Dr. Strome proposes to continue her ongoing investigations of control mechanisms that are essential for development and perpetuation of germ cells in the nematode C. elegans, taking advantage of its sophisticated genetics and sequenced genome. The proposal focuses on two separate but related aspects of germ cell function: 1) germ-line-specific RNA-rich granules called P granules and 2) a set of proteins required for survival of the germ line (MES proteins). The proposed studies of of P granules are based on the discovery in Dr. Stome's lab of PGL-1 protein a putative RNA binding P-granule protein that is required for germ-line development. In preliminary studies she has identified two new P-granule proteins that are similar in sequence to PGL-1 and bind to PGL-1 suggesting that a family of PGL proteins multimerize in P granules. She will determine which regions of the PGL proteins recruit them to P granules, and which germline processes require the PGL complex. She has also found that PGL-1 can bind to a germline-enriched isoform of eukaryotic translation initiation factor 4E (IFE1), raising the possibility that P granules control the translation of specific mRNAs. She will determine whether IFE-1 is a component of P granules and which germline processes fail when IFE-1 is eliminated. She will also identify P-granule-associated RNAs and use them to test models of P-granule function, namely delivery, stabilization, and control of translation of germline mRNAs. Four MES proteins, also discovered in Strome's lab, are required for normal early development and survival of the germline. Based on the similarity of three of the MES proteins to members of the Polycomb Group of transcriptional regulators, on molecular epistasis results, and on assays of transgene expression in the germline, she proposes that the MES proteins operate in complexes to control chromatin organization and gene expression in the early germline, and that death of the germline in mes mutants is due to aberrant patterns of gene expression. She will test this hypothesis by determining whether mes mutant germlines show altered patterns of gene expression, and by investigating whether MES proteins function through control of histone deacetylation. A novel genetic screen will also be used to identify new components involved in MES-mediated regulation.
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