In this mini-review, the role of heat shock proteins in susceptability to induction of atrial fibrillation (AF) or in the process of AF is discussed. AF is the most common arrhythmia in humans, is self-perpetuating in nature and hence tends to become more persistent in time. Some studies show a correlation between high Hsp70 (HspA1A) expression in cardiac tissue and a reduced susceptability to induction of postoperative AF. Expression of Hsp70, Hsc70 (HspA8), Hsp40 (DnaJB1), Hsp60 (HspD1), Hsp90 (HspC1) was not associated with progression of AF. However, both correlative studies in human and experimental studies suggest that Hsp27 (HspB1) may delay progression of AF to the more permanent forms and hence Hsp27 might be referred to as a “Beat shock protein”.

Forty-five Sprague-Dawley rats (60–80 days old) were randomly placed into one of three groups: sedentary pregnant control (PC); prepregnancy trained animals that exercised throughout pregnancy (PR); and nonpregnant trained animals (NPR). Each exercising animal ran at approximately 60–70% aerobic capacity (V̇O_2max) for 1 hour/day up to and including day 18 of gestation (term = 21 days). On day 20 of gestation, fetuses were excised from each pregnant animal and scrutinized for gross abnormalities. In 3 randomly chosen fetuses from each litter, brain, heart, kidney, hind limb, and placental tissues were removed to assess the accumulation of the inducible isoform of the 70-kilodalton heat shock protein (Hsp 72i). No significant differences were detected between fetal hearts, hind limbs, or placental tissues of PC or PR groups. No Hsp 72i signal could be detected in fetal kidney or brain tissues from either pregnant group. Results indicate that maternal core temperature did not reach the threshold that would induce either gross fetal abnormalities or a fetal heat shock protein response. However, fetal and placental growth was reduced by the exercise protocol.

The meeting on “Investigating cellular stress responses—a multidisciplinary approach from basic science to therapeutics” was held in London on 13 October 2006. The purpose of this 1-day meeting was to bring together European scientists investigating the immune biology of stress proteins and their potential clinical applications. The main topics included: the role of heat shock proteins (Hsps) in bacterial infections; the role of Hsps with a molecular mass of about 70 kDa in cancer therapy and in prediction of the clinical outcome following allogeneic hematopoietic stem cell transplantation; the quality and duration of stress as a danger signal for the initiation of a stress response; the mechanism of Hsp-protein interaction; and Hsp export from tumor cells in secretory granules.

Heat shock proteins (Hsp) are families of highly conserved molecules and immunodominant antigens in some infections and in autoimmune diseases. Some reports suggest that different regions of the Hsp60 molecule induce distinct immune responses. However, there are no reports comparing physiological T-cell reactivity to Hsp60 in mice. In this study, we have analyzed T-cell proliferation and cytokine production induced by Hsp60, under physiological conditions, in three mouse strains bearing distinct major histocompatibility complex (MHC) backgrounds. Proliferative response predominantly was found in C57BL/6 mice, mostly induced by N-terminal and intermediate Hsp60 peptides (P < 0.0001). Interferon-γ (IFNγ) production was broadly induced by different regions of Hsp60 in all three mouse strains, although response was focused in different peptide groups in each strain. We did not observe an exclusive Th1 or Th2 cytokine profile induced by any particular region of Hsp60. However, we identified a strain hierarchy in IL-10 production induced by Hsp60 peptides from different regions, mostly detected in C3H/HePas, and in BALB/c, but not in C57BL/6 mice. In contrast, IL-4 production only was induced by the intermediate and C-terminal region peptides in both C3H/HePas and BALB/c mice. Our data give original information on physiological cellular reactivity to Hsp60. We also have identified peptides with the capacity to induce the production of anti-inflammatory cytokines, bringing perspectives for their use in immunotherapy of chronic inflammatory diseases and allograft rejection.

Many bacteria possess 2 or more genes for the chaperonin GroEL and the cochaperonin GroES. In particular, rhizobial species often have multiple groEL and groES genes, with a high degree of amino-acid similarity, in their genomes. The Rhizobium leguminosarum strain A34 has 3 complete groE operons, which we have named cpn.1, cpn.2 and cpn.3. Previously we have shown the cpn.1 operon to be essential for growth, but the two other cpn operons to be dispensable. Here, we have investigated the extent to which loss of the essential GroEL homologue Cpn60.1 can be compensated for by expression of the other two GroEL homologues (Cnp60.2 and Cpn60.3). Cpn60.2 could not be overexpressed to high levels in R. leguminosarum, and was unable to replace Cpn60.1. A strain that overexpressed Cpn60.3 grew in the absence of Cpn60.1, but the complemented strain displayed a temperature-sensitive phenotype. Cpn60.1 and Cpn60.3, when coexpressed in Escherichia coli, preferentially selfassembled rather than forming mixed heteroligomers. We conclude that, despite their high amino acid similarity, the GroEL homologues of R. leguminosarum are not functionally equivalent in vivo.

