%0 Journal Article %A Simon L. Currie %A Jedediah J. Doane %A Kathryn S. Evans %A Niraja Bhachech %A Bethany J. Madison %A Desmond K. W. Lau %A Lawrence P. McIntosh %A Jack J. Skalicky %A Kathleen A. Clark %A Barbara J. Graves %T ETV4 and AP1 transcription factors form multivalent interactions with three sites on the MED25 activator-interacting domain %D 2017 %R 10.1101/126458 %J bioRxiv %P 126458 %X The recruitment of transcriptional cofactors by sequence-specific transcription factors challenges the basis of high affinity and selective interactions. Extending previous studies that the N-terminal activation domain (AD) of ETV5 interacts with Mediator subunit 25 (MED25), we establish that similar, aromatic-rich motifs located both in the AD and in the DNA-binding domain (DBD) of the related ETS factor ETV4 interact with MED25. These ETV4 regions bind MED25 independently, display distinct kinetics, and combine to contribute to a high-affinity interaction of full-length ETV4 with MED25. Within the ETS family, high-affinity interactions with MED25 are specific for the ETV1/4/5 subfamily as other ETS factors display weaker or no detectable binding. The AD binds to a single site on MED25 and the DBD interacts with three MED25 sites, allowing for simultaneous binding of both domains in full-length ETV4. MED25 also stimulates the in vitro DNA binding activity of ETV4 by relieving autoinhibition. ETV1/4/5 factors are often overexpressed in prostate cancer and genome-wide studies in a prostate cancer cell line indicate that ETV4 and MED25 occupy enhancers that are enriched for ETS-binding sequences and are both functionally important for the transcription of genes regulated by these enhancers. AP1-binding sequences were observed in MED25-occupied regions and JUN/FOS also contact MED25; FOS strongly binds to the same MED25 site as ETV4 AD and JUN interacts with the other two MED25 sites. In summary, we describe features of the multivalent ETV4- and AP1-MED25 interactions, thereby implicating these factors in the recruitment of MED25 to transcriptional control elements.ADactivation domainACIDactivator-interacting domainChIP-Seqchromatin immunoprecipitation followed by deep sequencingDBDDNA-binding domainETSE26-transformation specificMED25Mediator subunit 25HSQCheteronuclear single quantum coherenceAP1activator protein 1ARandrogen receptor %U https://www.biorxiv.org/content/biorxiv/early/2017/04/11/126458.full.pdf