SRSF3 maintains transcriptome integrity in oocytes by regulation of alternative splicing and transposable elements

Dang Vinh Do,Ruiz De Los Mozos Igor,Iain Macaulay,John Marioni,Tim Xiaoming Hu,Engin Cukuroglu,Keng Boon Wee,M Azim Surani,Jonathan Göke,Caroline Lee,Bernhard Strauss,Ule Jernej,Sabine Dietmann,Richard Butler,Christopher W J Smith,Andrew Harrison

doi:10.1038/s41421-018-0032-3

Abstract

The RNA-binding protein SRSF3 (also known as SRp20) has critical roles in the regulation of pre-mRNA splicing. Zygotic knockout of Srsf3 results in embryo arrest at the blastocyst stage. However, SRSF3 is also present in oocytes, suggesting that it might be critical as a maternally inherited factor. Here we identify SRSF3 as an essential regulator of alternative splicing and of transposable elements to maintain transcriptome integrity in mouse oocyte. Using 3D time-lapse confocal live imaging, we show that conditional deletion of Srsf3 in fully grown germinal vesicle oocytes substantially compromises the capacity of germinal vesicle breakdown (GVBD), and consequently entry into meiosis. By combining single cell RNA-seq, and oocyte micromanipulation with steric blocking antisense oligonucleotides and RNAse-H inducing gapmers, we found that the GVBD defect in mutant oocytes is due to both aberrant alternative splicing and derepression of B2 SINE transposable elements. Together, our study highlights how control of transcriptional identity of the maternal transcriptome by the RNA-binding protein SRSF3 is essential to the development of fertilized-competent oocytes.

Highlights

Development of fertilization-competent oocytes includes completion of meiosis, cytoplasmic maturational events that provide competence for fertilization and embryogenesis, and maintenance of genomic integrity by protection against disruptive factors such as retrotransposon activation[1]
We first assessed whether SRSF3 protein and mRNA are present in oocytes and preimplantation embryos using immunofluorescence (IF) and single cell quantitative PCR (Fig. 1a, b)
We found that oocytes injected with B2 short interspersed nuclear element (SINE) RNAs showed a significant reduction in the percentage of germinal vesicle breakdown (GVBD) (40% reduction) as compared to those injected with water (Fig. 6f, g)

Summary

Introduction

Development of fertilization-competent oocytes includes completion of meiosis, cytoplasmic maturational events that provide competence for fertilization and embryogenesis, and maintenance of genomic integrity by protection against disruptive factors such as retrotransposon activation[1]. These important processes are largely dependent on mRNA and proteins that are synthesized and stored in oocytes as maternally inherited factors during their growth phase[2, 3]. Growing oocytes with an intact germinal vesicle (GV) are arrested at prophase I (referred to as fully grown GV oocytes) at the end of their growth phase. Because there is a transition from the transcriptionally active state in growing GV oocytes, to a transcriptionally inactive state in the fully grown GV and MII oocytes[4], it is necessary to generate a sufficient pool of maternal transcripts, while maintaining the transcriptome integrity in the oocyte

Methods

Results

Conclusion