A mouse homolog of the Escherichia coli recA and Saccharomyces cerevisiae RAD51 genes.

Abstract

Analysis of mitotic and meiotic recombination in mammalian cells has been hampered by the complexity of the reactions involved as well as lack of mutants. Furthermore, none of the genes involved in the process has yet been identified. In budding yeast, Saccharomyces cerevisiae, the RADSI gene is essential along with other genes of the RAD52 epistasis group for mitotic and meiotic recombination and DNA repair. The Rad5l protein is structurally similar toEscherichia coi RecA protein, which is required in homologous recombination and SOS responses in bacteria. Here we report the isolation of a mouse homolog of the yeast RADSI gene. The amino acid sequence predicted from the gene shows 83% and 55% homology with those of the yeast RADSI and the E. coli recA product, respectively. The mouse gene complemented a rdSl mutation of S. cerevisiae with sensitivity to methylmethanesulfonate, which produces double-strand breaks of DNA. This gene is expressed in the thymus, testis, ovary, spleen, and intestine, suggesting that its product is involved in mitotic and meiotic recombination in addition to DNA repair. MATERIALS AND METHODSIsolation of Mouse Rad5l cDNA Clone and DNA Sequencing Analysis. Two oligonucleotides (5'-AGTTGGATCCGAA/ GTTC/TA/CGIACIGGIAAA/G-3' and 5'-AAGGAAGCT-TGCIAA/GA/GTGCATT/CTGICT/GA/G/C/TGC-3', where I refers to inosine) were designed to code for two peptides in the conserved regions of Saccharomyces cerevisiae, RAD51, and its Schizosaccharomyces pombe homolog. The amino acid sequences used were EFRTGKS, corresponding to residues 186-192, and ARQMHLA, corresponding to residues 298-304 of the RAD51 gene of S. cerevisiae (6), encompassing the two ATP binding motifs. The oligonucleotides were used in a reverse transcriptase polymerase chain reaction with total RNA from mouse testis as a template. The 360-bp fragment generated was subcloned and used as a probe to screen a AZAP cDNA library from 129/Sv mouse testes. A clone, pMR51, was isolated and both strands were sequenced.Complementation

Sublethal gamma-radiation induces differentiation of CD4-/CD8- into CD4+/CD8+ thymocytes without T cell receptor beta rearrangement in recombinase activation gene 2-/- mice.

“…Although partial differentiation of thymocytes and, to a lesser degree, bone marrow cells is promoted by irradiation, the lack of mature peripheral lymphocytes is no doubt due to the inability of radiation to reconstitute the recombinatorial assembly of antigen receptor genes. Thus, it appears that DNA-repair genes, although capable of recombinatorial activity (13)(14)(15)23), do not compensate for the lack of RAG-2 function. Moreover, the genetic alterations that we hypothesize may account for a partial lymphoid maturation and do not appear to act as second-site suppressors to deficiencies in the RAG-2 gene.…”

Mitochondria-related male infertility

“…It has been reported that synapsis of homozygous chromosomes in the mammalian spermatogenic meiotic process begins at the zygotene stage through chromosome movement and attachment (for reviews, see refs. 21 and 22) and that several factors for SCs, such as RAD51, DMC1, and cohesin, contain functional ATP-binding domains (23)(24)(25). In yeast, it has been demonstrated that mitochondrial respiratory function is essential for meiosis (13)(14)(15) and that mitochondrial dynamics determined by opposing mitochondrial fission and fusion events are important in the meiotic sporulation process (16).…”

Gonadotropin-regulated testicular RNA helicase (GRTH/Ddx25) is essential for spermatid development and completion of spermatogenesis

“…It is conceivable that GRTH either directly or indirectly influences the apoptosis pathway and the dynamic changes of chromosome events through the regulation of protein expression of relevant genes. It is interesting to note that DNA repair proteins that are necessary for the maintenance of chromosome integrity, such as Rad51 and Dmc1 (14,15), were increased in GRTH Ϫ/Ϫ testis. Rad51 is involved in mitotic and meiotic double-strand DNA break repair and Dmc1 is a germline meiotic-specific gene essential for chromosome synapsis in the first meiotic prophase.…”