The Transcription Regulation Unit applies a transcriptional approach to the study of mechanisms of signal transduction in activate T-cells. Work in this laboratory has established that the interleukin-2 (IL-2) promoter is major target of regulation by the p300/CBP nuclear co-activator proteins. A central component of the action of p300/CBP at the interleukin 2 promoter is its sequence specific recruitment to composite elements within the 300 bp proximal IL-2 promoter by the coordinate action of nuclear transcription factors. One of the key target sequences in the promoter is the CD28RE-TRE element, which mediates the assembly of a multi-protein complex at the IL-2 promoter containing c-rel/kappa B and ATF/CREB transcription factors. Recently we have defined this assembled complex as a central target for the integration of molecular signaling events during T-cell activation. Moreover we have established that this complex is targeted for down-regulation by the p53 tumor suppressor gene and the immunosuppreseve action of the glucocorticoid receptor. Surpringly the p53-dependent repression of the IL-2 promoter is not rescued by the action of the mdm2 oncogene. Instead we have found the mdm2 targets the IL-2 promoter for repression though a direct interaction with p300. The effect is in direct contrast to the action of the T-cell oncogene Tax which both reverses p53 induced repression of IL-2 expression as well as superinducing IL-2 expression in the absence of p53. We have also found that p300 activity can be differentially regulated in a promoter specific fashion by the glucorticoid receptor co-activator , SRC-1, in a manner that suggests a hormone driven molecular signaling pathway that acts at many levels to modulate cytokine gene expression in activated T-cells. Chromatin precipitation experiments to identify genes linked to p300 during T-cell activation suggest that the p300 occupancy at the IL-2 promoter is transient and subject to modification by the action of the factors and components described above. Through detailed studies of the biochemistry of p300 in activated T-cells and its functional interaction at the interleukin-2 promoter we have demonstrated that p300 undergoes significant post-translational modification in mitogen activated T-cells. Phospopeptide map analysis of p300/CBP indicate that there are changes in both the level of phosphorylation and the level of acetate incorporation including both acetylation and fatty acid modification. Interestingly, these modifications show correlation with the subnuclear localization of p300 in a manner that is distinct from it homologue CBP. Current studies are focussed on identifying the structural correlates of p300 that dictate its subnuclear localization and how this partitioning of p300 in activated T-cell influences it ability to regulate gene expression in response to molecular signaling events.
