Heat shock proteins in autoimmune diseases

Heat shock proteins (hsp's) are among the most conserved proteins in evolution. They have been identified as important pathogen-related antigens as well as autoantigens suitable for construction of novel vaccines. The high evolutionary homology of hsp's has raised the question about the safety of such vaccines. Experimental and clinical observations have confirmed that hsp proteins are involved in the regulation of some autoimmune disease such as autoimmune arthritis, type 1 diabetes mellitus, atherosclerosis, multiple sclerosis, and other autoimmune reactions. It has been shown in experimental animals that some hsp proteins (especially hsp60, hsp70, and hsp10) can either induce or prevent autoimmune reactions depending on the circumstances. This article discusses the involvement of hsp proteins in the etiology of autoimmune diseases and it presents promising experimental data on the effects of immunization with hsp proteins in the prevention and therapy of autoimmune diseases.     

Cellular localization of Drosophila 83-kilodalton heat shock protein in normal, heat-shocked, and recovering cultured cells with a specific antibody

To gain insight on the possible functions of heat shock proteins (hsp's) in Drosophila, we have purified the 83-kilodalton hsp (hsp 83) from cultured cells and studied its intracellular localization by immunofluorescence in normal, heat-shocked, and recovering cells. The specificity of the antibody was assessed by one- and two-dimensional gel immunoblotting and by partial proteolytic digestion. The anti-hsp 83 antibody does not show any significant cross-reactivity with hsp's of different avian or mammalian cell lines, but cross-reacts with hsp's of similar molecular masses in other dipteran insects. The partial proteolytic peptide maps of Drosophila hsp 83 differ from those of mouse hsp 89 and chicken hsp 84. Immunoblotting of Drosophila Kc cells heat shocked at different temperatures indicates a maximal expression of hsp 83 at 33 degrees C. By immunofluorescence, hsp 83 is shown to have a strictly cytoplasmic localization. In unstressed cells, it is distributed in the entire cytoplasm with a slight enrichment in the perinuclear region. After heat shock, it seems to concentrate at the cell periphery close to the plasma membrane and it gradually redistributes to the whole cytoplasm during cellular recovery at normal temperatures.     

Expression of the murine small heat shock proteins hsp 25 and alpha B crystallin in the absence of stress

Stress induces the synthesis of several large and small heat shock proteins (hsp's). Two related small hsp's, hsp25 and alpha B crystallin exist in mice. alpha B crystallin is an abundant protein in several tissues even in the absence of stress. Particularly high amounts accumulate in the eye lens. Here we show that hsp25 is likewise constitutively expressed in many normal adult tissues. In the absence of stress the protein is most abundant in the eye lens, heart, stomach, colon, lung, and bladder. The stress-independent expression pattern of the two small hsp's is distinct. In several tissues the amount of hsp25 exceeds that accumulating in NIH 3T3 fibroblasts in response to heat stress. hsp25, like alpha B crystallin, exists in a highly aggregated form in the eye lens. The expression of hsp25 and alpha B crystallin in normal tissues suggests an essential, but distinct function of the two related proteins under standard physiological conditions.     

Heat shock and developmental regulation of the Drosophila melanogaster hsp83 gene.

In contrast to the hsp70 gene, whose expression is normally at a very low level and increases by more than 2 orders of magnitude during heat shock, the hsp83 gene in Drosophila melanogaster is expressed at high levels during normal development and increases only severalfold in response to heat shock. Developmental expression of the hsp83 gene consists of a high level of tissue-general, basal expression and a very high level of expression in ovaries. We identified regions upstream of the hsp83 gene that were required for its developmental and heat shock-induced expression by assaying beta-galactosidase activity and mRNA levels in transgenic animals containing a series of 5' deletion and insertion mutations of an hsp83-lacZ fusion gene. Deletion of sequences upstream of the overlapping array of a previously defined heat shock consensus (HSC) sequence eliminated both forms of developmental expression of the hsp83 gene. As a result, the hsp83 gene with this deletion mutation was regulated like that of the hsp70 gene. Moreover, an in vivo polymer competition assay revealed that the overlapping HSC sequences of the hsp83 gene and the nonoverlapping HSC sequences of the hsp70 gene had similar affinities for the factor required for heat induction of the two heat shock genes. We discuss the functional similarity of hsp70 and hsp83 heat shock regulation in terms of a revised view of the heat shock-regulatory sequence.     

Time course and magnitude of synthesis of heat-shock proteins in congeneric marine snails (Genus tegula) from different tidal heights

The time course and magnitude of the heat-shock response in relation to severity of thermal stress are important, yet poorly understood, aspects of thermotolerance. We examined patterns of protein synthesis in congeneric marine snails (genus Tegula) that occur at different heights along the subtidal to intertidal gradient after a thermal exposure (30 degrees C for 2.5 h, followed by 50 h recovery at 13 degrees C) that induced the heat-shock response. We monitored the kinetics and magnitudes of protein synthesis by quantifying incorporation of 35S-labeled methionine and cysteine into newly synthesized proteins and observed synthesis of putative heat-shock proteins (hsp's) of size classes 90, 77, 70, and 38 kDa. In the low- to mid-intertidal species, Tegula funebralis, whose body temperature frequently exceeds 30 degrees C during emersion, synthesis of hsp's commenced immediately after heat stress, reached maximal levels 1-3 h into recovery, and returned to prestress levels by 6 h, except for hsp90 (14 h). In contrast, in the low-intertidal to subtidal species, Tegula brunnea, for which 2.5 h at 30 degrees C represents a near lethal heat stress, synthesis of hsp's commenced 2-14 h after heat stress; reached maximal levels after 15-30 h, which exceeded magnitudes of synthesis in T. funebralis; and returned to prestress levels in the case of hsp90 (50 h) and hsp77 (30 h) but not in the case of hsp70 and hsp38. Exposures to 30 degrees C under aerial (emersion) and aquatic (immersion) conditions resulted in differences in hsp synthesis in T. brunnea but not in T. funebralis. The different time courses and magnitudes of hsp synthesis in these congeners suggest that the vertical limits of their distributions may be set in part by thermal stress.     

