Characterization of the Neurospora crassa mus-25 mutant: the gene encodes a protein which is homologous to the Saccharomyces cerevisiae Rad54 protein

Characterization of the Neurospora crassa mus-25 mutant suggests that it is defective in recombination repair and belongs to the uvs-6 epistasis group . It shows a high sensitivity to the alkylating agents methyl methanesulfon ate (MMS) and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), but not to UV ra diation. It is barren (i.e. does not produce ascospores) in homozygous cros ses. The frequency of MMS-induced mutations at the ad-3 loci is approximate ly three times higher than in the wild type. The ratio of homologous to non homologous integration of the pMTR::HYG plasmid is much lower than in wild type. The mus-25 mutant is epistatic to the mei-3 mutant for MMS sensitivit y, mei-3, which is a homololog of the Saccharomyces cerevisiae gene RAD51, is a member of the uvs-6 epistasis group which contains several genes that are homologous to recombination repair genes in other organisms. The mus-25 gene was cloned by identifying a genomic DNA fragment which complements th e MMS sensitivity of the mutant. The amino acid sequence deduced from the c loned DNA showed a high degree of homology to the Rad54 protein. which is i nvolved in recombinational repair in S, cerevisiae. Comparison of the nucle otide sequences of the genomic and cDNAs of the mus-25 gene revealed an ORF of 2505 by with a single 118-bp intron beginning immediately after the sec ond nucleotide of the AUG start codon. The molecular weight of the deduced gene product was 93.5 kDa. The transcript level was raised within 60 min af ter UV irradiation or MMS treatment, as also observed for the expression of the other N. crassa recombinational repair genes, suggesting the existence of a common mechanism which induces expression of the recombinational repa ir genes in response to DNA damage.