Physical Interaction between Aldolase and Vacuolar H+-ATPase Is Essential for the Assembly and Activity of the Proton Pump

Abstract

Vacuolar proton-translocating ATPases (V-ATPases) are a family of highly conserved proton pumps that couple hydrolysis of cytosolic ATP to proton transport out of the cytosol. Although V-ATPases are involved in a number of cellular processes, how the proton pumps are regulated under physiological conditions is not well understood. We have reported that the glycolytic enzyme aldolase mediates V-ATPase assembly and activity by physical association with the proton pump (Lu, M., Holliday, L. S., Zhang, L., Dunn, W. A., and Gluck, S. L. (2001) J. Biol. Chem. 276, 30407–30413 and Lu, M., Sautin, Y., Holliday, L. S., and Gluck, S. L. (2004) J. Biol. Chem. 279, 8732–8739). In this study, we generate aldolase mutants that lack binding to the B subunit of V-ATPase but retain normal catalytic activities. Functional analysis of the aldolase mutants shows that disruption of binding between aldolase and the B subunit of V-ATPase results in disassembly and malfunction of V-ATPase. In contrast, aldolase enzymatic activity is not required for V-ATPase assembly. Taken together, these findings strongly suggest an important role for physical association between aldolase and V-ATPase in the regulation of the proton pump. Vacuolar proton-translocating ATPases (V-ATPases) are a family of highly conserved proton pumps that couple hydrolysis of cytosolic ATP to proton transport out of the cytosol. Although V-ATPases are involved in a number of cellular processes, how the proton pumps are regulated under physiological conditions is not well understood. We have reported that the glycolytic enzyme aldolase mediates V-ATPase assembly and activity by physical association with the proton pump (Lu, M., Holliday, L. S., Zhang, L., Dunn, W. A., and Gluck, S. L. (2001) J. Biol. Chem. 276, 30407–30413 and Lu, M., Sautin, Y., Holliday, L. S., and Gluck, S. L. (2004) J. Biol. Chem. 279, 8732–8739). In this study, we generate aldolase mutants that lack binding to the B subunit of V-ATPase but retain normal catalytic activities. Functional analysis of the aldolase mutants shows that disruption of binding between aldolase and the B subunit of V-ATPase results in disassembly and malfunction of V-ATPase. In contrast, aldolase enzymatic activity is not required for V-ATPase assembly. Taken together, these findings strongly suggest an important role for physical association between aldolase and V-ATPase in the regulation of the proton pump. Vacuolar proton-translocating ATPases (V-ATPases) 2The abbreviations used are: V-ATPasevacuolar proton-translocating ATPaseaaamino acid(s)GSTglutathione S-transferaseRAVEregulator of the (H+)-ATPase of the vacuolar and endosomal membranes are a family of highly conserved proton pumps that couple hydrolysis of cytosolic ATP to proton transport out of the cytosol. They reside on intracellular membranes of all eukaryotic cells and function to acidify a variety of intracellular compartments, including secretory vesicles, endosomes, lysosomes, the trans-Golgi network, and the central vacuole of yeast (1Stevens T.H. Forgac M. Annu. Rev. Cell Dev. Biol. 1997; 13: 779-808Crossref PubMed Scopus (527) Google Scholar, 2Forgac M. J. Biol. Chem. 1999; 274: 12951-12954Abstract Full Text Full Text PDF PubMed Scopus (265) Google Scholar, 3Nishi T. Forgac M. Nat. Rev. Mol. Cell Biol. 2002; 3: 94-103Crossref PubMed Scopus (1022) Google Scholar, 4Kane P.M. Microbiol. Mol. Biol. Rev. 2006; 70: 177-191Crossref PubMed Scopus (337) Google Scholar). Acidification of the vacuolar compartments plays an important role in a number of cellular processes, including receptor-mediated endocytosis, intracellular targeting of newly synthesized lysosomal enzymes, macromolecular processing and degradation, and coupled transport of small molecules. In mammals, V-ATPases also reside at high levels on the plasma membrane of some specialized cells such as kidney epithelial cells and osteoclasts, where they are responsible for transepithelial or cellular proton transport required for normal acid-base homeostasis of the body or bone remodeling (5Gluck S. Underhill D. Iyori M. Holliday L. Kostrominova T. Lee B.S. Annu. Rev. Physiol. 