What is the association between FBF-1 Mutant function and P-granule Localization in C. elegans Germlines?

Stem cells are essential to tissue maintenance through differentiation and proliferation to generate new cells, FBF homologs are key regulatory proteins that control their meiotic entry. In the Voronina Lab, we use the germ-lines of Caenorhabditis elegans to study how germ-cell RNA regulators like FBF-1 impact the substructure of P-granules, specialized germ-granules within the germline cytoplasm that preserve germline identity. The localization of these granules is generally conserved among species, but the importance of the specific substructure is unknown. Recent developments suggest that FBF-1 facilitates epigenetic small-RNA inheritance associated with p-granules, and functional differences can be observed in mutated strains where FBF-1 is present. We hypothesize that the changes in function observed in FBF-1 mutants are directly associated with substructural changes within p-granules. Using a standard cross strategy, we generated two mutated strains of c.elegans, fbf-1(ok91);gfp::flag::wago-4 and fbf-1(ok91);mut-16::gfp (each associated with different compartments), harvested the germlines, and immunostained them with antibodies to identify the granules of interest. A graduate student imaged the germ-lines, and we observed that both associated granules formed properly, but their relative distances from the P-granules differ from the WT, and abnormal puncta formed. This suggests that in the presence of FBF-1, mut-16 and wago-4 affected localization of their associated granules, and FBF-1 may directly or indirectly control the assembly of some of these foci or orientation of granules relative to one another. Further, FBF homologs may control small-RNA regulation in other species in which this mechanism is conserved.