Roles of Mediator subunit MED26 in Regulation of Post‐initiation Events in RNA Pol II Transcription

Abstract

Mediator is a conserved multi‐protein complex that transduces regulatory signals from gene‐specific transcription factors to RNA polymerase II and its regulatory factors to control transcription. Mediator subunit MED26 has been implicated in control of post‐initiation steps in transcription. Its N‐terminal domain binds two types of complexes containing elongation factor ELL, the Little Elongation Complexes (LECs) and Super Elongation Complexes (SECs) ( Takahashi et al., 2011). MED26 regulates expression of non‐polyadenylated snRNA genes, at least in part by recruiting LECs ( Takahashi et al., 2015). In assays using PRO‐seq to measure the location of transcribing Pol II in MED26 mutant cells, we observed an increase in mapped reads extending past the normal transcription termination sites of LEC regulated snRNA genes as well as genes encoding replication‐dependent histone transcripts, which, like snRNAs, are not polyadenylated ( Takahashi, Ranjan, et al., 2020). These observations suggest a new role for MED26‐containing Mediator in control of 3′‐end processing and transcription termination at these genes.MED26‐containing Mediator also helps recruit SECs to c‐myc and several other protein coding genes to regulate phosphorylation of the Rpb1 CTD and, most likely, transcription elongation ( Takahashi et al., 2011); however, whether MED26 contributes to these processes globally is unknown. In this study, we used genome‐wide approaches to examine the consequences of MED26 mutation in regulation of post‐initiation events including CTD phosphorylation and elongation, assessed by measuring the distribution of transcriptionally active Pol II across gene bodies. We observed that MED26 mutation alters Pol II distribution and reduces CTD Ser2 phosphorylation (Ser2P) at many protein‐coding genes, mostly towards their 3′‐ends. Moreover, MED26 mutation is associated with a remarkable shift of Ser2P towards 5′‐ends at many protein‐coding genes. Notably, 5′‐enrichment of Ser2P in MED26 mutant cells appeared independent of changes in gene expression. We hypothesize that because of a defect in Mediator’s ability to recruit elongation complexes, MED26 mutation could generally slow Pol II elongation and trigger premature termination.Support or Funding InformationThis research was supported by the Stowers Institute for Medical Research and the Helen Nelson Medical Research Fund of the Greater Kansas City Community Foundation