Ca²⁺ Transport by Mitochondria from L1210 Mouse Ascites Tumor Cells

Abstract

Mitochondria isolated from the ascites form of L1210 mouse leukemia cells readily accumulate Ca²⁺ from the suspending medium and eject H⁺ during oxidation of succinate in the presence of phosphate and Mg²⁺, with normal stoichiometry between Ca²⁺ uptake and electron transport. Ca²⁺ loads up to 1600 ng-atoms per mg of protein are attained. As is the case in mitochondria from normal tissues, Ca²⁺ uptake takes precedence over oxidative phosphorylation. However, Ca²⁺ transport by the L-1210 mitochondria is unusual in other respects, which may possibly have general significance in tumor cells. The apparent affinity of the L1210 mitochondria for Ca²⁺ in stimulation of oxygen uptake is about 3-fold greater than in normal liver mitochondria; moreover, the maximal rate of Ca²⁺ transport is also considerably higher. Furthermore, when Ca²⁺ pulses are added to L1210 mitochondria in the absence of phosphate or other permeant anions, much larger amounts of Ca²⁺ are bound and H⁺ ejected per atom of oxygen consumed than in the presence of phosphate; up to 7 Ca²⁺ ions are bound per pair of electrons passing each energy-conserving site of the electron-transport chain. Such “superstoichiometry” of Ca²⁺ uptake can be accounted for by two distinct types of respiration-dependent interaction of Ca²⁺ with the L1210 mitochondria. One is the stimulation of oxygen consumption, which is achieved by relatively low concentrations of Ca²⁺ (K_m ≅ 8 μM) and is accompanied by binding of Ca²⁺ up to 40 ng-atoms per mg of protein. The second process, also dependent on electron transport, is the binding of further Ca²⁺ from the medium in exchange with previously stored membrane-bound protons, in which the affinity for Ca²⁺ is much lower (K_m ≅ 120 μM).

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