Crystal Structure of Human REV7 in Complex with a Human REV3 Fragment and Structural Implication of the Interaction between DNA Polymerase ζ and REV1
2010; Elsevier BV; Volume: 285; Issue: 16 Linguagem: Inglês
10.1074/jbc.m109.092403
ISSN1083-351X
AutoresKodai Hara, Hiroshi Hashimoto, Yoshiki Murakumo, Shunsuke Kobayashi, Toshiaki Kogame, Satoru Unzai, Satoko Akashi, Shunichi Takeda, Toshiyuki Shimizu, Mamoru Sato,
Tópico(s)Genomics and Chromatin Dynamics
ResumoDNA polymerase ζ (Polζ) is an error-prone DNA polymerase involved in translesion DNA synthesis. Polζ consists of two subunits: the catalytic REV3, which belongs to B family DNA polymerase, and the noncatalytic REV7. REV7 also interacts with REV1 polymerase, which is an error-prone Y family DNA polymerase and is also involved in translesion DNA synthesis. Cells deficient in one of the three REV proteins and those deficient in all three proteins show similar phenotype, indicating the functional collaboration of the three REV proteins. REV7 interacts with both REV3 and REV1 polymerases, but the structure of REV7 or REV3, as well as the structural and functional basis of the REV1-REV7 and REV3-REV7 interactions, remains unknown. Here we show the first crystal structure of human REV7 in complex with a fragment of human REV3 polymerase (residues 1847–1898) and reveal the mechanism underlying REV7-REV3 interaction. The structure indicates that the interaction between REV7 and REV3 creates a structural interface for REV1 binding. Furthermore, we show that the REV7-mediated interactions are responsible for DNA damage tolerance. Our results highlight the function of REV7 as an adapter protein to recruit Polζ to a lesion site. REV7 is alternatively called MAD2B or MAD2L2 and also involved in various cellular functions such as signal transduction and cell cycle regulation. Our results will provide a general structural basis for understanding the REV7 interaction. DNA polymerase ζ (Polζ) is an error-prone DNA polymerase involved in translesion DNA synthesis. Polζ consists of two subunits: the catalytic REV3, which belongs to B family DNA polymerase, and the noncatalytic REV7. REV7 also interacts with REV1 polymerase, which is an error-prone Y family DNA polymerase and is also involved in translesion DNA synthesis. Cells deficient in one of the three REV proteins and those deficient in all three proteins show similar phenotype, indicating the functional collaboration of the three REV proteins. REV7 interacts with both REV3 and REV1 polymerases, but the structure of REV7 or REV3, as well as the structural and functional basis of the REV1-REV7 and REV3-REV7 interactions, remains unknown. Here we show the first crystal structure of human REV7 in complex with a fragment of human REV3 polymerase (residues 1847–1898) and reveal the mechanism underlying REV7-REV3 interaction. The structure indicates that the interaction between REV7 and REV3 creates a structural interface for REV1 binding. Furthermore, we show that the REV7-mediated interactions are responsible for DNA damage tolerance. Our results highlight the function of REV7 as an adapter protein to recruit Polζ to a lesion site. REV7 is alternatively called MAD2B or MAD2L2 and also involved in various cellular functions such as signal transduction and cell cycle regulation. Our results will provide a general structural basis for understanding the REV7 interaction. IntroductionLarge numbers of DNA lesions occur daily in every cell, and the majority