Activation of plasma membrane H(+)-ATPase and expression of PMA1 and PMA2 genes in Saccharomyces cerevisiae cells grown at supraoptimal temperatures.

During exponential growth at temperatures of 30 to 39 degrees C, the specific activity of H(+)-ATPase in the plasma membrane of Saccharomyces cerevisiae (assayed at the standard temperature 30 degrees C) increased with increases in growth temperature. In addition, the optimal temperature for in vitro activity of this ATPase was 42 degrees C. Therefore, the maximum values of ATPase activity were expected to occur in cells that grew within the supraoptimal range of temperatures. Activation induced by supraoptimal temperatures was not the result of increased synthesis of this membrane enzyme. When the growth temperature increased from 30 to 40 degrees C, expression of the essential PMA1 gene, monitored either by the level of PMA1 mRNA or the beta-galactosidase activity of the lacZ-PMA1 fusion, was reduced. Consistently, quantitative immunoassays showed that the ATPase content in the plasma membrane decreased. Like ATPase activity, the efficiency of the PMA2 promoter increased with increases in growth temperature in cells that had been grown at 30 to 39 degrees C, but its level of expression was several hundred-fold lower than that of PMA1. These results suggest that the major PMA1 ATPase is activated at supraoptimal temperatures.     

Effects of Supraoptimal Temperatures on Merulius lacrymans

The growth rate of Merulius lacrymans (Jacq.) Fr. decreases very rapidly at temperatures above optimum (about 22°C). No growth occurred above 28°C. The thermal death-time of the fungus at different temperatures above maximum was determined. At 37.5°C the thermal death-time is 4 hours. At this temperature, the respiration is not directly affected, since an appreciable respiration occurred even after 6 hours. On the basis of ¹⁴C-experiments and analysis of UV-absorbing material excreted from the mycelium at supraoptimal temperatures, it is suggested that the detrimental effects of supraoptimal temperatures on this fungus include a degradation of the nucleic acids and a subsequent leakage of the nucleotides out of the cells.     

Effect of temperature on the physiology and cytology of the methylotrophic yeastCandida boidinii growing in methanol-limited chemostat

All deviations from optimum cultivation temperature affect strongly the physiology and morphology of cells ofCandida boidinii strain 2 during growth in methanol-limited chemostat. The optimum cultivation temperature was 28–30 °C at which maximum cell concentration and maximum cell yield (Y _S 0.4 g/g) were achieved. At suboptimal growth temperatures the cells were rich in cell protein, RNA, alcohol oxidase (AO) and in peroxisomes. Formation of cubic peroxisomes and a 20 % decrease of budding cells in the population was observed. At supraoptimal growth temperatures (>30 °C) a sharp decrease in AO activity was accompanied by degradation of peroxisomes in the cells. The culture forms pseudomycelium: at 34 °C the cells stop growing and they are washed out of the bioreactor.     

Physiological Responses of Apple Trees to Supraoptimal Root Temperature

Ungrafted apple rootstocks were grown in sand cultures at constant root temperatures between 20°C to 40°C. Temperatures of 30°C and above reduced root and shoot growth. Serious damage to the leaves occurred at 35°C and above. The O₂ consumption, CO₂ evolution and respiratory quotient (RQ) of the roots showed maximum values at 35°C. Different rootstock cultivars varied greatly in their susceptibility to damage by supraoptimal root temperatures apparently due to anaerobic respiration. The more susceptible ones differed from resistant types in the larger amount of ethanol they accumulated in their roots at supraoptimal root temperature, and the more severe reduction in the malic acid content of the roots at such temperature. Acetaldehyde was also found in roots and leaves at supraoptimal root temperatures, whereas the organic acid content of the leaves tended to decrease. Supraoptimal root temperature also caused a reduction of cytokinins in both roots and leaves accompanied by a reduction in the leaf chlorophyll content. This could be prevented by the application of kinetin or benzyladenine to the leaves. In a short experiment a rise in root temperature up to 40°C caused an increase in transpiration and a decrease in the resistance of the leaves to the passage of water vapor, whereas in prolonged experiments transpiration reached a maximum and leaf resistance a minimum at 30°C. The leaf water potential increased also with increasing root temperature. Leaf temperature increased with increasing root temperature, irrespective of increasing or decreasing transpiration rates.     

