Small RNA (sRNA) pathways are critical regulators of gene expression, germ cell integrity, and fertility. At the core of these pathways are effector complexes composed of sRNAs and their binding partners, the Argonaute (AGO) proteins. C. elegans expresses 19 different AGOs. Our lab has systematically characterized all 19 AGO expression patterns and observed that eight AGOs localize to germ granules. Germ granules are conserved, phase separated RNA/protein rich organelles that are only present in germline cells. Four different types of germ granules, P granules, Z granules, Mutator foci, and SIMR foci, are present in C. elegans and appear to perform separate functions. Germ granules can either be distributed in the cytoplasm, or associated with nuclear pores. In fact, up to 75% of nuclear pores are associated with P granules, therefore a extensive proportion of germline transcripts pass through germ granules upon exiting the nucleus. Disruption of germ granules leads to defects in RNA processing and stability, transgenerational RNA interference (RNAi), and fertility. We aim to identify which AGOs and germline transcripts localize to each of the granules, and how AGOs and germ granules interact to carry out their functions. For example, CSR-1b is an essential AGO which is known to localize to P granules, thus we examined the localization of one of its known target transcripts,
pgl-3, using smFISH to identify its localization. We observed that the
pgl-3 transcript localized to germ granules with partial overlap with CSR-1, and that this localization was disrupted in sRNA pathway, germ granule, and nuclear pore mutants. In parallel, our recent quantitative co-localization studies with AGO-tagged and germ granule-tagged strains reveal distinct patterns of AGO/germ granule overlap that strongly correlate with sRNA pathway function-that is, AGOs that target overlapping sets of genes via their sRNAs also display overlap with the same types of germ granules. Collectively, our systematic studies will uncover the sRNA and gene regulatory functions of each type of germ granule that are important for epigenetic inheritance and fertility.