A NOVEL N-TERMINAL SPLICE VARIANT OF THE RAT H-K+-ATPASE ALPHA-2 SUBUNIT - CLONING, FUNCTIONAL EXPRESSION, AND RENAL ADAPTIVE RESPONSE TO CHRONIC HYPOKALEMIA()

The H+-K+-ATPase of renal collecting duct mediates K+ conservation dur ing chronic hypokalemia, K+ deprivation promotes H+-K+-ATPase alpha 2 (HK alpha 2) gene expression in the medullary collecting duct, the pri ncipal site of active K+ reabsorption, suggesting that this isozyme co ntributes to renal K+ reclamation, We report here that alternative tra nscriptional initiation and mRNA splicing give rise to distinct N-term inal variants of the HK alpha 2 subunit, Sequence analysis and in vitr o translation revealed that HK alpha 2a corresponds to the known HK al pha 2 cDNA (Crowson, M. S., and Shull, G. E. (1992) J. Biol. Chem. 267 , 13740-13748), whereas HK alpha 2b represents a novel variant truncat ed by 108 amino acids at its N terminus, HK alpha 2b mRNA contains a c omplex 5'-untranslated region with eight upstream open reading frames, features implicated in translational regulation of other genes, Heter ologous expression of HK alpha 2b with and without the gastric H+-K+-A TPase beta subunit in HEK 293 cells indicated that this variant encode s a KC uptake mechanism that is relatively Sch 28080-resistant, partia lly sensitive to ouabain, and appears to require coexpression with the gastric H+-K+-ATPase beta subunit for optimal functional activity, No rthern analysis demonstrated that both subtypes (HK alpha 2b > HK alph a 2a) are expressed abundantly in distal colon and modestly in proxima l colon and kidney, Moreover, the abundance of the two mRNAs increases coordinately among the renal zones, but not in colon, with chronic K deprivation. These results demonstrate the potential for complex cont rol of HK alpha 2 gene expression by transcriptional and posttranscrip tional mechanisms not recognized in other members of the Na+-K+-ATPase /H+-K+-ATPase family.