The transcription factor SRF (serum response factor) recruits two families of coactivators, the MRTFs (myocardinrelated transcription factors) and the TCFs (ternary complex factors), to couple gene transcription to growth factor signaling. Here we investigated the role of the SRF network in the immediate transcriptional response of fibroblasts to serum stimulation. SRF recruited its cofactors in a gene-specific manner, and virtually all MRTF binding was directed by SRF. Much of SRF DNA binding was serum-inducible, reflecting a requirement for MRTF-SRF complex formation in nucleosome displacement. We identified 960 serum-responsive SRF target genes, which were mostly MRTF-controlled, as assessed by MRTF chromatin immunoprecipitation (ChIP) combined with deep sequencing (ChIP-seq) and/or sensitivity to MRTF-linked signals. MRTF activation facilitates RNA polymerase II (Pol II) recruitment or promoter escape according to gene context. MRTF targets encode regulators of the cytoskeleton, transcription, and cell growth, underpinning the role of SRF in cytoskeletal dynamics and mechanosensing. Finally, we show that specific activation of either MRTFs or TCFs can reset the circadian clock.

SummaryThe ERK-regulated ternary complex factors (TCFs) act with the transcription factor serum response factor (SRF) to activate mitogen-induced transcription. However, the extent of their involvement in the immediate-early transcriptional response, and their wider functional significance, has remained unclear. We show that, in MEFs, TCF inactivation significantly inhibits over 60% of TPA-inducible gene transcription and impairs cell proliferation. Using integrated SRF ChIP-seq and Hi-C data, we identified over 700 TCF-dependent SRF direct target genes involved in signaling, transcription, and proliferation. These also include a significant number of cytoskeletal gene targets for the Rho-regulated myocardin-related transcription factor (MRTF) SRF cofactor family. The TCFs act as general antagonists of MRTF-dependent SRF target gene expression, competing directly with the MRTFs for access to SRF. As a result, TCF-deficient MEFs exhibit hypercontractile and pro-invasive behavior. Thus, competition between TCFs and MRTFs for SRF determines the balance between antagonistic proliferative and contractile programs of gene expression.

In peripheral tissues circadian gene expression can be driven either by local oscillators or by cyclic systemic cues controlled by the master clock in the brain's suprachiasmatic nucleus. In the latter case, systemic signals can activate immediate early transcription factors (IETFs) and thereby control rhythmic transcription. In order to identify IETFs induced by diurnal blood-borne signals, we developed an unbiased experimental strategy, dubbed Synthetic TAndem Repeat PROMoter (STAR-PROM) screening. This technique relies on the observation that most transcription factor binding sites exist at a relatively high frequency in random DNA sequences. Using STAR-PROM we identified serum response factor (SRF) as an IETF responding to oscillating signaling proteins present in human and rodent sera. Our data suggest that in mouse liver SRF is regulated via dramatic diurnal changes of actin dynamics, leading to the rhythmic translocation of the SRF coactivator Myocardin-related transcription factor-B (MRTF-B) into the nucleus.

In vitro, without Mediator, the association of general transcription factors (GTF) and RNA polymerase II (Pol II) in preinitiation complexes (PIC) occurs in an orderly fashion. In this work, we explore the in vivo function of Mediator in GTF recruitment to PIC. A direct interaction between Med11 Mediator head subunit and Rad3 TFIIH subunit was identified. We explored the significance of this interaction and those of Med11 with head module subunits Med17 and Med22 and found that impairing these interactions could differentially affect the recruitment of TFIIH, TFIIE, and Pol II in the PIC. A med11 mutation that altered promoter occupancy by the TFIIK kinase module of TFIIH genome-wide also reduced Pol II CTD serine 5 phosphorylation. We conclude that the Mediator head module plays a critical role in TFIIH and TFIIE recruitment to the PIC. We identify steps in PIC formation that suggest a branched assembly pathway.