Availability of splicing factors in the nucleoplasm can regulate the release of mRNA from the gene after transcription

PLoS Genet. 2019 Nov 25;15(11):e1008459. doi: 10.1371/journal.pgen.1008459. eCollection 2019 Nov.

Abstract

Gene expression dynamics can be measured in single living cells. Using a detectable transcriptionally active gene in living cells, we previously found that an mRNA undergoing several splicing events was retained at this gene after transcription until completion of mRNA processing. To determine the reason for this delay in release and whether mRNA retention on the gene might depend on splicing factor availability, we modulated the levels of splicing factors in the nucleus. Increasing the abundance of the diffusing fraction of splicing factors by their overexpression or by Clk1 kinase overexpression to disassemble nuclear speckles, led to a reduction in splicing factor residence times on the active gene, and the retained mRNA was rapidly released from the gene. Other treatments such as overexpression of a mutant inactive Clk1, the downregulation of MALAT1 lncRNA or of the Son protein, or the overexpression of the splicing factor import factor TNPO3, did not affect the dynamics of mRNA release from the gene. We found that the faster release of the mRNA from the gene mediated by increased availability of splicing factors, was dependent on the RS domain of the splicing factors and its phosphorylation state. We propose that the relative abundancies of splicing factors in the nucleoplasm can affect their availability for the splicing events taking place, and regulate the kinetics of mRNA release from the gene after processing.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • DNA-Binding Proteins / genetics
  • Gene Expression Regulation / genetics
  • HeLa Cells
  • Humans
  • Introns / genetics
  • Minor Histocompatibility Antigens / genetics
  • Phosphorylation
  • Protein Binding / genetics
  • Protein Serine-Threonine Kinases / genetics
  • Protein-Tyrosine Kinases / genetics
  • RNA Precursors / genetics
  • RNA Splicing / genetics*
  • RNA Splicing Factors / genetics*
  • RNA, Long Noncoding / genetics
  • RNA, Messenger / genetics
  • RNA, Small Interfering / genetics
  • Transcription, Genetic*
  • beta Karyopherins / genetics

Substances

  • DNA-Binding Proteins
  • MALAT1 long non-coding RNA, human
  • Minor Histocompatibility Antigens
  • RNA Precursors
  • RNA Splicing Factors
  • RNA, Long Noncoding
  • RNA, Messenger
  • RNA, Small Interfering
  • SON protein, human
  • TNPO3 protein, human
  • beta Karyopherins
  • Clk dual-specificity kinases
  • Protein-Tyrosine Kinases
  • Protein Serine-Threonine Kinases

Grants and funding

This work was supported by the German-Israeli Foundation (929/06) and the European Research Council (GENEXP). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.