EXPRESSION OF NA-K+-ATPASE ALPHA-SUBUNIT AND BETA-SUBUNIT ALONG RAT NEPHRON - ISOFORM SPECIFICITY AND RESPONSE TO HYPOKALEMIA()

The activity of Na+-K+-adenosinetriphosphatase (Na+-K+-ATPase), the so dium pump, which drives active Na+ reabsorption along the nephron, var ies over an order of magnitude, depending on the nephron segment, and activity is increased in the outer medullary collecting tubule (MCT) d uring hypokalemia. The aims of the present study were to assess abunda nce of sodium pump alpha(1)- and beta(1)-subunits in dissected nephron segments of the rat by immunoblotting, to determine if alpha(2)- or a lpha(3)-protein could be detected in the collecting tubules, as sugges ted by Barlet-Bas et al. (C. Barlet-Bas, E. Arystarkhova, L. Cheval, S . Marsy, K. Sweadner, N. Modyanov, and A. Doucet. J. Biol. Chem. 268: 11512-11515, 1993) for rabbit, and to determine if alpha(1) and beta(1 ) were increased in MCT by hypokalemia. Tubules from the rat were reso lved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (12- 100 mm/lane), blotted, and probed with subunit-specific antisera. alph a(1) and beta(1), detected in all tubule segments assayed, were highes t in cortical and medullary thick ascending limbs and proximal convolu ted tubule (PCT), lower in the MCT and barely detectable in proximal s traight tubule. In the cortical collecting tubule (CCT), alpha(1) abun dance was equivalent to that in PCT, whereas beta(1) and enzymatic act ivity were both less than one-half of that in PCT. After 2 wk of a K+- deficient diet, alpha(1)- and beta(1)-subunit levels in MCT increased 3.4 +/- 0.6- and 11.7 +/- 4.0-fold, respectively, associated with a 5- fold increase in activity. alpha(2) and alpha(3) were not detected in the CCT or MCT. In conclusion, Na+-K+-ATPase subunit levels can be mon itored at the protein level in dissected tubules, alpha(1)- and beta(1 )-isoform subunits were detected in all tubules assayed, whereas alpha (2) and alpha(3) could not be detected, and the increases in Na+-K+-AT Pase activity in rat MCT from hypokalemic rat can be accounted for by increases in pool sizes of alpha(1)- and beta(1)-subunits.