The <i>rad51-K191R</i> ATPase-Defective Mutant Is Impaired forPresynaptic Filament Formation

Fung, Cindy W.; Fortin, Gary S; Peterson, Shaun; Symington, Lorraine S.

doi:10.1128/mcb.00599-06

Cited by 42 publications

(49 citation statements)

References 65 publications

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“…However, in contrast to the weak suppression of the UV sensitivity of the rad55 mutant by MAT heterozygosity, expression of both mating-type alleles in the rad55 mutant rescued YFP-Rad51 formation in response to UV (P ¼ 0.0001). As reported previously, the Rad51 foci formed in response to UV or IR in the rad55 mutant were less bright than those observed in wild-type cells (Fung et al 2006). The dimmer foci suggest Rad51 is still able to nucleate in the absence of the Rad51 paralogs, but is unable to form extensive filaments or the filaments are less stable.…”

Section: Resultsmentioning

confidence: 49%

“…Consistent with a role in Rad51 recruitment, Rad51 foci are not observed in rad55 or rad57 mutants during meiosis (Gasior et al 1998). However, Rad51 is still able to associate with double-strand breaks (DSBs) in rad55 mutants during vegetative growth although recruitment of Rad51 is slower and less extensive in rad55 mutants than in wild type (Sugawara et al 2003;Lisby et al 2004;Fung et al 2006). The role of the Rad51 paralogs as accessory proteins for Rad51 is also supported by the observation that overexpression of RAD51 partially suppresses the radiation or mitomycin C sensitivity of cell lines with mutations in any of the Rad51 paralog-encoding genes (Hays et al 1995;Johnson and 1 Symington 1995;Takata et al 2001).…”

mentioning

confidence: 73%

“…The IR sensitivity of rad55 or rad57 mutants is also suppressed by expression of both mating-type alleles in haploids. It has been suggested that this suppression acts at the level of Rad51 activity because MAT heterozygosity, deletion of SRS2, or overexpression of Rad51 also suppresses other mutants known to have mediator defects, such as rad51-K191R (Fung et al 2006) and rad52-20 (Schild 1995), but does not suppress rad51 or rad52 null mutants. In budding and fission yeasts, rad55 or rad57 null mutants exhibit cold sensitivity for DSB repair (DSBR) (Symington 2002).…”

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confidence: 99%

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Role of the Saccharomyces cerevisiae Rad51 Paralogs in Sister Chromatid Recombination

2008

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Rad51 requires a number of other proteins, including the Rad51 paralogs, for efficient recombination in vivo. Current evidence suggests that the yeast Rad51 paralogs, Rad55 and Rad57, are important in formation or stabilization of the Rad51 nucleoprotein filament. To gain further insights into the function of the Rad51 paralogs, reporters were designed to measure spontaneous or double-strand break (DSB)-induced sister or nonsister recombination. Spontaneous sister chromatid recombination (SCR) was reduced 6000-fold in the rad57 mutant, significantly more than in the rad51 mutant. Although the DSBinduced recombination defect of rad57 was suppressed by overexpression of Rad51, elevated temperature, or expression of both mating-type alleles, the rad57 defect in spontaneous SCR was not strongly suppressed by these same factors. In addition, the UV sensitivity of the rad57 mutant was not strongly suppressed by MAT heterozygosity, even though Rad51 foci were restored under these conditions. This lack of suppression suggests that Rad55 and Rad57 have different roles in the recombinational repair of stalled replication forks compared with DSB repair. Furthermore, these data suggest that most spontaneous SCR initiates from single-stranded gaps formed at stalled replication forks rather than DSBs.

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Section: Resultsmentioning

confidence: 49%

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confidence: 73%

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confidence: 99%

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Role of the Saccharomyces cerevisiae Rad51 Paralogs in Sister Chromatid Recombination

2008

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“…or by stabilizing the filament once it is assembled. In rad55 mutants, Rad51 is recruited to DSBs with slower kinetics and forms dimmer IR-induced foci compared to wild-type cells (Sugawara et al 2003;Lisby et al 2004;Fung et al 2006). The IR sensitivity of rad55 and rad57 mutants is partially bypassed by overexpression of Rad51, which increases the availability of the protein for filament formation, or by RAD51 gain-of-function alleles, such as rad51-I345T, that encode proteins with a higher affinity for DNA than wild-type Rad51 (Hays et al 1995;Johnson and Symington 1995;Fortin and Symington 2002;Malik and Symington 2008).…”

