Regulation of Rad51 Recombinase Presynaptic Filament Assembly via Interactions with the Rad52 Mediator and the Srs2 Anti-recombinase

Abstract

Homologous recombination represents an important means for the error-free elimination of DNA double-strand breaks and other deleterious DNA lesions from chromosomes. The Rad51 recombinase, a member of the RAD52 group of recombination proteins, catalyzes the homologous recombination reaction in the context of a helical protein polymer assembled on single-stranded DNA (ssDNA) that is derived from the nucleolytic processing of a primary lesion. The assembly of the Rad51-ssDNA nucleoprotein filament, often referred to as the presynaptic filament, is prone to interference by the single-strand DNA-binding factor replication protein A (RPA). The Saccharomyces cerevisiae Rad52 protein facilitates presynaptic filament assembly by helping to mediate the displacement of RPA from ssDNA. On the other hand, disruption of the presynaptic filament by the Srs2 helicase leads to a net exchange of Rad51 for RPA. To understand the significance of protein-protein interactions in the control of Rad52- or Srs2-mediated presynaptic filament assembly or disassembly, we have examined two rad51 mutants, rad51 Y388H and rad51 G393D, that are simultaneously ablated for Rad52 and Srs2 interactions and one, rad51 A320V, that is differentially inactivated for Rad52 binding for their biochemical properties and also for functional interactions with Rad52 or Srs2. We show that these mutant rad51 proteins are impervious to the mediator activity of Rad52 or the disruptive function of Srs2 in concordance with their protein interaction defects. Our results thus provide insights into the functional significance of the Rad51-Rad52 and Rad51-Srs2 complexes in the control of presynaptic filament assembly and disassembly. Moreover, our biochemical studies have helped identify A320V as a separation-of-function mutation in Rad51 with regards to a differential ablation of Rad52 interaction. Homologous recombination represents an important means for the error-free elimination of DNA double-strand breaks and other deleterious DNA lesions from chromosomes. The Rad51 recombinase, a member of the RAD52 group of recombination proteins, catalyzes the homologous recombination reaction in the context of a helical protein polymer assembled on single-stranded DNA (ssDNA) that is derived from the nucleolytic processing of a primary lesion. The assembly of the Rad51-ssDNA nucleoprotein filament, often referred to as the presynaptic filament, is prone to interference by the single-strand DNA-binding factor replication protein A (RPA). The Saccharomyces cerevisiae Rad52 protein facilitates presynaptic filament assembly by helping to mediate the displacement of RPA from ssDNA. On the other hand, disruption of the presynaptic filament by the Srs2 helicase leads to a net exchange of Rad51 for RPA. To understand the significance of protein-protein interactions in the control of Rad52- or Srs2-mediated presynaptic filament assembly or disassembly, we have examined two rad51 mutants, rad51 Y388H and rad51 G393D, that are simultaneously ablated for Rad52 and Srs2 interactions and one, rad51 A320V, that is differentially inactivated for Rad52 binding for their biochemical properties and also for functional interactions with Rad52 or Srs2. We show that these mutant rad51 proteins are impervious to the mediator activity of Rad52 or the disruptive function of Srs2 in concordance with their protein interaction defects. Our results thus provide insights into the functional significance of the Rad51-Rad52 and Rad51-Srs2 complexes in the control of presynaptic filament assembly and disassembly. Moreover, our biochemical studies have helped identify A320V as a separation-of-function mutation in Rad51 with regards to a differential ablation of Rad52 interaction. Regulation of Rad51 recombinase presynaptic filament assembly via interactions with the Rad52 mediator and the Srs2 anti-recombinase.Journal of Biological ChemistryVol. 