The effect of sodium malonate on yeast thermotolerance

The study of the effect of malonate (an inhibitor of the succinate dehydrogenase complex of the respiratory chain of mitochondria) on the thermotolerance of the fermentative Saccharomyces cerevisiae and nonfermentative Rhodotorula rubra yeasts showed that malonate augmented the damaging effect of heat shock on the yeasts utilizing glucose (or other sugars) by means of oxidative phosphorylation. At the same time, malonate did not influence and sometimes even improved the thermotolerance of the yeasts utilizing glucose through fermentation. The suggestion is made that cell tolerance to heat shock depends on the normal functioning of mitochondria. On the other hand, their increased activity at elevated temperatures may accelerate the formation of cytotoxic reactive oxygen species and, hence, is not beneficial to cells.     

The Effect of Cytochrome Oxidase Inhibitors on the Thermotolerance of Yeasts

The investigation of the effect of the cytochrome oxidase inhibitors sodium cyanide and sodium azide on the thermotolerance of the yeasts Rhodotorula rubra, Debaryomyces vanriji, and Saccharomyces cerevisiae showed that these inhibitors diminish the thermotolerance of R. rubraand D. vanriji, but do not affect the thermotolerance of S. cerevisiae. Taking into account the fact that, unlike the latter yeast, R. rubra and D. vanriji are nonfermentative yeasts, the difference in the effects of the inhibitors on the yeast thermotolerance can be readily explained by the different types of glucose utilization (either oxidative or fermentative) in these yeasts. The data obtained also provide evidence that there is a correlation between the functional activity of mitochondria and the thermotolerance of yeast cells.     

The Effect of Sodium Azide on the Thermotolerance of the Yeasts Saccharomyces cerevisiae and Candida albicans

The addition of sodium azide (a mitochondrial inhibitor) at a concentration of 0.15 mM to glucose-grown Saccharomyces cerevisiae or Candida albicans cells before exposing them to heat shock increased cell survival. At higher concentrations of azide, its protective effect on glucose-grown cells decreased. Furthermore, azide, even at low concentrations, diminished the thermotolerance of galactose-grown yeast cells. It is suggested that azide exerts a protective effect on the thermotolerance of yeast cells when their energy requirements are met by the fermentation of glucose. However, when cells obtain energy through respiratory metabolism, the azide inhibition of mitochondria enhances the damage inflicted on the cells by heat shock.     

The Absence of a Direct Relationship between the Ability of Yeasts to Grow at Elevated Temperatures and Their Survival after Lethal Heat Shock

The study of the growth of the yeasts Rhodotorula rubra, Saccharomyces cerevisiae, and Debaryomyces vanriji at elevated temperatures and their survival after transient lethal heat shock showed that the ability of these yeasts to grow at supraoptimal temperatures (i.e., their thermoresistance) and their ability to tolerate lethal heat shocks (i.e., their thermotolerance) are determined by different mechanisms. It is suggested that the thermotolerance of the yeasts is mainly determined by the division rate of cells before their exposure to heat shock.     

Effect of Sodium Azide on the Thermotolerance of the Yeasts Saccharomyces cerevisiaeand Debaryomyces vanriji

The pretreatment of Saccharomyces cerevisiaeand Debaryomyces vanrijiwith sodium azide was found to induce thermotolerance in both yeasts, whereas sodium azide used in combination with heat shock enhanced the thermotolerance of S. cerevisiaeand substantially decreased the thermotolerance of D. vanriji.It is suggested that the different responses of the yeasts to sodium azide during heat shock are due to the different functional organizations of their mitochondrial apparatus.     

