SEARCH · PubMed Health

Results for “Hsp70”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources

PubMed1992

Cloning of the HSP70 gene from Halobacterium marismortui: relatedness of archaebacterial HSP70 to its eubacterial homologs and a model for the evolution of the HSP70 gene.

Heat shock induces the synthesis of a set of proteins in Halobacterium marismortui whose molecular sizes correspond to the known major heat shock proteins. By using the polymerase chain reaction and degenerate oligonucleotide primers for conserved regions of the 70-kDa heat shock protein (HSP70) family, we have successfully cloned and sequenced a gene fragment containing the entire coding sequence for HSP70 from H. marismortui. HSP70 from H. marismortui shows between 44 and 47% amino acid identity with various eukaryotic HSP70s and between 51 and 58% identity with its eubacterial and archaebacterial homologs. On the basis of a comparison of all available HSP70 sequences, we have identified a number of unique sequence signatures in this protein family that provide a clear distinction between eukaryotic organisms and prokaryotic organisms (archaebacteria and eubacteria). The archaebacterial (viz., H. marismortui and Methanosarcina mazei) HSP70s have been found to contain all of the signature sequences characteristic of eubacteria (particularly the gram-positive bacteria), which suggests a close evolutionary relationship between these groups. In addition, detailed analyses of HSP70 sequences that we have carried out have revealed a number of additional novel features of the HSP70 protein family. These include (i) the presence of an insertion of about 25 to 27 amino acids in the N-terminal quadrants of all known eukaryotic and prokaryotic HSP70s except those from archaebacteria and the gram-positive group of bacteria, (ii) significant sequence similarity in HSP70 regions comprising its first and second quadrants from organisms lacking the above insertion, (iii) highly significant similarity between a protein, MreB, of Escherichia coli and the N-terminal half of HSP70s, (iv) significant sequence similarity between the N-terminal quadrant of HSP70 (from gram-positive bacteria and archaebacteria) and the m-type thioredoxin of plant chloroplasts. To account for these and other observations, a model for the evolution of HSP70 proteins involving gene duplication is proposed. The model proposes that HSP70 from archaebacteria (H. marismortui and M. mazei) and the gram-positive group of bacteria constitutes the ancestral form of the protein and that all other HSP70s (viz., other eubacteria as well as eukaryotes) containing the insert have evolved from this ancient protein.

Amino Acid Sequence↗

PubMed1992

Effects of low culture temperature on the induction of hsp70 mRNA and the accumulation of hsp70 and hsp105 in mouse FM3A cells.

We have shown that heat shock does not induce the synthesis of hsp70 in FM3A cells maintained at a low culture temperature of 33 degrees C although it does so in cells maintained at 37 degrees C [T. Hatayama et al. (1991) Biochem. Int. 24, 467-474]. In this paper, we show that FM3A cells maintained at 37 degrees C produced hsp70 mRNA during continuous heating at 42 degrees C or during postincubation at either 37 or 33 degrees C after being heated at 45 degrees C for 15 min, whereas cells maintained at 33 degrees C did not produce hsp70 mRNA during continuous heating at 37, 39, 42, or 45 degrees C, or during postincubation after being heated at any temperature. Thus the lack of hsp70 synthesis in cells maintained at 33 degrees C seemed to be due to the absence of hsp70 mRNA induction. Also, hsp70 was accumulated in cells maintained at 37 degrees C during continuous heating at 42 degrees C and during postincubation at 37 degrees C after heat shock at 45 degrees C, but not during postincubation at 33 degrees C. The cellular level of the constitutive hsp73 as well as the mRNA level were both similar in cells maintained at 33 and 37 degrees C. On the other hand, the cellular level of the constitutive hsp105 in cells maintained at 33 degrees C was only half of that in cells maintained at 37 degrees C. These hsp105 levels increased significantly in both types of cells after continuous heating at 39 degrees C. These findings indicate that the culture temperature affects not only the induction of hsp70 mRNA but also the accumulation of hsp70 and hsp105 in the cells.

Animals↗

PubMed1992

Syntenic conservation of HSP70 genes in cattle and humans.

