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Glioblastoma multiforme (GBM) is a primary brain tumor that is very aggressive, resistant to treatment, and characterized by a high degree of anaplasia and proliferation. Routine treatment includes ablative surgery, chemotherapy, and radiotherapy. However, GMB rapidly relapses and develops radioresistance. Here, we briefly review the mechanisms underpinning radioresistance and discuss research to stop it and install anti-tumor defenses. Factors that participate in radioresistance are varied and include stem cells, tumor heterogeneity, tumor microenvironment, hypoxia, metabolic reprogramming, the chaperone system, non-coding RNAs, DNA repair, and extracellular vesicles (EVs). We direct our attention toward EVs because they are emerging as promising candidates as diagnostic and prognostication tools and as the basis for developing nanodevices for delivering anti-cancer agents directly into the tumor mass. EVs are relatively easy to obtain and manipulate to endow them with the desired anti-cancer properties and to administer them using minimally invasive procedures. Thus, isolating EVs from a GBM patient, supplying them with the necessary anti-cancer agent and the capability of recognizing a specified tissue-cell target, and reinjecting them into the original donor appears, at this time, as a reachable objective of personalized medicine.

Mutations in genes encoding molecular chaperones, for instance the genes encoding the subunits of the chaperonin CCT (chaperonin containing TCP-1, also known as TRiC), are associated with rare neurodegenerative disorders. Using a classical molecular dynamics approach, we investigated the occurrence of conformational changes and differences in physicochemical properties of the CCT5 mutations His147Arg and Leu224Val associated with a sensory and a motor distal neuropathy, respectively. The apical domain of both variants was substantially but differently affected by the mutations, although these were in other domains. The distribution of hydrogen bonds and electrostatic potentials on the surface of the mutant subunits differed from the wild-type molecule. Structural and dynamic analyses, together with our previous experimental data, suggest that genetic mutations may cause different changes in the protein-binding capacity of CCT5 variants, presumably within both hetero- and/or homo-oligomeric complexes. Further investigations are necessary to elucidate the molecular pathogenic pathways of the two variants that produce the two distinct phenotypes. The data and clinical observations by us and others indicate that CCT chaperonopathies are more frequent than currently believed and should be investigated in patients with neuropathies.

Salivary gland tumors represent a serious medical problem and new tools for differential diagnosis and patient monitoring are needed. Here, we present data and discuss the potential of molecular chaperones as biomarkers and therapeutic targets, focusing on Hsp10 and Hsp90. The salivary glands are key physiological elements but, unfortunately, the information and the means available for the management of their pathologies, including cancer, are scarce. Progress in the study of carcinogenesis has occurred on various fronts lately, one of which has been the identification of the chaperone system (CS) as a physiological system with presence in all cells and tissues (including the salivary glands) that plays a role in tumor-cell biology. The chief components of the CS are the molecular chaperones, some of which belong to families of evolutionarily related molecules named heat shock protein (Hsp). We are quantifying and mapping these molecular chaperones in salivary glands to determine their possible role in the carcinogenetic mechanisms in these glands and to assess their potential as diagnostic biomarkers and therapeutic targets. Here, we report recent findings on Hsp10 and Hsp90 and show that the quantitative and topographic patterns of tissue Hsp90 are distinctive of malignant tumors and differentiate benign from malignant lesions. The Hsp90 results show a correlation between quantity of chaperone and tumor progression, which in turn calls for negative chaperonotherapy, namely, elimination/inhibition of the chaperone to stop the tumor. We found that in vitro, the Hsp90 inhibitor Ganetespib is cytotoxic for the salivary gland UM-HACC-2A cell line. The drug, by interfering with the pro-survival NF-κB pathway, hampers cellular proliferation and migration, and favors apoptosis, and can, therefore, be considered a suitable candidate for future experimentation to develop a treatment for salivary gland tumors.

