Dynactin interaction with AP-2 adaptor complex requires CLIP-170 and autophagy

Abstract

AbstractThe endocytic adaptor protein 2 (AP-2) complex binds dynactin as part of its noncanonical function, which is necessary for dynein-driven autophagosome transport along microtubules in neuronal axons. The absence of this AP-2-dependent transport causes neuronal morphology simplification and neurodegeneration. The mechanisms that lead to formation of the AP-2– dynactin complex have not been studied to date. However, inhibition of the mammalian/mechanistic target of rapamycin complex 1 (mTORC1) enhances the transport of newly formed autophagosomes, by influencing the biogenesis and protein interactions of Rab- interacting lysosomal protein (RILP), another dynein cargo adaptor. We tested the effects of mTORC1 inhibition on interactions between the AP-2 and dynactin complexes, with a focus on their two essential subunits, AP-2β and p150Glued. We found that the mTORC1 inhibitor rapamycin enhanced AP-2–dynactin complex formation in both neurons and non-neuronal cells. The live imaging of neuronal axons revealed that when combined with brain-derived neurotrophic factor (BDNF), an agonist of tropomyosin receptor kinase B (TrkB), rapamycin also increased the number of retrogradely moving mobile AP-2β-p150Glued complexes. Additional analysis revealed that the AP-2β–p150Glued interaction was indirect and required integrity of the dynactin complex. Rapamycin-driven enhancement of the AP-2–dynactin interaction also required the presence of cytoplasmic linker protein 170 (CLIP-170) and activation of autophagy. The latter was sufficient to enhance the AP-2β interaction with p150Glued, even when mTORC1 was active. Altogether, our results show that autophagy regulates the AP-2–dynactin interaction to coordinate sufficient motor-adaptor availability for newly generated autophagosomes.