Function Of Hsps Research Articles

Comprehensive genome‑wide analysis of the chicken heat shock protein family: identification, genomic organization, and expression profiles in indigenous chicken with highly pathogenic avian influenza infection

BackgroundHeat shock proteins (HSPs) function as molecular chaperones with critical roles in chicken embryogenesis, immune response to infectious diseases, and response to various environmental stresses. However, little is known on HSP genes in chicken. In this study, to understand the roles of chicken HSPs, we performed genome-wide identification, expression, and functional analyses of the HSP family genes in chicken.ResultsA total of 76 HSP genes were identified in the chicken genome, which were further classified into eight distinct groups (I-VIII) based on phylogenetic tree analysis. The gene-structure analysis revealed that the members of each clade had the same or similar exon-intron structures. Chromosome mapping suggested that HSP genes were widely dispersed across the chicken genome, except in chromosomes 16, 18, 22, 25, 26, and 28–32, which lacked chicken HSP genes. On the other hand, the interactions among chicken HSPs were limited, indicating that the remaining functions of HSPs could be investigated in chicken. Moreover, KEGG pathway analysis showed that the HSP gene family was involved in the regulation of heat stress, apoptotic, intracellular signaling, and immune response pathways. Finally, RNA sequencing data revealed that, of the 76 chicken HSP genes, 46 were differentially expressed at 21 different growth stages in chicken embryos, and 72 were differentially expressed on post-infection day 3 in two indigenous Ri chicken lines infected with highly pathogenic avian influenza.ConclusionsThis study provides significant insights into the potential functions of HSPs in chicken, including the regulation of apoptosis, heat stress, chaperone activity, intracellular signaling, and immune response to infectious diseases.

Transcriptome Analysis Reveals the Heat Stress Response Genes by Fire Stimulation in Michelia macclurei Dandy

Heat stress due to external heat sources such as fire is an ecological problem for plants. When forest plants suffer from fire, high temperatures cause an array of morphological, physiological, and biochemical changes, which affect growth and development. Michelia macclurei Dandy is an evergreen broad-leaved tree species with the characteristics of fast growth, strong adaptability, and good fire-resistance. Some studies have improved the understanding of how fire behavior affects physiology, function and mortality, but the extreme heat response genes and mechanisms need improved understanding. In this study, we conducted a fire experiment (slight and severe) and RNA-Seq in M. macclure. The de novo assembly obtained 104,052 unigenes, and 48.46% were annotated in at least one public database. Specifically, 4458 and 4810 differentially expressed genes (DEGs) were identified in slight and severe fire treatment groups, respectively. In two treatment groups, 612 unigenes were differentially expressed, which were enriched in ‘oxidoreductase activity’ in the molecular function (MF) category of Gene Ontology (GO) enrichment analysis, suggesting the core role of oxidoreductase activity in response to extremely high temperatures in M. macclurei. In KEGG enrichment analysis of DEGs, the ‘plant hormone signal transduction’ is overrepresented, suggesting that this process plays an important role during heat response in M. macclurei. In the pathways of cytokinine and salicylic acid, some vital DEGs were enriched, which were related to cell division, shoot initiation, and disease resistance, and the potential interactions during heat stress were discussed. Moreover, the DEGs linked to heat stress response were identified, including heat shock factors, stress enhanced protein, signal transduction, photosystem, and major transcription factors. The qRT-PCR examination of various tissues, expression dynamics, and treatments revealed that the genes coding for the heat shock protein HSF30, stress enhanced protein, and photosystem I reaction center subunit II exhibited particularities in leaf tissue. Genes coding for heat shock proteins displayed a distinct expression pattern between fire treatment and conventional heat stress, which could signify the distinctive function of HSPs and the mechanism of heat responses. Altogether, these may interact to respond to fire stress through alterations in cellular processes, signaling transduction, and the synthesis and degradation of response proteins in M. macclurei. The results of this study provide a crucial transcriptional profile influenced by heat stress in M. macclurei, and could be of great use to explore the fire prevention mechanisms of fire-resistant tree species.

