, despite the great number of genetic variants described so far in this gene, seven new mutations were identified. The two hotspots at codons 1061 and 1309 of the APC gene accounted for 9,4% of the APC-positive families, although they were underrepresented in Galician samples. The deletion at codon 1061 was not found in 19 APC-positive Galician patients but represented 23% of the Catalonian positive families (p = 0,058). The same trend was observed at codon 1309, even though statistical analysis showed no significance between populations. Twenty-four percent of the APC-negative patients carried biallelic MUTYH germline mutations, and showed an attenuated polyposis phenotype generally without extracolonic manifestations. New genetic variants were found, as well as the two hotspots already reported (p.Tyr165Cys and p.Gly382Asp). CONCLUSION The results we present indicate that in Galician patients the frequency of the hotspot at codon 1061 in APC differs significantly from the Catalonian and also other Caucasian populations. Similar results had already been obtained in a previous study and could be due to the genetic isolation of the Galician population. MUTYH analysis is also recommended for all APC-negative families, even if a recessive inheritance is not confirmed.
Collapse
Affiliation(s)
- Nuria Gómez-Fernández
- Fundación Pública Galega de Medicina Xenómica, Grupo de Medicina Xenómica-USC, CIBERER, Santiago de Compostela, Galicia, Spain.
| | | | | | | | | | | | | | | | | | | | | | | |
Collapse
|
|
36
|
Sidorenko VS, Zharkov DO. Role of base excision repair DNA glycosylases in hereditary and infectious human diseases. Mol Biol 2008. [DOI: 10.1134/s0026893308050166] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/23/2022]
|
|
37
|
Lin JC, Singh RRP, Cox DL. Theoretical study of DNA damage recognition via electron transfer from the [4Fe-4S] complex of MutY. Biophys J 2008; 95:3259-68. [PMID: 18599627 PMCID: PMC2547449 DOI: 10.1529/biophysj.108.132183] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/26/2008] [Accepted: 06/11/2008] [Indexed: 11/18/2022] Open
Abstract
The mechanism of site-specific recognition of DNA by proteins has been a long-standing issue. The DNA glycosylase MutY, for instance, must find the rare 8-oxoguanine-adenine mismatches among the large number of basepairs in the DNA. This protein has a [4Fe-4S] cluster, which is highly conserved in species as diverse as Escherichia Coli and Homo sapiens. The mixed-valent nature of this cluster suggests that charge transfer may play a role in MutY's function. We have studied the energetics of the charge transfer in Bacillus stearothermophilus MutY-DNA complex using multiscale calculation including density functional theory and molecular dynamics. The [4Fe-4S] cluster in MutY is found to undergo 2+ to 3+ oxidation when coupling to DNA through hole transfer, especially when MutY is near an oxoguanine modified base (oxoG). Employing the Marcus theory for electron transfer, we find near optimal Frank-Condon factors for electron transfer from MutY to oxoguanine modified base. MutY has modest selectivity for oxoguanine over guanine due to the difference in oxidation potential. The tunneling matrix element is significantly reduced with the mutation R149W, whereas the mutation L154F reduces the tunneling matrix element as well as the Frank-Condon factor. Both L154F and R149W mutations are known to dramatically reduce or eliminate repair efficiency. We suggest a scenario where the charge transfer leads to a stabilization of the specific binding conformation, which is likely the recognition mode, thus enabling it to find the damaged site efficiently.