287Issue 15PreviewVOLUME 284 (2009) PAGES 24363–24371 Full-Text PDF Open Access Homologous recombination (HR) 3The abbreviations used are: HRhomologous recombinationRPAreplication protein Asssingle-strandeddsdouble-strandedNi-NTAnickel-nitrilotriacetic acidDTTdithiothreitol. helps maintain genomic stability by eliminating DNA double-strand breaks induced by ionizing radiation and chemical reagents, by restarting damaged or collapsed DNA replication forks, and by elongating shortened telomeres especially when telomerase is dysfunctional (1.Symington L.S. Microbiol. Mol. Biol. Rev. 2002; 66: 630-670Crossref PubMed Scopus (820) Google Scholar, 2.Sung P. Klein H. Nat. Rev. Mol. Cell Biol. 2006; 7: 739-750Crossref PubMed Scopus (499) Google Scholar, 3.San Filippo J. Sung P. Klein H. Annu. Rev. Biochem. 2008; 77: 229-257Crossref PubMed Scopus (1188) Google Scholar). Accordingly, defects in HR invariably lead to enhanced sensitivity to genotoxic agents, chromosome aberrations, and tumor development (4.Jasin M. Oncogene. 2002; 21: 8981-8993Crossref PubMed Scopus (296) Google Scholar, 5.D'Andrea A.D. Grompe M. Nat. Rev. Cancer. 2003; 3: 23-34Crossref PubMed Scopus (672) Google Scholar). In meiosis also, HR helps mediate the linkage of homologous chromosome pairs via arm cross-overs, thus ensuring the proper segregation of chromosomes at the first meiotic division (6.Bishop D.K. Zickler D. Cell. 2004; 117: 9-15Abstract Full Text Full Text PDF PubMed Scopus (273) Google Scholar). Accordingly, HR mutants exhibit a plethora of meiotic defects, including early meiotic cell cycle arrest, aneuploidy, and inviability. homologous recombination replication protein A single-stranded double-stranded nickel-nitrilotriacetic acid dithiothreitol. Much of the knowledge regarding the mechanistic basis of HR has been derived from studies of model organisms, such as the budding yeast Saccharomyces cerevisiae. Genetic analyses in S. cerevisiae have led to the identification of the RAD52 group of genes, namely, RAD50, RAD51, RAD52, RAD54, RAD55, RAD57, RAD59, RDH54, MRE11, and XRS2 (1.Symington L.S. Microbiol. Mol. Biol. Rev. 2002; 66: 630-670Crossref PubMed Scopus (820) Google Scholar), that are needed for the successful execution of HR. Each member of the RAD52 group of genes has an orthologue in higher eukaryotes, including humans, and mutations in any of these genes cause defects in HR and repair of double-strand breaks. The DNA pairing and strand invasion step of the HR reaction is mediated by RAD51-encoded protein, which is orthologous to the Escherichia coli recombinase RecA (2.Sung P. Klein H. Nat. Rev. Mol. Cell Biol. 2006; 7: 739-750Crossref PubMed Scopus (499) Google Scholar). Like RecA, Rad51 polymerizes on ssDNA, derived from the nucleolytic processing of a primary lesion such as a double-strand break, to form a right-handed nucleoprotein filament, often referred to as the presynaptic filament (3.San Filippo J. Sung P. Klein H. Annu. Rev. Biochem. 2008; 77: 229-257Crossref PubMed Scopus (1188) Google Scholar, 7.Bianco P.R. Tracy R.B. Kowalczykowski S.C. Front. Biosci. 1998; 3: D570-D603Crossref PubMed Google Scholar). The presynaptic filament engages dsDNA, conducts a search for homology in the latter, and catalyzes DNA joint formation between the recombining ssDNA and dsDNA partners upon the location of homology (1.Symington L.S. Microbiol. Mol. Biol. Rev. 2002; 66: 630-670Crossref PubMed Scopus (820) Google Scholar, 3.San Filippo J. Sung P. Klein H. Annu. Rev. Biochem. 