Germ granules share features with many other membraneless organelles, and recent evidence suggests that a liquid-liquid phase separation (LLPS) process drives their formation. The C. elegans adult germ line is an excellent model to study LLPS and its role in RNP granule assembly due to the dramatic cellular changes that occur in response to meiotic arrest or stress. During prolonged meiotic arrest that occurs in middle-aged hermaphrodites depleted of sperm and in females, germ granules recruit additional mRNAs and proteins to form large complexes termed RNP granules that are 20 times larger than typical germ granules. To identify regulators of RNP granule assembly, we performed an RNAi screen that identified 319 positives, including two interesting families of chaperone proteins, the the TCP-1 Ring Complex (TriC)/ chaperone containing Tcp-1 (CCT) family and the DnaJ/Heat shock protein 40 kDa (Hsp40) family (Wood et al., 2016). We are currently exploring the specificity of these chaperone proteins in regulating different types of RNP granules as well as the related Hsp70 chaperone family. Environmental stresses such as heat stress induce the assembly of large, germ line RNP granules that are similar, but not identical, to those induced by meiotic arrest. We have begun investigating the cellular response to glucose in the germ line and have determined that high concentrations of glucose induce the assembly of RNP granules. Interestingly, the RNP granules are maintained for up to three hours; however, they dissociate after longer periods of stress. Based on several lines of evidence, the germ line response to glucose largely appears to be an osmotic stress response, thus identifying osmotic stress as a trigger of LLPS. Although RNP granules are not maintained beyond three hours of osmotic stress, the quality of oocytes does not appear to decrease after longer periods of stress, suggesting a secondary adaptation in the germ line. We used an indirect marker of glycerol and observed high levels after five and twenty hours of glucose exposure. Moreover, in
gpdh-1;
gpdh-2 germ lines glycerol levels are reduced concomitant with RNP granules being maintained for an extended period. We speculate that increased glycerol levels may function as a secondary osmoregulatory adaptive response in the germ line, following a primary response of RNP granule assembly.