of the DNA lesions stall replicative DNA polymerases. This results in the arrest of DNA replication, which causes lethal effects including genome instability and cell death. Translesion DNA synthesis (TLS) 2The abbreviations used are: TLStranslesion DNA synthesisPolpolymeraseWTwild typeGSTglutathione S-transferaseGFPgreen fluorescent protein. releases this replication blockage by replacing the stalled replicative polymerase with a DNA polymerase specialized for TLS (TLS polymerase). It is generally considered that TLS includes two steps performed by at least two types of TLS polymerases, namely inserter and extender polymerases (reviewed in Refs. 1Moldovan G.L. Pfander B. Jentsch S. Cell. 2007; 129: 665-679Abstract Full Text Full Text PDF PubMed Scopus (1307) Google Scholar and 2Friedberg E.C. Lehmann A.R. Fuchs R.P. Mol. Cell. 2005; 18: 499-505Abstract Full Text Full Text PDF PubMed Scopus (338) Google Scholar). In the first step, the stalled replicative polymerase is switched to an inserter polymerase such as Polη, Polκ, Polι, or REV1, which are classified as Y family DNA polymerases (3Ohmori H. Friedberg E.C. Fuchs R.P. Goodman M.F. Hanaoka F. Hinkle D. Kunkel T.A. Lawrence C.W. Livneh Z. Nohmi T. Prakash L. Prakash S. Todo T. Walker G.C. Wang Z. Woodgate R. Mol. Cell. 2001; 8: 7-8Abstract Full Text Full Text PDF PubMed Scopus (731) Google Scholar) and have different lesion specificity (reviewed in Refs. 4Vaisman A. Lehmann A.R. Woodgate R. Wei Y. Adv. Protein Chem. 2004; 69: 205-228Crossref PubMed Scopus (26) Google Scholar, 5Ohmori H. Ohashi E. Ogi T. Adv. Protein Chem. 2004; 69: 265-278Crossref PubMed Scopus (16) Google Scholar, 6Lawrence C.W. Adv. Protein Chem. 2004; 69: 167-203Crossref PubMed Scopus (116) Google Scholar, 7Prakash S. Johnson R.E. Prakash L. Annu. Rev. Biochem. 2005; 74: 317-353Crossref PubMed Scopus (820) Google Scholar, 8Guo C. Kosarek-Stancel J.N. Tang T.S. Friedberg E.C. Cell Mol. Life Sci. 2009; 66: 2363-2381Crossref PubMed Scopus (113) Google Scholar), and an inserter polymerase incorporates nucleotides opposite the DNA lesion instead of the stalled replicative polymerase. In the second step, an inserter polymerase is switched to the extender polymerase DNA polymerase ζ (Polζ), and then Polζ extends a few additional nucleotides before a replicative polymerase restarts DNA replication.Polζ consists of the catalytic REV3 and the noncatalytic REV7 subunits. REV3 is classified as a B family DNA polymerase on the basis of the primary sequence. The catalytic activity of yeast REV3 is stimulated by yeast REV7 (9Nelson J.R. Lawrence C.W. Hinkle D.C. Science. 1996; 272: 1646-1649Crossref PubMed Scopus (595) Google Scholar). Biochemical analysis has been done only for yeast REV3 but not mammalian REV3, because the molecular mass of human REV3 is larger (∼350 kDa) than that of yeast REV3 (∼150 kDa). Disruption of the mouse REV3 gene causes embryonic lethality accompanied by massive apoptosis (10Bemark M. Khamlichi A.A. Davies S.L. Neuberger M.S. Curr. Biol. 2000; 10: 1213-1216Abstract Full Text Full Text PDF PubMed Scopus (134) Google Scholar, 11Wittschieben J. Shivji M.K. Lalani E. Jacobs M.A. Marini F. Gearhart P.J. Rosewell I. Stamp G. Wood R.D. Curr. Biol. 2000; 10: 1217-1220Abstract Full Text Full Text PDF PubMed Scopus (152) Google Scholar, 12Esposito G. Godindagger I. Klein U. Yaspo M.L. Cumano A. Rajewsky K. Curr. Biol. 2000; 10: 1221-1224Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar), suggesting