Vacuolar cation/H+ antiporters of Saccharomyces cerevisiae.
We previously demonstrated that Saccharomyces cerevisiae vnx1Δ mutant strains displayed an almost total loss of Na(+) and K(+)/H(+) antiporter activity in a vacuole-enriched fraction. However, using different in vitro transport conditions, we were able to reveal additional K(+)/H(+) antiporter activity. By disrupting genes encoding transporters potentially involved in the vnx1 mutant strain, we determined that Vcx1p is responsible for this activity. This result was further confirmed by complementation of the vnx1Δvcx1Δ nhx1Δ triple mutant with Vcx1p and its inactivated mutant Vcx1p-H303A. Like the Ca(2+)/H(+) antiporter activity catalyzed by Vcx1p, the K(+)/H(+) antiporter activity was strongly inhibited by Cd(2+) and to a lesser extend by Zn(2+). Unlike as previously observed for NHX1 or VNX1, VCX1 overexpression only marginally improved the growth of yeast strain AXT3 in the presence of high concentrations of K(+) and had no effect on hygromycin sensitivity. Subcellular localization showed that Vcx1p and Vnx1p are targeted to the vacuolar membrane, whereas Nhx1p is targeted to prevacuoles. The relative importance of Nhx1p, Vnx1p, and Vcx1p in the vacuolar accumulation of monovalent cations will be discussed.
Cagnac
Olivier
O
Departamento de Bioquímica, Biología Celular y Molecular de Plantas, Estacion Experimental del Zaidín, Consejo Superior de Investigaciones Científicas, Apartado 419, E-18080 Granada, Spain. olivier.cagnac@eez.csic.es
Aranda-Sicilia
Maria Nieves
MN
Leterrier
Marina
M
Rodriguez-Rosales
Maria-Pilar
MP
Venema
Kees
K
D017920:Antiporters / Q000737:chemistry; D002104:Cadmium / Q000737:chemistry; D002412:Cations / Q000737:chemistry*; D002934:Cinnamates / Q000737:chemistry; D006921:Hygromycin B / Q000031:analogs & derivatives / Q000737:chemistry; D008856:Microscopy, Fluorescence / Q000379:methods; D009154:Mutation; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D011188:Potassium / Q000737:chemistry; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry; D017923:Sodium-Hydrogen Exchangers / Q000737:chemistry; D013347:Subcellular Fractions / Q000737:chemistry; D014617:Vacuoles / Q000378:metabolism*; D015032:Zinc / Q000737:chemistry
D017920:Antiporters; D002412:Cations; D002934:Cinnamates; C492183:NHX1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D017923:Sodium-Hydrogen Exchangers; C099946:VCX1 protein, S cerevisiae; C523520:Vnx1 protein, S cerevisiae; D002104:Cadmium; D006921:Hygromycin B; C026273:hygromycin A; D015032:Zinc; D011188:Potassium