Biochemical characterization of dephosphorylation of the plasma membrane H⁺-ATPase

Abstract

Plant plasma membrane H⁺-ATPase creates an electrochemical gradient of H⁺ across the plasma membrane. Recent studies have demonstrated that the H⁺-ATPase is activated via phosphorylation on a penultimate threonine residue in the C-terminus and subsequent binding of 14-3-3 protein to the phosphorylated C-terminus. However, biochemical properties of protein kinase and protein phosphatase, which regulate phosphorylation level of the H⁺-ATPase, are largely unknown. In this study, we investigated in vitro dephosphorylation of the H⁺-ATPase. Dephosphorylation of the H⁺-ATPase was detected in the plasma membrane from etiolated seedlings of Arabidopsis. Furthermore, the dephosphorylation was inhibited by EDTA, a chelating agent for divalent cations, but not by calyculin A, an inhibitor of type 1 and type 2A protein phosphatases. These results suggest that a divalent cation-dependent protein phosphatase localized in the plasma membrane is involved in dephosphorylation of the H⁺-ATPase. We will report analysis of the H⁺-ATPase complex.