that the function of mammalian REV3 is essential for embryogenesis. Although REV7 is a smaller protein with a molecular mass of 24–28 kDa compared with REV3, the function of REV7 is less understood. REV7 is a member of the HORMA (Hop1, Rev7, and Mad2) family of proteins (13Aravind L. Koonin E.V. Trends Biochem. Sci. 1998; 23: 284-286Abstract Full Text Full Text PDF PubMed Scopus (159) Google Scholar). REV7, which is alternatively called MAD2B or MAD2L2, appears to be involved in multiple cellular functions including not only TLS but also cell cycle regulation (14Chen J. Fang G. Genes Dev. 2001; 15: 1765-1770Crossref PubMed Scopus (110) Google Scholar), bacterial infection (15Iwai H. Kim M. Yoshikawa Y. Ashida H. Ogawa M. Fujita Y. Muller D. Kirikae T. Jackson P.K. Kotani S. Sasakawa C. Cell. 2007; 130: 611-623Abstract Full Text Full Text PDF PubMed Scopus (122) Google Scholar), and signal transduction (16Zhang L. Yang S.H. Sharrocks A.D. Mol. Cell. Biol. 2007; 27: 2861-2869Crossref PubMed Scopus (35) Google Scholar, 17Hong C.F. Chou Y.T. Lin Y.S. Wu C.W. J. Biol. Chem. 2009; 284: 19613-19622Abstract Full Text Full Text PDF PubMed Scopus (33) Google Scholar).In this study, we investigated the function of REV7 in TLS from structural analysis. Previous studies have reported that human REV7 interacts with the central region (residues 1847–1892) of human REV3 by yeast two-hybrid and in vitro interaction assays (18Murakumo Y. Roth T. Ishii H. Rasio D. Numata S. Croce C.M. Fishel R. J. Biol. Chem. 2000; 275: 4391-4397Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar). Interestingly, human REV7 also interacts with the C-terminal region (residues 1130–1251) of human REV1 polymerase as shown by yeast two-hybrid, in vitro interaction and co-immunoprecipitation assays (19Murakumo Y. Ogura Y. Ishii H. Numata S. Ichihara M. Croce C.M. Fishel R. Takahashi M. J. Biol. Chem. 2001; 276: 35644-35651Abstract Full Text Full Text PDF PubMed Scopus (187) Google Scholar, 20Guo C. Fischhaber P.L. Luk-Paszyc M.J. Masuda Y. Zhou J. Kamiya K. Kisker C. Friedberg E.C. EMBO J. 2003; 22: 6621-6630Crossref PubMed Scopus (296) Google Scholar, 21Ohashi E. Murakumo Y. Kanjo N. Akagi J. Masutani C. Hanaoka F. Ohmori H. Genes Cells. 2004; 9: 523-531Crossref PubMed Scopus (222) Google Scholar). Furthermore, human REV7 and human REV1 were co-expressed by Escherichia coli, and the REV7-REV1 complex was purified, whereas REV7 does not affect the polymerase activity of REV1 (22Masuda Y. Ohmae M. Masuda K. Kamiya K. J. Biol. Chem. 2003; 278: 12356-12360Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar). The three yeast rev mutants and the triple mutant show very similar sensitivity to various genotoxic treatments (23Lawrence C.W. Christensen R.B. Genetics. 1979; 92: 397-408Crossref PubMed Google Scholar, 24Lawrence C.W. Christensen R.B. Mol. Gen. Genet. 1982; 186: 1-9Crossref PubMed Scopus (34) Google Scholar, 25Lawrence C.W. Das G. Christensen R.B. Mol. Gen. Genet. 