α-Arrestins maintain phospholipid balance and Atg18 distribution to permit efficient autophagy

Abstract

Cells selectively reorganize their membrane proteome in response to stressors via selective protein trafficking. The α-arrestins, a family of conserved protein trafficking adaptors, bind to select membrane proteins and interact with the ubiquitin ligase Rsp5. The α-arrestins recruit Rsp5 to its membrane protein substrates, permitting their ubiquitination and endocytosis. To identify new α-arrestin functions, we performed a genetic screen to isolate mutants that alter α-arrestin-mediated resistance to rapamycin, a drug that inhibits TORC1. Interestingly, loss of many of the ATG genes, which encode the machinery needed for the self-degradative process of autophagy, disrupted α-arrestins’ ability to promote growth on rapamycin. Herein we define a genetic network linking α-arrestins to autophagy. We show autophagy impairment in the absence of select α-arrestins, with increased autophagosome lifetimes and delayed/reduced delivery of autophagosomes to the vacuole. The α-arrestin mutants that impeded autophagy had vacuole morphology defects and increased vacuolar retention of Atg18, a member of the PROPPIN family that is needed to maintain vacuole shape and facilitate lipid transfer to expanding autophagosomes. Atg18 binds phosphatidylinositol 3 phosphate (PI3P) and phosphatidylinositol 3,5-bisphosphate (PI(3,5)P2), and we observed increased PI3P on the vacuole membrane in α-arrestin mutants. The levels of Vps34 and Fab1, the kinases responsible for the generation of PI3P and PI(3,5)P2, respectively, were also elevated at vacuole membranes in cells lacking α-arrestins. We posit that altered phospholipids in the vacuolar membrane form the basis for the Atg18-Atg2 mislocalization and autophagy defect. These data demonstrate a previously unappreciated link between the α-arrestins and autophagy, expanding the functional impact of these trafficking adaptors in responding to nutrient stress.Author SummaryCells survive nutrient starvation by degrading parts of themselves through the process of autophagy. During autophagy, cells make a double membrane, known as an autophagosome (AP), around bits of cytoplasm or organelles. The AP and its engulfed material are delivered to the vacuole, an organelle that helps break down proteins and lipids. These materials can then be used as building blocks to generate the essential components needed for the cell to survive starvation. For cells to undergo efficient autophagy, they need α-arrestins, a group of proteins important for deciding where membrane proteins localize. In cells lacking α-arrestins, the AP forms slowly, likely due to a problem in growing the AP membrane. This results in less material being delivered to the vacuole via APs when cells do not have α-arrestins. This study defines a new role for α-arrestins in promoting AP formation and starvation survival.