Interaction of a small heat shock protein with light-harvesting cyanobacterial phycocyanins under stress conditions

Phycobiliproteins such as phycocyanins are the most abundant proteins found in cyanobacteria which are assembled to form the phycobilisome. Here, we showed that a small heat shock protein, HspA, interacts directly with phycocyanins from the cyanobacterium Synechococcus sp. strain PCC 7942 in vitro and suppresses inactivation of their light-harvesting functions due to heat denaturation in the presence of hydrogen peroxide. Under the denaturing conditions, phycobilisomes were de-assembled to lighter complexes and then aggregated. HspA associated with phycocyanins in the dissociated complexes, and suppressed the aggregation. The specific interaction between a small heat shock protein and phycocyanins was further supported by the fact that HspA and alpha-crystallin protected isolated phycocyanins from denaturation, while HtpG and lysozyme did not. The maximum protection was observed at a molar ratio of four HspA monomer per one phycocyanin (alpha beta) monomer.     

A limited turbidostat yeast culture under heat shock conditions

A response of limited turbidostat S. cerevisiae 14 culture on rapid increase in temperature from optimal 30 degrees to supraoptimal 37.5 degrees C. The temporal thermotolerance was absent in glucose and phosphate-limited cultures. Limitation of nitrogen, Mg, betaalanine, biotin, and, to a certain extent, potassium did not decrease the thermotolerance. The pattern of changes in age composition and specific optical density was found to be similar in limited and unlimited cultures. The glucose and phosphate expenditures at 37.5 degrees C were estimated. The response to heat shock was found to depend on the nature of the limiting factor and the extent of limiting.     

Lipid metabolism in Aspergillus niger under conditions of heat shock

The processes of lipid synthesis and decomposition in Aspergillus niger under conditions of heat shock (HS) were studied in a pulse-chase experiment with ¹⁴C-labeled sodium acetate. HS (60 min) resulted in the synthesis of phospholipids and sphingolipids intensified compared to the control, as was evident from incorporation of the labeled substrate. The same pattern was observed for neutral lipids, especially for triacylglycerides, while incorporation of the label into sterols remained almost the same. Further cultivation for 3 h in the medium without the labeled substrate resulted in a significant decrease of the label content in the membrane lipids of both the control and the experiment, although under HS conditions this decrease was much more pronounced, especially for phosphatidylcholines and phosphatidylethanolamines. A threefold increase of the label content in phosphatidic acids was observed only under HS conditions. These results indicate more intense metabolism of the membrane lipids under heat shock and suggest the degradation of the major cell phospholipids as the factor responsible for the increased level of phosphatidic acids in A. niger mycelium.     

Accelerated protein aggregation induced by macrophage migration inhibitory factor under heat stress conditions

Kinetics of thermal aggregation of model protein substrates (glycogen phosphorylase b from rabbit skeletal muscle and yeast alcohol dehydrogenase) were investigated under heat stress conditions (41-48 degrees C) in the presence of macrophage migration inhibitory factor (MIF), a heat-stable hydrophobic protein (12.5 kD). Anti-chaperone MIF activity found by turbidimetry manifests itself in significantly accelerated protein aggregation and increased limiting value of apparent optical absorption at 360 nm and t --> infinity in the sub-stoichiometric range of MIF concentrations. The aggregation kinetics is shown to have cooperative character. Possible reversibility of aggregation after removal of denaturing conditions was demonstrated using alcohol dehydrogenase aggregation at a temperature close to the physiological level (41.5 degrees C). This reversibility is caused by solubility of aggregates and stabilization of oligomeric structure of the substrate as a result of MIF binding to the partially denatured protein. The data suggest that in spite of distinct anti-chaperone effect, the chaperone-like activity of MIF can be observed in the case of heat stress removal and restoration of the system to normal conditions.     

Cell cycle changes in transformed cells growing under serum-free conditions.

Polyoma transformed hamster cells (PyBHK) and SV40 transformed mouse cells (SV3T3) were transferred in culture using crystalline trypsin followed by neutralisation with soybean trypsin inhibitor. Such cells were able to proliferate freely in defined medium without any serum supplement and without any intervening period of adaptation. However, growth rates were reduced under serum-free conditions. Re-establishment of rapid growth rates could be achieved by addition of serum, with the rate attained being proportional to the serum concentration. Irrespective of the prevailing rates of growth, percentages of cells synthesising DNA were the same. However, the rate at which DNA was being synthesised was found to change proportionately with the changes in overall growth rate.     

Proteome analysis of the Escherichia coli heat shock response under steady-state conditions

In this study a proteomic approach was used to investigate the steady-state response of Escherichia coli to temperature up-shifts in a cascade of two continuously operated bioreactors. The first reactor served as cell source with optimal settings for microbial growth, while in the second chemostat the cells were exposed to elevated temperatures. By using this reactor configuration, which has not been reported to be used for the study of bacterial stress responses so far, it is possible to study temperature stress under well-defined, steady-state conditions. Specifically the effect on the cellular adaption to temperature stress using two-dimensional gel electrophoresis was examined and compared at the cultivation temperatures of 37°C and 47.5°C. As expected, the steady-state study with the double bioreactor configuration delivered a different protein spectrum compared to that obtained with standard batch experiments in shaking flasks and bioreactors. Setting a high cut-out spot-to-spot size ratio of 5, proteins involved in defence against oxygen stress, functional cell envelope proteins, chaperones and proteins involved in protein biosynthesis, the energy metabolism and the amino acid biosynthesis were found to be differently expressed at high cultivation temperatures. The results demonstrate the complexity of the stress response in a steady-state culture not reported elsewhere to date.     

