Heat stress screening of peanut (<Emphasis Type="Italic">Arachis hypogaea</Emphasis> L.) seedlings for acquired thermotolerance

Heat can be one of the major abiotic stresses that adversely affect crop production worldwide at different stages of development. As field screening for heat tolerance can be inconsistent and seasonally-limited, it is important to develop a reliable protocol under controlled conditions that allows simultaneous screening of multiple genotypes. The objective of this research was to develop a straightforward laboratory protocol using acquired thermotolerance (ATT) in peanut seedlings as a measure of one mechanism of heat stress tolerance. Sixteen genotypes, including selected accessions of the US peanut minicore collection along with standard checks, were evaluated for acquired themotolerance in two independent experiments. A change in the temperature sensitivity of chlorophyll accumulation was used as an indicator of acquired thermotolerance. Pre-incubation at 38°C for 4 h before the 30-min 50°C challenge triggered the acquired thermotolerance system of the leaf disks, resulting in chlorophyll accumulation upon exposure to light. There was considerable variation among genotypes for ATT in both experiments. Genotypic ranking for mean ATT values were highly correlated (0.949) in both experiments. The effect of seed weight on ATT was not significant. This method is relatively simple and inexpensive and can be used to screen a large number of genotypes.     

Isolation of Arabidopsis mutants lacking components of acquired thermotolerance

Acquired thermotolerance is a complex physiological phenomenon that enables plants to survive normally lethal temperatures. This study characterizes the temperature sensitivity of Arabidopsis using a chlorophyll accumulation bioassay, describes a procedure for selection of acquired thermotolerance mutants, and provides the physiological characterization of one mutant (AtTS02) isolated by this procedure. Exposure of etiolated Arabidopsis seedlings to 48 degrees C or 50 degrees C for 30 min blocks subsequent chlorophyll accumulation and is eventually lethal. Arabidopsis seedlings can be protected against the effects of a 50 degrees C, 30-min challenge by a 4-h pre-incubation at 38 degrees C. By the use of the milder challenge, 44 degrees C for 30 min, and protective pretreatment, mutants lacking components of the acquired thermotolerance system were isolated. Putative mutants isolated by this procedure exhibited chlorophyll accumulation levels (our measure of acquired thermotolerance) ranging from 10% to 98% of control seedling levels following pre-incubation at 38 degrees C and challenge at 50 degrees C. The induction temperatures for maximum acquired thermotolerance prior to a high temperature challenge were the same in AtTS02 and RLD seedlings, although the absolute level of chlorophyll accumulation was reduced in the mutant. Genetic analysis showed that the loss of acquired thermotolerance in AtTS02 was a recessive trait. The pattern of proteins synthesized at 25 degrees C and 38 degrees C in the RLD and AtTS02 revealed the reduction in the level of a 27-kD heat shock protein in AtTS02. Genetic analysis showed that the reduction of this protein level was correlated with the acquired thermotolerance phenotype.     

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°C, with either a slope (40 min) or a shock (10 s), both followed by a 1-h plateau at 50°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°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.     

Thermotolerance of IVM‐derived bovine oocytes and embryos after short‐term heat shock

A series of experiments were designed to study the effect of elevated temperatures on developmental competence of bovine oocytes and embryos produced in vitro. In experiment 1, the effect of heat shock (HS) by a mild elevated temperature (40.5°C) for 0, 30, or 60 min on the viability of in vitro matured (IVM) oocytes was tested following in vitro fertilization (IVF) and culture. No significant difference was observed between the control (39°C) and the heat‐treated groups in cleavage, blastocyst formation, or hatching (P > 0.05). In experiment 2, when the HS temperature was increased to 41.5°C, neither the cleavage rate nor blastocyst development was affected by treatment. However, the rate of blastocyst hatching appeared lower in the HS groups (13% in control group vs. 3.9% and 5.6% in 30 min and 60 min, respectively; P < 0.05). When IVM oocytes were treated at 43°C prior to IVF (experiment 3), no difference was detected in blastocyst and expanded blastocyst development following heat treatment for 0, 15, or 30 min, but heat treatment of oocytes for 45 or 60 min significantly reduced blastocyst and expanded blastocyst formation (P < 0.05). In experiment 4, the thermotolerance of day 3 and day 4 bovine IVF embryos were compared. When embryos were pre‐treated with a mild elevated temperature (40.5°C) for 1 hr, and then with a higher temperature (43°C) for 1 hr, no improvement in thermotolerance of the embryos was observed as compared to those treated at 43°C alone. However, a higher thermotolerance was observed in day 4 than day 3 embryos. In conclusion, treatment at 43°C, but not 40.5°C or 41.5°C significantly reduced oocyte developmental competence. An increase in thermotolerance was observed from day 3 to day 4 of in vitro embryonic development, which corresponds to the maternal to zygotic transition of gene expression in bovine embryos. Mol. Reprod. Dev. 53:336–340, 1999. © 1999 Wiley‐Liss, Inc.     

