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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 post-transcriptional 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 RNA-protein interactions during vertebrate MZT and shows that Hnrnpa1 RNA-binding activities are spatially and temporally coordinated to regulate RNA metabolism during early development.

SUBMITTER: Despic V

PROVIDER: S-EPMC5495070 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

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

Despic Vladimir V Dejung Mario M Gu Mengting M Krishnan Jayanth J Zhang Jing J Herzel Lydia L Straube Korinna K Gerstein Mark B MB Butter Falk F Neugebauer Karla M KM

Genome research 20170405 7

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 post-transcriptional 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 ...[more]

PMID: 28381614

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Project description:During early stages of embryonic development the genome is transcriptionally inactive and cells are under the control of maternally provided mRNA and proteins. At a key point in development, known as the maternal to zygotic transition (MZT), the genome becomes activated and the maternally provided mRNAs begin to degrade. In the early zebrafish embryo, when under maternal control, cells divide rapidly and synchronously with cell cycles that lack gap phases. At cell cycle ten the introduction of gap phases lengthens the cell cycle and synchronised division is lost. During the MZT, zygotic activation of microRNAs leads to the targeted degradation of a number of maternally provided mRNAs, thus linking genome activation to maternal mRNA degredation.While the MZT has been studied in several different organisms the molecular mechanisms that coordinate genome activity and mRNA degradation remain largely unknown. For example, while the bulk of zygotic transcription occurs at cell cycle ten we do not understand why there is a minor wave of transcription before this time point. Similarly maternal mRNAs degrade at different rates, with only a percentage undergoing microRNA-mediated degredation. The aparant different rates of maternal mRNA degredation may be obscured by zygotic transcription.In order to gain an understanding of the MZT I intend to establish a precise understanding of transcription in the early embryo by using solexa sequencing to perform transcript counting at five different developmental stages that span the MZT. Specifically I intend to use crosses from two different zebrafish strains SAT (Sequenced AB and Tbingen, Zv9) and WIK. This project will allow one to understand the overall transcription profiles of genes in the early embryo, but importantly, the SNPs between the two different strains will determine if transcripts are maternal or zygotic (paternal). As a proof of principle we will first use one lane of sequencing to identify transcripts from a cross of an SAT and a WIK fish. This will allow us to observe SNPs between the two different strains. This will be run over one solexa lane. We will then sequence from five different time points on four different crosses SAT male and WIK female, WIK male and SAT female, WIK male and WIK female and SAT male with SAT female. We will prepare the libraries, which will be sequenced paired-end 54 bp over a total of seven lanes of solexa.. This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/

Project description:BackgroundRNA modification-induced ovarian dysgenesis appears to be necessary for ovary development. However, how m5 C (5-methylcytosine)-coordinating modificatory transcripts are dynamically regulated during oogenesis, and ovarian development is unknown. The purpose of this study was to determine whether NOP2/Sun RNA methyltransferase 5 (Nsun5) deletion leads to suppression of ovarian function and arrest of embryonic development. The regulation of mRNA decay and stability by m5 C modification is essential at multiple stages during the maternal-to-zygotic (MZT) transition.MethodsMouse ovaries and oocytes with Nsun5KO and the KGN cell line were subjected to m5 C identification, alternative splicing analysis and protein expression. BS-m5 C-seq, real-time polymerase chain reaction, Western blot, immunofluorescence and actinomycin D treatment assays were used. In particular, BS-m5 C-seq revealed a dynamic pattern of m5 C sites and genes in the ovaries between Nsun5KO and WT mice at the 2-month and 6-month stages. Diverse bioinformatic tools were employed to identify target genes for Nsun5.ResultsHere, a maternal mRNA stability study showed that deletion of the m5 C methyltransferase Nsun5 obstructs follicular development and ovarian function, which leads directly to inhibition of embryogenesis and embryo development. Dynamic analysis of m5 C revealed that the level of m5 C decreased in a time-dependent manner after Nsun5 knockout. Regarding the molecular mechanism, we found that Nsun5 deficiency caused a m5 C decline in the exon and 3'UTR regions that influenced the translation efficiency of Mitotic arrest deficient 2 like 2 (MAD2L2) and Growth differentiation factor 9 (GDF9) in the ovary. Mechanistic investigation of alternative splicing indicated that Nsun5KO triggers aberrant events in the exon region of Brd8.ConclusionsNsun5 loss arrests follicular genesis and development in ovarian aging, indicating that Nsun5/m5 C-regulated maternal mRNA stabilization is essential for MZT transition.

Dataset Information

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

Publications

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

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