Hurthle cell (HC), anaplastic (AC), and medullary (MC) carcinomas are low frequency thyroid tumors that pose several challenges for physicians and pathologists due to the scarcity of cases, information, and histopathological images, especially in the many areas around the world in which sophisticated molecular and genetic diagnostic facilities are unavailable. It is, therefore, cogent to provide tools for microscopists to achieve accurate diagnosis, such as histopathological images with reliable biomarkers, which can help them to reach a differential diagnosis. We are investigating whether components of the chaperone system (CS), such as the molecular chaperones, can be considered dependable biomarkers, whose levels and distribution inside and outside cells in the tumor tissue could present a distinctive histopathological pattern for each tumor type. Here, we report data on the chaperones Hsp27, Hsp60, and Hsp90. They presented quantitative levels and distribution patterns that were different for each tumor and differed from those of a benign thyroid pathology, goiter (BG). Therefore, the reported methodology can be beneficial when the microscopist must differentiate between HC, AC, MC, and BG.

Breast cancer (BC) is a major public health problem, with key pieces of information needed for developing preventive and curative measures still missing. For example, the participation of the chaperone system (CS) in carcinogenesis and anti-cancer responses is poorly understood, although it can be predicted to be a crucial factor in these mechanisms. The chief components of the CS are the molecular chaperones, and here we discuss four of them, Hsp27, Hsp60, Hsp70, and Hsp90, focusing on their pro-carcinogenic roles in BC and potential for developing anti-BC therapies. These chaperones can be targets of negative chaperonotherapy, namely the elimination/blocking/inhibition of the chaperone(s) functioning in favor of BC, using, for instance, Hsp inhibitors. The chaperones can also be employed in immunotherapy against BC as adjuvants, together with BC antigens. Extracellular vesicles (EVs) in BC diagnosis and management are also briefly discussed, considering their potential as easily accessible carriers of biomarkers and as shippers of anti-cancer agents amenable to manipulation and controlled delivery. The data surveyed from many laboratories reveal that, to enhance the understanding of the role of the CS in BS pathogenesis, one must consider the CS as a physiological system, encompassing diverse members throughout the body and interacting with the ubiquitin–proteasome system, the chaperone-mediated autophagy machinery, and the immune system (IS). An integrated view of the CS, including its functional partners and considering its highly dynamic nature with EVs transporting CS components to reach all the cell compartments in which they are needed, opens as yet unexplored pathways leading to carcinogenesis that are amenable to interference by anti-cancer treatments centered on CS components, such as the molecular chaperones.

Hsp60 belongs to the subgroup of molecular chaperones named chaperonins and, typically, resides and functions in the mitochondria but it is also present in extramitochondrial sites. It chaperones client peptides as they fold to achieve the native conformation and also displays anti-stress roles by helping stress-damaged proteins regain a functional shape. Thus, Hsp60 is central to the integrity and functionality of mitochondria and energy production. All cells in the nervous system depend on Hsp60 so when the chaperonin malfunctions the consequences on nervous tissues are usually devastating, causing diverse diseases. These are the Hsp60 chaperonopathies, which can be genetic or acquired with the former caused by gene variants and the latter by various post-transcriptional mechanisms. All forms of chaperonopathies, i.e., by defect, by excess, and by mistake, associated with Hsp60 have been described, and some illustrative examples are discussed here. It is clear that this chaperonin is key to neuromuscular physiology but, when qualitatively and/or quantitatively abnormal causes diseases, often very serious.

