The TRAP (thyroid hormone receptor-associated proteins) transcription coactivator complex (also known as Mediator) was first isolated as a group of proteins that facilitate the function of the thyroid hormone receptor. This complex interacts physically with several nuclear receptors through the TRAP220 subunit, and with diverse activators through other subunits. TRAP220 has been reported to show ligand-enhanced interaction with peroxisome proliferator-activated receptor gamma(2) (PPAR gamma(2)), a nuclear receptor essential for adipogenesis. Here we show that Trap220(-/-) fibroblasts are refractory to PPAR gamma(2)-stimulated adipogenesis, but not to MyoD-stimulated myogenesis, and do not express adipogenesis markers or PPAR gamma(2) target genes. These defects can be restored by expression of exogenous TRAP220. Further indicative of a direct role for TRAP220 in PPAR gamma(2) function via the TRAP complex, TRAP functions directly as a transcriptional coactivator for PPAR gamma(2) in a purified in vitro system and interacts with PPAR gamma(2) in a ligand- and TRAP220-dependent manner. These data indicate that TRAP220 acts, via the TRAP complex, as a PPAR gamma(2)-selective coactivator and, accordingly, that it is specific for one fibroblast differentiation pathway (adipogenesis) relative to another (myogenesis).

The yeast SWI/SNF complex is required for the transcription of several yeast genes and has been shown to alter nucleosome structure in an ATP-dependent reaction. In this study, we show that the complex stimulated in vitro transcription from nucleosome templates in an activation domain-dependent manner. Transcription stimulation by SWI/SNF required an activation domain with which it directly interacts. The acidic activation domains of VP16, Gcn4, Swi5, and Hap4 interacted directly with the purified SWI/SNF complex and with the SWI/SNF complex in whole-cell extracts. The similarity of activation domain interactions and transcriptional stimulation between SWI/SNF and the SAGA histone acetyltransferase complex may account for their apparent overlapping functions in vivo.

Transcriptional coactivators showing physical and functional interactions with PPARgamma include the protein acetyl transferase p300, the TRAP/Mediator complex that interacts with the general transcription machinery, and the highly regulated PGC-1alpha. We show that PGC-1alpha directly interacts with TRAP/Mediator, through the PPARgamma-interacting subunit TRAP220, and stimulates TRAP/Mediator-dependent function on DNA templates. Further, while ineffective by itself, PGC-1alpha stimulates p300-dependent histone acetylation and transcription on chromatin templates in response to PPARgamma. These functions are mediated by largely independent PPARgamma, p300, and TRAP220 interaction domains in PGC-1alpha, whereas p300 and TRAP220 show ligand-dependent interactions with a common region of PPARgamma. Apart from showing PGC-1alpha functions both in chromatin remodeling and in preinitiation complex formation or function (transcription), these results suggest a key role for PGC-1alpha, through concerted but dynamic interactions, in coordinating these steps.

Ligand activation of Notch receptors leads to release of the intracellular receptor domain (Notch IC), which translocates to the nucleus and interacts with the DNA-binding protein RBP-J to control expression of specific target genes. A number of proteins have been shown to interact with Notch ICs and to modulate target gene activation, but the precise function of and interplay between these factors is not known. This report investigates the Notch IC-interacting proteins, p300, PCAF, and Mastermind-like 1 (MAML1), in an in vitro transcription system with purified factors and naked DNA or chromatin templates. MAML1, RBP-J, and Notch IC are all required for optimal transcription from DNA, whereas transcription from chromatin requires, in addition, p300, which interacts with MAML1. The transcriptional activity of p300 requires acetyl coenzyme A, indicating that it functions as a histone acetyltransferase when mediating Notch IC function. PCAF is unable to promote transcription on its own but enhances Notch IC-mediated transcription from chromatin in conjunction with p300. These data define a critical role for p300 in the potentiation of Notch IC function by MAML1 and PCAF, provide the first evidence for cooperativity between PCAF and p300 in Notch IC function, and also indicate direct effects of RBP-J, Notch IC, and MAML1 on the general transcription machinery.