The effects of population and thermal acclimation on the growth,condition and cold responsive mRNA expression of age-0 lake sturgeon (Acipenser fulvescens)

In Manitoba, Canada, wild lake sturgeon (Acipenser fulvescens) populations exist along a latitudinal gradient and are reared in hatcheries to bolster threatened populations. We reared two populations of lake sturgeon, one from each of the northern and southern ends of Manitoba and examined the effects of typical hatchery temperatures (16°C) as well as 60-day acclimation to elevated rearing temperatures (20°C) on mortality, growth and condition throughout early development. Additionally, we examined the cold shock response, which may be induced during stocking, through the hepatic mRNA expression of genes involved in the response to cold stress and homeoviscous adaptation (HSP70, HSP90a, HSP90b, CIRP and SCD). Sturgeon were sampled after 1 day and 1 week following stocking into temperatures of 8, 6 and 4°C in a controlled laboratory environment. The southern population showed lower condition and higher mortality during early life than the northern population while increased rearing temperature impacted the growth and condition of developing northern sturgeon. During the cold shock, HSP70 and HSP90a mRNA expression increased in all sturgeon treatments as stocking temperature decreased, with higher expression observed in the southern population. Expression of HSP90b, CIRP and SCD increased as stocking temperature decreased in northern sturgeon with early acclimation to 20°C. Correlation analyses indicated the strongest molecular relationships were in the expression of HSP90b, CIRP and SCD, across all treatments, with a correlation between HSP90b and body condition in northern sturgeon with early acclimation to 20°C. Together, these observations highlight the importance of population and rearing environment throughout early development and on later cellular responses induced by cold stocking temperatures.     

Isolation of four heat shock protein cDNAs from grapefruit peel tissue and characterization of their expression in response to heat and chilling temperature stresses

In order to continuously supply horticultural products for long periods, it is essential to store them after harvest in low temperatures. However, many tropical and subtropical fruits and vegetables, such as citrus, are sensitive to chilling. In previous studies, the authors have shown that a short hot water rinsing treatment (at 62°C for 20 s) increased chilling tolerance in grapefruit. In order to gain more insight into the molecular mechanisms involved in heat‐induced chilling tolerance, PCR cDNA subtraction analysis was performed which isolated four different PCR fragments whose expression was enhanced 24 h after the heat treatment, and that showed high sequence homology with various plant HSP18‐I, HSP18‐II, HSP22 and HSP70 genes. It was found that the short hot water treatment given at 62°C for 20 s, but not at lower temperatures of 20 or 53°C, increased the expression of the various HSP cDNAs in grapefruit peel tissue. However, when the fruits were kept at ambient temperatures, the increases in HSP mRNA levels following the hot water treatment were temporary and lasted only between 6 and 48 h. Similar temporary increases in the HSP mRNA levels were detected following exposure of the fruit to a hot air treatment at 40°C for 2 h. Nevertheless, when the fruits were treated with hot water but afterwards stored at chilling temperatures of 2°C, the mRNA levels of the various HSP18‐I, HSP18‐II, HSP22 and HSP70 cDNAs increased and remained high and stable during the entire 8‐week cold‐storage period, suggesting their possible involvement in heat‐induced chilling‐tolerance responses. The chilling treatment by itself increased the expression of the HSP18‐I cDNA, but had no effect on the mRNA levels of any of the other HSP cDNAs. Exposure of fruit to other stresses, such as wounding, UV irradiation, anaerobic conditions and exposure to ethylene, had no effect on the expression of the various HSPs. Overall, the study explored the correlation between the expression and persistence of various HSP cDNAs in grapefruit peel tissue during cold storage, on the one hand, and the acquisition of chilling tolerance, on the other hand, and the results suggest that HSPs may play a general role in protecting plant cells under both high‐ and low‐temperature stresses.     

<Emphasis Type="Italic">Arabidopsis</Emphasis> immunophilins ROF1 (AtFKBP62) and ROF2 (AtFKBP65) exhibit tissue specificity,are heat-stress induced,and bind HSP90

