Involvement of protein synthesis in the development of thermotolerance using a CHO temperature-sensitive mutant.

A Chinese hamster ovary temperature-sensitive mutant (CHO-tsH1) with defective leucyl-tRNA synthetase at temperatures greater than 39 degrees C was used to examine the importance of protein synthesis in the development of thermotolerance. Its wild-type parent CHO-SC1 was used as the control. At temperatures of 41.5 degrees C, 42 degrees C and 42.5 degrees C, SC1 showed the classical biphasic thermotolerant response while tsH1 showed no thermotolerance. When both cell lines were heated for 15 min at 45 degrees C, then allowed to incubate at the permissive temperature of 35 degrees C and finally challenged with another 25 min treatment at 45 degrees C, tolerance was expressed in both cell lines. When the development incubation temperature was raised from 35 degrees C to the non-permissive temperature of 40 degrees C, tolerance was also observed. Although both cell lines expressed tolerance under these conditions, the magnitude and duration of response of the mutant cell line were reduced. Heat-shock protein analysis using one-dimensional gel electrophoresis showed that, under permissive conditions, the mutant cell was able to express the full spectrum of heat-shock proteins as seen in the wild-type cells. Under non-permissive conditions, little or no detectable proteins were synthesized in the mutant cell. We therefore postulate that the synthesis of new cytosol proteins is not required for the initial onset of thermotolerance but is necessary for the sustenance of tolerance.