Thermotolerance induced by heat, sodium arsenite, or puromycin: Its inhibition and differences between 43°C and 45°C

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Abstract
When CHO cells were treated either for 10 min at 45–45.5°C or for 1 hr with 100 μM sodium arsenite (ARS) or for 2 hr with 20 μg/ml puromycin (PUR-20), they became thermotolerant to a heat treatment at 45–45.5°C administered 4–14 hr later, with thermotolerance ratios at 10−3 isosurvival of 4–6, 2–3.2, and 1.7, respectively. These treatments caused an increase in synthesis of HSP families (70, 87, and 110 kDa) relative to total protein synthesis. However, for a given amount of thermotolerance, the ARS and PUR-20 treatments induced 4 times more synthesis than the heat treatment. This decreased effectiveness of the ARS treatment may occur because ARS has been reported to stimulate minimal redistribution of HSP-70 to the nucleus and nucleolus. Inhibiting protein synthesis with cycloheximide (CHM, 10 μg/ml) or PUR (100 μg/ml) after the initial treatments greatly inhibited thermotolerance to 45–45.5°C in all cases. However, for a challenge at 43°C, thermotolerance was inhibited only for the ARS and PUR-20 treatments. CHM did not suppress heat-induced thermotolerance to 43°C, which was the same as heat protection observed when CHM was added before and during heating at 43°C without the initial heat treatment. These differences between the initial treatments and between 43 and 45°C may possibly be explained by reports that show that heat causes more redistribution of HSP-70 to the nucleus and nucleolus than ARS and that redistribution of HSP-70 can occur during heating at 42°C with or without the presence of CHM. Heating cells at 43°C for 5 hr after thermotolerance had developed induced additional thermotolerance, as measured with a challenge at 45°C immediately after heating at 43°C. Compared to the nonthermotolerant cells, thermotolerance ratios were 10 for the ARS treatment and 8.5 for the initial heat treatment. Adding CHM (10 μg/ml) or PUR (100 μg/ml) to inhibit protein synthesis during heating at 43°C did not greatly reduce this additional thermotolerance. If, however, protein synthesis was inhibited between the initial heat treatment and heating at 43°C, protein synthesis was required during 43°C for the development of additional thermotolerance to 45°C. These data suggest that if a considerable amount of synthesis of HSP families occurred after the initial treatment before heating at 43°C, redistribution during 43°C of the previously synthesized HSP families could lead to the additional thermotolerance to 45°C.