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Mol Cell Biol. Oct 1996; 16(10): 5604–5615.
PMCID: PMC231560

The Saccharomyces cerevisiae Msh2 and Msh6 proteins form a complex that specifically binds to duplex oligonucleotides containing mismatched DNA base pairs.

Abstract

The yeast Saccharomyces cerevisiae encodes six proteins, Msh1p to Msh6p, that show strong amino acid sequence similarity to MutS, a central component of the bacterial mutHLS mismatch repair system. Recent studies with humans and S. cerevisiae suggest that in eukaryotes, specific MutS homolog complexes that display unique DNA mismatch specificities exist. In this study, the S. cerevisiae 109-kDa Msh2 and 140-kDa Msh6 proteins were cooverexpressed in S. cerevisiae and shown to interact in an immunoprecipitation assay and by conventional chromatography. Deletion analysis of MSH2 indicated that the carboxy-terminal 114 amino acids of Msh2p are important for Msh6p interaction. Purified Msh2p-Msh6p selectively bound to duplex oligonucleotide substrates containing a G/T mismatch and a +1 insertion mismatch but did not show specific binding to +2 and +4 insertion mismatches. The mismatch binding specificity of the Msh2p-Msh6p complex, as measured by on-rate and off-rate binding studies, was abolished by ATP. Interestingly, palindromic substrates that are poorly repaired in vivo were specifically recognized by Msh2p-Msh6p; however, the binding of Msh2p-Msh6p to these substrates was not modulated by ATP. Taken together, these studies suggest that the repair of a base pair mismatch by the Msh2p-Msh6p complex is dependent on the ability of the Msh2p-Msh6p-DNA mismatch complex to use ATP hydrolysis to activate downstream events in mismatch repair.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.
  • Aaltonen LA, Peltomäki P, Leach FS, Sistonen P, Pylkkänen L, Mecklin JP, Järvinen H, Powell SM, Jen J, Hamilton SR, et al. Clues to the pathogenesis of familial colorectal cancer. Science. 1993 May 7;260(5109):812–816. [PubMed]
  • Alani E, Chi NW, Kolodner R. The Saccharomyces cerevisiae Msh2 protein specifically binds to duplex oligonucleotides containing mismatched DNA base pairs and insertions. Genes Dev. 1995 Jan 15;9(2):234–247. [PubMed]
  • Alani E, Reenan RA, Kolodner RD. Interaction between mismatch repair and genetic recombination in Saccharomyces cerevisiae. Genetics. 1994 May;137(1):19–39. [PMC free article] [PubMed]
  • Au KG, Welsh K, Modrich P. Initiation of methyl-directed mismatch repair. J Biol Chem. 1992 Jun 15;267(17):12142–12148. [PubMed]
  • Bishop DK, Andersen J, Kolodner RD. Specificity of mismatch repair following transformation of Saccharomyces cerevisiae with heteroduplex plasmid DNA. Proc Natl Acad Sci U S A. 1989 May;86(10):3713–3717. [PMC free article] [PubMed]
  • Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. [PubMed]
  • Bronner CE, Baker SM, Morrison PT, Warren G, Smith LG, Lescoe MK, Kane M, Earabino C, Lipford J, Lindblom A, et al. Mutation in the DNA mismatch repair gene homologue hMLH1 is associated with hereditary non-polyposis colon cancer. Nature. 1994 Mar 17;368(6468):258–261. [PubMed]
  • Chi NW, Kolodner RD. Purification and characterization of MSH1, a yeast mitochondrial protein that binds to DNA mismatches. J Biol Chem. 1994 Nov 25;269(47):29984–29992. [PubMed]
