Calorimetric characterization of critical targets for killing and acquiredthermotolerance in yeast

We characterized thermal behaviours of cellular components by differential scanning calorimetry (DSC) in order to investigate how Saccharomyces cerevi siae cells acquire thermotolerance after heat shock or in stationary phase. Whole-cell DSC profiles consisted of at least five endothermic components over the range 45-75 degrees C for exponentially growing, heat-shocked and stationary-phase cells. In these profiles, we attempted to localize the end othermic profiles due to denaturation of the two critical targets which wer e predicted by using the Arrhenius parameters of hyperthermic killing of th e cells (Obuchi et al., 1998). This prediction indicated that (a) the heat shock stabilized one family of targets and destabilized the other, while (b ) arrest in stationary phase stabilized both targets. Therefore, the heat-s hock response does not stabilize all cellular components, and arrest in sta tionary phase appears to stabilize cellular components in a different manne r from the heat-shock response. It was not possible unambiguously to resolv e the profiles of the critical targets in the DSC scans of whole cells. Com ponents I (T-m=49.7 degrees C) and II (T-m=56.1 degrees C) may both include denaturations of critical targets 1 (T-m=55.4 degrees C) and 2 (T-m=53.0 d egrees C) in exponential cells. Components I and II were both stabilized (T -m=53.5 and 57.2 degrees C, respectively) in heat-shocked cells. Subcellula r fractions suspended in 1.2 M trehalose solution, which mimics the cytosol in tolerant cells, were more stable than those in 0.6 M KCl, which mimics the cytosol in sensitive cells. The microsomal fractions in KCl and trehalo se had endothermic profiles in similar temperature ranges to those predicte d for sensitive and tolerant cells, respectively. This agreement suggests t hat the microsomal fraction may contain critical targets, and that trehalos e accumulation in the heat-shocked and in the stationary phase yeast cells is a stabilizer of cellular components. Copyright (C) 2000 John Wiley & Son s, Ltd.