Two eIF4E paralogs occupy separate germ granule mRNPs that mediate mRNA repression and translational activation

We studied translation factor eIF4E paralogs that regulate germline mRNAs. Translational control of mRNAs is essential for germ cell differentiation and embryogenesis. Messenger ribonucleoprotein (mRNP) complexes assemble on mRNAs in the nucleus, as they exit via perinuclear germ granules, and in the cytoplasm. Bound mRNP proteins including eIF4Es exert both positive and negative post-transcriptional regulation. In C. elegans, germ granules are surprisingly dynamic mRNP condensates that remodel during development. Two eIF4E paralogs (IFE-1 and IFE-3), their cognate eIF4E-Interacting Proteins (4EIPs), and polyadenylated mRNAs are present in germ granules. Affinity purification of IFE-1 and IFE-3 mRNPs allowed mass spectrometry and mRNA-Seq to identify other proteins and the mRNAs that populate stable eIF4E complexes. We find translationally repressed mRNAs (e.g. pos-1, mex-3, spn-4, etc.) enriched with IFE-3, but excluded from IFE-1. Identified mRNAs overlap substantially with mRNAs previously described to be IFE-1-dependent for translation. The findings suggest that oocytes and embryos utilize the two eIF4Es for opposite purposes on critically regulated germline mRNAs. Biochemical composition of isolated mRNPs indicates opposing yet cooperative roles for the two eIF4Es. We propose that the IFEs accompany controlled mRNAs in the repressed or activated state during transit to the cytoplasm. Copurification of IFE-1 with IFE-3 suggests they may interact to move repressed mRNAs to ribosomes. RNA-Seq analysis of mRNA population precipitated with each of two eIF4E paralogs from C elegans. The eIF4Es (IFE-1 and IFE-3) were individually tagged by CRISPR and co-expressed in the same worm strain. Each was purified from extracts by Trap (IP) using unique N-terminal tags. RNA-IP enrichment was compared to total mRNA-Seq from the same strain. RIP-Seq from strains expressing tags alone were also included but not used in the analysis because of poor mRNA representation.