CONTROL OF CL- EFFLUX IN CHARA-CORALLINA BY CYTOSOLIC PH, FREE CA2-MEMBRANE ANION CHANNELS IN CYTOSOLIC PH REGULATION(, AND PHOSPHORYLATIONINDICATES A ROLE OF PLASMA)

Enhanced Cl- efflux during acidosis in plants is thought to play a rol e in cytosolic pH (pH(c)) homeostasis by short-circuiting the current produced by the electrogenic H+ pump, thereby facilitating enhanced H efflux from the cytosol. Using an intracellular perfusion technique, which enables experimental control of medium composition at the cytoso lic surface of the plasma membrane of charophyte algae (Chara corallin a), we show that lowered pH(c) activates Cl- efflux via two mechanisms . The first is a direct effect of pH, on Cl- efflux; the second mechan ism comprises a pH(c)-induced increase in affinity for cytosolic free Ca2+ ([Ca2+](c)), which also activates Cl- efflux. Cl- efflux was cont rolled by phosphorylation/dephosphorylation events, which override the responses to both pH(c) and [Ca2+](c). Whereas phosphorylation (perfu sion with the catalytic subunit of protein kinase A in the presence of ATP) resulted in a complete inhibition of Cl- efflux, dephosphorylati on (perfusion with alkaline phosphatase) arrested Cl- efflux at 60% of the maximal level in a manner that was both pH(c) and [Ca2+](c) indep endent. These findings imply that plasma membrane anion channels play a central role in pH(c) regulation in plants, in addition to their est ablished roles in turgor/volume regulation and signal transduction.