RadB acts in homologous recombination in the archaeon Haloferax volcanii , consistent with a role as recombination mediator

Abstract

Homologous recombination plays a central role in the repair of double-strand DNA breaks, the restart of stalled replication forks and the generation of genetic diversity. Regulation of recombination is essential since defects can lead to genome instability and chromosomal rearrangements. Strand exchange is a key step of recombination – it is catalysed by RecA in bacteria, Rad51/Dmc1 in eukaryotes and RadA in archaea. RadB, a paralogue of RadA, is present in many archaeal species. RadB has previously been proposed to function as a recombination mediator, assisting in RadA-mediated strand exchange. In this study, we use the archaeon Haloferax volcanii to provide evidence to support this hypothesis. We show that RadB is required for efficient recombination and survival following treatment with DNA-damaging agents, and we identify two point mutations in radA that suppress the ΔradB phenotype. Analysis of these point mutations leads us to propose that the role of RadB is to act as a recombination mediator, which it does by inducing a conformational change in RadA and thereby promoting its polymerisation on DNA.

Role of RadA and DNA Polymerases in Recombination-Associated DNA Synthesis in Hyperthermophilic Archaea

“…However, any function of this protein, called RadB, has not been identified. In Haloferax volcanii, RadB is supposed to induce conformational change in RadA to promote its polymerization on DNA [68]. In P. furiosus, a direct interaction between RadA and RadB has been detected [11], but functionally the binding of RadB on DNA negatively interfered with the formation of the RadA nucleoprotein filament.…”

Differences in homologous recombination and maintenance of heteropolyploidy between Haloferax volcanii and Haloferax mediterranei

“…WARDELL et al 2017) expression was noted in H. mediterranei stationary phase cells. This difference could result in divergent processing of DNA breaks and explain the decreased homologous recombination rate in H. mediterranei.DiscussionH.…”

Haloferax volcanii —a model archaeon for studying DNA replication and repair

“…Archaeal RadA is more similar to eukaryotic Rad51 than to bacterial RecA [408]. Like bacterial cells lacking RecA, RadA-deficient H. volcanii cells show growth defects, a lack of homologous recombination, and increased sensitivity to DNA damage agents [409][410][411][412], indicating an important role for HR in archaea. In thermophilic archaea, RadA is essential [111], suggesting these organisms are critically reliant on HR for repair and/or replication.…”

“…In archaea, RadB is a paralogue of RadA that is present only in Euryarchaea [ 422 ]. In H. volcanii, RadB (HVO_2383) acts as a recombination mediator that interacts with RadA and promotes its polymerization on ssDNA [ 412 ]. Strains in which radB has been deleted show a phenotype similar to cells lacking radA ; defects include growth retardation, low levels of recombination and DNA damage sensitivity [ 411 , 412 ].…”

“…In H. volcanii, RadB (HVO_2383) acts as a recombination mediator that interacts with RadA and promotes its polymerization on ssDNA [ 412 ]. Strains in which radB has been deleted show a phenotype similar to cells lacking radA ; defects include growth retardation, low levels of recombination and DNA damage sensitivity [ 411 , 412 ]. Two suppressor mutations that alleviate the ΔradB phenotype have been identified in H. volcanii RadA, S101P and A196 V, which suggest that RadB induces a conformational change in RadA, promoting its polymerization on ssDNA [ 412 ].…”