Strand-specific Mismatch Repair in Mammalian Cells (1997)

Mistakes occurring during DNA transactions necessarily produce mismatched base pairs. DNA biosynthetic errors that escape the polymerase editing function yield mispaired bases, but mismatches also populate a recombination heteroduplex produced by strand transfer between related sequences that have diverged at the nucleotide level. Study of microbial systems has demonstrated the importance of mismatch correction in the reversal of such errors and consequent stabilization of the genome (1Kolodner R.D. Alani E. Curr. Opin. Biotechnol. 1994; 5: 585-594Crossref PubMed Scopus (58) Google Scholar, 2Matic I. Taddei F. Radman M. Trends Microbiol. 1996; 4: 69-72Abstract Full Text PDF PubMed Scopus (130) Google Scholar, 3Kolodner R. Genes Dev. 1996; 10: 1433-1442Crossref PubMed Scopus (544) Google Scholar, 4Modrich P. Lahue R. Annu. Rev. Biochem. 1996; 65: 101-133Crossref PubMed Scopus (1343) Google Scholar). 1Review articles. Interest in this process has increased dramatically over the past few years with the demonstration that mismatch repair plays an equally important role in mutation avoidance in mammalian cells and the finding that inactivation of the system by inherited or somatic mutation predisposes cells to tumor development and renders them resistant to the cytotoxic effects of certain DNA-damaging agents. This minireview will emphasize the molecular features of the mammalian strand-specific mismatch repair pathway. Work in other systems will be alluded to only as necessary for clarity or in those instances where compelling supporting data is available. The best understood function of mismatch repair is its role in the correction of DNA biosynthetic errors. Since mismatches are comprised of normal Watson-Crick bases, mismatch repair systems rely on secondary signals within the helix to distinguish daughter and parental DNA strands, with the former by definition containing the incorrect nucleotide of a mispair. The nature of the strand signal has been established in Escherichia coli, where repair is directed by the state of adenine methylation of d(GATC) sequences. Since this modification is postsynthetic, d(GATC) sequences in newly synthesized DNA exist in a transiently unmodified state, and it is the absence of methylation that directs repair to the daughter strand. Initiation of the bacterial repair reaction occurs via mismatch-provoked incision of the unmodified strand at a hemimethylated d(GATC) site, and it is the nick in the new strand that actually directs repair (4Modrich P. Lahue R. Annu. Rev. Biochem. 1996; 65: 101-133Crossref PubMed Scopus (1343) Google Scholar).1 Although the nature of the strand signal has not been defined in any eukaryotic organism, heteroduplex DNAs containing a site-specific, strand-specific incision are subject to mismatch-provoked, strand-specific repair by human cell extracts in a reaction that requires ATP, the four dNTPs, and a divalent cation (5Holmes J. Clark S. Modrich P. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 5837-5841Crossref PubMed Scopus (345) Google Scholar, 6Thomas D.C. Roberts J.D. Kunkel T.A. J. Biol. Chem. 1991; 266: 3744-3751Abstract Full Text PDF PubMed Google Scholar). All eight base-base mismatches are subject to nick-directed correction, as are small insertion/deletion (ID) 2The abbreviations used are: ID, insertion/deletion; dCTPαS, 2′-deoxycytidine-5′-O-(1-thiotriphosphate); PCNA, proliferating cell nuclear antigen; pol, polymerase; HNPCC, hereditary nonpolyposis colon cancer; MNNG,N-methyl-N′-nitro-N-nitrosoguanidine; MNU, N-methyl-N-nitrosourea; O6meG, O6-methylguanine. mispairs in which one strand contains a consecutive run of a few extra nucleotides (5Holmes J. Clark S. Modrich P. