ACTIVE CALCIUM TRANSPORTERS IN ISOLATED MEMBRANES OF WHEAT ROOT-CELLS

Plant endomembranes carry both primary and secondary ATP-dependent Ca2 + transporters, but conflicting results have been obtained as to their subcellular localization and means of regulation. This study describe s how these transporters can be given a respective characterization, b y proper choice of pH and inhibitors/activators, even under conditions when they cannot physically be separated from each other. The charact erization was done with a light endomembrane fraction from the outer 1 -1.5 cm of wheat root tips (Triticum aestivum L.), free from plasma me mbranes. The endomembrane secondary active ATP-driven Ca2+ transport w as totally inhibited by bafilomycin, was increased several-fold by oxa late and had a pH optimum at 7.4. The primary active ATP-dependent Ca2 + transport was inhibited by vanadate and erythrosin B, was not increa sed by oxalate, and had a relatively lower pH optimum (less than or eq ual to pH 6.8). The two endomembrane Ca2+ transporters were located in different cellular compartments, the secondary H+/Ca2+ anti-port loca ted in the vacuolar membranes, and the primary Ca2+ ATPase located in the endoplasmic reticulum. A Ca2+ ATPase in wheat root plasma membrane s was characterized in an earlier study (OIbe and Sommarin, 1991). Thu s, two primary Ca2+ ATPases with different locations can be distinguis hed in wheat root cells, The endomembrane primary Ca2+ transport showe d many similarities to that of the plasma membranes, e.g. sensitivity to vanadate and to erythrosin B (although higher erythrosin B was need ed to inhibit the endomembrane activity), and stimulation by calmoduli n (provided the plasma membranes had been extensively washed), Both en zymes showed affinity for Ca2+ in the low micromolar range, but differ ed in pH preference and substrate specificity. Taken together, they ar e clearly distinct enzymes located in different subcellular compartmen ts.