A phage library of bovine genomic DNA was screened for hybridization with a human HSP70 cDNA probe, and 21 positive plaques were identified and isolated. Restriction mapping and blot hybridization analysis of DNA from the recombinant plaques demonstrated that the cloned DNAs were derived from three different regions of the bovine genome. One region contains two tandemly arrayed HSP70 sequences, designated HSP70-1 and HSP70-2, separated by approximately 8 kb of DNA. Single HSP70 sequences, designated HSP70-3 and HSP70-4, were found in two other genomic regions. Locus-specific probes of unique flanking sequences from representative HSP70 clones were hybridized to restriction endonuclease-digested DNA from bovine-hamster and bovine-mouse somatic cell hybrid panels to determine the chromosomal location of the HSP70 sequences. The probe for the tandemly arrayed HSP70-1 and HSP70-2 sequences mapped to bovine chromosome 23, syntenic with glyoxalase 1, 21 steroid hydroxylase, and major histocompatibility class I loci. HSP70-3 sequences mapped to bovine chromosome 10, syntenic with nucleoside phosphorylase and murine osteosarcoma viral oncogene (v-fos), and HSP70-4 mapped to bovine syntenic group U6, syntenic with amylase 1 and phosphoglucomutase 1. On the basis of these data, we propose that bovine HSP70-1,2 are homologous to human HSPA1 and HSPA1L on chromosome 6p21.3, bovine HSP70-3 is the homolog of an unnamed human HSP70 gene on chromosome 14q22-q24, and bovine HSP70-4 is homologous to one of the human HSPA-6,-7 genes on chromosome 1.

Animals↗

PubMed1992

The human heat shock protein hsp70 interacts with HSF, the transcription factor that regulates heat shock gene expression.

Transcriptional regulation of the human hsp70 gene in response to heat shock and other forms of physiological stress occurs through the activation of heat shock transcription factor (HSF). Exposure of cells to a heat shock temperature of 42 degrees C results in transient activation of HSF; its DNA-binding activity increases rapidly, plateaus, and attenuates, during which the intracellular levels of hsp70 increase. In an effort to understand whether HSF is regulated negatively by hsp70, we have examined whether HSF associates with hsp70. We show that activated HSF associates with hsp70 and that the interaction is detected as the levels of hsp70 increase in the cell. Addition of ATP and other hydrolyzable nucleotides results in the dissociation of hsp70 from HSF while nonhydrolyzable nucleotide analogs do not disrupt the complex. We demonstrate that exogenous recombinant wild-type hsp70 can associate with activated HSF, whereas no association is observed with an amino-terminal or a carboxy-terminal deletion mutant of hsp70. We also show that hsp70 blocks the in vitro activation of HSF from its cryptic non-DNA-binding state to a DNA-binding form; this inhibitory effect of hsp70 is abolished by ATP. We suggest that hsp70 may negatively regulate the activation of HSF.

Adenosine Triphosphate↗

PubMed1990

Structure and expression of the three MHC-linked HSP70 genes.

A duplicated locus encoding the major heat shock-induced protein HSP70 is located in the major histocompatibility complex (MHC) class III region 92 kilobases (kb) telomeric to the C2 gene. Nucleotide sequence analysis of the two intronless genes, HSP70-1 and HSP70-2, has shown that they encode an identical protein product of 641 amino acids. A third intronless gene, HSP70-Hom, has also been identified 4 kb telomeric to the HSP70-1 gene. This encodes a more basic protein of 641 amino acids which has 90% sequence similarity with HSP70-1. In order to investigate the expression of the three (MHC)-linked HSP70 genes individually by northern blot analysis, we have isolated locus-specific probes from the 3' untranslated regions of the genes. The HSP70-1 and HSP70-2 genes have been shown to be expressed at high levels as a approximately 2.4 kb mRNA in cells heat-shocked at 42 degrees C. HSP70-1 is also expressed constitutively at very low levels. The HSP70-Hom gene, which has no heat shock consensus sequence in its 5' flanking sequence, is expressed as a approximately 3 kb mRNA at low levels both constitutively and following heat shock.

Amino Acid Sequence↗

PubMed1992

The consequences of expressing hsp70 in Drosophila cells at normal temperatures.