Pyrazolyl‐Benzoxazinone Derivatives as Dual Hsp Inhibitors in Human Breast Cancer

AbstractHeat Shock Proteins (Hsps) play major role on the onset of several cancers. Metabolic rates of cancer cells are higher compared to that of untransformed cells. This accelerated rate force functional substrate proteins to fold faster than normal folding rate. Although, the process leads cell cycle halting and eventually induces apoptosis, Hsps help cell survival and inhibit apoptosis and fold substrate proteins especially signaling proteins. When cancer cells accelerate the metabolism for invasion and metastasis, substrate proteins must fold to their native state rapidly. Since, functional forms of the proteins must be folded properly, cancer cells overexpress Hsps to fold substrate proteins and avoid apoptosis. Hsp90 and Hsp70 play key role in these processes. Inhibition of either Hsp90 or Hsp70 display complementary function. Therefore, dual inhibition of Hsp70 and Hsp90 potentially provides anticancer affect. In silico studies showed that pyrazolyl‐benzoxazine derivatives display binding activity for both Hsps. For this purpose, pyrazole‐3‐carbonyl chloride were converted to pyrazolyl‐benzoxazine derivatives via reactions of anthranilic acids in good yields (68–83 %). The structures of the newly synthesized compounds were elucidated by IR‐NMR spectroscopy, elemental analysis, and single‐crystal X‐ray diffraction. Binding of the compounds inhibit function of Hsps and cause cytotoxic effect over MCF‐7 cells. The compounds display potential anticancer effects.

Genomic and Transcriptomic Landscape and Evolutionary Dynamics of Heat Shock Proteins in Spotted Sea Bass (Lateolabrax maculatus) under Salinity Change and Alkalinity Stress.

Simple SummaryHeat shock proteins (Hsps) are ubiquitous and conserved in almost all living organisms and are involved in a wide spectrum of cellular responses against diverse environmental stresses. However, our knowledge about the coordinated Hsp co-chaperon interaction is still limited, especially in aquatic animals facing dynamic water environments. In this study, we provided the systematic analysis of 95 Hsp genes (LmHsps) in spotted sea bass (Lateolabrax maculatus), an important aquaculture species in China, under salinity change and alkalinity stress through in silico analysis. The coordinated expression of LmHsps in response to salinity change and alkalinity stress in the gills was determined. Our results confirmed the diverse regulated expression of Hsps in L. maculatus, and that the responses to alkalinity stress may have arisen through the adaptive recruitment of LmHsp40-70-90 co-chaperons. Our results provide vital insights into the function and adaptation of aquatic animal Hsps in response to salinity-alkalinity stress.The heat shock protein (Hsp) superfamily has received accumulated attention because it is ubiquitous and conserved in almost all living organisms and is involved in a wide spectrum of cellular responses against diverse environmental stresses. However, our knowledge about the Hsp co-chaperon network is still limited in non-model organisms. In this study, we provided the systematic analysis of 95 Hsp genes (LmHsps) in the genome of spotted sea bass (Lateolabrax maculatus), an important aquaculture species in China that can widely adapt to diverse salinities from fresh to sea water, and moderately adapt to high alkaline water. Through in silico analysis using transcriptome and genome database, we determined the expression profiles of LmHsps in response to salinity change and alkalinity stress in L. maculatus gills. The results revealed that LmHsps were sensitive in response to alkalinity stress, and the LmHsp40-70-90 members were more actively regulated than other LmHsps and may also be coordinately interacted as co-chaperons. This was in accordance with the fact that members of LmHsp40, LmHsp70, and LmHsp90 evolved more rapidly in L. maculatus than other teleost lineages with positively selected sites detected in their functional domains. Our results revealed the diverse and cooperated regulation of LmHsps under alkaline stress, which may have arisen through the functional divergence and adaptive recruitment of the Hsp40-70-90 co-chaperons and will provide vital insights for the development of L. maculatus cultivation in alkaline water.

The HSP Immune Network in Cancer

Heat shock proteins are molecular chaperones which support tumor development by regulating various cellular processes including unfolded protein response, mitochondrial bioenergetics, apoptosis, autophagy, necroptosis, lipid metabolism, angiogenesis, cancer cell stemness, epithelial-mesenchymal transition and tumor immunity. Apart from their intracellular activities, HSPs have also distinct extracellular functions. However, the role that HSP chaperones play in the regulation of immune responses inside and outside the cell is not yet clear. Herein, we explore the intracellular and extracellular immunologic functions of HSPs in cancer. A broader understanding of how HSPs modulate immune responses may provide critical insights for the development of effective immunotherapies.

Characterization, and Functional Analysis of Hsp70 and Hsp90 Gene Families in Glyphodes pyloalis Walker (Lepidoptera: Pyralidae).

