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Aspects Of Eukaryotic Dna Replication, Repair And Recombination
The mechanism that replicates, maintains, and sometimes alters the DNA is most fundamental and important for life. Three processes (DNA Replication, Repair, and Recombination) that are involved in this mechanism are closely related and well conserved in evolution. In my Ph.D. studies, I have used the cross model-organism approach to investigate the molecular mechanisms of DNA replication stress induced cancer as well as meiosis disruption caused infertility. Mcm4Chaos3, which encodes a mutant subunit of the hexameric MCM helicase, was previously reported to cause genetic instability (GIN) in mice and predispose homozygous female mice to mammary adenocarcinomas. My results show that homozygous diploid yeast carrying the equivalent mutation are defective in replicating long terminal repeat (LTR) elements. The replication stress at these interspersed repeat sequences coupled with error prone repair is the source of GIN, which is the driving force for the acquisition of an accelerated proliferation (AP) phenotype with aneuploidy as byproducts. I showed that mutations unrelated to aneuploidy are the cause of the AP phenotype. Moreover, the fragility of LTR regions is dependent on ploidy. The LTRs are not vulnerable to replication stress in haploid yeast and mcm4Chaos3 haploids use other repair pathways without generating GIN. Therefore, the dichotomy of consequences of DNA replication stress and repair pathway choice stems from cell-type specific regulation of fragile sites. In mammalian meiosis, homologous chromosome synapsis is coupled with recombination. As in most eukaryotes, mammalian meiocytes have checkpoints that monitor the fidelity of these processes. I reported that the mouse ortholog (Trip13) of pachytene checkpoint 2 (PCH2), an essential component of the synapsis checkpoint in S. cerevisiae and C. elegans, is required after strand invasion for completing a subset of recombination events, but possibly not those destined to be crossovers (Li and Schimenti 2007). TRIP13-deficient mice exhibit spermatocyte death in pachynema and loss of oocytes around birth. The chromosomes of mutant spermatocytes synapse fully, yet retain several markers of recombination intermediates. This is the first model to separate recombination defects from asynapsis in mammalian meiosis. Surprisingly, we found no evidence for checkpoint function, suggesting different pachytene checkpoint mechanisms may be involved in different species (Li et al. 2008).
dissertation or thesis