Intracellular protein degradation in Neurospora crassa.

In exponentially growing cultures of Neurospora crassa, the basal rate of protein degradation increases as the constant of the rate of growth decreases, so that in slow growing cells (mu = 0.13) the rate of protein degradation is about 25% of the rate of protein accumulation. During glucose starvation and shift-down transition of growth, the rate of protein degradation is greatly enhanced, and a moderate reduction (about 30%) of the ATP level is observed. Treatment of glucose-starved cells with 2-deoxyglucose reduces the ATP content by 70% and blocks protein degradation. The addition of cycloheximide, given at the onset of glucose starvation, prevents the enhancement of protein degradation; instead cycloheximide is without effect if added when proteolysis has already started. At a supraoptimal temperature (42 degrees C) the basal rate of protein degradation is not stimulated, contrary to the behavior observed in bacteria. Guanosine nucleotides, which appear to have a regulatory role for protein degradation in bacteria, are not found in N. crassa.     

The effect of temperature on the lipid composition of Botryococcus

The lipid composition of the green alga Botryococcus was studied at three different cultivation temperatures: suboptimal, optimal, and supraoptimal. Cultivation at the supraoptimal temperature was found to considerably inhibit the synthesis of nearly all intracellular lipids, except for triacylglycerides, and to influence their fatty acid composition. In particular, the content of trienoic fatty acids was significantly lower at the supraoptimal than at the optimal cultivation temperature. At the same time, the fatty acid composition of the extracellular lipids of the alga virtually did not depend on cultivation temperature.     

Immune suppression with supraoptimal doses of antigen in contact sensitivity. I. Demonstration of suppressor cells and their sensitivity to cyclophosphamide.

Immunologic suppression was induced in a mouse model of contact sensitization to DNFB by using supraoptimal doses of antigen. In these studies, in vivo measurement of ear swelling as an indication of immunologic responsiveness correlated well with measurement of in vitro antigen-induced cell proliferation. This unresponsiveness was specific, since supraoptimal doses of DNFB did not interfere with the development of contact sensitivity to another contactant, oxazolone. The decrease in responsiveness is a form of active suppression, as lymphoid cells from supraoptimally sensitized donors transferred suppression to normal recipients. Furthermore, pretreatment with cyclophosphamide (Cy) reversed the suppression seen in supraoptimally sensitized animals but had no effect on the optimal sensitization regimen. These results indicate that supraoptimal doses of contactants can activate suppressor cells and that precursors of these cells are sensitive to Cy. Such suppressors regenerate within 7 to 14 days after Cy treatment. The ability of Cy pretreatment to affect supraoptimal sensitization without affecting optimal sensitization confirms other reports indicating that the observed results of Cy treatment depend critically upon the dose of antigen used.     

The Effect of Temperature on the Lipid Composition of the Green Alga Botryococcus

The lipid composition of the green alga Botryococcus was studied at three different cultivation temperatures: suboptimal (18°C), optimal (25°C), and supraoptimal (32°C). Cultivation at the supraoptimal temperature was found to considerably inhibit the synthesis of nearly all intracellular lipids, except for triacylglycerides, and to influence their fatty acid composition. In particular, the content of trienoic fatty acids was significantly lower at the supraoptimal than at the optimal cultivation temperature. At the same time, the fatty acid composition of the extracellular lipids of the alga virtually did not depend on cultivation temperature.     

Effect of vitamin B12 and folate on biosynthesis of methionine from homocysteine in the nematode Caenorhabditis briggsae.

