The ncBAF complex regulates transcription in AML through H3K27ac sensing by BRD9 (TT-Seq)
The non-canonical BAF complex (ncBAF) subunit BRD9 is essential for acute myeloid leukemia (AML) cell viability but has an unclear role in leukemogenesis. Because BRD9 is required for ncBAF complex assembly through its DUF3512 domain, precise bromodomain inhibition is necessary to parse the role of BRD9 as a transcriptional regulator from that of a scaffolding protein. To understand the specific role of BRD9 bromodomain function in regulating AML, we selected a panel of five AML cell lines with distinct driver mutations, disease classifications, and genomic aberrations and subjected them to short-term BRD9 bromodomain inhibition. We characterized the bromodomain-dependent growth of these cell lines, identifying a dependency in AML cell lines but not HEK293T cells. To define a mechanism through which BRD9 maintains AML cell survival, we examined nascent transcription, chromatin accessibility, and ncBAF complex binding genome-wide after bromodomain inhibition. We identified extensive regulation of transcription by BRD9 bromodomain activity, including repression of myeloid maturation factors and tumor suppressor genes, while standard AML chemotherapy targets (Bcl2, Btk, Kit) were repressed by inhibition of the BRD9 bromodomain. BRD9 bromodomain activity maintained accessible chromatin at both gene promoters and gene-distal putative enhancer regions, in a manner that qualitatively correlated with enrichment of BRD9 binding. Furthermore, we identified reduced chromatin accessibility at GATA, ETS, and AP-1 motifs and increased chromatin accessibility at SNAIL-, HIC-, and TP53-recognized motifs after BRD9 inhibition. We interpret these data as signifying a role for BRD9 in regulating the differentiation block that is common to all AML subtypes through modulation of accessibility at hematopoietic transcription factor binding sites. Overall design: Nascent RNA-seq (TT-seq) samples, treated for 6 hours with either the BRD9 bromodomain inhibitor  I-BRD9 or vehicle control (DMSO). Five different AML cell lines are profiled in duplicate (Kasumi-1, KG-1, ML-1, MV-4-11, and U937).