1996; 58: 427-445Crossref PubMed Scopus (124) Google Scholar). Mutations in V-ATPase subunits have been shown to cause renal tubular acidosis (6Karet F.E. Finberg K.E. Nelson R.D. Nayir A. Mocan H. Sanjad S.A. Rodriguez-Soriano J. Santos F. Cremers C.W. Di Pietro A. Hoffbrand B.I. Winiarski J. Bakkaloglu A. Ozen S. Dusunsel R. Goodyer P. Hulton S.A. Wu D.K. Skvorak A.B. Morton C.C. Cunningham M.J. Jha V. Lifton R.P. Nat. Genet. 1999; 21: 84-90Crossref PubMed Scopus (598) Google Scholar, 7Smith A.N. Skaug J. Choate K.A. Nayir A. Bakkaloglu A. Ozen S. Hulton S.A. Sanjad S.A. Al-Sabban E.A. Lifton R.P. Scherer S.W. Karet F.E. Nat. Genet. 2000; 26: 71-75Crossref PubMed Scopus (5) Google Scholar) and osteopetrosis (8Frattini A. Orchard P.J. Sobacchi C. Giliani S. Abinun M. Mattsson J.P. Andersson A.K. Wallbrandt P. Zecca L. Notarangelo L.D. Vezzoni P. Villa A. Nat. Genet. 2000; 25: 343-346Crossref PubMed Scopus (582) Google Scholar). vacuolar proton-translocating ATPase amino acid(s) glutathione S-transferase regulator of the (H+)-ATPase of the vacuolar and endosomal membranes Although V-ATPases play important physiological roles in a variety of cellular processes, how the V-ATPase-directed proton transport is coupled to cellular metabolism remains poorly understood. Disassembly of V-ATPase in Saccharomyces cerevisiae was first reported to occur in the absence of extracellular glucose (9Kane P.M. J. Biol. Chem. 1995; 270: 17025-17032Abstract Full Text Full Text PDF PubMed Google Scholar). Glucose-dependent assembly and regulation of V-ATPase was also observed in mammalian cells (10Sautin Y. Lu M. Gaugler A. Zhang L. Gluck S.L. Mol. Cell. Biol. 2005; 25: 575-589Crossref PubMed Scopus (192) Google Scholar). However, V-ATPase was found to assemble normally in a panel of yeast mutants with deficiencies in the known glucose-sensing pathways, including the Ras-cAMP, Snf1p, protein kinase C, and Rts1p pathways (11Parra K.J. Kane P.M. Mol. Cell. Biol. 1998; 18: 7064-7074Crossref PubMed Google Scholar). The glucose-induced assembly of V-ATPase is, therefore, independent of these pathways. In a recent report, the glycolytic enzyme phosphofructokinase has been shown to interact with the “a” subunit of human V-ATPase (12Su Y. Zhou A. Al-Lamki R.S. Karet F.E. J. Biol. Chem. 2003; 278: 20013-20018Abstract Full Text Full Text PDF PubMed Scopus (103) Google Scholar). It remains to be determined whether the interaction between phosphofructose kinase and V-ATPase is physiologically relevant to V-ATPase regulation in either mammalian or yeast cells. In a search for protein partners that interact with V-ATPase, we discovered that the glycolytic enzyme aldolase physically associated with three distinct subunits of V-ATPase (13Lu M. Holliday L.S. Zhang L. Dunn W.A. Gluck S.L. J. Biol. Chem. 2001; 276: 30407-30413Abstract Full Text Full Text PDF PubMed Scopus (169) Google Scholar, 14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). We examined the growth pattern of yeast mutant cells deficient in aldolase and observed a growth phenotype similar to that previously reported in the V-ATPase subunit deletion mutant cells (14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). Our data suggest that aldolase deficiency in yeast cells results in V-ATPase malfunction by disrupting the physical association between aldolase and V-ATPase. Furthermore, when the wild-type aldolase gene was introduced and expressed in aldolase deletion mutant cells, abnormalities in V-ATPase assembly and protein expression were restored to normal levels (14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). As a result, ATP hydrolysis and proton transport activities of V-ATPase were also restored to normal levels by aldolase complementation (14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). Taken together, these findings support the notion that the ATP-generating glycolytic pathway is directly coupled to the ATP-hydrolyzing proton pump by physical interaction between aldolase and V-ATPase. An important issue that remains to be addressed is whether the abnormalities in V-ATPase observed in aldolase-deficient cells are caused by disruption of the aldolase-V-ATPase interaction or by attenuation of cellular metabolism mediated by aldolase as a glycolytic enzyme. In this study, we generate aldolase mutants that lack binding to the B subunit of V-ATPase but retain normal enzymatic activities. Functional analysis of the aldolase mutants shows that disruption of binding between aldolase and the B subunit of