title: Mapping the H(+) (V)-ATPase interactome: identification of proteins involved in trafficking, folding, assembly and phosphorylation.
abstract: V-ATPases (H(+) ATPases) are multisubunit, ATP-dependent proton pumps that regulate pH homeostasis in virtually all eukaryotes. They are involved in key cell biological processes including vesicle trafficking, endosomal pH sensing, membrane fusion and intracellular signaling. They also have critical systemic roles in renal acid excretion and blood pH balance, male fertility, bone remodeling, synaptic transmission, olfaction and hearing. Furthermore, V-ATPase dysfunction either results in or aggravates various other diseases, but little is known about the complex protein interactions that regulate these varied V-ATPase functions. Therefore, we performed a proteomic analysis to identify V-ATPase associated proteins and construct a V-ATPase interactome. Our analysis using kidney tissue revealed V-ATPase-associated protein clusters involved in protein quality control, complex assembly and intracellular trafficking. ARHGEF7, DMXL1, EZR, NCOA7, OXR1, RPS6KA3, SNX27 and 9 subunits of the chaperonin containing TCP1 complex (CCT) were found to interact with V-ATPase for the first time in this study. Knockdown of two interacting proteins, DMXL1 and WDR7, inhibited V-ATPase-mediated intracellular vesicle acidification in a kidney cell line, providing validation for the utility of our interactome as a screen for functionally important novel V-ATPase-regulating proteins. Our data, therefore, provide new insights and directions for the analysis of V-ATPase cell biology and (patho)physiology.
authors.affiliation: MGH Center for Systems Biology, Program in Membrane Biology &Division of Nephrology, Richard B. Simches Research Center, Massachusetts General Hospital and Department of Medicine, Harvard Medical School, Boston, MA 02114, USA.
authors.forename: Maria
authors.initials: M
authors.lastname: Merkulova
authors.affiliation: MGH Center for Systems Biology, Program in Membrane Biology &Division of Nephrology, Richard B. Simches Research Center, Massachusetts General Hospital and Department of Medicine, Harvard Medical School, Boston, MA 02114, USA.
authors.forename: Teodor G
authors.initials: TG
authors.lastname: Păunescu
authors.affiliation: MGH Center for Systems Biology, Program in Membrane Biology &Division of Nephrology, Richard B. Simches Research Center, Massachusetts General Hospital and Department of Medicine, Harvard Medical School, Boston, MA 02114, USA.
authors.forename: Anie
authors.initials: A
authors.lastname: Azroyan
authors.affiliation: MGH Center for Systems Biology, Program in Membrane Biology &Division of Nephrology, Richard B. Simches Research Center, Massachusetts General Hospital and Department of Medicine, Harvard Medical School, Boston, MA 02114, USA.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Marshansky
authors.affiliation: MGH Center for Systems Biology, Program in Membrane Biology &Division of Nephrology, Richard B. Simches Research Center, Massachusetts General Hospital and Department of Medicine, Harvard Medical School, Boston, MA 02114, USA.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Breton
authors.affiliation: MGH Center for Systems Biology, Program in Membrane Biology &Division of Nephrology, Richard B. Simches Research Center, Massachusetts General Hospital and Department of Medicine, Harvard Medical School, Boston, MA 02114, USA.
authors.forename: Dennis
authors.initials: D
authors.lastname: Brown
mesh_terms: D000818:Animals; D047468:Immunoprecipitation; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008822:Mice, Transgenic; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D017510:Protein Folding; D025941:Protein Interaction Mapping / Q000379:methods; D060066:Protein Interaction Maps; D021381:Protein Transport; D025262:Vacuolar Proton-Translocating ATPases / Q000235:genetics / Q000378:metabolism*
chemical_list: C576980:DMXL2 protein, mouse; D009419:Nerve Tissue Proteins; C480628:Atp6v1b1 protein, mouse; D025262:Vacuolar Proton-Translocating ATPases