Glyphodes pyloalis Walker (Lepidoptera: Pyralidae) is a destructive mulberry pest, causing great damage to mulberry in China. Heat shock proteins (Hsps) are involved in various signal pathways and regulate lots of physiological processes in insects. The function of Hsps in G. pyloalis, however, has still received less attention. Here, we identified five Hsp genes from G. pyloalis transcriptome dataset including two Hsp70 family genes (GpHsp71.3 and GpHsp74.9) and three Hsp90 family genes (GpHsp82.4, GpHsp89, and GpHsp93.4). Quantitative Real-time PCR validation revealed that all Hsps of G. pyloalis have significant expression in pupal and diapause stage, at which the larvae arrest the development. Expressions of GpHsp71.3 and GpHsp82.4 were increased significantly after thermal treatment at 40°C, and this upregulation depended on heat treatment duration. Furthermore, silencing GpHsp82.4 by RNA interference led to a significant increase in mortality of G. pyloalis larvae under the heat stress compared to the control group. After starvation stress, the expression levels of GpHsp82.4 and GpHsp93.4 were significantly increased. At last, after being parasitized by the parasitoid wasp Aulacocentrum confusum, Hsp70 and Hsp90 genes of G. pyloalis were decreased significantly in the early stage of parasitization and this moderation was affected by time post-parasitization. This study highlights the function of G. pyloalis Hsps in response to environmental stress and provides a perspective for the control of this pest.

The Implication Inferred from the Expression of Small Heat-Shock Protein Genes in Dinoflagellate Resting Cysts Buried in Marine Sediment

Dinoflagellates are unicellular eukaryotic microalgae, occupying pivotal niches in aquatic ecosystems with great ecological, biological, and economic significance. Small heat shock proteins (sHsps) are the most omnipresent, but the least conserved, family of molecular chaperones found in all domains of life. Although their common name (small Hsp) implies to exclusively stress their heat shock-responsive function, many sHsps in fact engage in a variety of physiological processes, from cell growth and proliferation to embryogenesis, development, differentiation, apoptosis, and even to human disease prevention. Recent years have greatly expanded our understanding of sHsps in higher plants; however, comprehensive study aiming to delineate the composition and expression pattern of dinoflagellate sHsp gene family has not yet been performed. In this study, we constructed dinoflagellate-specific environmental cDNA library from marine sediment and sequenced using the third-generation sequencing technique. Screening of sHsp genes from the library returned 13 entries with complete coding regions, which were considered to be transcriptionally activated in the natural community of dinoflagellate resting cysts. All the 13 dinoflagellate sHsps consisted of a solely characteristic α-crystallin domain, covering 88–123 amino acid residues with the typical A-X-X-X-N-G-V-L motif, flanked by variable N- and C-terminal extensions. Multiple alignment revealed considerable amino acid divergence (~26.7% average similarity) among them. An unexpected close relationship was revealed between dinoflagellate and green algal sHsps in the phylogenetic tree, seemingly reflecting a close evolutionary relationship of these sHsps themselves. We confirmed that sHsp mRNAs are expressed during dormancy of the resting cyst assemblages of dinoflagellates that were buried in marine sediment, which raised the possibility that the sHsp expression is part of the machinery of maintaining the dormancy or/and the adaptation to ambient conditions of dinoflagellate resting cysts. Our results, although preliminary, gained an important glance on the universal presence of sHsps in dinoflagellates and their active expressions in the assemblage of resting cysts that were buried in the marine sediment. The essentiality of sHsps functioning in resting cysts necessitate more intensive and extensive investigations on all possible functions of Hsps in dinoflagellates, a group of protists with vital ecological and biological importance.

Genome-wide identification of heat shock proteins in harpacticoid, cyclopoid, and calanoid copepods: Potential application in marine ecotoxicology

Constant evolution of omics-technologies has provided access to identification of various important gene families. Recently, genome assemblies on widely used ecotoxicological model species, including rotifers and copepods have been completed and representative detoxification-related gene families have been discovered for biomarker genes. However, despite ubiquitous presence of stress-response proteins, limited information on full genome-wide report on heat shock proteins (Hsps) is available. Various studies have demonstrated multiple cellular functions of Hsps in living organisms as an important biomarker in response to abiotic and biotic stressors, however, full genome-wide identification of Hsps, particularly in aquatic invertebrates, has not been reported. This is the first study to report the entire Hsps and basal gene expression levels in three regional-specific copepods: Tigriopus japonicus and kingsejongensis, Paracyclopina nana, and Eurytemora affnis, and how each Hsp family gene is regulated at a basal level.