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 5837-5841Crossref PubMed Scopus (345) Google Scholar, 6Thomas D.C. Roberts J.D. Kunkel T.A. J. Biol. Chem. 1991; 266: 3744-3751Abstract Full Text PDF PubMed Google Scholar, 7Fang W.-h. Modrich P. J. Biol. Chem. 1993; 268: 11838-11844Abstract Full Text PDF PubMed Google Scholar, 8Parsons R. Li G.M. Longley M.J. Fang W.H. Papadopoulos N. Jen J. de la Chapelle A. Kinzler K.W. Vogelstein B. Modrich P. Cell. 1993; 75: 1227-1236Abstract Full Text PDF PubMed Scopus (973) Google Scholar, 9Umar A. Boyer J.C. Thomas D.C. Nguyen D.C. Risinger J.I. Boyd J. Ionov Y. Perucho M. Kunkel T.A. J. Biol. Chem. 1994; 269: 14367-14370Abstract Full Text PDF PubMed Google Scholar, 10Umar A. Boyer J.C. Kunkel T.A. Science. 1994; 266: 814-816Crossref PubMed Scopus (125) Google Scholar). However, some mismatches are better substrates than others, and sequence context effects have been observed. In general, Pur-Pyr and Pur-Pur mismatches are among the best substrates, and Pyr-Pyr mispairs are processed with reduced efficiency. The C-C mismatch is the weakest substrate, repaired in some sequence contexts but not others (5Holmes J. Clark S. Modrich P. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 5837-5841Crossref PubMed Scopus (345) Google Scholar, 6Thomas D.C. Roberts J.D. Kunkel T.A. J. Biol. Chem. 1991; 266: 3744-3751Abstract Full Text PDF PubMed Google Scholar, 8Parsons R. Li G.M. Longley M.J. Fang W.H. Papadopoulos N. Jen J. de la Chapelle A. Kinzler K.W. Vogelstein B. Modrich P. Cell. 1993; 75: 1227-1236Abstract Full Text PDF PubMed Scopus (973) Google Scholar). ID heterologies of 1–5 nucleotides are subject to nick-directed correction with efficiencies comparable with those of Pur-Pyr and Pur-Pur mispairs (8Parsons R. Li G.M. Longley M.J. Fang W.H. Papadopoulos N. Jen J. de la Chapelle A. Kinzler K.W. Vogelstein B. Modrich P. Cell. 1993; 75: 1227-1236Abstract Full Text PDF PubMed Scopus (973) Google Scholar, 10Umar A. Boyer J.C. Kunkel T.A. Science. 1994; 266: 814-816Crossref PubMed Scopus (125) Google Scholar, 11Drummond J.T. Li G.-M. Longley M.J. Modrich P. Science. 1995; 268: 1909-1912Crossref PubMed Scopus (541) Google Scholar). The upper limit on ID heterology size for correction by the mismatch repair system appears to lie between 8 and 12 nucleotides, with larger heterologies processed by a distinct system (10Umar A. Boyer J.C. Kunkel T.A. Science. 1994; 266: 814-816Crossref PubMed Scopus (125) Google Scholar). 3S. Littman, W.-h. Fang, and P. Modrich, unpublished experiments. The mechanism of nick-directed mismatch repair, deduced from study of the reaction in human nuclear extracts, is shown in Fig.1. The strand break that directs correction can be located either 3′ or 5′ to the mispair, with mismatch-provoked excision removing that portion of the incised strand spanning the two DNA sites (7Fang W.-h. Modrich P. J. Biol. Chem. 1993; 268: 11838-11844Abstract Full Text PDF PubMed Google Scholar). This mechanism is largely based on study of the fate of nicked heteroduplex DNA in nuclear extracts under conditions of repair DNA synthesis inhibition by aphidicolin (5Holmes J. Clark S. Modrich P. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 5837-5841Crossref PubMed Scopus (345) Google Scholar, 6Thomas D.C. Roberts J.D. Kunkel T.A. J. Biol. Chem. 1991; 266: 3744-3751Abstract Full Text PDF PubMed Google Scholar) or by omission of exogenous dNTPs (7Fang W.-h. Modrich P. J. Biol. Chem. 1993; 268: 11838-11844Abstract Full Text PDF PubMed Google Scholar). Mismatch-free, nicked homoduplex DNA is ligated under these conditions, but heteroduplex molecules are subject to mismatch-provoked excision to yield a gap extending from the site of the strand break and terminating at a number of discrete sites 90–170 nucleotides beyond the original location of the mispair. This scheme is also consistent with the location of repair DNA synthesis tracts visualized by incorporation of radiolabeled nucleotides (5Holmes J. Clark S. Modrich P. