title: Cellular vacuolization caused by overexpression of the PIKfyve-binding deficient Vac14L156Ris rescued by starvation and inhibition of vacuolar-ATPase.
abstract: Phosphoinositides (PI) and converting enzymes are crucial determinants of organelle identity and morphology. One important endolysosomal specific PI is PI(3,5)P2, generated by the PIKfyve kinase, which orchestrates in combination with Vac14 and Fig4. Dysfunction of this complex leads to large intracellular vacuoles in various cell types and is linked to neurological diseases. Here, we characterize the vacuolization phenotype caused by overexpression of the PIKfyve binding deficient mutant Vac14L156Rin podocytes, which represent specialized cells of the kidney. Vacuolization of podocytes, which was associated with strong maturation defects in the endolysosomal system, could be completely rescued by starvation or treatment of cells with the v-ATPase inhibitor Bafilomycin A1. Moreover, we elucidated a strong and reversible de-vacuolization effect of the cholesterol export inhibitor U18666A, which was accompanied by increased basification of the lysosomal pH values. Taken together, our data give new hints to potential therapeutic targets in the treatment of disease linked to intracellular vacuolization.
authors.affiliation: Internal Medicine D, University Hospital of Muenster, Albert-Schweitzer-Campus 1, A14, D-48149 Muenster, Germany. Electronic address: ulfsch@uni-muenster.de.
authors.forename: Ulf
authors.initials: U
authors.lastname: Schulze
authors.affiliation: Internal Medicine D, University Hospital of Muenster, Albert-Schweitzer-Campus 1, A14, D-48149 Muenster, Germany.
authors.forename: Beate
authors.initials: B
authors.lastname: Vollenbröker
authors.affiliation: Institute of Medical Biochemistry, Center for Molecular Biology of Inflammation, and Interdisciplinary Clinical Research Centre, University of Muenster, Von-Esmarch-Str. 56, D-48149 Muenster, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: Kühnl
authors.affiliation: Internal Medicine D, University Hospital of Muenster, Albert-Schweitzer-Campus 1, A14, D-48149 Muenster, Germany.
authors.forename: Daniel
authors.initials: D
authors.lastname: Granado
authors.affiliation: Internal Medicine D, University Hospital of Muenster, Albert-Schweitzer-Campus 1, A14, D-48149 Muenster, Germany.
authors.forename: Samet
authors.initials: S
authors.lastname: Bayraktar
authors.affiliation: Institute of Medical Biochemistry, Center for Molecular Biology of Inflammation, and Interdisciplinary Clinical Research Centre, University of Muenster, Von-Esmarch-Str. 56, D-48149 Muenster, Germany.
authors.forename: Ursula
authors.initials: U
authors.lastname: Rescher
authors.affiliation: Internal Medicine D, University Hospital of Muenster, Albert-Schweitzer-Campus 1, A14, D-48149 Muenster, Germany.
authors.forename: Hermann
authors.initials: H
authors.lastname: Pavenstädt
authors.affiliation: Internal Medicine D, University Hospital of Muenster, Albert-Schweitzer-Campus 1, A14, D-48149 Muenster, Germany. Electronic address: weidet@uni-muenster.de.
authors.forename: Thomas
authors.initials: T
authors.lastname: Weide
mesh_terms: D019943:Amino Acid Substitution / Q000235:genetics; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free / Q000494:pharmacology*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005502:Food; D057809:HEK293 Cells; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D018942:Macrolides / Q000494:pharmacology*; D008565:Membrane Proteins / Q000235:genetics*; D050199:Podocytes / Q000187:drug effects* / Q000378:metabolism / Q000648:ultrastructure; D015854:Up-Regulation / Q000235:genetics; D025262:Vacuolar Proton-Translocating ATPases / Q000037:antagonists & inhibitors; D014617:Vacuoles / Q000187:drug effects* / Q000235:genetics*
chemical_list: D016895:Culture Media, Serum-Free; D004791:Enzyme Inhibitors; D047908:Intracellular Signaling Peptides and Proteins; D018942:Macrolides; D008565:Membrane Proteins; C494631:VAC14 protein, human; C057620:bafilomycin A; D025262:Vacuolar Proton-Translocating ATPases