Recruitment of mSin3A and Histone Deacetylase 2 (HDAC2) by the Chromatin Remodeling Protein BRG1 Mediates Transcriptional Repression in Erythroid Progenitors.

The Swi/Snf chromatin remodeling complexes are critical for activating or repressing transcription of specific genes and require one of two homologous ATPases, Brg1 and Brm. Although Brg1 is also known to be involved in ß-globin gene transcription and a recent report showed that Brg1-containing complexes could be recruited to the ß-globin promoter and its locus control region (LCR), the exact roles of Brg1 in erythroid gene expression and differentiation have not been directly investigated. We previously showed that Brg1, but not Brm, is recruited to the Protein 4.2 (P4.2) promoter by a multi-protein DNA-binding complex containing TAL1/SCL, E47, GATA-1, LMO2, and Ldb1. This complex transactivates P4.2 gene expression through tandem E box-GATA elements in its proximal promoter. We report here that overexpression of Brg1 in murine erythroleukemia (MEL) cells significantly reduced the proportion of benzidine-stained cells and dramatically inhibited expression of mRNA for P4.2 and ß-globin but not glycophorin A and erythroid Kruppel-like factor in MEL cells induced to differentiate with dimethyl sulfoxide (DMSO). Further, overexpression of Brg1 repressed P4.2 promoter activity in a dose-dependent manner in DMSO-treated MEL cells, while treatment of cells with trichostatin A, a specific HDAC inhibitor, attenuated Brg1-directed inhibition of reporter activity in transient transfection analysis. In addition, coimmunoprecipitation analysis showed that Brg1 interacted stably with the corepressor mSin3A and HDAC2 in erythroid cells, suggesting that HDAC activity is involved in Brg1-mediated transcriptional repression. Quantitative chromatin immunoprecipitation (ChIP) analysis demonstrated that Brg1 occupancy of the P4.2 and ß-globin promoters and hypersensitive site 2 (HS2) of the ß-globin LCR decreased upon erythroid differentiation. Importantly, the selective transcriptional repression of P4.2 and ß-globin gene expression by Brg1 correlated with increased loading of mSin3A and HDAC2 on the P4.2 promoter and ß-globin HS2, respectively. Together, these results suggest that Brg1-mediated transcriptional repression of select erythroid genes is characterized by the localized recruitment of a corepressor complex containing mSin3A and HDAC2 in a process of active repression.