  • Chi NW, Kolodner RD. The effect of DNA mismatches on the ATPase activity of MSH1, a protein in yeast mitochondria that recognizes DNA mismatches. J Biol Chem. 1994 Nov 25;269(47):29993–29997. [PubMed]
  • Christianson TW, Sikorski RS, Dante M, Shero JH, Hieter P. Multifunctional yeast high-copy-number shuttle vectors. Gene. 1992 Jan 2;110(1):119–122. [PubMed]
  • Datta A, Adjiri A, New L, Crouse GF, Jinks Robertson S. Mitotic crossovers between diverged sequences are regulated by mismatch repair proteins in Saccaromyces cerevisiae. Mol Cell Biol. 1996 Mar;16(3):1085–1093. [PMC free article] [PubMed]
  • Detloff P, Petes TD. Measurements of excision repair tracts formed during meiotic recombination in Saccharomyces cerevisiae. Mol Cell Biol. 1992 Apr;12(4):1805–1814. [PMC free article] [PubMed]
  • de Wind N, Dekker M, Berns A, Radman M, te Riele H. Inactivation of the mouse Msh2 gene results in mismatch repair deficiency, methylation tolerance, hyperrecombination, and predisposition to cancer. Cell. 1995 Jul 28;82(2):321–330. [PubMed]
  • Drummond JT, Li GM, Longley MJ, Modrich P. Isolation of an hMSH2-p160 heterodimer that restores DNA mismatch repair to tumor cells. Science. 1995 Jun 30;268(5219):1909–1912. [PubMed]
  • Fishel R, Ewel A, Lee S, Lescoe MK, Griffith J. Binding of mismatched microsatellite DNA sequences by the human MSH2 protein. Science. 1994 Nov 25;266(5189):1403–1405. [PubMed]
  • Fishel R, Lescoe MK, Rao MR, Copeland NG, Jenkins NA, Garber J, Kane M, Kolodner R. The human mutator gene homolog MSH2 and its association with hereditary nonpolyposis colon cancer. Cell. 1993 Dec 3;75(5):1027–1038. [PubMed]
  • Gietz RD, Schiestl RH. Applications of high efficiency lithium acetate transformation of intact yeast cells using single-stranded nucleic acids as carrier. Yeast. 1991 Apr;7(3):253–263. [PubMed]
  • Grilley M, Welsh KM, Su SS, Modrich P. Isolation and characterization of the Escherichia coli mutL gene product. J Biol Chem. 1989 Jan 15;264(2):1000–1004. [PubMed]
  • Haber LT, Walker GC. Altering the conserved nucleotide binding motif in the Salmonella typhimurium MutS mismatch repair protein affects both its ATPase and mismatch binding activities. EMBO J. 1991 Sep;10(9):2707–2715. [PMC free article] [PubMed]
  • Ho SN, Hunt HD, Horton RM, Pullen JK, Pease LR. Site-directed mutagenesis by overlap extension using the polymerase chain reaction. Gene. 1989 Apr 15;77(1):51–59. [PubMed]
  • Hollingsworth NM, Ponte L, Halsey C. MSH5, a novel MutS homolog, facilitates meiotic reciprocal recombination between homologs in Saccharomyces cerevisiae but not mismatch repair. Genes Dev. 1995 Jul 15;9(14):1728–1739. [PubMed]
  • Holmes J, Jr, Clark S, Modrich P. Strand-specific mismatch correction in nuclear extracts of human and Drosophila melanogaster cell lines. Proc Natl Acad Sci U S A. 1990 Aug;87(15):5837–5841. [PMC free article] [PubMed]
  • Hughes MJ, Jiricny J. The purification of a human mismatch-binding protein and identification of its associated ATPase and helicase activities. J Biol Chem. 1992 Nov 25;267(33):23876–23882. [PubMed]
  • Hunter N, Chambers SR, Louis EJ, Borts RH. The mismatch repair system contributes to meiotic sterility in an interspecific yeast hybrid. EMBO J. 1996 Apr 1;15(7):1726–1733. [PMC free article] [PubMed]
  • Ionov Y, Peinado MA, Malkhosyan S, Shibata D, Perucho M. Ubiquitous somatic mutations in simple repeated sequences reveal a new mechanism for colonic carcinogenesis. Nature. 1993 Jun 10;363(6429):558–561. [PubMed]
  • Jiricny J, Su SS, Wood SG, Modrich P. Mismatch-containing oligonucleotide duplexes bound by the E. coli mutS-encoded protein. Nucleic Acids Res. 1988 Aug 25;16(16):7843–7853. [PMC free article] [PubMed]
  • Johnson AW, Kolodner RD. Strand exchange protein 1 from Saccharomyces cerevisiae. A novel multifunctional protein that contains DNA strand exchange and exonuclease activities. J Biol Chem. 1991 Jul 25;266(21):14046–14054. [PubMed]
  • Johnson RE, Kovvali GK, Prakash L, Prakash S. Requirement of the yeast MSH3 and MSH6 genes for MSH2-dependent genomic stability. J Biol Chem. 1996 Mar 29;271(13):7285–7288. [PubMed]
  • Kolodner RD. Mismatch repair: mechanisms and relationship to cancer susceptibility. Trends Biochem Sci. 1995 Oct;20(10):397–401. [PubMed]
  • Kolodziej PA, Young RA. Epitope tagging and protein surveillance. Methods Enzymol. 1991;194:508–519. [PubMed]
  • Kramer B, Kramer W, Williamson MS, Fogel S. Heteroduplex DNA correction in Saccharomyces cerevisiae is mismatch specific and requires functional PMS genes. Mol Cell Biol. 1989 Oct;9(10):4432–4440. [PMC free article] [PubMed]
  • Kramer W, Kramer B, Williamson MS, Fogel S. Cloning and nucleotide sequence of DNA mismatch repair gene PMS1 from Saccharomyces cerevisiae: homology of PMS1 to procaryotic MutL and HexB. J Bacteriol. 1989 Oct;171(10):5339–5346. [PMC free article] [PubMed]
  • Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. [PubMed]
  • Lahue RS, Au KG, Modrich P. DNA mismatch correction in a defined system. Science. 1989 Jul 14;245(4914):160–164. [PubMed]
  • Landschulz WH, Johnson PF, McKnight SL. The leucine zipper: a hypothetical structure common to a new class of DNA binding proteins. Science. 1988 Jun 24;240(4860):1759–1764. [PubMed]
  • Leach FS, Nicolaides NC, Papadopoulos N, Liu B, Jen J, Parsons R, Peltomäki P, Sistonen P, Aaltonen LA, Nyström-Lahti M, et al. Mutations of a mutS homolog in hereditary nonpolyposis colorectal cancer. Cell. 1993 Dec 17;75(6):1215–1225. [PubMed]
  • Levinson G, Gutman GA. High frequencies of short frameshifts in poly-CA/TG tandem repeats borne by bacteriophage M13 in Escherichia coli K-12. Nucleic Acids Res. 1987 Jul 10;15(13):5323–5338. [PMC free article] [PubMed]
  • Li GM, Modrich P. Restoration of mismatch repair to nuclear extracts of H6 colorectal tumor cells by a heterodimer of human MutL homologs. Proc Natl Acad Sci U S A. 1995 Mar 14;92(6):1950–1954. [PMC free article] [PubMed]
  • Manivasakam P, Rosenberg SM, Hastings PJ. Poorly repaired mismatches in heteroduplex DNA are hyper-recombinagenic in Saccharomyces cerevisiae. Genetics. 1996 Feb;142(2):407–416. [PMC free article] [PubMed]
  • Marsischky GT, Filosi N, Kane MF, Kolodner R. Redundancy of Saccharomyces cerevisiae MSH3 and MSH6 in MSH2-dependent mismatch repair. Genes Dev. 1996 Feb 15;10(4):407–420. [PubMed]
