Structural studies of transcriptional regulation at the human c-fos serum response element

The c-fos proto-oncogene is one of the best-studied immediately early genes that can be transiently induced by many extracellular signals. The serum response element (SRE) is an important cis-acting DNA element at the c-fos promoter region that is involved in regulating c-fos gene transcription. Two distinct transcription factors, serum response factor (SRF) and a ternary complex factor (TCF), can cooperatively bind to their adjacent DNA target sites at the SRE to achieve “combinatorial control” of c-fos gene transcription. The TCF proteins form a subfamily of the ETS-domain DNA-binding proteins that include SAP-1 and Elk-1, whose DNA-binding properties are very different despite their high similarity in the ETS DNA-binding domain. The structural studies of the SAP-1 ETS-domain, which has a less stringent DNA-binding specificity, were carried out for its complexes with two distinct DNA target sites, E74DNA, a high affinity site for all ETS-domain proteins, and c-fosDNA, a natural target in the c-fos SRE, which cannot be bound by the nascent Elk-1 ETS-domain. Together, the high-resolution crystal structures of the SAP-1/E74DNA and SAP-1/c-fosDNA complexes reveal that a set of conserved residues contact a GGA core DNA sequence. Discrimination for sequences outside this core is mediated by DNA contacts from conserved and non-conserved protein residues and sequence-dependent DNA structural properties characteristic of A-from DNA structure. The structural studies of the Elk-1 ETS-domain were carried out for its complex with an E74DNA site. A comparison of the Elk-1/E74 DNA structure with the SAP-1/E74DNA structure reveals that the differential DNA-binding properties of these proteins are nucleated by non-conserved residues distal from the DNA-binding surface by orienting residues in the DNA recognition helix for sequence-specific DNA contact. As a result, nearly one-third of the protein/DNA interactions mediated by the recognition helix are different between the two proteins. Together, the structures of SAP-1/E74DNA, SAP-1/c-fosDNA and Elk-1/E74 DNA complexes provide a detailed account for how the highly conserved ETS-domain achieves differential DNA-binding specificity. A ternary complex structure of the SAM ETS-domain and the SRF core domain homodimer bound to c-fos SRE DNA was determined with X-ray crystallography as well. The structure reveals that the DNA recognition helices of these proteins interact with each other when they bind to their adjacent target sites in the SRE DNA, thus stabilizing the ternary complex. Strikingly, the residues in SAP-1 that interact with SRF in the ternary complex were implicated from structural and biochemical studies to be important for DNA-binding specificity determination. Thus, the formation of the ternary complex may provide another way to modulate the DNA-binding specificity of the TCF proteins. In addition to the direct SRF-SAP-1 interactions at the SRE, there is a bend of 82° at the DNA region encompassing the target site for SRF. This bend is 10° larger than the bend observed in the SRF/DNA binary complex, suggesting that DNA-bending may also facilitate the formation of the SAP-1/SRF/SRE ternary complex