Collapse
Affiliation(s)
- Jong-Chin Lin
- Department of Physics, University of California, Davis, California 95616, USA
| | | | | |
Collapse
|
|
38
|
Ali M, Kim H, Cleary S, Cupples C, Gallinger S, Bristow R. Characterization of mutant MUTYH proteins associated with familial colorectal cancer. Gastroenterology 2008; 135:499-507. [PMID: 18534194 PMCID: PMC2761659 DOI: 10.1053/j.gastro.2008.04.035] [Citation(s) in RCA: 80] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/14/2008] [Revised: 04/07/2008] [Accepted: 04/30/2008] [Indexed: 01/04/2023]
Abstract
BACKGROUND & AIMS The human mutyh gene encodes a base excision repair protein that prevents G:C to T:A transversions in DNA. Biallelic mutations in this gene are associated with recessively inherited familial colorectal cancer. The aim of this study was to characterize the functional activity of mutant-MUTYH and single-nucleotide polymorphism (SNP)-MUTYH proteins involving familial colorectal cancer. METHODS MUTYH variants were cloned and assayed for their glycosylase and DNA binding activities using synthetic double-stranded oligonucleotide substrates by analyzing cleavage products by polyacrylamide gel electrophoresis. RESULTS In this study, we have characterized 9 missense/frameshift mutants and 2 SNPs for their DNA binding and repair activity in vitro. Two missense mutants (R260Q and G382D) were found to be partially active in both glycosylase and DNA binding, whereas 3 other missense mutants (Y165C, R231H, and P281L) were severely defective in both activities. All of the frameshift mutants (Y90X, Q377X, E466X, and 1103delC) were completely devoid of both glycosylase and DNA binding activities. One SNP (V22M) showed the same activity as wild-type MUTYH protein, but the other SNP (Q324H) was partially impaired in adenine removal. CONCLUSIONS This study of MUTYH mutants suggests that certain SNPs may be as partially dysfunctional in base excision repair as missense-MUTYH mutants and lead to colorectal carcinogenesis.
Collapse
Affiliation(s)
- Mohsin Ali
- Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, ON, Canada, Applied Molecular Oncology, Ontario Cancer Institute/Princess Margaret Hospital (University Health Network) and Departments of Radiation Oncology and Medical Biophysics, University of Toronto, Toronto, ON, Canada
| | - Hyeja Kim
- Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, ON, Canada
| | - Sean Cleary
- Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, ON, Canada
| | - Claire Cupples
- Dept. of Biochemistry and Microbiology, University of Victoria, Victoria, BC, Canada
| | - Steven Gallinger
- Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, ON, Canada,Corresponding author: Robert Bristow MD PhD FRCPC, Radiation Medicine Program, Princess Margaret Hospital (UHN), 610 University Avenue, Toronto, Ontario, CANADA M5G2M9, Tel: 416-946-2223; Fax: 416-946-4586;
| | - Robert Bristow
- Applied Molecular Oncology, Ontario Cancer Institute/Princess Margaret Hospital (University Health Network) and Departments of Radiation Oncology and Medical Biophysics, University of Toronto, Toronto, ON, Canada,Corresponding author: Robert Bristow MD PhD FRCPC, Radiation Medicine Program, Princess Margaret Hospital (UHN), 610 University Avenue, Toronto, Ontario, CANADA M5G2M9, Tel: 416-946-2223; Fax: 416-946-4586;
| |
Collapse
|
|
39
|
Slean MM, Panigrahi GB, Ranum LP, Pearson CE. Mutagenic roles of DNA "repair" proteins in antibody diversity and disease-associated trinucleotide repeat instability. DNA Repair (Amst) 2008; 7:1135-54. [PMID: 18485833 DOI: 10.1016/j.dnarep.2008.03.014] [Citation(s) in RCA: 37] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/25/2022]
Abstract
While DNA repair proteins are generally thought to maintain the integrity of the whole genome by correctly repairing mutagenic DNA intermediates, there are cases where DNA "repair" proteins are involved in causing mutations instead. For instance, somatic hypermutation (SHM) and class switch recombination (CSR) require the contribution of various DNA repair proteins, including UNG, MSH2 and MSH6 to mutate certain regions of immunoglobulin genes in order to generate antibodies of increased antigen affinity and altered effector functions. Another instance where "repair" proteins drive mutations is the instability of gene-specific trinucleotide repeats (TNR), the causative mutations of numerous diseases including Fragile X mental retardation syndrome (FRAXA), Huntington's disease (HD), myotonic dystrophy (DM1) and several spinocerebellar ataxias (SCAs) all of which arise via various modes of pathogenesis. These healthy and deleterious mutations that are induced by repair proteins are distinct from the genome-wide mutations that arise in the absence of repair proteins: they occur at specific loci, are sensitive to cis-elements (sequence context and/or epigenetic marks) and transcription, occur in specific tissues during distinct developmental windows, and are age-dependent. Here we review and compare the mutagenic role of DNA "repair" proteins in the processes of SHM, CSR and TNR instability.
