SRSF3 facilitates reprogramming and coordinates gene expression in pluripotent cells through multiple RNA processing machineries — ASN Events
  1. Schedule
  2. Session 8B: RNA Regulation, Disease & Development
  3. SRSF3 facilitates reprogramming and coordinates gene expression in pluripotent cells through multiple RNA processing machineries

SRSF3 facilitates reprogramming and coordinates gene expression in pluripotent cells through multiple RNA processing machineries (#51)

Madara Ratnadiwakara 1 2 , Stuart Archer 3 , Jernej Ule 4 , Anja Knaupp 1 2 , Jose M Polo 1 2 , Traude Beilharz 1 5 , Minna-Liisa Anko 1 2
  1. Development and Stem Cells Program, Monash Biomedicine Discovery Institute, Monash University, Clayton, VIC, Australia
  2. Department of Anatomy and Developmental Biology, Monash University, Clayton, VIC, Australia
  3. Monash University, Clayton, VIC, Australia
  4. Department of Molecular Neuroscience, UCL, London, UK
  5. Department of Biochemistry and Molecular Biology, Monash University, Clayton, VIC, Australia

RNA binding proteins are key mediators of RNA processing and enable gene regulation at the post-transcriptional level. RNA processing comprises versatile regulatory mechanisms that are essential to expand and fine-tune transcriptomes during cell fate transitions. The establishment, maintenance and exit of pluripotency depend on precise coordination of gene expression. While the transcriptional and epigenetic control of pluripotent cells has been the primary focus, the regulation at the level of RNA remains poorly understood. Serine-arginine rich splicing factor 3 (SRSF3) is an RNA binding protein previously implicated in controlling somatic reprogramming, but the underlying molecular mechanisms are not known. We found that SRSF3 expression increases in two distinct phases during reprogramming, coinciding with the two major transcriptional transitions. By modulating the expression of SRSF3 during reprogramming and in pluripotent cells, we show that SRSF3 is essential for both the early cell proliferative response of reprogramming and the late commitment to pluripotency. The determination of SRSF3 binding sites using iCLIP (UV crosslinking and immunoprecipitation followed by sequencing) and RNA-seq analysis of reprogramming cells revealed that SRSF3 directly regulates the core pluripotency network. Firstly, SRSF3 binds to the essential pluripotency factor Nanog mRNA and facilitates its export from the nucleus to the cytoplasm. In SRSF3 depleted cells, Nanog mRNA is retained in the nucleus, leading to dramatic reduction in NANOG protein levels. Secondly, SRSF3 binds to multiple mRNAs encoding the polycomb repressive complex 2 (PRC2) which is crucial for epigenetic modulation during cell fate transitions. In SRSF3 depleted cells, Ezh2 mRNA encoding the histone methyltransferase enzyme gets degraded by nonsense mediated decay, ultimately resulting in reduced H3K27me3 levels. Finally, SRSF3 binds to ~50% of recently identified new class of introns, called detained introns, in pluripotent cells. The splicing of these introns is slower than the other introns within the pre-mRNA and constitutes a rate limiting step in the processing of the mRNA. This slow splicing affects the expression of several genes that play a key role during reprogramming, including other RNA regulators. In conclusion, SRSF3 is the key coordinator of gene expression in pluripotent cells. It controls the expression of chromatin modifiers, transcription factors and splicing regulators essential for the establishment and maintenance of pluripotency through several RNA processing machineries both in the nucleus and the cytoplasm.

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