Abstract

The Mediator complex regulates transcription by connecting enhancers to promoters. High Mediator binding density defines super enhancers, which regulate cell-identity genes and oncogenes. Protein interactions of Mediator may explain its role in these processes but have not been identified comprehensively. Here, we purify Mediator from neural stem cells (NSCs) and identify 75 protein-protein interaction partners. We identify super enhancers in NSCs and show that Mediator-interacting chromatin modifiers colocalize with Mediator at enhancers and super enhancers. Transcription factor families with high affinity for Mediator dominate enhancers and super enhancers and can explain genome-wide Mediator localization. We identify E-box transcription factor Tcf4 as a key regulator of NSCs. Tcf4 interacts with Mediator, colocalizes with Mediator at super enhancers and regulates neurogenic transcription factor genes with super enhancers and broad H3K4me3 domains. Our data suggest that high binding-affinity for Mediator is an important organizing feature in the transcriptional network that determines NSC identity.

Highlights

  • There is debate on whether super enhancers (SEs) act mechanistically different from typical enhancers[12]

  • We were interested in proteins that can bind to the Mediator complex relying solely on protein–protein interactions and not being mediated via chromatin, which may co-purify with a chromatin-binding factor such as the Mediator complex

  • We reasoned that proteins interacting with Mediator by protein–protein interaction would not show a reduced interaction efficiency when treating the nuclear extract with the DNA–RNA digesting enzyme Benzonase or with ethidium bromide (EtBr), which intercalates in the DNA and disrupts protein-DNA interactions, as compared to untreated nuclear extracts (Fig. 1a)

Read more

Summary

Introduction

There is debate on whether SEs act mechanistically different from typical enhancers[12]. In a recently proposed model, the constituent enhancers of an SE and their regulated promoter(s) would group together to form a phaseseparated assembly[14]. Such an assembly would rely on interactions between transcriptional and chromatin regulators[14]. Mediator was shown to interact with SE-enriched chromatin modifier Crebbp[17] and the Cohesin complex[18], suggesting that Mediator could, in principle, provide an anchoring role at enhancers, SEs and the proposed phase-separated assemblies. High-Mediator affinity appears an important characteristic of master TFs. Our Mediator interactome contains many known enhancer-binding chromatin modifiers and we show that Mediator-interacting chromatin modifiers Jmjd1c and. Together this suggests that high-Mediator-binding affinity selects proteins that play important roles in establishing and maintaining enhancers and SEs to facilitate the regulation of cell identity

Methods
Results
Conclusion

Talk to us

Join us for a 30 min session where you can share your feedback and ask us any queries you have

Schedule a call

Disclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.