Induce Heat Shock Protein Synthesis Research Articles

Synthesis and approbation of new neuroprotective chemicals of pyrrolyl- and indolylazine classes in a cell model of Alzheimer's disease.

One of the major causes of neurodegeneration in the pathogenesis of Alzheimer's disease is the accumulation of cytotoxic amyloid species within the intercellular compartments of the brain. The efficacy of the anti-proteotoxic mechanism based on the molecular chaperones Hsp70 and Hsp90 in numerous types of neurons is often low, while its pharmacological enhancement has been shown to ameliorate the physiological and cognitive functions of the brain. Suggesting that the chemicals able to induce heat shock protein synthesis and therefore rescue neural cells from cytotoxicity associated with amyloid, we have synthesized a group of pyrrolyl- and indolylazines that cause the accumulation of heat shock proteins, using a novel method of photocatalysis that is employed in green chemistry. The selected compounds were tested in a cell model of Alzheimer's disease and demonstrated a pronounced neuroprotective effect. These substances increased the survival of neurons, blocked the activation of β-galactosidase, and prevented apoptosis in neurons cultured in the presence of β-amyloid.

Mild heat induces a distinct \u201ceustress\u201d response in Chinese Hamster Ovary cells but does not induce heat shock protein synthesis

The current research on cellular heat stress management focuses on the roles of heat shock proteins (HSPs) and the proteostasis network under severe stress conditions. The mild, fever-type stress and the maintenance of membrane homeostasis are less well understood. Herein, we characterized the acute effect of mild, fever-range heat shock on membrane organization, and HSP synthesis and localization in two mammalian cell lines, to delineate the role of membranes in the sensing and adaptation to heat. A multidisciplinary approach combining ultrasensitive fluorescence microscopy and lipidomics revealed the molecular details of novel cellular “eustress”, when cells adapt to mild heat by maintaining membrane homeostasis, activating lipid remodeling, and redistributing chaperone proteins. Notably, this leads to acquired thermotolerance in the complete absence of the induction of HSPs. At higher temperatures, additional defense mechanisms are activated, including elevated expression of molecular chaperones, contributing to an extended stress memory and acquired thermotolerance.

Decreased levels of heat shock proteins in gut epithelial cells after exposure to plant lectins

BACKGROUNDThe enterocytes of the intestinal epithelium are regularly exposed to potentially harmful substances of dietary origin, such as lectins. Expression of heat shock proteins (HSPs) by this epithelium may be...

Chemical Stress does not Induce Heat Shock Protein Synthesis in Leishmania donovani

The cellular heat shock response in kinetoplastid protozoa is regulated exclusively at a post-transcriptional level. The heat-inducibility of heat shock protein synthesis is retained under actinomycin C(1) which indicates an inducible translation of heat shock mRNAs. We have also assessed the ability of various chemicals known to be effective triggers of the heat shock response in higher eukaryotes to induce heat shock protein synthesis in Leishmania donovani. None of the tested chemicals elicited a stress response. We propose that the lack of transcription regulation in the kinetoplastida precludes a stress response under chemical stress.

Basal Synthesis of Heat Shock Protein 70 Increases with Age in Rat Kidneys

The heat shock protein (HSP) system is a mechanism of cell defense induced by stress, constitutively expressed during basal conditions and essential to the maintenance of cellular integrity. Acutely induced HSP synthesis decreases with aging, but the effect of age on the basal expression of HSP70 has not been specifically addressed so far. The aim of this work is to study the age-dependent basal concentrations of HSP70 mRNA in rat kidneys. In 8 young (2–3 months), 6 adult (6–11 months) and 6 old male Wistar rats (22–27 months), steady-state concentrations of HSP70 and γ-actin mRNA and of rRNA were measured. Pentosidine was measured by HPLC. The basal, unstimulated HSP70 mRNA is increased in young and old rats compared with adult subjects [young: 182% of adult levels (100–299), old: 167% of adults (142–209); p < 0.005]. The amount of pentosidine increases with age (young: 0.6 ± 0.1; adult: 1.65 ± 0.15; old: 2.3 ± 0.3 pmol/mg of protein; p < 0.0001). It seems likely that different mechanisms are responsible for the increased HSP70 basal synthesis in both the young and old animals. The prevalence of anabolic activity can trigger the increased basal production of HSP70 in young rats. The accumulation of posttranslational modified proteins, documented by pentosidine, can chronically enhance HSP70 synthesis in aged animals. The suppression of the synthesis of other proteins accompanying HSP-selective production might contribute to the impairment of specific cell functions in aging.