(i) Omission of L-methionine from the medium resulted in an 80% population reduction. Substitution of D,L-homocysteine corrected methionine deficiency in C. briggsae in the presence of supraoptimal vitamin B12 and folic acid. (ii) An absolute vitamin B12 requirement in C. briggsae developed in the medium containing homocysteine at the second subculture. Concentration of 6 ng/ml of vitamin B12 (at 100 ng/ml of folic acid) was sufficient to support maximum growth of C. briggsae in the medium containing homocysteine. (iii) It was found that either supraoptimal folic acid (2000 ng/ml) or supraoptimal vitamin B12 (3750 ng/ml), with homocysteine, supported very little population growth of C. briggsae. However, supraoptimal folic acid and supraoptimal vitamin B12 together supported a maximum population growth. Therefore, it was concluded that both vitamin B12 and folic acid were required for the biosynthesis of methionine from homocysteine. Studies also showed that the two vitamins spared each other for population growth in the medium containing homocysteine.     

Phosphorylation-dephosphorylation changes in yeast ribosomal proteins S2 and S6 during growth under normal and hyperthermal conditions

Dynamic changes were observed in the phosphorylation level of a basic ribosomal protein S6 under different growth conditions of yeast culture. The maximum level of S6 phosphorylation occurred within 30 min of 32P-labelling after the transfer of cells into fresh nutrient medium. The elevation of temperature to supraoptimal level (38 degrees C and 41 degrees C) led to extensive dephosphorylation of S6 protein, and the recovery from heat shock was characterized by its rephosphorylation. Contrary to S6, phosphorylation of ribosomal protein S2 remained on unchanged level, irrespective of growth conditions.     

Selective diminishment of virus-specific translatable mRNA in the densonucleosis virus-infected midgut of the silkworm, Bombyx mori, reared at a supraoptimal temperature

Effect of a supraoptimal temperature on the accumulation of virus-specific translatable mRNAs was examined in the larvae of the silkworm. Bombyx mori, infected with Bombyx densonucleosis virus type 2. The results showed that incubation of infected larvae at 35 degrees C resulted in a rapid and selective decrease in the virus-specific translatable mRNAs which preexisted in the infected midgut. Upon temperature-shift from 35 degrees to 25 degrees C, the virus-specific translatable mRNAs became clearly observed within 12 hr. These results indicate that a supraoptimal temperature restricts the accumulation of virus-specific translatable mRNAs. Both rapid decay and suppressed synthesis might be responsible for the selective restriction of virus-specific translatable mRNA accumulation at a supraoptimal temperature.     

Temperature profiles of cellular growth and exopolysaccharide synthesis by Botryococus braunii Kütz. UC 58

Temperature profiles (range 20–33 °C) were obtained for growth and exopolysaccharide (EPS) biosynthesis of the microalga Botryococcus braunii strain UC 58 under photoautotrophic conditions. The maximum temperature for growth was 32 °C and the temperature dependence of the specific growth rate was described by the Hinshelwood equation based on the Arrhenius relationship. The optimal range of temperatures for growth and extracellular EPS synthesis (25–30 °C) concurred and production of 4.5–5 g l⁻¹ of EPS was obtained routinely, leading to high broth viscosities. Below 23 °C EPS biosynthesis was negligible, although the specific growth rate maintained high values. At supraoptimal temperatures EPS biosynthesis decreased, accompanying the increase in doubling time. The polymers formed at temperatures within the optimal range for production, when dissolved in water, produced solutions (2 gl⁻¹) with the highest viscosity, suggesting that their molecular weight showed the highest values. The degree of polymerization of the EPS synthesized at suboptimal and supraoptimal temperatures was significantly below the values within the optimal range.     

