Degree Of Thermotolerance Research Articles

Dissecting the heat stress-induced alterations in the leaf ultrastructure and some antioxidant network components in interspecific (Cucurbita maxima × Cucurbita moschata) inbred line of squash ‘Maxchata’ as to its parents possessing variable heat tolerance

We previously found ‘C. moschata’ to be heat-tolerant’, ‘C. maxima’ to be heat-susceptible, and their interspecific (Cucurbita maxima × Cucurbita moschata) inbred line ‘Maxchata’ to exhibit intermediary thermotolerance through its photosynthetic response. In this work, we investigated the ultramorphological, biochemical, and transcriptional changes induced in the leaves of ‘Maxchata’ and its parents upon exposure to moderate (37 °C) and severe (42 °C) heat stress to obtain insights into heat-tolerance indicators. The results showed that heat stress caused a relatively lower degree of membrane damage with lower malondialdehyde level and electrolyte leakage and higher proline contents in ‘C. moschata’ and ‘Maxchata’. The electron microscopy highlighted the maximum degradation of the leaf ultrastructure of ‘C. maxima’ upon heat exposure. In contrast, ‘C. moschata’ and ‘Maxchata’ exhibited lower degree of subcellular injury. The enzymatic activities of superoxide dismutase (SOD), catalase (CAT), ascorbate peroxidase (APX), and guaiacol peroxidase (POD) were found to be highest in ‘C. moschata’, moderate in ‘Maxchata’, and lowest in ‘C. maxima’. In addition, the gene expressions of the CAT (CAT1, CAT2, and CAT3), SOD (Cu/ZnSOD, FeSOD, and MnSOD), and APX (APX1 and APX2) isozymes were unique in each of the studied squashes with increase in temperature indicating their differential contribution to heat tolerance. Although the selected genes were genetically similar in ‘C. maxima’ and ‘Maxchata’, most of the genes were more highly induced at elevated temperatures in ‘Maxchata’ compared with ‘C. maxima’. Hence, the heat stress-induced gene expression in ‘Maxchata’ improved the efficiency of its antioxidant network and provided it some degree of thermotolerance.

Identification of a gene, FMP21, whose expression levels are involved in thermotolerance in Saccharomyces cerevisiae.

Elucidation of the mechanism of high temperature tolerance in yeasts is important for the molecular breeding of high temperature-tolerant yeasts that can be used in bioethanol production. We identified genes whose expression is correlated with the degree of thermotolerance in Saccharomyces cerevisiae by DNA microarray analysis. Gene expression profiles of three S. cerevisiae strains showing different levels of thermotolerance were compared, and we chose three of them as candidate genes. Among these genes, FMP21 was investigated as a thermotolerance-related gene in S. cerevisiae by comparing the growth at high temperature with the gene expression in eight strains. The expression ratio of FMP21 at 37°C was correlated with the doubling time ratio at a coefficient of determination of 0.787. The potential involvement of the Fmp21 in the thermotolerance of yeasts was evaluated. The FMP21 deletion variant showed a decreased respiratory growth rate and increased thermosensitivity. Furthermore, the overexpression of FMP21 improved thermotolerance in yeasts. In conclusion, the function of Fmp21 is important for thermotolerance in yeasts.

Improvement of protein stability and enzyme recovery under stress conditions by using a small HSP (tpv-HSP 14.3) from Thermoplasma volcanium

In this study we cloned and expressed a small heat shock protein, tpv-HSP 14.3, from thermoacidophilic archaeon Thermoplasma volcanium. This novel recombinant small heat shock protein was purified to homogeneity and produced a protein band of 14.3kDa on SDS-polyacrylamide gel. Transmission electron microscopy images of the negatively stained tpv-HSP 14.3 samples showed spherical particles of 13nm diameter. E. coli cells over expressing tpv-HSP 14.3 endowed the cells with some degree of thermotolerance. After exposure to 52°C for 120min, survivability of the E. coli cells expressing tpv-HSP 14.3 was approximately 2.5-fold higher than the control cells. As a molecular chaperone tpv-HSP 14.3 enhanced the thermal stabilization of substrate proteins, pig heart citrate synthase and bovine l-glutamic dehdyrogenase, considerably. The highest protection effect of tpv-HSP 14.3 was observed at 47°C for pig heart citrate synthase; the remaining activity was 5-fold higher than that of the sample without tpv-HSP 14.3. The tpv-sHSP 14.3 prevented inactivation of bovine l-glutamic dehdyrogenase the most effectively at 53°C; the residual activity was approximately 2-fold higher than that of the sample heated without tpv-HSP 14.3. However, refolding activity of the tpv-HSP 14.3 was relatively weak for the chemically denatured substrate proteins.