Heat shock protein induction by certain chemical stressors is correlated with their cytotoxicity, lipophilicity and protein-denaturing capacity

Seven agents were analyzed with respect to their ability to induce heat shock protein (HSP) synthesis in C6 rat glioma cells. Induction of HSP synthesis was correlated with cytotoxicity and lipophilicity of the substances. In addition to the first four n-alcohols (methanol, ethanol, propanol and butanol) and phenol, whose capacity to induce HSP was analyzed earlier (Neuhaus-Steinmetz et al., 1994. Mol. Pharmacol. 45, 36–41), isopropanol, 1,4-dinitrophenol (DNP), diethylstilbestrol (DES), carbonylcyanide- m-chlorophenylhydrazone (CCCP), rotenone, paracetamol and acetyl salicylic acid (ASA) induced HSP synthesis after a 1-h incubation at a substance-specific concentration. The maximal induction of HSPs was closely correlated with the cytotoxicity of all substances and occurred when cell viability was reduced to 75±11% of the controls. Cytotoxicity and the ability to induce HSP were correlated with the lipophilicity of the alcohols, phenol, rotenone and paracetamol. Calculation of the hypothetical membrane concentrations of these compounds yielded a nearly equal value (0.54±0.13 M), indicating that interaction of substances with lipophilic cellular compounds, such as membranes or lipophilic core regions of proteins, is a critical step leading to HSP induction. This assumption is supported by a correlation between HSP induction and protein denaturation by the different alcohols (Herskovits et al., 1970. J. Biol. Chem. 245, 2588–2598). We assume that the amount of misfolded proteins induced by these lipophilic agents is responsible for the induction of HSP synthesis. ASA, DNP and CCCP induced HSP at lower concentrations than substances with a similar lipophilicity, which may be due to effects which add to the misfolding of proteins or to other signal pathways.

Helium-neon laser irradiation is not a stressful treatment: a study on heat-shock protein (HSP70) level.

Helium-neon (He-Ne) laser irradiation has been clinically used to reduce chemotherapy-induced mucositis. This work was designed to find out if this treatment is stressful at the cellular level by studying its effects on the level of the stress-inducible heat shock proteins. Human desmodontal and mouse L929 fibroblasts were irradiated using a 60 mW laser by a single application of 1.5 and 3J/cm2 in continuous mode. Heat shock protein level was studied by gel electrophoresis and Western blotting using monoclonal antibodies. He-Ne treatment does not induce heat shock protein synthesis in human desmodontal nor in mouse fibroblasts at the energy densities used in this study. These results indicate that the treatment is not stressful at the cellular level.

Heat shock protein synthesis is affected by intracellular pH: inhibition by monensin-induced alkalosis in C6 rat glioma cells

The effect of intracellular pH (pH;) on heat shock protein (HSP) synthesis was investigated in C6 rat glioma cells. pH i changes were analysed by means of fluorescence spectroscopy in a perfused monitoring system allowing continuous measurements before, during and after treatments. HSP induction was determined by means of Western blots and autoradiographs. A 20 min heat shock (HS) of 44°C decreased the pH i from 7.36 to 7.05 during exposure [171 and elicited the synthesis of heat shock proteins 2–8 h later. A pH i decrease, brought about by low extracellular pH (pH e) of 4.5 and 5.0 or 5.5, induced HSP synthesis after 1 h or 3 h, respectively. During these treatments, pH i decreased to values significantly lower than that caused by HS. Three h exposure to pH e 6.2, however, was not inductive. These results indicate that the heat shock-induced pH i decrease alone is not sufficient to stimulate HSP synthesis. In order to investigate the effect of alkaline pH i on the induction of HSP by heat, pH i was increased prior to HS treatments. Preincubation of cells at pH e ranging from 6.8 to 8.0 had little effect on pH i and on HSP synthesis. A shift of pH i to more alkaline values was achieved by adding the H +/Na + exchanger monensin at alkaline pH e. Twenty μM monensin raised the pH i and inhibited the HSP induction depending on the pH e values: as pH e was increased from pH 7.2 to 8.0 HSP synthesis was increasingly inhibited. Monensin also diminished the HS-induced drop of pH i particularly at higher pH e. The result showed that neither a lower pH i nor a drop of pH i during HS is a necessary prerequisite for the induction, whereas alkalosis inhibits the synthesis of HSP.