SoHSC70 positively regulates thermotolerance by alleviating cell membrane damage, reducing ROS accumulation, and improving activities of antioxidant enzymes

High temperature is a major environmental factor affecting plant growth. Heat shock proteins (Hsps) are molecular chaperones that play important roles in improving plant thermotolerance during heat stress. Spinach (Spinacia oleracea) is very sensitive to high temperature; however, the specific function of Hsps in spinach is unclear. In this study, cytosolic heat shock 70 protein (SoHSC70), which was induced by heat stress, was cloned from spinach. Overexpressing SoHSC70 in spinach calli and Arabidopsis enhanced their thermotolerance. In contrast, spinach seedlings with silenced SoHSC70 by virus-induced gene silencing (VIGS) showed more sensitivity to heat stress. Further analysis revealed that overexpressing SoHSC70 altered relative electrical conductivity (REC), malondialdehyde (MDA) content, photosynthetic rate, reactive oxygen species (ROS) accumulation and the activities of antioxidant enzymes, such as superoxide dismutase (SOD), peroxidase (POD), ascorbate peroxidase (APX), and catalase (CAT) after the heat treatment. Taken together, our results suggest that overexpressing SoHSC70 positively affects heat tolerance by reducing membrane damage and ROS accumulation and improving activities of antioxidant enzymes.

Comprehensive analysis of BpHSP genes and their expression under heat stresses in Betula platyphylla

HSPs are a large family of proteins, including HSP100, HSP90, HSP70, DnaJ and various sHSPs, which are important for plants to resist abiotic stress. However, the precise functions of HSPs involved in heat stress have not been thoroughly elucidated in birch (Betula platyphylla). In this study, 21 HSP genes in birch (named BpHSP1-BpHSP21) were cloned and characterized. The expression patterns of BpHSPs in response to high temperature stress were monitored by quantitative real time-polymerase chain reaction (qRT-PCR). The results showed that most of the BpHSPs were significantly upregulated or downregulated in response to high temperature, indicating that these BpHSPs may be involved in birch heat stress. Among the BpHSPs, the transcription of BpHSP9 was upregulated in stems and leaves under heat stress throughout all study periods, suggesting that this protein may play an important role under heat stress. Furthermore, BpHSP9-overexpressing, RNAi-silencing and empty pROKII vector (as the control) birch plants were generated by transient transformation. Under heat stress, overexpression BpHSP9 transgentic birch plants increased reactive oxygen species (ROS) scavenging, the activities of some antioxidant enzymes and non-enzymatic antioxidants. These effects led to diminished O2− and H2O2 content and reduced electrolyte leakage rates. However, the results were reversed when BpHSP9 was transiently silenced by RNAi. Taken together, these results strongly suggest that BpHSP9 may plays an important physiological role in the heat stress tolerance of birch plants.

Transcriptomic profiling of mitochondrial dysfunction induced apoptosis in accelerated cardiovascular disease

