Characterization Of Heat Shock Protein Research Articles

Molecular characterization of heat shock protein 70 gene in Deoni cattle (Bos indicus)

Molecular characterization of heat shock protein 70 gene in Deoni cattle (Bos indicus)

Insilico-based genome-wide identification and structural characterization of heat shock proteins 70 (HSP70) in cowpea (Vigna unguiculata L.)

Insilico-based genome-wide identification and structural characterization of heat shock proteins 70 (HSP70) in cowpea (Vigna unguiculata L.)

Computational analysis of heat shock protein (HSP) in Citrus X Sinensis

Heat shock proteins (HSPs) are molecular chaperones and one of the cell’s most important regulatory proteins present in all species. HSPs are a multigene family classified into six families according to their molecular weight range between 8KDa to 110KDa: HSP100, HSP90, HSP70, HSP60, small heat shock proteins (sHSPs), and ubiquitin. Citrus X Sinensis is very economical, and widely grown in the world. It is one of the best sources of nutrition, vitamin C, fiber, thiamine, folate, and antioxidants. It has a variety of health benefits i.e., protects against cell damage, boosts the immune system, heals wounds, etc. For highlighting the aspect of HSPs of citrus in computing protein, this work has great significance. Present studies are based on the identification and characterization of heat shock proteins in Citrus sinuses. In silico research was performed; Arabidopsis thaliana protein sequences of HSP60, HSP70 and HSP90 were used as a reference to identify these molecular chaperones in Citrus sinuses through Blast Phytozome. Furthermore, physiochemical properties, multiple sequence alignment, Multiple Expectation Maximization for Motif Elicitation (MEME) and phylogeny were constructed for 44 sequences of HSP60, 22 sequences of HSP70 and 23 for HSP90. The best expression of HSPs was found in Endoplasmic Reticulum. The findings of the present studies provide a good understanding of the HSP60, HSP70 and HSP90 families' function and expression site in the cell of Citrus sinuses. It also provides the basic knowledge for further investigations of the putative structure of dynamic proteins.

Bioinformatics Analysis, Expression Profiling, and Functional Characterization of Heat Shock Proteins in Wolfi-poria cocos

Heat shock proteins (HSPs) play critical roles in regulating different mechanisms under high-temperature conditions. HSPs have been identified and well-studied in different plants. However, there is a lack of information about their genomic organization and roles in medicinal plants and fungi, especially in Wolfi-poria cocos (W. cocos). We identified sixteen heat shock proteins (HSPs) in W. cocos and analyzed in terms of phylogenetic analysis, gene structure, motif distribution patterns, physiochemical properties, and expression comparison in different strains. Based on phylogenetic analysis, HSPs were divided into five subgroups (WcHSP100, WcHSP90, WcHSP70, WcHSP60, and WcsHSP). Subgroups WcHSP100s, WcHSP90s, WcHSP70s, WcHSP60, and WcsHSPs were further divided into 3, 2, 3, 1, and 6 subfamilies, respectively. Moreover, the expression profiling of all HSP genes in five strains of W. cocos under different temperature extremes revealed that expression of most HSPs were induced by high temperature. However, every subfamily showed different expression suggesting distinctive role in heat stress tolerance. WcHSP70-4, WcHSP90-1, and WcHSP100-1 showed the highest response to high temperature stress. Heterologous expression of WcHSP70-4, WcHSP90-1, and WcHSP100-1 genes in Escherichia coli enhanced survival rate of E. coli during heat stress. These findings suggest the role of W. cocos heat shock genes in the high temperature stress tolerance.

Genome-Wide Identification and Characterization of Heat Shock Protein 20 Genes in Maize.

