Variation in heat shock proteins within tropical and desert species of poeciliid fishes

The 70-kilodalton heat shock protein (hsp70) family of molecular chaperones, which contains both stress-inducible and normally abundant constitutive members, is highly conserved across distantly related taxa. Analysis of this protein family in individuals from an outbred population of tropical topminnows, Poeciliopsis gracilis, showed that while constitutive hsp70 family members showed no variation in protein isoforms, inducibly synthesized hsp70 was polymorphic. Several species of Poeciliopsis adapted to desert environments exhibited lower levels of inducible hsp70 polymorphism than the tropical species, but constitutive forms were identical to those in P. gracilis, as they were in the confamilial species Gambusia affinis. These differences suggest that inducible and constitutive members of this family are under different evolutionary constraints and may indicate differences in their function within the cell. Also, northern desert species of Poeciliopsis synthesize a subset of the inducible hsp70 isoforms seen in tropical species. This distribution supports the theory that ancestral tropical fish migrated northward and colonized desert streams; the subsequent decrease in variation of inducible hsp70 may have been due to genetic drift or a consequence of adaptation to the desert environment. Higher levels of variability were found when the 30- kilodalton heat shock protein (hsp30) family was analyzed within different strains of two desert species of Poeciliopsis and also in wild-caught individuals of Gambusia affinis. In both cases the distribution of hsp30 isoform diversity was similar to that seen previously with allozyme polymorphisms.      

Effect of Temperature on the Activity of Carboxylases in Tropical and Temperate Gramineae

Temperate Gramineae show maximal net photosynthesis at 20–5°C, whereas tropical Gramineae have maxima between 30 and35 °C. Moreover, it has been suggested that different carboxylationreactions are involved in the two groups. The present studyof the temperature dependance of in vitro ribulose-1,5- diphosphate(RuDP), and phosphopyruvate (PEP) carboxylases indicates thatthe two enzymes have clearly marked differences in temperaturesensitivity. RuDP carboxylase, present in the temperate andtropical species studied, showed maximal activity around 20–5°C except in Zea. By contrast, PEP carboxylase activityin all species was maximal between 30 and 35 °C. The dataimply that activity and temperature sensitivity of the relevantcarboxylase enzymes may well be a significant limiting factorin leaf photosynthesis, even at light saturation.     

Plankton composition, abundance and dynamics in a severely stressed cooling reservoir

The effects of severe thermal stress imposed by the intermittentoperation of a nuclear reactor on plankton abundance and dynamicswere investigated in Pond C, a cooling reservoir on the SavannahRiver Site in South Carolina, USA. Temperatures in Pond C rangedup to 58°C during reactor operation. The thermal effluenteliminated zooplankton from regions where the temperature exceeded45°C, reduced zooplankton abundance by 1–3 ordersof magnitude and typically halved the number of taxa. Reactoroperation also reduced phytoplankton biovolume, often by >70%.During intermittent reactor operation, the rotifer Filinia longisetadominated the zooplankton and two cladocerans of the genus Moinawere abundant. These species were not abundant during extendedreactor shutdowns. The success of Filinia and Moina was dueprimarily to their tolerance of high temperatures. Sparse phytoplanktonprobably limited some zooplankton taxa, although other taxa,such as Filinia may have utilized bacterial resources. Reactoroperation may have intensified predation on crustacean zooplanktonwhen fish were concentrated in refuge areas with zooplankton.Processes by which zooplankton repopulated the reservoir afterreactor shutdown were inferred from zooplankton distributionpatterns, and population growth and birth rates. Repopulationtypically occurred within a few days due to rapid growth ofpopulations from refuge areas within the reservoir and colonistsbrought in through a tributary canal. Mechanisms of zooplanktonrepopulation in Pond C suggest that refuges or colonizationcorridors should be maintained when the re-establishment ofcommunities following cessation of stresses is desired.     