IntroductionVascular calcification is a significant contributor to accelerated cardiovascular disease and mortality in patients with chronic kidney disease (CKD). We previously reported that heat shock protein 70 (HSP70) induction can inhibit human aortic smooth muscle cell (HA‐SMC) calcification in CKD. In addition, our data indicated mitochondrial complexes II, IV, NADH cytochrome c reductase (NCCR), succinate cytochrome c reductase (SCCR), and cytochrome c oxidase (CCO) activities were preserved by HSP70 induction leading to reduced apoptosis. Apoptosis occurs with either receptor mediated extrinsic pathway or the mitochondrial‐dependent intrinsic pathway. The molecular mechanisms of HSP70 vasculo‐protective effects involving mitochondrial dysfunction induced apoptosis are currently unknown.HypothesisThe goal of the present study was to investigate the transcriptomic profile of genes involved in mitochondrial function and vasculo‐protective mechanisms of HSPs in apoptosis induced vascular calcification.MethodsHuman arteries were collected from healthy and CKD patients. Human aortic endothelial cells (HAECs) was treated in calcification medium (CM: 5mM β‐glycerolphosphate and 5mM CaCl2) in time‐course experiments (0–48 hours), in vitro. Total RNA from human arteries and HAECs was extracted using the Nucleospin RNA isolation kit. cDNA was synthesized and fragmented using SMART‐seq2 and Nextera XT (Illumina), respectively to generate libraries for sequencing on Hi‐seq 2000 (Illumina). Five million single‐ended raw reads from each sample were mapped to the human reference genome consortium GRCh38 by Burrows‐Wheeler Aligner (BWA). Gene analysis were performed by the combinations of fold‐changes on log 2 ratio, and p value < 0.05.ResultsIn healthy human arteries, we found that HSPA5 (fold change, FC, 3.85), HSPA1 (FC=3.64), and HSPA9 (FC=3.85) expression was preserved and Bcl2 associated Athanogene 1 (BAG1) (FC=7.44) was upregulated. However, mitochondrial fusion regulation genes MFN1 (FC=1.83) and OPA1 (FC=2.33) were down‐regulated in CKD arteries together with downregulation of downstream anti‐apoptotic gene Bcl2 (FC=4.52) and up‐regulation of pro‐apoptotic gene BAX (FC=2.77) and P53 (FC=5.35). Lastly, caspase 3 (FC=2.6) and cytochrome c (FC=2.29) expression were significantly increased in CKD arteries compared to control. In the primary HAECs, our data indicates that BAG1 was significantly increased at 6 hours prior to upregulation of Bcl2 at 12 hours in HAECs after CM treatment. BAG1 is a co‐chaperone for heat‐shock protein 70 (HSP70) that coordinates signals for cell growth in response to stress. Stress respondent genes, DNAJB6 (HSP40) and HSPA5 (HSP70 member5) was significantly increased at 24 hours and 12 hours respectively in CM‐treated HAECs.ConclusionComplex differential gene regulation involving preservation of mitochondrial function and cell apoptosis may be involved in the pathogenesis of accelerated vascular calcification. Decreased HSPs in CKD arteries may play the key role in mitochondrial dysfunction induced endothelial cells' apoptosis, which further leading to vascular calcification. Our findings may serve to inform the rational development of therapeutic strategies for vascular calcification involving therapeutic targeting of HSPs.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Heat shock protein (Hsp) regulation by muscarinic acetylcholine receptor (mAChR) activation in the rat hippocampus.

The cholinergic system plays a crucial role in modulating in the central nervous system physiological responses such as neurogenesis, neuronal differentiation, synaptic plasticity, and neuroprotection. In a recent study, we showed that Oxotremorine-M, a non-selective muscarinic acetylcholine receptor agonist, is able to transactivate the fibroblast growth factor receptor and to produce a significant increase in the hippocampal primary neurite outgrowth. In the present study we aimed to explore in the rat hippocampus the possible effect of acute or chronic treatment with Oxotremorine-M on some heat shock proteins (Hsp60, Hsp70, Hsp90) and on activation of related transcription factor heat shock factor 1 (HSF1). Following single injection of Oxotremorine-M (0.4 mg/kg) all Hsps examined were significantly increased in at least one of the time points studied (24, 48, and 72 hr). Treatment with Oxotremorine-M significantly increased the level of phosphorylated HSF1 in all time points studied, without change of protein levels. Similar pattern of Hsps changes was obtained following chronic Oxotremorine-M treatment (0.2 mg/kg) for 5 days. Surprisingly, following chronic treatment for 10 days no changes were observed in Hsps. The muscarinic acetylcholine receptor antagonist scopolamine (1 mg/kg) was able to completely block Oxotremorine-M effects on Hsps. In conclusion, considering the function of Hsps in protecting neuronal cells from deleterious proteotoxic stress, for example, protein mis-folding and aggregation, the results obtained indicate that muscarinic acetylcholine receptor activation may have implications in potential treatment of neurodegenerative disorders linked to protein aggregation, such as Alzheimer disease.

Plasmodium falciparum Hop (PfHop) Interacts with the Hsp70 Chaperone in a Nucleotide-Dependent Fashion and Exhibits Ligand Selectivity.