2008; 77: 229-257Crossref PubMed Scopus (1188) Google Scholar). As such, the timely and efficient assembly of the presynaptic filament is indispensable for the successful execution of HR. Because the nucleation of Rad51 onto ssDNA is a rate-limiting process, presynaptic filament assembly is prone to interference by the single-strand DNA-binding protein replication protein A (RPA) (1.Symington L.S. Microbiol. Mol. Biol. Rev. 2002; 66: 630-670Crossref PubMed Scopus (820) Google Scholar, 3.San Filippo J. Sung P. Klein H. Annu. Rev. Biochem. 2008; 77: 229-257Crossref PubMed Scopus (1188) Google Scholar, 7.Bianco P.R. Tracy R.B. Kowalczykowski S.C. Front. Biosci. 1998; 3: D570-D603Crossref PubMed Google Scholar). In reconstituted biochemical systems, the addition of Rad52 counteracts the inhibitory action of RPA (8.New J.H. Sugiyama T. Zaitseva E. Kowalczykowski S.C. Nature. 1998; 391: 407-410Crossref PubMed Scopus (501) Google Scholar, 9.Shinohara A. Ogawa T. Nature. 1998; 391: 404-407Crossref PubMed Scopus (408) Google Scholar). Consistent with the biochemical results, in both mitotic and meiotic cells, the recruitment of Rad51 to double-strand breaks is strongly dependent on Rad52 (10.Sugawara N. Wang X. Haber J.E. Mol. Cell. 2003; 12: 209-219Abstract Full Text Full Text PDF PubMed Scopus (296) Google Scholar, 11.Wolner B. van Komen S. Sung P. Peterson C.L. Mol. Cell. 2003; 12: 221-232Abstract Full Text Full Text PDF PubMed Scopus (164) Google Scholar, 12.Lisby M. Barlow J.H. Burgess R.C. Rothstein R. Cell. 2004; 118: 699-713Abstract Full Text Full Text PDF PubMed Scopus (738) Google Scholar). This effect of Rad52 on Rad51 presynaptic filament assembly has been termed a “recombination mediator” function (13.Sung P. Krejci L. Van Komen S. Sehorn M.G. J. Biol. Chem. 2003; 278: 42729-42732Abstract Full Text Full Text PDF PubMed Scopus (343) Google Scholar). Interestingly, genetic studies have shown that the Srs2 helicase fulfills the role of an anti-recombinase. Specifically, mutations in Srs2 often engender a hyper-recombinational phenotype and can also help suppress the DNA damage sensitivity of rad6 and rad18 mutants, because of the heightened HR proficiency being able to substitute for the post-replicative DNA repair defects of these mutant cells (2.Sung P. Klein H. Nat. Rev. Mol. Cell Biol. 2006; 7: 739-750Crossref PubMed Scopus (499) Google Scholar, 14.Schiestl R.H. Prakash S. Prakash L. Genetics. 1990; 124: 817-831Crossref PubMed Google Scholar). Importantly, in reconstituted systems, Srs2 exerts a strong inhibitory effect on Rad51-mediated reactions in a manner that is potentiated by RPA. Biochemical and electron microscopic analyses have provided compelling evidence that Srs2 acts by disassembling the presynaptic filament, to effect the replacement of Rad51 by RPA (15.Veaute X. Jeusset J. Soustelle C. Kowalczykowski S.C. Le Cam E. Fabre F. Nature. 2003; 423: 309-312Crossref PubMed Scopus (489) Google Scholar, 16.Krejci L. Van Komen S. Li Y. Villemain J. Reddy M.S. Klein H. Ellenberger T. Sung P. Nature. 2003; 423: 305-309Crossref PubMed Scopus (505) Google Scholar). The ability of Srs2 to dissociate the presynaptic filament relies on its ATPase activity, revealed using mutant variants, K41A and K41R, that harbor changes in the Walker A in Accordingly, the K41A and mutants are L. M. Li Y. Van Komen S. Villemain J. Ellenberger T. Klein H. Sung P. J. Biol. Chem. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar). In both yeast and biochemical a of Rad51 with Rad52 or Srs2 can (1.Symington L.S. Microbiol. Mol. Biol. Rev. 2002; 66: 630-670Crossref PubMed Scopus (820) Google Scholar, 16.Krejci L. Van Komen S. Li Y. Villemain J. Reddy M.S. Klein H. Ellenberger T. Sung P. Nature. 