Thermotolerance and trehalose accumulation induced by heat shock in yeast cells of Candida albicans

Candida albicans yeast cells growing exponentially on glucose are extremely sensitive to severe heat shock treatments (52.5°C for 5 min). When these cultures were subjected to a mild temperature preincubation (42°C), they became thermotolerant and displayed higher resistance to further heat stress. The intracellular content of trehalose was very low in exponential cells, but underwent a marked increase upon non-lethal heat exposure. The accumulation of trehalose is likely due to heat-induced activation of the trehalose-6-phosphate synthase complex, whereas the external trehalase remained practically unmodified. After a temperature reversion shift (from 42°C to 28°C), the pool of trehalose was rapidly mobilized without any concomitant change in trehalase activity. These results support an important role of trehalose in the mechanism of acquired thermotolerance in C. albicans and seem to exclude the external trehalase as a key enzyme in this process.     

Thermotolerance of the photosynthetic light reactions in two <Emphasis Type="Italic">Phaseolus</Emphasis> species: a comparative study

The common bean (Phaseolus vulgaris L.) is sensitive to high temperature, while an ecologically contrasting species (Phaseolus acutifolius A. Gray) is cultivated successfully in hot environments. In this study, the two bean species were respectively acclimated to a control temperature of 25 °C and a moderately elevated temperature of 35 °C in order to compare the thermotolerance capabilities of their photosynthetic light reactions. Growth at 35 °C appeared to have no obvious adverse effect on the photosynthetic activities of the two beans, but changed their thermotolerance. After a short period of heat shock (40 °C for up to 4 h), the photosynthetic activities of 25 °C-grown P. vulgaris declined more severely than those of P. acutifolius grown at 25 °C, implying that the basal thermotolerance of P. vulgaris is inferior to that of P. acutifolius. But after acclimating to 35 °C, the thermotolerances of the two species were both greatly enhanced to about the same level, clearly demonstrating the induction of acquired thermotolerance in their chloroplasts, and P. vulgaris could be as good as P. acutifolius. Temperature acclimation also changed plants’ resistance to photoinhibition in a manner similar to those toward heat stress. In addition, acquisition of tolerance to heat and strong irradiance would reduce the dependency of the two beans on xanthophyll pigments to dissipate heat, and also seemed irrelevant to the agents with antioxidant activities such as SOD.     

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.     

The heat shock response and acquired thermotolerance in three strains of cyanobacteria

The heat shock response of three cyanobacterial strains,Anabaena sp. Strain PCC (paris Culture Collection) 7120,Plectonema boryanum Strain PCC 6306, andSynechococcus sp. Strain PCC 7942, was characterized by polyacrylamide gel electrophoresis.Anabaena produced 33 heat shock proteins,P. boryanum 35 proteins, andSynechoccus 19 proteins. The rapid response to heat shock was consistent for all three strains, although the number of time-dependent proteins varied. All strains developed thermotolerance when first pretreated with a sublethal heat shock and then challenged with a previously lethal temperature. A 30-min 30°C incubation was required between the heat shock and challenge forSynechococcus, but not forAnabaena andP. boryanum. Synechococcus cells required a higher challenge temperature (51° vs. 49°C) than the other two strains to destroy control cells that were not pretreated with a heat shock.     

Heat Shock Responses of Closely Related Species of Tropical and Desert Fish

Over the past twelve years, we have studied heat shock proteinsin two tropical species, a half dozen desert species and a numberof hemiclones of viviparous fishes in the genus Poeciliopsis.Heat shock protein (Hsp) isoform patterns were determined usinghigh resolution two-dimensional polyacrylamide gels. Two familiesof Hsps were studied in detail, the nucleocytoplasmic 70 kilodaltonHsp70 family and the 30 kilodalton Hsp30 family related to -crystallin.The temperature dependence of Hsp accumulation was investigatedusing both intact fish and cultured cells. When the thresholdtemperatures were mapped onto thermal preference profiles, itwas apparent that the Hsp70 threshold (33°C) was closelylinked to the most frequently selected temperatures and theHsp30 threshold (37°C) was closely linked to high temperaturesthat fish rarely selected, indicating that fish deploy thesetwo molecular chaperones differently. One tropical species P.gracilis is a genetic reservoir for most of the Hsp70 isoformsof the desert species. Acquired resistance to 41°C was stronglycorrelated with Hsp70 abundance for gracilis that containedHsp70 isoform 3 whereas fish lacking this isoform showed similarlevels of acquired thermotolerance which did not correlate withHsp70 abundance, suggesting multiple, compensating mechanismsof acquired resistance. Isoform 3 was degraded in cultured cellsfrom a desert species during several hours of recovery at normaltemperature following heat shock whereas two other Hsp70 isoformswere stable. The implications of this property of isoform 3are discussed.     