Interactive Effects of CO2, Temperature,Irradiance, and Nutrient Limitation on the Growth and Physiology of the Marine Diatom Thalassiosira pseudonana (Coscinodiscophyceae)

The marine diatom Thalassiosira pseudonana was grown in continuous culture systems to study the interactive effects of temperature, irradiance, nutrient limitation, and the partial pressure of CO₂ (pCO₂) on its growth and physiological characteristics. The cells were able to grow at all combinations of low and high irradiance (50 and 300 μmol photons · m⁻² · s⁻¹, respectively, of visible light), low and high pCO₂ (400 and 1,000 μatm, respectively), nutrient limitation (nitrate-limited and nutrient-replete conditions), and temperatures of 10–32°C. Under nutrient-replete conditions, there was no adverse effect of high pCO₂ on growth rates at temperatures of 10–25°C. The response of the cells to high pCO₂ was similar at low and high irradiance. At supraoptimal temperatures of 30°C or higher, high pCO₂ depressed growth rates at both low and high irradiance. Under nitrate-limited conditions, cells were grown at 38 ± 2.4% of their nutrient-saturated rates at the same temperature, irradiance, and pCO₂. Dark respiration rates consistently removed a higher percentage of production under nitrate-limited versus nutrient-replete conditions. The percentages of production lost to dark respiration were positively correlated with temperature under nitrate-limited conditions, but there was no analogous correlation under nutrient-replete conditions. The results suggest that warmer temperatures and associated more intense thermal stratification of ocean surface waters could lower net photosynthetic rates if the stratification leads to a reduction in the relative growth rates of marine phytoplankton, and at truly supraoptimal temperatures there would likely be a synergistic interaction between the stresses from temperature and high pCO₂ (lower pH).     

Effects of Visible Light and UV Radiation on Photosynthesis in a Population of a Hot Spring Cyanobacterium, a Synechococcus sp., Subjected to High-Temperature Stress

Assays of photosynthesis were conducted with a biofilm population of a cyanobacterium, a Synechococcus sp., growing at ~70°C in a Yellowstone National Park hot spring to test whether cells growing near the upper temperature limit of photosynthetic life are optimally adapted to their mean environmental temperature. Cell suspensions were assayed at 70, 65, and 55°C while being simultaneously exposed to modified solar environments, including reduction of total irradiance and exclusion of UV radiation. Carbon fixation was greatest at 65°C, while 70 and 55°C were always supraoptimal and suboptimal for photosynthesis, respectively. The degree of temperature stress was dependent upon light intensity, and this light-dependent temperature effect may involve both reduced quantum efficiency at subsaturating irradiances and a lower saturating irradiance at both supraoptimal and suboptimal temperatures. The Synechococcus sp. was also more susceptible to UV inhibition of photosynthesis at nonoptimal temperatures. These results suggest that this population is persisting at a nearly lethal temperature and is consequently subject to greater damage by both visible and UV radiation, but it is speculated that these cells may be avoiding competition with other photoautotrophs under these nonoptimal conditions. In separate experiments monitoring diurnal patterns of photosynthesis, cells exhibited peak productivity during the morning, followed by an afternoon decline. No recovery of photosynthesis was observed during the remaining daytime, and carbon fixation was always UV inhibited under conditions of photosynthetically saturating light.     

SHIP down-regulates FcepsilonR1-induced degranulation at supraoptimal IgE or antigen levels

