Weerasooriya, C. | bioRxiv Science

bioRxiv2026

Pumilio and Brain tumor regulate the expression of Smaug during the Drosophila maternal-to-zygotic transition

During the Drosophila maternal-to-zygotic transition, maternally deposited RNAs are cleared in a temporally controlled manner. The RNA-binding protein Smaug is a major regulator of maternal mRNA decay and its expression at both the RNA and protein level is limited to a narrow temporal window during the MZT. Here we show that Pumilio RNA-binding protein promotes degradation of smaug mRNA at the end of the MZT through Pumilio-binding cis-elements in the smaug 3 untranslated region, and that disruption of this regulation leads to ectopic Smaug protein expression beyond its normal developmental window. We further find that another RNA-binding protein, Brain tumor, also contributes to repression of ectopic Smaug expression. Transcriptome-wide analyses of embryos lacking Pumilio reveal that it directly regulates hundreds of maternal mRNAs after zygotic genome activation, with many of these Pumilio targets also regulated by Brain tumor. The ectopic Smaug protein that results from loss of either Pumilio or Brain tumor causes widespread downregulation of mRNAs that contain Smaug binding sites. Together, these findings define a post-transcriptional regulatory pathway in which Pumilio and Brain tumor ensure orderly progression of the Drosophila maternal-to-zygotic by clearing smaug mRNA and preventing ectopic Smaug activity.

developmental biology↗

bioRxiv2025

The ORB2 RNA-binding protein represses translation of its target transcripts during the Drosophila maternal-to-zygotic transition via its functionally conserved Zinc binding 'ZZ' domain

Post-transcriptional regulation is particularly prominent during the maternal-to-zygotic transition (MZT), a developmental phase during which a large proportion of maternally provided mRNAs is repressed and cleared from metazoan embryos. RNA-binding proteins (RBPs) are key components of the post-transcriptional regulatory machinery. We show that the ORB2 RBP, the Drosophila ortholog of human Cytoplasmic Polyadenylation Element Binding Protein (hCPEB) 2-4 protein subfamily, binds to hundreds of maternally provided, rare-codon-enriched mRNAs in early embryos; that ORB2 targets are translationally repressed and unstable during the MZT; identify a U-rich motif enriched in ORB2 targets 3'UTRs; and show that this motif confers ORB2 binding and repression to a luciferase reporter mRNA in S2 tissue culture cells. When tethered to a luciferase reporter, ORB2 and hCPEB2 (but not ORB and hCPEB1) repress translation; the C-terminal Zinc-binding ( ZZ) domain of ORB2 is necessary and sufficient for repression. ORB2 interacts with a suite of post-transcriptional regulators in early embryos; a subset of these interactions is lost upon deletion of the ZZ domain, notably with the Cup repressive complex. ORB2-targets significantly overlap with those previously identified for the repressive RBP, Smaug (SMG). Analysis of the early embryos translatome in the presence or absence of the endogenous ZZ domain shows that mRNAs bound by ORB2 but not by SMG move onto polysomes upon ZZ domain deletion whereas co-bound transcripts do not, consistent with co-regulation of the latter set of transcripts by both RBPs. Our results assign a function to the ZZ domain and position ORB2 in the post-transcriptional network that regulates maternal transcripts during the Drosophila MZT. ARTICLE SUMMARYWe show that Drosophila ORB2, the ortholog of the human CPEB2 RNA-binding protein, negatively regulates its target mRNAs during the maternal-to-zygotic transition via its C-terminal Zinc-binding ( ZZ) domain.

genetics↗