Collapse
Affiliation(s)
- Meghan M Slean
- Program of Genetics & Genome Biology, The Hospital for Sick Children, Toronto, Ontario, Canada M5G 1L7
| | | | | | | |
Collapse
|
|
40
|
Küry S, Buecher B, Robiou-Du-Pont S, Scoul C, Colman H, Lelièvre B, Olschwang S, Houérou CL, Neel TL, Faroux R, Ollivry J, Lafraise B, Chupin LD, Bézieau S. The Thorough Screening of the MUTYH Gene in a Large French Cohort of Sporadic Colorectal Cancers. ACTA ACUST UNITED AC 2007; 11:373-9. [DOI: 10.1089/gte.2007.0029] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/22/2023]
Affiliation(s)
- Sébastien Küry
- Laboratoire d'Etude de l'ADN, Faculté de Médecine de Nantes, 44035 France
- Institut des Maladies de l'Appareil Digestif et CIC INSERM, Centre Hospitalier Universitaire de Nantes 44093, France
| | - Bruno Buecher
- Institut des Maladies de l'Appareil Digestif et CIC INSERM, Centre Hospitalier Universitaire de Nantes 44093, France
- Service d'Hépato-Gastroentérologie, Centre Hospitalier Universitaire de Nantes 44093, France
| | - Sébastien Robiou-Du-Pont
- Laboratoire d'Etude de l'ADN, Faculté de Médecine de Nantes, 44035 France
- Institut des Maladies de l'Appareil Digestif et CIC INSERM, Centre Hospitalier Universitaire de Nantes 44093, France
| | - Catherine Scoul
- Laboratoire d'Etude de l'ADN, Faculté de Médecine de Nantes, 44035 France
- Institut des Maladies de l'Appareil Digestif et CIC INSERM, Centre Hospitalier Universitaire de Nantes 44093, France
| | - Hélène Colman
- Laboratoire d'Etude de l'ADN, Faculté de Médecine de Nantes, 44035 France
- Institut des Maladies de l'Appareil Digestif et CIC INSERM, Centre Hospitalier Universitaire de Nantes 44093, France
| | - Bénédicte Lelièvre
- Laboratoire d'Etude de l'ADN, Faculté de Médecine de Nantes, 44035 France
- Institut des Maladies de l'Appareil Digestif et CIC INSERM, Centre Hospitalier Universitaire de Nantes 44093, France
| | - Sylviane Olschwang
- Département d'Oncologie Génétique, Prévention et Dépistage, Institut Paoli-Calmettes, Marseille 13273, France
| | - Claire Le Houérou
- Institut des Maladies de l'Appareil Digestif et CIC INSERM, Centre Hospitalier Universitaire de Nantes 44093, France
| | - Tanguy Le Neel
- Laboratoire d'Etude de l'ADN, Faculté de Médecine de Nantes, 44035 France
- Biofortis, Nantes 44200, France
| | - Roger Faroux
- Service d'Hépato-Gastroentérologie, Centre Hospitalier Universitaire de la Roche-Sur-Yon, 85000 France
| | - Jean Ollivry
- Association des Gastroentérologues de Vendée, 85300 Challans, France
| | | | | | - Stéphane Bézieau
- Laboratoire d'Etude de l'ADN, Faculté de Médecine de Nantes, 44035 France
- Institut des Maladies de l'Appareil Digestif et CIC INSERM, Centre Hospitalier Universitaire de Nantes 44093, France
| |
Collapse
|
|
41
|
Unnatural substrates reveal the importance of 8-oxoguanine for in vivo mismatch repair by MutY. Nat Chem Biol 2007; 4:51-8. [PMID: 18026095 DOI: 10.1038/nchembio.2007.40] [Citation(s) in RCA: 33] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/11/2007] [Accepted: 09/08/2007] [Indexed: 11/09/2022]
Abstract
