Epigenetic dysregulation is a major driver of tumorigenesis. The most commonly mutated epigenetic modifier in human cancer is the BAF complex (Brahma Associated Factors). BAF is an evolutionarily-conserved multi- subunit ATP-dependent chromatin remodeling complex that is mutated in one fifth of all cancers. BAF mobilizes nucleosomes to expose the underlying DNA in order to regulate access of DNA binding sites to transcriptional machinery, and thus functions as a major regulator of transcription. The enzymatic activity of the complex is provided by one of two mutually exclusive catalytic ATPases, BRG1 or BRM, with additional subunits regulating the targeting and enzymatic activity of the complex. Despite the frequency of BAF mutations in cancer, little is known about how cancer-associated BAF mutations affect the nucleosome remodeling activity of the complex or contribute to tumorigenesis. Determining the effects of these mutations is hindered by our lack of knowledge regarding the normal activity and regulation of the complex. This study will address this gap in knowledge by using purified recombinant BAF complexes to assess the normal regulation of the complex as well as its misregulation in cancer. This study will leverage the known regulatory features of RSC, a yeast homolog of BAF, and test whether they are conserved in humans. The function of the Actin/BAF53 module as well as the roles of the Post-HSA and Protrusion 1 domains of BRG1 and BRM in the catalytic activity of the complex will be assessed. The Post-HSA and Protrusion 1 domains of BRG1 and BRM are frequently mutated in cancer; the effects of these mutations on the remodeling activity of the complex will be examined as well. The most clearly oncogenic BAF alterations are the uniform loss of the SNF5 subunit in malignant rhabdoid tumor (MRT) and the fusion between the SS18 BAF subunit and the SSX protein in synovial sarcoma (SS), which is reported to evict SNF5 from the BAF complex. MRT and SS are among the most genetically simple tumors, each typically bearing only the single genetic alteration listed above. This lack of additional mutations suggests that SNF5 loss and the SS18-SSX fusion protein promote tumor formation through an epigenetic mechanism. Importantly, SNF5 affects the catalytic activity of BRG1, suggesting that aberrant enzymatic activity of the complex is the epigenetic driver of tumorigenesis in MRT and SS. The tumor specificity of these mutations indicates that SS18-SSX fusions and SNF5 loss are not equivalent. The nucleosome remodeling activities of recombinant BAF complexes containing or lacking SNF5 and SS18-SSX fusions will be assessed to determine whether the tissue specificity of these mutations may be attributed to a difference in catalytic activity. The results of this study will provide insight into the normal function of a major epigenetic modifier as well as its misregulation in cancer, and may inform new therapeutic strategies in a wide variety of tumors. !
The BAF (Brahma Associated Factors) ATP-dependent chromatin remodeling complex is a major regulator of transcription that is mutated in one fifth of human cancers. It is not known how these mutations affect the activity of the complex or contribute to tumorigenesis. This study aims to elucidate the regulatory logic of the BAF complex and its misregulation in cancer, deepening our understanding of the mechanisms of epigenetic dysregulation in cancer and potentially informing new therapeutic strategies for a wide variety of cancers.