EVIDENCE FOR K+ CHANNEL CONTROL IN VICIA GUARD-CELLS COUPLED BY G-PROTEINS TO A 7TMS RECEPTOR MIMETIC

To explore the involvement of a class of seven-transmembrane-span (7TMS) receptors in cellular signalling, a synthetic analogue (mas7) of the amphipathic tetradecapeptide mastoparan was used to mimic hormonal stimulus in guard cells of Vicia faba. The ability for mas7 to substitute for an activated receptor complex was assayed by the effect on guard cell ion channel activities in the absence of any hormonal stimulus. Currents carried by inward(I-K,I-in and outward- (I-K,I-out) rectifying potassium channels were determined under voltage clamp conditions before, during, and after exposure to mas7. The dominant effect of mas7 was to inactivate I-K,I-in within 30 sec of application. By contrast, I-K,I-out was largely unaffected under these conditions. The effect of mas7 on I-k,I-in was both concentration and voltage-dependent. At any one clamp voltage, mas7 inactivation showed Michaelian behaviour, with a mean K-i of 0.05 +/- 0.02 mu M at -240 mV. Increasing mas7 concentration also shifted the voltage for half-maximal activation of the current negative, with 0.5 mu M mas7 effecting a -13 +/- 2 mV displacement and lengthening the halftime for activation of the current by up to threefold. By contrast, the non-amphipathic analogue of mas7, masCP, had no appreciable effect on the steady-state current or its activation kinetics; nor was the poly-cation polylysine able to substitute for mas7 in its action on the K+ channels. Application of the non-hydrolysable analogue of GDP, GDP-beta-S, either by iontophoresis or by diffusion from the microelectrode, effectively blocked mas7-induced inactivation of I-K,I-in. These, and additional results provide in vivo evidence for the involvement of G-protein-linked 7TMS receptors in the regulation of membrane transport in a higher plant cell.