Low input ribosome profiling reveals stage-specific translational regulation during early C. elegans embryogenesis

Translation of maternal mRNAs guides early embryonic development. C. elegans embryos exhibit an invariant cell lineage from the first division, offering a unique opportunity to trace the impact of post-transcriptional gene regulation on subsequent embryonic development. Although prior work has explored the translation of a select few transcripts, we lack a transcriptome-wide view of translation control during early embryogenesis. We developed a method for low-input ribosome profiling and combined it with low-input RNA sequencing to measure new protein translation during the first four cell cycles of C. elegans embryogenesis. Strikingly, our results show a poor correlation between RNA abundance and ribosome occupancy in this system compared to other model systems, emphasizing the predominant role of variation in translation efficiency in this system. We found numerous translationally regulated transcripts that had not been previously identified. Housekeeping genes involved in ribosome function and biogenesis were stably translated throughout the studied stages, while genes involved in mRNA processing and mRNA binding are dynamically regulated, consistent with roles in cell fate specification. A gain-of-function mutation in a known translational repressor protein, OMA-1, stalls the developmental program by preventing the translational remodeling that should occur at the 2-cell stage. These insights advance our understanding of the post-transcriptional regulatory mechanisms that shape the embryonic proteome and direct cell differentiation, and provide a valuable resource studies of C. elegans development. Ribosome profiling via isotachophoresis (RiboITP) and RNA-seq were performed on pooled Caenorhabditis elegans embryos at distinct early developmental stages (1-cell, 2-cell, 4-cell, and 8-cell) in both wild-type (N2) and OMA-1 mutant (TX20 strain, zu405 allele) backgrounds. For wild-type, samples were collected at all four stages, while the OMA-1 mutant was analyzed at 1-cell, 2-cell, and 4-cell stages. Each stage-specific sample consisted of pooled embryos to ensure adequate material for both RiboITP and RNA-seq analysis. The RiboITP and RNA-seq measurements enabled calculation of translational efficiency at each developmental stage, allowing comparison of translational regulation between wild-type and the OMA-1 mutant during early embryogenesis.