1985; 200: 80-85Crossref PubMed Scopus (94) Google Scholar). Furthermore, chicken DT40 cells deficient in one of the three REV proteins and those deficient in all three proteins show hypersensitivity to various genotoxic treatment including cisplatin (cis-diaminedichloroplatinum (II)), indicating the functional collaboration of the three REV proteins (26Okada T. Sonoda E. Yoshimura M. Kawano Y. Saya H. Kohzaki M. Takeda S. Mol. Cell. Biol. 2005; 25: 6103-6111Crossref PubMed Scopus (96) Google Scholar). However, these previous analyses failed to determine the mechanism underlying the protein-protein interactions on the atomic revel, because they tried to analyze without data of the three-dimensional structures. In addition, it remains unclear whether mammalian REV1, REV3, and REV7 can form the Polζ-REV1 ternary complex. It has been considered that switching of DNA polymerase occurred at least twice in TLS: the switching from a stalled replicative polymerase to an inserter polymerase and from an inserter polymerase to the extender polymerase. Recently, the structural implications of the first polymerase switching have been reported (27Hishiki A. Hashimoto H. Hanafusa T. Kamei K. Ohashi E. Shimizu T. Ohmori H. Sato M. J. Biol. Chem. 2009; 284: 10552-10560Abstract Full Text Full Text PDF PubMed Scopus (112) Google Scholar). However, the mechanism underlying the recruitment of the extender polymerase to the lesion site and the second polymerase switching as well as the physical and functional interactions of REV1, REV3, and REV7 remains unclear. Here we report the first crystal structure of human REV7 in complex with a fragment of human REV3 (residues 1847–1898). The structure reveals the mechanism underlying Polζ formation and shows that the REV7-REV3 interaction unexpectedly provides a structural interface for REV1 binding. Furthermore, we show that these REV7-mediated interactions with REV1 and REV3 are responsible for DNA damage tolerance. Lastly, we propose a model of the structural interplay of REV1, REV3, and REV7 in TLS. Our results will provide a general structural basis for understanding the REV7 interaction in various cellular functions. IntroductionLarge numbers of DNA lesions occur daily in every cell, and the majority of the DNA lesions stall replicative DNA polymerases. This results in the arrest of DNA replication, which causes lethal effects including genome instability and cell death. Translesion DNA synthesis (TLS) 2The abbreviations used are: TLStranslesion DNA synthesisPolpolymeraseWTwild typeGSTglutathione S-transferaseGFPgreen fluorescent protein. releases this replication blockage by replacing the stalled replicative polymerase with a DNA polymerase specialized for TLS (TLS polymerase). It is generally considered that TLS includes two steps performed by at least two types of TLS polymerases, namely inserter and extender polymerases (reviewed in Refs. 1Moldovan G.L. Pfander B. Jentsch S. Cell. 2007; 129: 665-679Abstract Full Text Full Text PDF PubMed Scopus (1307) Google Scholar and 2Friedberg E.C. Lehmann A.R. Fuchs R.P. Mol. Cell. 2005; 18: 499-505Abstract Full Text Full Text PDF PubMed Scopus (338) Google Scholar). In the first step, the stalled replicative polymerase is switched to an inserter polymerase such as Polη, Polκ, Polι, or REV1, which are classified as Y family DNA polymerases (3Ohmori H. Friedberg E.C. Fuchs R.P. Goodman M.F. Hanaoka F. Hinkle D. Kunkel T.A. Lawrence C.W. Livneh Z. Nohmi T. Prakash L. Prakash S. Todo T. Walker G.C. Wang Z. Woodgate R. Mol. Cell. 2001; 8: 7-8Abstract Full Text Full Text PDF PubMed Scopus (731) Google Scholar) and have different lesion specificity (reviewed in Refs. 4Vaisman A. Lehmann A.R. Woodgate R. Wei Y. Adv. Protein Chem. 2004; 69: 205-228Crossref PubMed Scopus (26) Google Scholar, 5Ohmori H. Ohashi E. Ogi T. Adv. Protein Chem. 2004; 69: 265-278Crossref PubMed Scopus (16) Google Scholar, 6Lawrence C.W. Adv. Protein Chem. 2004; 69: 167-203Crossref PubMed Scopus (116) Google Scholar, 7Prakash S. Johnson R.E. Prakash L. Annu. Rev. Biochem. 