2003; 423: 305-309Crossref PubMed Scopus (505) Google Scholar). yeast rad51 mutant A320V, and G393D, that engender a in the yeast with Rad52 have been L. J. B. M. C. Mol. Cell. Biol. 21: PubMed Scopus Google Scholar). we our biochemical studies the of these rad51 mutant proteins to and with Interestingly, we that two of these rad51 mutants, namely, Y388H and G393D, are also in Srs2 interaction. Accordingly, these mutant rad51 proteins form presynaptic that are to the disruptive action of Srs2. Our results thus the role of Rad51-Rad52 and Rad51-Srs2 interactions in the of Rad51 presynaptic filament assembly and and also the of Rad52 and Srs2 interaction in In these our biochemical studies have the A320V as a separation-of-function mutation in of the in protein at The rad51 A320V, rad51 and rad51 proteins in yeast cells and to as for the P. Cell. Full Text PDF PubMed Scopus Google Scholar). Rad52 with a at its and Srs2 with a at its in E. coli and to as L. Van Komen S. Li Y. Villemain J. Reddy M.S. Klein H. Ellenberger T. Sung P. Nature. 2003; 423: 305-309Crossref PubMed Scopus (505) Google Scholar, C. Sehorn M.G. B. P. Sung P. Krejci L. J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar). and a at its from cells, as M. Van Komen S. P. Ellenberger T. Sung P. J. Biol. Chem. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar, P. Y. C. A. Sung P. Klein J. Biol. Chem. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). the and of a DNA acid of Rad52 into cells at to at which The for at and the cells by Cell from of cells to of by the cell in of cell and and for The onto a and the and onto a The proteins with a from to in and The onto a and with a from to in The onto and with a from to in The and to with The of Rad51 or rad51 mutant with Rad52 or Srs2 at for in of To the effect of Rad52 on Rad51-Srs2 Rad51 or rad51 A320V with or and Srs2 as The reactions at for with of which the on or to protein the with of the the proteins with The and by and with The form DNA and strand DNA from The dsDNA by the form DNA with the DNA by the form DNA with as Van Komen S. S. Klein H. Sung P. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). the DNA the with and The to its by the two at for by to The DNA from a as P. Y. C. A. Sung P. Klein J. Biol. Chem. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar). the a to of DNA used Komen S. S. S. Sung P. Mol. Cell. Full Text Full Text PDF PubMed Scopus Google Scholar). The used as DNA in the electron microscopic of presynaptic filament Rad51 or rad51 mutant with strand DNA in of at A of the reaction at the and with an of to the and used to the of as S. Sung P. Nature. 1998; PubMed Scopus Google Scholar). The ssDNA or dsDNA with the of Rad51 or rad51 mutant in of for at The reaction in in The on a of and to To DNA induced by Rad51 or the rad51 mutants, the of the or mutant protein with dsDNA for at in of by the of of in and a To the disruption of the presynaptic filament by Rad51 or rad51 mutant with ssDNA for with an and in of by the of RPA in and Srs2 or in and a dsDNA and of in and by an The reaction with and for at being on in DNA with and the results in a of the reaction at the the of Rad51 or rad51 mutant with ssDNA in of at for the addition of RPA in dsDNA in and in into the by a the reaction by and as To the of the rad51 Rad51 or rad51 mutant RPA and the of Rad52 on for in ssDNA and a these reactions and in the manner as The with Rad51 or rad51 mutant for at in of and the to the Rad51-ssDNA nucleoprotein filament, by the of RPA in and Srs2 or in and a at in and the