DnaK/DnaJ chaperone system reactivates endogenous E coli thermostable FBP aldolase in vivo and in vitro; the effect is enhanced by GroE heat shock proteins

At least four proteins of 70,000 to 75,000 molecular weight (70-75K) were synthesized from mRNA which hybridized with a cloned heat shock gene previously shown to be localized to the 87A and 87C heat shock puff sites. These in vitro-synthesized proteins were indistinguishable from in vivo-synthesized heat shock-induced proteins when analyzed on sodium dodecyl sulfate-polyacrylamide gels. A comparison of the pattern of this group of proteins synthesized in vivo during a 5-min pulse or during continuous labeling indicates that the 72-75K proteins are probably not kinetic precursors to the major 70K heat shock protein. Partial digestion products generated with V8 protease indicated that the 70-75K heat shock proteins are closely related, but that there are clear differences between them. The partial digestion patterns obtained from heat shock proteins from the Kc cell line and from the Oregon R strain of Drosophila melanogaster are very similar. Genetic analysis of the patterns of 70-75K heat shock protein synthesis indicated that the genes encoding at least two of the three 72-75K heat shock proteins are located outside of the major 87A and 87C puff sites.

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Protective Effect of Post-Ischaemic Viral Delivery of Heat Shock Proteins in vivo

Journal of Cerebral Blood Flow & Metabolism ◽

10.1038/jcbfm.2008.106 ◽

2008 ◽

Vol 29 (2) ◽

pp. 254-263 ◽

Cited By ~ 20

Author(s):

Romina A Badin ◽

Michael Modo ◽

Mike Cheetham ◽

David L Thomas ◽

David G Gadian ◽

...

Keyword(s):

Heat Shock ◽

Heat Shock Proteins ◽

Viral Vector ◽

Lesion Size ◽

In Vivo Studies ◽

Current Evidence ◽

Time Points ◽

Shock Proteins

Heat shock proteins (HSPs) function as molecular chaperones involved in protein folding, transport and degradation and, in addition, they can promote cell survival both in vitro and in vivo after a range of stresses. Although some in vivo studies have suggested that HSP27 and HSP70 can be neuroprotective, current evidence is limited, particularly when HSPs have been delivered after an insult. The effect of overexpressing HSPs after transient occlusion of the middle cerebral artery in rats was investigated by delivering an attenuated herpes simplex viral vector (HSV-1) engineered to express HSP27 or HSP70 30 mins after tissue reperfusion. Magnetic resonance imaging scans were used to determine lesion size and cerebral blood flow at six different time points up to 1 month after stroke. Animals underwent two sensorimotor tests at the same time points to assess the relationship between lesion size and function. Results indicate that post-ischaemic viral delivery of HSP27, but not of HSP70, caused a statistically significant reduction in lesion size and induced a significant behavioural improvement compared with controls. This is the first evidence of effective post-ischaemic gene therapy with a viral vector expressing HSP27 in an experimental model of stroke.

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Exertional heat injury and gene expression changes: a DNA microarray analysis study

Journal of Applied Physiology ◽

10.1152/japplphysiol.00886.2003 ◽

2004 ◽

Vol 96 (5) ◽

pp. 1943-1953 ◽

Cited By ~ 75

Author(s):

Larry A. Sonna ◽

C. Bruce Wenger ◽

Scott Flinn ◽

Holly K. Sheldon ◽

Michael N. Sawka ◽

...

Keyword(s):

Gene Expression ◽

Stress Response ◽

Heat Shock ◽

Heat Shock Proteins ◽

Molecular Evidence ◽

Peripheral Blood Mononuclear ◽

Heat Injury ◽

Shock Proteins

This study examined gene expression changes associated with exertional heat injury (EHI) in vivo and compared these changes to in vitro heat shock responses previously reported by our laboratory. Peripheral blood mononuclear cell (PBMC) RNA was obtained from four male Marine recruits (ages 17-19 yr) who presented with symptoms consistent with EHI, core temperatures ranging from 39.3 to 42.5°C, and elevations in serum enzymes such as creatine kinase. Controls were age- and gender-matched Marines from whom samples were obtained before and several days after an intense field-training exercise in the heat (“The Crucible”). Expression analysis was performed on Affymetrix arrays (containing ∼12,600 sequences) from pooled samples obtained at three times for EHI group (at presentation, 2-3 h after cooling, and 1-2 days later) and compared with control values (average signals from two chips representing pre- and post-Crucible samples). After post hoc filtering, the analysis identified 361 transcripts that had twofold or greater increases in expression at one or more of the time points assayed and 331 transcripts that had twofold or greater decreases in expression. The affected transcripts included sequences previously shown to be heat-shock responsive in PBMCs in vitro (including both heat shock proteins and non-heat shock proteins), a number of sequences whose changes in expression had not previously been noted as a result of in vitro heat shock in PBMCs (including several interferon-induced sequences), and several nonspecific stress response genes (including ubiquitin C and dual-specificity phosphatase-1). We conclude that EHI produces a broad stress response that is detectable in PBMCs and that heat stress per se can only account for some of the observed changes in transcript expression. The molecular evidence from these patients is thus consistent with the hypothesis that EHI can result from cumulative effects of multiple adverse interacting stimuli.