Heat shock increases levels of reactive oxygen species,autophagy and apoptosis

Hyperthermia is a promising anticancer treatment used in combination with radiotherapy and chemotherapy. Temperatures above 41.5 °C are cytotoxic and hyperthermia treatments can target a localized area of the body that has been invaded by a tumor. However, non-lethal temperatures (39–41 °C) can increase cellular defenses, such as heat shock proteins. This adaptive survival response, thermotolerance, can protect cells against subsequent cytotoxic stress such as anticancer treatments and heat shock (>41.5 °C). Autophagy is another survival process that is activated by stress. This study aims to determine whether autophagy can be activated by heat shock at 42 °C, and if this response is mediated by reactive oxygen species (ROS). Autophagy was increased during shorter heating times (<60 min) at 42 °C in cells. Levels of acidic vesicular organelles (AVO) and autophagy proteins Beclin-1, LC3-II/LC-3I, Atg7 and Atg12-Atg5 were increased. Heat shock at 42 °C increased levels of ROS. Increased levels of LC3 and AVOs at 42 °C were inhibited by antioxidants. Therefore, increased autophagy during heat shock at 42 °C (<60 min) was mediated by ROS. Conversely, heat shock at 42 °C for longer times (1?3 h) caused apoptosis and activation of caspases in the mitochondrial, death receptor and endoplasmic reticulum (ER) pathways. Thermotolerant cells, which were developed at 40 °C, were resistant to activation of apoptosis at 42 °C. Autophagy inhibitors 3-methyladenine and bafilomycin sensitized cells to activation of apoptosis by heat shock (42 °C). Improved understanding of autophagy in cellular responses to heat shock could be useful for optimizing the efficacy of hyperthermia in the clinic.     

Physiological implication of intracellular trehalose and mannitol changes in response of entomopathogenic fungus <Emphasis Type="Italic">Beauveria bassiana</Emphasis> to thermal stress

To explore possible role of intracellular trehalose accumulation in fungal tolerance to summer-like thermal stress, 3-day colonies of Beauveria bassiana grown on a glucose-free medium at 25°C were separately exposed to 35, 37.5 and 40°C for 1–18 h, respectively. Trehalose accumulation in stressed mycelia increased from initial 4.2 to 88.3, 74.7 and 65.5 mg g⁻¹ biomass after 6-h stress at 35, 37.5 and 40°C, respectively, while intracellular mannitol level generally declined with higher temperatures and longer stress time. The stress-enhanced trehalose level was significantly correlated to decreased trehalase activity (r ² = 0.73) and mannitol content (r ² = 0.38), which was inversely correlated to the activity of mannitol dehydrogenase (r ² = 0.41) or mannitol 1-phosphate dehydrogenase (r ² = 0.30) under the stresses. All stressed cultures were successfully recovered at 25°C but their vigor depended on stressful temperature, time length and the interaction of both (r ² = 0.98). The highest level of 6-h trehalose accumulation at 35°C was found enhancing the tolerance of the stressed cultures to the greater stress of 48°C. The results suggest that the trehalose accumulation result partially from metabolized mannitol and contribute to the fungal thermotolerance. Trehalase also contributed to the thermotolerance by hydrolyzing accumulated trehalose under the conditions of thermal stress and recovery.     

Heat shock protein induction and the acquisition of thermotolerance in the psychrotrophic yeastTrichosporon pullulans

Two-dimensional gel electrophoretic analysis of the heat shock response in the psychrotrophic yeastTrichosporon pullulans revealed the induction of 11 heat shock proteins (hsps) after a 5° to 21°C heat shock, 12 hsps after a 5° to 26°C heat shock, and 12 hsps after a 5° to 29°C heat shock. Heat shock from 5° to 26° or 29°C resulted in a statistically significant increase in thermotolerance to a lethal heat challenge at 45°C for 5 min. When the protein synthesis inhibitor, cycloheximide, was added prior to the heat shock, no statistically significant thermotolerance was acquired. To confirm the correlation between the synthesis of hsps and the acquisition of thermotolerance, protein extracts of cells that had been heat shocked in the presence or absence of cycloheximide were electrophoretically analyzed. Addition of the same concentration of cycloheximide that prevented the acquisition of thermotolerance also inhibited the synthesis of any hsps.     