The Effect of Sodium Azide on Basic and Induced Thermotolerance in Saccharomyces cerevisiae

The action mechanism of the mitochondrial inhibitor sodium azide on thermotolerance in Saccharomyces cerevisiae was studied. At ambient growth temperature, pretreatment with sodium azide was shown to improve the thermotolerance of parent cells and the hsp104 mutant. Treating with the inhibitor during a mild heat shock suppressed the development of induced thermotolerance due to the inhibition of heat shock protein (Hsp104) synthesis. Treating with the inhibitor immediately before lethal heat shock produced a variety of effects on thermotolerance depending on whether the yeast metabolism was oxidative or fermentative. The conclusions are: (1) the protective effect of sodium azide on the thermotolerance of S. cerevisiae cells grown on glucose-containing medium is not related to Hsp104 functioning, and (2) the mechanisms of basic and induced thermotolerance differ considerably.     

Thermotolerance,cell filamentation,and induced protein synthesis in psychrophilic and psychrotrophic bacteria

Both the psychrophile Aquaspirillum arcticum and the psychrotroph Bacillus psychrophilus were found to acquire thermotolerance when either heat shocked or treated with nalidixic acid; two conditions which also resulted in the induction of heat shock proteins and/or stress proteins and also cell filamentation. The possible relatedness of acquisition of thermotolerance and cell filamentation was examined by inhibiting cell filamentation with 1.5% KCl. A. arcticum cells which were heat shocked in the presence of KCl did not become filamentous nor acquire thermotolerance suggesting that these two responses may be related. On the other hand, when cells of B. psychrophilus were treated in a similar fashion, they also were prevented from cell filamentation but their ability to become thermotolerant was unaffected. When A. arcticum cells were heat shocked in the presence of chloramphenicol, heat shock protein synthesis was inhibited but not the acquistion of thermotolerance. Similar experiments with B. psychrophilus revealed that partial induction of heat shock proteins still occurred; however, no thermotolerance was exhibited.Abbreviations hsp(s) heat shock proteins(s) - SEM standard error of the mean     

Impact of mild heat treatments on induction of thermotolerance in the biocontrol yeast Candida sake CPA-1 and viability after spray-drying

Aims: The objective of this study was to examine the induction of thermotolerance in the biocontrol agent Candida sake CPA‐1 cells by mild heat treatments to enhanced survival of formulations using spray‐drying. The possible role of heat‐shock proteins (HSPs) biosynthesis in induced thermotolerance and the role of sugars and sugar alcohols were also determined. Methods and Results: Studies were conducted on C. sake cells grown in molasses medium and exposed to mild temperatures of 30 and 33°C during mid‐ (16 h), late‐exponential (24 h), early‐ (30 h) and mid‐stationary (36 h) growth phases. The effect on viability was determined both before and after spray‐drying. Cycloheximide and chloramphenicol were used to examine the role of HSPs and HPLC was used to analyse the accumulation of sugar and sugar alcohols. The results indicate that both temperatures induced thermotolerance in cells of C. sake. Mild heat‐adapted cells at 33°C in the early‐ or mid‐stationary phases had survival values after spray‐drying significantly higher (P ≤ 0·05) than nonadapted cells. However, viabilities were not high enough to be considered for commercial use with values up to 17%. HSPs were not implicated in thermotolerance acquired by mild heat‐adapted cells as similar viabilities were obtained in the presence of protein inhibitors. Little change was observed in sugar and sugar alcohols with an increase in glucose and arabitol in some treatments. Conclusions: This study suggests that it is possible to induce thermotolerance in biocontrol yeasts such as C. sake. However, this does not improve survival of cells exposed to spray‐drying sufficiently to consider this a suitable formulation method for this biocontrol agent. HSPs, sugars and sugar polyols were not directly responsible for induced thermotolerance in yeast cells. Significance and Impact of the Study: This type of information can be effectively applied to improve the viability of cells in the process of formulation.     