Escherichia coli MutY has an important role in preventing mutations associated with the oxidative lesion 7,8-dihydro-8-oxo-2'-deoxyguanosine (OG) in DNA by excising adenines from OG.A mismatches as the first step of base excision repair. To determine the importance of specific steps in the base pair recognition and base removal process of MutY, we have evaluated the effects of modifications of the OG.A substrate on the kinetics of base removal, mismatch affinity and repair to G-C in an E. coli-based assay. Notably, adenine modification was tolerated in the cellular assay, whereas modification of OG resulted in minimal cellular repair. High affinity for the mismatch and efficient base removal required the presence of OG. Taken together, these results suggest that the presence of OG is a critical feature that is necessary for MutY to locate OG.A mismatches and select the appropriate adenines for excision to initiate repair in vivo before replication.
Collapse
|
|
42
|
Abstract
Maintaining the chemical integrity of DNA in the face of assault by oxidizing agents is a constant challenge for living organisms. Base-excision repair has an important role in preventing mutations associated with a common product of oxidative damage to DNA, 8-oxoguanine. Recent structural studies have shown that 8-oxoguanine DNA glycosylases use an intricate series of steps to locate and excise 8-oxoguanine lesions efficiently against a high background of undamaged bases. The importance of preventing mutations associated with 8-oxoguanine is shown by a direct association between defects in the DNA glycosylase MUTYH and colorectal cancer. The properties of other guanine oxidation products and the associated DNA glycosylases that remove them are now also being revealed.
Collapse
Affiliation(s)
- Sheila S David
- Department of Chemistry, University of California at Davis, 1 Shields Avenue, Davis, California 95616, USA.
| | | | | |
Collapse
|
|
43
|
Abstract
The base excision repair carried out by the bacterial MutY DNA glycosylase and eukaryotic MutY homolog (MYH) is responsible for removing adenines misincorporated into DNA opposite 7,8-dihydro-8-oxo-guanines (8-oxoG), thereby preventing G:C to T:A mutations. MutY and MYH can also remove adenines from A/G and A/C and can remove guanines from G/8-oxoG mismatches at reduced rates. Biallelic germline mutations in the human MYH gene predispose individuals to multiple colorectal adenomas and carcinoma. Four functional assays are usually employed to characterize the MutY and MYH. Gel mobility shift or fluorescence anisotropy assays measures DNA-binding affinity and the apparent dissociation constants. Glycosylase assay determines the catalytic parameters of the enzyme. By using a trapping assay in the presence of sodium borohydride, the protein-DNA covalent intermediate can be identified. The in vivo activity of MutY or MYH can be measured by complementation in Escherichia coli mutY mutants or fission yeast Schizosaccharomyces pombe MYH knockout cells. MutY and MYH interacting proteins can be analyzed by the glutathione S-transferase pull-down assay, Far-western, and coimmunoprecipitation. The in vitro and in vivo activities of MYH can be modulated by several proteins, including mismatch recognition enzymes MSH2/MSH6, proliferating cell nuclear antigen, and apurinic/apyrimidinic endonuclease.