In Drosophila cells, regulatory mechanisms not only act to provide rapid induction of hsp70 during heat shock but also to prevent expression at normal temperatures. To determine whether expression of hsp70 is detrimental to cells growing at normal temperatures, we used heterologous promoters to force expression of the protein in tissue culture cells and in larval salivary glands. Initially, constitutive expression of hsp70 substantially reduces the rate of cell growth. With continued expression, however, growth rates recover. At the same time, the intracellular distribution of hsp70 changes. Immediately after induction, the protein is diffusely distributed throughout the cell, but as growth resumes it coalesces into discrete points of high concentration, which we term hsp70 granules. hsp70 granules are also observed both in wild-type Drosophila tissue culture cells and in salivary glands after extended periods of recovery from heat shock. The protein in these granules appears to be irreversibly inactivated. It cannot be dispersed with a second heat shock, and cells containing these granules do not show thermotolerance. Only partial overlap between hsp70 granules and lysosomes indicates that the granules form independently of lysosomes. We conclude that expression of hsp70 is detrimental to growth at normal temperatures. We suggest that the change in hsp70 distribution, from diffuse to granular, represents a mechanism for controlling the protein's activity by sequestration.

Animals↗

PubMed1992

Analysis of topoisomerase I and II cleavage sites on the Drosophila actin and Hsp70 heat shock genes.

We have compared topoisomerase I and II cleavage sites on the actin 5C and 57A genes and the hsp70 genes in Drosophila Kc cells using the inhibitors camptothecin (topoisomerase I specific) and VM-26 (topoisomerase II specific) to assess the role of these enzymes in transcriptional regulation. Topoisomerase I cleavage sites were localized to the transcribed regions of the actin 5C and hsp70 genes and were present only when these genes were active. The actin 57A gene, shown previously to be inactive in Kc cells, had no detectable topoisomerase I cleavage sites. In contrast to topoisomerase I, topoisomerase II cleavage sites could be detected on transcriptionally active and inactive actin and hsp70 DNA sequences. Topoisomerase II cleavage sites on the inactive hsp70 gene were primarily localized to the very 5' end of the transcribed region of the gene. However, upon heat-induced activation of hsp70 transcription, topoisomerase II cleavage rapidly shifted from the 5' to the 3' end of the gene. Then, during the shutdown of hsp70 expression, there was a gradual reappearance of topoisomerase II cleavage at the 5' end of the gene that temporally correlated with the repression of hsp70 transcription. There was a similar preferential association of topoisomerase II with the 5' ends of transcriptionally repressed actin 5C and 57A genes. These results demonstrate that there are marked differences in how topoisomerases I and II interact with transcriptionally active and inactive regions of chromatin. In addition, we have identified an unusual type of topoisomerase II binding site that is preferentially associated with the 5' ends of inactive hsp70 and actin genes, suggesting that this enzyme may facilitate changes in chromatin structure that are associated with repression of gene transcription.

Actins↗

PubMed1992

Different thresholds of HSP70 and HSC70 heat shock mRNA induction in post-ischemic gerbil brain.

Thresholds of induction of heat shock protein (HSP) 70 and heat shock cognate protein (HSC) 70 mRNAs after transient global ischemia in gerbil brain were investigated by in situ hybridization using cloned cDNA probes selective for each mRNA species. In sham control brain, HSP70 mRNA was little present, while HSC70 mRNA was present in most cell populations. A 0.5-min occlusion of bilateral common carotid arteries did not affect the amount of HSP70 and HSC70 mRNAs. The selective induction of HSC70 mRNA was observed in dentate granule cells at 1 h, and in most cells of hippocampus especially dentate gyrus at 3 h after 1 min of ischemia when induction of HSP70 mRNA was not evident in the identical brain. The selective induction diminished by 2 days. However, after 2 min of ischemia, HSP70 and HSC70 mRNAs were induced together in hippocampal cells from 1 h of the reperfusion, and the co-induction prolonged in CA1 cells until 2 days. Body temperatures monitored at rectum increased after the reperfusion with a peak at 30 min. The degree of increase of the body temperature was significantly higher in the case after 2-min ischemia than in the cases after 0.5- and 1-min ischemia. Although HSP70 and HSC70 mRNAs are generally co-induced in stressful conditions, our results suggest the different thresholds of the induction between HSP70 and HSC70 mRNAs after transient brain ischemia. The selective induction of HSC70 mRNA which is not accompanied by the induction of HSP70 mRNA may relate to the differences of the duration of ischemia and the degree of the increase of body temperature after ischemia.