The plant co-chaperones FK506-binding proteins (FKBPs) are peptidyl prolyl cis-trans isomerases that function in protein folding, signal transduction and chaperone activity. We report the characterization of the Arabidopsis large FKBPs ROF1 (AtFKBP62) and ROF2 (AtFKBP65) expression and protein accumulation patterns. Transgenic plants expressing ROF1 promoter fused to GUS reporter gene reveal that ROF1 expression is organ specific. High expression was observed in the vascular elements of roots, in hydathodes and trichomes of leaves and in stigma, sepals, and anthers. The tissue specificity and temporal expression of ROF1 and ROF2 show that they are developmentally regulated. Although ROF1 and ROF2 share 85% identity, their expression in response to heat stress is differentially regulated. Both genes are induced in plants exposed to 37 °C, but only ROF2 is a bonafide heat-stress protein, undetected when plants are grown at 22 °C. ROF1/ROF2 proteins accumulate at 37 °C, remain stable for at least 4 h upon recovery at 22 °C, whereas, their mRNA level is reduced after 1 h at 22 °C. By protein interaction assays, it was demonstrated, that ROF1 is a novel partner of HSP90. The five amino acids identified as essential for recognition and interaction between the mammalian chaperones and HSP90 are conserved in the plant ROF1-HSP90. We suggest that ROF/HSP90 complexes assemble in vivo. We propose that specific complexes formation between an HSP90 and ROF isoforms depends on their spatial and temporal expression. Such complexes might be regulated by environmental conditions such as heat stress or internal cues such as different hormones. Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users.     

Regulation of heat shock proteins in the apple maggot Rhagoletis pomonella during hot summer days and overwintering diapause

Abstract Developing larvae of the apple maggot Rhagoletis pomonella are frequently exposed to summertime apple temperatures that exceed 40 °C and, during their overwintering diapause, pupae are exposed to sub‐zero soil temperatures for prolonged periods. To investigate the potential involvement of heat shock proteins (Hsps) in response to these environmental extremes, the genes encoding Hsp70 and Hsp90 in R. pomonella are cloned and expression monitored during larval feeding within the apple and during overwintering pupal diapause. Larvae reared in the laboratory at constant temperatures of 25, 28 or 35 °C express Hsp90 but very little Hsp70. Larvae do not survive rearing at 40 °C. The temperature cycles to which larvae were exposed inside apples in the field, ranging 16–46.9 °C over a 24‐h period, elicit strong Hsp70 and Hsp90 expression, which begins at mid‐day and reaches a peak in late afternoon, coinciding with peak air and apple temperatures. Heat shock proteins are also expressed strongly by pupae during their overwintering diapause. Hsp70 is not expressed in nondiapausing pupae but is highly expressed throughout diapause. Hsp90 is constitutively expressed in both diapausing and nondiapausing pupae. Rhagoletis pomonella thus strongly expresses its Hsps during pupal diapause, presumably as a protection against low temperature injury, and during larval development to cope with natural temperature cycles prevailing in late summer.     

The time-profile of the PBMC HSP70 response to in vitro heat shock appears temperature-dependent

Summary. Heat shock proteins (HSPs) are synthesised by cells subsequent to a stress exposure and are known to confer protection to the cell in response to a second challenge. HSP induction and decay are correlated to thermotolerance and may therefore be used as a biomarker of thermal history. The current study tested the temperature-dependent nature of the heat shock response and characterised its time profile of induction. Whole blood from 6 healthy males (Age: 26 ± (SD) 2 yrs; Body mass 74.2 ± 3.8 kgs; VO_2max: 49.1 ± 4.0 ml·kg⁻¹·min⁻¹) were isolated and exposed to in vitro heat shock (HS) at 37, 38, 39, 40, and 41 °C for a period of 90 min. After HS the temperature was returned to 37 °C and intracellular HSP70 was quantified from the leukocytes at 0, 2, 4, and 6 h after heat treatment. The concentration of HSP70 was not different between temperatures (P > 0.05), but the time-profile of HSP70 synthesis appeared temperature-dependent. At control (37 °C) and lower temperatures (38–39 °C) the mean HSP70 concentration increased up to 4 h post HS (P < 0.05) and then returned towards baseline values by 6 h post HS. With in vitro hyperthermic conditions (40–41 °C), the time-profile was characterised by a sharp rise in HSP70 levels immediately after treatment (P < 0.05 for 40 °C at 0 h), followed by a progressive decline over time. The results suggest a temperature-dependent time-profile of HSP70 synthesis. In addition, the temperature at which HSP70 is inducted might be lower than 37 °C.     

induction of heat shock response and acquisition of thermotolerance in callus cultures of Gerbera jamesonii

Summary The heat shock (HS) response in callus cultures of the ornamental plant Gerbera jamesonii H. Bolus var. hybrida was analyzed. A HS at 35° C or 40° C for 4 h induced (a) the synthesis of several heat shock proteins (HSPs), especially in the small molecular weight range and some spots corresponding to HSP70 components, and (b) an increase in the steady state levels of some specific mRNAs. At the nonstressing temperature (26° C), a sustainable level of translation for HSP70 was indeed carried out, as confirmed by immunological analysis with a monoclonal antibody against cotton HSP70. The steady state levels of mRNAs measured before and after a HS by Northern hybridization showed an increase with the heterologous probes HSP17.4, HSP17.6, and HSP21, whereas the probes HSC70 and HSP70 did not show any difference between the levels of control and HS-mRNAs. A pretreatment at 35° C, which induced a set of HSPs in the callus cultures, decreased the cell damage upon exposure to a temperature of 45° C as determined either with a regrowth test or by the tetrazolium reduction assay. Typically, as with the whole plants, callus of Gerbera jamesonii possessed the ability to respond to HS both by inducing HSPs and by developing an acquired thermotolerance.     