Physiological responses of Escherichia coli exposed to different heat-stress kinetics

The effects of heat-stress kinetics on the viability of Escherichia coli were investigated. Cells were exposed to heat-stress treatments extending from 30 to 50 degrees C, with either a slope (40 min) or a shock (10 s), both followed by a 1-h plateau at 50 degrees C in nutritive medium. A higher survival rate was observed after the slope than after the shock, when both were followed by a plateau, so the heat slope induced a certain degree of thermotolerance. This tolerance was partly (i) linked to de novo protein synthesis during the subsequent plateau phase, and (ii) abolished after rapid cooling from 50 to 30 degrees C, which means that cellular components with rapidly reversible thermal properties are involved in this type of thermotolerance. The heat-slope-induced thermotolerance was chiefly linked to the maintenance of the plasma membrane integrity (preservation of structure, fluidity, and permeability), and not to GroEL or DnaK overexpression. Moreover, the high level of cell mortality induced by the heat shock could be related to changes in the membrane integrity.

Effects of hyperthermia on the central nervous system: What was learnt from animal studies?

Animal studies show that nervous tissue is sensitive to heat. Although inter-species variations may play a role, the data indicate that the maximum heat dose without obvious complications after localized hyperthermia in regions of the central nervous system (CNS) lies in the range of 40–60 min at 42–42.5°C or 10–30 min at 43°C. Expression of thermotolerance after a ‘conditioning’ heat dose was clearly observed in the spinal cord of rodents and the thermotolerance ratio's (ratio between heat doses with and without conditioning required to obtain a certain defined effect) were high, ∼2. The thermotolerant state of CNS is shown to protect also against other types of injury as well: pre-treatment of rats with hyperthermia protected against spinal cord ischemic injury. During the rather long period required for temperature elevation which is inherent to WBH, some degree of thermotolerance may develop. The correlation between thermotolerance and hsp70 induction in CNS is obvious. Heat, at least if applied shortly after X-rays, enhances the response of nervous tissue to radiation. Data on the combined effects of X-ray irradiation and hyperthermia on rodent spinal cord clearly show that the radiation response can be enhanced with a factor of 1.1–1.3. There are no clear experimental data indicating an increase in adverse effects specific to the CNS after localized or whole body hyperthermia as a result of combined treatment with chemotherapy.

