title: Phosphorylation of Atg9 regulates movement to the phagophore assembly site and the rate of autophagosome formation.
abstract: Macroautophagy is primarily a degradative process that cells use to break down their own components to recycle macromolecules and provide energy under stress conditions, and defects in macroautophagy lead to a wide range of diseases. Atg9, conserved from yeast to mammals, is the only identified transmembrane protein in the yeast core macroautophagy machinery required for formation of the sequestering compartment termed the autophagosome. This protein undergoes dynamic movement between the phagophore assembly site (PAS), where the autophagosome precursor is nucleated, and peripheral sites that may provide donor membrane for expansion of the phagophore. Atg9 is a phosphoprotein that is regulated by the Atg1 kinase. We used stable isotope labeling by amino acids in cell culture (SILAC) to identify phosphorylation sites on this protein and identified an Atg1-independent phosphorylation site at serine 122. A nonphosphorylatable Atg9 mutant showed decreased autophagy activity, whereas the phosphomimetic mutant enhanced activity. Electron microscopy analysis suggests that the different levels of autophagy activity reflect differences in autophagosome formation, correlating with the delivery of Atg9 to the PAS. Finally, this phosphorylation regulates Atg9 interaction with Atg23 and Atg27.
authors.affiliation: a Life Sciences Institute and the Department of Molecular, Cellular and Developmental Biology, University of Michigan , Ann Arbor , MI , USA.
authors.forename: Yuchen
authors.initials: Y
authors.lastname: Feng
authors.affiliation: a Life Sciences Institute and the Department of Molecular, Cellular and Developmental Biology, University of Michigan , Ann Arbor , MI , USA.
authors.forename: Steven K
authors.initials: SK
authors.lastname: Backues
authors.affiliation: b Research Institute for Science and Technology, Kogakuin University , Tokyo , Japan.
authors.forename: Misuzu
authors.initials: M
authors.lastname: Baba
authors.affiliation: c Department of Cell Biology , Harvard Medical School , Boston , MA USA.
authors.forename: Jin-mi
authors.initials: JM
authors.lastname: Heo
authors.affiliation: c Department of Cell Biology , Harvard Medical School , Boston , MA USA.
authors.forename: J Wade
authors.initials: JW
authors.lastname: Harper
authors.affiliation: a Life Sciences Institute and the Department of Molecular, Cellular and Developmental Biology, University of Michigan , Ann Arbor , MI , USA.
authors.forename: Daniel J
authors.initials: DJ
authors.lastname: Klionsky
mesh_terms: D000595:Amino Acid Sequence; D000071182:Autophagosomes / Q000378:metabolism* / Q000648:ultrastructure; D001343:Autophagy; D000071183:Autophagy-Related Proteins / Q000737:chemistry / Q000378:metabolism*; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism*; D010766:Phosphorylation; D010768:Phosphoserine / Q000378:metabolism; D011485:Protein Binding; D021381:Protein Transport; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: C434065:ATG9 protein, S cerevisiae; D000071183:Autophagy-Related Proteins; D008565:Membrane Proteins; D029701:Saccharomyces cerevisiae Proteins; D010768:Phosphoserine
keywords: autophagy; lysosome; stress; vacuole; yeast