Sodium azide reduces the thermotolerance of respiratively grown yeasts

The effect of sodium azide in heat shock-induced cell death was studied in Debaryomyces vanrijiae, Candida albicans, and Saccharomyces cerevisiae yeasts. The results presented demonstrate that the azide addition induced a drastic decrease in the thermotolerance of glucose-grown D. vanrijiae. In contrast, glucose-grown S. cerevisiae and C. albicans cells treated with NaN₃ became more resistant to heat shock than control cells. Nevertheless, in galactose medium the decrease of thermotolerance of S. cerevisiae and C. albicans cells was observed in the presence of sodium azide. It was suggested that the decreasing effect of sodium azide on thermotolerance takes place only when the yeast cell is incapable of using fermentation for ATP synthesis and obtains energy via oxidative phosphorylation. Received: 27 December 2001 / Accepted: 27 February 2002     

Effect of cytochrome oxidase inhibitors on the yeast thermotolerance

The investigation of the effect of the cytochrome oxidase inhibitors sodium cyanide and sodium azide on the thermotolerance of the yeasts Rhodotorula rubra, Debaryomyces vanriji, and Saccharomyces cerevisiae showed that these inhibitors diminish the thermotolerance of R. rubra and D. vanriji, but do not affect the thermotolerance of S. cerevisiae. Taking into account the fact that, unlike the latter yeast, R. rubra and D. vanriji are nonfermentative yeasts, the difference in the effects of the inhibitors on the yeast thermotolerance can be readily explained by the different types of glucose utilization (either oxidative or fermentative) in these yeasts. The data obtained also provide evidence that there is a correlation between the functional activity of mitochondria and the thermotolerance of yeast cells.     

The Induction of Saccharomyces cerevisiae Hsp104 Synthesis by Heat Shock Is Controlled by Mitochondria

Heat shock protein Hsp104 of Saccharomyces cerevisiae functions as a protector of cells against heat stress. When yeast are grown in media containing nonfermentable carbon sources, the constitutive level of this protein increases, which suggests an association between the expression of Hsp104 and yeast energy metabolism. In this work, it is shown that distortions in the function of mitochondria appearing as a result of mutation petite or after exposure of cells to the mitochondrial inhibitor sodium azide reduce the induction of Hsp104 synthesis during heat shock. Since the addition of sodium azide suppressed the formation of induced thermotolerance in the parent type and in mutant hsp104,the expression of gene HSP104 and other stress genes during heat shock is apparently regulated by mitochondria.     

The effect of sodium azide on the thermotolerance of the yeast Saccharomyces cerevisiae and Candida albicans

The addition of sodium azide (a mitochondrial inhibitor) at a concentration of 0.15 mM to glucosegrown Saccharomyces cerevisiae or Candida albicans cells before exposing them to heat shock increased cell survival. At higher concentrations of azide, its protective effect on glucose-grown cells decreased. Furthermore, azide, even at low concentrations, diminished the thermotolerance of galactose-grown yeast cells. It is suggested that azide exerts a protective effect on the thermotolerance of yeast cells when their energy requirements are met by the fermentation of glucose. However, when cells obtain energy through respiratory metabolism, the azide inhibition of mitochondria enhances damage inflicted on the cells by heat shock.     

The absence of direct relationship between the ability of yeasts to grow at elevated temperatures and their survival after lethal heat shock

The study of the growth of the yeasts Rhodotorula rubra, Saccharomyces cerevisiae, and Debaryomyces vanriji at elevated temperatures and their survival after transient lethal heat shock showed that the ability of these yeasts to grow at supraoptimal temperatures (i.e., their thermoresistance) and their ability to tolerate lethal heat shocks (i.e., their thermotolerance) are determined by different mechanisms. The thermotolerance of the yeasts is suggested to be mainly determined by the division rate of cells before their exposure to heat shock.     

Stress proteins and stress tolerance in an Antarctic, psychrophilic yeast, Candida psychrophila

Conditions are described for the heat shock acquisition of thermotolerance, peroxide tolerance and synthesis of heat shock proteins (hsps) in the Antarctic, psychrophilic yeast Candida psychrophila. Cells grown at 15°C and heat shocked at 25°C (3 h) acquired tolerance to heat (35°C) and hydrogen peroxide (100 mM). Novel heat shock inducible proteins at 80 and 110 kDa were observed as well as the presence of hsp 90, 70 and 60. The latter hsps were not significantly heat shock inducible. The absence of hsp 104 was intriguing and it was speculated that the 110 kDa protein may play a role in stress tolerance in psychrophilic yeasts, similar to that of hsp 104 in mesophilic species.     

