Identification of the phosphorylation site for cAMP-dependent protein kinase on Na+,K(+)-ATPase and effects of site-directed mutagenesis.
- G Fisone,
- S X Cheng,
- A C Nairn,
- A J Czernik,
- H C Hemmings, Jr,
- J O Höög,
- A M Bertorello,
- R Kaiser,
- T Bergman and
- H Jörnvall
- Laboratory of Molecular and Cellular Neuroscience, Rockefeller University, New York, New York 10021.
Abstract
Phosphorylation of purified Na+,K(+)-ATPase by cAMP-dependent protein kinase (protein kinase A) decreases the activity of this enzyme. We have now shown, using several experimental approaches, that a highly conserved seryl residue on the catalytic (alpha) subunit of Na+,K(+)-ATPase, corresponding to Ser943 of the rat alpha 1 isoform, is the phosphorylation site for protein kinase A. cDNAs corresponding to wild-type Na+,K(+)-ATPase and Na+,K(+)-ATPase in which Ser943 was mutated to Ala were transfected into COS cells. Treatment of the transfected cells with forskolin plus 3-isobutyl-1-methylxanthine resulted in a decrease in the activity of the wild-type enzyme but not in that of the mutated enzyme. The results suggest that, in intact cells, the activity of the Na+,K(+)-ATPase is regulated in part by signal transduction pathways that use protein kinase A-dependent phosphorylation of the Na+,K(+)-ATPase alpha subunit.
