The mRNA-bound proteome of the early fly embryo

Early embryogenesis is characterized by the maternal to zygotic transition (MZT), in which maternally deposited messenger RNAs are degraded while zygotic transcription begins. Before the MZT, post-transcriptional gene regulation by RNA-binding proteins (RBPs) is the dominant force in embryo patterning. We used two mRNA interactome capture methods to identify RBPs bound to polyadenylated transcripts within the first two hours of embryogenesis. We identified a high-confidence set of 476 putative RBPs and confirmed RNA-binding activities for most of 24 tested candidates. Most proteins in the interactome are known RBPs or harbor canonical RBP features, but 99 were previously characterized as DNA-binding transcription factors (TFs), signaling proteins, etc., and were not known to bind RNA. mRNA-bound RBPs and TFs exhibit distinct expression dynamics, in which the newly identified RBPs dominate the first two hours of embryonic development. In situ hybridization data from existing databases showed that RNAs encoding RBPs are enriched in posterior regions of the early embryo, suggesting their general importance in posterior patterning and germ cell maturation. Protein on ecdysone puffs (PEP) and CG3800 CLIPseq small RNA sequencing data in Drosophila S2 cells stably transfected with FLAG/HA-tagged PEP or CG3800. CLIP experiments were conducted in biological duplicates, including rRNA-depleted, size selected, matched total RNA inputs