Germline mutations of EXO1 gene in patients with hereditary nonpolyposis colorectal cancer (HNPCC) and atypical HNPCC forms

doi:10.1053/gast.2001.25117

Gastroenterology

Volume 120, Issue 7, June 2001, Pages 1580-1587

https://doi.org/10.1053/gast.2001.25117 Get rights and content

Abstract

Background & Aims: Germline mutations in one of four mismatch repair genes have been found in the majority of families with hereditary nonpolyposis colorectal cancer (HNPCC), but only in a small part of families with atypical HNPCC. The recently cloned EXO1 gene might be involved in the pathogenesis of HNPCC because the EXO1 protein strongly interacts with the MSH2 protein. To determine its role in HNPCC, EXO1 was scanned for germline mutations. Methods: All 14 exons of EXO1 were scanned for mutations in index patients from 33 families with HNPCC fulfilling the Amsterdam criteria and in 225 index patients suspected of HNPCC. Results: Germline variants of EXO1 were detected in 14 patients, including one splice-site mutation in a family with HNPCC and 13 missense mutations in patients with atypical HNPCC. These variants did not occur in more than 200 control individuals. From 13 of these 14 patients, tumors were available for analysis of microsatellite instability and loss of heterozygosity. Six of the tumors showed microsatellite instability. Heterozygosity analysis showed one case without EXO1 allelic loss and 12 tumors with loss of the mutant allele and retention of the normal one. Conclusions: The results indicate a possible association of germline EXO1 variants with HNPCC and atypical HNPCC.

GASTROENTEROLOGY 2001;120:1580-1587

Section snippets

Patients

Patients were included into the study if they fulfilled at least one of the following criteria: (1) diagnosis of colorectal cancer or endometrial cancer before the age of 50 years; (2) diagnosis of any HNPCC-associated tumor (gastric, small bowel, ovarian, urothelial, pancreatic, or bile duct) and a first-degree relative with a diagnosis of colorectal cancer or endometrial cancer, or vice versa, with one of them diagnosed before the age of 50 years; or (3) diagnosis of more than one

Mutation analysis

Germline variants of EXO1 were detected in 14 patients (Table 2), including 1 patient from a family with HNPCC with a splice-site mutation and 13 index patients suspected of HNPCC, all with a missense mutation.These variants did not occur in more than 200 anonymous control individuals from the Dutch population. In 15 of the 33 unrelated families with HNPCC and in 23 of the 225 index patients suspected of HNPCC, a germline MSH2, MLH1, or MSH6 mutation had been identified previously (our

Discussion

Scanning EXO1 for germline mutations in patients with HNPCC and patients suspected of HNPCC, 9 different germline EXO1 variants (1 splice-site mutation and 8 missense mutations) were detected in 14 index patients (Table 2). These variants did not occur in more than 200 control individuals from the Dutch population. All 14 patients had been screened for germline MSH2, MLH1, or MSH6 mutations. No such mutations had been detected. The splice-site mutation found in a family with HNPCC leads to a

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Cited by (121)