Transcript analysis and expression profiling of three heat shock protein 70 genes in the ectoparasitoid Habrobracon hebetor （Hymenoptera： Braconidae）

Heat shock proteins （HSPs） are known as chaperones that help with folding of other proteins when cells are under environmental stresses. The upregulation of HSPs is essential for cold survival during insect diapause. The ectoparasitoidHabrobracon hebetor, a potential biological control agent, can enter reproductive diapause when reared at low temperature and short photoperiod. However, the expression of HSPs during diapause of H. hebetor has not been studied. In this study, we sequenced and characterized the full-length complementary DNAs of three Hsp70 genes （HhHsp70I, HhHsp70II and HhHsp70IIl） from 11. hebetor. Their deduced amino acid sequences showed more than 80% identities to their counterparts from other insect species. However, the multiple se- quence alignment among the three deduced amino acid sequences of HhHsp70s showed only 46% identities. A phylogenetic analysis of the three HhHsp70s and all other known Hsp70 sequences from Hymenoptera clustered all the Hsp70s into four groups, and the three HhHsp70s were distributed into three different groups. Real-time quantitative poly- merase chain reaction analysis showed that the expression of the three HhHspTO genes in H. hebetor reared at different conditions was quite different. HhHspTOI showed higher relative expression when H. hebetor were reared at 27.5℃ than at two lower temperatures （17.5℃ and 20℃） regardless of the photoperiod, whereas HhHspTOII showed higher ex- pression when H. hebetor were reared at 20℃ and 10 ： 14 L ： D than when reared at 17.5℃ and either 16 ： 8 L ： D or 10 ： 14 L ： D. In contrast, HhHSP7OIIIwas expressed at similar levels regardless of the rearing conditions. These results may suggest functional differences among the three HhHspTO genes in H. hebetor.     

Effects of temperature and fish oil supplementation on ovarian development and foxl2 mRNA expression in spotted scat Scatophagus argus

In this study, the complete foxl2 complementary (c)DNA sequence was isolated by simple modular‐architecture research tool (SMART)er rapid amplification of cDNA ends (RACE). Two year‐old female spotted scat, Scatophagus argus, were reared at different temperatures (23, 26 and 29° C) for 6 weeks, or fed with different concentrations of dietary fish oil (0, 2 or 6%) for 8 weeks. Ovarian development, serum oestradiol‐17β (E₂) levels, as well as ovarian foxl2 expression were measured. At the end of experiment, ovarian foxl2 messenger (m)RNA expression in fish reared at 23 and 26° C was significantly higher than that in fish reared at 29° C, and that in 2 and 6% fish oil groups was also significantly higher than that in control group (P < 0·05). Serum E₂ levels exhibited the same trend with foxl2 mRNA expression in temperature treatment groups and fish oil fed groups. There was a significant positive correlation between stage of oocytes and foxl2 expressions. Results showed that from 23 to 29° C, the optimal temperature for ovarian development in S. argus was 23–26° C, and 6% fish oil supplementation could effectively promote ovarian development. Optimal temperature and fish oil supplement might increase ovarian foxl2 mRNA expressions to promote ovarian development in S. argus.     

HSP 70 gene expression in Trypanosoma cruzi is regulated at different levels

The level of HSP 70 mRNA is altered in Trypanosoma cruzi cells incubated at supra-optimal temperatures: the total amount of this RNA per cell is increased at 37°C, and slightly decreased at 40°C relative to its level at 29°C. However, its amount is greater in the polysomes at either temperature. The relative increase of this RNA is larger in the polysomes fraction than it is in the total RNA. In addition the level of HSP 70 protein in heat-shocked cells is greater than would be expected from the recruitment of HSP 70 mRNA in the polysomal fraction. Taken together the data are interpreted as indicating that at 37°C and 40°C the HSP 70 gene regulation in T. cruzi involves both the selective accumulation of the HSP 70 mRNA in the polysomes and its preferential translation. At 37°C, in addition, an increase in the total amount of this template is observed in the cells.     