Heat shock proteins (Hsps) play an important role in the development and pathogenicity of malaria parasites. One of the most prominent functions of Hsps is to facilitate the folding of other proteins. Hsps are thought to play a crucial role when malaria parasites invade their host cells and during their subsequent development in hepatocytes and red blood cells. It is thought that Hsps maintain proteostasis under the unfavourable conditions that malaria parasites encounter in the host environment. Although heat shock protein 70 (Hsp70) is capable of independent folding of some proteins, its functional cooperation with heat shock protein 90 (Hsp90) facilitates folding of some proteins such as kinases and steroid hormone receptors into their fully functional forms. The cooperation of Hsp70 and Hsp90 occurs through an adaptor protein called Hsp70-Hsp90 organising protein (Hop). We previously characterised the Hop protein from Plasmodium falciparum (PfHop). We observed that the protein co-localised with the cytosol-localised chaperones, PfHsp70-1 and PfHsp90 at the blood stages of the malaria parasite. In the current study, we demonstrated that PfHop is a stress-inducible protein. We further explored the direct interaction between PfHop and PfHsp70-1 using far Western and surface plasmon resonance (SPR) analyses. The interaction of the two proteins was further validated by co-immunoprecipitation studies. We observed that PfHop and PfHsp70-1 associate in the absence and presence of either ATP or ADP. However, ADP appears to promote the association of the two proteins better than ATP. In addition, we investigated the specific interaction between PfHop TPR subdomains and PfHsp70-1/ PfHsp90, using a split-GFP approach. This method allowed us to observe that TPR1 and TPR2B subdomains of PfHop bind preferentially to the C-terminus of PfHsp70-1 compared to PfHsp90. Conversely, the TPR2A motif preferentially interacted with the C-terminus of PfHsp90. Finally, we observed that recombinant PfHop occasionally eluted as a protein species of twice its predicted size, suggesting that it may occur as a dimer. We conducted SPR analysis which suggested that PfHop is capable of self-association in presence or absence of ATP/ADP. Overall, our findings suggest that PfHop is a stress-inducible protein that directly associates with PfHsp70-1 and PfHsp90. In addition, the protein is capable of self-association. The findings suggest that PfHop serves as a module that brings these two prominent chaperones (PfHsp70-1 and PfHsp90) into a functional complex. Since PfHsp70-1 and PfHsp90 are essential for parasite growth, findings from this study are important towards the development of possible antimalarial inhibitors targeting the cooperation of these two chaperones.

The double-edged sword: conserved functions of extracellular hsp90 in wound healing and cancer.

Heat shock proteins (Hsps) represent a diverse group of chaperones that play a vital role in the protection of cells against numerous environmental stresses. Although our understanding of chaperone biology has deepened over the last decade, the “atypical” extracellular functions of Hsps have remained somewhat enigmatic and comparatively understudied. The heat shock protein 90 (Hsp90) chaperone is a prototypic model for an Hsp family member exhibiting a duality of intracellular and extracellular functions. Intracellular Hsp90 is best known as a master regulator of protein folding. Cancers are particularly adept at exploiting this function of Hsp90, providing the impetus for the robust clinical development of small molecule Hsp90 inhibitors. However, in addition to its maintenance of protein homeostasis, Hsp90 has also been identified as an extracellular protein. Although early reports ascribed immunoregulatory functions to extracellular Hsp90 (eHsp90), recent studies have illuminated expanded functions for eHsp90 in wound healing and cancer. While the intended physiological role of eHsp90 remains enigmatic, its evolutionarily conserved functions in wound healing are easily co-opted during malignancy, a pathology sharing many properties of wounded tissue. This review will highlight the emerging functions of eHsp90 and shed light on its seemingly dichotomous roles as a benevolent facilitator of wound healing and as a sinister effector of tumor progression.

Four heat shock protein genes of the endoparasitoid wasp, Cotesia vestalis, and their transcriptional profiles in relation to developmental stages and temperature.

