Heat shock response in mycoplasmas, genome-limited organisms.

We have measured the effect of heat shock on three mycoplasmas (Acholeplasma laidlawii K2 and JA1 and Mycoplasma capricolum Kid) and demonstrated the induction of mycoplasma heat shock proteins under these conditions. Increased synthesis of at least 5 heat shock proteins in A. laidlawii K2, 11 heat shock proteins in A. laidlawii JA1, and 7 heat shock proteins in M. capricolum was observed by electrophoretic analysis of proteins from heat-shocked cells in sodium dodecyl sulfate-polyacrylamide gels. In all three strains, major heat shock proteins (66 to 68 and 26 to 29 kilodaltons [kDa]) were found. The 66- to 68-kDa protein cross-reacted with antibody to Escherichia coli DnaK protein, suggesting that this heat shock protein has been conserved in spite of major reductions in genetic complexity during mycoplasma evolution. A. laidlawii also contained a 60-kDa protein that cross-reacted with eubacterial GroEL protein and a 40-kDa protein that cross-reacted with E. coli RecA protein. Unlike with coliphages, the mycoplasma virus L2 progeny yield was not increased when virus was plated on heat-shocked A. laidlawii host cells. However, UV-irradiated L2 virus could be host cell reactivated by both A. laidlawii SOS repair and heat shock systems.     

Carotenogenic response in photosynthetic organisms: a colorful story

Carotenoids are a diverse group of terpenoid pigments ubiquitous in and essential for functioning of phototrophs. Most of the researchers in the field are focused on the primary carotenoids serving light harvesting, photoprotection, and supporting the structural integrity of the photosynthetic apparatus (PSA) within the thylakoid membranes. A distinct group of the pigments functionally and structurally uncoupled from the PSA and accumulating outside of the thylakoids is called secondary carotenoids. Induction of the biosynthesis and massive accumulation of the latter termed as secondary carotenogenesis and carotenogenic response (CR), respectively, is a major though insufficiently studied stress response discovered in many phototrophic organisms ranging from single-celled algae to terrestrial higher plants. The CR protects cell by means of optical shielding of cell structures vulnerable photodamage, consumption of potentially harmful dioxygen, augmenting sink capacity of photoassimilates, and exerting an antioxidant effect. The secondary carotenoids exhibit a remarkable photostability in situ. Therefore, the CR-based photoprotective mechanism, unlike, e.g., antioxidant enzyme-based protection in the chloroplast, does not require continuous investment of energy and metabolites making it highly suitable for long-term stress acclimation in phototrophs. Capability of the CR determines the strategy of acclimation of photosynthetic organisms to different stresses such as excessive irradiance, drought, extreme temperatures, and salinities. Build-up of the CR might be accompanied by gradual disengagement of ‘classical’ active (energy-dependent) photoprotective mechanisms such as non-photochemical quenching. In addition to that, the CR has great ecological significance. Illustrious examples of this are extremely stress-tolerant ‘snow’ algae and conifer species developing red coloration during winter. The CR has also considerable practical implications since the secondary carotenoids exert a plethora of beneficial effects on human and animal health. The carotenogenic microalgae are the richest biotechnological sources of natural value-added carotenoids such as astaxanthin and β-carotene. In the present review, we summarize current functional, mechanistic, and ecological insights into the CR in a broad range of organisms suggesting that it is obviously more widespread and important stress response than it is currently thought to be.     

Quantitative imaging of membrane lipid order in cells and organisms

It is now recognized that lipids and proteins in cellular membranes are not homogenously distributed. A high degree of membrane order is the biophysical hallmark of cholesterol-enriched lipid rafts, which may induce the lateral sorting of proteins within the membrane. Here we describe a quantitative fluorescence microscopy technique for imaging localized lipid environments and measuring membrane lipid order in live and fixed cells, as well as in intact tissues. The method is based on the spectral ratiometric imaging of the polarity-sensitive membrane dyes Laurdan and di-4-ANEPPDHQ. Laurdan typically requires multiphoton excitation, making it suitable for the imaging of tissues such as whole, living zebrafish embryos, whereas di-4-ANEPPDHQ imaging can be achieved with standard confocal microscopes. This approach, which takes around 4 h, directly examines the organization of cellular membranes and is distinct from alternative approaches that infer membrane order by measuring probe partitioning or dynamics.     

