A Homolog ofScRAD5Is Involved in DNA Repair and Homologous Recombination in Arabidopsis  

Abstract

Rad5 is the key component in the Rad5-dependent error-free branch of postreplication repair in yeast (Saccharomyces cerevisiae). Rad5 is a member of the Snf2 ATPase/helicase family, possessing as a characteristic feature, a RING-finger domain embedded in the Snf2-helicase domain and a HIRAN domain. Yeast mutants are sensitive to DNA-damaging agents and reveal differences in homologous recombination. By sequence comparisons we were able to identify two homologs (AtRAD5a and AtRAD5b) in the Arabidopsis thaliana genome, sharing about 30% identity and 45% similarity to yeast Rad5. AtRad5a and AtRad5b have the same kind of domain organization with a higher degree of similarity to each other than to ScRad5. Surprisingly, both genes differ in function: whereas two independent mutants of Atrad5a are hypersensitive to the cross-linking agents mitomycin C and cis-platin and to a lesser extent to the methylating agent, methyl methane sulfonate, the Atrad5b mutants did not exhibit any sensitivity to all DNA-damaging agents tested. An Atrad5a/Atrad5b double mutant resembles the sensitivity phenotype of the Atrad5a single mutants. Moreover, in contrast to Atrad5b, the two Atrad5a mutants are deficient in homologous recombination after treatment with the double-strand break-inducing agent bleomycin. Our results suggest that the RAD5-dependent error-free branch of postreplication repair is conserved between yeast and plants, and that AtRad5a might be functionally homologous to ScRad5.

RAD5a and REV3 function in two alternative pathways of DNA-damage tolerance in Arabidopsis

“…As discussed above, uev1d (Wen et al, 2008) and rad5a (Chen et al, 2008) single mutants are sensitive to DNA damaging agents. Since genes involved in yeast DDT are conserved in Arabidopsis (Kunz and Xiao, 2007) and mammals (Pastushok and Xiao, 2004), it suggests that the two-branch model of DDT is also conserved; however, how DDT operates in plants is unknown prior to this study.…”

“…While this investigation was in progress, Chen et al reported that only AtRAD5a (At5g22750), but not AtRAD5b (At5g43530), is required for the DNA damage response. More importantly, the Atrad5a Atrad5b double mutant behaves same as the Atrad5a single mutant (Chen et al, 2008), suggesting that AtRAD5b does not provide a backup function for AtRAD5a. This report prompted us to focus on the genetic interactions between TLS represented by AtREV3 and the putative error-free bypass pathway represented by AtRAD5a.…”

“…Both of them contain the conserved RING finger domain, characteristic of ubiquitin ligases and SWI/SNF helicase domains. Inactivation of AtRAD5a results in an enhanced sensitivity to cross-linking agents and MMS (Chen et al, 2008). Surprisingly, Atrad5b mutant plants did not show such sensitivity and the Atrad5a Atrad5b double mutant plants act like the Atrad5a mutant alone.…”

“…According to the result of searching database via blasting the yeast Rad5 sequence, sequences related to yeast Rad5 have been identified in Arabidopsis and these genes were grouped previously into a putative RAD5/RAD16-like gene family (Shaked et al, 2006). Among them, AtRad5a and AtRad5b have the most significant identity and similarity (Chen et al, 2008). Both of them contain the conserved RING finger domain, characteristic of ubiquitin ligases and SWI/SNF helicase domains.…”

“…is involved in homologous recombination while Atrad5b is not (Chen et al, 2008). More work is needed to determine whether AtRad5b is still a functional homologue of yeast Rad5.…”