Exponential and plateau phase Chinese hamster ovary cells were maintained for 3 days at 32, 37, 39 or 41 degrees C. The effect of growth temperature on the fluidity and composition of the cellular membranes, and on the ability of the cells to resist a subsequent heat treatment at 43 degrees C, was measured. Cells grown at temperatures above 37 degrees C displayed increased resistance or tolerance to a 43 degree C heat treatment, whereas cells grown at 32 degrees C were sensitized to heat. Extensive cell division was not required for expression of heat tolerance. Membrane fluidity, as determined by the degree of rotational mobility of the fluorescent probe diphenylhexatriene, decreased with increasing growth temperatures, but the relationship did not hold in exponential phase cells grown at 32 degrees C. The cholesterol : phospholipid molar ratio correlated with the fluorescence polarization values, suggesting that the cells are able to adjust membrane fluidity by varying the concentration of cholesterol. The results are compatible with the concept of homeoviscous adaptation: that organisms strive to maintain an optimal level of membrane fluidity and when grown at a different temperature will alter the lipid composition in order to maintain this level. Up until now, cholesterol has not been implicated in this process.