Heat shock response in Lactobacillus plantarum

Heat stress resistance and response were studied in strains of Lactobacillus plantarum. Stationary-phase cells of L. plantarum DPC2739 had decimal reduction times (D values) (D value was the time that it took to reduce the number of cells by 1 log cycle) in sterile milk of 32.9, 14.7, and 7.14 s at 60, 72, and 75 degrees C, respectively. When mid-exponential-phase cells were used, the D values decreased. The temperature increases which caused a 10-fold reduction in the D value ranged from 9 to 20 degrees C, depending on the strain. Part of the cell population treated at 72 degrees C for 90 s recovered viability during incubation at 7 degrees C in sterile milk for 20 days. When mid-exponential- or stationary-phase cells of L. plantarum DPC2739 were adapted to 42 degrees C for 1 h, the heat resistance at 72 degrees C for 90 s increased ca. 3 and 2 log cycles, respectively. Heat-adapted cells also showed increased growth at pH 5 and in the presence of 6% NaCl. Two-dimensional gel electrophoresis of proteins expressed by control and heat-adapted cells revealed changes in the levels of expression of 31 and 18 proteins in mid-exponential- and stationary-phase cells, respectively. Twelve proteins were commonly induced. Nine proteins induced in the heat-adapted mid-exponential- and/or stationary-phase cells of L. plantarum DPC2739 were subjected to N-terminal sequencing. These proteins were identified as DnaK, GroEL, trigger factor, ribosomal proteins L1, L11, L31, and S6, DNA-binding protein II HlbA, and CspC. All of these proteins have been found to play a role in the mechanisms of stress adaptation in other bacteria. Antibodies against GroES detected a protein which was induced moderately, while antibodies against DnaJ and GrpE reacted with proteins whose level of expression did not vary after heat adaptation. This study showed that the heat resistance of L. plantarum is a complex process involving proteins with various roles in cell physiology, including chaperone activity, ribosome stability, stringent response mediation, temperature sensing, and control of ribosomal function. The physiological mechanisms of response to pasteurization in L. plantarum are fundamental for survival in cheese during manufacture.     

Heat shock response in Actinobacillus actinomycetemcomitans

Abstract The heat shock response in Actinobacillus actinomycetemcomitans , a capnophilic Gram-negative bacterial species that is implicated in the development of certain forms of periodontitis, was characterized. Different strains of A. actinomycetemcomitans were grown at 37, 42 and 48°C in the presence of ³⁵S-methionine. The bacterial cells were lysed, run on SDS-PAGE and subsequently blotted on nitrocellulose paper. After autoradiography of the blots, several protein bands from the cultures at 42°C showed an increased intensity; major bands were observed at 90, 70, and 60 kDa, but increased protein synthesis was also detected at 54, 28 and 17 kDa. Nitrocellulose blots were also incubated with a panel of monoclonal and polyclonal antibodies directed to epitopes on different heat shock proteins. Strong reactivity was found with several antibodies at the position corresponding to a molecular mass of 60 kDa. The protein is probably the GroEL homologue in A. actinomycetemcomitans , a member of the ‘common bacterial antigen’ family.     

Heat shock response and ageing: mechanisms and applications

Ageing is associated with a decrease in the ability of cells to cope with environmental challenges. This is due partly to the attenuation of a primordial stress response, the so-called heat shock (HS) response, which induces the expression of heat shock proteins (HSPs), composed of chaperones and proteases. The attenuation of the HS response during ageing may be responsible for the accumulation of damaged proteins as well as abnormal regulation of cell death. Maintenance of the HS response by repeated mild heat stress causes anti-ageing hormetic effects on cells and organisms. Here, we describe the molecular mechanism and the state of the HS response as well as the role of specific HSPs during ageing, and discuss the possibility of hormetic modulation of ageing and longevity by repeated mild stress.     