Collapse
Affiliation(s)
- A-Lien Lu-Chang
- University of Maryland, Department of Biochemistry, Baltimore, USA
| |
Collapse
|
|
44
|
Russo MT, De Luca G, Degan P, Bignami M. Different DNA repair strategies to combat the threat from 8-oxoguanine. Mutat Res 2007; 614:69-76. [PMID: 16769088 DOI: 10.1016/j.mrfmmm.2006.03.007] [Citation(s) in RCA: 73] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2005] [Revised: 03/08/2006] [Accepted: 03/10/2006] [Indexed: 05/10/2023]
Abstract
Oxidative DNA damage is one of the most common threats to genome stability and DNA repair enzymes provide protection from the effects of oxidized DNA bases. In mammalian cells, base excision repair (BER) mediated by the OGG1 and MYH DNA glycosylases prevents the accumulation of 8-oxoguanine (8-oxoG) in DNA. When steady-state levels of DNA 8-oxoG were measured in myh(-/-) and myh(-/-)/ogg1(-/-) mice, an age-dependent accumulation of the oxidized purine was found in lung and small intestine of doubly defective myh(-/-)/ogg1(-/-) mice. Since there is an increased incidence of lung and small intestinal cancer in myh(-/-)/ogg1(-/-) mice, these findings are consistent with a causal role for unrepaired oxidized DNA bases in cancer development. We previously presented in vitro evidence that mismatch repair (MMR) participates in the repair of oxidative DNA damage and msh2(-/-) mouse embryo fibroblasts also have increased steady state levels of DNA 8-oxoG. To investigate whether DNA 8-oxoG also accumulates in vivo, basal levels were measured in several organs of 4-month-old msh2(-/-) mice and their wild-type counterparts. Msh2(-/-) mice had significantly increased levels of DNA 8-oxoG in spleen, heart, liver, lung, kidney and possibly small intestine but not in bone marrow, thymus or brain. The tissue-specificity of DNA 8-oxoG accumulation in msh2(-/-) and other DNA repair defective mice suggests that DNA protection of different organs is mediated by different combinations of repair pathways.
Collapse
Affiliation(s)
- Maria Teresa Russo
- Department of Environment and Primary Prevention, Istituto Superiore di Sanità, Viale Regina Elena 299, 00161 Rome, Italy
| | | | | | | |
Collapse
|
|
45
|
Shi G, Chang DY, Cheng CC, Guan X, Venclovas Č, Lu AL. Physical and functional interactions between MutY glycosylase homologue (MYH) and checkpoint proteins Rad9-Rad1-Hus1. Biochem J 2006; 400:53-62. [PMID: 16879101 PMCID: PMC1635441 DOI: 10.1042/bj20060774] [Citation(s) in RCA: 73] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/25/2006] [Revised: 07/24/2006] [Accepted: 08/01/2006] [Indexed: 01/02/2023]
Abstract
The MYH (MutY glycosylase homologue) increases replication fidelity by removing adenines or 2-hydroxyadenine misincorporated opposite GO (7,8-dihydro-8-oxo-guanine). The 9-1-1 complex (Rad9, Rad1 and Hus1 heterotrimer complex) has been suggested as a DNA damage sensor. Here, we report that hMYH (human MYH) interacts with hHus1 (human Hus1) and hRad1 (human Rad1), but not with hRad9. In addition, interactions between MYH and the 9-1-1 complex, from both the fission yeast Schizosaccharomyces pombe and human cells, are partially interchangeable. The major Hus1-binding site is localized to residues 295-350 of hMYH and to residues 245-293 of SpMYH (S. pombe MYH). Val315 of hMYH and Ile261 of SpMYH play important roles for their interactions with Hus1. hHus1 protein and the 9-1-1 complex of S. pombe can enhance the glycosylase activity of SpMYH. Moreover, the interaction of hMYH-hHus1 is enhanced following ionizing radiation. A significant fraction of the hMYH nuclear foci co-localizes with hRad9 foci in H2O2-treated cells. These results reveal that the 9-1-1 complex plays a direct role in base excision repair.
