IGF2BP3 remodels the microRNA targeting landscape in MLL-AF4 leukemia

Insulin-like growth factor 2 mRNA binding protein 3 (IGF2BP3/I3) is a multi-domain RNA-binding protein required for MLL-AF4-driven leukemogenesis, but its mechanism of action remains enigmatic. We hypothesized from our previous work that I3 amplifies oncogenic gene expression by modulating RNA induced silencing complex (RISC) mRNA interactions. To test this, we performed miR-eCLIP of AGO2, the catalytic RISC subunit, in I3 knock-out (I3KO) as well as control B-cell acute lymphoblastic cell lines (B-ALL) and identified I3-dependent AGO2 binding sites on 111 3'UTRs. Analyzing chimeric miRNA-mRNA reads, we observed differential miRNA occupancy in the I3KO compared to control, including increased targeting by miR-181a, a regulator of leukocyte differentiation. Notably, miR-181a overexpression phenocopied I3 loss, implicating I3 in restricting miR-181a-mediated repression. Biochemical assays confirmed direct competition between I3 and AGO2-miRNA complexes for 3'-UTR binding. Taken together, our results provide a model for how I3 promotes leukemogenesis by antagonizing RISC-mediated repression of oncogenic mRNAs.