The long term goal of this project is to learn about mechanisms regulating the meiotic stage of spermatogenesis in mice. This is important because successful completion of meiosis is essential in the production of normal sperm. This goal is achieved by studying structure and behavioral dynamics of chromosomes during the first meiotic prophase. Work conducted during the current funding period demonstrated that Chromatin with the potential for recombination is more nuclease sensitive than nonrecombining chromatin. This work on meiotic chromatin conformation leads to fundamental questions about early meiotic prophase. When do the initiating events of recombination occur? What is the nature of these events? How are these events related to meiotic condensation of the sex chromosomes? What Proteins mediate them? The specific aims are designed to fill these gaps in our knowledge.
The first aim i s to determine whether the initiating events for mammalian meiotic recombination are DNA double-strand or single-strand breaks, and to determine the time of appearance of the recombination-associated DNA breaks during meiotic prophase in spermatocytes.
The second aim constitutes an investigation of the timing of meiotic condensation of sex chromosomes to relate it to the initiating events of recombination and is also a test of the hypothesis that condensation is related to failure of gene sequences to pair.
The final aim i s to utilize the yeast two- hybrid technology to screen mouse leptotene/zygotene and pachytene spermatocyte cDNA libraries to identify mouse proteins associated with a pivotal protein of recombination, the mouse Rad5l. By defining temporal orders events and by identifying key regulatory molecules, these studies will provide much-needed information about mechanisms of those meiotic prophase events that are important for production of genetically normal sperm capable of activating development.
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