Addition of lipid to the photosynthetic membrane: effects on membrane structure and energy transfer

We have carried out a series of experiments in which the lipid composition of the photosynthetic membrane has been altered by the addition of lipid from a defined source under experimental conditions. Liposomes prepared by sonication are mixed with purified photosynthetic membranes obtained from spinach chloroplasts and are taken through cycles of freezing and thawing. Several lines of evidence, including gel electrophoresis and freeze-fracture electron microscopy, indicate that an actual addition of lipid has taken place. Structural analysis by freeze-fracture shows that intramembrane particles are widely separated after the addition of large amounts of lipid, with one exception: large hexagonal lattices of particles appear in some regions of the membrane. These lattices are identical in appearance with lattices formed from a single purified component of the membrane known as chlorophyll-protein complex II. The suggestion that the presence of such lattices in lipid-enriched membranes reflects a profound rearrangement of photosynthetic structures has been confirmed by analysis of the fluorescence emission spectra of natural and lipid- enriched membranes. Specifically, lipid addition in each of the cases we have studied results in the apparent detachment of chlorophyll- protein complex II from photosynthetic reaction centers. It is concluded that specific arrangements of components in the photosynthetic membrane, necessary for the normal functioning of the membrane in the light reaction of photosynthesis, can be regulated to a large extent by the lipid content of the membrane.     

Heat shock response of Dictyostelium

In response to a shift from 22 to 30°C the relative rate of synthesis of a small number of proteins is dramatically increased in Dictyostelium discoideum. The cells neither grow nor develop at this temperature but die slowly with a half-life of 18 hr. The major protein synthesized in response to a heat shock to 30°C in either growing cells or developing cells has an apparent molecular weight of 70,000 (70K). An increase in the relative rate of synthesis of 70K can be seen as early as 20 min following heat shock. Synthesis of 70K remains high for 4 hr at 30°C and then decreases. Similar kinetics of 70K synthesis occur during recovery at 22°C following a 1-hr heat shock. RNA synthesis during the first half-hour of heat shock is essential for the high rate of 70K measured 2 hr later. By isoelectric focusing the 70K protein can be separated into two spots, one of which overlaps one of the major heat shock proteins of Drosophila melanogaster. The relative rate of synthesis of several other proteins (82K, 60K, 43K) increases less dramatically in Dictyostelium during heat shock at 30°C. A heat shock to 34°C results in rapid synthesis of these proteins but not of 70K. The relative rates of synthesis of most other proteins made at 22°C decreases, most notably that of actin. Synthesis of heat shock proteins at 30°C does not significantly affect viability at 30°C but dramatically prolongs the period of time the cells can survive at 34°C. Thus, 30°C appears to be a stasis condition for Dictyostelium which elicits a response essential for protection from lethal temperatures. The similarity of the heat shock response in Dictyostelium to that in Drosophila and vertebrate cells suggests that certain aspects of the response may be universal in eukaryotes.     

Heat shock response: hsp70 in environmental monitoring

Heat shock proteins (Hsps) are a ubiquitous feature of cells in which these proteins cope with stress-induced denaturation of other proteins. Among the different families of Hsps, the 70 kDa family (hsp70) is the most highly conserved and has been most extensively studied. Apart from their primary role in cellular defense under stress condition, a number of studies in recent years have shown the immense potential of hsp70 in pollution monitoring using even transgenic approach both in vivo and in vitro. This article reviews the recent developments in the widespread application of hsp70 in environmental risk assessment.     

Tocopherol functions in photosynthetic organisms

During the past decade, the genes required for tocopherol (vitamin E) synthesis in plants and cyanobacteria have been identified. A series of mutants in which specific pathway steps are disrupted have been generated, providing new insights into tocopherol functions in photosynthetic organisms. Tocopherols are essential for controlling non-enzymatic lipid peroxidation during seed dormancy and seedling germination. Their absence results in elevated levels of malondialdehyde and phytoprostanes, and in inappropriate activation of plant defense responses. Surprisingly, tocopherol deficiency in mature leaves has limited consequences under most abiotic stresses, including high intensity light stress. The cell wall development of phloem transfer cells under cold conditions is, however, severely impaired in mature leaves of tocopherol-deficient mutants, indicating that tocopherols are required for proper adaptation of phloem loading at low temperatures.     