Animals↗

PubMed1992

Polymorphic analysis of the three MHC-linked HSP70 genes.

Three genes encoding members of the M(r) 70,000 heat shock protein family (HSP70) are known to lie in the class III region of the human major histocompatibility complex. In order to determine whether these genes or their protein products exhibit any polymorphism the three genes have been specifically amplified from genomic DNA and sequenced. The HSP70-1 and HSP70-2 genes encode the major heat-inducible HSP70. A comparison of the nucleotide sequences of these genes from B8, SC01, DR3, B18, F1C30, DR3, and B7, SC30, DR2 haplotypes has revealed only very limited sequence variation which is not associated with any amino acid polymorphism. The HSP70-Hom gene encodes a protein that is highly related to HSP70-1, but which is not heat-inducible. Nucleotide sequence analysis of this gene from different haplotypes has revealed a Met----Thr amino acid substitution at residue 493 in a number of the haplotypes tested. This variable amino acid lies in the proposed peptide-binding site of the HSP70-Hom protein.

Base Sequence↗

PubMed1992

Nuclear colocalization of c-myc protein and hsp70 in cells transfected with human wild-type and mutant c-myc genes.

Using immunofluorescence and electron microscopy we have studied the localization of wild-type and mutant c-myc proteins transiently expressed in CV-1 cells. In agreement with our previous observations, wild-type c-myc protein accumulated in large amorphous globules in the nucleus. All mutant proteins tested accumulated in the nucleus as well, but gave rise to morphologically different inclusion bodies. Many small globules appeared in cells transfected with D145-262 (deletion of amino acids 145-262), while cells transfected with D371-412 or D414-433 generated structures looking like a fine network or like beads on a string. In addition, a particulate cytoplasmic staining appeared in some cells transfected with the wild-type gene and in cells transfected with mutants D145-262 or D414-433. Since the c-myc protein has been reported to stimulate expression of exogenous hsp70 protein, we also examined the intracellular distribution of hsp70 in the transfected cells. Double immunofluorescence microscopy revealed that hsp70 codistributed with the c-myc protein in distinct globules in the nucleus of many but not all myc-positive cells. However, the levels of hsp70 transcripts were not significantly raised compared to nontransfected and vector-transfected cells. Likewise, the levels of hsp70 protein did not vary significantly. These findings indicate that overexpression of c-myc stimulates translocation of preexisting hsp70 from the cytoplasm into the nucleus, rather than influencing hsp70 expression. Conceivably, this may represent one of several mechanisms whereby the cell deals with excessive amounts of c-myc protein.

Animals↗

PubMed1992

Heat shock protein hsp70 protects cells from thermal stress even after deletion of its ATP-binding domain.

Retroviral-mediated gene transfer experiments show that rodent cells become heat resistant when stably and constitutively expressing a cloned human gene encoding an intact human 70-kDa heat shock protein (hsp70). Cells expressing higher levels of the hsp70 protein generally tolerate thermal stress better, whereas cells expressing either of two mutated hsp70-encoding genes, one with a 4-base pair out-of-frame deletion and one with an in-frame deletion of codons 438-618, are heat sensitive. These results provide strong evidence that expression of hsp70 leads directly to thermal tolerance. Surprisingly, cells expressing a mutant hsp70 of a human gene missing codons 120-428 are, nevertheless, heat resistant. Because the deleted region of this mutant contains the ATP-binding domain of human hsp70, this domain appears dispensable in the hsp70-mediated protection of cells from thermal stress.

Adenosine Triphosphate↗

PubMed1991

Human major HSP70 protein complements the localization and functional defects of cytoplasmic mutant SV40 T antigen in Swiss 3T3 mouse fibroblast cells.

