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 a-subunit of the colonic H+,K+-ATPase (alpha(C)) were studied in Xenopus laevis oocytes. alpha(C) was injected with different rat beta-subunits, the beta-subunit of the gastric H+,K+-ATPase (beta(G), the only H+,K+-ATPase beta-subunit identified in rat), or the beta(1)-subunit of the Na+,K+-ATPase (beta(1)) (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 Rb-86(+) uptake (a K+ congener) in the presence or absence of Sch-28080 and ouabain. Significant Na+-independent Rb-86(+) uptake was observed only when alpha(C) was coexpressed with one of the beta-subunits. The expressed alpha(C) beta(1) and alpha(C) beta(G) complexes were not inhibited by Sch-28080, were only partially sensitive to ouabain (IC50 = 400-600 mu M, in the presence of external 1 mm KCl), and exhibited comparable K+ activation kinetics. Coexpression of alpha(C) with epitope-tagged beta(G) or beta(1), followed by immunopurification of the alpha beta complexes, confirmed stable assembly of alpha(C) beta(G) and alpha(C) beta(1) complexes. Since the beta(1)-subunit, but not the alpha(1)-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 beta(1)-subunit may play a surrogate role as the beta-subunit for the colonic H+,K+-ATPase.