The WASH complex, an endosomal Arp2/3 activator, interacts with the Hermansky-Pudlak syndrome complex BLOC-1 and its cargo phosphatidylinositol-4-kinase type IIα

Vesicle biogenesis machinery components such as coat proteins can interact with the actin cytoskeleton for cargo sorting into multiple pathways. It is unknown, however, whether these interactions are a general requirement for the diverse endosome traffic routes. In this study, we identify actin cytoskeleton regulators as previously unrecognized interactors of complexes associated with the Hermansky-Pudlak syndrome. Two complexes mutated in the Hermansky-Pudlak syndrome, adaptor protein complex-3 and biogenesis of lysosome-related organelles complex-1 (BLOC-1), interact with and are regulated by the lipid kinase phosphatidylinositol-4-kinase type II alpha (PI4KII alpha). We therefore hypothesized that PI4KII alpha interacts with novel regulators of these complexes. To test this hypothesis, we immunoaffinity purified PI4KII alpha from isotope-labeled cell lysates to quantitatively identify interactors. Strikingly, PI4KII alpha isolation preferentially coenriched proteins that regulate the actin cytoskeleton, including guanine exchange factors for Rho family GTPases such as RhoGEF1 and several subunits of the WASH complex. We biochemically confirmed several of these PI4KII alpha interactions. Of importance, BLOC-1 complex, WASH complex, RhoGEF1, or PI4KII alpha depletions altered the content and/or subcellular distribution of the BLOC-1-sensitive cargoes PI4KII alpha, ATP7A, and VAMP7. We conclude that the Hermansky-Pudlak syndrome complex BLOC-1 and its cargo PI4KII alpha interact with regulators of the actin cytoskeleton.