Identifying and characterizing proteins that detect unpaired DNA during meiosis.

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DESCRIPTION (provided by applicant): Meiosis is the process by which chromosomes are replicated, recombined, and segregated to gametes. Problems during meiosis can produce gametes with incorrect numbers of chromosomes, leading to disorders such as trisomy 21 (Down syndrome), trisomy 18 (Edwards syndrome), and trisomy 13 (Patau syndrome). Abnormal meiosis can also cause infertility in adults. Yet, despite the importance of meiosis, there are significant gaps in our understanding of its fundamental processes. For example, the 46 chromosomes in a human cell must first be organized into pairs of homologs in order for meiosis to be successful. However, the mechanism that identifies the homologs so that they can be paired is largely obscure. A phenomenon called meiotic silencing in the model eukaryote Neurospora crassa may help shed light on homolog pairing. During meiotic silencing, unidentified protein complexes scan DNA sequences between homolog pairs. When they find regions of DNA that do not match (i.e., that are unpaired), genes within these regions are prevented from being expressed. Because such protein complexes must have the ability to check for DNA homology between homologs, it is possible that they are also involved in the homology search that identifies the homologs in the first place. Additionally, these same protein complexes may also contribute to homology searches required by some DNA repair pathways. Their identification could thus have implications for cancer research. Therefore, the first specifc aim of this project is to identify and characterize the protein complexes that detect unpaired DNA during meiotic silencing. Proteins will be identified with a high-throughput genetic screen and characterized with respect to their 1) cellular localization patterns, 2) requirement for norma growth and development, 3) role in producing meiotic silencing-specific RNA, and 4) protein-interaction partners. The second aim is to identify a theoretical RNA predicted to be linked to unpaired DNA detection. This will be achieved by identifying transcriptionally quiescent regions in the N. crassa genome, unpairing those regions during meiosis, and characterizing the RNA transcripts that result from this unpairing. Completion of these aims should provide the knowledge necessary to develop a specific model of N. crassa unpaired DNA detection, which could help answer the broader question of homolog identification during meiosis, as well as shed light on DNA homology searching in general.