2005; 74: 317-353Crossref PubMed Scopus (820) Google Scholar, 8Guo C. Kosarek-Stancel J.N. Tang T.S. Friedberg E.C. Cell Mol. Life Sci. 2009; 66: 2363-2381Crossref PubMed Scopus (113) Google Scholar), and an inserter polymerase incorporates nucleotides opposite the DNA lesion instead of the stalled replicative polymerase. In the second step, an inserter polymerase is switched to the extender polymerase DNA polymerase ζ (Polζ), and then Polζ extends a few additional nucleotides before a replicative polymerase restarts DNA replication.Polζ consists of the catalytic REV3 and the noncatalytic REV7 subunits. REV3 is classified as a B family DNA polymerase on the basis of the primary sequence. The catalytic activity of yeast REV3 is stimulated by yeast REV7 (9Nelson J.R. Lawrence C.W. Hinkle D.C. Science. 1996; 272: 1646-1649Crossref PubMed Scopus (595) Google Scholar). Biochemical analysis has been done only for yeast REV3 but not mammalian REV3, because the molecular mass of human REV3 is larger (∼350 kDa) than that of yeast REV3 (∼150 kDa). Disruption of the mouse REV3 gene causes embryonic lethality accompanied by massive apoptosis (10Bemark M. Khamlichi A.A. Davies S.L. Neuberger M.S. Curr. Biol. 2000; 10: 1213-1216Abstract Full Text Full Text PDF PubMed Scopus (134) Google Scholar, 11Wittschieben J. Shivji M.K. Lalani E. Jacobs M.A. Marini F. Gearhart P.J. Rosewell I. Stamp G. Wood R.D. Curr. Biol. 2000; 10: 1217-1220Abstract Full Text Full Text PDF PubMed Scopus (152) Google Scholar, 12Esposito G. Godindagger I. Klein U. Yaspo M.L. Cumano A. Rajewsky K. Curr. Biol. 2000; 10: 1221-1224Abstract Full Text Full Text PDF PubMed Scopus (147) Google Scholar), suggesting that the function of mammalian REV3 is essential for embryogenesis. Although REV7 is a smaller protein with a molecular mass of 24–28 kDa compared with REV3, the function of REV7 is less understood. REV7 is a member of the HORMA (Hop1, Rev7, and Mad2) family of proteins (13Aravind L. Koonin E.V. Trends Biochem. Sci. 1998; 23: 284-286Abstract Full Text Full Text PDF PubMed Scopus (159) Google Scholar). REV7, which is alternatively called MAD2B or MAD2L2, appears to be involved in multiple cellular functions including not only TLS but also cell cycle regulation (14Chen J. Fang G. Genes Dev. 2001; 15: 1765-1770Crossref PubMed Scopus (110) Google Scholar), bacterial infection (15Iwai H. Kim M. Yoshikawa Y. Ashida H. Ogawa M. Fujita Y. Muller D. Kirikae T. Jackson P.K. Kotani S. Sasakawa C. Cell. 2007; 130: 611-623Abstract Full Text Full Text PDF PubMed Scopus (122) Google Scholar), and signal transduction (16Zhang L. Yang S.H. Sharrocks A.D. Mol. Cell. Biol. 2007; 27: 2861-2869Crossref PubMed Scopus (35) Google Scholar, 17Hong C.F. Chou Y.T. Lin Y.S. Wu C.W. J. Biol. Chem. 2009; 284: 19613-19622Abstract Full Text Full Text PDF PubMed Scopus (33) Google Scholar).In this study, we investigated the function of REV7 in TLS from structural analysis. Previous studies have reported that human REV7 interacts with the central region (residues 1847–1892) of human REV3 by yeast two-hybrid and in vitro interaction assays (18Murakumo Y. Roth T. Ishii H. Rasio D. Numata S. Croce C.M. Fishel R. J. Biol. Chem. 2000; 275: 4391-4397Abstract Full Text Full Text PDF PubMed Scopus (160) Google Scholar). Interestingly, human REV7 also interacts with the C-terminal region (residues 1130–1251) of human REV1 polymerase as shown by yeast two-hybrid, in vitro interaction and co-immunoprecipitation assays (19Murakumo Y. Ogura Y. Ishii H. Numata S. Ichihara M. Croce C.M. Fishel R. Takahashi M. J. Biol. Chem. 2001; 276: 35644-35651Abstract Full Text Full Text PDF PubMed Scopus (187) Google Scholar, 20Guo C. Fischhaber P.L. Luk-Paszyc M.J. Masuda Y. Zhou J. Kamiya K. Kisker C. Friedberg E.C. EMBO J. 2003; 22: 6621-6630Crossref PubMed Scopus (296) Google Scholar, 21Ohashi E. Murakumo Y. Kanjo N. Akagi J. Masutani C. Hanaoka F. Ohmori H. Genes Cells. 