Enhanced H+/ATP coupling ratio of H+-ATPase and increased 14-3-3 protein content in plasma membrane of tomato cells upon osmotic shock

Modulation of proton extrusion and ATP-dependent H+ transport through the plasma membrane in relation to the presence of 14-3-3 proteins in this membrane in response to osmotic shock was studied in tomato (Lycopersicon esculentum Mill. cv. Pera) cell cultures. In vivo H+ extrusion by cells was activated rapidly and significantly after adding 100 mM NaCl, 100 mM KCl, 50 mM Na-2 SO4 , 1.6% sorbitol or 2 muM fusicoccin to the medium. The increase in H+ extrusion by cells treated with 100 mM NaCl was correlated with an increase of H+ transport by the plasma membrane H+ -ATPase (EC 3.6.1.35), but not with changes in ATP hydrolytic activity of this enzyme, suggesting an increased coupling ratio of the enzyme. Immunoblot experiments showed increased amounts of 14-3-3 proteins in plasma membrane fractions isolated from tomato cells treated with 100 mM NaCl as compared to control cells without changing the amount of plasma membrane H+-ATPase. Together, these data indicate that in tomato cells an osmotic shock could enhance coupling between ATP hydrolysis and proton transport at the plasma membrane through the formation of a membrane 14-3-3/H+-ATPase complex.