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Mammalian lymphocytes: stress-induced synthesis of heat-shock proteins in vitro and in vivo

Canadian Journal of Biochemistry and Cell Biology ◽

10.1139/o85-089 ◽

1985 ◽

Vol 63 (7) ◽

pp. 711-722 ◽

Cited By ~ 23

Author(s):

David Rodenhiser ◽

Jack H. Jung ◽

Burr G. Atkinson

Keyword(s):

Heat Shock ◽

Heat Shock Proteins ◽

White Blood Cells ◽

Sodium Dodecyl ◽

Molecular Masses ◽

Febrile Episodes ◽

Shock Proteins ◽

Mouse Lymphocytes

Mammalian (human, mouse, and rabbit) white blood cells (lymphocytes) maintained in culture respond to a brief incubation at an elevated temperature (at or above 41 °C) by (i) the new and (or) enhanced synthesis of a small number of proteins (the so-called heat-shock proteins; HSPs) having molecular masses of approximately 110 000, 100 000, 90 000, 70 000, 65 000, and 26 000 daltons and (ii) the depressed synthesis of proteins normally made at 37 °C. The HSPs synthesized in culture by human, rabbit, and mouse (peripheral and splenic) lymphocytes are similar in number, molecular mass, and distribution on two-dimensional (isoelectric focusing and sodium dodecyl sulfate – polyacrylamide) electrophoretic gels to those synthesized in vivo by lymphocytes in hyperthermic mice. Since the level of hyperthermia used to induce HSP synthesis in mouse lymphocytes in vitro and in vivo is of a magnitude (41 °C) also used to promote thermotolerance in mice and is similar to temperatures attained during febrile episodes in rabbits and in humans, we suggest that the in vitro and in vivo synthesis of HSPs by mouse lymphocytes, demonstrated in this study, represents a relevant, physiological response which mammalian lymphocytes may normally use to survive periods of thermal stress.

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Multiple layers of control govern expression of the Escherichia coli ibpAB heat-shock operon

Microbiology ◽

10.1099/mic.0.043802-0 ◽

2011 ◽

Vol 157 (1) ◽

pp. 66-76 ◽

Cited By ~ 35

Author(s):

Lena C. Gaubig ◽

Torsten Waldminghaus ◽

Franz Narberhaus

Keyword(s):

Escherichia Coli ◽

Heat Shock ◽

Transcriptional Control ◽

Rnase E ◽

Translation Control ◽

E Coli ◽

Complex Process ◽

Shock Proteins

The Escherichia coli ibpAB operon encodes two small heat-shock proteins, the inclusion-body-binding proteins IbpA and IbpB. Here, we report that expression of ibpAB is a complex process involving at least four different layers of control, namely transcriptional control, RNA processing, translation control and protein stability. As a typical member of the heat-shock regulon, transcription of the ibpAB operon is controlled by the alternative sigma factor σ 32 (RpoH). Heat-induced transcription of the bicistronic operon is followed by RNase E-mediated processing events, resulting in monocistronic ibpA and ibpB transcripts and short 3′-terminal ibpB fragments. Translation of ibpA is controlled by an RNA thermometer in its 5′ untranslated region, forming a secondary structure that blocks entry of the ribosome at low temperatures. A similar structure upstream of ibpB is functional in vitro but not in vivo, suggesting downregulation of ibpB expression in the presence of IbpA. The recently reported degradation of IbpA and IbpB by the Lon protease and differential regulation of IbpA and IbpB levels in E. coli are discussed.

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Identification of mRNAs encoding low molecular mass heat-shock proteins in maize (Zea mays L.)