DNA-damage-induced degradation of EXO1 exonuclease limits DNA end resection to ensure accurate DNA repair
2017, Journal of Biological Chemistry
Citation Excerpt :
It is, therefore, clear that EXO1 stability is probably regulated by an interplay of post-translational modifications (PTM)–phosphorylation, SUMOylation, and ubiquitination–but the exact choreography of these PTMs and the exact roles of specific modifications in EXO1 regulation still need to be elucidated. Regulatory mechanisms impinging on EXO1 are of significance from a cancer standpoint, as multiple SNPs in EXO1 have been identified in patients with colorectal cancers (89–91), and a specific polymorphism, K589E (rs1047840), has been linked with an increased risk of breast (92), gastric (93), oral (94), lung (95), and brain (96) cancer development. Another polymorphism, N279S (rs4149909), was recently linked to an increased risk of breast cancer in a genome-wide association analysis of >120,000 individuals (97).
End resection of DNA double-strand breaks (DSBs) to generate 3′-single-stranded DNA facilitates DSB repair via error-free homologous recombination (HR) while stymieing repair by the error-prone non-homologous end joining (NHEJ) pathway. Activation of DNA end resection involves phosphorylation of the 5′ to 3′ exonuclease EXO1 by the phosphoinositide 3-kinase-like kinases ATM (ataxia telangiectasia-mutated) and ATR (ATM and Rad3-related) and by the cyclin-dependent kinases 1 and 2. After activation, EXO1 must also be restrained to prevent over-resection that is known to hamper optimal HR and trigger global genomic instability. However, mechanisms by which EXO1 is restrained are still unclear. Here, we report that EXO1 is rapidly degraded by the ubiquitin-proteasome system soon after DSB induction in human cells. ATR inhibition attenuated DNA-damage-induced EXO1 degradation, indicating that ATR-mediated phosphorylation of EXO1 targets it for degradation. In accord with these results, EXO1 became resistant to degradation when its SQ motifs required for ATR-mediated phosphorylation were mutated. We show that upon the induction of DNA damage, EXO1 is ubiquitinated by a member of the Skp1-Cullin1-F-box (SCF) family of ubiquitin ligases in a phosphorylation-dependent manner. Importantly, expression of degradation-resistant EXO1 resulted in hyper-resection, which attenuated both NHEJ and HR and severely compromised DSB repair resulting in chromosomal instability. These findings indicate that the coupling of EXO1 activation with its eventual degradation is a timing mechanism that limits the extent of DNA end resection for accurate DNA repair.
Mismatch Repair and Colon Cancer: Mechanisms and Therapies Explored
2016, Trends in Molecular Medicine
Colorectal cancer (CRC) remains one of the most prevalent cancers worldwide. In sporadic CRC, mutations frequently occur in the DNA mismatch repair (MMR) pathway. In addition, germline MMR mutations have been linked to Lynch syndrome, the most common form of hereditary CRC. Although genetic mutations, diet, inflammation, and the gut microbiota can influence CRC, it is unclear how MMR deficiency relates to these factors to modulate disease. In this review, the association of MMR to the etiology of CRC is examined, particularly in the context of microRNAs (miRNAs), inflammation, and the microbiome. We also discuss the most current targeted therapies, methods of prevention, and molecular biomarkers against MMR-deficient CRC, all of which are encouraging advancements in the field.
Exo1 phosphorylation status controls the hydroxyurea sensitivity of cells lacking the Pol32 subunit of DNA polymerases delta and zeta
2014, DNA Repair
Exo1 belongs to the Rad2 family of structure-specific nucleases and possesses 5′–3′ exonuclease activity on double-stranded DNA substrates. Exo1 interacts physically with the DNA mismatch repair (MMR) proteins Msh2 and Mlh1 and is involved in the excision of the mispaired nucleotide. Independent of its role in MMR, Exo1 contributes to long-range resection of DNA double-strand break (DSB) ends to facilitate their repair by homologous recombination (HR), and was recently identified as a component of error-free DNA damage tolerance pathways. Here, we show that Exo1 activity increases the hydroxyurea sensitivity of cells lacking Pol32, a subunit of DNA polymerases δ and ζ. Both, phospho-mimicking and dephospho-mimicking exo1 mutants act as hypermorphs, as evidenced by an increase in HU sensitivity of pol32Δ cells, suggesting that they are trapped in an active form and that phosphorylation of Exo1 at residues S372, S567, S587, S692 is necessary, but insufficient, for the accurate regulation of Exo1 activity at stalled replication forks. In contrast, neither phosphorylation status is important for Exo1's role in MMR or in the suppression of genome instability in cells lacking Sgs1 helicase. This ability of an EXO1 deletion to suppress the HU hypersensitivity of pol32Δ cells is in contrast to the negative genetic interaction between deletions of EXO1 and POL32 in MMS-treated cells as well as the role of EXO1 in DNA-damage treated rad53 and mec1 mutants.
Nucleases in homologous recombination as targets for cancer therapy
2014, FEBS Letters
Genomic DNA is constantly challenged from endogenous as well as exogenous sources. The DNA damage response (DDR) mechanism has evolved to combat these challenges and ensure genomic integrity. In this review, we will focus on repair of DNA double-strand breaks (DSB) by homologous recombination and the role of several nucleases and other recombination factors as suitable targets for cancer therapy. Their inactivation as well as overexpression have been shown to sensitize cancer cells by increasing toxicity to DNA-damaging agents and radiation or to be responsible for resistance of cancer cells. These factors can also be used in targeted cancer therapy by taking advantage of specific genetic abnormalities of cancer cells that are not present in normal cells and that result in cancer cell lethality.
MRE11A: a novel negative regulator of human DNA mismatch repair
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WRN Is a Promising Synthetic Lethal Target for Cancers with Microsatellite Instability (MSI)
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^*: Address requests for reprints to: Robert M. W. Hofstra, Ph.D., Department of Medical Genetics, Ant. Deusinglaan 4, 9713 AW Groningen, The Netherlands. E-mail: [email protected]; fax: (31) 50-363-2947.

