Abstract
The vacuolar H(+)-ATPase of the yeast Saccharomyces cerevisiae is composed of a complex of peripheral subunits (the V1 sector) attached to an integral membrane complex (the V0 sector). In the experiments described here, attachment of the V1 to the V0 sector was assessed in wild-type cells under a variety of growth conditions. Depriving the yeast cells of glucose, even for as little as 5 min, caused dissociation of approximately 70% of the assembled enzyme complexes into separate V1 and V0 subcomplexes. Restoration of glucose induced rapid and efficient reassembly of the enzyme from the previously synthesized subcomplexes. Indirect immunofluorescence microscopy and subcellular fractionation revealed detachment of the peripheral subunits from the vacuolar membrane in the absence of glucose, followed by reattachment in the presence of glucose. Rapid dissociation of vacuolar H(+)-ATPases could also be triggered by shifting cells into a variety of other carbon sources, and reassembly could be generated by addition of glucose. Disassembly and reassembly of vacuolar H(+)-ATPases in vivo may be a means of regulating organelle acidification in response to extracellular conditions, or a mechanism for assembling alternate complexes of vacuolar H(+)-ATPases in different intracellular compartments.
Highlights
From the Department of Biochemistry and Molecular Biology, State University of New York Health Science Center, Syracuse, New York 13210
The vacuolar H+ -ATPase of the yeast Saccharomyces cerevisiae is composed of a complex of peripheral subunits attached to an integral membrane complex
And reassembly of vacuolar H+ ATPases in vivo may be a means of regulating organelle acidification in response to extracellular conditions, or a mechanism for assembling alternate complexes of vacuolar H+-ATPases in different intracellular compartments
Summary
Vol 270, No 28, Issue of July 14, pp. 17025-17032, 1995 Printed in U.S.A. From the Department of Biochemistry and Molecular Biology, State University of New York Health Science Center, Syracuse, New York 13210. The vacuolar H+ -ATPase of the yeast Saccharomyces cerevisiae is composed of a complex of peripheral subunits (the VI sector) attached to an integral membrane complex (the Vo sector). Unlike the F IFo-ATPases, which can be separated into a soluble F I complex that is capable of ATP hydrolysis [6], and a membrane-bound F 0 complex that appears to function as a proton channel [7], a variety of biochemical studies indicate that the V I sector of the vacuolar H+ -ATPases does not retain. This paper describes the reversible dissociation of the wild-type VI complex from the Vo complex in vivo Both the disassembly and subsequent reassembly of the complex occur rapidly and efficiently in response to changes in growth conditions, suggesting that these processes could playa role in regulation of vacuolar. H+-ATPase activity or in cellular distribution of the active enzyme
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