The CT3 cytoplasmic localization mutant of SV40 T antigen is neither properly transported to the nucleus nor is it functional in rodent cells. Human precrisis cells are able to complement this mutation, as they are fully transformed by CT3 with wild-type efficiency. The human-specific factors responsible for this species-specific difference in response to CT3 were localized to human chromosome 6 by synteny in a panel of six somatic cell hybrids. A major human HSP70 heat shock protein located on chromosome 6 is expressed constitutively in human cells. Hsp70 proteins have been reported to play a role in intracellular movement of newly synthesized proteins. To test whether human HSP70 played a role in the complementation by human cells of the defect of CT3, we constructed a series of mouse cell lines expressing human HSP70 and tested them for their ability to localize CT3 T antigen in the nucleus and for their ability to be transformed by CT3 DNA. Mouse cell lines expressing human HSP70 protein were able to translocate mutant CT3 T antigen into the nucleus and were transformed by CT3 at rates comparable with wild-type SV40. Mouse-inducible HSP70 protein was not able to translocate cytoplasmic T antigen in Swiss 3T3 mouse fibroblast cells, even after heat shock. Apparently human HSP70 is capable of complementing directly or indirectly the structural and functional alterations in SV40 T antigen introduced by the CT3 mutation.

3T3 Cells↗

PubMed1992

Borrelia burgdorferi HSP70 homolog: characterization of an immunoreactive stress protein.

The gene encoding an immunoreactive Borrelia burgdorferi HSP70 homolog was isolated and characterized. The predicted amino acid sequence of this spirochetal protein confirms that this gene encodes a member of the HSP70 family of proteins. Although there appears to be a single copy of this gene on the spirochetal chromosome, two distinct transcripts hybridizing to the hsp70 probe are detected in RNA isolated from B. burgdorferi. The amount of spirochetal HSP70 RNA transcripts is shown to be thermally regulated. Antibodies in the serum of three Lyme arthritis patients and cloned T-cell lines isolated from one patient with Lyme arthritis recognize the expressed recombinant HSP70, indicating that it is an immunologically important spirochetal antigen. Antibodies in a rabbit antiserum, as well as antibodies in the serum of two of three Lyme arthritis patients examined, bound to expressed truncated recombinant HSP70s with 250 amino acids deleted from either the amino or carboxy terminus of the protein. However, antibodies in the serum of three Lyme arthritis patients, which were reactive with spirochetal HSP70, did not cross-react with human HSP70 proteins.

Adolescent↗

PubMed1991

Analysis of the specificity and mechanism of transcriptional activation of the human hsp70 gene during infection by DNA viruses.

We have examined the transcriptional regulation of the 70-kDa (70K) heat shock gene family following infection of human and monkey cells with four different DNA viruses: adenovirus type 5 (Ad5), herpes simplex virus type 1 (HSV-1), simian virus 40, and vaccinia virus. Our results indicate that induction of these genes is not a general response to the stress of viral infection but is instead a highly specific response, both with regard to the inducing virus and with regard to the target gene. Of three 70K heat shock genes examined, only hsp70 was induced during viral infection, and induction occurred only after infection by Ad5 and HSV-1. As revealed by genomic footprinting analysis, the mechanism of transcriptional activation of hsp70 during Ad5 or HSV-1 infection does not involve changes in the avidity of binding of basal transcription factors to the hsp70 promoter. In HSV-1-infected HeLa cells, transcriptional activation of hsp70 was quite transient, following which transcription was rapidly repressed; this was accompanied by the release of bound factors from the hsp70 promoter. In addition to the selectivity which characterizes the viral activation of hsp70 transcription, our results indicate that the consequences of this activation, as measured by changes in hsp70 mRNA levels and protein synthesis, are also virus specific.

Adenovirus Early Proteins↗

PubMed1991

Maturation of polycistronic pre-mRNA in Trypanosoma brucei: analysis of trans splicing and poly(A) addition at nascent RNA transcripts from the hsp70 locus.

