Differential localization of the vacuolar H+ pump with G subunit isoforms (G1 and G2) in mouse neurons.
Vacuolar H(+)-ATPases (V-ATPases), a family of multimeric proton pumps, are involved in a wide variety of physiological processes. We have identified two mouse genes, Atp6g1 and Atp6g2, encoding the G1 and G2 isoforms of the V-ATPase G subunit, respectively. G1 was distributed ubiquitously in the tissues examined, whereas G2 was specifically distributed in central nervous system neurons. G1 was expressed at an early embryonic stage, whereas G2 transcription was significantly induced at 10.5 days postcoitus (embryonic day 10.5, i.e. 2 days before axon outgrowth). Both G1 and G2 were strongly expressed in cortical and hippocampal neurons, cerebellar granule cells, and Purkinje cells. Immunohistochemistry with isoform-specific antibodies revealed that G2 was localized in cell bodies, dendrites, and axons. In addition, electron microscopy and subcellular fractionation indicated that G2 was localized in synaptic vesicles, whereas G1 was not detectable. G1 and G2 exhibit 62% identity, and both isoforms were immunoprecipitated with the c and A subunits of V-ATPase. G2 could complement the yeast deletion mutant Deltavma10, which lacks the G subunit. The V-ATPases containing the G1 and G2 isoforms, respectively, showed similar K(m)((ATP)) values and maximal velocity. These results indicate that G1 and G2 are bona fide subunits of V-ATPases and that the enzyme with the G2 isoform is involved in synaptic vesicle acidification.
Division of Biological Sciences, Institute of Scientific and Industrial Research, Osaka University, Osaka 567-0047, Japan.
Yoshiko
Y
Murata
Ge-Hong
GH
Sun-Wada
Takao
T
Yoshimizu
Akitsugu
A
Yamamoto
Yoh
Y
Wada
Masamitsu
M
Futai
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D002462:Cell Membrane / Q000378:metabolism; D002531:Cerebellum / Q000166:cytology; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D017353:Gene Deletion; D005816:Genetic Complementation Test; D006624:Hippocampus / Q000166:cytology; D007150:Immunohistochemistry; D007700:Kinetics; D008099:Liver / Q000378:metabolism; D008247:Lysosomes / Q000378:metabolism; D051379:Mice; D008813:Mice, Inbred ICR; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D011233:Precipitin Tests; D020033:Protein Isoforms; D011522:Protons; D012333:RNA, Messenger / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013347:Subcellular Fractions; D016708:Synaptophysin / Q000378:metabolism; D013997:Time Factors; D014018:Tissue Distribution; D014158:Transcription, Genetic; D025262:Vacuolar Proton-Translocating ATPases / Q000096:biosynthesis* / Q000737:chemistry* / Q000378:metabolism
D018076:DNA, Complementary; D020033:Protein Isoforms; D011522:Protons; D012333:RNA, Messenger; D016708:Synaptophysin; D000251:Adenosine Triphosphatases; C467738:V-ATPase G subunit, mouse; D025262:Vacuolar Proton-Translocating ATPases