Arabidopsis HSP70‐16 is required for flower opening under normal or mild heat stress temperatures

Sepals play important roles in protecting inner floral organs from various stresses and in guaranteeing timely flower opening. However, the exact role of sepals in coordinating interior and exterior signals remains elusive. In this study, we functionally characterized a heat shock protein gene, Arabidopsis HSP70‐16, in flower opening and mild heat stress response, using combined genetics with anatomic, physiological, chemical, and molecular analyses. We showed that HSP70‐16 is required for flower opening and mild heat response. Mutation of HSP70‐16 led to a significant reduction in seed setting rate under 22°C, which was more severe at 27°C. Mutation of HSP70‐16 also caused postgenital fusion at overlapping tips of two lateral sepals, leading to failed flower opening, abnormal floral organ formation, and impaired fertilization and seed setting. Chemical and anatomic analyses confirmed specific chemical and morphological changes of cuticle property in mutant lateral sepals, and qRT‐PCR data indicated that expression levels of different sets of cuticle regulatory and biosynthetic genes were altered in mutants grown at both 22°C and 27°C temperatures. This study provides a link between thermal and developmental perception signals and expands the understanding of the roles of sepal in plant development and heat response.     

Ambient temperature influences aging in an annual fish (Nothobranchius rachovii)

Extending lifespan by lowering ambient temperature in the habitat has been shown in a variety of organisms. Its mechanism, however, remains elusive. In this study, we examined the survivorship and the aging process of the annual fish (Nothobranchius rachovii) reared under high (30 °C), moderate (25 °C) and low (20 °C) ambient temperatures. The results showed that low ambient temperatures prolong survivorship, whereas high ambient temperatures shorten survivorship. At low ambient temperature, expression of senescence‐associated β‐galactosidase, lipofuscin, reactive oxygen species, lipid peroxidation, protein oxidation, mitochondrial density and ADP/ATP ratio were reduced compared with those reared at high and moderate temperatures, whereas catalase activity, Mn‐superoxide dismutase activities, mitochondrial membrane potential and the levels of ATP, ADP, Sirt1 and Forkhead box O expression were elevated. The expression levels of Hsp70 and CIRP showed no significant difference under any of the ambient temperatures tested. We concluded that cellular metabolism, energy utilization and gene expression are altered at lower ambient temperature, which is associated with the extension of lifespan of the annual fish.     

Responses of the bed bug,Cimex lectularius,to temperature extremes and dehydration: levels of tolerance,rapid cold hardening and expression of heat shock proteins

This study of the bed bug, Cimex lectularius, examines tolerance of adult females to extremes in temperature and loss of body water. Although the supercooling point (SCP) of the bed bugs was approximately −20°C, all were killed by a direct 1 h exposure to −16°C. Thus, this species cannot tolerate freezing and is killed at temperatures well above its SCP. Neither cold acclimation at 4°C for 2 weeks nor dehydration (15% loss of water content) enhanced cold tolerance. However, bed bugs have the capacity for rapid cold hardening, i.e. a 1‐h exposure to 0°C improved their subsequent tolerance of −14 and −16°C. In response to heat stress, fewer than 20% of the bugs survived a 1‐h exposure to 46°C, and nearly all were killed at 48°C. Dehydration, heat acclimation at 30°C for 2 weeks and rapid heat hardening at 37°C for 1 h all failed to improve heat tolerance. Expression of the mRNAs encoding two heat shock proteins (Hsps), Hsp70 and Hsp90, was elevated in response to heat stress, cold stress and during dehydration and rehydration. The response of Hsp90 was more pronounced than that of Hsp70 during dehydration and rehydration. Our results define the tolerance limits for bed bugs to these commonly encountered stresses of temperature and low humidity and indicate a role for Hsps in responding to these stresses.     

Cloning of heat shock protein genes from the brown planthopper,Nilaparvata lugens,and the small brown planthopper,Laodelphax striatellus,and their expression in relation to thermal stress

Three heat shock protein (HSP) genes (hsp70, hsc70, hsp90) were partially cloned from the brown planthopper Nilaparvata lugens and the small brown planthopper Laodelphax striatellus (Homoptera: Delphacidae), which are serious pests of the rice plant. Sequence comparisons at the deduced amino acid level showed that the three HSPs of planthoppers were most homologous to corresponding HSPs of dipteran and lepi‐dopteran species. Identities of both heat shock cognate 70 and HSP90 were higher than HSP70 in both species. Identity of the HSP70 between the two planthopper species was only 81%, a value much lower than seen among fly and moth groups. Effects of heat and cold shocks were demonstrated on expression of the three hsp genes in the two planthopper species. Heat shock (40 °C) upregulated the hsp90 level but did not change the hsc70 level in either the nymph and adult stages of either species. On the other hand, the hsp70 level was only upregulated in L. striatellus. This heat shock response was prompt and lasted only for 1 h after treatment. In contrast, cold shock at 4°C did not change the expression levels of any hsp in either species.