ERcalcistorin/protein-disulfide isomerase acts as a calcium storage protein in the endoplasmic reticulum of a living cell: Comparison with calreticulin and calsequestrin

ERcalcistorin/protein-disulfide isomerase (ECaSt/PDI), a high capacity low affinity Ca²⁺-binding protein in the endoplasmic reticulum of sea urchin eggs (Lebeche, D., and Kaminer, B. (1992) Biochem. J. 287, 741-747), shares 55% sequence identity with mammalian PDI and has PDI activity (Lucero, H., Lebeche, D., and Kaminer, B. (1994) J. Biol. Chem. 269, 23112-23119). We report on ECaSt/PDI functioning as a Ca²⁺ storage protein in the endoplasmic reticulum (ER) of a living cell and compare it with calsequestrin and calreticulin, high capacity low affinity Ca²⁺-binding proteins in the sarcoplasmic reticulum and ER, respectively. Stably transfected Chinese hamster ovary cell clones expressed these proteins, which were localized in the ER of the cell. Microsomes from cells expressing ECaSt/PDI, calreticulin, and calsequestrin accumulated 17.2 ± 0.27, 20.0 ± 0.82, and 38.0 ± 0.28 nmol of Ca²⁺/mg of protein, respectively; control microsomes accumulated from 2.6 ± 0.17 to 2.9 ± 0.14 nmol of Ca²⁺/mg of protein. The initial rate of Ca²⁺ uptake was similar in microsomes from transfected and control cells. Microsomes containing an ECaSt/PDI mutant in which 45% of the acidic residue pairs in the C terminus were truncated had a reduced Ca²⁺ storage capacity. This supports our previous hypothesis that the degree of low affinity Ca²⁺ binding is dependent on the number of pairs of carboxyl groups in the molecule. The maximal Ca²⁺ accumulation by microsomes containing the expressed ECaSt/PDI, C-terminally truncated ECaSt/PDI, calreticulin, or calsequestrin correlates approximately with the Ca²⁺ binding capacity of the respective proteins.