TCF3::HLF Orchestrates an Enhancer-Promoter Network with Activation of MEF2C to Promote Immature HSC Gene Expression in Leukemia [HiChIP]

Oncogenic fusion transcription factors (TFs) frequently drive hematopoietic malignancies by altering gene expression in key developmental programs. TCF3::HLF is a fusion TF that characterizes a rare, treatment-resistant subtype of B-cell acute lymphoblastic leukemia (t(17;19) TCF3::HLF-positive B-ALL). Despite its clinical significance, the mechanisms by which TCF3::HLF induces leukemia are unclear. We used HiChIP mapping and genetic interference to analyze TCF3::HLF at the 3D-genome level, revealing enhancer-promoter interactions that control gene activation or repression. Notably, TCF3::HLF directly regulates MEF2C expression through its enhancer, as interference disrupted MEF2C transcription and inhibited leukemia propagation. This disruption also diminished embryonal hematopoietic stem cell (HSC) gene signatures and restored mature HSC and B-lymphoid markers. These findings highlight MEF2C as a critical component of the transcriptional network reprogrammed by TCF3::HLF. Our study provides insight into how TCF3::HLF rewires the 3D genome to drive leukemia and serves as a resource for further exploration of the TCF3::HLF regulome. Overall design: To delineate the 3D-genomic interactions orchestrated by the fusion protein TCF3::HLF in t(17;19) ALL cells, we performed MNase-HiChIP with two independent replicates using the TCF3::HLF positive cell line HAL-01, with the TCF3 wild-type allele knocked out, thereby enabling selective immunoprecipitation of the fusion protein when using antibodies against TCF3. We performed the HiChIP following the Dovetail®-HiChIP-MNase-Kit-User-Guide-Version-1.3. with modefications using E2A (CST #12258) antibody for pull-down. Interaction events were analyzed using FitHiChIP.