Proliferation arrest and induction of CDK inhibitors p21 and p27 by depleling the calcium store in cultured C6 glioma cells

In this study we investigated the role of the intracellular calcium store in modulating the cellular proliferation and the expression of cell cycle regulatory proteins in cultured C6 glioma cells. By means of microspectrofluorimetry and Ca2+ sensitive indicator fura-2, we found that the intracellular Ca2+ pump inhibitors, thapsigargin (TG) irreversibly and 2,5-di-tert-butyl-hydroquinone (DBHQ) reversibly depleted the Ca2+-store accompanied with the induction of G(0)/G(1) arrest, an increase in glial fibrillary acidic protein (GFAP) expression and morphological changes from a round flat shape to a differentiated spindle-shaped cell. The machinery underlying these changes induced by Ca2+-store depletion was investigated, The results indicated that Ca2+-store depletion caused an increased expression of p21 and p27 proteins (cyclin-dependent kinase inhibitors), with unchanged mutant p53 protein of C6 cells but reduced amounts of the cell cycle regulators: cyclin-dependent kinase 2 (CDK2), cdc2, cyclin C, cyclin D1, cyclin D3 and proliferating cell nuclear antigen (PCNA) in a time-dependent manner. These findings indicate a new function of the endoplasmic reticulum (ER) Ca2+ store in regulating cellular proliferation rate through altering the expression of p21 and p27 proteins. Moreover, cellular differentiation as revealed by spindle-shaped morphology and induced GFAP expression were also modulated by the ER Ca2+ store. The implication of this finding is that the abnormal growth of cancer cells such as C6 glioma cells may be derived from a signalling of the ER which can be manipulated by depleting the Ca2+ store.