Introduction: Heat Shock Protein 60 (Hsp60) is a member of the chaperoning system that assists protein folding inside mitochondria and plays other roles beyond these organelles. It is implicated in the carcinogenic processes in various types of cancer. In human salivary glands, Hsp60 has not yet been measured or mapped in detail and its role in gland development and functioning is virtually unknown. Consequently, its potential as biomarker for gland diseases, including malignancies cannot be assessed. The S-100 protein, a known marker for schwannomas, has been found also in myoepithelial-cell carcinomas of the salivary glands. Here, we present our initial findings on the anatomic-histological distribution of Hsp60 in human submandibular salivary gland (SMG) at various stages of development and its changes during tumorigenesis, in parallel with changes of S-100 in salivary gland tumors. Methods: Adult human submandibular gland (normal and tumoral) and embryonic head tissue samples were processed by standard methods for routine histological analysis. Additionally, these same sections underwent immunohistochemical staining using antibodies against Hsp60 and S- 100. Specimens from patients were obtained from the archives of the Human Pathology Section, Department of Health Science, University of Palermo, Italy. All procedures were in accordance with the Helsinki Declaration. We determined the percentages of cells immunopositive for Hsp60 or S-100 and made comparative evaluations applying the one way ANOVA. Results: Hsp60 was present in the acini and ducts of embryonic salivary glands but had a different distribution pattern in adult glands: it occurred only in the ducts and in a few acini. In contrast, Hsp60 was not detected in Pleomorphic Adenoma (PA) or Warthin’s tumor (WT) , whereas its levels were high in Adenoid Cystic Adenoma (ACC) . S-100 was present in the nuclei and/or in the cytoplasm in PA and ACC and its levels in the nuclei in ACC were higher than in the PA nuclei . Conclusions: Since the chaperonin is abundant in acini and ducts of embryonic salivary glands, it can be hypothesized that it actively participates in the developmental process leading to the formation of a wholly functional adult, mature organ. Hsp60 and S-100 immunopositivities were high in the malignant tumor implying their involvement in neoplasm formation and progression. These results foreshadow the diagnostic and prognostic potential of Hsp60 and S-100 when measured side by side as biomarkers useful for distinguishing between benign and malignant tumors.

Heat Shock Proteins (HSPs) role in development and function of human salivary glands have not yet been measured or mapped in detail, in particular, HSP60 role is unknown. They can be potential biomarkers for glandular diseases and malignancies. We present our initial findings on HSP10, HSP27, HSP60, HSP70 and HSP90 distribution in human submandibular salivary glands (SMG) at various stages of development and tumorigenesis.

Chaperonopathies are diseases in which abnormal chaperones play an etiopathogenic role. A chaperone is mutated or otherwise abnormal (e.g., modified by an aberrant posttranslational modification) in structure/function. To understand the pathogenic mechanisms of chaperonopathies, it is necessary to elucidate the impact of the pathogenic mutation or posttranslational modification on the chaperone molecule's properties and functions. This impact is usually subtle because if it were more than subtle the overall effect on the cell and organism would be catastrophic, lethal. This is because most chaperones are essential for life and, if damaged in structure/function too strongly, there would be death of the cell/organism, and no phenotype, i.e., there would be no patients with chaperonopathies. Consequently, diagnostic procedures and analysis of defects of the abnormal chaperones require a multipronged method for assessing the chaperone molecule from various angles. Here, we present such a method that includes assessing the intrinsic properties and the chaperoning functions of chaperone molecules.

Hsp60 (also called Cpn60) is a chaperonin with essential functions for cell physiology and survival. Additionally, its involvement in the pathogenesis of a variety of diseases (e.g., some autoimmune disorders and cancer) is becoming evident with new research. For example, the distribution and levels of Hsp60 in cells and tissues have been found altered in many pathologic conditions, and the significance of these alterations is being investigated in a number of laboratories. The aim of this ongoing research is to determine the meaning of these Hsp60 alterations with regard to pathogenetic mechanisms, diagnosis, classification of lesions, and assessing prognosis and response to treatment.Hsp60 occurs in the mitochondria, i.e., its typical residence according to classic knowledge, and also in other locales, such as the cytosol, the cell membrane, the intercellular space, and biological fluids (e.g., blood and cerebrospinal fluid). Detection and quantitative determinations in all these locations are becoming essential components of laboratory pathology in clinics and research. Consequently, immunohistochemistry targeting Hsp60 is also becoming essential for pathologists and researchers interested in disorders involving this chaperonin.In this chapter, we summarize some recent discoveries on the participation of Hsp60 in the pathogenesis of human diseases, and describe in detail how to perform immunohistochemical reactions for detecting the chaperonin, determining its location, and measuring its quantitative levels.

