The long term goal of this proposal is to understand the biological roles of DNA helicase and helicase-related genes in recognizing and repairing spontaneous damage that occurs during the mitotic cell cycle and in meiosis. The mitotic DNA damage most likely occurs during DNA replication, but may be processed by homologous recombination. Defects in recombination, particularly in helicase genes, can result in blocked substrates that trigger a cell cycle checkpoint. The DNA lesions that occur in meiosis are of two types. Again it is possible that damage arises as a result of replication and this damage is repaired via recombination. It is not known how damage that occurs during meiotic replication is recognized and repaired and distinguished from meiotic DSBs, or whether failure to repair this damage can arrest cells before the meiotic DSBs are formed. This proposal is aimed at examining some of these issues, with focus on the RAD54 and RDH54 recombination/repair genes and the SRS2 DNA helicase gene. Results from these experiments will have implications for spontaneous chromosome loss, which can result in loss of heterozygosity. Loss of heterozygosity unveils recessive mutations and is associated with many cancers. This type of genomic instability will be studied through a careful examination of arrested mutant strains, to determine when in the cell cycle the arrest occurs and whether the arrested cells have accumulated novel replication or recombination intermediates. The mechanism of chromosome loss will be studied by determining the chromosome number in the sister cells of the cells undergoing chromosome loss. The biological consequences of mutations in yeast RAD54 and RDH54, modeled on RAD54 mutations found in human tumors and yeast SWI2/SNF2 mutations that impair function of this chromatin remodeling protein will be studied.
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