Heat Shock Proteins of Cyanobacterium Anacystis nidulans

Cells of Anacystis nidulans grown at 30°C were incubatedwith ¹⁴C-Chlorella protein hydrolysate at the elevated temperatures(30–55°C) and the effect of heat shock treatment onprotein synthesis was studied. Incubation temperatures higherthan 45°C caused a significant decrease in the incorporationof amino acids into proteins. Further, the heat shock treatmentinduced significant changes in the fluorographic profile ofthe newly synthesized proteins. (Received October 25, 1985; Accepted December 4, 1985)     

Spore Germination in Two Tropical Mosses: Fissidenssp. and Racopilum sp

Optimum germination of spores of a Fissidens species occurredat a relatively higher temperature (30 °C) than the optimumgermination of those of a Racopilum species (25 °C). Subsequentprotonemal growth of the two mosses also showed the same differentialtemperature optima. The high-temperature requirement for germinationof Fissidens spores happens to coincide with the maturationand dispersal of the spores in the dry season, and apparentlyfavours the establishment of new shoots. Fissidens sp, Racopilum sp, tropical mosses, spore germination, temperature adaptation     

Starvation reduces the heat shock protein responses in white sturgeon larvae

This study investigates the responses of white sturgeon larvae (Acipenser transmontanus) to starvation and thermal stress, through the measurement of nutritional status (i.e. growth performances) and cellular biomarkers: heat shock proteins (Hsp) 70 and 90. White sturgeon larvae (25 day post hatch; initial weight 179.0 ± 5.1 mg) were fed (20% body weight per day) or starved for 24, 48 or 72 hrs. Every 24 hrs, five larvae from each of the starved or fed treatment replicates were exposed to heat shock resulting from an increase in water temperature from 19°C to 26°C, at a rate of 1°C per 15 min, and maintained at 26°C for 4 hrs. No mortality was observed in this study. Starvation significantly (p < 0.05) decreased the body weight and body contents of energy, protein, and lipid of the experimental larvae, compared to the fed larvae. Heat shock induced the expressions of Hsp70 and Hsp90 in both the fed and starved group; however, starvation reduced the induction at all sampling points. The current study demonstrates that poor larval nutritional status, assessed by the aforementioned parameters, reduced heat shock responses to thermal stress, as measured by heat shock protein levels. Furthermore, Hsp70 and 90 are more sensitive to heat shock and starvation, respectively. This may be, in part, a result of the different functioning of the heat shock proteins in cellular stress response and warrants further study.     

Effects of Temperature on the In vivo Assay of Nitrate Reductase in some C3 and C4Species

Nitrate reductase (NR) activity was measuredin vivo in differentcrop species at 25–60 °C. Highest NR activity wasobserved at 40 °C in pigeon pea, cowpea, sunflower, sesameand sorghum, 45 °C in maize and 50 °C in bajra. At higherincubation temperatures NR activity declined in all species.In mung bean the optimum incubation temperature varied withthe season. In the summer crop of mung beans, NR activity wasmaximum at 50 °C while in the rainy season crop 30 °Cwas the optimum incubation temperature. NR in sesame was relativelyheat-tolerant. The results indicate that 30 °C is not theoptimum incubation temperature for all crop species. Nitrate reductase, temperature, in vivo assay     

Seed Germination and Radicle Growth of a Halophyte, Kalidium caspicum(Chenopodiaceae)

Effects of temperature, light, NaCl and polyethylene glycol(PEG)-6000 on seed germination and radicle growth in a halophyticshrub, Kalidium caspicum(L.) Ung.-Sternb. were investigated.When seeds were incubated in deionized water at constant temperaturesbetween 10 and 30°C, the percentage germination in the darkexceeded 75%; light suppressed seed germination at alternatingtemperatures. Incubating seeds with a hypersaline solution ofNaCl for 30 d had no adverse effect on their germinability.The percentage germination of seeds incubated with a –0.8MPa NaCl solution was 73, 80 and 54% at 10, 20 and 30°C,respectively, but all radicles died before their length exceeded5 mm. In contrast, when seeds were incubated with a –0.8MPa PEG solution at 20°C, 68% of seeds germinated, and 95%of the emerging radicles survived beyond 5 mm. The high sensitivityof small radicles of this species to salinity indicated thatsalt must be removed from the soil surface for seedling establishment.Copyright2000 Annals of Botany Company Chinese desert, radicle growth, germination, halophyte, Kalidium caspicum, salinity     

