Alterations in BRG1 gene and epigenetic silencing of BRM inactivate the SWI/SNF complex in lung cancer

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SWI/SNF is a multimeric chromatin remodeling complex which plays a critical role in transcription. This complex specifically alters chromatin by shifting the position of the histones thereby changing the accessibility of specific DNA regions to key cellular proteins. This process can either facilitate or suppress gene expression. In yeast, the SWI/SNF complex is known to regulate approximately 5-7% of the genome. A number of tumor suppressor proteins and cell cycle regulatory proteins including p53, RB, BRCA1, and steroid receptors interact with and functionally require the SWI/SNF complex. These interactions are likely to be compromised when expression of one or more of the SWI/SNF subunits are lost. As we have found that loss of BRG1 and BRM occurs primarily in lung, head/neck, esophageal, ovarian, and prostate cancers, loss of SWI/SNF complex function may be important in the development of a number of tumor types. To understand the mechanism of BRG1 and BRM loss, we sequenced the BRG1 and BRM transcripts from ten cell lines deficient in BRG1 and BRM protein expression. In these cell lines, we found that the BRG1 gene had acquired either nonsense mutations, indels (frameshifting mutations) or defects in BRG1 splicing. In contrast, the BRM gene was essentially unaltered. We next investigated if epigenetic mechanisms were suppressing BRM expression, and treated these cell lines with either HDAC inhibitors or 5-aza-deoxycytidine (5-azaCdR). By semi-quantitative PCR, we found that both the BRG1 and BRM mRNAs were induced by HDAC inhibition; but by western blotting, only the BRM protein was upregulated. Treatment of these cell lines with 5-azaCdR had no appreciable effect on either BRG1 or BRM mRNA expression. To further investigate the mechanism of BRM regulation, we cloned the BRM promoter and measured promoter activity in BRG1/BRM positive and negative cell lines. In our dual luciferase assay system, we did not detect a significant change in relative transcriptional activity after treatment with HDAC inhibitors. These results show that BRG1 and BRM expressions are lost by different mechanisms. BRM mRNA is suppressed by epigenetic mechanisms and blocking HDAC activity restores BRM protein expression. As a number of growth control signaling pathways require the SWI/SNF complex for their function, reactivation of BRM containing SWI/SNF complexes may have important clinical implications.