Binding of 14-3-3 protein to the plasma membrane H+-ATPase AHA2 involves the three C-terminal residues Tyr(946)-Thr-Val and requires phosphorylation of Thr(947)

14-3-3 proteins play a regulatory role in a diverse array of cellular funct ions such as apoptosis, regulation of the cell cycle, and regulation of gen e transcription. The phytotoxin fusicoccin specifically induces association of virtually any 14-3-3 protein to plant plasma membrane Hf-ATPase. The 14 -3-3 binding site in the Arabidopsis plasma membrane H+-ATPase AHA2 was loc alized to the three C-terminal residues of the enzyme (Tyr(946)-Thr-Val). f inding of 14-3-3 protein to this target was induced by phosphorylation of T hr(947) (K-D = 88 nM) and was in practice irreversible in the presence of f usicoccin (K-D = 7 nM). Mass spectrometry analysis demonstrated that AHA2 e xpressed in yeast was phosphorylated at Thr(947). We conclude that the extr eme end of AHA2 contains an unusual high-affinity binding site for 14-3-3 p rotein.