Physiological and pharmacological inhibitors of luteinizing hormone- dependent steroidogenesis induce heat shock protein-70 in rat luteal cells

Heat shock protein (HSP) synthesis increases in cells with a broad range of stress conditions. We recently showed that induction of HSP-70 is associated with inhibition of hormone-sensitive steroidogenesis, but not hormone-sensitive cAMP accumulation, in rat luteal cells by a mechanism associated with interruption of cholesterol translocation in mitochondria. As HSP induction may be an early mediator of luteal regression, we investigated whether physiological and pharmacological inhibitors of luteal function would induce HSP-70 in rat luteal cells. Both [35S]methionine labeling and Western blotting with antibodies against the inducible form of HSP-70 revealed HSP induction in rat luteal cells by 1 microM prostaglandin F2 alpha (PGF2 alpha) coincident with inhibition of progesterone synthesis. In contrast, PGE2 (1 microM) failed to increase HSP-70 synthesis. Phorbol 12-myristate 13 acetate (3 microM), tumor necrosis factor-alpha (100 ng/ml), and ionomycin (1 microM) also induced HSP synthesis. Induction of HSP-70 was preceded by the rapid activation of heat shock transcription factor, which binds to the heat shock transcriptional control element. Gel retardation assays demonstrated heat shock transcription factor activation within 15 min of PGF2 alpha treatment. Northern analysis with an oligonucleotide probe specific for inducible HSP-70 showed induction at the transcriptional level by the above agents within 30 min. As functional luteal regression is known to display elements of a stress response, the finding that a number of factors that inhibit hormone-sensitive progesterone synthesis rapidly activate the heat shock response further implicates HSPs as possible mediators of luteolysis.

Physiological and pharmacological inhibitors of luteinizing hormone-dependent steroidogenesis induce heat shock protein-70 in rat luteal cells.

Heat shock protein (HSP) synthesis increases in cells with a broad range of stress conditions. We recently showed that induction of HSP-70 is associated with inhibition of hormone-sensitive steroidogenesis, but not hormone-sensitive cAMP accumulation, in rat luteal cells by a mechanism associated with interruption of cholesterol translocation in mitochondria. As HSP induction may be an early mediator of luteal regression, we investigated whether physiological and pharmacological inhibitors of luteal function would induce HSP-70 in rat luteal cells. Both [35S]methionine labeling and Western blotting with antibodies against the inducible form of HSP-70 revealed HSP induction in rat luteal cells by 1 microM prostaglandin F2 alpha (PGF2 alpha) coincident with inhibition of progesterone synthesis. In contrast, PGE2 (1 microM) failed to increase HSP-70 synthesis. Phorbol 12-myristate 13 acetate (3 microM), tumor necrosis factor-alpha (100 ng/ml), and ionomycin (1 microM) also induced HSP synthesis. Induction of HSP-70 was preceded by the rapid activation of heat shock transcription factor, which binds to the heat shock transcriptional control element. Gel retardation assays demonstrated heat shock transcription factor activation within 15 min of PGF2 alpha treatment. Northern analysis with an oligonucleotide probe specific for inducible HSP-70 showed induction at the transcriptional level by the above agents within 30 min. As functional luteal regression is known to display elements of a stress response, the finding that a number of factors that inhibit hormone-sensitive progesterone synthesis rapidly activate the heat shock response further implicates HSPs as possible mediators of luteolysis.

