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Abstract We studied translation factor eukaryotic initiation factor 4E (eIF4E) paralogs that regulate germline mRNAs. Translational control of mRNAs is essential for germ cell differentiation and embryogenesis. Messenger ribonucleoprotein complexes assemble on mRNAs in the nucleus, as they exit via perinuclear germ granules, and in the cytoplasm. Bound messenger ribonucleoproteins including eIF4E exert both positive and negative posttranscriptional regulation. In Caenorhabditiselegans, germ granules are surprisingly dynamic messenger ribonucleoprotein condensates that remodel during development. Two eIF4E paralogs (IFE-1 and IFE-3), their cognate eIF4E–interacting proteins, and polyadenylated mRNAs are present in germ granules. Affinity purification of IFE-1 and IFE-3 messenger ribonucleoproteins allowed mass spectrometry and mRNA-Seq to identify other proteins and the mRNAs that populate stable eukaryotic initiation factor 4E 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 2 eukaryotic initiation factor 4E paralogs for opposite purposes on critically regulated germline mRNAs. Sublocalization within adult perinuclear germ granules suggests an architecture in which Vasa/GLH-1, PGL-1, and the IFEs are stratified, which may facilitate sequential remodeling of messenger ribonucleoproteins leaving the nucleus. Biochemical composition of isolated messenger ribonucleoproteins indicates opposing yet cooperative roles for the 2 eukaryotic initiation factor 4E paralogs. 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. 

Cellular identity and fate are determined by the proteins synthesized. Initiation of mRNA translation requires an important translation factor, eIF4G (ifg-1 in C. elegans). Embryos use mRNA translational control for spatial and temporal regulation of protein synthesis. Using CRISPR engineering, we added in-frame epitope and fluorescent tags (V5, Myc, Flag, GFP, and mCherry) to IFG-1. Tagged forms containing the V5 epitope caused embryonic arrest. Internal disruption of the V5 tag restored viability at 25°C. This study demonstrates that the molecular nature of a small epitope tag is sufficient to disrupt C. elegans embryogenesis.