Effects of Cycloheximide on Thermotolerance Expression, Heat Shock Protein Synthesis, and Heat Shock Protein mRNA Accumulation in Rat Fibroblasts

Abstract

A single hyperthermic exposure can render cells transiently resistant to subsequent high temperature stresses. Treatment of rat embryonic fibroblasts with cycloheximide for 6 h after a 20-min interval at 45 degrees C inhibits protein synthesis, including heat shock protein (hsp) synthesis, and results in an accumulation of hsp 70 mRNA, but has no effect on subsequent survival responses to 45 degrees C hyperthermia. hsp 70 mRNA levels decreased within 1 h after removal of cycloheximide but then appeared to stabilize during the next 2 h (3 h after drug removal and 9 h after heat shock). hsp 70 mRNA accumulation could be further increased by a second heat shock at 45 degrees C for 20 min 6 h after the first hyperthermic exposure in cycloheximide-treated cells. Both normal protein and hsp synthesis appeared increased during the 6-h interval after hyperthermia in cultures which received two exposures to 45 degrees C for 20 min compared with those which received only one treatment. No increased hsp synthesis was observed in cultures treated with cycloheximide, even though hsp 70 mRNA levels appeared elevated. These data indicate that, although heat shock induces the accumulation of hsp 70 mRNA in both normal and thermotolerant cells, neither general protein synthesis nor hsp synthesis is required during the interval between two hyperthermic stresses for Rat-1 cells to express either thermotolerance (survival resistance) or resistance to heat shock-induced inhibition of protein synthesis.