Maize is an important cereal crop worldwide and is sensitive to abiotic stresses in fluctuant environments that seriously affect its growth, yield, and quality. The small heat shock protein (HSP20) plays a crucial role in protecting plants from abiotic stress. However, little is known about HSP20 in maize (ZmHSP20). In this study, 44 ZmHSP20s were identified, which were unequally distributed over 10 chromosomes, and 6 pairs of ZmHSP20s were tandemly presented. The gene structure of ZmHSP20s was highly conserved, with 95% (42) of the genes having no more than one intron. The analysis of the cis-element in ZmHSP20s promoter demonstrated large amounts of elements related to hormonal and abiotic stress responses, including abscisic acid (ABA), high temperature, and hypoxia. The ZmHSP20s protein had more than two conserved motifs that were predictably localized in the cytoplasm, nucleus, endoplasmic reticulum, peroxisome, mitochondria, and plasma. Phylogenetic analysis using HSP20s in Arabidopsis, rice, maize, and Solanum tuberosum indicated that ZmHSP20s were classified into 11 categories, of which each category had unique subcellular localization. Approximately 80% (35) of ZmHSP20 were upregulated under heat stress at the maize seedling stage, whereas the opposite expression profiling of 10 genes under 37 and 48 °C was detected. A total of 20 genes were randomly selected to investigate their expression under treatments of ABA, gibberellin (GA), ethylene, low temperature, drought, and waterlogging, and the results displayed that more than half of these genes were downregulated while ZmHSP20-3, ZmHSP20-7, ZmHSP20-24, and ZmHSP20-44 were upregulated under 1 h treatment of ethylene. A yeast-one-hybrid experiment was conducted to analyze the binding of four heat stress transcription factors (ZmHSFs) with eight of the ZmHSP20s promoter sequences, in which ZmHSF3, ZmHSF13, and ZmHSF17 can bind to most of these selected ZmHSP20s promoters. Our results provided a valuable resource for studying HSP20s function and offering candidates for genetic improvement under abiotic stress.

Molecular characterization of heat shock protein 20 (hsp20) in goldfish (Carassius auratus) and expression analysis in response to environmental stresses

Heat shock proteins (Hsps) are described as molecular chaperones that were synthesized rapidly in response to various stress conditions in many organisms. However, information of small heat shock protein (sHsp) in response to environmental stresses is lacking in fish. In this study, the full length cDNA of hsp20 in goldfish, Carassius auratus, with a 65 bp 5′-untranslated region (UTR), a 563 bp 3′-UTR, and a 450 bp open reading frame encoding 149 amino acids, was obtained. gfhsp20 contained a highly conserved α-crystallin domain. Consistent with humans and zebrafish, goldfish HSP20 gene had three exons and two introns. gfhsp20 mRNA were detected by quantitative real-time PCR in all of the tissues examined, with the highest level in the muscle, followed by the skin, heart, intestine, liver, spleen, brain, kidney, gill and head kidney. gfhsp20 mRNA was significantly induced with heat shock in liver and spleen, but decreased in brain. Under H2O2-mediated oxidative stress, the expression level of gfhsp20 was significantly up-regulated in brain, liver and spleen. After LPS and PolyI:C infection, the expression level of gfhsp20 in spleen was significantly up-regulated with a higher increase under PolyI:C infection than that of LPS injection. These results revealed the potential function of gfHsp20 in response to heat shock, oxidative stress and infectious pathogens.

Characterization of Heat Shock Protein 60 as an Interacting Partner of Superoxide Dismutase 2 in the Silkworm, Bombyx mori, and Its Response to the Molting Hormone, 20-Hydroxyecdysone.

Oxidative stress promotes pupation in some holometabolous insects. The levels of superoxide, a reactive oxygen species (ROS), are increased and superoxide dismutase 1 (BmSod1) and superoxide dismutase 2 (BmSod2) are decreased during metamorphic events in silkworm (Bombyx mori). These observations strongly suggest that pupation is initiated by oxidative stress via the down-regulation of BmSod1 and BmSod2. However, the molecular mechanisms underlying ROS production during metamorphic events in silkworm remain unknown. To investigate these molecular mechanisms, the peripheral proteins of BmSod1 and BmSod2 were identified and characterized using dry and wet approaches in this study. Based on the results, silkworm heat shock protein 60 (BmHsp60) was identified as an interacting partner of BmSod2, which belongs to the Fe/MnSOD family. Furthermore, the present study results showed that BmHsp60 mRNA expression levels were increased in response to oxidative stress caused by ultraviolet radiation and that BmHsp60 protein levels (but not mRNA levels) were decreased during metamorphic events, which are regulated by the molting hormone 20-hydroxyecdysone. These findings improve our understanding of the mechanisms by which holometabolous insects control ROS during metamorphosis.