Collapse
Key Words
- base excision repair
- dna damage checkpoint
- dna glycosylase
- fission yeast
- hus1
- muty glycosylase homologue (myh)
- the 9-1-1 complex, rad9, rad1 and hus1 heterotrimer complex
- ap, apurinic/apyrimidinic
- ape1, ap endonuclease 1
- atm, ataxia telangiectasia mutated
- atr, atm- and rad3-related protein
- atrip, atr-interacting protein
- brca1, breast-cancer susceptibility gene 1
- dapi, 4′,6′-diamidino-2-phenylindole
- fen1, flap endonuclease 1
- go, 7,8-dihydro-8-oxo-guanine
- gst, glutathione s-transferase
- ha, haemagglutinin
- hhus1, human hus1
- hrad1, human rad1
- msh2, muts homologue 2
- mlh1, mutl homologue 1
- myh, muty glycosylase homologue
- hmyh, human myh
- mmyh, mouse myh
- pcna, proliferating-cell nuclear antigen
- hpcna, human pcna
- rfc, replication factor c
- rmyh, rat myh
- rpa, replication protein a
- sphus1, s. pombe hus1
- spmyh, s. pombe myh
- xpa, xeroderma pigmentosum group a
- xpf, xeroderma pigmentosum group f
Collapse
Affiliation(s)
- Guoli Shi
- *Department of Biochemistry and Molecular Biology and Greenebaum Cancer Center, School of Medicine, University of Maryland, Baltimore, MD 21201, U.S.A
| | - Dau-Yin Chang
- *Department of Biochemistry and Molecular Biology and Greenebaum Cancer Center, School of Medicine, University of Maryland, Baltimore, MD 21201, U.S.A
| | - Chih-Chien Cheng
- *Department of Biochemistry and Molecular Biology and Greenebaum Cancer Center, School of Medicine, University of Maryland, Baltimore, MD 21201, U.S.A
| | - Xin Guan
- *Department of Biochemistry and Molecular Biology and Greenebaum Cancer Center, School of Medicine, University of Maryland, Baltimore, MD 21201, U.S.A
| | | | - A-Lien Lu
- *Department of Biochemistry and Molecular Biology and Greenebaum Cancer Center, School of Medicine, University of Maryland, Baltimore, MD 21201, U.S.A
| |
Collapse
|
|
46
|
Bai H, Grist S, Gardner J, Suthers G, Wilson TM, Lu AL. Functional characterization of human MutY homolog (hMYH) missense mutation (R231L) that is linked with hMYH-associated polyposis. Cancer Lett 2006; 250:74-81. [PMID: 17081686 PMCID: PMC1907362 DOI: 10.1016/j.canlet.2006.09.016] [Citation(s) in RCA: 33] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/15/2006] [Accepted: 09/22/2006] [Indexed: 12/23/2022]
Abstract
The MutY homolog (MYH) can excise adenines misincorporated opposite to guanines or 7,8-dihydro-8-oxo-guanines (8-oxoG) during DNA replication; thereby preventing G:C to T:A transversions. Germline mutations in the human MYH gene are associated with recessive inheritance of colorectal adenomatous polyposis (MAP). Here, we characterize one newly identified MAP-associated MYH missense mutation (R231L) that lies adjacent to the putative hMSH6 binding domain. The R231L mutant protein has severe defects in A/GO binding and in adenine glycosylase activities. The mutant fails to complement mutY-deficiency in Escherichia coli, but does not affect binding to hMSH6. These data support the role of the hMYH pathway in carcinogenesis.
