INACTIVATION OF EUKARYOTIC INITIATION-FACTOR 2B IN-VITRO BY HEAT-SHOCK

Protein synthesis in rat H35 Reuber hepatoma cells is rapidly inhibite d on heat shock. At mild heat-shock temperatures the main cause for in hibition is the inactivation of the guanine nucleotide exchange factor eukaryotic initiation factor 2B (eIF2B); under more severe heat-shock conditions the activity of several initiation factors is compromised. eIF2B is required for GDP/GTP exchange on eIF2, which delivers methio nyl-tRNA to the 40 S ribosomal subunit. We have tried to elucidate the mechanism underlying the inactivation of eIF2B by assays in vitro. In cubation of cell extracts at 41 degrees C or higher led to the inactiv ation of eIF2B. In agreement with observations in cells exposed to mil d heat shocks, the thermal inactivation of eIF2B could be ascribed to neither eIF2 alpha phosphorylation nor the induction of another inhibi tor. With the use of glycerol gradients we show that eIF2B forms aggre gates in heat-treated extracts. Furthermore eIF2B activity is protecte d against heat shock in thermotolerant cells. Taken together, these re sults suggest a role for chaperones in the control of eIF2B activity.