V-ATPase results in disassembly and malfunction of V-ATPase. Our data indicate that physical association between aldolase and V-ATPase is essential for V-ATPase assembly and activity. Materials—The Taq DNA polymerase was purchased from Promega (Madison, The enzyme was from and DNA were from The expression was from The yeast expression was from and the and B subunits of V-ATPase, were purchased from was purchased from and were from for deletion mutants from the of human aldolase B were by high for the human aldolase B was the and the at the of human aldolase B was the and the at the of human aldolase B was the and the at the of human aldolase B was the and the The aldolase were with the on the yeast expression The DNA were by by DNA and with the V-ATPase B subunit as (13Lu M. Holliday L.S. Zhang L. Dunn W.A. Gluck S.L. J. Biol. Chem. 2001; 276: 30407-30413Abstract Full Text Full Text PDF PubMed Scopus (169) Google Scholar). of yeast cells and of were as previously (13Lu M. Holliday L.S. Zhang L. Dunn W.A. Gluck S.L. J. Biol. Chem. 2001; 276: 30407-30413Abstract Full Text Full Text PDF PubMed Scopus (169) Google Scholar). was out to amino in the binding The on of by Taq polymerase as well as of and number of to the of 1997; PubMed Scopus Google Scholar). of the mutants were found to to the at the of human aldolase B was the and the to the at the of yeast aldolase was the and the The mutant were and with the of the aldolase previously by was to generate human aldolase B the and the yeast aldolase mutants were by the and the was out to for that on but not on amino in the binding were by DNA human aldolase B P. J. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google was by the were by high The were and used as for the the and the the yeast aldolase A. K.E. PubMed Scopus Google was by and the was by a the and the The human and yeast aldolase mutants were the yeast expression and yeast cells for aldolase mutants from were expressed in wild-type yeast cells or in yeast cells deficient in The levels of expression of these mutant were determined by as previously (13Lu M. Holliday L.S. Zhang L. Dunn W.A. Gluck S.L. J. Biol. Chem. 2001; 276: 30407-30413Abstract Full Text Full Text PDF PubMed Scopus (169) Google Scholar). aldolase mutant were expressed in and examined for enzymatic activities. The coupled enzyme was used to aldolase enzymatic activity as previously (13Lu M. Holliday L.S. Zhang L. Dunn W.A. Gluck S.L. J. Biol. Chem. 2001; 276: 30407-30413Abstract Full Text Full Text PDF PubMed Scopus (169) Google Scholar). aldolase mutant were in and enzymatic activity was determined by at a whether the aldolase mutants V-ATPase we out analysis as previously (13Lu M. Holliday L.S. Zhang L. Dunn W.A. Gluck S.L. J. Biol. Chem. 2001; 276: 30407-30413Abstract Full Text Full Text PDF PubMed Scopus (169) Google Scholar). yeast cells were in in the of glucose and were by with with and with subunit by of protein The were by with the subunit of the was a (13Lu M. Holliday L.S. Zhang L. Dunn W.A. Gluck S.L. J. Biol. Chem. 2001; 276: 30407-30413Abstract Full Text Full Text PDF PubMed Scopus (169) Google Scholar). of Vacuolar vacuolar membrane were as previously M. S. Zhang L. Holliday L.S. J. Gluck S.L. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). yeast cells were to and with The were in a and in a The from and were Vacuolar membrane were by the ATP whether the aldolase mutants V-ATPase ATP was ATP hydrolysis of V-ATPase was by the of as previously M. S. Zhang L. Holliday L.S. J. Gluck S.L. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). The of whether the aldolase mutants proton we determined the of proton transport in yeast cells the mutant as we previously M. S. Zhang L. Holliday L.S. J. Gluck S.L. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). Vacuolar membrane were with and transport was by the of a with at and at as previously M. S. Zhang L. Holliday L.S. J. Gluck S.L. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). of to the B of the yeast and we the interaction between the glycolytic enzyme aldolase and the subunit of human V-ATPase (13Lu M. Holliday L.S. Zhang L. Dunn W.A. Gluck S.L. J. Biol. Chem. 2001; 276: 30407-30413Abstract Full Text Full Text PDF PubMed Scopus (169) Google Scholar). was to the a and B subunits of human V-ATPase that also interact with aldolase (14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). the three highly aldolase in mammals, the human aldolase B in renal tubular acidosis found in with P. J. