BLAP75/RMI1 promotes the BLM-dependent dissolution of homologous recombination intermediates

Abstract

BLM encodes a member of the highly conserved RecQ DNA helicase family, which is essential for the maintenance of genome stability. Homozygous inactivation of BLM gives rise to the cancer predisposition disorder Bloom's syndrome. A common feature of many RecQ helicase mutants is a hyperrecombination phenotype. In Bloom's syndrome, this phenotype manifests as an elevated frequency of sister chromatid exchanges and interhomologue recombination. We have shown previously that BLM, together with its evolutionarily conserved binding partner topoisomerase III␣ (hTOPO III␣), can process recombination intermediates that contain double Holliday junctions into noncrossover products by a mechanism termed dissolution. Here we show that a recently identified third component of the human BLM͞hTOPO III␣ complex, BLAP75͞ RMI1, promotes dissolution catalyzed by hTOPO III␣. This activity of BLAP75͞RMI1 is specific for dissolution catalyzed by hTOPO III␣ because it has no effect in reactions containing either Escherichia coli Top1 or Top3, both of which can also catalyze dissolution in a BLM-dependent manner. We present evidence that BLAP75͞RMI1 acts by recruiting hTOPO III␣ to double Holliday junctions. Implications of the conserved ability of type IA topoisomerases to catalyze dissolution and how the evolution of factors such as BLAP75͞RMI1 might confer specificity on the execution of this process are discussed.Bloom's syndrome ͉ Holliday junction dissolution ͉ topoisomerase III ͉ sister chromatid exchanges T he RecQ family of DNA helicases is essential for the maintenance of genome stability (1). The human genome contains five RecQ helicase genes. Mutations in three of these genes give rise to clinically defined cancer predisposition disorders (2). One of these disorders is Bloom's syndrome (BS), which is caused by biallelic mutations in the BLM gene (3). The BLM protein is a 3Ј-5Ј DNA helicase that processes a broad range of structurally diverse DNA substrates (4-7). These substrates include DNA structures that arise during homologous recombination, such as D-loops and Holliday junctions (5, 6). These structures are of particular relevance to the BS phenotype because BS cells display elevated levels of homologous recombination (8). This hyperrecombination phenotype is also a feature of Saccharomyces cerevisiae and Schizosaccharomyces pombe mutants defective in their respective BLM orthologs, SGS1 and rqh1 ϩ (9-11). In the case of BS cells, recombination events are particularly apparent between sister chromatids, and such recombination events are termed sister chromatid exchanges (SCEs) (8). These exchanges arise primarily as a consequence of crossing-over during the processing of recombination intermediates (12).BLM exists in a complex with topoisomerase III␣ (hTOPO III␣), a type IA topoisomerase (13,14). This complex is evolutionarily conserved, and functional and͞or physical interactions between RecQ helicases and type IA topoisomerases have also been demonstrated in bacteria and yeast (9, 15-17). Two type IA topoisomerases ar...

Topoisomerase IIIα and Bloom’s helicase can resolve a mobile double Holliday junction substrate through convergent branch migration

“…However, the ability of type IA topoisomerases to pass one ssDNA through the other makes these enzymes an ideal agent in removing the topological linkages in between dHJ (5). Recent work has shown that several type IA topoisomerases, in association with BLM, are capable of removing the last two linkages from an oligonucleotide-based substrate (24,31). In this work, we have shown that DmTopo III␣, in association with DmBlm, is capable of catalyzing the earlier steps of convergent branch migration of the HJs.…”

Yeast Mph1 helicase dissociates Rad51-made D-loops: implications for crossover control in mitotic recombination

“…Mechanistically, the human BLM helicase works together with topoisomerase IIIa and the BLAP75/Rmi1 protein to branch migrate and decatenate dHJs into noncrossovers (Wu and Hickson 2003;Raynard et al 2006;Wu et al 2006). It is expected that Sgs1, the likely BLM ortholog in Saccharomyces cerevisiae, also functions in this manner to resolve dHJs into noncrossover during DSB repair.…”

Shu Proteins Promote the Formation of Homologous Recombination Intermediates That Are Processed by Sgs1-Rmi1-Top3

“…This is exemplified by the cancer-predisposition disorder, Bloom's syndrome, which is caused by mutation in the human BLM gene (reviewed in German, 1993). Because the BLM protein, in conjunction with its associated proteins, hTOPOIII␣ and hRMI1 (Johnson et al, 2000;Wu et al, 2000;Yin et al, 2005), can catalyze dissolution of HRR intermediates in vitro Plank et al, 2006;Raynard et al, 2006;Wu et al, 2006), it is likely that unprocessed and/or aberrantly processed HRR intermediates at least partly contribute to the cellular defects in Bloom's syndrome. Indeed, Bloom's syndrome cells classically demonstrate elevated levels of sister chromatid exchanges, mitotic recombination, and genome instability.…”

“…It should be noted, however, that we do not know presently if the X-molecules arising in MMS-treated sgs1, rmi1, and TOP3 Y356F strains are identical or if they represent different types of HRR intermediates that cannot readily be distinguished by the 2D gel technique. One possibility, based on the known enzymatic functions of BLM, hTOPOIII and RMI1 Plank et al, 2006;Raynard et al, 2006;Wu et al, 2006), is that X-molecules arising in sgs1 mutants are double Holliday junctions, whereas those arising in TOP3 Y356F , or rmi1, mutants are hemicatenanes formed by the Sgs1-dependent convergent branch migration of double Holliday junctions. Surprisingly, we found that although mutation of RAD51 prevented persistent MMS-induced HRR intermediates in sgs1, rmi1, and TOP3 Y356F cells, mutation of SHU1 did not.…”