Heat shock proteins (Hsps) are molecular chaperones that aid in protein synthesis and trafficking and have been shown to protect cells/tissues from various protein damaging stressors. To determine the extent to which a single heat stress and the concurrent accumulation of Hsps influences the early events of skeletal muscle hypertrophy, Sprague-Dawley rats were heat stressed (42°C, 15 minutes) 24 hours prior to overloading 1 plantaris muscle by surgical removal of the gastrocnemius muscle. The contralateral plantaris muscles served as controls. Heat-stressed and/or overloaded plantaris muscles were assessed for muscle mass, total muscle protein, muscle protein concentration, Type I myosin heavy chain (Type I MHC) content, as well as Hsp72 and Hsp25 content over the course of 7 days following removal of the gastrocnemius muscle. As expected, in non–heat-stressed animals, muscle mass, total muscle protein and MHC I content were significantly increased (P < 0.05) following overload. In addition, Hsp25 and Hsp72 increased significantly after 2 and 3 days of overload, respectively. A prior heat stress–elevated Hsp25 content to levels similar to those measured following overload alone, but heat stress–induced Hsp72 content was increased significantly greater than was elicited by overload alone. Moreover, overloaded muscles from animals that experienced a prior heat stress showed a lower muscle mass increase at 5 and 7 days; a reduced total muscle protein elevation at 3, 5, and 7 days; reduced protein concentration; and a diminished Type I MHC content accumulation at 3, 5, and 7 days relative to non– heat-stressed animals. These data suggest that a prior heat stress and/or the consequent accumulation of Hsps may inhibit increases in muscle mass, total muscle protein content, and Type I MHC in muscles undergoing hypertrophy.

Atrial fibrillation (AF) is the most common sustained arrhythmia. Because heat shock proteins (Hsp) can protect cells from stress, we compared the levels of Hsp60, Hsp72, Hsc73, and Hsp27 in atrial myocardium from 17 patients with AF (8 paroxysmal and 9 persistent) and 7 controls in sinus rhythm (SR). Hsp60, Hsp72, and Hsc73 levels were not significantly different among the 3 groups. Hsp27 expression was slightly higher in paroxysmal AF than in SR and in persistent AF, and a borderline significant difference (P = 0.064) was seen between the paroxysmal and persistent AF subgroups. Hsp60 levels in the moderate, severe, and profound myolysis groups were significantly lower than the light myolysis group, but no differences were found in other Hsps. In summary, the data indicate that expression of Hsp27 and Hsc73 may be associated with different stages of AF and that Hsp60 also may be associated with the degree of atrial myolysis.

Induction of heat shock proteins (Hsps) is under investigation as treatment for neurodegenerative disorders, yet many types of neurons, including motor neurons that degenerate in amyotrophic lateral sclerosis (ALS), have a high threshold for activation of the major transcription factor mediating stress-induced Hsp upregulation, heat shock transcription factor 1 (Hsf1). Hsf1 is tightly regulated by a series of inhibitory checkpoints that include sequestration in multichaperone complexes governed by Hsp90. This study examined the role of multichaperone complexes in governing the heat shock response in motor neurons. Hsp90 inhibitors induced expression of Hsp70 and Hsp40 and transactivation of a human inducible hsp70 promoter–green fluorescent protein (GFP) reporter construct in motor neurons of dissociated spinal cord–dorsal root ganglion (DRG) cultures. On the other hand, overexpression of activator of Hsp90 adenosine triphosphatase ([ATPase 1], Aha1), which should mobilize Hsf1 by accelerating turnover of mature, adenosine triphosphate–(ATP) bound Hsp90 complexes, and death domain–associated protein (Daxx), which in cell lines has been shown to promote transcription of heat shock genes by relieving inhibition exerted by interactions between nuclear Hsp90/multichaperone complexes and trimeric Hsf1, failed to induce Hsps in the absence or presence of heat shock. These results indicate that disruption of multichaperone complexes alone is not sufficient to activate the neuronal heat shock response. Furthermore, in motor neurons, induction of Hsp70 by Hsp90-inhibiting drugs was prevented by overexpression of wild-type Hsf1, contrary to what would be expected for a classical Hsf1-mediated pathway. These results point to additional differences in regulation of hsp genes in neuronal and nonneuronal cells.

