Angiotensin II stimulates elution of Na-K-ATPase from a digoxin-affinity column by increasing the kinetic response to ligands that trigger the decay of E2-P.
We earlier observed that treating rat proximal tubules with concentrations of angiotensin II (ANG II) that directly stimulate Na-K-ATPase activity changed how Na-K-ATPase subsequently eluted from an ouabain-affinity column. In this study we tested whether ANG II increases the rate of elution in response to ligands that trigger the decay of E(2)-P, which implies a change in functional properties of Na-K-ATPase, or by decreasing the amount subsequently eluted with SDS, which suggests a change in how Na-K-ATPase interacts with other proteins. We utilized a new digoxin-affinity column and novel lines of opossum kidney (OK) cells that coexpress the rat AT(1a) receptor and either the wild-type rat alpha(1)-isoform of Na-K-ATPase or a truncation mutant missing the first 32 amino acids of its NH(2) terminus. We characterized how rat kidney microsomes bind to and elute from the digoxin-affinity column and demonstrated that they are heterogeneous in the rate at which they release digoxin in response to ligands that trigger the decay of E(2)-P. Incubating OK cells with ANG II stimulated the ensuing elution of wild-type rat alpha(1)-subunit by increasing the kinetic response to ligands that cause a decay of E(2)-P without affecting the amount later eluted with SDS. In contrast, ANG II had no effect on the kinetic response of the truncation mutant but decreased the amount eluted with SDS. These data suggest that ANG II regulates both the kinetic properties of Na-K-ATPase and its interaction with other proteins by a mechanism(s) involving its NH(2) terminus.
Yingst
Douglas R
DR
Dept. of Physiology, Wayne State Univ. School of Medicine, 540 E. Canfield Ave., Detroit, MI 48201, USA. dyingst@med.wayne.edu
Doci
Tabitha M
TM
Massey
Katherine J
KJ
Rossi
Noreen F
NF
Rucker
Ebony
E
Mattingly
Raymond R
RR
D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D002846:Chromatography, Affinity; D004077:Digoxin; D015232:Dinoprostone / Q000494:pharmacology; D007668:Kidney / Q000201:enzymology; D007687:Kidney Tubules, Proximal / Q000201:enzymology; D007700:Kinetics; D008861:Microsomes / Q000201:enzymology; D051381:Rats; D000254:Sodium-Potassium-Exchanging ATPase / Q000302:isolation & purification / Q000378:metabolism*
D000804:Angiotensin II; D004077:Digoxin; D000254:Sodium-Potassium-Exchanging ATPase; D015232:Dinoprostone