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The Pol II Pre-Initiation Complex (PIC) Influences Mediator Binding but Not Promoter-Enhancer Looping [promoter_capture_Hi-C]

ABSTRACT: The Pol II Pre-Initiation Complex (PIC) Influences Mediator Binding but Not Promoter-Enhancer Looping [promoter_capture_Hi-C]

PROVIDER: PRJNA741245 | ENA |

REPOSITORIES: ENA

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The Pol II Pre-Initiation Complex (PIC) Influences Mediator Binding but Not Promoter-Enhancer Looping [RNA-seq]

Project description:Knowledge of how Mediator and TFIID crosstalk contributes to promoter-enhancer (P-E) communication is important for elucidating the mechanism of enhancer function. We conducted an shRNA knockdown screen in murine embryonic stem cells to identify the functional overlap between Mediator and TFIID subunits on gene expression. Auxin-inducible degrons were constructed for TAF12 and MED4, the subunits eliciting the greatest overlap. Degradation of TAF12 led to a dramatic genomewide decrease in gene expression accompanied by destruction of TFIID, loss of Pol II pre-initiation complex (PIC) at the promoter, and significantly decreased Mediator binding to promoters and enhancers. Interestingly, loss of the PIC elicited only a mild effect on P-E looping by Promoter Capture Hi-C (PCHi-C). Degradation of MED4 had a minor effect of Mediator integrity but led to a consistent 2-fold loss in gene expression, decreased binding of Pol II to Mediator, and decreased recruitment of Pol II to the promoters, but no effect on the other PIC components. PCHi-C revealed no consistent effect of Med4 degradation on P-E looping. Collectively, our data show that TAF12 and MED4 contribute mechanistically in different ways to P-E communication but neither factor appears to directly control P-E looping, thereby dissociating P-E communication from physical looping.

2021-07-22 | GSE178849 | GEO