Section: H Omologous Recombination Is Required Formentioning

confidence: 99%

Suppression of the Double-Strand-Break-Repair Defect of the Saccharomyces cerevisiae rad57 Mutant

Fung¹,

Mozlin²,

Symington

2009

Genetics

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The Rad51 paralogs Rad55 and Rad57 form a heterodimer required to mediate the formation and/or stabilization of the Rad51 filament. To further characterize the function of Rad55-Rad57, we used a combination of rad57 partial suppressors to determine whether the DNA repair and recombination defects of the rad57 mutant could be completely suppressed. The combination of all suppressors, elevated temperature, srs2, rad51-I345T, and mating-type (MAT) heterozygosity resulted in almost complete suppression of the rad57 mutant defect in the recruitment of Rad51 to DNA-damaged sites, as well as survival in response to ionizing radiation and camptothecin. In a physical assay to monitor the kinetics of double-strand-break (DSB)-induced gene conversion, the rad57 mutant defect was effectively suppressed by srs2 and MAT heterozygosity, but these same suppressors failed to suppress the spontaneous recombination defect. Thus the Rad55-Rad57 heterodimer appears to have a unique function in spontaneous recombination that is not essential for DSB repair. Furthermore, we investigated the currently unknown mechanism of rad57 suppression by MAT heterozygosity and found that it is independent of DNL4. H OMOLOGOUS recombination is required forthe faithful repair of DNA double-strand breaks (DSBs) that arise during normal cellular processes or from exposure of cells to DNA-damaging agents. Central to the process of homologous recombination is the Rad51 protein, which facilitates synapsis and strand invasion into homologous duplex DNA (San Filippo et al. 2008). Rad51 belongs to the RecA family of homologous pairing proteins (Aboussekhra et al. 1992;Basile et al. 1992;Shinohara et al. 1992). Yeast and humans have two RecA homologs: Rad51 and the meiosis-specific Dmc1 (Bishop et al. 1992;San Filippo et al. 2008). In addition, the Saccharomyces cerevisiae RAD55 and RAD57 genes encode proteins with sequence similarity to RecA and Rad51 and are considered to be Rad51 paralogs (Kans and Mortimer 1991;Lovett 1994). Mutation of RAD51, RAD55, or RAD57 confers sensitivity of ionizing radiation (IR) and defects in mitotic and meiotic recombination, indicating that their functions are not redundant (Symington 2002). rad51 mutants generally exhibit more severe defects than rad55 or rad57 mutants in DSB-induced recombination assays; however, rad55 and rad57 mutants are more defective than rad51 in some assays that measure spontaneous recombination between repeated sequences (Rattray and Symington 1995;Mozlin et al. 2008).The molecular details of homologous recombination are largely based on genetic, physical, and cytological studies of DSB repair (DSBR) and on biochemical characterization of purified proteins (Paques and Haber 1999;Sugawara et al. 2003;Lisby et al. 2004;San Filippo et al. 2008). The single-stranded DNA (ssDNA)-binding protein, replication protein A (RPA), initially binds ssDNA that forms by nucleolytic processing of DNA ends at DSBs. In vitro, RPA has been shown to be inhibitory to Rad51 binding to ssDNA, but this inhibition c...

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“…Mutations in functional regions of Rad51 decrease efficiency of double strand break repair and increase lethality 10,11 . In vitro, isolated Rad51 is capable of catalyzing strand exchange in the absence of recombination mediators.…”

Section: Nucleoprotein Filament: Structure and Functionmentioning

confidence: 99%

Homologous recombination: Single-molecule experiments and their lessons for the in vivo situation

Franker¹

2016

JOSHA

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The rad51-K191R ATPase-Defective Mutant Is Impaired forPresynaptic Filament Formation

Cited by 42 publications

References 65 publications

Role of the Saccharomyces cerevisiae Rad51 Paralogs in Sister Chromatid Recombination

Role of the Saccharomyces cerevisiae Rad51 Paralogs in Sister Chromatid Recombination

Suppression of the Double-Strand-Break-Repair Defect of the Saccharomyces cerevisiae rad57 Mutant

Homologous recombination: Single-molecule experiments and their lessons for the in vivo situation

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