Characterization of the effects of Ca2+ depletion on the synthesis, phosphorylation and secretion of caseins in lactating mammary epithelial cells.

We have examined the effects of depleting lumenal Ca2+ on the synthesis, phosphorylation and secretion of caseins in lactating mouse mammary cells by using inhibitors of the endoplasmic reticulum Ca(2+)-ATPase or the ionophore ionomycin in the absence of external Ca2+. Treatment with these drugs resulted in a transient increase in the cytosolic Ca2+ concentration due to Ca2+ mobilization. Protein synthesis over a 1 h period was substantially inhibited by Ca2+ depletion, but in a pulse-chase protocol secretion of pre-synthesized proteins was unaffected by Ca2+ depletion. Analysis of polysome profiles showed that Ca2+ depletion resulted in a loss of polysomes, consistent with an inhibition of initiation of protein synthesis. Neither treatment with Ca(2+)-ATPase inhibitors to deplete endoplasmic reticulum Ca2+ nor treatment with ionomycin/EGTA had any effect on an early phase of phosphorylation of alpha- or beta/gamma-caseins, but Ca2+ depletion resulted in a decrease in a late phase of casein phosphorylation. These results indicate that lumenal Ca2+ is required to maintain protein synthesis in lactating mammary cells but is not required for protein secretion, and that Ca2+ accumulation in the Golgi cisternae is required for a late but not for an early phase of casein phosphorylation.