Aging entails progressive deterioration of molecules and supramolecular structures, including Hsp chaperones and their complexes, paralleled by functional decline. Recent research has changed our views on Hsp chaperones. They work inside and outside cells in many locations, alone or forming teams, interacting with cells, receptors, and molecules that are not chaperones, in roles that are not typically attributed to chaperones, such as protein folding. Hsp chaperones form a physiological system with a variety of functions and interactions with other systems, for example, the immune system. We propose that chaperone malfunctioning due to structural damage or gene dysregulation during aging has an impact on the immune system, creating the conditions for an overall malfunction of both systems. Pathological chaperones cannot interact with the immune system as normal ones do, and this leads to an overall readjustment of the interactions that is apparent during senescence and is likely to cause many of its manifestations.

The effect of supraoptimal concentrations of ammonia on cell growth and stress genes of the dnaK locus was investigated in Methanosarcina mazei S-6, a methanogen important for anaerobic digestion in bioreactors. When the concentration of NH4Cl was increased 25 times above the optimal for growth, the lag phase lengthened, growth occurred at a decreased rate, and a stress response was induced – the transcripts from the stress genes grpE, dnaK, and dnaJ increased, whereas that of the adjacent genes orf11-trkA decreased. In contrast, a 10-fold elevation of the NH4Cl concentration, which had only a slight stimulating effect on the stress genes, augmented the orf11-trkA transcript. These findings show that ammonia at certain levels is a cell stressor for M. mazei S-6.

A procedure was developed for real-time measurement of the interaction between an archaeal TATA-binding protein (TBP) with stress-gene promoters from the archaeon Methanosarcina mazeii using surface plasmon resonance (SPR), the BIACORE 3000 equipment, and the SA (streptavidin) Sensor Chip. Measurements were based on the SPR optical phenomenon, which resulted in light extinction when TBP bound a promoter. This process, detected as a change in a particular angle, was recorded in a sensorgram. The BIA-evaluation program allowed the calculation of the equilibrium constant (K(A)) of the interaction of M. mazeii TBP with the promoters of the stress genes grpE, hsp70(dnaK), and hsp40(dnaJ) (0.47, 0.26, and 1.21x10(7)M(-1), respectively) and, for comparison, with the promoter of a non-heat-shock gene, orf16 (0.08x10(7)M(-1)). The association rate (k(a)) of the non-heat-shock gene orf16 was 0.4x10(4)M(-1)s(-1) and those for the stress genes, grpE, hsp70(dnaK), and hsp40(dnaJ) were higher: 2.8, 1.5, and 3.5x10(4)M(-1)s(-1), respectively. The new procedure will allow a comparative analysis of different TPBs and promoters (wild type and mutants) under physiologic and stress conditions, and a correlation of TBP binding parameters with constitutive and stress-induced gene expression.

The molecular chaperones Hsp60 and Hsp10 are, according to recent reports, involved in cancer development and progression. We, for instance, have found that their expression varies with distinctive patterns in different malignancies: they are overexpressed in colorectal, exocervical and prostate carcinogenesis, and colorectal cancer progression, but they are downregulated during bronchial carcinogenesis. There is also evidence showing that Hsp60 and Hsp10 can be used as therapeutic agents, for example in rheumatoid arthritis. In view of these findings we want now to call attention to the potential of Hsp60 and Hsp10 in cancer therapy.

Hsp60 (also called Cpn60) is a chaperonin with essential functions for cell physiology and survival. Additionally, its involvement in the pathogenesis of a number of diseases (e.g., some autoimmune disorders and cancer) is becoming evident with new research. For example, the distribution and levels of Hsp60 in cells and tissues have been found altered in many pathologic conditions, and the significance of these alterations is being investigated in a number of laboratories. The aim of this ongoing research is to determine the meaning of these Hsp60 alterations with regard to pathogenetic mechanisms, diagnosis, classification of lesions, and assessing of prognosis and response to treatment. Hsp60 occurs in the mitochondria, i.e., its typical residence according to classic knowledge, and also in other locales, such as the cytosol, the cell membrane, the intercellular space, and biological fluids (e.g., blood and cerebrospinal fluid). Detection and quantitative determinations in all these locations are becoming essential components of laboratory pathology in clinics and research. Consequently, immunohistochemistry targeting Hsp60 is also becoming essential for pathologists and researchers interested in disorders involving this chaperonin. In this chapter, we briefly summarize some recent discoveries on the participation of Hsp60 in the pathogenesis of human diseases and describe in detail how to perform immunohistochemical reactions for detecting the chaperonin, determining its location, and measuring its levels of expression.

