Protontranslocating activity of plasma membrane vesicles from the tubers of initial and transgenic potato Solanum tuberosum L. plants

The protontranslocating activity of plasma membrane vesicles (PMV) from the tubers of initial potato plants was compared with that of transformed plants carrying the defensine gene from Amaranthus caudatus. The ATP-dependent accumulation of H+ into PW was assessed by the initial rate of acridine orange (AO) fluorescence quenching. The addition of ATP to the incubation medium led to the instantaneous raise in the AO fluorescence intensity followed by its decrease. The fluorescence quenching was observed neither in the presence of the protonophore ClCCP, nor when inhibitors of the membrane-bound HI-ATPase were added to the medium. We conclude that the ATP-induced quenching of AO fluorescence resulted from the H+ accumulation in PMV due to the functioning of the plasma membrane-bound H(+)ATPase. In the PMV samples from transgenic and control plants, the patterns of the initial rate of the AO fluorescence quenching were similar. They differed neither in dependence on the concentration of plasma membrane protein and Mg+2, nor in sensitivity to ClCCP, DCC, and Na3VO4. The rate of passive H+ efflux from PMV of initial and transformed plants also was the same. We also studied the effect of synthetic growth regulators - epibrassinolide-694 and ambiol on proton translocation across the membrane of PMV from the tubers of initial and transformed plants. Both regulators stimulated H+ accumulation in PMV from the tubers of initial plants. In the samples form transformed plants, epibrassinolide did not influence ATP-driven H+ accumulation, and sensitivity to ambiol was lesser than in control plants. It is proposed that introduction of defensine gene from Amaranthus caudatus in potato plant does not affect directly the activity of plasma membrane proton pump, but influences its sensitivity to growth regulators.