^**: Supported by grant RUG 1997-1544 from the Dutch Cancer Society.

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Gastroenterology

Abstract

Section snippets

Patients

Mutation analysis

Discussion

References (20)

New clinical criteria for hereditary nonpolyposis colorectal cancer (HNPCC, Lynch syndrome) proposed by the International Collaborative Group on HNPCC

Gastroenterology

Association of hereditary nonpolyposis colorectal cancer-related tumours displaying low microsatellite instability with MSH6 germline mutations

Am J Hum Genet

Human exonuclease 1 functionally complements its yeast homologues in DNA recombination, RNA primer removal, and mutation avoidance

J Biol Chem

Purification of DNA from formaldehyde fixed and paraffin embedded human tissue

Biochem Biophys Res Commun

Inactivation of germline mutant APC alleles by attenuated somatic mutations: a molecular genetic mechanism for attenuated familial adenomatous polyposis

Am J Hum Genet

The international collaborative group on hereditary non-polyposis colorectal cancer (ICG-HNPCC)

Dis Colon Rectum

Conversion of diploidy to haploidy—individuals susceptible to multigene disorders may now be spotted more easily

Nature

Suspected hereditary nonpolyposis colorectal cancer

Dis Colon Rectum

Identification of a human gene encoding a homologue of Saccharomyces cerevisiae EXO1, an exonuclease implicated in mismatch repair and recombination

Cancer Res

Hex1: a new human Rad2 nuclease family member with homology to yeast exonuclease 1

Nucleic Acids Res

Cited by (121)

DNA-damage-induced degradation of EXO1 exonuclease limits DNA end resection to ensure accurate DNA repair

Mismatch Repair and Colon Cancer: Mechanisms and Therapies Explored

Exo1 phosphorylation status controls the hydroxyurea sensitivity of cells lacking the Pol32 subunit of DNA polymerases delta and zeta

Nucleases in homologous recombination as targets for cancer therapy

MRE11A: a novel negative regulator of human DNA mismatch repair

WRN Is a Promising Synthetic Lethal Target for Cancers with Microsatellite Instability (MSI)

Rapid CommunicationsGermline mutations of EXO1 gene in patients with hereditary nonpolyposis colorectal cancer (HNPCC) and atypical HNPCC forms*,**

Abstract

Section snippets

Patients

Mutation analysis

Discussion

Gastroenterology

Am J Hum Genet

J Biol Chem

Biochem Biophys Res Commun

Am J Hum Genet

The international collaborative group on hereditary non-polyposis colorectal cancer (ICG-HNPCC)

Dis Colon Rectum

Conversion of diploidy to haploidy—individuals susceptible to multigene disorders may now be spotted more easily

Nature

Suspected hereditary nonpolyposis colorectal cancer

Dis Colon Rectum

Identification of a human gene encoding a homologue of Saccharomyces cerevisiae EXO1, an exonuclease implicated in mismatch repair and recombination

Cancer Res

Hex1: a new human Rad2 nuclease family member with homology to yeast exonuclease 1

Nucleic Acids Res

Rapid Communications
Germline mutations of EXO1 gene in patients with hereditary nonpolyposis colorectal cancer (HNPCC) and atypical HNPCC forms*,**