Chromatin, Gene, and RNA Regulation SNF5 Reexpression in Malignant Rhabdoid Tumors Regulates Transcription of Target Genes by Recruitment of SWI/SNF Complexes and RNAPII to the Transcription Start Site of Their Promoters

Malignant rhabdoid tumor (MRT), a highly aggressive cancer of young children, displays inactivation or loss of the hSNF5/INI1/SMARCB1 gene, a core subunit of the SWI/SNF chromatin-remodeling complex, in primary tumors and cell lines. We have previously reported that reexpression of hSNF5 in someMRT cell lines causes a G1 arrest via p21 (p21) mRNA induction in a p53-independent manner. However, the mechanism(s) by which hSNF5 reexpression activates gene transcription remains unclear. We initially searched for other hSNF5 target genes by asking whether hSNF5 loss altered regulation of other consensus p53 target genes. Our studies show that hSNF5 regulates only a subset of p53 target genes, including p21 andNOXA, inMRT cell lines.We also show that hSNF5 reexpression modulates SWI/SNF complex levels at the transcription start site (TSS) at both loci and leads to activation of transcription initiation through recruitment of RNApolymerase II (RNAPII) accompanied by H3K4 and H3K36 modifications. Furthermore, our results show lower NOXA expression in MRT cell lines compared with other human tumor cell lines, suggesting that hSNF5 loss may alter the expression of this important apoptotic gene. Thus, one mechanism for MRT development after hSNF5 loss may rely on reduced chromatinremodeling activity of the SWI/SNF complex at the TSS of critical gene promoters. Furthermore, because we observe growth inhibition after NOXA expression in MRT cells, the NOXA pathway may provide a novel target with clinical relevancy for treatment of this aggressive disease. Mol Cancer Res; 11(3); 1–10. 2013 AACR. Introduction Malignant rhabdoid tumor (MRT) is a rare and extremely aggressive childhood cancer, originally described as an unfavorable histologic variant of the pediatric renalWilms' tumor (1). The most common locations are in the kidney and central nervous system, although MRTs may arise in almost any site (2, 3). Despite significant advances in treatment, for MRTs diagnosed before the age of 6months, patient survival at 4 years drops to approximately 8.8% (4). Therefore, improved patient outcome requires a better understanding ofMRT and the development of novel therapeutic strategies. The common genetic alteration in MRTs is inactivation of hSNF5 (also known as SMARCB1, INI1, and BAF47), located in chromosome band 22q11.2 (5), which implicates the loss of hSNF5 function as the primary cause of these tumors (6). Therefore, the elucidation of hSNF5 function should lead to the identification of the key molecular steps necessary for MRT tumorigenesis. hSNF5 is a component of the SWI/SNF chromatinremodeling complex. SWI/SNF complexes are ATP-dependent chromatin-remodeling complexes that regulate gene transcription by causing conformational changes in chromatin structure (7). SWI/SNF subunits can be subclassified into 3 categories: (i) ATPase subunit (either BRG1or BRM), (ii) core subunits (hSNF5, BAF155, and BAF170), and (iii) accessory subunits (BAF53, BAF57, BAF180, and others; ref. 8). How loss of the core SNF5 subunit leads to the development of a rare pediatric cancer remains one of the most challenging questions in the cancer epigenetic field. Our previous study has shown that while hSNF5 reexpression in MRT cells increases both p21 and p16 expression during the induction of G1 cell-cycle arrest, p21 upregulation precedes p16. Furthermore, we have showed that p21 transcription shows both p53-dependent and -independent mechanisms of induction after hSNF5 reexpression. hSNF5 was confirmed to bind to the p21 promoter by chromatin immunoprecipitation (ChIP) analysis (9). However, little is known about how hSNF5 activates transcription at its target promoters. Authors' Affiliations: Lineberger Comprehensive Cancer Center; Curriculums in Toxicology and Genetics andMolecular Biology; Department of Pathology and Laboratory Medicine, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina; and Department of Pediatrics, Kyoto Prefectural University of Medicine, Kyoto, Japan Corresponding Author: Bernard E. Weissman, Lineberger Comprehensive Cancer Center, Room 32-048, University of North Carolina, 450 West Drive, Chapel Hill, NC 27599. Phone: 919-966-7533; Fax: 919-966-3015; E-mail: [email protected] doi: 10.1158/1541-7786.MCR-12-039