  • Matic I, Rayssiguier C, Radman M. Interspecies gene exchange in bacteria: the role of SOS and mismatch repair systems in evolution of species. Cell. 1995 Feb 10;80(3):507–515. [PubMed]
  • McEntee K, Weinstock GM, Lehman IR. recA protein-catalyzed strand assimilation: stimulation by Escherichia coli single-stranded DNA-binding protein. Proc Natl Acad Sci U S A. 1980 Feb;77(2):857–861. [PMC free article] [PubMed]
  • Miret JJ, Milla MG, Lahue RS. Characterization of a DNA mismatch-binding activity in yeast extracts. J Biol Chem. 1993 Feb 15;268(5):3507–3513. [PubMed]
  • Miret JJ, Parker BO, Lahua RS. Recognition of DNA insertion/deletion mismatches by an activity in Saccharomyces cerevisiae. Nucleic Acids Res. 1996 Feb 15;24(4):721–729. [PMC free article] [PubMed]
  • Modrich P. Mechanisms and biological effects of mismatch repair. Annu Rev Genet. 1991;25:229–253. [PubMed]
  • Nag DK, Petes TD. Seven-base-pair inverted repeats in DNA form stable hairpins in vivo in Saccharomyces cerevisiae. Genetics. 1991 Nov;129(3):669–673. [PMC free article] [PubMed]
  • New L, Liu K, Crouse GF. The yeast gene MSH3 defines a new class of eukaryotic MutS homologues. Mol Gen Genet. 1993 May;239(1-2):97–108. [PubMed]
  • Nicolaides NC, Papadopoulos N, Liu B, Wei YF, Carter KC, Ruben SM, Rosen CA, Haseltine WA, Fleischmann RD, Fraser CM, et al. Mutations of two PMS homologues in hereditary nonpolyposis colon cancer. Nature. 1994 Sep 1;371(6492):75–80. [PubMed]
  • Palombo F, Gallinari P, Iaccarino I, Lettieri T, Hughes M, D'Arrigo A, Truong O, Hsuan JJ, Jiricny J. GTBP, a 160-kilodalton protein essential for mismatch-binding activity in human cells. Science. 1995 Jun 30;268(5219):1912–1914. [PubMed]
  • Papadopoulos N, Nicolaides NC, Liu B, Parsons R, Lengauer C, Palombo F, D'Arrigo A, Markowitz S, Willson JK, Kinzler KW, et al. Mutations of GTBP in genetically unstable cells. Science. 1995 Jun 30;268(5219):1915–1917. [PubMed]
  • Papadopoulos N, Nicolaides NC, Wei YF, Ruben SM, Carter KC, Rosen CA, Haseltine WA, Fleischmann RD, Fraser CM, Adams MD, et al. Mutation of a mutL homolog in hereditary colon cancer. Science. 1994 Mar 18;263(5153):1625–1629. [PubMed]
  • Parker BO, Marinus MG. Repair of DNA heteroduplexes containing small heterologous sequences in Escherichia coli. Proc Natl Acad Sci U S A. 1992 Mar 1;89(5):1730–1734. [PMC free article] [PubMed]
  • Parsons R, Li GM, Longley MJ, Fang WH, Papadopoulos N, Jen J, de la Chapelle A, Kinzler KW, Vogelstein B, Modrich P. Hypermutability and mismatch repair deficiency in RER+ tumor cells. Cell. 1993 Dec 17;75(6):1227–1236. [PubMed]
  • Prolla TA, Christie DM, Liskay RM. Dual requirement in yeast DNA mismatch repair for MLH1 and PMS1, two homologs of the bacterial mutL gene. Mol Cell Biol. 1994 Jan;14(1):407–415. [PMC free article] [PubMed]
  • Prolla TA, Pang Q, Alani E, Kolodner RD, Liskay RM. MLH1, PMS1, and MSH2 interactions during the initiation of DNA mismatch repair in yeast. Science. 1994 Aug 19;265(5175):1091–1093. [PubMed]
  • Rayssiguier C, Thaler DS, Radman M. The barrier to recombination between Escherichia coli and Salmonella typhimurium is disrupted in mismatch-repair mutants. Nature. 1989 Nov 23;342(6248):396–401. [PubMed]
