PROJECT SUMMARY/ABSTRACT Recent whole-exome sequencing studies in human cancer have unmasked frequent mutations in genes encoding chromatin regulatory proteins. Specifically, genes encoding subunits of the mammalian SWI/SNF ATP- dependent chromatin remodeling complexes (also called mSWI/SNF or BAF complexes) are perturbed in over 20% of human malignancy, underscoring their critical roles in the maintenance of timely and appropriate gene expression. A major question lies in the mechanisms by which mSWI/SNF complexes are targeted on chromatin. Indeed, studies by our group and others seek to systematically evaluate the role for each of the 29 subunits pieced together combinatorically in to 12-15 subunit entities, as well as the interfacial surfaces and domains that contribute specific binding interactions on chromatin, either by direct engagement with the histone landscape or via tethering to transcription factors. Human synovial sarcoma (SS) is uniformly driven by the t(X;18) chromosomal translocation, which results in the fusion of 78 amino acids of SSX to the C-terminus of the BAF complex subunit, SS18. The SS18-SSX fusion oncoprotein dominantly integrates in to BAF complexes, displacing the product of the remaining wild-type allele. This results in altered genome-wide distribution of BAF complexes on chromatin, suggesting that SS18-SSX actively recruits BAF complexes to specific features on the chromatin landscape. The mechanism underpinning this highly cancer-specific targeting of BAF complexes in synovial sarcoma remains unknown and represents a major barrier to progress in the field. Indeed, understanding the molecular basis for SSX-driven targeting of BAF complexes may facilitate the identification of new strategies for therapeutic intervention. As such, the goals of this proposal are to: (1) define the genomic targets of SS18-SSX-containing BAF complexes and the impact of this cancer-specific targeting on chromatin accessibility and gene expression in SS cell lines and primary tumors; (2) Identify SS18-SSX binding partners and histone landscape targets on mammalian nucleosomes that engage the 78aa SSX tail; (3) determine the features of the SSX 78aa tail required for SS-specific targeting on chromatin, gene expression, and maintenance of SS cell proliferation. Taken together, successful completion of these aims will provide a major and highly needed advance at the intersection of the chromatin regulation and sarcoma biology fields, and contribute important knowledge regarding the mechanism of targeting of BAF complexes across a range of both normal and oncogenic states. A major impediment to the development of on-target inhibitory agents of SS18-SSX function lie in the lack of biological understanding of SS18-SSX-mediated targeting. The results of this proposal are likely to provide the foundation for a new set of approaches toward targeted disruption of the interaction between SS18-SSX-containing BAF complexes and chromatin.