Heat shock protein synthesis is affected by intracellular pH: inhibition by monensin-induced alkalosis in C6 rat glioma cells

Abstract

The effect of intracellular pH (pH;) on heat shock protein (HSP) synthesis was investigated in C6 rat glioma cells. pH i changes were analysed by means of fluorescence spectroscopy in a perfused monitoring system allowing continuous measurements before, during and after treatments. HSP induction was determined by means of Western blots and autoradiographs. A 20 min heat shock (HS) of 44°C decreased the pH i from 7.36 to 7.05 during exposure [171 and elicited the synthesis of heat shock proteins 2–8 h later. A pH i decrease, brought about by low extracellular pH (pH e) of 4.5 and 5.0 or 5.5, induced HSP synthesis after 1 h or 3 h, respectively. During these treatments, pH i decreased to values significantly lower than that caused by HS. Three h exposure to pH e 6.2, however, was not inductive. These results indicate that the heat shock-induced pH i decrease alone is not sufficient to stimulate HSP synthesis. In order to investigate the effect of alkaline pH i on the induction of HSP by heat, pH i was increased prior to HS treatments. Preincubation of cells at pH e ranging from 6.8 to 8.0 had little effect on pH i and on HSP synthesis. A shift of pH i to more alkaline values was achieved by adding the H +/Na + exchanger monensin at alkaline pH e. Twenty μM monensin raised the pH i and inhibited the HSP induction depending on the pH e values: as pH e was increased from pH 7.2 to 8.0 HSP synthesis was increasingly inhibited. Monensin also diminished the HS-induced drop of pH i particularly at higher pH e. The result showed that neither a lower pH i nor a drop of pH i during HS is a necessary prerequisite for the induction, whereas alkalosis inhibits the synthesis of HSP.