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 5837-5841Crossref PubMed Scopus (345) Google Scholar, 6Thomas D.C. Roberts J.D. Kunkel T.A. J. Biol. Chem. 1991; 266: 3744-3751Abstract Full Text PDF PubMed Google Scholar) or by mapping incorporated α-thionucleotides when dCTP was partially substituted with dCTPαS (12Fang, W.-h. (1993) Strand-specific Mismatch Repair in Human Cells. Ph.D. thesis, Duke UniversityGoogle Scholar). Since the repaired DNA strand is recovered from nuclear extracts largely as a full-length, covalently continuous species (7Fang W.-h. Modrich P. J. Biol. Chem. 1993; 268: 11838-11844Abstract Full Text PDF PubMed Google Scholar), the last step of the reaction presumably involves ligase closure of the strand break that remains after repair synthesis. The nature of the excision step in the human repair reaction has not been established, but the mechanism of the overall reaction shown in Fig. 1 is similar to that of E. coli methyl-directed mismatch correction, in which excision is exonucleolytic and initiates at the strand break (4Modrich P. Lahue R. Annu. Rev. Biochem. 1996; 65: 101-133Crossref PubMed Scopus (1343) Google Scholar).1 Since each of the base-base mismatches and a dinucleotide ID mispair provoke a similar gap formation reaction in human cell extracts (7Fang W.-h. Modrich P. J. Biol. Chem. 1993; 268: 11838-11844Abstract Full Text PDF PubMed Google Scholar, 8Parsons R. Li G.M. Longley M.J. Fang W.H. Papadopoulos N. Jen J. de la Chapelle A. Kinzler K.W. Vogelstein B. Modrich P. Cell. 1993; 75: 1227-1236Abstract Full Text PDF PubMed Scopus (973) Google Scholar), nick-directed repair of these different heterologies has been attributed to a single pathway. This broad mismatch specificity is similar to that of the bacterial methyl-directed system. Initiation of bacterial mismatch repair occurs via mismatch recognition by a MutS homodimer followed by binding of MutL, with the latter protein probably serving to interface mismatch recognition by MutS to other components of the repair system (4Modrich P. Lahue R. Annu. Rev. Biochem. 1996; 65: 101-133Crossref PubMed Scopus (1343) Google Scholar, 13Allen, D. J., Makhov, A., Grilley, M., Taylor, J., Thresher, R., Modrich, P., and Griffith, J. D. (1997) EMBO J., Vol. 16 in pressGoogle Scholar).1Human mismatch repair has not been reconstituted in a defined system, but five activities have been implicated in the reaction, including several that are structural and functional homologs of MutS and MutL. As discussed below, certain tumor cell lines, as well as hypermutable cell lines resistant to the cytotoxic action of certain DNA-damaging agents, are defective in nick-directed mismatch repair. These cell lines define several in vitro complementation groups (9Umar A. Boyer J.C. Thomas D.C. Nguyen D.C. Risinger J.I. Boyd J. Ionov Y. Perucho M. Kunkel T.A. J. Biol. Chem. 1994; 269: 14367-14370Abstract Full Text PDF PubMed Google Scholar,11Drummond J.T. Li G.-M. Longley M.J. Modrich P. Science. 1995; 268: 1909-1912Crossref PubMed Scopus (541) Google Scholar), and activities that restore repair to extracts of two of these have been isolated. Human MutSα, a heterodimer of the MutS homologs MSH2 and MSH6 (MSH6 has also been called GTBP or p160), binds to base-base mismatches and small ID heterologies and restores nick-directed repair on both types of substrate to extracts of cell lines that are genetically deficient in MSH2 (11Drummond J.T. Li G.-M. Longley M.J. Modrich P. Science. 1995; 268: 1909-1912Crossref PubMed Scopus (541) Google Scholar,14Palombo F. Gallinari P. Iaccarino I. Lettieri T. Hughes M. D'Arrigo A. Truong O. Hsuan J.J. Jiricny J. Science. 