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In order to characterize the property of Ca^(2+) transport in plant cell, microsomes were prepared from the roots of tomato and microsomal ^(45)Ca^(2+) uptake was measured. When 1 mM vanadate, a selective inhibitor of P-type ATPases, 50 mM NO₃^-, a specific inhibitor of vacuolar H^+-ATPase, and both of these inhibitors were treated, the microsomal ^(45)Ca^(2+) uptakes were inhibited by 20, 33 and 47%, respectively. The inhibitory effects of these two inhibitors were investigated by using a protonophore, gramicidin. When the chemical gradient of H^+ was relieved by gramicidin, the uptake was decreased by 30%, implying the presence of Ca^(2+)/H^+ antiporter in the microsomal membrane. In the ^(45)Ca^(2+) uptake experiment, the effect of gramicidin was independent of vanadate-induced inhibition. However, when the activity of vacuolar H^+-ATPase was inhibited by NO₃^-, the effect of gramicidin was severely decreased. Meanwhile, thapsigargin, a specific antagonist of ER/SR-type Ca^(2+)-ATPase, inhibited the microsomal ^(45)Ca^(2+) uptake and the maximum inhibitory effect was obtained at 10 μM. The effect of thapsigargin was blocked by NO₃^- and gramicidin, but not by vanadate. These results imply that vanadate directly inhibits the activity of Ca^(2+)-ATPase; however, NO₃^- and thapsigargin block the activity of Ca^(2+)/H^+ antiporter by inhibiting the vacuolar H^+-ATPase. In conclusion, the microsomal ^(45)Ca^(2+) uptakes are mediated by two major enzymes, Ca^(2+)-ATPase and Ca^(2+)/H^+ antiporter in tomato root tissue.
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