  • Reenan RA, Kolodner RD. Isolation and characterization of two Saccharomyces cerevisiae genes encoding homologs of the bacterial HexA and MutS mismatch repair proteins. Genetics. 1992 Dec;132(4):963–973. [PMC free article] [PubMed]
  • Reenan RA, Kolodner RD. Characterization of insertion mutations in the Saccharomyces cerevisiae MSH1 and MSH2 genes: evidence for separate mitochondrial and nuclear functions. Genetics. 1992 Dec;132(4):975–985. [PMC free article] [PubMed]
  • Ross-Macdonald P, Roeder GS. Mutation of a meiosis-specific MutS homolog decreases crossing over but not mismatch correction. Cell. 1994 Dec 16;79(6):1069–1080. [PubMed]
  • Selva EM, New L, Crouse GF, Lahue RS. Mismatch correction acts as a barrier to homeologous recombination in Saccharomyces cerevisiae. Genetics. 1995 Mar;139(3):1175–1188. [PMC free article] [PubMed]
  • Shen P, Huang HV. Effect of base pair mismatches on recombination via the RecBCD pathway. Mol Gen Genet. 1989 Aug;218(2):358–360. [PubMed]
  • Strand M, Earley MC, Crouse GF, Petes TD. Mutations in the MSH3 gene preferentially lead to deletions within tracts of simple repetitive DNA in Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1995 Oct 24;92(22):10418–10421. [PMC free article] [PubMed]
  • Strand M, Prolla TA, Liskay RM, Petes TD. Destabilization of tracts of simple repetitive DNA in yeast by mutations affecting DNA mismatch repair. Nature. 1993 Sep 16;365(6443):274–276. [PubMed]
  • Streisinger G, Okada Y, Emrich J, Newton J, Tsugita A, Terzaghi E, Inouye M. Frameshift mutations and the genetic code. This paper is dedicated to Professor Theodosius Dobzhansky on the occasion of his 66th birthday. Cold Spring Harb Symp Quant Biol. 1966;31:77–84. [PubMed]
  • Su SS, Lahue RS, Au KG, Modrich P. Mispair specificity of methyl-directed DNA mismatch correction in vitro. J Biol Chem. 1988 May 15;263(14):6829–6835. [PubMed]
  • Thibodeau SN, Bren G, Schaid D. Microsatellite instability in cancer of the proximal colon. Science. 1993 May 7;260(5109):816–819. [PubMed]
  • Thomas DC, Roberts JD, Kunkel TA. Heteroduplex repair in extracts of human HeLa cells. J Biol Chem. 1991 Feb 25;266(6):3744–3751. [PubMed]
  • Umar A, Boyer JC, Thomas DC, Nguyen DC, Risinger JI, Boyd J, Ionov Y, Perucho M, Kunkel TA. Defective mismatch repair in extracts of colorectal and endometrial cancer cell lines exhibiting microsatellite instability. J Biol Chem. 1994 May 20;269(20):14367–14370. [PubMed]
  • Varlet I, Radman M, Brooks P. DNA mismatch repair in Xenopus egg extracts: repair efficiency and DNA repair synthesis for all single base-pair mismatches. Proc Natl Acad Sci U S A. 1990 Oct;87(20):7883–7887. [PMC free article] [PubMed]
  • Welsh KM, Lu AL, Clark S, Modrich P. Isolation and characterization of the Escherichia coli mutH gene product. J Biol Chem. 1987 Nov 15;262(32):15624–15629. [PubMed]
  • Winston F, Dollard C, Ricupero-Hovasse SL. Construction of a set of convenient Saccharomyces cerevisiae strains that are isogenic to S288C. Yeast. 1995 Jan;11(1):53–55. [PubMed]
  • Worth L, Jr, Clark S, Radman M, Modrich P. Mismatch repair proteins MutS and MutL inhibit RecA-catalyzed strand transfer between diverged DNAs. Proc Natl Acad Sci U S A. 1994 Apr 12;91(8):3238–3241. [PMC free article] [PubMed]

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