Cells of higher organisms must respond to a variety of different molecular signals impinging on their
surface. This information regulates the cell division and differentiation as well as the metabolic state.
In most cases, signals are transmitted through the cytoplasm where cross talk occurs, and then pass
into the nucleus to affect gene activity. Genes are turned on and off via proteins which bind
specifically to regulatory regions along the DNA, thereby enabling or disabling gene readout by the
DNA transcription machinery. These transcription factor proteins work in concert by assembling in
multi-protein complexes on adjacent and overlapping DNA sites. They may undergo chemical
modification such as phosphorylation due to the activation of a signal transduction kinase. One
aspect of our research focuses on providing atomic resolution, three-dimensional images of
transcription factor complexes, and showing how their molecular structures change after modification
in response to signal transduction. We use X-ray crystallography and synchrotron radiation for these
studies which provide mechanistic details of how the activation of DNA transcription is regulated
through signal transduction.
We determined the first atomic structure of a member of the MADS family of transcription factors,
the human serum response factor (SRF) bound to DNA (Nature 376, 490, 1995). We then extend
our investigation in the first part of the TMR project to a second member of the MADS family, yeast
MCM1 in a complex with the homeodomain protein MATalpha2 and DNA (Nature., 391,
660-666). In the last year we have completed the X-ray structures of a third family member, human
MEF2A bound to DNA (J. Mol. Biol. 297, 437), and of SRF in an important complex with an
ets-factor protein, SAP-1, and DNA (in preparation).
Myocyte enhancer factor-2 (MEF2) transcription factors regulate transcription of muscle-specific
genes important for myogenic development by directly binding the appropriate gene promoters. We
have now solved the X-ray structure at 1.5 Å resolution of the MADS and MEF domains of
MEF2A bound to its DNA recognition site. The structure reveals that while the DNA binding mode
is similar to that for SRF and MCM1, but the lack of amino acids N-terminal to the MADS domain
in MEF2 is crucial for its DNA binding specificity. The structure shows why the DNA bound to
MEF2A is straight instead of highly bent as seen in the SRF and MCM1 complexes. Furthermore,
the MEF domain, which is C-terminal to the MADS-box and interacts with coregulator proteins, has
a conformation considerably different from the same region in SRF and MCM1.
The c-fos promoter is paradigmatic for transcriptional regulation in human and mouse cells. The
serum response element (SRE) is found in many immediate-early genes and is necessary and
sufficient for rapid induction of the c-fos proto-oncogene in response to cell stimuli such as serum
agonists, growth factors and phorbol esters. Mitogenic signal transduction targets a protein complex
at the SRE containing the serum response factor (SRF) and one of several ETS-family factors such
as SRF-associated protein (SAP1). Formation of a functional ternary complex of SAP1/SRF/DNA
requires the conserved B-box and DNA-binding region of SAP1 and the DNA binding and
dimerization MADS-domain of SRF. We have solved the X-ray structure of the N-terminal core
domains and B-box linker of SAP1 and SRF bound to DNA revealing the basis of cooperative
interaction within the SAP1/SRF/DNA complex. Our results point to general principles for the
recognition of associating transcription factors.