Dynamic RNA–protein interactions underlie the zebrafish maternal-to-zygotic transition

Abstract

During the maternal-to-zygotic transition (MZT), transcriptionally silent embryos rely on post-transcriptional regulation of maternal mRNAs until zygotic genome activation (ZGA). RNA-binding proteins (RBPs) are important regulators of posttranscriptional RNA processing events, yet their identities and functions during developmental transitions in vertebrates remain largely unexplored. Using mRNA interactome capture, we identified 227 RBPs in zebrafish embryos before and during ZGA, hereby named the zebrafish MZT mRNA-bound proteome. This protein constellation consists of many conserved RBPs, some of which are potential stage-specific mRNA interactors that likely reflect the dynamics of RNA-protein interactions during MZT. The enrichment of numerous splicing factors like hnRNP proteins before ZGA was surprising, because maternal mRNAs were found to be fully spliced. To address potentially unique roles of these RBPs in embryogenesis, we focused on Hnrnpa1. iCLIP and subsequent mRNA reporter assays revealed a function for Hnrnpa1 in the regulation of poly(A) tail length and translation of maternal mRNAs through sequence-specific association with 3 ′ UTRs before ZGA. Comparison of iCLIP data from two developmental stages revealed that Hnrnpa1 dissociates from maternal mRNAs at ZGA and instead regulates the nuclear processing of pri-mir-430 transcripts, which we validated experimentally. The shift from cytoplasmic to nuclear RNA targets was accompanied by a dramatic translocation of Hnrnpa1 and other pre-mRNA splicing factors to the nucleus in a transcription-dependent manner. Thus, our study identifies global changes in RNAprotein interactions during vertebrate MZT and shows that Hnrnpa1 RNA-binding activities are spatially and temporally coordinated to regulate RNA metabolism during early development.

Discovering the RNA-Binding Proteome of Plant Leaves with an Improved RNA Interactome Capture Method

“…Since 2012, RIC has been applied to multiple organisms including Trypanosoma brucei [16], Leishmania [17,18], Plasmodium falciparum [19], Saccharomyces cerevisiae [20][21][22][23], Caenorhabditis elegans [23], Drosophila melanogaster [2,24], Danio rerio [25], Mus musculus [26][27][28][29] and different Homo sapiens cell lines [3,[12][13][14]21,30,31]. This has greatly expanded the repertoire of RBPs, and with it our knowledge of RNA biology [32].…”

Cohesin facilitates zygotic genome activation in zebrafish

“…Other maternal RNAs are N6-methyladenosine (m6A) modified, and are cleared by a m6A-binding protein, Ythdf2 (Zhao et al, 2017). Unique RNAbinding proteins also play a role in controlling RNA metabolism and turnover during MZT (Despic et al, 2017). Therefore, clearance of maternal RNA is essential for transition to the zygotic transcription program.…”

Transcriptome-wide discovery of coding and noncoding RNA-binding proteins

“…Furthermore, the oligo(dT) capture enables detection of low-abundance crosslinked proteins, overcoming the limited efficiency of UV cross-linking. This method has has been applied to profile poly(A) RNA interactome in various mammalian cells (13)(14)(15)(16)(17)(18); Saccharomyces cerevisiae (14,19,20); Caenorhabditis elegans (20); zebrafish (21); the early embryo of Drosophila melanogaster (22,23); Arabidopsis thaliana seedlings, leaves, and cultured cells (24)(25)(26); and human parasites (27)(28)(29). However, the poly(A) tails mostly exist on mature mRNAs, leaving RBPs on nonpoly(A) RNAs, including most ncRNAs and pre-mRNAs, largely undiscovered.…”