Altered regulation of heat shock gene expression in heat resistant mouse cells

Purpose : The differences in the heat shock gene regulation between the RIF-1 cell line and its heat resistant derivative TR4 are further characterized. Methods and Materials : In vitro gel retardation assays were used to assess the presence of activated heat shock transcription factor in the two cell lines. The levels of the heat-inducible HSP 70.1, the constitutive HSC 70, the germ line-specific HSP 70.2, and the HSP 28 mRNAs in both untreated and iso-heated RIF-1 and TR4 cells were determined using the polymerase chain reaction coupled with the reverse transcriptase reaction. Induction and decay of induced heat shock protein synthesis was measured by 35S-methionine labeling of proteins. Results : Unheated TR4 cells display characteristics of heat shocked RIF-1 cells. TR4 cells have a constitutively activated heat shock transcription factor and elevated levels of the HSP 70.1, HSC 70, and the HSP 28 mRNAs. Upon an equal heat dose of 45 C, 15 min, the TR4 cells exhibited a more rapid onset in heat shock mRNA and protein induction than did the RIF-1 cells. During the recovery from heat shock, the activated heat shock transcription factor and the induced HSP70 mRNAs decayed more slowly in the TR4 cells, although the protein synthesis pattern of the TR4 cells returned to control levels more rapidly following heat shock than did protein synthesis of the RIF-1 cells. Conclusion : Unheated TR4 cells are similar to heat shocked RIF-1 cells at the transcriptional level. Induced HSP70 expression is modulated by the severity of the heat treatment (or the degree of heat damage) perceived by the cells rather than by the absolute heat dose given. We propose that the unheated TR4 cells are locked into the “ON” state of the heat shock response.

Rhizobium meliloti suhR suppresses the phenotype of an Escherichia coli RNA polymerase sigma 32 mutant

sigma 32, the product of the Escherichia coli rpoH locus, is an alternative RNA polymerase sigma factor utilized to express heat shock genes upon a sudden rise in temperature. E. coli K165 [rpoH165(Am) supC(Ts)] is temperature sensitive for growth and does not induce heat shock protein synthesis. We have isolated a locus from Rhizobium meliloti called suhR that allows E. coli K165 to grow at high temperature and induce heat shock protein synthesis. R. meliloti suhR mutants were viable and symbiotically effective. suhR was found to have no DNA or derived amino acid sequence similarity to the genes of previously sequenced sigma factors or other data base entries, although a helix-turn-helix DNA-binding protein motif is present. suhR did not restore the phenotypic defects of delta rpoH E. coli; suppression of the E. coli K165 phenotype is thus likely to involve E. coli sigma 32. Western immunoblots showed that suhR caused an approximately twofold elevation of sigma 32 levels in K165; RNA blots indicated that rpoH mRNA level and stability were not altered. Stabilization of sigma 32 protein and increased rpoH mRNA translation are thus the most probable mechanisms of suppression.

Expression of the Heat Shock Response in a Tomato Interspecific Hybrid Is Not Intermediate between the Two Parental Responses

While it is apparent that the heat shock response is ubiquitous, variabilities in the nature of the heat shock response between closely related species have not been well characterized. The heat shock response of three genotypes of tomato, Lycopersicon esculentum, Lycopersicon pennellii, and the interspecific sexual hybrid was characterized. The two parental genotypes differed in the nature of the heat shock proteins synthesized; the speciesspecific heat shock proteins were identified following in vivo labeling of leaf tissue with [(35)S]methionine and cysteine. The duration of, and recovery from, heat shock varied between the two species: L. esculentum tissue recovered more rapidly and protein synthesis persisted longer during a heat shock than in the wild species, L. pennellii. Both species induced heat shock protein synthesis at 35 degrees C and synthesis was maximal at 37 degrees C. The response of the F1 to heat shock was intermediate to the parental responses for duration of, and recovery from, heat shock. In other aspects, the response of the F1 to heat shock was not intermediate to the parental responses: the F1 induced only half of the L. esculentum specific heat shock proteins, and all of the L. pennellii specific heat shock proteins. A discussion of the inheritance of the regulation of the heat shock response is presented.

Heat shock mRNA accumulation during recovery from cold shock in Drosophila melanogaster

Heat shock protein synthesis is induced in response to a variety of chemical and physical stresses. Among these are heating above normal growing temperatures, treatment with heavy metals, amino acid analogues, steroid hormones and a variety of other chemicals ( CRC Crit. Rev. Biochem. 18, 239–280). We have shown previously that heat shock proteins are also synthesized during recovery from prolonged 0°C treatment in Drosophila larval salivary glands. In this paper we describe the cold treatments which induce heat shock protein synthesis in more detail, and show that heat shock mRNA does not accumulate during the cold treatment, but rather during the recovery period when the larvae are returned to 25°C. The implications of these results for the regulation of heat shock mRNA levels, and for the role of heat shock proteins in recovery from cold shock are discussed.