reaction for at DNA pairing by form DNA in and the reaction at for The reaction and to as and the and to To the presynaptic filament, the and Rad51 or rad51 for in of and an at the addition of RPA in and Srs2 in the reaction for at a with a of the reaction to a with which been in The with for and with The examined with a electron with a filament at with a at a of A namely, A320V, and G393D, that the interaction of Rad51 with Rad52 with or in the yeast L. J. B. M. C. Mol. Cell. Biol. 21: PubMed Scopus Google Scholar). their biochemical we the rad51 mutant proteins the in yeast cells for the and these proteins to using the for the protein P. Cell. Full Text PDF PubMed Scopus Google Scholar). The mutants to the as the protein, and mutants the protein Moreover, a of these mutants to that of the protein the rad51 mutants for their and functional interactions with Rad52 and we to first that the biochemical of the protein with regards to DNA and the ability to mediate homologous DNA pairing and strand exchange in with RPA. the ATPase activity with DNA as and the results that rad51 mutant proteins a of ATPase activity with that of the protein a DNA with ssDNA or dsDNA as we also that of the rad51 mutants has any DNA binding of the of Rad51 is the ability to form helical nucleoprotein on The of nucleoprotein filament formation can by a The are that rad51 mutants are as as the protein in the formation of We the rad51 mutant proteins for their ability to the homologous DNA pairing and DNA strand exchange In the in ssDNA is first with Rad51 to for presynaptic filament and RPA by homologous dsDNA are to the these RPA the of the by helping in the ssDNA and also the ssDNA as a of DNA strand As shown in rad51 mutants the of reaction as rad51 mutants are as as Rad51 in the of homologous DNA pairing and strand the results from the analyses as to that the rad51 A320V, and mutants biochemical with that of We used to the rad51 mutants are in interaction with Rad52 with a at its the rad51 mutants and Rad51 with and any protein that on which the on proteins with and of rad51 mutants with as a of Rad51 with Rad52 Rad51 any of the rad51 mutants the in the of Rad52 rad51 mutants are in formation with In rad51 mutants the of ability to with and Because of the ability of RPA to with Rad51 for on ssDNA, of RPA with Rad51 to the ssDNA strongly the homologous DNA pairing and strand exchange Importantly, the reaction can by the of an of Rad52 to that of Rad51 C. Sehorn M.G. B. P. Sung P. Krejci L. J. Biol. Chem. 2008; Full Text Full Text PDF PubMed Scopus Google Scholar). We used the to for the of the rad51 mutant proteins to As RPA strongly the of homologous pairing and strand exchange mediated by Rad51 or any of the mutants with and Importantly, Rad52 able to help the inhibitory action of RPA with Rad51 and with at a Rad51 to Rad52 of and the by RPA at by Rad52 when any of the rad51 mutant proteins examined and results thus provide evidence that interaction with Rad51 is for the of Rad52 recombination mediator As and L. J. B. M. C. Mol. Cell. Biol. 21: PubMed Scopus Google Scholar), the rad51 A320V, rad51 and rad51 mutants are in interaction with and are in Rad52 We these rad51 mutants the ability to with Srs2. To Rad51 and rad51 mutants with Srs2 and with to protein which from the with and Rad51 with Srs2 as L. Van Komen S. Li Y. Villemain J. Reddy M.S. Klein H. Ellenberger T. Sung P. Nature. 2003; 423: 305-309Crossref PubMed Scopus (505) Google Scholar). Interestingly, rad51 A320V also rad51 Y388H or rad51 with Srs2 Specifically, of the Rad51 or rad51 A320V with of rad51 Y388H or of rad51 by Srs2 the As Rad51 any of the rad51 mutants on the Srs2 thus revealed an Srs2 interaction in the rad51 Y388H and rad51 proteins and provide evidence that the rad51 A320V mutation differentially Rad52 interaction. As an Srs2 with Rad51-mediated homologous DNA pairing and strand exchange by the presynaptic filament L. Van Komen S. Li Y. Villemain J. Reddy M.S. Klein H. Ellenberger T. Sung P. Nature. 