Heat shock and cold shock in Deinococcus radiodurans

On the basis of acquired thermotolerance and cryotolerance, the optimal heat shock and cold shock temperatures have been determined for Deinococcus radiodurans. A heat shock at 42°C maximized survival at the lethal temperature of 52°C and a cold shock at 20°C maximized survival after repeated freeze-thawing. Enhanced survival from heat shock was found to be strongly dependent on growth stage, with its greatest effect shortly after phase. Increased synthesis of a total of 67 proteins during heat shock and 42 proteins during cold shock were observed by two-dimensional polyacrylamide gel electrophoresis (2D PAGE) and autoradiography. Eight of the most highly induced heat shock proteins shown by 2D PAGE were identified by MALDI-MS as Hsp20, GroEL, DnaK, SodA, Csp, Protease I and two proteins of unknown function.     

Acquisition of thermotolerant yeast Saccharomyces cerevisiae by breeding via stepwise adaptation

A thermotolerant Saccharomyces cerevisiae yeast strain, YK60‐1, was bred from a parental strain, MT8‐1, via stepwise adaptation. YK60‐1 grew at 40°C, a temperature at which MT8‐1 could not grow at all. YK60‐1 exhibited faster growth than MT8‐1 at 30°C. To investigate the mechanisms how MT8‐1 acquired thermotolerance, DNA microarray analysis was performed. The analysis revealed the induction of stress‐responsive genes such as those encoding heat shock proteins and trehalose biosynthetic enzymes in YK60‐1. Furthermore, nontargeting metabolome analysis showed that YK60‐1 accumulated more trehalose, a metabolite that contributes to stress tolerance in yeast, than MT8‐1. In conclusion, S. cerevisiae MT8‐1 acquired thermotolerance by induction of specific stress‐responsive genes and enhanced intracellular trehalose levels. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:1116–1123, 2013     

Blood heat shock proteins evoked by some <Emphasis Type="Italic">Salmonella</Emphasis> strains infection in ducks

Bacterial heat-shock response is a global regulatory system required for effective adaptation to changes (stress) in the environment. An in vitro study was conducted to investigate the impact of a sublethal temperature (42°C) on heat shock protein (HSP) expression in 6 Salmonella strains (Salmonella Enteritidis, S. Typhimurium, S. Virchow, S. Shubra, S. Haifa and S. Eingedi). The 6 Salmonella strains were isolated from the tissues of ducklings that had died from avian salmonellosis. To determine the induction of HSP in the 6 Salmonella strains, they were exposed to the selected temperature level for 24 h and further kept for 48 h at culturing condition of 42°C. Growth under a sublethal temperature of 42°C increased the expression of several proteins of Salmonella, including a 63 kDa protein in addition to the generation and/or overexpression of 143 proteins which were specific to heat shock, concurrent to this acquired thermotolerance. The 6 Salmonella strains responded to 24 h of thermal stress at an elevated temperature 42°C by synthesizing different heat shock proteins (HSP) with molecular weights ranging between 13.62 and 96.61 kDa. At 48 h, the 6 Salmonella strains synthesized different HSPs with molecular weights ranging between 14.53 and 103.43 kDa. It follows that salmonellae would produce HSPs during the course of the infectious process. Salmonellosis produced several proteins after 24 and 48 h of infection. Seven of these proteins (100, 80, 60, 40, 30, 20 and 10 kDa) were recognized in the serum obtained from the ducklings infected with S. Enteritidis, S. Typhimurium, S. Virchow, S. Shubra, S. Haifa and S. Eingedi after 24 h of infection. After 48 h, the 1–7 kDa HSP became more evident and indicated their de novo generation.     

Behavioural and physiological responses of juvenile geoduck (Panopea zelandica) following acute thermal stress