Collapse
Affiliation(s)
- Haibo Bai
- Department of Biochemistry and Molecular Biology, University of Maryland, Baltimore, MD 21201, USA
| | - Scott Grist
- Department of Haematology and Genetic Pathology, Flinders Medical Centre and Flinders University, Adelaide, South Australia, 5042
| | - Justin Gardner
- Department of Haematology and Genetic Pathology, Flinders Medical Centre and Flinders University, Adelaide, South Australia, 5042
| | - Graeme Suthers
- South Australian Familial Cancer Service, Women’s and Children’s Hospital, Adelaide, South Australia, 5042
| | - Teresa M. Wilson
- Department of Radiation Oncology, University of Maryland, Baltimore, MD 21201, USA
- The University of Maryland Greenebaum Cancer Center, Baltimore, MD 21201, USA
| | - A-Lien Lu
- Department of Biochemistry and Molecular Biology, University of Maryland, Baltimore, MD 21201, USA
- The University of Maryland Greenebaum Cancer Center, Baltimore, MD 21201, USA
- * Corresponding author. Tel.: +1-410-706-4356; Fax: 410-706-1787. E-mail address: (A-L. Lu)
| |
Collapse
|
|
47
|
Sokhansanj BA, Wilson DM. Estimating the effect of human base excision repair protein variants on the repair of oxidative DNA base damage. Cancer Epidemiol Biomarkers Prev 2006; 15:1000-8. [PMID: 16702383 DOI: 10.1158/1055-9965.epi-05-0817] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022] Open
Abstract
Epidemiologic studies have revealed a complex association between human genetic variance and cancer risk. Quantitative biological modeling based on experimental data can play a critical role in interpreting the effect of genetic variation on biochemical pathways relevant to cancer development and progression. Defects in human DNA base excision repair (BER) proteins can reduce cellular tolerance to oxidative DNA base damage caused by endogenous and exogenous sources, such as exposure to toxins and ionizing radiation. If not repaired, DNA base damage leads to cell dysfunction and mutagenesis, consequently leading to cancer, disease, and aging. Population screens have identified numerous single-nucleotide polymorphism variants in many BER proteins and some have been purified and found to exhibit mild kinetic defects. Epidemiologic studies have led to conflicting conclusions on the association between single-nucleotide polymorphism variants in BER proteins and cancer risk. Using experimental data for cellular concentration and the kinetics of normal and variant BER proteins, we apply a previously developed and tested human BER pathway model to (i) estimate the effect of mild variants on BER of abasic sites and 8-oxoguanine, a prominent oxidative DNA base modification, (ii) identify ranges of variation associated with substantial BER capacity loss, and (iii) reveal nonintuitive consequences of multiple simultaneous variants. Our findings support previous work suggesting that mild BER variants have a minimal effect on pathway capacity whereas more severe defects and simultaneous variation in several BER proteins can lead to inefficient repair and potentially deleterious consequences of cellular damage.
Collapse
Affiliation(s)
- Bahrad A Sokhansanj
- School of Biomedical Engineering, Science and Health Systems, Drexel University, 3141 Chestnut Street, Philadelphia, PA 19104, USA.
| | | |
Collapse
|
|
48
|
Di Gregorio C, Frattini M, Maffei S, Ponti G, Losi L, Pedroni M, Venesio T, Bertario L, Varesco L, Risio M, Ponz de Leon M. Immunohistochemical expression of MYH protein can be used to identify patients with MYH-associated polyposis. Gastroenterology 2006; 131:439-44. [PMID: 16890597 DOI: 10.1053/j.gastro.2006.05.049] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/05/2005] [Accepted: 05/11/2006] [Indexed: 12/22/2022]
Abstract
BACKGROUND & AIMS MYH-associated polyposis is a recently described, autosomal-recessive disease characterized by multiple colorectal adenomas and cancer. There are only few immunohistochemical studies of the MYH protein. We investigated the expression pattern of the MYH protein to evaluate whether a immunohistochemical approach could be used in clinical practice to screen patients for germline mutations in the MYH gene. METHODS The expression of MYH, MSH2, MLH1, and MSH6 proteins was studied by immunohistochemistry in 20 samples (colorectal adenomas or cancer) from 18 patients with biallelic MYH mutation, in 11 samples from patients with germline adenomatous polyposis coli (APC) mutations, in 20 samples from patients with sporadic colorectal cancers, and in 10 samples from patients with normal colonic mucosa without malignancies. RESULTS In all cases the mismatch repair proteins were expressed normally. Nuclear and cytoplasmic immunoreactivity for the MYH protein were observed in normal colorectal mucosa, in sporadic colorectal carcinomas, and in adenomas and carcinomas from patients carrying APC germline mutations. Adenomas and carcinomas from patients with MYH biallelic mutation showed a different pattern of expression: a strong granular cytoplasmic staining was observed without any nuclear expression. The same immunophenotype was observed in the surrounding normal mucosa. CONCLUSIONS Patients with biallelic MYH mutations showed disappearance of staining from the nucleus, and segregation of immunoreactivity in the cytoplasm, both in neoplastic and surrounding healthy mucosa. Because this pattern of expression seems to be specific for biallelic mutations, it follows that immunohistochemistry might be used in clinical practice to screen patients at risk for MYH-associated polyposis.