Numerous protein-coding genes of the protozoan Trypanosoma brucei are arranged in tandem arrays that are transcribed polycistronically. The pre-mRNA transcripts are processed by trans splicing, leading to the addition of a capped 39-nucleotide (nt) miniexon and by poly(A) addition. We wished to determine the order of the RNA processing events at the hsp70 locus and address the potential occurrence of cotranscriptional RNA processing. We determined the rate of transcriptional elongation at the hsp70 locus in isolated nuclei, which measured between 20 and 40 nt/min. This low rate of RNA chain elongation allowed us to label the 3' end of hsp70 nascent RNA with a short (about 180-nt) 32P tail. The structure of the labeled nascent hsp70 RNA could then be analyzed by RNase T1 and RNase T1/RNase A mapping. We show that the trans splicing of hsp70 pre-mRNA did not occur immediately after the synthesis of the 3' splice acceptor site, and nascent RNA molecules that contained about 550 nt of RNA beyond the 3' splice acceptor site still had not acquired a miniexon. In contrast, nascent RNA with a 5' end that mapped to the polyadenylation site of the hsp70 genes could be detected, indicating that maturation of the pre-mRNA in trypanosomes involves a rapid cleavage of the nascent hsp70 RNA (within seconds after synthesis of the site) for poly(A) addition. Our data suggest that polycistronic pre-mRNA is unlikely to be synthesized in toto and rather appears to be processed cotranscriptionally by cleavage for poly(A) addition.

Animals↗

PubMed1991

The characterization and use of different antibodies against the hsp70 major heat shock protein family for the development of an immunoassay.

The hsp70 family of major stress proteins is composed of several different members exhibiting similar structural and functional properties. In order to obtain an antiserum with wide epitope reactivity, rabbits were immunized with a mixture of native and denatured hsp70 purified from bovine muscle by ATP-affinity chromatography. Screening for antibody specificity was performed by a "sandwich" enzyme linked immunosorbent assay (ELISA). Immunoprecipitation and immunoblotting analyses demonstrated that the polyclonal antiserum obtained by us and a monoclonal antibody raised against a different preparation of antigen recognized the same determinant on the native hsp70 molecule (inducible form). With a different specificity the polyclonal antiserum recognized only the denatured monomers of the other members of the hsp70 family. These results are discussed in relation to the immunological features of the hsp70 molecule and to the development of an immunoassay for the detection of hsp70 in cell and tissue extracts.

Antibody Specificity↗

PubMed1992

Major heat shock protein hsp70 protects tumor cells from tumor necrosis factor cytotoxicity.

Heat treatment and various other stresses render tumor cells resistant to cytotoxicity mediated by tumor necrosis factors (TNFs). Here, we elucidate the molecular basis of this phenomenon by demonstrating that the major heat shock protein, hsp70, protects tumor cells from TNF cytotoxicity even in the absence of stress. The human hsp70 gene was stably introduced into highly TNF-sensitive WEHI-S tumor cells both in the sense and antisense orientation. All clones constitutively expressing the exogenous human hsp70 gene were protected from TNF-mediated killing approximately 1000-fold. Remarkably, the growth of one clone was actually stimulated by low concentrations of TNF. Moreover, a clone expressing antisense hsp70 RNA was rendered extremely sensitive to TNFs. Hsp70-mediated protection from TNF cytotoxicity was confirmed in transient expression experiments employing retroviral vectors. Changes in cellular sensitivity to TNF were not associated with alterations in the binding of TNF to its receptors. Neither the transfection procedure itself nor overexpression of the low molecular weight heat shock protein, hsp27, had any effect on cellular susceptibility to TNFs. Our data suggest that hsp70 may increase the oncogenic potential of some tumor cells by providing them with an escape mechanism from immunological defense.

Animals↗

PubMed1992

Expression of heat shock genes (hsp70) in the rabbit spinal cord: localization of constitutive and hyperthermia-inducible mRNA species.

We have previously reported that hyperthermia induces the expression of a heat shock gene in the rabbit brain (Sprang and Brown, Mol Brain Res 3:89-93, 1987). Striking regional and cell type differences in the pattern of induction of the hsp70 mRNA were noted. Tissue injury also induces the rapid induction of hsp70 mRNA in the mammalian brain (Brown et al., Neuron 2:1559-1564, 1989). In the present study, in situ hybridization with 35S-labelled riboprobes specific for constitutive and inducible hsp70 mRNA species was employed to investigate the effect of fever-like temperatures on hsp70 gene expression in the rabbit spinal cord. Expression of constitutive hsp70 mRNA was detected in large motor neurons of both control and hyperthermic animals. Within 1 hr after hyperthermia, a massive induction of inducible hsp70 mRNA was noted in fibre tracts of the spinal cord, a pattern consistent with a strong glial response to heat shock. Induction was not observed in the large motor neurons.

Animals↗