Respiratory activity of yeast Yarrowia lipolytica under oxidative stress and heat shock

Heat shock (45 degrees C) and the effect of oxidants (H2O2) resulted in a decrease of the respiratory activity of yeast cells and their survival rate. Increased resistance to stress effects after mild heat treatment (37 degrees C) or treatment with a nonlethal dose of oxidants (0.5 mM H2O2 for 60 min) was accompanied by appearance of an alternative (cyanide-resistant) oxidative pathway in the mitochondria, which promotes survival due to retention of the capacity for ATP synthesis in the first coupling point at the level of endogenous NADH dehydrogenase. The alternative oxidative pathway is more resistant to the effect of stressors that disrupt electron transfer in the cytochrome site of the respiratory chain.     

Respiratory activity of yeast Yarrowia lipolytica under oxidative stress and heat shock

Heat shock (45°C) and the effect of oxidants (H₂O₂) resulted in a decrease of the respiratory activity of yeast cells and their survival rate. Increased resistance to stress effects after mild heat treatment (37°C) or treatment with a nonlethal dose of oxidants (0.5 mM H₂O₂) for 60 min) was accompanied by appearance of an alternative (cyanide-resistant) oxidative pathway in the mitochondria, which promotes survival due to retention of the capacity for ATP synthesis in the first coupling point at the level of endogenous NADH dehydrogenase. The alternative oxidative pathway is more resistant to the effect of stressors that disrupt electron transfer in the cytochrome site of the respiratory chain.     

Adaptive responses to fescue toxicosis under thermoneutral and heat stress conditions

This study determined the potential for short-term adaptation to fescue toxicosis and heat stress in rats. Male CD outbred rats (n=24) were implanted with temperature transmitters (Respironics, Bend, OR) to measure core temperature (Tc) and general activity. All rats were initially fed diets with ground, uninfected tall fescue seed (E−) and exposed to 21 °C (thermoneutral, TN) to establish baseline values. In Period 1, all groups were maintained at TN for 7 days, with one group fed a diet containing ground, endophyte-infected tall fescue seed (E+, approximately 165 μg ergovaline/kg BW/d) and two groups fed E− diet. Ergovaline is thought to be the primary toxin responsible for many symptoms associated with fescue toxicosis. Period 1 was followed by 7 days at 31 °C (heat stress, HS, Period 2) on the same diets. All animals were fed E− diet during the second 7 day of HS (Period 3). In the final 7 day (Period 4), E+ diet was returned to the original group and fed to one of the previously E− groups, with the third group remaining on E− diet. A 40% decrease in FI occurred with E+ treatment at TN (P<0.05), with a comparable BW reduction (P<0.05) after 4 day. Both responses worsened during HS. Treatment with E+ in Period 4 indicated that FI and BW had not adapted to fescue toxicosis. A reduction in daily Tc occurred with E+ treatment at TN (P<0.05) followed by hyperthermia during the initial stage of HS (P<0.05). Although feed intake and growth rate showed no change over time, there was a reduction in fescue toxicosis-induced hyperthermia in the heat with repeat treatment. Conditioning animals to fescue toxicosis and heat stress prior to exposure may be beneficial in reducing impacts on thermal status of the animal.     

Daily rumination time of lactating dairy cows under heat stress conditions

The dairy industry in regions with moderate climates, such as Central Europe, will be increasingly challenged in the future by climate change. The problem of heat stress will especially affect dairy husbandry in naturally ventilated barns (NVB). The approach of the study was to determine a heat stress threshold of the average daily temperature-humidity index (THI) that results in changes in the daily rumination time (RT) of lactating, high-yielding cows. The data set was composed of a high sample size of 183 cows and long-duration measurements of 21240 daily observations over two years from June 2015 to May 2017, which were collected in an NVB in Groβ Kreutz, Germany. The THI was calculated in 5-min intervals by data from several sensors in different positions inside the barn. Additionally, every cow from the herd of an average of 53 cows in the experimental procedure was wearing a neck collar with a Lely Qwes HR system that provided the RT 24 h a day (12 2-h recordings were summarized). The study showed that heat stress also negatively influenced RT in moderate climates. The heat stress threshold of 52 THI was determined by broken-stick regression and indicated changes of RT of lactating dairy cows in Germany. During the experimental period, the heat stress threshold for RT was reached from April to September for up to 720 h per month. The changes in RT to the heat stress threshold will be affected by cows' characteristics. Therefore, we considered several cow-related factors, such as milk yield (MY), lactation number (LN), lactation stage (days in milk, or DIM) and pregnancy stage (P) to better understand cows’ individual reactions to heat stress. Multiparous, high-yielding cows in later lactation stages are potentially more strongly affected than other cows.