Molecular characterization of heat shock protein 70 gene in Iraqi buffalo

The heat shock protein 70 (HSP 70) has important roles in protecting cells and keeping them alive when exposed to different stress conditions. The polymorphism of the hsp 70 gene could be linked with the ability of stress tolerance. This study aimed to determine the polymorphism of the hsp 70 gene in Iraqi buffaloes and study bits effects on the resistance to stress. This study was conducted during from November 2018 to February 2019. The number of buffalo females used was 35 at the age of 4 - 6 years, which belonged to the local farmers from Basra city, Iraq. The DNA was extraction from the blood samples then the polymerase chain reaction (PCR) amplification was performed. The DNA sequences were analyzed by using bioinformatics analysis. The results of the molecular analysis showed that there were two groups of the hsp 70 gene as a compare with the same genes in GenBank due to silent and missense mutations. Based on these findings, it can be concluded that the Iraqi buffaloes have adapted to the surrounding environmental conditions as a result of the action of HSP 70 proliferation. The hsp 70 gene was a useful biomarker of stress tolerance in buffaloes.

Molecular characterization of heat shock protein 90 from the dinoflagellate Prorocentrum donghaiense and its transcriptional response to thermal, copper and nutrient stresses

ABSTRACTIn this study, a novel HSP90 gene (PdHSP90) was cloned from Prorocentrum donghaiense. The full length of PdHSP90 is 2443 bp, with 5′-UTR of 111 bp, 3′-UTR of 199 bp, and open reading frame (ORF) of 2133 bp. The ORF encodes a polypeptide of 710 amino acid residues, with molecular weight of approximately 85.81 kDa and isoelectric point of 4.95. Phylogenetic analysis revealed that PdHSP90 belongs to the dinoflagellate clade and is closely related to HSP90s from P. minimum and P. micans. PdHSP90 expression increased under the thermal stress (p < 0.05), with the highest changes of 7.6-fold at 10°C 2 h post treatment compared to the control. PdHSP90 expression was significantly upregulated (1- to 10.8-fold change) (p < 0.05) with increasing concentrations of copper from 0 µM to 31.25 µM. PdHSP90 expression level under 176 µM nitrogen was nearly equal to that under 88 µM nitrogen, which were 4.5- and 4.6-fold higher, respectively, compared with the control (p < 0.05). Upon treatment with phosphorus limitation from 36 µM to 3.6 µM, PdHSP90 transcription level increased 11.4-fold (p < 0.05) and then decreased. These results provide new insights into the molecular mechanism underlying adaptation of P. donghaiense and other harmful dinoflagellates in response to diverse environmental factors.

Expression status of heat shock proteins in response to cold, heat, or insecticide exposure in the Colorado potato beetle Leptinotarsa decemlineata.