Heat shock proteins (Hsps) play important roles in the environmental adaptation of various organisms. To explore the functions of Hsps in relation to heat stress and development in Cotesia vestalis, a solitary larval endoparasitoid of Plutella xylostella, four heat shock protein genes, CvHsp40, CvHsc70, CvHsp70 and CvHsp90, were cloned and sequenced from C. vestalis by real-time quantitative PCR and RACE. The cDNA sequence of CvHsp40, CvHsc70, CvHsp70 and CvHsp90 were 1473 bp, 2316 bp, 2279 bp and 2663 bp long, which encode proteins with calculated molecular weights (MW) of 39.1 kDa, 71.2 kDa, 70.1 kDa and 83.3 kDa, respectively. Furthermore, the analysis of genomic DNA confirmed that no introns existed in CvHsp40, CvHsp70 and CvHsp90 while two introns were present in CvHsc70. The amino acid sequence analysis of CvHsps indicated that CvHsp40 is a Type II Hsp40 homolog, CvHsp70 and CvHsc70 are the eukaryotic cytoplasmic Hsp70s, and CvHsp90 is the β-isoform of Hsp90. The divergent transcriptional patterns of CvHsp40, CvHsp70 and CvHsp90 in the different developmental stages suggested that CvHsp transcripts were under different mechanisms of regulation during the development of parasitoid larvae. The dramatic increase of transcripts of CvHsp70 at the third-instar larva coincided with its developmental change in this stage, that is, from inside host to outside host. CvHsp40, CvHsc70 and CvHsp70 showed a trend of sex-specific differences of transcript abundance in the adult stage. All CvHsp transcripts in different developmental stages were significantly induced by heat stress, and the lowest transcript abundances appeared around the temperature 27°C, which probably suggest that this is the most favorable temperature for the development of C. vestalis. Our results suggest that the expression of heat shock proteins reflects to some extent the developmental changes and environmental requirements of insects.

Heat Shock Proteins In Cancer: Targeting the ‘Chaperones’

Heat shock proteins (Hsps) are highly conserved proteins working as molecular chaperones for several cellular proteins essential for normal cell viability and growth, and have numerous cytoprotective roles. The expression of Hsps is induced in response to a wide variety of physiological and environmental stress insults, including anticancer chemotherapy, thus allowing the cell to survive lethal conditions. Cancer cells experience high levels of proteotoxic stress and rely upon stress-response pathways for survival and proliferation, thereby becoming dependent on proteins such as stress-inducible Hsps. Owing to the implication of Hsps in cancer, Hsp inhibition has recently emerged as an interesting potential anticancer strategy. Many natural and synthetic Hsp inhibitors molecular compounds are in development and many are being evaluated as potential cancer therapies. One of the Hsps in particular, Hsp90, has several client proteins and is emerging as a particularly exciting cancer target due to the prospect of simultaneously inhibiting chaperoning of numerous oncogenic proteins. This review describes the function of Hsps focusing on current efforts in exploiting the attributes of Hsps as potential targets for anticancer therapy.

Heat shock protein 70 purification and characterization from Cyprinion macrastomus macrastomus and Garra rufa obtusa

Organisms resist stress factors by common mechanisms at cellular and physiological levels. Heat shock proteins (Hsps) play an essential role as a protective mechanism against stressors, and Hsp70 has a central role in the heat shock response. Although several investigations have been made on the cellular functions of Hsps, very little is known about extreme temperature effects on organisms like fish. To address this point, we have studied two fish species, Cyprinion macrostomus macrostomus and Garra rufa obtuse, whose habitat is the Kangal hot spring in Turkey. The hot spring has multiple stress factors; extremely hot temperature, food deprivation, infectious medium and low levels of oxygen. Like other Hsp70s, two Cyprinidae species Hsp70 showed similar ATPase and substrate protein folding activities in vitro. These proteins also showed relatively higher thermal stability. Biochemical characterization of two Hsp70, suggested a Hsp mechanism capable of protecting the cell against stressors even under adverse conditions and in the presence of multiple stress factors.

Heat shock proteins and cancer therapy: The trail grows hotter!