2003; 423: 305-309Crossref PubMed Scopus (505) Google Scholar). The of the rad51 Y388H and rad51 mutant proteins that the functional of Rad51 are in Srs2 interaction the to the functional significance of the Rad51-Srs2 in the function of Srs2. Because Srs2 exerts a strong effect on the reaction mediated by the protein L. Van Komen S. Li Y. Villemain J. Reddy M.S. Klein H. Ellenberger T. Sung P. Nature. 2003; 423: 305-309Crossref PubMed Scopus (505) Google Scholar), we the reaction mediated by rad51 A320V, rad51 or rad51 in with is also prone to the inhibitory action of Srs2. Consistent with their proficiency in interaction rad51 mutants a of with that with Rad51 the rad51 reaction to a as in the of with rad51 Y388H or rad51 results thus revealed a of Srs2 function on its ability to with Rad51 We a to the of presynaptic of the rad51 A320V, rad51 and rad51 mutants to Srs2. In Rad51 from ssDNA by Srs2 is on dsDNA, to of the DNA that can as a DNA upon with The dsDNA is by L. Van Komen S. Li Y. Villemain J. Reddy M.S. Klein H. Ellenberger T. Sung P. Nature. 2003; 423: 305-309Crossref PubMed Scopus (505) Google Scholar). We that the rad51 mutants are as as the protein in the when to to dsDNA and and In the presynaptic by the mutant proteins are in the of Srs2 and The addition of Srs2 to the Rad51 presynaptic filament induced the of and the displacement of Rad51 from the ssDNA and of the Rad51 on the when Srs2 with presynaptic that rad51 A320V and In a of DNA when presynaptic of rad51 Y388H or rad51 with Srs2 and results provide evidence that disruption of Rad51 nucleoprotein filament by Srs2 the interaction of Rad51 with Srs2. to the effect of Srs2 on the Rad51 or rad51 presynaptic with the that the to Srs2. of Rad51 or rad51 with a presynaptic with the and and complexes with a and As the addition of Srs2 to Rad51 presynaptic led to their such that the complexes the of nucleoprotein Importantly, the presynaptic of rad51 to the disruptive action of because a in their upon the of Srs2 these results also the that the of presynaptic filament by Srs2 is dependent on the interaction between Rad51 and Srs2. Because rad51 Y388H and rad51 are simultaneously for Rad52 and Srs2 that the in Rad51 that mediate its interaction with Rad52 and Srs2 the Rad52 with Srs2 for Rad51 To Srs2 on with Rad51 and an of a Rad52 termed the L. B. Rothstein R. Sung P. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar, 7: PubMed Scopus Google Scholar). the with proteins with and to the of As shown in on the upon the of and effect dependent on the of Importantly, the reaction the with Srs2 and rad51 A320V which we have shown to in Rad52 interaction to the of the results from the the that the Rad52 and Srs2 interaction in Rad51 also provide that the of in Rad51 to represents a separation-of-function mutation with regards to the differential of Rad52 interaction. The inhibitory effect of RPA on Rad51 presynaptic filament assembly can by Rad52 (3.San Filippo J. Sung P. Klein H. Annu. Rev. Biochem. 2008; 77: 229-257Crossref PubMed Scopus (1188) Google Scholar). including yeast and biochemical yeast Rad51 and Rad52 have been shown to form a (1.Symington L.S. Microbiol. Mol. Biol. Rev. 2002; 66: 630-670Crossref PubMed Scopus (820) Google Scholar, 3.San Filippo J. Sung P. Klein H. Annu. Rev. Biochem. 