Induction of a heat shock-like response by unfolded protein in Escherichia coli: dependence on protein level not protein degradation.

To test the idea that unfolded protein might act as an intracellular signal for induction of the heat shock response in Escherichia coli, we examined the synthesis of several heat shock proteins after expression of an unfolded variant of the amino-terminal domain of lambda repressor. These experiments show that expression of a single mutant protein, and not its wild-type counterpart, is sufficient to induce a heat shock-like response. In addition, by measuring the abilities of unfolded variants of differing proteolytic susceptibilities to induce heat shock protein synthesis and by monitoring heat shock protein synthesis as a function of the amount of a single unfolded protein, we show that it is the concentration of unfolded protein in the cell, and not its degradation, that is important for inducing the heat shock-like response.

Oxidative Stress and Heat Shock Protein Induction in Human Cells

Agents which induce heat shock protein synthesis in cultured monolayers of Hela cells such as hyperthermia, ethanol and sodium arsenite can also cause increases in the levels of lipid peroxidation as determined by the formation of TBA-products. The heat induced increases may be diminished by addition to the medium of mannitol or EGTA. These compounds are known to depress heat shock protein synthesis. Following hyperthermia there is also a decrease in protein synthesis. In vitro studies indicate possible damage to ribosomes, and since the heat induced loss of protein synthetic capacity can be increased by superoxide dismutase inhibitors, and prevented by mannitol, such effects may be linked to the increases observed in lipid peroxidation. It is suggested that a connection exists between lipid peroxidation and heat shock protein gene activation.

Heat shock protects germinating conidiospores of Neurospora crassa against freezing injury.

Germinating conidiospores of Neurospora crassa that were exposed to 45 degrees C, a temperature that induces a heat shock response, were protected from injury caused by freezing in liquid nitrogen and subsequent thawing at 0 degrees C. Whereas up to 90% of the control spores were killed by this freezing and slow thawing, a prior heat shock increased cell survival four- to fivefold. Survival was determined by three assays: the extent of spore germination in liquid medium, the number of colonies that grew on solid medium, and dry-weight accumulation during exponential growth in liquid culture. The heat shock-induced protection against freezing injury was transient. Spores transferred to normal growth temperature after exposure to heat shock and before freezing lost the heat shock-induced protection within 30 min. Spores subjected to freezing and thawing stress synthesized small amounts of the heat shock proteins that are synthesized in large quantities by cells exposed to 45 degrees C. Pulse-labeling studies demonstrated that neither chilling the spores to 10 degrees C or 0 degrees C in the absence of freezing nor warming the spores from 0 degrees C to 30 degrees C induced heat shock protein synthesis. The presence of the protein synthesis inhibitor cycloheximide during spore exposure to 45 degrees C did not abolish the protection against freezing injury induced by heat shock. Treatment of the cells with cycloheximide before freezing, without exposure to heat shock, itself increased spore survival.

Induction of thermotolerance and enhanced heat shock protein synthesis in chinese hamster fibroblasts by sodium arsenite and by ethanol

Synthesis of a family of proteins called "heat shock" proteins is enhanced in cells in response to a wide variety of environmental stresses. This suggests that these proteins may have functions essential to cell survival under stressful conditions. A causative relationship between heat shock protein synthesis and development of thermotolerance would imply that agents known to induce heat shock protein synthesis, such as sodium arsenite, also induce thermotolerance. Conversely, agents known to induce thermotolerance, such as ethanol, would also enhance heat shock protein synthesis. To test this hypothesis, I have examined the effect of sodium arsenite or ethanol treatment on protein synthesis and cell survival in Chinese hamster ovary HA-1 cells. After either sodium arsenite or ethanol treatment, the synthesis of heat shock proteins was greatly enhanced over that of untreated cells. In parallel, cell survival was increased as much as 10(4)-fold when cells exposed to either agent were challenged by a subsequent heat treatment. The synthesis of heat shock proteins correlated well with the development of thermotolerance. A qualitative analysis of individual proteins suggests that the synthesis of 70,000 and 87,000 molecular weight proteins most closely mirrored the development of thermotolerance. The results, therefore, strongly reinforce the hypothesis that a causal relationship exists between the enhanced synthesis of heat shock protein and cell survival under specific stresses.