Genome-wide Impact of the BRG1 SWI/SNF Chromatin Remodeler on the Transforming Growth Factor β Transcriptional Program

Qiaoran Xi,Wei He,Xiang H.-F Zhang,Hong-Van Le,Joan Massagué

doi:10.1074/jbc.m707479200

Abstract

The transcription factors Smad2 and Smad3 mediate a large set of gene responses induced by the cytokine transforming growth factor beta (TGFbeta), but the extent to which their function depends on chromatin remodeling remains to be defined. We observed interactions between these two Smads and BRG1, BAF250b, BAF170, and BAF155, which are core components of the SWI/SNF chromatin-remodeling complex. Smad2 and Smad3 have similar affinity for these components in vitro, and their interactions are primarily mediated by BRG1. In vivo, however, BRG1 predominantly interacts with Smad3, and this interaction is enhanced by TGFbeta stimulation. Our results suggest that BRG1 is incorporated into transcriptional complexes that are formed by activated Smads in the nucleus, on target promoters. Using BRG1-deficient cell systems, we defined the BRG1 dependence of the TGFbeta transcriptional program genome-wide. Most TGFbeta gene responses in human epithelial cells are dependent on BRG1 function. Remarkably, BRG1 is not required for the TGFbeta-mediated induction of SMAD7 and SNON, which encode key mediators of negative feedback in this pathway. Our results provide a genome-wide scope of the participation of BRG1 in TGFbeta action and suggest a widespread yet differential involvement of BRG1 SWI/SNF remodeler in the transcriptional response of many genes to this cytokine.

Highlights

Recent identification of interactions between Smad transcription factors and a SWI/SNF remodeler complex [10, 11] provides an opportunity to investigate this question
Smad2 and Smad3 Bind a BRG1 SWI/SNF Remodeler Complex—We used affinity purification followed by mass spectrometry to identify proteins that bind to Smad2 and Smad3
Mass spectrometry analysis of the excised bands identified these proteins as the SWI/SNF complex subunits BRG1, BAF155, BAF170, and BAF250b in both eluates (Fig. 1A)

Summary

The abbreviations used are

TGF␤, transforming growth factor ␤; HA, hemagglutinin; ChIP, chromatin immunoprecipitation; siRNA, small interfering RNA; qRT, quantificative reverse transcription; shRNA, short hairpin RNA; T␤R-II, TGF␤ type II receptor; CBP, cAMP response element-binding protein (CREB)-binding protein. In a recent search for Smad-binding proteins, we noticed an interaction between Smad and several SWI/SNF components [10] In vitro this interaction was independent of the activation state of Smad, sharing this property with the interaction of Smad with the histone acetyl-transferase CBP [10]. An interaction between Smad and BRG1 in cell culture and a requirement for BRG1 in the activation of two TGF␤ target genes (lefty and nodal) were recently reported by others [11]. These observations suggested that Smad proteins may bring to DNA histone acetylating enzymes to relax the chromatin structure and SWI/SNF complexes to provide chromatin remodeling activity. The results provide a global view of the dependence of the TGF␤ transcriptional program on BRG1 SWI/SNF function

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION