ATP-Dependent Chromatin Remodeling in Human Malignancy

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DESCRIPTION (provided by applicant): Recent screens for driving mutations in human malignancy have repeatedly identified subunits of mammalian SWI/SNF-like BAF complexes as tumor suppressors. Biochemical studies indicate that the identified subunits: BAF250a, Brg (BAF190), BAF155, BAF60b, BAF53a, and BAF47 (hSNF5) are dedicated to these complexes and not found as individual proteins or as parts of other complexes. In addition, we have recently found that SS18 (mutated in Synovial Sarcoma), Bcl7 and Bcl11 appear to be dedicated, stable subunits of BAF complexes. These studies suggest that SWI/SNF-like BAF complexes might be one of the most commonly mutated chromatin regulators in human cancer. BAF complexes regulate chromatin structure and are composed of about 14 subunits that are combinatorially assembled from the products of gene families encoding the subunits. The mechanisms underlying their frequent mutation in cancer are unclear. We have found that conditional deletion or depletion of the oncogenic subunits leads to stalling in mitosis and anaphase bridge formation, strongly implicating a failure to decatenate DNA during M phase. Importantly, Topoisomerase IIa (Topo IIa), which resolves catenated DNA at M phase associates with BAF complexes and Brg is essential for chromatin binding by Topo IIa. Furthermore, purified BAF complexes are required for optimal decatenation by purified Topo IIa in vitro. These observations suggest that a failure of decatenation by Topo IIa contributes to the genesis of human cancers, which is the central hypothesis of this application. This hypothesis is supported by the high frequency of concurrent mutations in other genes or aneuploidy in tumors bearing apparent initiating mutations in BAF subunits. To study these oncogenic BAF mutations, the SS18 translocation to SSX is particularly useful since it produces a sterotypic in-frame fusion of the SS18 BAF subunit to a member of the SSX gene family located on the X chromosome. This precise translocation attaches 78aa of SSX to SS18 and is almost certainly the driving event in synovial sarcoma (SS), which account for about 8% of sarcomas. Remarkably, the translocation of one allele leads to partial dissolution of BAF complexes. We will begin our studies by determining whether the SS18-SSX fusion causes a loss or gain of function for BAF complexes. We will define the consequences of this translocation for BAF complex binding over the genome to determine if target genes are lost or gained. We will then determine why the wildtype SS18 allele is repressed and leads to the formation of little protein compared to the translocated allele. We will examine the potentially oncogenic role of Topo IIa's dependence upon BAF complexes. Finally, we will attempt to understand how activating mutations in a- catenin and PI3K apparently cooperate with loss-of-function mutations in BAF subunits to lead to cancer. At the conclusion of our work we expect to have gained insight into the mechanism of transformation by mutations of the subunits of BAF complexes, which are emerging as major contributors to human cancer.