Climate extremes, such as heatwaves, are expected to become more intense and of longer duration in the near future. These climatic conditions may have a significant impact on the prospects of establishing a new aquaculture industry for the endemic New Zealand geoduck, Panopea zelandica. This study focused on characterising animal behaviour, haemocytes , and heat shock protein (HSP70 & HSP90) mRNA expression following exposure to elevated temperatures, such as those encountered during marine heatwaves around 20 °C and an extreme scenario of 25 °C, contrasted to an ambient temperature of 17 °C. After 24 h of heat challenge, P. zelandica were found to be significantly influenced by the thermal changes, as there were differences recorded in all the responses examined. With increasing temperatures, juvenile geoduck were observed to fully emerge from the sediment a behaviour that has not previously been quantified nor associated with stress in this species. The ability of P. zelandica juveniles to re-bury still warrants further investigation, as adults are unable to do so. Haemocyte analyses revealed an increase in the abundance of granulocytes, cellular aggregations, and size of these aggregations at the highest temperature exposure. Increased expression of the hsp70 gene in the haemolymph after exposure at 25 °C for 24 h was detected and attributed to attempts to mitigate protein denaturation caused by thermal stress. The inter-individual variability in the response of heat shock proteins recorded could aid in future selective breeding programs if it is reflected in net thermotolerance. P. zelandica shows great potential for growing in subtidal habitats around New Zealand, and this study highlights the importance of temperature considerations when selecting potential farm and reseeding locations.     

Involvement of phospholipase D in the low temperature acclimation-induced thermotolerance in grape berry

Both phospholipase D (PLD, EC 3.1.4.4) and salicylic acid (SA) play important roles in response to external stimulation and activating defense system in plants. However, roles of the two signals in plants during the development of thermotolerance induced by low temperature acclimation remain unclear. In the experiment presented in the paper, grape berries (Vitis vinifera L. cv. Chardonnay) were pretreated at 8 °C for 3 h and then transferred to 45 °C for heat stress. Compared with the control without low temperature pretreatment, membrane permeability and malondialdehyde (MDA) contents were reduced and the expression of HSP73 increased in the low temperature-pretreated berries under heat stress. During low temperature acclimation, PLD, SA and HSP73 could be activated. Meanwhile, the expression of HSP73 and the accumulation of free SA induced by low temperature can be inhibited by PLD activity inhibitor. All these results suggest that the activation of PLD is an early response to low temperature, and it is involved in the accumulation of free SA and the development of thermotolerance induced by low temperature acclimation.     

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     

Induced thermotolerance during early development of murine and bovine embryos

During early development, elevated temperatures have deleterious effects on embryonic viability and development. The primary objective of the current study was to determine the ontogeny of induced thermotolerance during early murine embryonic development. Embryos were either retrieved from superovulated ICR female mice at the 2 cell and 4 cell stages and cultured thereafter or were retrieved from oviducts or uterine horns at the desired stage of development. Induction of thermotolerance was detected by evaluating viability and further development after embryos were exposed to homeothermic temperature (37°C), mild heat shock (40°C for 1 h), severe heat shock (42°C for 1 h or 43°C for 2 h), or mild heat shock followed by severe heat shock (to induce thermotolerance). Induction of thermotolerance was observed beginning at the 8 cell stage when embryos were developed in culture from the 2 cell to 4 cell stage. When embryos were developed in vivo (i.e., were retrieved from the reproductive tract at the desired stage of development), thermotolerance was not induced until the blastocyst stage of development. The induction of thermotolerance was dependent on serum supplementation since induction of thermotolerance was not observed when embryos were placed in medium without serum. Induced thermotolerance could also be demonstrated in bovine blastocysts. In conclusion, embryos acquire the ability to undergo thermotolerance as they progress through development. The timing of processes leading to acquisition of thermotolerance can, however, be hastened by exposure of embryos to in vitro conditions.     

Simultaneous exposure of Xenopus A6 kidney epithelial cells to concurrent mild sodium arsenite and heat stress results in enhanced hsp30 and hsp70 gene expression and the acquisition of thermotolerance

In this study, we examined the effect of concurrent low concentrations of sodium arsenite and mild heat shock temperatures on hsp30 and hsp70 gene expression in Xenopus A6 kidney epithelial cells. RNA blot hybridization and immunoblot analysis revealed that exposure of A6 cells to 1–10 µM sodium arsenite at a mild heat shock temperature of 30 °C enhanced hsp30 and hsp70 gene expression to a much greater extent than found with either stress individually. In cells treated simultaneously with 10 µM sodium arsenite and different heat shock temperatures, enhanced accumulation of HSP30 and HSP70 protein was first detected at 26 °C with larger responses at 28 and 30 °C. HSF1 activity was involved in combined stress-induced hsp gene expression since the HSF1 activation inhibitor, KNK437, inhibited HSP30 and HSP70 accumulation. Immunocytochemical analysis revealed that HSP30 was present in a granular pattern primarily in the cytoplasm in cells treated simultaneously with both stresses. Finally, prior exposure of A6 cells to concurrent sodium arsenite (10 µM) and heat shock (30 °C) treatment conferred thermotolerance since it protected them against a subsequent thermal challenge (37 °C). Acquired thermotolerance was not observed with cells treated with the two mild stresses individually.