1995; 268: 1912-1914Crossref PubMed Scopus (483) Google Scholar, 15Palombo F. Iaccarino I. Nakajima E. Ikejima M. Shimada T. Jiricny J. Curr. Biol. 1996; 6: 1181-1184Abstract Full Text Full Text PDF PubMed Scopus (304) Google Scholar, 16Acharya S. Wilson T. Gradia S. Kane M.F. Guerrette S. Marsischky G.T. Kolodner R. Fishel R. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 13629-13634Crossref PubMed Scopus (476) Google Scholar). 4J. Drummond, J. Genschel, E. Wolf, and P. Modrich, manuscript in preparation; J. Genschel, S. Littman, and P. Modrich, manuscript in preparation. The affinity of MutSα for an oligonucleotide heteroduplex is markedly reduced in the presence of ATP (11Drummond J.T. Li G.-M. Longley M.J. Modrich P. Science. 1995; 268: 1909-1912Crossref PubMed Scopus (541) Google Scholar), an effect that has also been observed with bacterial MutS. In the case of the bacterial protein, this effect has been attributed to departure of the MutS dimer from the mismatch by translocation or linear diffusion along the DNA contour, with movement of the protein along the helix presumably involved in the search for the strand signal that directs repair (13Allen, D. J., Makhov, A., Grilley, M., Taylor, J., Thresher, R., Modrich, P., and Griffith, J. D. (1997) EMBO J., Vol. 16 in pressGoogle Scholar). In addition to its ability to complex with MSH6, MSH2 also forms a heterodimer with a second MutS homolog called MSH3 (15Palombo F. Iaccarino I. Nakajima E. Ikejima M. Shimada T. Jiricny J. Curr. Biol. 1996; 6: 1181-1184Abstract Full Text Full Text PDF PubMed Scopus (304) Google Scholar, 16Acharya S. Wilson T. Gradia S. Kane M.F. Guerrette S. Marsischky G.T. Kolodner R. Fishel R. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 13629-13634Crossref PubMed Scopus (476) Google Scholar). This MutSβ complex binds to ID mispairs but displays little if any affinity for base-base mismatches. Furthermore, cell lines genetically defective in the MSH6 subunit of MutSα are deficient in base-base mismatch correction but retain partial activity on ID mispairs, and this residual activity has been shown to be due to the MutSβ complex (11Drummond J.T. Li G.-M. Longley M.J. Modrich P. Science. 1995; 268: 1909-1912Crossref PubMed Scopus (541) Google Scholar).4 MutSα thus functions in base-base and ID mismatch repair, but MutSβ is apparently restricted to processing of the latter class of heterology. Whereas these findings may suggest redundant functions for MutSα and MutSβ in the recognition and processing of ID mispairs, there is some evidence that MutSα and MutSβ may function in a complementary fashion, differentially recognizing ID heterologies depending on size and sequence context (15Palombo F. Iaccarino I. Nakajima E. Ikejima M. Shimada T. Jiricny J. Curr. Biol. 1996; 6: 1181-1184Abstract Full Text Full Text PDF PubMed Scopus (304) Google Scholar). Recombinant MSH2 has also been reported to bind to mismatched base pairs (17Fishel R. Ewel A. Lescoe M.K. Cancer Res. 1994; 54: 5539-5542PubMed Google Scholar), but the heteroduplex affinity of this protein is considerably less than that of MutSα or MutSβ (11Drummond J.T. Li G.-M. Longley M.J. Modrich P. Science. 1995; 268: 1909-1912Crossref PubMed Scopus (541) Google Scholar, 14Palombo F. Gallinari P. Iaccarino I. Lettieri T. Hughes M. D'Arrigo A. Truong O. Hsuan J.J. Jiricny J. Science. 1995; 268: 1912-1914Crossref PubMed Scopus (483) Google Scholar, 15Palombo F. Iaccarino I. Nakajima E. Ikejima M. Shimada T. Jiricny J. Curr. Biol. 1996; 6: 1181-1184Abstract Full Text Full Text PDF PubMed Scopus (304) Google Scholar, 16Acharya S. Wilson T. Gradia S. Kane M.F. Guerrette S. Marsischky G.T. Kolodner R. Fishel R. