Lack of correlation between the activation of the Ca 2+-phosphatidylinositol cascade and the regulation of DNA synthesis in the dog thyrocyte

Abstract

Changes in the [Ca 2+] i and/or activation of phospholipase C are thought to participate in the control by several growth factors of the mammalian cell proliferation. It has even been claimed that activation of the Ca 2+-phosphatidylinositol cascade is sufficient to elicit cell proliferation [Jackson et al. (1988) Nature 335, 437–440; Julius et al. (1989) Science 244, 1057–1062]. In this work, we have evaluated the control of DNA synthesis by this cascade in a differentiated epithelial cell model: the dog thyrocyte in primary culture. We first observed that potent activators of the dog thyrocyte Ca 2+-phosphatidylinositol cascade such as carbachol or bradykinin failed to promote the onset of DNA synthesis in these cells. Moreover, carbachol inhibited the mitogenic effect of thyroid stimulating hormone (TSH) and of epidermal growth factor (EGF). The mitogenic effect of EGF was also reduced by bradykinin. Nevertheless, carbachol enhanced the expression of the protooncogenes c-fos and c-myc mRNAs. The time course of this enhancement was identical to the time course for the induction of c-fos and c-myc mRNAs by phorbol esters or EGF. On the other hand, in most experiments, TSH and EGF were able to trigger the onset of dog thyrocyte DNA synthesis without affecting their intracellular free Ca 2+ concentration [Ca 2+] i, 45Ca 2+ efflux, or inositol phosphate generation. In several experiments, TSH increased the dog thyrocyte 45Ca 2+ release and promoted a rise in the [Ca 2+] i or the inositol phosphate accumulation but these effects were weak. In contrast to the effect of carbachol, the TSH effects on the [Ca 2+] i and the 45Ca 2+ efflux appeared slowly, were sustained, and were extremely sensitive to extracellular Ca 2+ depletion. They were observed at hormone concentrations higher than the concentration achieving maximal stimulation of DNA synthesis. Similarly, in a few experiments, a slight increase in the [Ca 2+] i or in the inositol trisphosphate generation were provoked by EGF. However, these modifications were not associated with an increased mitogenic potency of EGF. Finally, in all experiments, fetal calf serum slightly accelerated the dog thyrocyte 45Ca 2+ efflux and increased their inositol phosphate generation. However, the acceleration of the 45Ca 2+ release and the triggering of the dog thyrocyte DNA synthesis were maximal at different concentrations (5 and 20%, respectively). Our results therefore suggest that an early activation of the dog thyrocyte Ca 2+-phosphatidylinositol cascade and the enhancement of the mRNA expression of the protooncogenes c-fos and c-myc are not correlated with the late control of DNA synthesis in these cells.