2004; 9: 523-531Crossref PubMed Scopus (222) Google Scholar). Furthermore, human REV7 and human REV1 were co-expressed by Escherichia coli, and the REV7-REV1 complex was purified, whereas REV7 does not affect the polymerase activity of REV1 (22Masuda Y. Ohmae M. Masuda K. Kamiya K. J. Biol. Chem. 2003; 278: 12356-12360Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar). The three yeast rev mutants and the triple mutant show very similar sensitivity to various genotoxic treatments (23Lawrence C.W. Christensen R.B. Genetics. 1979; 92: 397-408Crossref PubMed Google Scholar, 24Lawrence C.W. Christensen R.B. Mol. Gen. Genet. 1982; 186: 1-9Crossref PubMed Scopus (34) Google Scholar, 25Lawrence C.W. Das G. Christensen R.B. Mol. Gen. Genet. 1985; 200: 80-85Crossref PubMed Scopus (94) Google Scholar). Furthermore, chicken DT40 cells deficient in one of the three REV proteins and those deficient in all three proteins show hypersensitivity to various genotoxic treatment including cisplatin (cis-diaminedichloroplatinum (II)), indicating the functional collaboration of the three REV proteins (26Okada T. Sonoda E. Yoshimura M. Kawano Y. Saya H. Kohzaki M. Takeda S. Mol. Cell. Biol. 2005; 25: 6103-6111Crossref PubMed Scopus (96) Google Scholar). However, these previous analyses failed to determine the mechanism underlying the protein-protein interactions on the atomic revel, because they tried to analyze without data of the three-dimensional structures. In addition, it remains unclear whether mammalian REV1, REV3, and REV7 can form the Polζ-REV1 ternary complex. It has been considered that switching of DNA polymerase occurred at least twice in TLS: the switching from a stalled replicative polymerase to an inserter polymerase and from an inserter polymerase to the extender polymerase. Recently, the structural implications of the first polymerase switching have been reported (27Hishiki A. Hashimoto H. Hanafusa T. Kamei K. Ohashi E. Shimizu T. Ohmori H. Sato M. J. Biol. Chem. 2009; 284: 10552-10560Abstract Full Text Full Text PDF PubMed Scopus (112) Google Scholar). However, the mechanism underlying the recruitment of the extender polymerase to the lesion site and the second polymerase switching as well as the physical and functional interactions of REV1, REV3, and REV7 remains unclear. Here we report the first crystal structure of human REV7 in complex with a fragment of human REV3 (residues 1847–1898). The structure reveals the mechanism underlying Polζ formation and shows that the REV7-REV3 interaction unexpectedly provides a structural interface for REV1 binding. Furthermore, we show that these REV7-mediated interactions with REV1 and REV3 are responsible for DNA damage tolerance. Lastly, we propose a model of the structural interplay of REV1, REV3, and REV7 in TLS. Our results will provide a general structural basis for understanding the REV7 interaction in various cellular functions.
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