Two germ granule eIF4E isoforms reside in different mRNPs to hand off C elegans mRNAs from translational repression to activation.

We studied the function of translation factor eIF4E isoforms in regulating mRNAs in germ cell granules/condensates. Translational control of mRNAs plays an essential role in germ cell gene regulation. Messenger ribonucleoprotein (mRNP) complexes assemble on mRNAs as they move from the nucleus into perinuclear germ granules to exert both positive and negative post-transcriptional regulation in the cytoplasm. In C. elegans , germ granules are surprisingly dynamic mRNP condensates that remodel during development. Two eIF4E isoforms (called IFE-1 and IFE-3), eIF4E-Interacting Proteins (4EIPs), RBPs, DEAD-box helicases, polyadenylated mRNAs, Argonautes and miRNAs all occupy positions in germ granules. Affinity purification of IFE-1 and IFE-3 allowed mass spectrometry and mRNA-Seq to identify the proteins and mRNAs that populate stable eIF4E mRNPs. We find translationally controlled mRNAs (e.g. pos-1, mex-3, spn-4, etc.) enriched in IFE-3 mRNPs, but excluded from IFE-1 mRNPs. These mRNAs also require IFE-1 for efficient translation. The findings support a model in which oocytes and embryos utilize the two eIF4Es for opposite purposes on critically regulated germline mRNAs. Careful colocalization of the eIF4Es with other germ granule components suggests an architecture in which GLH-1, PGL-1 and the IFEs are stratified to facilitate sequential interactions for mRNAs. Biochemical characterization demonstrates opposing yet cooperative roles for IFE-3 and IFE-1 to hand-off of translationally controlled mRNAs from the repressed to the activated state, respectively. The model involves eIF4E mRNPs shuttling mRNAs through nuclear pore-associated granules/condensates to cytoplasmic ribosomes.

GLH-1/Vasa represses neuropeptide expression and drives spermiogenesis in the C. elegans germline.

Germ granules harbor processes that maintain germline integrity and germline stem cell capacity. Depleting core germ granule components in C. elegans leads to the reprogramming of germ cells, causing them to express markers of somatic differentiation in day-two adults. Somatic reprogramming is associated with complete sterility at this stage. The resulting germ cell atrophy and other pleiotropic defects complicate our understanding of the initiation of reprogramming and how processes within germ granules safeguard the totipotency and immortal potential of germline stem cells. To better understand the initial events of somatic reprogramming, we examined total mRNA (transcriptome) and polysome-associated mRNA (translatome) changes in a precision full-length deletion of glh-1, which encodes a homolog of the germline-specific Vasa/DDX4 DEAD-box RNA helicase. Fertile animals at a permissive temperature were analyzed as young adults, a stage that precedes by 24 â€‹h the previously determined onset of somatic reporter-gene expression in the germline. Two significant changes are observed at this early stage. First, the majority of neuropeptide-encoding transcripts increase in both the total and polysomal mRNA fractions, suggesting that GLH-1 or its effectors suppress this expression. Second, there is a significant decrease in Major Sperm Protein (MSP)-domain mRNAs when glh-1 is deleted. We find that the presence of GLH-1 helps repress spermatogenic expression during oogenesis, but boosts MSP expression to drive spermiogenesis and sperm motility. These insights define an early role for GLH-1 in repressing somatic reprogramming to maintain germline integrity.