벼 엽록체 small HSP의 과발현에 의한 형질전환 식물체의 내열성 증가

엽록체 small HSP의 기능을 조사하기 위하여 벼로부터 분리한 엽록체 small HSP를 구성적으로 발현하는 형질전환 식물체를 제작하였다. 먼저 고온 스트레스 조건하에서의 형질전환 식물체의 내열성을 chlorophyll 형광으로 측정하여 분석하였다. Leaf disc를 고온 스트레스 조건에서 5분간 처리한 후, 광화학계 II의 불활성화를 나타내는 Fo 값과 증가치를 조사하였다. 형질전환 식물체는 고온 스트레스 하에서의 Fo 값의 증가가 현저하게 감소하였다. 또한 무균적으로 Petri dish에서 재배한 유식물체를 치사온도인 <TEX>$52^{\circ}C$</TEX>에서 45분간 처리한 후, <TEX>$20^{\circ}C$</TEX> 에서 계속적으로 배양하였을 때, wild-type 식물체는 전부 고사하였으나, 형질전환 식물체의 약 80%는 정상적으로 생존하였다. 또한 과발현된 Oshsp26 단백질의 축적량이 많을수록 내열성의 정도도 증가하였다. 이러한 결과는 엽록체 small HSP가 고온 스트레스 하에서 광합성기구를 보호함으로서 식물체의 내열성을 증가시키는데 있어서 중요한 기능을 담당하고 있음을 나타낸다. To investigate the function of chloroplast small heat shock protein (HSP), transgenic tobacco plants (Nicotiana tabacum L, cv. SR-1) that constitutively overexpress the rice chloroplast small HSP (Oshsp26) were generated. Effects of constitutive expression of the Oshsp26 on thermotolerance were investigated with the chlorophyll fluorescence. After 5-min incubation of leaf discs at high temperatures, an increase in the Fo level, indication of separation of LHCII from PSII, was mitigated by constitutive expression of the chloroplast small HSP When tobacco plantlets grown in Petri dishes were incubated at <TEX>$20^{\circ}C$/TEX> for 45 min and subsequently incubated at <TEX>$20^{\circ}C$/TEX> leaf color of wild-type plant became gradually white and all plantlets were finally died. Under the conditions in which all the wild-type plants died, more than 80% of the transformants remained green and survived. It was also found that the levels of Oshsp26 protein accumulated in transgenic plants were correlated with the degree of thermotolerance. These results suggest that the chloroplast small HSP plays an important role in protecting photosynthetic machinery, as a results, increases thermotolerance of whole plant during heat stress.

Amino acid residue 184 of yeast Hsp104 chaperone is critical for prion-curing by guanidine, prion propagation, and thermotolerance.

Inactivation of Hsp104 by guanidine is contended to be the mechanism by which guanidine cures yeast prions. We now find an Hsp104 mutation (D184N) that confers resistance to guanidine-curing of the yeast [PSI(+)] prion. In an independent screen we isolated an HSP104 allele altered in the same residue (D184Y) that dramatically impairs [PSI(+)] propagation in a temperature-dependent manner. Directed mutagenesis of HSP104 produced additional alleles that conferred varying degrees of resistance to guanidine-curing or impaired [PSI(+)] propagation. The mutations similarly affected propagation of the [URE3] prion. Basal and induced abundance of all mutant proteins was normal. Thermotolerance of cells expressing mutant proteins was variably resistant to guanidine, and the degree of thermotolerance did not correlate with [PSI(+)] stability. We thus show that guanidine cures yeast prions by inactivating Hsp104 and identify a highly conserved Hsp104 residue that is critical for yeast prion propagation. Our data suggest that Hsp104 activity can be reduced substantially without affecting [PSI(+)] stability, and that Hsp104 interacts differently with prion aggregates than with aggregates of thermally denatured protein.

Phenotypic characterization of Listeria monocytogenes and Listeria innocua strains isolated from short-ripened cheeses

Listeria monocytogenes and Listeria innocua strains isolated from short-ripened cheeses and classified, respectively, into five (m1 to m5) and two (i1 and i2) REDP clusters (restriction enzyme digestion profiles by PFGE), were characterized on the basis of their carbohydrate fermentation profiles, growth in milk, haemolytic activity, sensitivity to sanitizing agents, tolerance to low pH, organic acids and NaCl, and resistance to pasteurization. All isolates fermented L -rhamnose and α-methyl- D -mannoside and most of them were lactose positive; melezitose was fermented by L. monocytogenes strains but not by L. innocua. The haemolytic activity of L. monocytogenes varied depending on the strain. The isolates were sensitive to all cleaning and disinfectant agents tested (acid, alkaline, chlorine and peroxide-containing agents), with the only exception of NaOH. Storage at refrigeration temperature enhanced the tolerance to NaCl of both species and the resistance of L. monocytogenes to the acid. Organic acids were more inhibitory than HCl and their effectiveness was greater at higher than at lower concentration and followed the order acetic>lactic>citric. M4, a group genetically related to certain pathogenic L. monocytogenes strains, was unable to ferment lactose although its growth in milk was not affected. The slightly reduced tolerance to NaCl of this group with respect to the others might have prevented a wider distribution in dairy environments. Although pasteurization was effective in most cases, two different L. monocytogenes strains isolated from the same cheese sample displayed a certain degree of thermotolerance; this resistance could have been induced by successive heat treatments of strains contaminating the dairy environment. All this data pointed to a relationship between several phenotypic characteristics and the ability of Listeria to colonize certain environments.

