title: Loss of vacuolar H+-ATPase activity in organelles signals ubiquitination and endocytosis of the yeast plasma membrane proton pump Pma1p.
abstract: Yeast mutants lacking the intracellular V-ATPase proton pump (vma mutants) have reduced levels of the Pma1p proton pump at the plasma membrane and increased levels in organelles including the vacuolar lumen. We examined the mechanism and physiological consequences of Pma1p mislocalization. Pma1p is ubiquitinated in vma mutants, and ubiquitination depends on the ubiquitin ligase Rsp5p and the arrestin-related adaptor protein Rim8p. vma mutant strains containing rsp5 or rim8 mutations maintain Pma1p at the plasma membrane, suggesting that ubiquitination is required for Pma1p internalization. Acute inhibition of V-ATPase activity with concanamycin A triggers Pma1p ubiquitination and internalization. In an endocytosis-deficient mutant (end4Δ) Pma1p is ubiquitinated but retained at the plasma membrane during concanamycin A treatment. Consistent with specificity in signaling loss of V-ATPase activity to Pma1p, another plasma membrane transporter, Mup1p, is not internalized in a vma mutant, and loss of the Mup1p adaptor Art1p does not prevent Pma1p internalization in a vma mutant. Very poor growth of vma2 rsp5-1 and vma2 rim8Δ double mutants suggests that Pma1p internalization benefits the vma mutants. We hypothesize that loss of V-ATPase-mediated organelle acidification signals ubiquitination, internalization, and degradation of a portion of Pma1p as a means of balancing overall pH homeostasis.
authors.affiliation: Department of Biochemistry and Molecular Biology, SUNY Upstate Medical University, Syracuse, New York 13210.
authors.forename: Anne M
authors.initials: AM
authors.lastname: Smardon
authors.affiliation: Department of Biochemistry and Molecular Biology, SUNY Upstate Medical University, Syracuse, New York 13210. Electronic address: kanepm@upstate.edu.
authors.forename: Patricia M
authors.initials: PM
authors.lastname: Kane
mesh_terms: D018797:Cell Cycle Proteins; D002462:Cell Membrane / Q000201:enzymology*; D004705:Endocytosis; D056827:Endosomal Sorting Complexes Required for Transport / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D047908:Intracellular Signaling Peptides and Proteins / Q000378:metabolism; D017136:Ion Transport; D018942:Macrolides / Q000737:chemistry; D008856:Microscopy, Fluorescence; D009154:Mutation; D009841:Oligonucleotides / Q000737:chemistry; D006180:Proton-Translocating ATPases / Q000235:genetics / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D025801:Ubiquitin / Q000378:metabolism*; D043743:Ubiquitin-Protein Ligase Complexes / Q000378:metabolism; D025262:Vacuolar Proton-Translocating ATPases / Q000378:metabolism*; D014617:Vacuoles / Q000378:metabolism
chemical_list: D018797:Cell Cycle Proteins; D056827:Endosomal Sorting Complexes Required for Transport; D047908:Intracellular Signaling Peptides and Proteins; D018942:Macrolides; D009841:Oligonucleotides; C547024:Rim8 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D025801:Ubiquitin; C036978:concanamycin A; D043743:Ubiquitin-Protein Ligase Complexes; C092477:PMA1 protein, S cerevisiae; D025262:Vacuolar Proton-Translocating ATPases; D006180:Proton-Translocating ATPases; C469287:RSP5 protein, S cerevisiae
keywords: Arrestin; Endocytosis; Plasma Membrane Proton Pumps; Ubiquitination; Vacuolar ATPase; Vacuolar Acidification; Yeast; pH Regulation