Bax gene expression alters Ca²⁺ signal transduction without affecting apoptosis in an epithelial cell line

Bax is an oncogene that has proapoptotic properties but not all cells that express Bax undergo apoptosis. Bax may have a function unrelated to apoptosis. To elucidate the role of Bax in cell signaling, an epithelial cell line called SMG-C6 was transfected with the human bax gene. Stable transfectants were studied for their response to carbachol, a muscarinic receptor agonist, by measuring the increase in intracellular free Ca²⁺ and Ca²⁺ influx. Carbachol-mediated release of Ca²⁺ from intracellular stores was significantly higher in Bax transfectants compared to control transfectants (empty vector). Ca²⁺ influx was also increased in Bax transfectants. Bax had no affect on the storage operated channels. However, the concentration of Ca²⁺ in the intracellular stores (i.e., mitochondria and granules) was 40-50% lower in the Bax transfectants. There was no significant difference in thapsigargin-mediated apoptosis in Bax transfectants compared to wild-type and control transfectants. Measurement of glutathione was reduced in the Bax transfectant. Restoration of glutathione levels with glutathione monoethyl ester partially normalized Ca²⁺ mobilization and storage capacity in the mitochondria to control levels. This study shows that sub-apoptotic levels of Bax can reduce Ca²⁺ content in intracellular stores and Ca²⁺ homeostasis. Bax may mediate these effects by reducing the levels of antioxidants resulting in mild oxidative stress.