Characterization of the heat shock protein response of Atlantic salmon (Salmo salar)

The heat shock protein (hsp) response of juvenile Atlantic salmon, Salmo salar was investigated in isolated tissues subjected to various degrees of thermal shock. Distinct but overlapping arrays of proteins from the major hsp families (hsps 100, 90, 70, 60 and small hsps) were induced in branchial lamellae, hepatic tissue and erythrocytes. The two most prominent proteins induced by heat shock (MW ≅ 65 and 66 kDa) were found to be antigenic homologues of mammalian hsps72/73. A 2.6 kb transcript upregulated by the same conditions hybridized with cDNA probes to both human and salmon hsp70. Branchial lamellae exhibited the greatest degree of thermotolerance and mounted the most significant heat shock response. Moderate thermal shock induced more species of proteins in branchial lamellae than in hepatic tissue or erythrocytes, with the rate of hsp65/66 synthesis increased by as much as five fold. Thermal shock induced hsp65/66 eight fold in erythrocytes. In contrast, hepatic tissue which was least tolerant of thermal shock, lacked the inducible hsp65 and exhibited minimal induction of hsp66. Persistence of hsps was tested in erythrocytes, where elevated levels remained in the cells for at least 48 h after heat shock. The temporal pattern and magnitude of the hsp response in the stenothermal Atlantic salmon differed from that previously reported for eurythermal species. Also notable was the limited hsp response mounted by salmon tissues exposed to sodium arsenite, a known inducer of hsps. The characteristics of the hsp response to thermal shock support the significance of these proteins in adaptation of Atlantic salmon to environmental insult.

Seed Developmental Temperature Regulation of Thermotolerance in Lettuce

Lettuce (Lactuca sativa L.) seeds can fail to germinate at temperatures above 24 °C. The degree of thermotolerance is thought to be at least partly related to the environment under which the seed developed. In order to study the effects of temperature during seed development on subsequent germination, various lettuce genotypes were screened for their ability to germinate at temperatures ranging from 20 to 38 °C. Seeds of the selected genotypes `Dark Green Boston' and `Valmaine' (thermosensitive), `Floricos 83', `Everglades', and PI 251245 (thermotolerant) were produced at 20/10, 25/15, 30/20, and 35/25 °C day/night temperature regimes in plant growth chambers. Seeds were germinated on a thermogradient bar from 24 to 36 °C under 12 h light/dark cycles. As germination temperature increased, the number of seeds that failed to germinate increased. Above 27 °C, seeds matured at 20/10 or 25/15 °C exhibited a lower percent germination than seeds that matured at 30/20 or 35/25 °C. Seeds of `Dark Green Boston' and `Everglades' that matured at 30/20 °C exhibited improved thermotolerance over those that matured at lower temperatures. Seeds of `Valmaine' produced at 20/10 °C exhibited 40% germination at 30 °C, but seeds that matured at higher temperatures exhibited over 95% germination. Germination of `Valmaine' at temperatures above 30 °C was not affected by seed maturation temperature. The upper temperature limit for germination of lettuce seed could thus be modified by manipulating the temperature during seed production. The potential thermotolerance of seed thereby increased, wherein thermosensitive genotypes became thermotolerant and thermotolerant genotypes (e.g., PI251245) germinated fully at 36 °C. This information is useful for improving lettuce seed germination during periods of high soil temperature, and can be used to study the biology of thermotolerance in lettuce.