Cross-linking of the IgE-loaded high-affinity IgE receptor (FcepsilonR1) by multivalent Ags results in mast cell activation and subsequent release of multiple proinflammatory mediators. The dose-response curve for FcepsilonR1-mediated degranulation is bell-shaped, regardless of whether the IgE or the Ag concentration is varied. Although overall calcium influx follows this bell-shaped curve, intracellular calcium release continues to increase at supraoptimal IgE or Ag concentrations. As well, overall calcium mobilization adopts more transient kinetics when stimulations are conducted with supraoptimal instead of optimal Ag concentrations. Moreover, certain early signaling events continue to increase whereas degranulation drops under supraoptimal conditions. We identified SHIP, possibly in association with the FcepsilonR1 beta-chain, as a critical negative regulator acting within the inhibitory (supraoptimal) region of the dose-response curve that shifts the kinetics of calcium mobilization from a sustained to a transient response. Consistent with this, we found that degranulation of SHIP-deficient murine bone marrow-derived mast cells was not significantly reduced at supraoptimal Ag levels. A potential mediator of SHIP action, Bruton's tyrosine kinase, did not seem to play a role within the supraoptimal suppression of degranulation. Interestingly, SHIP was found to colocalize with the actin cytoskeleton (which has been shown previously to mediate the inhibition of degranulation at supraoptimal Ag doses). These results suggest that SHIP, together with other negative regulators, restrains bone marrow-derived mast cell activation at supraoptimal IgE or Ag concentrations in concert with the actin cytoskeleton.     

Effect of continuous heat stress on cell growth and protein synthesis in Aedes albopictus

Aedes albopictus (clone C6/36) cells, which normally grow at 28 degrees C, were maintained at a supraoptimal temperature of 37 degrees C. The effect of continuous heat stress (37 degrees C) on cell growth was analyzed as were the modifications occurring with protein synthesis during short- and long-term heat stress. We observed that cells in lag or exponential growth phase, present inhibition of cell growth, and cells in the lag phase showed more sensitivity to death than cells growing exponentially. During the first hour of exposing the cells to 37 degrees C, they synthesized two heat shock proteins (hsps) of 82 kd and 70 kd, respectively, concomitant with inhibition of normally produced proteins at control temperature (28 degrees C). However, for incubations longer than 2 hr at 37 degrees C, a shift to the normal pattern of protein synthesis occurred. During these transitions, two other hsps of 76 kd and 90 kd were synthesized. Pulse chase experiments showed that the 70-kd hsp is stable at least for 18 hr, when the cells are returned to 28 degrees C. However, if cells were incubated at 37 degrees C, the 70-kd hsp is stable for at least 48 hr. The 70-kd hsp was localized in the cytoplasmic and in the nuclear compartment. Our results indicate a possible role of hsp 70-kd protein in the regulation of cell proliferation.     

Characteristics of in vivo isolation of free fatty acids by prokaryotic and eukaryotic algae under elevated and reduced temperature]

The paper describes the composition of extracellular free fatty acids (FFAs) and intracellular fatty acids (FAs) in the enrichment cultures of the prokaryotic alga Spirulina platensis and the eukaryotic alga Chlorella vulgaris grown at optimal, supraoptimal, and suboptimal growth temperatures. With increasing growth temperature, the degree of unsaturation of the intracellular FAs of both algae decreased, while that of the extracellular FFAs of S. platensis increased. The composition of the extracellular FFAs of C. vulgaris practically did not depend on the growth temperature.     

Submicroscopic morphology of trichophyton mentagrophytes grown at different temperatures

Several modifications were observed in Trichophyton mentagrophytes cultivated at 19° and 37 °C, i.e. nine degrees below and above the optimum of 28 °C. The phenomena included inhibition of the growth rate, changes in the gross aspects of the cultures as well as of the microscopic and submicroscopic morphology of the hyphal cells. At the ultrastructural level, in particular, it was shown that, at the suboptimal temperature, although the organelle structure in both young and aged hyphal cells remained nearly unchanged, unusual bodies of probable storage significance and plasmalemmasomes were formed.At the supraoptimal temperature, the youngest cells showed a normal organization but were richer in glycogen clusters and enveloped by a cell wall thicker than the ones at the optimal condition. In the cells far from the apex, the endomembrane integrity was lost and consequently an autolytic activity occurred. Degradation phenomena were detectable also at cell wall level. The cytological changes observed were tentatively correlated with a possible different sensitivity of the membrane system at the experimented temperature conditions.Investigation supported by a grant from Consiglio Nazionale delle Ricerche of Italy (Contract No. 7500536).