Studies on heat shock proteins in sea urchin development

Work on stress proteins in sea urchin embryos carried out over the last 20 years is reviewed and the following major results are described. Entire sea urchin embryos, if subjected to a rise in temperature at any postblastular stage undergo a wave of heat shock protein (hsp) synthesis and survive. If subjected to the same rise between fertilization and blastula formation, they are not yet able to synthesize hsp and die. Four clones coding for the major hsp, hsp70, have been isolated and sequenced; evidence for the existence of a heat shock factor has been provided, and a mechanism for the developmental regulation of hsp synthesis discussed. Intraembryonic and intracellular hsp location has been described; and a mechanism for achievement of thermotolerance proposed. A chaperonine role for a constitutive mitochondrial hsp56 has been suggested, as well as a role for the constitutive hsp70 in cell division. Heat shock, if preceded by 12-O-tetradecanoylphorbol-12-acetate (TPA) treatment causes apoptosis.     

Expression and localization of Drosophila melanogaster hsp70 cognate proteins.

Monoclonal antibodies have been used to identify three proteins in Drosophila melanogaster that share antigenic determinants with the major heat shock proteins hsp70 and hsp68. While two of the proteins are major proteins at all developmental stages, one heat shock cognate protein, hsc70, is especially enriched in embryos. hsc70 is shown to be the product of a previously identified gene, Hsc4. We have examined the levels of hsp70-related proteins in adult flies and larvae during heat shock and recovery. At maximal induction in vivo, hsp70 and hsp68 never reach the basal levels of the major heat shock cognate proteins. Monoclonal antibodies to hsc70 have been used to localize it to a meshwork of cytoplasmic fibers that are heavily concentrated around the nucleus.     

Hsp 70 expression and function during gametogenesis

The dramatic transformations in nuclear content and cellular organization that occur during gametogenesis require unique regulation and execution of the mitotic and meiotic cell cycle, apoptotic cell death, DNA recombination and repair, and cellular differentiation. These processes are accompained by the constitutive and developmentally regulated expression of a number of hsp70 genes encoding 70 kDa heat shock proteins (Hsp70), including several hsp70s whose expression is unique to male germ cells. Examining the expression and function of Hsp70s in germ cells has provided significant insights into mechanisms of hsp70 gene regulation and Hsp70 protein function, as well as the developmental processes of gametogenesis.     

Cloning of sponge heat shock proteins: evolutionary relationships between the major kingdoms

In the present study we have cloned from sponges (Porifera) those molecules which are involved in the protection of organisms against physiological and stress conditions; the inducible heat shock protein M_r 70,000, hsp70, from the marine sponge Geodia cydonium , its interacting hsp40, a DnaJ-like protein (from G. cydonium ) and the constitutively expressed counterpart the glucose-regulated protein M_r 78,000, GRP78 from Suberites domuncula . Alignments of the sequences revealed that the deduced aa sequences of all sponge hsp's share high homology to other metazoan sequences, and are separated from related sequences from plants and fungi (hsp70, GRP78, DnaJ) as well as Bacteria (DnaK, the hsp70 homologoue and the DnaJ) and Archaea (DnaK, the hsp70 homologoue and the DnaJ). One comparison based on nt sequences (hsp70/DnaK) showed a less pronounced grouping. From these data we conclude, that for phylogenetic analyses of deep branches in the metazoan evolution, not only 'characteristic'metazoan genes, but also 'housekeeping genes'e.g. are suitable for evolutionary inference.
The sequences reported here have been submitted to the EMBL/GenBank data base (accession no. GCDNAJ Y09037, GCHSP70 X94985, SDGR78 Y09500).     

Aging affects expression of 70-kDa heat shock proteins in Drosophila

We examined the effect of cellular aging on adult mortality and hsp70 gene expression in Drosophila melanogaster under thermal stress. The results showed that flies exposed to 37 degrees C for various time intervals had reduced survival rate with age. The level of hsp70 mRNA increases in flies up to 23-28 days of age, but then declines as they get older. When flies are shifted to 25 degrees C after 30 min of heat stress, the time-dependent decrease in hsp70 mRNA levels occurs more rapidly in young flies than in old ones. The hsp70 mRNA present during this recovery period is translated into protein, and senescent flies continue to synthesize this protein for up to 5 h after heat shock. The prolonged expression of hsp70 RNA during recovery from heat shock was also observed in young flies fed canavanine, an arginine analogue. These data suggest that in old insects, the accumulation of conformationally altered proteins plays a role in the regulation of hsp70 RNA expression. These results are discussed in relation to the finding that old flies are more sensitive to thermal stress than young ones.