Influence of Temperature on the Effects of Water Stress on Quercus Species

Quercus coccifera L. and Q. ilex L. leaves, collected in winter,when the plants were in full turgor, were treated with polyethyleneglycol (PEG) with a water potential of –48 and –65bar, at 15 °C and at 30 °C, for 2 days. The responseof each species differs with temperature as far as the plastidpigment, anthocyanin and soluble sugar content is concerned.Thus Q. ilex is affected more at 15 °C rather than at 30°C, while in Q. coccifera, apart from a small increase inanthocyanins, no significant change was observed at 15 °C.On the contrary, at 30 °C significant changes were observedin all studied parameters. Chloroplast pigments decreased generallyon the first day and their content either dropped further orincreased to reach the control values on the second day. Alltreatments resulted in an increase in anthocyanin content. Solublesugar content raised in Q. ilex at 15 °C and decreased inboth species at 30 °C. Quercus coccifera, Quercus ilex, oak, temperature, water stress, anthocyanin, chlorophyll, carotenoids     

Sugar Content and Metabolic Activities in Cold-Stored Fragmented Xylopodium of Ocimum nudicaule Benth. var. anisifolia Giul. (Labiatae)

Sugar content and hydrolytic activities were measured in fragmentsof xylopodia of Ocimum nudicaule Benth. stored for one weekat 5 °C and 5.28 °C. At 5 °C the soluble carbohydratecontent first decreased and at 5.28 °C it increased duringstorage and then declined drastically during cultivation afterplanting. Galactose, glucose and fructose levels also rose at5–28 °C whereas at 5 °C only glucose increased.These changes in sugar level were found to be correlated withchanges in the activity of certain hydrolytic enzymes: at 5–28°C there was a rapid rise in the activities of -galactosidaseand hemicellulase followed by an increase in the activity ofinvertase and ß-galactosidase. The rapid changes inenzymes and carbohydrate levels are discussed in terms of adaptationof the plant to tropical and subtropical environments. Key words: Carbohydrates, Carbohydrate hydrolases, Underground organ (xylopodium), Cold storage     

Pigment Production by Cultured Florets of Chrysanthemum morifolium

Individual florets (4–5 mm long) of a purple cultivar(Fandango) of the horticultural chrysanthemum (Chrysanthemummorifolium Ramat) were taken from flower buds just prior toopening and cultured in a sterile liquid medium (containinginorganic salts and sucrose) at 15 °C under a 12-h day.For the first 14 days increase in wet weight was exponential.Anthocyanin appeared on the third day and was then synthesizedrapidly. Chlorophyll and carotenoid were present initially:carotenoid levels rose quickly while chlorophyll remained almostconstant. Highest pigment content and most growth were foundwhen the florets were grown on 3 per cent sucrose. However,the highest anthocyanin concentration was found with 4 per centsucrose, the highest carotenoid concentration with 0.6 per centsucrose. No anthocyanin was produced when the florets were grownat 6 or 30 °C; maximum yield was at 15 °C. Most carotenoidwas formed at 30 °C and most chlorophyll was found at 20–5°C. All florets from 1 to 7 mm long could be cultured. Theseresults are discussed in relation to flower colour and pigmentformation in vivo.     

In vitro heat shock of human monocytes results in a proportional increase of inducible Hsp70 expression according to the basal content

Heat shock proteins play an important role as molecular chaperones of the cell. Inducible heat shock protein 70 is rapidly synthesised in response to numerous stressors and monocytes are sensitive to changes in core temperature resulting in a circadian variation of Hsp70 expression. Monocytes were isolated via density centrifugation from nine healthy male volunteers at 5 am, 1 pm and 9 pm, representing the nadir (5 am), peak (9 pm) and intermediate (1 pm) of Hsp70 expression in the 24-h cycle. Analysis of freshly isolated monocytes for Hsp70 expression confirmed Hsp70 levels at the three selected time points. Monocytes were subjected to in vitro heat shock at 40°C (±0.1) for 90 min with a 90 min 37°C (±0.1) exposure acting as a control. A significant increase in Hsp70 was observed at 5 am (p < 0.001) and 1 pm (p = 0.028) at 40°C when compared to 37°C but not at 9 pm (p = 0.19). A significant increase was also observed from the basal levels of Hsp70, measured on freshly isolated monocytes and the levels detected after heat shock at 40°C at 5 am (p < 0.001) and 1 pm (p = 0.001), which was not observed at 9 pm (p = 0.15). Furthermore, a significant correlation was observed in the heat shock response at 40°C and that obtained at 37°C (p < 0.001). In conclusion, the heat shock response in monocytes is directly proportional to the amount of Hsp70 present in the cells and the stress response may be much higher at different times of the day.     

