Distinct retrograde microtubule motor sets drive early and late endosome transport

Although subcellular positioning of endosomes significantly impacts on their functions, the molecular mechanisms governing the different steady‐state distribution of early endosomes (EEs) and late endosomes (LEs)/lysosomes (LYs) in peripheral and perinuclear eukaryotic cell areas, respectively, are still unsolved. We unveil that such differences arise because, while LE retrograde transport depends on the dynein microtubule (MT) motor only, the one of EEs requires the cooperative antagonism of dynein and kinesin‐14 KIFC1, a MT minus end‐directed motor involved in cancer progression. Mechanistically, the Ser‐x‐Ile‐Pro (SxIP) motif‐mediated interaction of the endoplasmic reticulum transmembrane protein stromal interaction molecule 1 (STIM1) with the MT plus end‐binding protein 1 (EB1) promotes its association with the p150Glued subunit of the dynein activator complex dynactin and the distinct location of EEs and LEs/LYs. The peripheral distribution of EEs requires their p150Glued‐mediated simultaneous engagement with dynein and SxIP motif‐containing KIFC1, via HOOK1 and HOOK3 adaptors, respectively. In sum, we provide evidence that distinct minus end‐directed MT motor systems drive the differential transport and subcellular distribution of EEs and LEs in mammalian cells.