The in-frame internal tandem duplication (ITD) of the FLT3 gene is an important negative prognostic factor in acute myeloid leukemia (AML). FLT3-ITD is constitutive active and partially retained in the endoplasmic reticulum (ER). Recent reports show that 3'UTRs function as scaffolds that can regulate the localization of plasma membrane proteins by recruiting the HuR-interacting protein SET to the site of translation. Therefore, we hypothesized that SET could mediate the FLT3 membrane location and that the FLT3-ITD mutation could somehow disrupt the model, impairing its membrane translocation. Immunofluorescence and immunoprecipitation assays demonstrated that SET and FLT3 co-localize and interact in FLT3-WT cells but hardly in FLT3-ITD. SET/FLT3 interaction occurs before FLT3 glycosylation. Furthermore, RNA immunoprecipitation in FLT3-WT cells confirmed that this interaction occurs through the binding of HuR to the 3'UTR of FLT3. HuR inhibition and SET nuclear retention reduced FLT3 in the membrane of FLT3-WT cells, indicating that both proteins are involved in FLT3 membrane trafficking. Interestingly, the FLT3 inhibitor midostaurin increases FLT3 in the membrane and SET/FLT3 binding. Therefore, our results show that SET is involved in the transport of FLT3-WT to the membrane; however, SET barely binds FLT3 in FLT3-ITD cells, contributing to its retention in the ER.
Keywords: 3′-UTR; AML; FLT3; HuR; SET/I2PP2A.