Canadian Journal of Genetics and Cytology ◽

10.1139/g86-154 ◽

1986 ◽

Vol 28 (6) ◽

pp. 1106-1114 ◽

Cited By ~ 2

Author(s):

C. A. B. Rees ◽

N. C. Hogan ◽

D. B. Walden ◽

B. G. Atkinson

Keyword(s):

Heat Shock ◽

Heat Shock Proteins ◽

Molecular Mass ◽

Polyclonal Antibodies ◽

Relative Molecular Mass ◽

Temperature Heat ◽

Free Translation ◽

Shock Proteins

Subjecting 5-day-old maize seedlings to a rapid elevation in growth temperature (heat shock; 25–42 °C) results in a shift in the pattern of protein synthesis in maize plumules from the production of a broad spectrum of proteins to the new and (or) enhanced synthesis of a small number of heat-shock proteins (HSPs). The low relative molecular mass (Mr) HSPs, and more specifically an 18-kDa HSP with four major isoelectric variants, represent the majority of HSP synthesis following cell-free translation of total cellular poly (A)+ RNAs and polyribosomal RNAs extracted from heat-shocked plumules. Immunochemical studies, using polyclonal antibodies raised against the 18-kDa HSPs, show that the 18-kDa HSPs synthesized in vitro share immunochemical properties with HSPs of the same Mr synthesized in vivo by heat-shocked plumules. Furthermore, size fractionation and translation analyses of total cellular poly(A)+ RNAs extracted from heat-shocked plumules demonstrate that poly(A)+ RNAs encoding an 18-kDa HSP(s) have an estimated size of 0.6–0.95 kilobases. The observation that 18-kDa HSPs are absent among the translation products and immunoprecipitates of proteins synthesized in vitro by RNAs extracted from control plumules (25 °C) suggests that the mRNAs encoding 18-kDa HSPs are heat-shock induced.Key words: mRNA, maize, heat shock.

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Expression of heat shock proteins and heat shock protein messenger ribonucleic acid in human prostate carcinoma in vitro and in tumors in vivo

Cell Stress and Chaperones ◽

10.1379/csc-44r.1 ◽

2005 ◽

Vol 10 (1) ◽

pp. 46 ◽

Cited By ~ 112

Author(s):

Dan Tang ◽

Md Abdul Khaleque ◽

Ellen L. Jones ◽

Jimmy R. Theriault ◽

Cheng Li ◽

...

Keyword(s):

Heat Shock ◽

Heat Shock Proteins ◽

Heat Shock Protein ◽

Prostate Carcinoma ◽

Ribonucleic Acid ◽

Messenger Ribonucleic Acid ◽

Human Prostate Carcinoma ◽

Shock Proteins

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Identification of CHIP, a Novel Tetratricopeptide Repeat-Containing Protein That Interacts with Heat Shock Proteins and Negatively Regulates Chaperone Functions

Molecular and Cellular Biology ◽

10.1128/mcb.19.6.4535 ◽

1999 ◽

Vol 19 (6) ◽

pp. 4535-4545 ◽

Cited By ~ 612

Author(s):

Carol A. Ballinger ◽

Patrice Connell ◽

Yaxu Wu ◽

Zhaoyong Hu ◽

Larry J. Thompson ◽

...

Keyword(s):

Heat Shock ◽

Heat Shock Proteins ◽

Atpase Activity ◽

Substrate Binding ◽

Tetratricopeptide Repeat ◽

Forward Reaction ◽

Interacting Protein ◽

Shock Proteins

ABSTRACT The chaperone function of the mammalian 70-kDa heat shock proteins Hsc70 and Hsp70 is modulated by physical interactions with four previously identified chaperone cofactors: Hsp40, BAG-1, the Hsc70-interacting protein Hip, and the Hsc70-Hsp90-organizing protein Hop. Hip and Hop interact with Hsc70 via a tetratricopeptide repeat domain. In a search for additional tetratricopeptide repeat-containing proteins, we have identified a novel 35-kDa cytoplasmic protein, carboxyl terminus of Hsc70-interacting protein (CHIP). CHIP is highly expressed in adult striated muscle in vivo and is expressed broadly in vitro in tissue culture. Hsc70 and Hsp70 were identified as potential interaction partners for this protein in a yeast two-hybrid screen. In vitro binding assays demonstrated direct interactions between CHIP and both Hsc70 and Hsp70, and complexes containing CHIP and Hsc70 were identified in immunoprecipitates of human skeletal muscle cells in vivo. Using glutathione S-transferase fusions, we found that CHIP interacted with the carboxy-terminal residues 540 to 650 of Hsc70, whereas Hsc70 interacted with the amino-terminal residues 1 to 197 (containing the tetratricopeptide domain and an adjacent charged domain) of CHIP. Recombinant CHIP inhibited Hsp40-stimulated ATPase activity of Hsc70 and Hsp70, suggesting that CHIP blocks the forward reaction of the Hsc70-Hsp70 substrate-binding cycle. Consistent with this observation, both luciferase refolding and substrate binding in the presence of Hsp40 and Hsp70 were inhibited by CHIP. Taken together, these results indicate that CHIP decreases net ATPase activity and reduces chaperone efficiency, and they implicate CHIP in the negative regulation of the forward reaction of the Hsc70-Hsp70 substrate-binding cycle.

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