Effect of cycloheximide on the development of thermotolerance and the synthesis of 68-kilodalton heat shock protein in Chinese hamster V79 and mouse L cells in vitro.

Heat shock protein/Thermotolerance/Cycloheximide The relation of heat shock protein (HSP) synthesis and development of thermotolerance was compared in Chinese hamster V79 cells of low, and mouse L cells of high, thermosensitivities. Thermotolerance was induced by step-up heating (42•Ž, 2 hr and 44•Ž). Cycloheximide did not inhibit the development of ther motolerance in V79 cells. In contrast, no apparent thermotolerance was induced by step-up heating (42•Ž, 1 hr and 44•Ž) in L cells. In V79 cells, as shown by two-dimensional gel electrophoresis, two 68-kilodalton heat shock proteins (68-kDa HSPs) were synthesized constitutively and increased markedly when cells were heated at 42•Ž for 2 hr. Cycloheximide inhibited more than 90% of the total protein synthesis including the 68-kDa HSPs. In contrast, no 68-kDa HSP was detected in L cells at 37•Ž. When L cells were heated at 42•Ž for 1 hr, 68-kDa HSP was detected by fluorography but not by Coomassie blue staining. These results suggest 1) that cells constitutively synthesizing 68-kDa HSP had low thermosensitivity and could be thermotolerant without de novo synthesis of the protein, and 2) that cells not synthesizing 68-kDa HSP had high thermo sensitivity and could not be thermotolerant because the cellular 68-kDa HSP content was low even when the protein was newly synthesized.