Heat shock response and acute lung injury

All cells respond to stress through the activation of primitive, evolutionarily conserved genetic programs that maintain homeostasis and assure cell survival. Stress adaptation, which is known in the literature by a myriad of terms, including tolerance, desensitization, conditioning, and reprogramming, is a common paradigm found throughout nature, in which a primary exposure of a cell or organism to a stressful stimulus (e.g., heat) results in an adaptive response by which a second exposure to the same stimulus produces a minimal response. More interesting is the phenomenon of cross-tolerance, by which a primary exposure to a stressful stimulus results in an adaptive response whereby the cell or organism is resistant to a subsequent stress that is different from the initial stress (i.e., exposure to heat stress leading to resistance to oxidant stress). The heat shock response is one of the more commonly described examples of stress adaptation and is characterized by the rapid expression of a unique group of proteins collectively known as heat shock proteins (also commonly referred to as stress proteins). The expression of heat shock proteins is well described in both whole lungs and in specific lung cells from a variety of species and in response to a variety of stressors. More importantly, in vitro data, as well as data from various animal models of acute lung injury, demonstrate that heat shock proteins, especially Hsp27, Hsp32, Hsp60, and Hsp70 have an important cytoprotective role during lung inflammation and injury.     

Heat shock proteins: regulators of stress response and apoptosis

No Abstract Available     

Heat shock response of the rat lens

The sequence relationship between the small heat shock proteins and the eye lens protein alpha-crystallin (Ingolia, T. D., and E. E. Craig, 1982, Proc. Natl. Acad. Sci. USA, 79: 2360-2364) prompted us to subject rat lenses in organ culture to heat shock and other forms of stress. The effects on protein synthesis were followed by labeling with [35S]methionine and analysis by one- and two-dimensional gel electrophoresis and fluorography. Heat shock gave a pronounced induction of a protein that could be characterized as the stress protein SP71. This protein probably corresponds to the major mammalian heat shock protein hsp70. Also two minor proteins of 16 and 85 kD were induced, while the synthesis of a constitutive heat shock-related protein, P73, was considerably increased. The synthesis of SP71 started between 30 and 60 min after heat shock, reached its highest level after 3 h, and had stopped again after 8 h. In rat lenses that were preconditioned by an initial mild heat shock, a subsequent shock did not cause renewed synthesis of SP71. This effect resembles the thermotolerance phenomenon observed in cultured cells. The proline analogue azetidine-2-carboxylic acid, zinc chloride, ethanol, and calcium chloride did not, under the conditions used, induce stress proteins in the rat lens. Sodium arsenite, however, had very much the same effects as heat shock. Calcium ionophore A23187 specifically and effectively induced the synthesis of the glucose-regulated protein GRP78. No special response to stress on crystallin synthesis was noticed.     

Heat shock response ofChlorella protothecoides during greening

Heterotrophically grown cells ofChlorella protothecoides were transferred to autotrophic medium and allowed to green at 25°C. The protein synthetic activity of the greening cells measured in terms of incorporation of [³5S]-methionine showed a maximum around 20 h of greening and thereafter started declining. Similarly, an analysis of densitometric tracings of the fluorographic profile of the polypeptides associated with both total cellular fraction and membrane fractions during different hours of greening revealed that maximum number of polypeptides were getting labelled around 20 h of greening. At 20 h of greening, the cells were shifted to 40°C and the effect of heat shock on protein synthesis was studied. The heat shock treatment caused a definite decrease in the incorporation of [³⁵S]-methionine into proteins. Due to heat shock, the synthesis of total soluble proteins was affected much more than that of the thylakoid membrane bound proteins. When the cells were transferred back to 25°C after a brief period of heat shock at 40°C, there was a considerable recovery in the protein synthesis and this recovery was found to be significant in the case of soluble proteins, while there was no such definite recovery in the synthesis of thylakoid membrane bound proteins.