Effect of sodium azide on heat-shock resistance in Saccharomyces cerevisiae and Debaryomyces vanriji yeasts]

The pretreatment of Saccharomyces cerevisiae and Debaryomyces vanriji with sodium azide was found to induce thermotolerance in both yeasts, whereas sodium azide used in combination with heat shock enhanced the thermotolerance of S. cerevisiae and substantially decreased the thermotolerance of D. vanriji. It is suggested that the different responses of the yeasts to sodium azide during heat shock are due to the different functional organizations of their mitochondrial apparatus.     

Ethanol treatment triggers a heat shock-like response but no thermotolerance in soybean (Glycine max cv. Kaohsiung No.8) seedlings

Non‐lethal heat‐shock (HS) treatment has previously been shown to induce thermotolerance in soybean (Glycine max cv. Kaohsiung No.8) seedlings. This acquired thermotolerance correlates with the de novo synthesis of heat‐shock proteins (HSPs). Interestingly, we found that ethanol treatments also elicited HS‐like responses in aetiolated soybean seedlings at their normal growth temperature of 28 °C. Northern blot analyses revealed that the expression of HS genes hsp17.5, hsp70 and hsc 70 was induced by ethanol. Radioactive amino acids were preferentially incorporated into high molecular weight (HMW) HSPs rather than class I low molecular weight (LMW) HSPs during non‐lethal ethanol treatments. Immunoblot analysis confirmed that no accumulation of class I LMW HSPs occurred after non‐lethal ethanol treatment. Pre‐treatment with a non‐lethal dose of ethanol did not provide thermotolerance, as the aetiolated soybean seedlings could not survive a subsequent heat shock of 45 °C for 2 h. In contrast, non‐lethal HS pre‐treatment, 40 °C for 2 h, conferred tolerance on aetiolated soybean seedlings to otherwise lethal treatments of 7·5% ethanol for 8 h or 10% ethanol for 4 h. These results suggest that plant class I LMW HSPs may play important roles in providing both thermotolerance and ethanol tolerance.     

Heat shock response in psychrophilic and psychrotrophic yeast from Antarctica

The response to heat stress in six yeast species isolated from Antarctica was examined. The yeast were classified into two groups: one psychrophilic, with a maximum growth temperature of 20°C, and the other psychrotrophic, capable of growth at temperatures above 20°C. In addition to species-specific heat shock protein (hsp) profiles, a heat shock (15°C–25°C for 3 h) induced the synthesis of a 110-kDa protein common to the psychrophiles, Mrakia stokesii, M. frigida, and M. gelida, but not evident in Leucosporidium antarcticum. Immunoblot analyses revealed heat shock inducible proteins (hsps) corresponding to hsps 70 and 90. Interestingly, no proteins corresponding to hsps 60 and 104 were observed in any of the psychrophilic species examined. In the psychrotrophic yeast, Leucosporidium fellii and L. scottii, in addition to the presence of hsps 70 and 90, a protein corresponding to hsp 104 was observed. In psychrotrophic yeast, as observed in psychrophilic yeast, the absence of a protein corresponding to hsp 60 was noted. Relatively high endogenous levels of trehalose which were elevated upon a heat shock were exhibited by all species. A 10 Celsius degree increase in temperature above the growth temperature (15°C) of psychrophiles and psychrotrophs was optimal for heat shock induced thermotolerance. On the other hand, in psychrotrophic yeast grown at 25°C, only a 5 Celsius degree increase in temperature was necessary for heat shock induced thermotolerance. Induced thermotolerance in all yeast species was coincident with hsp synthesis and trehalose accumulation. It was concluded that psychrophilic and psychrotrophic yeast, although exhibiting a stress response similar to mesophilic Saccharomyces cerevisiae, nevertheless had distinctive stress protein profiles. Received: August 7, 1997 / Accepted: October 22, 1997