Collapse
|
|
49
|
Denver DR, Feinberg S, Steding C, Durbin MD, Lynch M. The relative roles of three DNA repair pathways in preventing Caenorhabditis elegans mutation accumulation. Genetics 2006; 174:57-65. [PMID: 16783005 PMCID: PMC1569771 DOI: 10.1534/genetics.106.059840] [Citation(s) in RCA: 41] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022] Open
Abstract
Mutation is a central biological process whose rates and spectra are influenced by a variety of complex and interacting forces. Although DNA repair pathways are generally known to play key roles in maintaining genetic stability, much remains to be understood about the relative roles of different pathways in preventing the accumulation of mutations and the extent of heterogeneity in pathway-specific repair efficiencies across different genomic regions. In this study we examine mutation processes in base excision repair-deficient (nth-1) and nucleotide excision repair-deficient (xpa-1) Caenorhabditis elegans mutation-accumulation (MA) lines across 24 regions of the genome and compare our observations to previous data from mismatch repair-deficient (msh-2 and msh-6) and wild-type (N2) MA lines. Drastic variation in both average and locus-specific mutation rates, ranging two orders of magnitude for the latter, was detected among the four sets of repair-deficient MA lines. Our work provides critical insights into the relative roles of three DNA repair pathways in preventing C. elegans mutation accumulation and provides evidence for the presence of pathway-specific DNA repair territories in the C. elegans genome.
Collapse
Affiliation(s)
- Dee R Denver
- Department of Biology, Indiana University, Bloomington, Indiana 47405, USA.
| | | | | | | | | |
Collapse
|
|
50
|
Stormorken A, Heintz KM, Andresen PA, Hovig E, Møller P. MUTYH Mutations Do Not Cause HNPCC or Late Onset Familial Colorectal Cancer. Hered Cancer Clin Pract 2006; 4:90-3. [PMID: 20223013 PMCID: PMC2837291 DOI: 10.1186/1897-4287-4-2-90] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/07/2006] [Accepted: 05/20/2006] [Indexed: 11/10/2022] Open
Abstract
Recently, carriers of biallelic mutations in the base excision repair gene MUTYH, have been demonstrated to have a predisposition for multiple adenomas and colorectal cancer. Still, many questions remain unanswered concerning MUTYH. We have addressed the following: Do biallelic MUTYH mutation carriers invariably demonstrate FAP, and may MUTYH be a gene causing HNPCC, HNPCC-like or dominantly inherited late onset colorectal cancer? We examined affecteds from our total series of HNPCC, HNPCC-like and dominantly inherited late onset colorectal cancer kindreds not demonstrated to have any MMR mutations. Bloodsamples from 96 patients were subjected to sequencing of exon 7 and exon 13 in the MUTYH gene. Two heterozygotes and one homozygote for the European founder mutations were found. The homozygous carrier did not meet criteria for FAP/AFAP. We conclude that MUTYH, when mutated, causes a rare recessively inherited disorder including colorectal- and duodenal cancers. It is not verified that heterozygous carriers of MUTYH mutations have an increased risk of cancer, and they do not explain the occurrence of familial colorectal cancer in the population.
Collapse
Affiliation(s)
- Astrid Stormorken
- Section for Inherited Cancer, Department for Medical Genetics, Rikshospitalet-Radiumhospitalet Medical Center, Oslo, Norway.
| | | | | | | | | |
Collapse
|