title: Vacuolar-type H(+)-ATPase with the a3 isoform is the proton pump on premature melanosomes.
abstract: The melanosome, an organelle specialized for melanin synthesis, is one of the lysosome-related organelles. Its lumen is reported to be acidified by vacuolar-type H(+)-ATPase (V-ATPase). Mammalian V-ATPase exhibits structural diversity in its subunit isoforms; with regard to membrane intrinsic subunit a, four isoforms (a1-a4) have been found to be localized to distinct subcellular compartments. In this study, we have shown that the a3 isoform is co-localized with a melanosome marker protein, Pmel17, in mouse melanocytes. Acidotropic probes (LysoSensor and DAMP) accumulate in non-pigmented Pmel17-positive melanosomes, and DAMP accumulation is sensitive to bafilomycin A1, a specific inhibitor of V-ATPase. However, none of the subunit a isoforms is associated with highly pigmented mature melanosomes, in which the acidotropic probes are also not accumulated. oc/oc mice, which have a null mutation at the a3 locus, show no obvious defects in melanogenesis. In the mutant melanocytes, the expression of the a2 isoform is modestly elevated, and a considerable fraction of this isoform is localized to premature melanosomes. These observations suggest that the V-ATPase keeps the lumen of premature melanosomes acidic, whereas melanosomal acidification is less significant in mature melanosomes.
authors.affiliation: Department of Biochemistry, Faculty of Pharmaceutical Sciences, Doshisha Women's College, Kyoto, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Tabata
authors.forename: Nobuyuki
authors.initials: N
authors.lastname: Kawamura
authors.forename: Ge-Hong
authors.initials: GH
authors.lastname: Sun-Wada
authors.forename: Yoh
authors.initials: Y
authors.lastname: Wada
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D017353:Gene Deletion; D006863:Hydrogen-Ion Concentration; D008544:Melanocytes / Q000648:ultrastructure; D020460:Melanosomes / Q000737:chemistry / Q000201:enzymology* / Q000648:ultrastructure; D008562:Membrane Glycoproteins / Q000032:analysis; D051379:Mice; D008810:Mice, Inbred C57BL; D008813:Mice, Inbred ICR; D010641:Phenotype; D010858:Pigmentation; D020033:Protein Isoforms / Q000032:analysis; D025262:Vacuolar Proton-Translocating ATPases / Q000032:analysis* / Q000235:genetics / Q000502:physiology; D058951:gp100 Melanoma Antigen
chemical_list: C484226:Atp6ap1 protein, mouse; D008562:Membrane Glycoproteins; C548642:Pmel protein, mouse; D020033:Protein Isoforms; D058951:gp100 Melanoma Antigen; D025262:Vacuolar Proton-Translocating ATPases