GLOCHIDIA OF THE FRESHWATER MUSSEL LAMPSILIS OVERWINTERING ON FISH HOSTS

Largemouth Bass were infected with glochidia of the freshwatermussel Lampsilis cardium. Three fishes each were held at 4.5,10, and 15.5°C; five fish were held at 21°C. By 64 days,metamorphosed juveniles were found in the 15.5 and 21°Ctrials but not in the 5.5 and 10°C trials, indicating thatthe lower threshold temperature for metamorphosis was between10 and 15.5°C for the duration. In a second experiment,Largemouth Bass were infected with glochidia of L. cardium andheld at 10°C. A sample of fishes was removed monthly andbrought to 21°C. Numbers of glochidia that metamorphosedafter being warmed were compared to the number that metamorphosedwithout warming. The percentage that metamorphosed after warmingdecreased linearly with time. At one month, 100% of the glochidiametamorphosed after warming. This decreased to 80% by two months,to 30% by four months and 3% by six months. Although this post-warmingpercentage decreased with time, the total percentage of metamorphosedjuveniles (at all temperatures) was not correlated with time.Controls kept at 21°C required three weeks to reach peakmetamorphosis, but test subjects subjected to 10°C requiredless than nine days to metamorphose once warmed. Many overwinteringglochidia therefore complete a portion of their developmenton the host at winter temperatures, but stop short of excystment.Some glochidia metamorphosed without being warmed, but thisphenomenon is not understood. This study confirms that glochidiamay overwinter on hosts, with some glochidia persisting formore than six months before metamorphosing when warmer conditionsreturn. (Received 29 September 1998; accepted 18 January 1999)     

Regulation of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis> Plasma membrane H<Superscript>+</Superscript>-ATPase (Pma1) by Dextrose and Hsp30 during Exposure to Thermal Stress

Pma1p is an essential plasma membrane H⁺-pump in Saccharomyces cerevisiae that pumps out H⁺ at the expense of cellular ATP. Its activity is induced by glucose at 30°C and is inhibited by Hsp30 during exposure to heat shock conditions. To further investigate the regulation of Pma1 function by glucose and Hsp30 during exposure to thermal stress, we estimated Pma1 activity, its protein levels and ser-phosphorylation status in membrane fractions isolated from BY4741 and hsp30Δ cells grown in dextrose and sorbitol at 30°C, and following exposure at 40°C for 30 min. Our results demonstrate that Pma1 activity and protein levels were reduced in Hsp30⁺ cells following exposure to thermal stress in dextrose media. The above was not observed in hsp30Δ cells wherein Pma1 activity did not decrease following exposure to similar conditions. Although Pma1p levels decreased in heat-shocked hsp30Δ cells, it was lower compared to that observed in Hsp30⁺ cells. Total ser-phosphorylation of Pma1 also showed a decrease following exposure to heat shock condition in dextrose media in both BY4741 and hsp30Δ cells. Its levels were also reduced in BY4741 cells upon heat shock treatment in sorbitol unlike that observed in hsp30Δ cells wherein it was increased. Taken together the above indicate that heat shock induced reduction in Pma1 activity and protein levels in dextrose media required Hsp30. To examine functional interactions between dextrose utilization, Hsp30 and the regulation of various aspects of Pma1, we determined if dextrose regulated other functions attributed to Hsp30. Results demonstrate that the deletion of HSP30 rendered cells dependent on dextrose utilization for survival during exposure to lethal heat stress. Our study has hence been able to establish a functional relationship between glucose utilization, Hsp30 function and the regulation of Pma1 activity. Finally, since the deletion of HSP30 renders Pma1p levels and its activity unresponsive to thermal stress in dextrose media, we concluded that Hsp30 is necessary to maintain Pma1 in a regulation competent conformation. Hsp30 may thus act as a chaperone in the S. cerevisiae plasma membrane.     