The physiological properties of the heat shock response were initially studied over three decades ago in Drosophila based on the observation that it had protective functions against teratogenic agents and heat induced animal death. While initially believed to be unique to flies, it was rapidly observed to be universal response common to all living organisms. Early mammalian studies (reviewed in ref. [1]) focused on the physiological role of heat shock proteins or HSPs in protecting the organism against heat, a phenomenon called thermotolerance. It was determined that HSPs were able to protect cellular proteins and therefore cells from proteotoxic stress such as heat shock. This general function of HSPs are not restricted to stress protection, but is an essential element in the natural operation of cellular machinery which can also lead to protein damage under non-stress conditions in a protein concentrated intracellular environment. Such interactions of the HSP with cellular proteins would protect the substrate protein from the damaging effects of heat and other proteotoxic conditions as well as to aid in the clearance of some damaged proteins via degradation. In cancer research, HSPs were initially perceived to play a role protecting tumors from hyperthermia when applied in multiple treatments, i.e. HSPs induced initially would protect the tumor from later treatments [2]. It took many more years to begin to appreciate that there are other and potentially more significant roles for HSPs in cancer. For example, it was recognized long ago that interference with HSP90, could revert tumor cells from a transformed to a normal phenotype due to the role of HSP90 in facilitating the functions of tyrosine kinases necessary in cell transformation. The ability to block HSP90 function has since been aggressively pursued as a cancer therapy using specific HSP90 inhibitors such as geldanamycin [3]. Later studies revealed the additional and more general role for HSPs in cancer. All proliferating cells require an increase in HSP chaperone concentrations due to an increase in macromolecular activities that occur compared to quiescent cells. This increase in activity can be expected to lead to an increase in various phenomena such as free radical formation and related protein damage in proliferating cells. In addition to this, the increased accumulation of mutated proteins expected in cancer also poses a further likelihood for cell damage compared to normal cells. Combined, these phenomena can result in the protein damage signals leading to the enhanced heat shock response and HSP levels, protecting the cancer cells from the accumulation of misfolded proteins. Thus, cancer cells can have an already activated heat shock response, at least in part, relative to normal cells. While a heat shock response can be robust, the pre-stress activated response in cancer suggests that cancer cells would be less capable of mounting a further strong response relative to normal cells. Therefore, additional proteotoxic stress such as hyperthermia may be more toxic to cancer cells relative to normal cells. A new paper by Neznanov et al [4] examines this hypothesis. It proposes that that proteotoxic stress such as hyperthermia might be more effective in killing cancer relative to normal tissues since a level of heat shock response is already partially occupied in cancer cells. Neznanov et al examined the combinations of three forms of proteotoxic stress, such as hyperthermia, puromycin treatment, and bortezomib treatment (which blocks protein degradation thereby increasing the accumulation of damaged proteins that would otherwise be eliminated). They show that these treatments caused substantial control of multiple myeloma cell growth, in vitro and in vivo. The increased cell death in cancer cells was found to be largely due to a p53-independent apoptotic process. This study provides tantalizing evidence that cancer cells are already addicted to the heat shock response for their own survival as hypothesized. The same group has previously used another approach to achieve a similar outcome. They have demonstrated that the inhibition of HSP expression or function through either inhibitors of the transcription factor heat shock factor 1or inhibitors of HSP70 function can also be toxic to cancer cells, again based on the same rationale, i.e. the enhanced need for HSPs in cancer cells [5]. Therefore the heat shock response has important survival functions in the development and progression of cancer. Importantly, studies from the Calderwood group also connect the heat shock response with carcinogenesis [6]. This information introduces new targets to exploit in cancer therapy. It is uplifting to see the very early and pioneering studies on the protective functions of HSPs in Drosophila being now applied to cancer therapy.

Expression and Functions of Heat Shock Proteins in the Normal and Pathological Mammalian Eye

Heat shock proteins (Hsps) are expressed in mammalian embryonic, adult and aging lens, cornea and retina. These proteins, particularly those belonging to the family of small Hsps, such as αA-crystallin (HspB4) and αB-crystallin (HspB5), play important roles in the differentiation of lens cells and are essential for the maintenance and protection of the supraorganization of proteins in differentiated corneal and lens fiber cells. Hsps are molecular chaperones characterized by their protective activity against different types of stress. They also have anti-apoptotic and anti-oxidant functions that help lens and corneal cells to better cope with the oxidative conditions that result from light induced injuries. They are also effective to protect the retina against the high rate of oxidative metabolism observed in this tissue. The goal of this review is to highlight recent works describing the expression and function(s) of the different Hsps as an attempt to better understand their roles in the normal and pathological eye. Particular emphasis is given to the α-crystallin polypeptides which, in addition to their protective functions, are key structural polypeptides that are essential for the refractive and light focusing properties of the lens, a property demonstrated by the caractogenic potential of their mutation.

Heat shock protein genes in fish

Heat shock proteins are a family of highly conserved cellular proteins present in all organisms including fish. Fish represent an ideal model organism to understand the regulation and functional significance of heat shock proteins (Hsps). The mechanism regulating the expression of Hsp genes in fish have not been studied in detail. In this review, the function, genomic structure and environmental adaptation of the major fish Hsps were discussed. Future research evaluating the functional genomics of Hsps in fish will provide substantial insight into the physiological and ecological roles of these highly conserved proteins.