The cytoskeleton has a unique property such that changes of conformation result in polymerization into a filamentous form. αB-Crystallin, a small heat shock protein (sHsp), has chaperone activities for various substrates, including proteins constituting the cytoskeleton, such as actin; intermediate filament; and tubulin. However, it is not clear whether the “α-crystallin domain” common to sHsps also has chaperone activity for the protein cytoskeleton. To investigate the possibility that the C-terminal α-crystallin domain of αB-crystallin has the aggregation-preventing ability for tubulin, we constructed an N-terminal domain deletion mutant of αB-crystallin. We characterized its structural properties and chaperone activities. Far-ultraviolet (UV) circular dichroism measurements showed that secondary structure in the α-crystallin domain of the deletion mutant is maintained. Ultracentrifuge analysis of molecular masses indicated that the deletion mutant formed smaller oligomers than did the full-length protein. Chaperone activity assays demonstrated that the N-terminal domain deletion mutant suppressed heat-induced aggregation of tubulin well. Comparison of chaperone activities for 2 other substrates (citrate synthase and alcohol dehydrogenase) showed that it was less effective in the suppression of their aggregation. These results show that αB-crystallin recognizes a variety of substrates and especially that α-crystallin domain binds free cytoskeletal proteins. We suggest that this feature would be advantageous in its functional role of holding or folding multiple proteins denatured simultaneously under stress conditions.

We have identified 24 members of the DnaK subfamily of heat shock 70 proteins (Hsp70s) in the complete genomes of 5 diverse photosynthetic eukaryotes. The Hsp70s are a ubiquitous protein family that is highly conserved across all domains of life. Eukaryotic Hsp70s are found in a number of subcellular compartments in the cell: cytoplasm, mitochondrion (MT), chloroplast (CP), and endoplasmic reticulum (ER). Although the Hsp70s have been the subject of intense study in model organisms, very little is known of the Hsp70s from early diverging photosynthetic lineages. The sequencing of the complete genomes of Thalassiosira pseudonana (a diatom), Cyanidioschyzon merolae (a red alga), and 3 green algae (Chlamydomonas reinhardtii, Ostreococcus lucimarinus, Ostreococcus tauri) allow us to conduct comparative genomics of the Hsp70s present in these diverse photosynthetic eukaryotes. We have found that the distinct lineages of Hsp70s (MT, CP, ER, and cytoplasmic) each have different evolutionary histories. In general, evolutionary patterns of the mitochondrial and endoplasmic reticulum Hsp70s are relatively stable even among very distantly related organisms. This is not true of the chloroplast Hsp70s and we discuss the distinct evolutionary patterns between “green” and “red” plastids. Finally, we find that, in contrast to the angiosperms Arabidopsis thaliana and Oryza sativa that have numerous cytoplasmic Hsp70, the 5 algal species have only 1 cytoplasmic Hsp70 each. The evolutionary and functional implications of these differences are discussed.

Cells are continuously exposed to environmental stresses and respond to them to maintain cellular homeostasis. Failure to respond to these stresses may cause pathological states such as renal failure, complications of diabetes, and autoimmune diseases. Signal transduction induced by osmotic and cold stresses is not fully understood. In addition, mechanisms of these stress responses are yet to be elucidated. Activation of many signaling pathways induces translocation of proteins into the nucleus to transduce signals and regulate nuclear functions. By using inducible translocation trap (ITT), a reporter gene–based screening technique, nuclear translocation of 2-amino-3-ketobutyrate coenzyme A ligase (KBL) was detected in response to cold and osmotic stresses. Rapid nuclear translocation of KBL was confirmed by biochemical analysis and fluorescent microscopy. A large region of KBL was required for stress-induced nuclear translocation. The KBL reporter system will be a useful tool for the investigation of cold and osmotic stress responses.

Previous work in our laboratory demonstrated the existence of an association between heat shock transcription factor 2 (HSF2) and the serine/threonine phosphatase 2A, which is mediated by interaction between HSF2 and the A subunit (also called PR65) of this protein phosphatase. In light of the importance of HSF2-PP2A association for HSF2 cellular function, in this study, we have sought to dissect the sequences within HSF2 that are important for interaction with the A subunit of PP2A. The results of these experiments indicate that the HSF2 region comprising amino acids 343–363 is important for A subunit interaction. This region includes part of the C-terminal leucine zipper motif of HSF2 called heptad repeat C (HR-C). The results of transfection/immunoprecipitation experiments also show that deletion of the 6 amino acids from 343 to 348 from HSF2 (HSF2 (Δ343–348)), is sufficient to prevent HSF2 from interacting with PP2A. These data provide insight into a new functional domain of HSF2, the PP2A A subunit-interacting region.