Influence of tonicity and chloramphenicol on hyperthermic cytotoxicity and cell permeability under various heating rates

The cell lethality and permeability induced in Escherichia coli E/r, Escherichia coli Bs-1 and Zygosaccharornyces bailii cells by high temperature (52°C) after heating at different rates (mean s 0.015, 0.25 and 150°C per s) and in media of different tonicity and content (isotonic YEP broth versus 0.01 M phosphate buffer, pH7.0 containing different concentrations of NaCl) and with versus without chloramphenicol (10pg/ml) have been investigated. Hyperthermic treatment in YEP broth or isotonic 0.01 M phosphate buffer resulted in markedly reduced cytotoxicity with decreasing heat rate. The heating rate effect was larger when the cells were treated in YEP broth. Chloramphenicol, which is known to inhibit expression of heat shock proteins in bacteria, did not affect the viability of cells or the development of thermotolerance in cells heated at different heating rates in isotonic phosphate buffer but prevented the development of an additional degree of thermotolerance in cells heated slowly in YEP broth. In contrast, the differential effect of heating rate on cytotoxicity and cell permeability was not demonstrated when cells were heated in hypertonic solution (1 M NaCl in 0.01 M phosphate buffer, pH 7.0). It is proposed that heat destabilization of the osmotic cell homeostasis, which is more profound after rapid heating, plays a major part in heat induced cellular lethality.

Hsp40, a possible indicator for thermotolerance of murine tumour in vivo

The relationship between Hsp40/Hsp70 synthesis and the development of thermotolerance was investigated using mouse squamous cell carcinoma in vivo. To examine the thermotolerance, tumours were heated at 44°C for 30 min as conditioning heating. After various intervals they were heated again at 44°C for 90 min as challenge heating. The tumour response to heat was evaluated by the growth delay. Thermotolerance rapidly developed with increasing interval and reached a maximum at 12 h interval. Subsequently, thermotolerance gradually decayed and almost disappeared at 120 h interval. Under this condition, synthesis of Hsp40/Hsp70 increased after conditioning heating, reached a maximum at 12 h interval, then gradually decreased thereafter within 120 h. The kinetics of accumulation and decay of both Hsp40 and Hsp70 were very similar. The extent of thermotolerance was well correlated with the relative amount of Hsp40/Hsp70. These results obtained in vivo were very similar to those in vitro (Kaneko et al. 1995). Our findings suggest that Hsp40 could be a useful indicator of the degree of thermotolerance in addition to Hsp70 in vivo as in vitro.

Induction of Heat-Shock Proteins and Accumulation of Trehalose by TPN in Saccharomyces cerevisiae

TPN [Tetrachloroisophthalonitrile], a kind of disinfectant, affected growth in Saccharomyces cervisiae. Exposed to TPN under no lethal conditions, 70-, 90-kDa protein and hsp104 were induced. Each of them was not uniformly induced; namely, the 70-kDa protein was more sensitive to TPN among other proteins. Trehalose was also accumulated depending on the concentration. The degree of thermotolerance in yeast cells pretreated with 1.0 mg/l TPN was about 100-fold greater than that in the control. Under this condition, TPN-inducible proteins were synthesized but trehalose was not accumulated. Although TPN-inducible proteins and trehalose were definitely synthesized with 10.0 or 100.0mg/l TPN treatment, thermotolerance was not acquired.

Heat shock protein expression in testis and bladder cancer cell lines exhibiting differential sensitivity to heat

Testis cancer cells are more sensitive than bladder and most other cancer cells to chemotherapeutic drugs both in the clinic and in vitro. In this study we show that they are also more sensitive than bladder cancer cells to heat. Since heat and drug sensitivity may be related to the ability of a cell to mount a stress response, constitutive and induced levels of heat shock proteins (HSPs) in three testis and three bladder human cancer cell lines were measured using Western blotting and scanning densitometry. No correlation between constitutive levels of HSP 90 or HSP 73/72 and cellular heat sensitivity was found. However, HSP 27 levels were much lower in the testis tumour cells, suggesting that low HSP 27 expression might contribute to heat sensitivity. Protein synthesis studies using [35S]methionine indicated that, for the same heat shocks, the kinetics of synthesis and decay of HSP 90 and HSP 73/72 in 833K (the most heat sensitive testis cells) was similar to or greater than that in HT1376 (the most heat-resistant bladder cells). Both 833K and HT1376 developed thermotolerance, and this followed an increase in synthesis of HSPs. These results indicate that, although there are differences in the constitutive levels of HSPs between testis and bladder cancer cells, both cell types are capable of mounting an induced heat shock response and can develop a similar degree of thermotolerance.