2008; 77: 229-257Crossref PubMed Scopus (1188) Google Scholar). The Rad51 interaction the of Rad52 7: PubMed Scopus Google Scholar), and using of Rad52 that the Rad51 interaction we and have provided evidence for a role of the Rad51-Rad52 in the of Rad51 to ssDNA to the assembly of the presynaptic filament A. Ogawa T. Nature. 1998; 391: 404-407Crossref PubMed Scopus (408) Google Scholar, L. B. Rothstein R. Sung P. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). To the of the Rad51-Rad52 in presynaptic filament we have and rad51 mutants that harbor a acid namely, rad51 A320V, rad51 and rad51 G393D, shown to for Rad52 interaction by the yeast L. J. B. M. C. Mol. Cell. Biol. 21: PubMed Scopus Google Scholar). We have provided of evidence that these rad51 mutants biochemical with regards to DNA and the ability to homologous DNA pairing and strand we have that rad51 mutants are in formation with the ability to with other recombination including and Consistent with their in Rad52 of the rad51 mutants is to the recombination mediator activity of our analyses with these rad51 mutants with studies the of Rad51 interaction of Rad52 A. Ogawa T. Nature. 1998; 391: 404-407Crossref PubMed Scopus (408) Google Scholar, L. B. Rothstein R. Sung P. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google provide for the that interaction with Rad51 is indispensable for the recombination mediator of of leads to of recombination including cell cycle arrest, and of in with other mutations L. M. Li Y. Van Komen S. Villemain J. Ellenberger T. Klein H. Sung P. J. Biol. Chem. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar, A. Klein Genetics. PubMed Google Scholar, D. Genetics. PubMed Google Scholar, S. Soustelle C. Fabre F. Nat. PubMed Scopus Google Scholar). in the have that Srs2 the role of an to or HR via the disruption of the Rad51 presynaptic (15.Veaute X. Jeusset J. Soustelle C. Kowalczykowski S.C. Le Cam E. Fabre F. Nature. 2003; 423: 309-312Crossref PubMed Scopus (489) Google Scholar, 16.Krejci L. Van Komen S. Li Y. Villemain J. Reddy M.S. Klein H. Ellenberger T. Sung P. Nature. 2003; 423: 305-309Crossref PubMed Scopus (505) Google Scholar). The ssDNA of Rad51 via Srs2 action by thus the DNA for Rad51 L. Van Komen S. Li Y. Villemain J. Reddy M.S. Klein H. Ellenberger T. Sung P. Nature. 2003; 423: 305-309Crossref PubMed Scopus (505) Google Scholar). In yeast and biochemical Srs2 a with and we have the Rad51 interaction to the of Srs2 L. Van Komen S. Li Y. Villemain J. Reddy M.S. Klein H. Ellenberger T. Sung P. Nature. 2003; 423: 305-309Crossref PubMed Scopus (505) Google Scholar). we that rad51 Y388H and rad51 G393D, rad51 A320V, are for Srs2 interaction. these rad51 mutant proteins, we have evidence derived from biochemical and electron microscopic analyses to the that the activity of Srs2 relies on formation with the Rad51 The interaction rad51 Y388H and rad51 mutants are to the activity of Srs2 is Rad51 or the interaction mutant rad51 Consistent with the that the Rad52 and Srs2 interactions in we have biochemical results that Srs2 is from with Rad51 when an of Rad52 is the that the rad51 A320V mutation differentially the ability to with Rad52 that the Rad52 an with Rad51 protein that is by Srs2. the rad51 A320V mutation the of Rad51 in such a as to Rad52 interaction Srs2 the of the rad51 mutations onto the of the yeast Rad51 filament We that and are on the of the Rad51 helical is in the of the We also that and are Rad51 including Rad51 to these to the of Rad51 proteins from other with their mediator proteins, such as the protein in cells, and the of the Rad51 with proteins, such as the helicase in cells H. Li J. Nature. PubMed Scopus Google Scholar, Filippo J. P. Sehorn M.G. J. Krejci L. Sung P. J. Biol. Chem. 2006; Full Text Full Text PDF PubMed Scopus Google Scholar).