ROLE OF HSP70 SYNTHESIS IN THE FATE OF THE INSULIN-RECEPTOR COMPLEX AFTER HEAT-SHOCK IN CULTURED FETAL HEPATOCYTES

The influence of a mild heat shock on the fate of the insulin-receptor complex was studied in cultured fetal rat hepatocytes whose insulin g lycogenic response is sensitive to heat [Zachayus and Plas (1995): J C ell Physiol 162:330-340]. After exposure from 15 min to 2 hr at 42.5 d egrees C, the amount of I-125-insulin associated with cells at 37 degr ees C was progressively decreased (by 35% after 1 hr), while the relea se of I-125-insulin degradation products into the medium was also inhi bited (by 75%), more than expected from the decrease in insulin bindin g. Heat shock did not affect the insulin-induced internalization of ce ll surface insulin receptors but progressively suppressed the recyclin g at 37 degrees C of receptors previously internalized at 42.5 degrees C in the presence of insulin. When compared to the inhibitory effects of chloroquine on insulin degradation and insulin receptor recycling, which were immediate (within 15 min), those of heat shock developed w ithin 1 hr of heating. The protein level of insulin receptors was not modified after heat shock and during recovery at 37 degrees C, while t hat of Hsp72/73 exhibited a transitory accumulation inversely correlat ed with variations in insulin binding, as assayed by Western immunoblo tting from whole cell extracts. Coimmunoprecipitation experiments reve aled a heat shock-stimulated association of Hsp72/73 with the insulin receptor. Affinity labeling showed an interaction between I-125-insuli n and Hsp72/73 in control cells, which was inhibited by heat shock. Th ese results suggest that increased Hsp72/73 synthesis interfered with insulin degradation and prevented the recycling of the insulin recepto r and its further thermal damage via a possible chaperone-like action in fetal hepatocytes submitted to heat stress. (C) 1996 Wiley-Liss, In c.