The Colorado potato beetle (Leptinotarsa decemlineata (Say)) is an agricultural pest that threatens the potato industry worldwide. This insect is widely regarded as one of the most difficult-to-control pests, as it can thrive in a wide range of temperature conditions and routinely develops resistance towards various insecticides. The molecular changes associated with response to these challenges have not been fully investigated in L. decemlineata. While differential expression and characterization of heat shock proteins (HSPs) in response to stress have been conducted in several insects, data regarding HSPs in L. decemlineata are limited. The overarching objective of this study consisted of evaluating the expression of various HSPs in L. decemlineata exposed to different temperatures or treated with the insecticides imidacloprid and chlorantraniliprole. Expression levels of HSP60, HSP70, HSP90, and HSP Beta-1 were evaluated by qRT-PCR and insect mortality was assessed using dsRNAs aimed at select HSP targets. Elevated HSP70 and HSP90 transcript levels were observed in heat-exposed L. decemlineata while downregulation of HSP70 transcript levels was measured in insects submitted to cold conditions. Chlorantraniliprole exposure was associated with reduced HSP Beta-1 transcript levels while no change in expression was monitored in insects exposed to imidacloprid. RNAi-based knockdown of HSP60 levels correlated with significant insect mortality 14days after dsRNA injection. These results highlight the modulation of HSPs that occur in L. decemlineata exposed to fluctuating temperatures and position HSPs as interesting candidates in the identification of novel molecular leads that could be targeted to control this insect.

Molecular Characterization of Heat Shock Protein 70-1 Gene of Capra aegagrus blythi

Molecular Characterization of Heat Shock Protein 70-1 Gene of Capra aegagrus blythi

Characterization of heat shock protein 27 in extracellular vesicles: a potential anti-inflammatory therapy.

Previously, we reported that elevated serum levels of heat shock protein 27 (HSP27) are predictive of a lower risk of having a heart attack, stroke, or death from cardiovascular disease. Moreover, augmenting HSP27 (or the murine ortholog, HSP25) attenuated experimental atherogenesis, reduced inflammation, and lowered cholesterol levels. Recently, we noted that HSP27 activates NF-κB via TLR-4, resulting in attenuation of plaque inflammation; however, the precise anti-atherosclerosis mechanisms mediated by extracellular HSP27 are incompletely understood. Our purpose in this study was to investigate the existence of HSP27 in extracellular vesicles (EVs) and whether HSP27 elicited atheroprotective effects on target cells. Here, we provide evidence that HSP27 localizes to EVs derived from THP-1 cells using transmission electron microscopy (TEM) and immunogold labeling, Western blotting, ELISA, and fluorescence-activated cell sorting. TEM imaging indicated that HSP27 is found at the exosomal membrane. Multiple reactor monitor-mass spectrometric analysis of large vesicles, which included microparticles and exosomes, isolated from human plasma, also led to detection of HSP27 using the unique signature peptide, R.LFDQAFGLPR.L. Studies using THP-1 and human embryonic kidney cells show that HSP27-laden exosomes significantly stimulated NF-κB activation ( P < 0.001) and release of IL-10 ( P < 0.0001), suggesting that HSP27 may be important exosomal cargo with beneficial anti-inflammatory effects.-Shi, C., Ulke-Lemée, A., Deng, J., Batulan, Z., O'Brien, E. R. Characterization of heat shock protein 27 in extracellular vesicles: a potential anti-inflammatory therapy.

Identification and characterization of heat shock protein 90 (HSP90) in the hard coral Acropora tenuis in response to Irgarol 1051

To elucidate the effects of the herbicide Irgarol 1051 on the gene expression of heat shock protein 90 (HSP90) in hard corals, we isolated a full-length cDNA encoding HSP90 from Acropora tenuis, which has a deduced open reading frame of 732-amino acid residues with a predicted molecular mass of 84.5 kDa. The amino acid sequence of A. tenuis HSP90 showed a high degree of similarity with the hermatypic-coral HSP90 families. After a 7-d exposure to 1 or 10 μg/L of Irgarol, the body colours of corals in the 10 μg/L treatment group were significantly whiter (bleached), whereas no such effects were observed in the corals in the 1 μg/L treatment group. However, the expression level of coral HSP90 was significantly downregulated after exposure to both 1 and 10 μg/L Irgarol. These results suggest that A. tenuis HSP90 may be a useful molecular biomarker to predict bleaching caused by herbicides.

