Cancer cells frequently depend on chromatin regulatory activities to maintain a malignant phenotype. Here, we show that leukemia cells require the mammalian SWI/SNF chromatin remodeling complex for their survival and aberrant self-renewal potential. While Brg1, an ATPase subunit of SWI/SNF, is known to suppress tumor formation in several cell types, we found that leukemia cells instead rely on Brg1 to support their oncogenic transcriptional program, which includes Myc as one of its key targets. To account for this context-specific function, we identify a cluster of lineage-specific enhancers located 1.7 Mb downstream from Myc that are occupied by SWI/SNF as well as the BET protein Brd4. Brg1 is required at these distal elements to maintain transcription factor occupancy and for long-range chromatin looping interactions with the Myc promoter. Notably, these distal Myc enhancers coincide with a region that is focally amplified in~3% of acute myeloid leukemias. Together, these findings define a leukemia maintenance function for SWI/SNF that is linked to enhancer-mediated gene regulation, providing general insights into how cancer cells exploit transcriptional coactivators to maintain oncogenic gene expression programs.

Acetylation of histones triggers association with bromodomaincontaining proteins that regulate diverse chromatin-related processes. Although acetylation of transcription factors has been appreciated for some time, the mechanistic consequences are less well understood. The hematopoietic transcription factor GATA1 is acetylated at conserved lysines that are required for its stable association with chromatin. We show that the BET family protein Brd3 binds via its first bromodomain (BD1) to GATA1 in an acetylation-dependent manner in vitro and in vivo. Mutation of a single residue in BD1 that is involved in acetyl-lysine binding abrogated recruitment of Brd3 by GATA1, demonstrating that acetylation of GATA1 is essential for Brd3 association with chromatin. Notably, Brd3 is recruited by GATA1 to both active and repressed target genes in a fashion seemingly independent of histone acetylation. Anti-Brd3 ChIP followed by massively parallel sequencing in GATA1-deficient erythroid precursor cells and those that are GATA1 replete revealed that GATA1 is a major determinant of Brd3 recruitment to genomic targets within chromatin. A pharmacologic compound that occupies the acetyl-lysine binding pockets of Brd3 bromodomains disrupts the Brd3-GATA1 interaction, diminishes the chromatin occupancy of both proteins, and inhibits erythroid maturation. Together these findings provide a mechanism for GATA1 acetylation and suggest that Brd3 "reads" acetyl marks on nuclear factors to promote their stable association with chromatin.hematopoiesis | posttranslational modifications | gene regulation P osttranslational acetylation of lysine residues is a widely used cellular signaling mechanism involving histone and nonhistone proteins. Typically, acetylated lysines (acK) serve as docking sites for bromodomains that are found in numerous chromatinrelated proteins (1, 2). Thus, bromodomains serve as "readers" of acK marks in a cascade of signaling events that influence diverse processes within chromatin. The hematopoietic zinc finger protein GATA1 was one of the earliest reported acetylated transcription factors (3, 4). GATA1 is a pivotal regulator of the erythroid, megakaryocyte, and mast cell lineages (5-9). GATA1 activates all known erythroid-specific genes, and mice lacking GATA1 succumb to anemia due to failed maturation and apoptosis of erythroid precursor cells (5, 10, 11). GATA1 also functions as a repressor, down-regulating genes associated with the proliferative, immature state. Mutations in the zinc finger region of GATA1 underlie congenital anemias and thrombocytopenias (12), whereas alterations at the N terminus of GATA1 are associated with megakaryoblastic leukemias in patients with Down syndrome (13). Previously, we found that GATA1 interacts with the acetyltransferase CREB binding protein (CBP) (14) to stimulate histone acetylation at GATA1 target genes (15, 16). CBP and its paralog p300 also acetylate GATA1 at two lysine-rich clusters C-terminal to each of its zinc fingers (Fig. 1A) (3, 4). CBPmediated acetylation of GATA1 c...

Facioscapulohumeral muscular dystrophy (FSHD) is a common, dominantly inherited disease caused by the epigenetic de-repression of the DUX4 gene, a transcription factor normally repressed in skeletal muscle. As targeted therapies are now possible in FSHD, a better understanding of the relationship between DUX4 activity, muscle pathology and muscle magnetic resonance imaging (MRI) changes is crucial both to understand disease mechanisms and for the design of future clinical trials. Here, we performed MRIs of the lower extremities in 36 individuals with FSHD, followed by needle muscle biopsies in safely accessible muscles. We examined the correlation between MRI characteristics, muscle pathology and expression of DUX4 target genes. Results show that the presence of elevated MRI short tau inversion recovery signal has substantial predictive value in identifying muscles with active disease as determined by histopathology and DUX4 target gene expression. In addition, DUX4 target gene expression was detected only in FSHD-affected muscles and not in control muscles. These results support the use of MRI to identify FSHD muscles most likely to have active disease and higher levels of DUX4 target gene expression and might be useful in early phase therapeutic trials to demonstrate target engagement in therapies aiming to suppress DUX4 expression.

Facioscapulohumeral dystrophy (FSHD) is caused by the mis-expression of DUX4 in skeletal muscle cells. DUX4 is a transcription factor that activates genes normally associated with stem cell biology and its mis-expression in FSHD cells results in apoptosis. To identify genes and pathways necessary for DUX4-mediated apoptosis, we performed an siRNA screen in an RD rhabdomyosarcoma cell line with an inducible DUX4 transgene. Our screen identified components of the MYC-mediated apoptotic pathway and the double-stranded RNA (dsRNA) innate immune response pathway as mediators of DUX4-induced apoptosis. Further investigation revealed that DUX4 expression led to increased MYC mRNA, accumulation of nuclear dsRNA foci, and activation of the dsRNA response pathway in both RD cells and human myoblasts. Nuclear dsRNA foci were associated with aggregation of the exon junction complex component EIF4A3. The elevation of MYC mRNA, dsRNA accumulation, and EIF4A3 nuclear aggregates in FSHD muscle cells suggest that these processes might contribute to FSHD pathophysiology.