Mechanisms of fusicoccin action: evidence for concerted modulations of secondary K+ transport in a higher plant cell

Fusicoccin (FC) has long been known to promote K+ uptake in higher plant cells, including stomatal guard cells, yet the precise mechanism behind this enhancement remains uncertain. Membrane hyperpolarization, thought to arise from primary H+ pumping stimulated in FC, could help drive K+ uptake, but the extent to which FC stimulates influx and uptake frequently exceeds any reasonable estimates from Constant Field Theory based on changes in the free-running membrane potential (Vm) alone; furthermore, unidirectional flux analyses have shown that in the toxin K+ (86Rb+) exchange plummets to 10% of the control (G.M. Clint and E.A.C. MacRobbie 1984, J. Exp. Bot.35 180–192). Thus, the activities of specific pathways for K+ movement across the membrane could be modified in FC. We have explored a role for K+ channels in mediating these fluxes in guard cells ofVicia faba L. The correspondence between FC-induced changes in chemical (86Rb+) flux and in electrical current under voltage clamp was followed, using the K+ channel blocker tetraethylammonium chloride (TEA) to probe tracer and charge movement through K+-selective channels. Parallel flux and electrical measurements were carried out when cells showed little evidence of primary pump activity, thus simplifying analyses. Under these conditions, outward-directed K+ channel current contributed appreciably to charge balance maintainingVm, and adding 10 mM TEA to block the current depolarized (positive-going)Vm; TEA also reduced86Rb+ efflux by 68–80%. Following treatments with 10 μM FC, both K+ channel current and86Rb+ efflux decayed, irreversbly and without apparent lag, to 10%–15% of the controls and with equivalent half-times (approx. 4 min). Fusicoccin also enhanced86Rb+ influx by 13.9-fold, but the influx proved largely insensitive to TEA. Overall, FC promotednet cation uptake in 0.1 mM K+ (Rb+), despite membrane potentials which were 30–60 mVpositive of the K+ equilibrium potential. These results tentatively link (chemical) cation efflux to charge movement through the K+ channels. They offer evidence of an energy-coupled mechanism for K+ uptake in guard cells. Finally, the data reaffirm early suspicions that FC alters profoundly the K+ transport capacity of the cells, independent of any changes in membrane potential.