Characterization of heat shock protein 70 transcript from Nilaparvata lugens (Stål): Its response to temperature and insecticide stresses

The brown planthopper, Nilaparvata lugens, possesses a strong adaptability to extreme temperature and insecticide stresses. Heat shock proteins (Hsps) are highly conserved molecular chaperones and play a pivotal role in response to various environmental stresses in insects. However, little is known about the response of Hsps to stresses in N. lugens. In the present study, an inducible Hsp70 (NlHsp70) was isolated from this insect and transcriptional expression patterns of NlHsp70 under temperature and insecticide stresses were analyzed. The full-length of NlHsp70 was 2805bp with an open reading frame (ORF) of 1896bp, showing high homology to its counterparts in other species. Expression of NlHsp70 was not altered by heat shock for 1h, nor following recovery from thermal stress. Conversely, decreased expression of NlHsp70 was observed in response to cold shock. In addition, the expression of NlHsp70 increased after imidacloprid exposure. RNA interference experiment combined with insecticide injury assay also demonstrated that NlHsp70 was essential for resistance against insecticide exposure. These observations indicated that NlHsp70 was an important gene involved in the resistance or tolerance to environmental stresses in N. lugens. Interestingly, weak changes in mRNA expression levels of two thermal-inducible Hsp genes, NlHsp90 and NlHsc70 were observed in imidacloprid-exposed N. lugens adults, suggesting that different Hsps may respond differential to the extreme temperature and insecticide stresses.

Characterization of Heat Shock Protein (HSP 70) Sequence in Leishmania donovani

Heat shock protein HSP70 is the highly conserved protein along the kinetoplastid phylogenetic spectrum. A full length hsp70 gene was amplified, cloned and sequenced. The protein sequence was aligned and phylogentic tree was constructed. The Leishmania donovani HSP70 protein sequence (653 amino acids) is slightly larger than the Human HSP70 protein sequence (641 amino acids) with 72% identity. The putative protein sequence is 98%, 97%, 96% and 92% identical to L. major, L. infantum, L. tarentolae and L. braziliensis respectively. Evolutionary relationship showed that kinetoplastid parasites are closely related to the human compared to prokaryotes, E. coli.Journal of Institute of Science and Technology, 2015, 20(1): 82-86

Molecular characterization of heat shock protein 70 cognate cDNA and its upregulation after diapause termination in Lycorma delicatula eggs

Since the sudden introduction of Lycorma delicatula into Korea in 2006, annual outbreaks of the species have seriously damaged various forest trees and horticultural plants, with grape vines being particularly affected. It is a univoltine species that overwinters during the egg stage by entering embryonic diapause. However, the mechanisms in relation to diapause of L. delicatula eggs remain unknown. Microscopic observation of diapausing eggs has shown that embryonic development was arrested at blastokinesis. Egg masses were collected monthly from a grape vineyard in the Daegu area, from December 2012 to April 2013, and incubated at 25°C until hatching. When the duration of overwintering was longer, the hatching rate was higher, the hatching duration was shorter, and the initial hatch date was occurred earlier. We elucidated the full cDNA sequence of the heat shock cognate 70 (hsc70) gene of L. delicatula. Quantitative real-time RT-PCR showed that the hsc70 expression level was very low in eggs collected from November to February but much higher in eggs collected in the field during March. Furthermore, hsc70 expression levels gradually increased postdiapause, until hatching at 25°C. Our results suggest that diapause termination in L. delicatula eggs requires a chilling period, which also facilitates postdiapause development. In addition, hsc70 may play a role in the resumption of embryonic development in eggs emerging from diapause.

Molecular characterization of heat shock protein 70 (HSP 70) promoter in Japanese flounder (Paralichthys olivaceus), and the association of Pohsp70 SNPs with heat-resistant trait

