Mediator binds to boundaries of chromosomal interaction domains and to proteins involved in DNA looping, RNA metabolism, chromatin remodeling, and actin assembly

Abstract

Mediator is a multi-unit molecular complex that plays a key role in transferring signals from transcriptional regulators to RNA polymerase II in eukaryotes. We have combined biochemical purification of the Saccharomyces cerevisiae Mediator from chromatin with chromatin immunoprecipitation in order to reveal Mediator occupancy on DNA genome-wide, and to identify proteins interacting specifically with Mediator on the chromatin template. Tandem mass spectrometry of proteins in immunoprecipitates of mediator complexes revealed specific interactions between Mediator and the RSC, Arp2/Arp3, CPF, CF 1A and Lsm complexes in chromatin. These factors are primarily involved in chromatin remodeling, actin assembly, mRNA 3′-end processing, gene looping and mRNA decay, but they have also been shown to enter the nucleus and participate in Pol II transcription. Moreover, we have found that Mediator, in addition to binding Pol II promoters, occupies chromosomal interacting domain (CID) boundaries and that Mediator in chromatin associates with proteins that have been shown to interact with CID boundaries, such as Sth1, Ssu72 and histone H4. This suggests that Mediator plays a significant role in higher-order genome organization.

MED12 exerts an emerging role in actin-mediated cytokinesis via LIMK2/cofilin pathway in NSCLC

“…Disrupted actin dynamics could induce aborted cytokinesis and decrease cell proliferation [18–20]. Recent study indicated that the Mediator complex subunit 8 (Med8) controls actin assembly via bond to Arp2/Arp3 complex [21]. However, the correlation of MED12 with actin assembly has not been explored.…”

The Med31 Conserved Component of the Divergent Mediator Complex in Tetrahymena thermophila Participates in Developmental Regulation

“…Mediator is a large protein complex required for basal and regulated expression of most RNA polymerase II (RNAP II)-transcribed genes, in part due to its interaction with and phosphorylation of the conserved C-terminal domain (CTD) of Rpb1 [1,2]. Mediator has been implicated in many aspects of gene expression including chromatin looping [3], higher-order chromatin folding [4], mRNA processing [5] and export [6], and transcriptional memory [7]. Mediator is thought to have played a major role during eukaryotic diversification [8,9], although its function remains unknown in evolutionarily deep branching eukaryotes lacking canonical CTD heptad repeats.…”

Functional interaction of human Ssu72 with RNA polymerase II complexes

“…Finally, Ssu72 may function to remove trace CTD phosphorylations on Pol II attempting to join the PIC, which may then serve to enhance the association of Mediator due to its inability to associate when the CTD is phosphorylated [ 57 ]. Mediator and Ssu72 are both important for the proper compaction of the yeast genome and both proteins co-precipitate from chromatin protein extracts [ 58 , 59 ]. This leads to the possibility that Ssu72 effects on chromatin are a result of its roles with Mediator and the PIC.…”