Expression patterns and organization of the hsp70 genes correlate with thermotolerance in two congener endemic amphipod species (Eulimnogammarus cyaneus and E. verrucosus) from Lake Baikal

We studied various aspects of heat‐shock response with special emphasis on the expression of heat‐shock protein 70 (hsp70) genes at various levels in two congener species of littoral endemic amphipods (Eulimnogammarus cyaneus and E. verrucosus) from Lake Baikal which show striking differences in their vertical distribution and thermal tolerance. Although both the species studied demonstrate high constitutive levels of Hsp70, the thermotolerant E. cyaneus exhibited a 5‐fold higher basal level of Hsp70 proteins under normal physiological conditions (7 °C) and significantly lower induction of Hsp70 after temperature elevation compared with the more thermosensitive E. verrucosus. We isolated the hsp70 genes from both species and analysed their sequences. Two isoforms of the cytosolic Hsp70/Hsc70 proteins were detected in both species under normal physiological conditions and encoded by two distinct hsp/hsc70 family members. While both Hsp70 isoforms were synthesized without heat shock, only one of them was induced by temperature elevation. The observed differences in the Hsp70 expression patterns, including the dynamics of Hsp70 synthesis and threshold of induction, suggest that the increased thermotolerance in E. cyaneus (compared with E. verrucosus) is associated with a complex structural and functional rearrangement of the hsp70 gene family and favoured the involvement of Hsp70 in adaptation to fluctuating thermal conditions. This study provides insights into the molecular mechanisms underlying the thermal adaptation of Baikal amphipods and represents the first report describing the structure and function of the hsp70 genes of endemic Baikal species dwelling in thermally contrasting habitats.     

Field and laboratory investigations of the thermal influence on tissue-specific Hsp70 levels in common carp (Cyprinus carpio)

Thermal discharge from power stations can affect normal environmental conditions and change in heat shock proteins expression of native fish with increasing temperature. In this study, we investigated levels of Hsp70 in the heart, kidney, brain and gill of the common carp Cyprinus carpio both in long-term heat discharge environment and after 24 h acute heat shock exposure. In laboratory exposure experiments, fish acclimated at 10 °C were exposed to various elevated temperatures (20, 24 and 28 °C). Hsp70 concentrations were determined in tissues by Western blotting analysis after one dimensional SDS-PAGE separation. In the field study, the level of Hsp70 in the gill of the carp remained at control values, and Hsp70 expression in the heart, kidney and brain underwent a 2.8 to 3.7-fold increase. A lower thermal sensitivity of the Hsp70 response of the brain, compared with the heart, kidney and gill, was observed in the laboratory experiments. Our data show that these tissues had different levels of Hsp70 responses to thermal influence both in acute exposure and long-term acclimation. The pattern of tissue Hsp70 expression may have a close relationship with the thermal tolerance of the carp and allows the fish to survive long-term thermal pollution.     

Molecular characterization and gene evolution of the heat shock protein 70 gene in snakehead fish with different tolerances to temperature

Temperature tolerances (including lethal limits) and associated rates of thermal acclimation of fish are critical information in predicting fish responses to global climate changes. In this study, a partial sequence of the heat shock protein 70 gene (HSP70) from the fish species Channa striatus was isolated and characterized. Evolutionary process that led to the diversity of HSP70 specific to vertebrates was also analysed. Results revealed that HSP70 is highly homologous in other fish families. The conservation of the HSP 70 gene among fish families could be driven by forces of natural selection due to climatic change. We exposed C. striatus to heat shock (32 °C) and cold shock (16 °C) respectively, in order to examine the differences of temperatures in influencing the expression patterns of HSP70. We revealed that expression of HSP70 was higher at 32 °C than at 16 °C in most of the organs. Specifically, occurrence of chaperone activity of HSP70 was found at low temperature. Therefore, this fish was postulated that to seems to be able to survive at lower temperature compared to higher temperature indicating there is force of natural selection acting towards this HSP 70 gene. This will demonstrate the effect of global warming towards the fish survivability.