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar, J. PubMed Scopus Google Scholar). the in aldolase B required for with the and subunits of human V-ATPase, we deletion mutants from the and of the human aldolase B the were with the we observed distinct on aldolase B for with the and subunits of V-ATPase (14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). binding of on human aldolase B was also to be essential for binding the B subunit of human V-ATPase not As a first to the regulation of V-ATPase, we out in a of human aldolase B that was previously to the binding required for interaction with the B subunit of V-ATPase. The were with the of human aldolase B by high and in the yeast was out to for that on of but not on of in human aldolase B was for lack of binding to the B subunit of human V-ATPase The on a of aldolase B that is distinct from the catalytic the the and involved in enzymatic P. J. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar, J. L. J. Biol. Chem. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar, J. Biol. Chem. 2003; 278: Full Text Full Text PDF PubMed Scopus Google Scholar). analysis also that the mutant binding to the a and subunits of human V-ATPase was also out to the lack of binding of the aldolase mutant to the B subunit of V-ATPase. The wild-type aldolase B was to the B subunit of V-ATPase In contrast, binding was observed between the aldolase mutant and the B subunit of V-ATPase association of human aldolase B mutants to the and subunits of B in a whether the mutant aldolase enzymatic we a on the mutant and expressed in The mutant protein was for expression by and enzymatic activity was the coupled enzyme as (14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). in aldolase enzymatic activity was between the aldolase B mutant and the wild-type aldolase B protein that the aldolase enzymatic activity was not by the 2The aldolase mutants retain normal enzymatic wild-type aldolase mutant wild-type aldolase aldolase B mutant in a a similar we out in the of the yeast aldolase yeast aldolase was for lack of binding to the B subunit of yeast V-ATPase. The mutant protein was expressed in and for enzymatic activity. aldolase enzymatic activity was observed of the on aldolase enzymatic activity. The of and on V-ATPase and that V-ATPase remains is in and proton transport activity in yeast cells deficient in aldolase (13Lu M. Holliday L.S. Zhang L. Dunn W.A. Gluck S.L. J. Biol. Chem. 2001; 276: 30407-30413Abstract Full Text Full Text PDF PubMed Scopus (169) Google Scholar). The abnormalities in V-ATPase be restored by aldolase complementation (14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). the mammalian and yeast are Genet. PubMed Scopus Google Scholar) and V-ATPases are conserved between human and yeast (1Stevens T.H. Forgac M. Annu. Rev. Cell Dev. Biol. 1997; 13: 779-808Crossref PubMed Scopus (527) Google we examined the of human and yeast aldolase mutants on V-ATPase assembly and activity in yeast cells deficient in yeast cells with the aldolase mutants were out on for of the human or yeast aldolase The yeast were and for expression of the mutant by the human or yeast The levels of mutant protein expression were found to be to that of the wild-type aldolase complementation of aldolase-deficient yeast with wild-type human aldolase B restored V-ATPase assembly ATPase activity and proton transport activity In contrast, of the aldolase-deficient yeast with the human aldolase B mutant to the in V-ATPase assembly observed in yeast cells deficient in aldolase with the on V-ATPase with the aldolase B mutant to ATP activity and proton transport activity we examined the of of aldolase-deficient yeast either with the wild-type yeast or with the on complementation of V-ATPase assembly and activity. As shown in expression of the wild-type yeast but not the B subunit restored V-ATPase assembly ATP activity and proton transport activity that the binding between aldolase and the B subunit of V-ATPase is essential for the assembly and activity of the proton we expressed the human and yeast aldolase mutants in wild-type yeast cells. V-ATPase ATPase and proton transport were of the aldolase B and activity a in V-ATPase assembly and in ATP activity and in proton transport activity in V-ATPase