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 13629-13634Crossref PubMed Scopus (476) Google Scholar). Inasmuch as free MSH2 has not been detected during fractionation of human nuclear extracts4 and since epistasis analysis suggests that Saccharomyces cerevisiae MSH2 functions in mitotic genetic stability only in conjunction with MSH3 or MSH6 (18Marsischky G.T. Filosi N. Kane M.F. Kolodner R. Genes Dev. 1996; 10: 407-420Crossref PubMed Scopus (506) Google Scholar,19Johnson R.E. Kovvali G.K. Prakash L. Prakash S. J. Biol. Chem. 1996; 271: 7285-7288Abstract Full Text Full Text PDF PubMed Scopus (167) Google Scholar), the biological function of free MSH2 must be regarded as uncertain. A human MutL activity has been by of its ability to restore strand-specific mismatch repair to extracts of one class of hypermutable tumor cell G.M. Modrich P. Proc. Natl. Acad. Sci. U. S. A. 1995; PubMed Scopus Google Scholar). This called is a heterodimer of the MutL homologs and Since cell lines deficient in are defective in repair of base-base and ID mismatches (8Parsons R. Li G.M. Longley M.J. Fang W.H. Papadopoulos N. Jen J. de la Chapelle A. Kinzler K.W. Vogelstein B. Modrich P. Cell. 1993; 75: 1227-1236Abstract Full Text PDF PubMed Scopus (973) Google Scholar, J.I. A. J.C. Kunkel T.A. J. Biol. Chem. 1995; Full Text Full Text PDF PubMed Scopus Google Scholar) and since the heterodimer restores repair on both types of heterology G.M. Modrich P. Proc. Natl. Acad. Sci. U. S. A. 1995; PubMed Scopus Google Scholar), presumably functions with MutSα in base-base mismatch repair or with either MutSα or MutSβ in ID heterology repair. Although binding of to or has not been in either or MutSα mismatch correction at or to the excision of repair (8Parsons R. Li G.M. Longley M.J. Fang W.H. Papadopoulos N. Jen J. de la Chapelle A. Kinzler K.W. Vogelstein B. Modrich P. Cell. 1993; 75: 1227-1236Abstract Full Text PDF PubMed Scopus (973) Google Scholar, A. Fang W.H. Longley M.J. Li G.M. Modrich P. Proc. Natl. Acad. Sci. U. S. A. 1993; PubMed Scopus Google Scholar), consistent with function of both activities at an step in repair. S. cerevisiae has been shown to bind to between MSH2 and heteroduplex or homoduplex DNA T.A. Alani E. Kolodner R.D. Science. 1994; PubMed Scopus Google Scholar), but this is subject to the the functional of MSH2 in the absence of MSH6 or cell nuclear a of the eukaryotic forms a the helix and the of DNA and S. P. J. 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Cell. 1996; 87: Full Text Full Text PDF PubMed Scopus Google Scholar) that PCNA, MutSα and functions in repair at or to the excision mismatch-provoked excision in human cell extracts is by or by a that binds to and also DNA S. B. 1994; PubMed Scopus Google Scholar). of in an step of mismatch repair may a role for the protein in the of the mismatch and the strand signal and in of an excision system A. Thomas D.C. Clark Kunkel T.A. Cell. 1996; 87: Full Text Full Text PDF PubMed Scopus Google Scholar) or may a between the mismatch repair system and the as has been for the bacterial (4Modrich P. Lahue R. Annu. Rev. Biochem. 1996; 65: 101-133Crossref PubMed Scopus (1343) Google Scholar).1 The repair DNA synthesis step of mismatch repair in human extracts is by aphidicolin (5Holmes J. Clark S. Modrich P. Proc. Natl. Acad. Sci. U. S. A. 1990; 87: 5837-5841Crossref PubMed Scopus (345) Google Scholar, 6Thomas D.C. Roberts J.D. Kunkel T.A. J. Biol. Chem. 1991; 266: 3744-3751Abstract Full Text PDF PubMed Google Scholar, 7Fang W.-h. Modrich P. J. Biol. Chem. 1993; 268: 11838-11844Abstract Full Text PDF PubMed Google Scholar) DNA polymerase in the of but containing residual and normal of have been shown to be in mismatch-provoked excision but defective in the repair DNA synthesis step M.J. Modrich P. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). This is by this DNA polymerase in the The presence of residual and in the extracts used in these for these but are to the repair synthesis In of the role of in mismatch correction, it is that this protein with M. B. PubMed Scopus Google Scholar) and probably with as well J. J. Biol. 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