Specificity of cyclin E-Cdk2, TFIIB, and E1A interactions with a common domain of the p300 coactivator.

The role of the p300 and CREB binding protein (CBP) transcriptional coactivators in the regulation of cellular gene expression is complex and involves the modulation of gene expression in cooperation with many transcription factors. The activation of many cellular transcription factors by the p300 and CBP coactivators, including CREB (23), Myb (9), Fos (3), Jun (5), Sap1α (19), NF-κB (34), p53 (15, 25), Stat 1 (44), Stat 2 (7), nuclear receptors (21), and myogenic transcription factors (11), has been widely demonstrated, but the mechanisms for coordinating the regulation of these different transcriptional activators are poorly understood. p300 and CBP transcriptional coactivators likely function, in part, through their histone acetylation property (4, 31), which is also shared by p300/CBP-associated factor, P/CAF (43). In addition, the p300/CBP transcriptional coactivators may provide a link between specific transcription factors and the general transcriptional machinery through binding to TFIIB and TATA binding protein (TBP) (1, 10, 23). In vitro experiments with the beta interferon enhanceosome have demonstrated that certain transcriptional activators recruit TFIIB and other general transcription factors to the enhanceosome, which allows for subsequent recruitment of a CBP-containing RNA polymerase II holoenzyme (22). Thus, multiple contacts between promoter-bound factors and the holoenzyme may be required for assembly of a stable transcription complex, and the interaction of p300/CBP with both specific and general transcription factors may facilitate this process. Proteins that regulate cellular signaling and proliferation have also been found in association with p300/CBP. For instance, pp90Rsk and p160Src have been shown to contribute to the control of p300/CBP function (28, 29). In addition, cyclin E–cyclin-dependent kinase 2 (cyclin E-Cdk2), required for the G1/S transition of the cell cycle, has been found in association with p300/CBP (34). These complexes also seem to regulate p300 function, since inhibition of cyclin E-Cdk2 by the p21 cyclin-dependent kinase inhibitor activates NF-κB function through p300 (34). The mechanism for regulation of p300 by cyclin E-Cdk2 complexes, however, has not been determined. The adenoviral E1A proteins also bind to and inhibit p300/CBP (41). In addition to disrupting cell proliferation, E1A controls gene expression of both early viral and certain cellular transcripts. The single E1A message is spliced to give rise to two products, the 13S and 12S proteins of E1A, which share two conserved regions, CR1 and CR2, while 13S E1A contains a third motif, CR3, that is not present in the 12S form (13, 20). The transcriptional activating potential of 13S E1A is well documented and is linked to the CR3 region, the portion of E1A that interacts with the TBP (13, 14, 20). 12S E1A, however, represses transcription of certain viral and cellular genes (8, 12, 17, 18, 38–40), although its mechanism remains unclear. Here we investigate the molecular mechanisms underlying the regulation of p300 function by E1A, cyclin E-Cdk2, and TFIIB. In particular, we analyze the requirements for p300 binding by cyclin E-Cdk2, the 12S and 13S proteins of E1A, and TFIIB and compare the specificities of and levels of competition between these interactions. We conclude that TFIIB, cyclin E-Cdk2, and E1A all bind within a common region of p300 and that competition for binding to p300 suggests a mechanism for integrating a wide variety of cellular signals.