assembly was also observed in wild-type yeast cells the yeast aldolase B and As ATP and proton transport activities were also and Our therefore, indicate that physical association between aldolase and V-ATPase is essential for the assembly and activity of the proton pump. how the aldolase mutants in a on V-ATPase assembly and we expressed the aldolase mutants in wild-type yeast cells. The expression levels of the aldolase mutant were determined to be to the aldolase expression not analysis was out the the B subunit of V-ATPase. The were by the yeast The binding between aldolase and V-ATPase in cells the human and yeast aldolase mutants was by and data indicate that of the aldolase binding of the yeast aldolase to V-ATPase. and assembly and of the yeast V-ATPase. In aldolase by the V-ATPase with a glycolytic of the aldolase mutants of the the on the assembly and activity of the proton pump. this we examined the of glucose and on wild-type yeast cells the wild-type aldolase analysis normal assembly of V-ATPase in the absence of glucose that of aldolase the normal to glucose for V-ATPase that glycolytic activity is not required for V-ATPase assembly aldolase is whether the enzymatic activity of aldolase is required for V-ATPase we the human aldolase B by and in the yeast expression The aldolase B found in a with was previously shown to have aldolase enzymatic activity P. J. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). We examined binding of the mutant to the and subunits of V-ATPase by the yeast The in the catalytic of aldolase B on the binding of aldolase B to of the three V-ATPase subunits the mutant was expressed in wild-type yeast cells, was found to be to that of the analysis normal assembly of V-ATPase in yeast the aldolase B mutant that the catalytic activity of aldolase is not required for V-ATPase assembly. the ATP and proton transport activities to be normal in cells the mutant and we the mutant of the yeast A. K.E. PubMed Scopus Google by and in the yeast expression The mutant was expressed in wild-type yeast cells, and expression was found to be as as that of the of V-ATPase in cells the mutant was to that in wild-type yeast cells the ATP and proton transport activities were also in cells the mutant and with human and yeast aldolase that B subunit but not the enzymatic activity of is required for V-ATPase assembly and the that yeast aldolase was responsible for V-ATPase assembly in cells we expressed either wild-type human aldolase B or the mutant in aldolase-deficient yeast cells and examined V-ATPase assembly by of the mutant in normal levels of V-ATPase assembly expression of the yeast aldolase V-ATPase assembly to that in cells wild-type yeast aldolase data that the B subunit binding of but not enzymatic is essential for normal V-ATPase assembly and strongly suggest that the for V-ATPase disassembly with glucose is of a of V-ATPase with from that the ATP-generating glycolytic pathway is directly coupled to the ATP-hydrolyzing proton pump by (13Lu M. Holliday L.S. Zhang L. Dunn W.A. Gluck S.L. J. Biol. Chem. 2001; 276: 30407-30413Abstract Full Text Full Text PDF PubMed Scopus (169) Google Scholar, 14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). In this study, we examined whether physical association between aldolase and V-ATPase was essential for the assembly and activity of the proton pump. We human and yeast aldolase mutants that to the B subunit of V-ATPase by It was that a a interaction between aldolase and V-ATPase with on the protein that the mutant is to retain aldolase enzymatic activity and binding for V-ATPase As the aldolase mutants were observed to retain normal enzymatic activities and binding to the a and subunits of V-ATPase the aldolase mutants were introduced and expressed in aldolase-deficient cells, abnormalities in V-ATPase assembly and activity a in the V-ATPase Furthermore, we expressed the human and yeast aldolase mutants in wild-type yeast cells and observed in V-ATPase assembly and activity In contrast, the aldolase enzymatic activity to be not essential for V-ATPase assembly and data strongly suggest an important role for physical association between aldolase and V-ATPase in the regulation of assembly and activity of the proton pump. Although aldolase enzymatic activity is not essential for V-ATPase assembly and analysis is required to whether V-ATPase function on It is that disruption of of the glycolytic pathway ATP to the ATP-hydrolyzing proton to malfunction of V-ATPase. the mutants of human aldolase B found in with three and have been P. J. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). They all the protein of aldolase but enzymatic activity P. J. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar). findings that aldolase enzymatic activity is required for V-ATPase Our analysis of of the three mutants that V-ATPase assembly and activity to be normal and The data well be by the of the in out with yeast cells. ATP was in the ATPase the of the mutant on V-ATPase activity be Taken together, data suggest that the aldolase enzymatic activity is not required for the assembly but be required for the activity of the proton pump. that ATP is used in ATP is required in for from and cellular membrane PubMed Scopus Google Scholar). of ATP by ATP synthesized by Annu. Rev. Physiol. PubMed Scopus Google Scholar). glycolytic enzyme have been observed in and that be involved in ATP R.S. J. Biol. Chem. Full Text PDF PubMed Google Scholar, L. A. A. P. S. A. 1997; PubMed Scopus Google Scholar). has been to be essential for and for normal A. D. T. J. Biol. Chem. 2003; 278: Full Text Full Text PDF PubMed Scopus Google Scholar). A. D. T. J. Biol. Chem. 2003; 278: Full Text Full Text PDF PubMed Scopus Google Scholar) that ATP is by the V-ATPase proton pump to generate an proton the membrane of vesicles, the for transport We have shown that the glycolytic enzyme aldolase mediates assembly and activity of V-ATPase (13Lu M. Holliday L.S. Zhang L. Dunn W.A. Gluck S.L. J. Biol. Chem. 2001; 276: 30407-30413Abstract Full Text Full Text PDF PubMed Scopus (169) Google Scholar, 14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). The of regulation of V-ATPase is by that ATP and proton transport are that as as to J.P. T. H. J. L. J. Biol. PubMed Google Scholar). these support the notion that cellular on ATP and that not be or to the ATP for these Glucose-dependent assembly of V-ATPase was reported to occur in yeast cells (9Kane P.M. J. Biol. Chem. 1995; 270: 17025-17032Abstract Full Text Full Text PDF PubMed Google Scholar). analysis in mammalian cells that assembly and activity of V-ATPase was also (10Sautin Y. Lu M. Gaugler A. Zhang L. Gluck S.L. Mol. Cell. Biol. 2005; 25: 575-589Crossref PubMed Scopus (192) Google Scholar). In this study, by yeast cells as a we observed that the human and yeast aldolase mutants in a similar in V-ATPase assembly and activity. findings suggest a highly conserved for regulation of V-ATPase by of to the proton pump. We previously reported that aldolase physically with the and subunits of V-ATPase (14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). analysis that in interaction between aldolase and V-ATPase in yeast cells in the of glucose (14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). findings suggest that aldolase as a glucose and V-ATPase assembly. In this study, we the of aldolase binding to the B subunit of V-ATPase in V-ATPase assembly and activity. the binding on aldolase for the a and subunits of V-ATPase in the aldolase assembly of V-ATPase was in wild-type yeast cells. The aldolase mutants that to the B subunit of V-ATPase were also found to be to the in V-ATPase assembly and activity in yeast cells in Our therefore, support the notion that aldolase mediates V-ATPase assembly by binding to subunits of V-ATPase. of a interaction between aldolase and V-ATPase the the of and In findings that physical association between aldolase and the B subunit of V-ATPase is important for the proton pump has been shown to the and subunits of V-ATPase (14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). It remains to be determined whether the subunit and subunit are also required for V-ATPase assembly and activity. Furthermore, aldolase in with for V-ATPase assembly and activity. aldolase with V-ATPase on the vacuolar membrane but not with the or (14Lu M. Sautin Y. Holliday L.S. Gluck S.L. J. Biol. Chem. 2004; 279: 8732-8739Abstract Full Text Full Text PDF PubMed Scopus (163) Google Scholar). In contrast, the to the is in the but not to or V-ATPase M. Kane P. J. Biol. Chem. 2002; Full Text Full Text PDF PubMed Scopus Google Scholar). and to V-ATPase in a It is, therefore, that the is to interact with from the vacuolar membrane glucose and when extracellular glucose is the to for V-ATPase