PGL granules formed under heat stress conditions contain components of the translation machinery, related to Fig. 1 . (A) Proteomics data from three independent replicates were analyzed by volcano plot. Blue dots indicate proteins significantly enriched in the GFP::PGL-3 IP-MS compared to GFP control (log2 fold change ≥1 and two-tailed, unpaired t test results P < 0.05). The top 10 enriched proteins are highlighted. Gray dots indicate proteins with no significant enrichment. (B) Brief description of the top 10 enriched proteins identified in the GFP::PGL-3 IP-MS compared the GFP control. GLH-1 and GLH-2 are the known components of germline P granules. (C) The 574 proteins that are significantly enriched in the GFP::PGL-3 IP-MS compared with GFP control were analyzed by DAVID. The top GO enrichment terms for molecular function are listed according to the number of proteins with P < 0.05 (automatically generated by DAVID). Black bars indicate proteins involved in RNA-related processes. (D and E) In atg-3(bp412) mutant embryos, IFE-1::GFP forms a large number of granules in somatic cells. Compared with embryos grown at 20°C (D), the number and intensity of IFE-1::GFP granules are increased, and the level of diffuse IFE-1::GFP is decreased, in embryos grown at 26°C (E). (F and G) The number and intensity of IFE-1::GFP granules are reduced in sepa-1(bp1726); atg-3(bp412) double mutant embryos at both 20°C (F) and 26°C (G). Maximum-intensity projections of Z-stack confocal images of comma-stage embryos are shown in D–G. (H) Quantification of the number of IFE-1::GFP granules in somatic cells. Data are shown as mean ± SEM (n = 3; n refers to the number of embryos analyzed for each genotype). Two-tailed, unpaired t test results: *P < 0.05. (I and J) Somatic IFE-1::GFP granules are partially colocalized with PGL granules labeled by TagRFP::SEPA-1 in atg-3(bp412) mutant embryos at 20°C (I) and 26°C (J). Some of the IFE-1::GFP granules are negative for SEPA-1. Comma-stage embryos are shown in I and J. (K and L) 3 µM purified TagRFP::IFE-1 protein fails to undergo LLPS during the observation time of 5 min. The reaction contained 150 mM NaCl. K is the DIC image of L. (M and N) TagRFP::IFE-1 is not evidently partitioned into droplets formed by 3 µM PGL-3/SEPA-1 proteins in an in vitro LLPS assay. M is the DIC image of N. Scale bars: 5 µm for D–G, I, and J; 2 µm for enlarged images in I and J; 20 µm for K–N.