Ambient temperature is one of the major abiotic environmental factors determining the main parameters of fish vital activity. HSP70 plays an essential role in heat response. In this investigation, the promoter and structure of Paralichthys olivaceus hsp70 (Pohsp70) gene was cloned and predicted. 2558 bp upstream regulatory region of Pohsp70 was annotated with four potential promoter elements and four putative binding sites of transcription factors heat shock elements (HSE, nGAAn) in the upstream of the transcription start site. In addition, one intron with 454 bp in the 5'-noncoding region was found. Quantitative Real Time PCR analysis indicated that the transcript level of Pohsp70 was raised markedly after 1 h by heat shocked. Furthermore, 25 SNPs were identified in Pohsp70 by resequencing, seven of which was associated with heat resistance. In addition, two of the seven SNPs, namely SNP14 and SNP16, were observed in strong linkage disequilibrium. The haplotype with association analysis showed TAGGAG haplotype was more represented in heat susceptible group while (DEL/T) GAATA haplotype was more frequent in heat resistant group. The heat resistant SNPs and haplotype could be candidate markers potentially serving for selective breeding programs of Japanese flounder aimed at improving anti-stress and production.

Characterization of heat-shock proteins in Escherichia coli strains under thermal stress in vitro

The aim of this study was to evaluate the effect of heat stress in in vitro conditions on the induction of heat-shock protein (Hsp)70 by Escherichia coli cells, and to determine the localization of Hsps in cell fractions. The material consisted of wild strains of E. coli isolated from the digestive tract of calves, suspended in an exponential-phase culture and subjected to 41.5 °C for 2 h. Individual fractions were analysed by SDS-PAGE and two-dimensional electrophoresis. Western blotting with mouse anti-Hsp70 and anti-Hsp60 mAbs was used to identify the proteins. Electrophoretic analysis of the heat-treated cells detected Hsp70 in all three fractions, cytoplasmic, periplasmic and membrane, which was confirmed by Western blotting. The proteins obtained had diverse localizations in the pH gradient in two-dimensional electrophoresis, which may indicate changes in their conformation and physical properties leading to stabilization and protection of intracellular structures in stress conditions. The presence of these Hsps in different cell fractions indicates a very strong protective adaptation in the bacteria in unfavourable conditions, which is critical for the organism infected by them.

Characterization of heat shock protein 90, 70 and their transcriptional expression patterns on high temperature in adult of Grapholita molesta (Busck)

Grapholita molesta (Busck) is a worldwide insect pest damaging stone and pome fruits. High temperature can significantly affect insect survival, development and fecundity. Heat shock protein (Hsp) genes were speculated to possess a pivotal function in response to high temperature stress. In this study, two full-length Hsp genes, Gmhsp90 and Gmhsp70, were cloned from G. molesta using rapid amplification of complementary DNA ends (RACE). The open reading frames of Gmhsp90 and Gmhsp70 obtained were 2 148 bp and 1 998 bp in length, respectively. Their deduced amino acids showed high homology to Hsp genes of other species. Subsequently, the transcriptional expression of Gmhsp90 and Gmhsp70 in G. molesta adults exposed at various temperatures (26, 29, 32, 35, 38, 41 and 44°C) for 1h and at 41°C for various time duration (0, 15, 30, 45, 60, 75, 90 and 105 min) were investigated via real time quantitative polymerase chain reaction (qPCR). The relative expression levels of Gmhsp90 and Gmhsp70 in G. molesta adults were both up-regulated with the rise of temperature and time duration. In addition, the Gmhsp70 usually showed a higher transcription accumulation than Gmhsp90. Interestingly, Gmhsp90 and Gmhsp70 in female adults could be induced much earlier than that in males, and the effective induction temperature in females was also lower than that in males. The distinct expression profiles of Gmhsp90 and Gmhsp70 indicated that Gmhsp90 and Gmhsp70 may play important roles in G. molesta adults responding to a thermal threat, and there is difference on induction between sexes.

Functional characterization of heat shock protein 90 targeted compounds

Heat shock protein 90 (Hsp90) is an attractive cancer target that possesses two potential binding domains at the C and N termini. Nevertheless, assays that can reliably distinguish between the C- and N-terminal Hsp90 inhibitors and quantitatively characterize candidate drugs are limited. Here we report an Hsp90 binding assay and ATPase inhibition assay that can not only separate the C- and N-terminal inhibitors but also quantitatively determine their respective IC50 values.