Meiotic roles of Mre11 and Rad50 in Coprinus cinereus.

摘要

The MRN complex, composed of Mre11, Rad50, and Nbs1, is important for a large number of processes involving the metabolism of DNA double-strand breaks (DSBs). One such process is that of meiosis, in which a programmed DNA DSB is created by the Spo11 protein and processed by the MRN complex. Processing results in single-stranded DNA that is used to initiate meiotic recombination, using Rad51 and other proteins. In this work I examined the ability of the MRN complex mutants of C. cinereus to undergo meiotic recombination. I used immunofluorescence staining with C. cinereus anti-Rad51 antibody as a marker for recombination activity. Two classes of MRN complex mutants were identified, those that do not undergo recombination, and those that exhibit aberrant recombination. Results support the hypothesis that the MRN complex is not required for meiotic DSB formation but is required for DSB processing, and indicate a role for the Rad50 coiled coil and ATPase domains in that processing step.;Most activities of the MRN complex have been shown to require all the complex members. In C. cinereus a difference is observed between the late meiotic phenotypes of mre11-1 and rad50-1. This difference in phenotype led me to investigate the epistasis relationship between mre11-1 and rad50-1, and the localization patterns of Mre11 and Rad50. I have shown that mre11-1 is epistatic to rad50-1 for their late meiotic phenotypes, indicating that Mre11 is required for the arrest seen in rad50-1. Examination of Mre11 and Rad50 localization has demonstrated that in C. cinereus an increase in localization is seen in meiotic prophase, concurrent with predicted MRN complex DSB processing activity. Localization of Mre11 and Rad50 in MRN complex mutants has aided in explaining the relationship between the coiled coil and the ATPase domains of Rad50 and the meiotic functions of the MRN complex.