Functional expression of the colonic H+,K+-ATPase alpha-subunit. Pharmacologic properties and assembly with X+,K+-ATPase beta-subunits.
The functional and pharmacological properties of the alpha-subunit of the colonic H+,K+-ATPase (alphaC) were studied in Xenopus laevis oocytes. alphaC was injected with different rat beta-subunits, the beta-subunit of the gastric H+,K+-ATPase (betaG, the only H+, K+-ATPase beta-subunit identified in rat), or the beta1-subunit of the Na+,K+-ATPase (beta1) (associated with the basolateral Na+, K+-ATPase, but also expressed in the epithelial apical membranes of rat distal colon) (Marxer, A., Stieger, B., Quarini, A., Kashgarian, M., and Hauri, H. P. (1989) J. Cell Biol. 109, 1057-1069). The effect of the different beta-subunits was studied by measuring 86Rb+ uptake (a K+ congener) in the presence or absence of Sch-28080 and ouabain. Significant Na+-independent 86Rb+ uptake was observed only when alphaC was coexpressed with one of the beta-subunits. The expressed alphaCbeta1 and alphaCbetaG complexes were not inhibited by Sch-28080, were only partially sensitive to ouabain (IC50 = 400-600 microM, in the presence of external 1 mM KCl), and exhibited comparable K+ activation kinetics. Coexpression of alphaC with epitope-tagged betaG or beta1, followed by immunopurification of the alphabeta complexes, confirmed stable assembly of alphaCbetaG and alphaCbeta1 complexes. Since the beta1-subunit, but not the alpha1-subunit, of Na+,K+-ATPase is expressed in the apical membrane of rat colonocytes, our data support the view that, in rat distal colon, the beta1-subunit may play a surrogate role as the beta-subunit for the colonic H+,K+-ATPase.
Codina
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Department of Internal Medicine, University of Texas Medical School at Houston, Houston, Texas 77030, USA. tdubose@heart.med.uth.tmc.edu
Kone
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Delmas-Mata
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DuBose
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D000818:Animals; D003001:Cloning, Molecular; D003106:Colon / Q000201:enzymology*; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D017506:H(+)-K(+)-Exchanging ATPase / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D009865:Oocytes; D051381:Rats; D014982:Xenopus laevis
D004791:Enzyme Inhibitors; D017506:H(+)-K(+)-Exchanging ATPase