Effects of ion gradients on H+ transport mediated by human MDR 1 protein.
In the previous paper we presented a variety of data consistent with significant perturbations in 9.3 yeast plasma membrane ion transport upon overexpression of the hu MDR 1 protein. Thus, in this paper, we compare formation of DeltapH for inside-out yeast plasma membrane vesicles (ISOV) prepared from control 9.3/pVT versus 9.3/hu MDR 1 yeast. Since MDR 1 ATPase activity has a broader, more alkaline pH profile relative to endogenous yeast H+ ATPase activity, we analyzed H+ pumping at pH >/= 8.0 in detail in order to selectively amplify hu MDR 1 contributions to H+ movement over those of the endogenous yeast H+ ATPase. We observed: (1) imposition of a Cl- gradient oriented outside to in enhances acidification for 9.3/pVT ISOV (as expected), but decreases acidification for 9.3/hu MDR 1 ISOV; (2) imposition of a Cl- gradient oriented inside to out decreases acidification for 9.3/pVT ISOV (as expected) but enhances acidification for 9.3/hu MDR 1 ISOV; (3) a Na+ gradient oriented in the same direction as the Cl- gradient amplifies the effects due to hu MDR 1 when both gradients are oriented inside to out, but not outside to in. The data are most easily explained by interesting Na+, Cl-, and ATP-dependent H+ transport mediated by hu MDR 1 protein as previously suggested [Hoffman and Roepe (1997) Biochemistry 36, 11153-11168]. These data may help to resolve a variety of conflicting reports in the literature regarding ion transport mediated by hu MDR 1 and have implications for the physiology of a number of polarized epithelia in which hu MDR 1 is endogenously expressed.
Santai
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Department of Chemistry, Lombardi Cancer Center Program in Tumor Biology, Georgetown University, Washington, D.C. 20057, USA.
Fritz
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Roepe
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D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics / Q000502:physiology*; D000165:Acridine Orange / Q000378:metabolism; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism / Q000502:physiology; D000818:Animals; D017920:Antiporters / Q000378:metabolism; D016466:CHO Cells; D002458:Cell Fractionation; D002462:Cell Membrane / Q000378:metabolism / Q000502:physiology; D006224:Cricetinae; D006801:Humans; D006863:Hydrogen-Ion Concentration; D017136:Ion Transport / Q000187:drug effects / Q000235:genetics; D011189:Potassium Chloride / Q000378:metabolism; D011522:Protons; D014162:Transfection; D014638:Vanadates / Q000378:metabolism
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D017920:Antiporters; D011522:Protons; C114260:hydrogen-chloride symporter; D014638:Vanadates; D011189:Potassium Chloride; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D000165:Acridine Orange