Endosomal membrane state governs trafficking and cell migration under TFEB control

Abstract

Transcription Factor EB (TFEB) regulates the biogenesis of lysosomes, which are acidic organelles of the endosomal network. While its role in cellular clearance is well established, the pleiotropic functions of TFEB across diverse cellular processes, as well as its activation in several cancer types, remain incompletely understood. Investigating TFEB in a bladder cancer model, we found that its depletion selectively impaired cell migration and adhesion. This phenotype was associated with the retention of the adhesion molecule integrin β5 (ITGβ5), in central intracellular compartments identified as multivesicular bodies (MVBs) upon TFEB knockdown (KD). Mechanistically, TFEB regulated cellular lipid composition and membrane fluidity of acidic endosomes that controlled ITGβ5 trafficking. Remarkably, exogenous supplementation with a monounsaturated fatty acid (MUFA) that increased MVB membrane fluidity was sufficient to phenocopy the intracellular trapping of ITGβ5 in 2D culture and 3D tumoroids. We further showed that TFEB-dependent maintenance of MVB membrane fluidity relies on ESCRT-0 subcomplex, HGS/Hrs, which retains cholesterol at MVBs. Together, these findings reveal how transcriptional programs shape endosomal membrane states to control cargo trafficking and cell behavior. Summary We uncover a previously unrecognized role for TFEB in controlling the lipid composition and membrane fluidity of multivesicular bodies, establishing endosomal membrane properties as a regulated output of transcriptional programs that govern cargo trafficking and cell behavior.