IMMUNOLOGIC STUDY OF GERM GRANULES IN C. ELEGANS EMBRY0S

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The establishment of asymmetry during early embryogenesis and the distribution of different developmental information to different embryonic cells are crucial processes in the development of every multicellular organism. Yet the nature of embryonic informational molecules and the mechanisms governing their differential distribution are not understood. This proposal delineates a combined immunologic, genetic, and biochemical approach to studying these problems in the nematode C. elegans. A set of monoclonal antibodies directed against germ-line-specific granules (P granules) has already been generated. In response to the asymmetry of the embryo, these granules are segregated specifically to the germ line or P lineage during early embryogenesis. P granules serve as an excellent marker for studying the mechanism of a symmetric distribution of cytoplasmic components to specific embryonic cells. Furthermore, they may act to specify germ-cell fates. The antibodies directed against P granules will be used in combination with molecular, cellular, and genetic techniques to address the following questions: 1) What is the nature of P granules? The antibodies will be utilized to purify P granules, identify their polypeptide components, and determine whether they contain nucleic acid. 2) Are P granules required for germ-cell development and, if so, how do they function? The antibodies will be used to specifically recover and characterize P-granule mutants. Antibodies will also be microinjected into living worms and embryos to try to functionally inactivate P granules. 3) How is P-granule gene expression regulated? The antibodies will be used to clone the genes encoding P-granule polypeptides, and the cloned sequences will be used to study the organization and transcription of the genes. 4) How are P granules segregated to the germ lineage? Because actin filaments appear to be involved in P-granule segregation, the organization of actin filaments appear to be involved in P-granule segregation, the organization of actin filaments in C. elegans embryos and the effects of actin inhibitors on granule segregation will be examined. The long range goal of the proposed experimental approach is to better understand how cell fates are specified early in embryonic development.