A genetic interaction network containing approximately 1000 genes and approximately 4000 interactions was mapped by crossing mutations in 132 different query genes into a set of approximately 4700 viable gene yeast deletion mutants and scoring the double mutant progeny for fitness defects. Network connectivity was predictive of function because interactions often occurred among functionally related genes, and similar patterns of interactions tended to identify components of the same pathway. The genetic network exhibited dense local neighborhoods; therefore, the position of a gene on a partially mapped network is predictive of other genetic interactions. Because digenic interactions are common in yeast, similar networks may underlie the complex genetics associated with inherited phenotypes in other organisms.

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...

We performed a systematic screen of the set of Ϸ5,000 viable Saccharomyces cerevisiae haploid gene deletion mutants and have identified 103 genes whose deletion causes sensitivity to the DNA-damaging agent methyl methanesulfonate (MMS). In total, 40 previously uncharacterized alkylation damage response genes were identified. Comparison with the set of genes known to be transcriptionally induced in response to MMS revealed surprisingly little overlap with those required for MMS resistance, indicating that transcriptional regulation plays little, if any, role in the response to MMS damage. Clustering of the MMS response genes on the basis of their cross-sensitivities to hydroxyurea, UV radiation, and ionizing radiation revealed a DNA damage core of genes required for responses to a broad range of DNA-damaging agents. Of particular significance, we identified a subset of genes that show a specific MMS response, displaying defects in S phase progression only in the presence of MMS. These genes may promote replication fork stability or processivity during encounters between replication forks and DNA damage. T he budding yeast Saccharomyces cerevisiae has been an invaluable tool for studying DNA damage-response pathways. Many S. cerevisiae DNA damage-response genes have human homologues, and mutations in a number of these genes have been implicated in human diseases. Although several screens for S. cerevisiae DNA damage-response genes have been conducted over the past 30-40 years, additional genes are still being identified. The set of viable S. cerevisiae deletion mutants (1) has allowed for genome-wide studies to identify genes required for resistance to various cellular insults (2-6). Here we report a systematic analysis of the complete set of Ϸ5,000 viable gene deletion mutants to identify genes that are required for resistance to the DNAdamaging agent methyl methanesulfonate (MMS).MMS is a monofunctional DNA alkylating agent and a known carcinogen (7,8) and primarily methylates DNA on N 7 -deoxyguanine and N 3 -deoxyadenine (9). Although the N 7 -methylguanine adduct may be nontoxic and nonmutagenic, N 3 -methyladenine is a lethal lesion that inhibits DNA synthesis and needs to be actively repaired (8, 10). The three pathways responsible for the removal of most N 3 -methyladenine lesions are bypass repair (or postreplication repair), recombination repair, and base excision repair (11). All three pathways are required for wild-type resistance to MMS-induced DNA damage (11). In addition, checkpoint proteins are required to maintain cell viability in the presence of MMS (12, 13).Several studies have found that cells are most sensitive to MMS during progression through S phase (13-15). Exposure to MMS causes a checkpoint-independent reduction in the rate of replication fork progression, likely due to a physical impediment of fork progression caused by alkylated DNA or some intermediate in lesion processing (13). rad53 and mec1 checkpoint mutants have high rates of replication fork termination, suggesting that damage-...