The structural and biochemical characterization of ETS transcription factors with relevance to prostate cancer

Appropriate regulation of gene expression is important for the development and homeostasis of multicellular organisms. DNA sequencespecific transcription factors play a central role in regulating the first step of gene expression, transcription. The aberrant expression of transcription factors is a common mechanism for the initiation and progression of many human cancers. The ETS family of transcription factors consists of twenty-eight human proteins that contain a conserved DNA-binding domain, termed the ETS domain. ETS factors have varied roles in organismal development and disease etiology. For example, ETS proteins from the ERG and ETV1/4/5 subfamilies are overexpressed in the majority of prostate cancers and contribute to cancer initiation and progression. In stark contrast, EHF and SPDEF are two ETS factors present in normal prostate tissue that have been characterized as tumor suppressors whose genes are often deleted during cancer progression. The phenotypic dichotomy displayed between these subclasses of ETS factors suggests that the understanding of the molecular mechanisms that underlie transcription factors' roles in normal and disease settings may provide additional opportunities for therapeutic intervention. Here we describe the DNA-binding autoinhibition of ETS factors ETV1, iv ETV4, and ETV5. An intrinsically disordered region and an α-helix cooperate to inhibit DNA-binding by altering the positioning of the DNA-recognition α-helix of the ETS domain. These inhibitory elements are distinct from those that have been previously described for other ETS factors. We also characterize the interaction of Mediator subunit 25 (MED25) with the transcriptional activation and DNA-binding domains of ETV4. The inhibitory α-helix of ETV4 provides a unique interaction surface for MED25, as compared to other ETS domains, and interaction with MED25 activates the DNA-binding of ETV4. We also demonstrate the differential ability of ETS factors to bind to DNA with JUN-FOS at composite DNA binding sites. These distinct intra- and intermolecular interactions distinguish ETS oncoproteins and tumor suppressors in prostate cancer and may, in part, underlie their phenotypic differences. Finally, we present an assay for ETS-DNA interactions that is amenable to high-throughput screening for small molecule inhibitors. This assay could be further modified to incorporate any of the previously described partnerships.