title: PEP3 overexpression shortens lag phase but does not alter growth rate in Saccharomyces cerevisiae exposed to acetic acid stress.
abstract: In fungi, two recognized mechanisms contribute to pH homeostasis: the plasma membrane proton-pumping ATPase that exports excess protons and the vacuolar proton-pumping ATPase (V-ATPase) that mediates vacuolar proton uptake. Here, we report that overexpression of PEP3 which encodes a component of the HOPS and CORVET complexes involved in vacuolar biogenesis, shortened lag phase in Saccharomyces cerevisiae exposed to acetic acid stress. By confocal microscopy, PEP3-overexpressing cells stained with the vacuolar membrane-specific dye, FM4-64 had more fragmented vacuoles than the wild-type control. The stained overexpression mutant was also found to exhibit about 3.6-fold more FM4-64 fluorescence than the wild-type control as determined by flow cytometry. While the vacuolar pH of the wild-type strain grown in the presence of 80 mM acetic acid was significantly higher than in the absence of added acid, no significant difference was observed in vacuolar pH of the overexpression strain grown either in the presence or absence of 80 mM acetic acid. Based on an indirect growth assay, the PEP3-overexpression strain exhibited higher V-ATPase activity. We hypothesize that PEP3 overexpression provides protection from acid stress by increasing vacuolar surface area and V-ATPase activity and, hence, proton-sequestering capacity.
authors.affiliation: Department of Biochemistry and Biophysics, Oregon State University, Corvallis, OR, USA.|Department of Food Science and Technology, Oregon State University, Corvallis, OR, USA.
authors.forename: Jun
authors.initials: J
authors.lastname: Ding
authors.affiliation: Department of Food Science and Technology, Oregon State University, Corvallis, OR, USA.|Department of Microbiology, Oregon State University, Corvallis, OR, USA.
authors.forename: Garrett
authors.initials: G
authors.lastname: Holzwarth
authors.affiliation: Department of Environmental & Molecular Toxicology, Oregon State University, Corvallis, OR, USA.
authors.forename: C Samuel
authors.initials: CS
authors.lastname: Bradford
authors.affiliation: Biological Sciences, Duquesne University, Pittsburgh, PA, USA.
authors.forename: Ben
authors.initials: B
authors.lastname: Cooley
authors.affiliation: Department of Food Science and Technology, Oregon State University, Corvallis, OR, USA.|Department of Microbiology, Oregon State University, Corvallis, OR, USA.
authors.forename: Allen S
authors.initials: AS
authors.lastname: Yoshinaga
authors.affiliation: Biological Sciences, Duquesne University, Pittsburgh, PA, USA.
authors.forename: Jana
authors.initials: J
authors.lastname: Patton-Vogt
authors.affiliation: Department of Biochemistry and Food Science, Hebrew University, Rehovot, Israel.
authors.forename: Hagai
authors.initials: H
authors.lastname: Abeliovich
authors.affiliation: Department of Food Science and Technology, Oregon State University, Corvallis, OR, USA.
authors.forename: Michael H
authors.initials: MH
authors.lastname: Penner
authors.affiliation: Department of Biochemistry and Biophysics, Oregon State University, Corvallis, OR, USA. alan.bakalinsky@oregonstate.edu.|Department of Food Science and Technology, Oregon State University, Corvallis, OR, USA. alan.bakalinsky@oregonstate.edu.|Department of Microbiology, Oregon State University, Corvallis, OR, USA. alan.bakalinsky@oregonstate.edu.
authors.forename: Alan T
authors.initials: AT
authors.lastname: Bakalinsky
mesh_terms: D019342:Acetic Acid / Q000633:toxicity*; D033942:Adaptor Proteins, Vesicular Transport / Q000096:biosynthesis* / Q000235:genetics; D015870:Gene Expression; D006863:Hydrogen-Ion Concentration; D012441:Saccharomyces cerevisiae / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development* / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins / Q000096:biosynthesis* / Q000235:genetics; D013312:Stress, Physiological; D025262:Vacuolar Proton-Translocating ATPases / Q000378:metabolism; D014617:Vacuoles / Q000378:metabolism
chemical_list: D033942:Adaptor Proteins, Vesicular Transport; C071657:PEP3 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D025262:Vacuolar Proton-Translocating ATPases; D019342:Acetic Acid
keywords: Acetic acid; CORVET; HOPS; PEP3; PEP5; STM1; Saccharomyces cerevisiae; V-ATPase; Vacuole; Yeast