Phase separation of RNA-binding protein promotes polymerase binding and transcription

An RNA-involved phase-separation model has been proposed for transcription control. Yet, the molecular links that connect RNA to the transcription machinery remain missing. Here we find RNA-binding proteins (RBPs) constitute half of the chromatin proteome in embryonic stem cells (ESCs), and some are colocalized with RNA polymerase (Pol) II at promoters and enhancers. Biochemical analyses of representative RBPs show that the paraspeckle protein PSPC1 inhibits the RNA-induced premature release of Pol II, and makes use of RNA as multivalent molecules to enhance the formation of transcription condensates and subsequent phosphorylation and release of Pol II. This synergistic interplay enhances polymerase engagement and activity via the RNA-binding and phase-separation activities of PSPC1. In ESCs, auxin-induced acute degradation of PSPC1 leads to genome-wide defects in Pol II binding and nascent transcription. We propose that promoter-associated RNAs and their binding proteins synergize the phase separation of polymerase condensates to promote active transcription. Overall design: Pol II ChIP-seq and TT-seq were used to chracterize the changes of Ser5 or Ser2 phosphorylated Pol II occupancy and transcription rate upon PSPC1 acute degradation. RBP ChIP-seq were used to charaterize the binding of different RBPs on chromatin. Pol II ChIP-seqs rescued with different forms of PSPC1 proteins were used to characterize whether the changes of Ser2 phosphorylated Pol II occupancy upon PSPC1 acute degradation is dependent on PSPC1 protein. ChIP-seqs of PSPC1 mutants were used to charaterize the binding of different PSPC1 mutants on chromatin. PSPC1 CLIP-seq was used to detect RNA targets of PSPC1. EU-seq was used to detect change of transcription level upon PSPC1 acute depletion.