RESUMO
Meiotic recombination and DNA repair are mediated by overlapping sets of genes. In the yeast Saccharomyces cerevisiae, many genes required to repair DNA double-strand breaks are also required for meiotic recombination. In contrast, mutations in genes required for nucleotide excision repair (NER) have no detectable effects on meiotic recombination in S. cerevisiae. The Drosophila melanogaster mei-9 gene is unique among known recombination genes in that it is required for both meiotic recombination and NER. We have analyzed the mei-9 gene at the molecular level and found that it encodes a homologue of the S. cerevisiae excision repair protein Rad1, the probable homologue of mammalian XPF/ERCC4. Hence, the predominant process of meiotic recombination in Drosophila proceeds through a pathway that is at least partially distinct from that of S. cerevisiae, in that it requires an NER protein. The biochemical properties of the Rad1 protein allow us to explain the observation that mei-9 mutants suppress reciprocal exchange without suppressing the frequency of gene conversion.
Assuntos
Reparo do DNA/genética , Proteínas de Ligação a DNA , Proteínas de Drosophila , Drosophila melanogaster/genética , Genes de Insetos , Proteínas Nucleares , Proteínas/genética , Recombinação Genética/genética , Saccharomyces cerevisiae/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteínas de Transporte/genética , Primers do DNA , Enzimas Reparadoras do DNA , Elementos de DNA Transponíveis , Drosophila melanogaster/efeitos dos fármacos , Endonucleases/genética , Proteínas Fúngicas/genética , Genes Fúngicos , Meiose , Metanossulfonato de Metila/farmacologia , Dados de Sequência Molecular , Mutagênese , Reação em Cadeia da Polimerase , Biossíntese de Proteínas , Proteínas/química , Receptores de Esteroides/metabolismo , Proteínas de Saccharomyces cerevisiae , Homologia de Sequência de AminoácidosRESUMO
The segregation of homologous chromosomes from one another is the essence of meiosis. In many organisms, accurate segregation is ensured by the formation of chiasmata resulting from crossing over. Drosophila melanogaster females use this type of recombination-based system, but they also have mechanisms for segregating achiasmate chromosomes with high fidelity. We describe a P-element mutagenesis and screen in a sensitized genetic background to detect mutations that impair meiotic chromosome pairing, recombination, or segregation. Our screen identified two new recombination-deficient mutations: mei-P22, which fully eliminates meiotic recombination, and mei-P26, which decreases meiotic exchange by 70% in a polar fashion. We also recovered an unusual allele of the ncd gene, whose wild-type product is required for proper structure and function of the meiotic spindle. However, the screen yielded primarily mutants specifically defective in the segregation of achiasmate chromosomes. Although most of these are alleles of previously undescribed genes, five were in the known genes alphaTubulin67C, CycE, push, and Trl. The five mutations in known genes produce novel phenotypes for those genes.
Assuntos
Elementos de DNA Transponíveis/genética , Drosophila melanogaster/genética , Genes de Insetos , Meiose/genética , Animais , Cromossomos/genética , DNA/genética , Feminino , Heterocromatina , Masculino , Metáfase , Mutação , Não Disjunção Genética , Fenótipo , Recombinação Genética , Projetos de Pesquisa , Cromossomo X/genéticaRESUMO
We investigated the roles of Caenorhabditis elegans MRE-11 in multiple cellular processes required to maintain genome integrity. Although yeast Mre11 is known to promote genome stability through several diverse pathways, inviability of vertebrate cells that lack Mre11 has hindered elucidation of the in vivo roles of this conserved protein in metazoan biology. Worms homozygous for an mre-11 null mutation are viable, allowing us to demonstrate in vivo requirements for MRE-11 in meiotic recombination and DNA repair. In mre-11 mutants, meiotic crossovers are not detected, and oocyte chromosomes lack chiasmata but appear otherwise intact. gamma-irradiation of mre-11 mutant germ cells during meiotic prophase eliminates progeny survivorship and induces chromosome fragmentation and other cytologically visible abnormalities, indicating a defect in repair of radiation-induced chromosome damage. Whereas mre-11 mutant germ cells are repair-deficient, they retain function of the meiotic G(2) DNA damage checkpoint that triggers germ cell apoptosis in response to ionizing radiation. Although mre-11/mre-11 animals derived from heterozygous parents are viable and produce many embryos, there is a marked drop both in the number and survivorship of embryos produced by succeeding generations. This progressive loss of fecundity and viability indicates that MRE-11 performs a function essential for maintaining reproductive capacity in the species.