Heat and cold shock responses in different strains of Thiobacillus ferrooxidans

Different strains of Thiobacillus ferrooxidans were examined for their ability to produce a heat shock and a cold shock response. Strain A1, heat shocked from 20° to 35°C, acquired thermotolerance, as it showed a 1000-fold reduction in cell mortality when exposed to the supermaximum temperature of 42°C, as compared to a non-heat-shocked control. A heat shock from 25° to 35°C yielded similar results, although a higher degree of thermotolerance was achieved for the shorter exposure times. Cultures heat shocked for 5 h showed a five-log reduction in viable counts after 41 h at 42°C, whereas non-heat-shocked cultures showed a similar reduction in viability in 28 h. Conferred thermotolerance was immediate and sustained for the duration of the exposure to 42°C. Heat-shocked cultures were not significantly protected against loss of viability due to freezing (-15°C for 24 h). Strain S2, cold shocked from 25° to 10°C, and strain D6, cold shocked from 25° to 5°C, were not protected against freezing at-15°C. An analysis of proteins extracted from heat-shocked cells of strain A1 showed the presence of at least one newly induced protein and eight hyper-induced proteins. The molecular weights of the heat shock proteins were in the range of 15–80.3 kDa.

Two-step Heat Treatment of Pineapple Crowns Increases Thermotolerance

Thermotolerance of pineapple crowns (`Champaka 153') to 50C and above was increased with a 30-min first treatment at 30, 35, or 40C. Pineapple crowns receiving a 30-min heat treatment, before a second heat treatment at 50 or 55C, exhibited significantly less leaf damage than controls receiving no first treatments (P ≤ 0.05). The degree of thermotolerance was dependent upon the season in which crowns were harvested; greater thermotolerance occurred in crowns harvested in April than those harvested in October. Maximum thermotolerance occurred after an interval of at least 8 h between the first treatment and the higher temperature heat treatment. Thermotolerance was stable for at least 24 h.

Effects of step-down and step-up heating on the development of thermotolerance in a C3H mammary carcinoma in vivo.

The effects of step-down (SDH) and step-up heating (SUH) on the development of thermotolerance were investigated in a C3H mammary carcinoma in vivo. The endpoint was tumour growth time, i.e. the time for a tumour to reach a volume five times that of the first treatment day. SDH consisted of 44.5 degrees C/5 min followed immediately by 41.0 degrees C/120 min. SUH consisted of the same heat treatments but in reverse sequence. Thermotolerance was detected by subsequent heating at 43.5 degrees C at variable intervals following the primary SDH or SUH. The degree of thermotolerance was quantified by the thermotolerance ratio (TTR) calculated as a ratio between the slope of the dose-response curve for tumours heated at 43.5 degrees C and tumours preheated with either SDH or SUH followed by 43.5 degrees C. Both SDH and SUH induced thermotolerance. However, the maximal degree of thermotolerance and the time interval to reach maximum thermotolerance were different. For SUH maximal thermotolerance was observed at 8 h with a TTR of 3.6. For SUH, thermotolerance peaked at 24-28 h with a TTR of 7.3. In both cases thermotolerance had decayed with a 120 h interval. The SDH priming induced about 2.5 times more heat damage than SUH. The results are therefore in agreement with previous data obtained in the same tumour model by single heating showing that both the degree and the time to reach maximal thermotolerance increases with pretreatment heat damage. In addition, the results indicate that thermotolerance and thermosensitization are independent phenomena.

