title: Methods to Assess Autophagy and Chronological Aging in Yeast.
abstract: Autophagy is a catabolic process that is crucial for cellular homeostasis and adaptive response to changing environments. Importantly, autophagy has been shown to be induced in many longevity-associated scenarios and to be required to maintain lifespan extension. Notably, autophagy is a highly conserved cellular process among eukaryotes, and the yeast Saccharomyces cerevisiae has become a universal model system for unraveling the molecular machinery underlying autophagic mechanisms. Here, we discuss different protocols to monitor survival and autophagy of yeast cells upon chronological aging. These include the use of propidium iodide to assess the loss of cell membrane integrity, as well as clonogenic assays to directly determine survival rates. Additionally, we describe methods to quantify autophagic flux, including the alkaline phosphatase activity or the GFP liberation assays, which measure the delivery of autophagosomal cargo to the vacuole. In sum, we have recapped established protocols used to evaluate a link between lifespan extension and autophagy in yeast.
authors.affiliation: Institute of Molecular Biosciences, NAWI Graz, University of Graz, Graz, Austria.
authors.forename: K
authors.initials: K
authors.lastname: Kainz
authors.affiliation: Institute of Molecular Biosciences, NAWI Graz, University of Graz, Graz, Austria.
authors.forename: J
authors.initials: J
authors.lastname: Tadic
authors.affiliation: Institute of Molecular Biosciences, NAWI Graz, University of Graz, Graz, Austria.
authors.forename: A
authors.initials: A
authors.lastname: Zimmermann
authors.affiliation: Institute of Molecular Biosciences, NAWI Graz, University of Graz, Graz, Austria.
authors.forename: T
authors.initials: T
authors.lastname: Pendl
authors.affiliation: Institute of Molecular Biosciences, NAWI Graz, University of Graz, Graz, Austria.
authors.forename: D
authors.initials: D
authors.lastname: Carmona-Gutierrez
authors.affiliation: Institute of Molecular Biosciences, NAWI Graz, University of Graz, Graz, Austria.
authors.forename: C
authors.initials: C
authors.lastname: Ruckenstuhl
authors.affiliation: Institute of Molecular Biosciences, NAWI Graz, University of Graz, Graz, Austria; BioTechMed-Graz, Graz, Austria.
authors.forename: T
authors.initials: T
authors.lastname: Eisenberg
authors.affiliation: Institute of Molecular Biosciences, NAWI Graz, University of Graz, Graz, Austria; BioTechMed-Graz, Graz, Austria. Electronic address: frank.madeo@uni-graz.at.
authors.forename: F
authors.initials: F
authors.lastname: Madeo
mesh_terms: D000469:Alkaline Phosphatase / Q000032:analysis / Q000378:metabolism; D001343:Autophagy; D000071190:Autophagy-Related Protein 8 Family / Q000032:analysis / Q000378:metabolism; D015153:Blotting, Western / Q000379:methods; D057075:Enzyme Assays / Q000379:methods; D049452:Green Fluorescent Proteins / Q000032:analysis / Q000378:metabolism; D008856:Microscopy, Fluorescence / Q000379:methods; D011419:Propidium / Q000378:metabolism; D059748:Proteolysis; D012441:Saccharomyces cerevisiae / Q000166:cytology* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000032:analysis / Q000378:metabolism
chemical_list: C113757:ATG8 protein, S cerevisiae; D000071190:Autophagy-Related Protein 8 Family; D029701:Saccharomyces cerevisiae Proteins; D049452:Green Fluorescent Proteins; D011419:Propidium; D000469:Alkaline Phosphatase
keywords: Alkaline phosphatase; Autophagy; CLS; GFP-Atg8; Longevity; Yeast; Yeast survival