Targeting the HBZ Viral Oncoprotein Transcriptional Network in Adult T-cell leukemia/lymphoma (ChIP-Seq). Homo sapiens

Adult T-cell Leukemia/Lymphoma (ATLL) is a frequently incurable disease associated with the human lymphotropic virus type I (HTLV-I). The transcription factor HBZ is the only virally encoded gene that is expressed in all ATLL cases, but it is unclear why it may be essential in ATLL and how it might be targeted therapeutically. Here we performed RNA interference screening of ATLL lines and discovered that their proliferation depends on the transcription factors BATF3 and IRF4, which regulate the identity of normal immune cells. These factors, which are highly expressed in ATLL, cooperatively bind and transactivate genes that distinguish ATLL from other T cell malignancies, including BATF3 itself. HBZ binds to an ATLL-specific BATF3 super-enhancer and thereby regulates the expression of BATF3 and its downstream targets, including the oncogene MYC. The BET protein inhibitor JQ1 collapsed the transcriptional network directed by HBZ and BATF3, and was consequently toxic for ATLL lines and patient samples in vitro, and blocked growth of ATLL xenografts. Our study demonstrates that the HTLV-I virus exploits a regulatory module that can potentially be attacked therapeutically with BET protein inhibitors. Overall design: For IRF4 ChIP-seq (n=4), KK1 and ST1 cell line chromatins were immuno-precipitated with normal rabbit IgG (Santa Cruz Biotechnology, sc2027) or IRF4 antibody (Santa Cruz Biotechnology, sc6059 and sc11450 (1:1 mix)). For BATF3 ChIP-seq (n=4), KK1 and ST1 cell line chromatins were immuno-precipitated with normal rabbit IgG (Santa Cruz Biotechnology, sc2027) or BATF3 antibody (R&D, AF7437). For BirA ChIP-seq (n=8), KK1 and ST1 cell lines were engineered to express BirA, transduced with Ctrl, HBZ (pRCMV/TO-flag-BIOTIN-HBZ-puro) or BATF3 (pRCMV/TO-flag-BIOTIN-BATF3-puro) Biotag constructs, and ChIP-seq performed using streptavidin to purify chromatin bound to these isoforms. For H3K27ac and BRD4 ChIP-seq (n=8), KK1 and ST1 cells were pre-treated with either DMSO or JQ1 before being cross-linked and chromatins were later immuno-precipitated with BRD4 antibody (Bethyl, Cat No. A301-985A) or H3K27ac antibody (Abcam, ab4729).