A Generalized Concept for Cell Killing by Heat: Effect of Acutely Induced Thermotolerance and Decay of Thermosensitization

Acute thermotolerance was induced in Chinese hamster ovary cells by heating at 43 degrees C for 45 min followed by incubation at 37 degrees C for up to 85 h. The degree of thermotolerance induced was quantified by a second heat exposure at 43 degrees C. To study the interaction of thermotolerance and thermosensitization, thermotolerant cells (43 degrees C/45 min-->37 degrees C/10 h) were subjected to step-down heating consisting of a priming heat treatment at 43 degrees C for 45 to 300 min followed immediately by a graded test treatment at 40 degrees C. Decay of thermosensitization was assayed by separating the priming treatment at 43 degrees C for 45 or 60 min and the test treatment at 40 degrees C by incubation at 37 degrees C for up to 24 h. The experimental data were analyzed by using a mathematical model published previously (H. Jung, Radiat. Res. 106, 56-72, 1986). According to this model, cell killing by heat is a two-step process. In a first step, heating produces nonlethal lesions at a rate p which, in a second step, are converted into lethal events at a rate c. Both rates were shown to depend on temperature. Data analysis, which was performed under the assumption that the rate constant c(T) is not altered by acute thermotolerance, showed that the rate constant p(43 degrees C) decreased exponentially (half-time 1.08 +/- 0.15 h) with increasing thermotolerance development, i.e., with increasing duration of the incubation interval at 37 degrees C. The rate p(43 degrees C) approached a minimum value by 14 +/- 3 h, where it was lower by a factor of 30 compared to normally sensitive cells. For incubation times at 37 degrees C exceeding 14 h, p(43 degrees C) increased exponentially with a doubling time of 13.9 +/- 0.9 h and approached the control level by 85 h. Decay of thermosensitization was quantified by calculating the number of nonlethal lesions from the data recorded for the sequences 43 degrees C-->37 degrees C-->43 degrees C and 43 degrees C-->37 degrees C-->40 degrees C. With increasing incubation interval at 37 degrees C, the number of nonlethal lesions induced per cell by a priming heat shock at 43 degrees C for 45 min remained constant for about 1.5 h and then decreased exponentially with a half-time of 2.0 +/- 0.5 h. Thus the two processes that occur simultaneously after a heat shock at 43 degrees C for 45 min, i.e., development of acute thermotolerance and decay of thermosensitization, proceed with different kinetics as could be separated by applying the model.

Heat induced protein denaturation in the particulate fraction of HeLa S3 cells: effect of thermotolerance.

In this study we investigated the effect of heat on the proteins of the particulate fraction (PF) of HeLa S3 cells using electron spin resonance (ESR) and thermal gel analysis (TGA). ESR detects overall conformational changes in proteins, while TGA detects denaturation (aggregation due to formation of disulfide bonds) in specific proteins. For ESR measurements the -SH groups of the proteins were labelled with a maleimido bound spin label (4-maleimido-tempo). The sample was heated inside the ESR spectrometer at a rate of 1 degree C/min. ESR spectra were made every 2-3 degrees C between 20 degrees C and 70 degrees C. In the PF of untreated cells conformational changes in proteins were observed in three temperature stretches: between 38 and 44 degrees C (transition A, TA); between 47 and 53 degrees C (transition B, TB); and above 58 degrees C (transition C, TC). With TGA, using the same heating rate, we identified three proteins (55, 70, and 90 kD) which denatured during TB. No protein denaturation was observed during TA, while during TC denaturation of all remaining proteins in the PF occurred. When the ESR and TGA measurements were done with the PF of (heat-induced) thermotolerant cells, TA was unchanged while TB and TC started at higher temperatures. The temperature shift for the onset of these transitions correlated with the degree of thermotolerance that was induced in the cells. These results suggest that protection against heat-induced denaturation of proteins in the PF is involved in heat induced thermotolerance.

Physiological and molecular studies of heat and drought tolerance in barley

SummaryComparison of three barley genotypes, two parentale, Onice and Georgie, and their F1 offspring, were made for protein synthesis modification upon exposure to high temperature. The two parentals showed different degree of thermotolerance and different sets of hsp's (heat shock proteins) induced. The F1 progeny showed high similarity with the winter type Onice. The relation between hsp's pattern and different stages of development was assessed by analysis of in Wira translation products of hs (heat shock) mRNAs from shoots and leaves of “Georgie”. The conclusions are that: 1) there is genotypic specificity in hsp induction that should be used to study inheritance of hsp's in the progeny; 2) there is a developmentally related regulation of hs genes.