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.

Public Health Relevance

Recent studies have found that many human tumors, including common tumors of the brain, lung, breast, ovary and kidney harbor mutations within the subunits of a complex that uses energy provided by ATP to control the packaging and unpackaging of DNA in the nucleus. These BAF complexes unveil genes for their expression at specific times in development and in specific tissues. At present there is only rudimentary knowledge of the function of these complexes in human cells. Furthermore, it is not known how the mutations affect the normal functions of the complexes and how they cause cells to become malignant. Our studies will shed light upon the molecular mechanisms involved in tumor formation and open new therapeutic strategies for control of specific cancers.

National Institute of Health (NIH)
National Cancer Institute (NCI)
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Cancer Genetics Study Section (CG)
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Okano, Paul
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Stanford University
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Stanton, Benjamin Z; Hodges, Courtney; Crabtree, Gerald R et al. (2017) A General Non-Radioactive ATPase Assay for Chromatin Remodeling Complexes. Curr Protoc Chem Biol 9:1-10
Kadoch, Cigall; Williams, Robert T; Calarco, Joseph P et al. (2017) Dynamics of BAF-Polycomb complex opposition on heterochromatin in normal and oncogenic states. Nat Genet 49:213-222
Miller, Erik L; Hargreaves, Diana C; Kadoch, Cigall et al. (2017) TOP2 synergizes with BAF chromatin remodeling for both resolution and formation of facultative heterochromatin. Nat Struct Mol Biol 24:344-352
Stanton, Benjamin Z; Hodges, Courtney; Calarco, Joseph P et al. (2017) Smarca4 ATPase mutations disrupt direct eviction of PRC1 from chromatin. Nat Genet 49:282-288
Buscarlet, Manuel; Krasteva, Veneta; Ho, Lena et al. (2014) Essential role of BRG, the ATPase subunit of BAF chromatin remodeling complexes, in leukemia maintenance. Blood 123:1720-8
Bao, Xiaomin; Tang, Jiong; Lopez-Pajares, Vanessa et al. (2013) ACTL6a enforces the epidermal progenitor state by suppressing SWI/SNF-dependent induction of KLF4. Cell Stem Cell 12:193-203
Takebayashi, Shin-Ichiro; Lei, Ienglam; Ryba, Tyrone et al. (2013) Murine esBAF chromatin remodeling complex subunits BAF250a and Brg1 are necessary to maintain and reprogram pluripotency-specific replication timing of select replication domains. Epigenetics Chromatin 6:42
Staahl, Brett T; Tang, Jiong; Wu, Wei et al. (2013) Kinetic analysis of npBAF to nBAF switching reveals exchange of SS18 with CREST and integration with neural developmental pathways. J Neurosci 33:10348-61
Kadoch, Cigall; Hargreaves, Diana C; Hodges, Courtney et al. (2013) Proteomic and bioinformatic analysis of mammalian SWI/SNF complexes identifies extensive roles in human malignancy. Nat Genet 45:592-601
Dykhuizen, Emily C; Hargreaves, Diana C; Miller, Erik L et al. (2013) BAF complexes facilitate decatenation of DNA by topoisomerase II?. Nature 497:624-7

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