Heat shock proteins (Hsps) are produced in response to various stressors, including ischemia-reperfusion, and they can exit cells and reach the blood. In this pilot study, we determined serum levels of Hsp60 and heme-oxygenase-1 (HO-1; also named Hsp32) in subjects with acute myocardial infarction (AMI) to assess their clinical significance and potential prognostic value. We also performed a bioinformatics analysis of the 2 molecules in search of structural clues on the mechanism of their release from cells. We studied 40 patients consecutively admitted for AMI (male:female patient ratio = 20:20, mean age: 64 ± 13 years) and 40 matched controls. A blood sample was drawn for biochemical analyses within 24 h of symptoms onset, and Hsp60 and HO-1 concentrations were determined by enzyme-linked immunosorbent assay (ELISA). All patients were followed up for 6 months to register adverse post-AMI cardiovascular events. A multivariate analysis demonstrated that elevated Hsp60 (P = 0.0361), creatine phosphokinase-muscle brain (CK-MB) (P = 0.0446), and troponin (P = 0.0490) were predictive of post-AMI adverse events. In contrast, increased HO-1 showed a significant association with less severity of coronary artery diseases (P = 0.0223). These findings suggest that Hsp60 and HO-1 play distinct roles in the pathogenesis of AMI and subsequent AMI-related pathology. The possibility that these proteins differ in their roles and mechanisms of action in AMI and post-AMI pathology was supported also by the bioinformatics estimates of probability of their localization in various subcellular compartments. The results clear the way for subsequent investigation on the pathogenetic role and clinical significance of Hsp60 and HO-1 in AMI.

Background Hsp60, a Group I mitochondrial chaperonin, is classically considered an intracellular chaperone with residence in the mitochondria; nonetheless, in the last few years it has been found extracellularly as well as in the cell membrane. Important questions remain pertaining to extracellular Hsp60 such as how generalized is its occurrence outside cells, what are its extracellular functions and the translocation mechanisms that transport the chaperone outside of the cell. These questions are particularly relevant for cancer biology since it is believed that extracellular chaperones, like Hsp70, may play an active role in tumor growth and dissemination. Methodology/Principal Findings Since cancer cells may undergo necrosis and apoptosis, it could be possible that extracellular Hsps are chiefly the result of cell destruction but not the product of an active, physiological process. In this work, we studied three tumor cells lines and found that they all release Hsp60 into the culture media by an active mechanism independently of cell death. Biochemical analyses of one of the cell lines revealed that Hsp60 secretion was significantly reduced, by inhibitors of exosomes and lipid rafts. Conclusions/Significance Our data suggest that Hsp60 release is the result of an active secretion mechanism and, since extracellular release of the chaperone was demonstrated in all tumor cell lines investigated, our observations most likely reflect a general physiological phenomenon, occurring in many tumors.

The study of cell walls has progressed considerably in recent years and has provided directions for future research that will soon result in practical applications. Accumulation of a variety of residues, which are toxic or at least space filling, along with threats of increased oil prices and the deleterious effects on the biosphere caused by the use of traditional fuels have stimulated the development of alternative resources of energy tied to technologies for waste elimination–reutilization and fuel production, using microbes, especially methanogenic archaea (in short, methanogens). Progress in the study of the microflora of man and animals has demonstrated the critical role of the resident microbes, including methanogens, in health and disease. These advances have boosted the study of methanogens in natural and manufactured ecosystems, including those in the gastrointestinal and urinary tracts, particularly concerning intermicrobial and microbial–host interactions that involve the cell wall. Consequently, the development of immunochemical methods for analysis, modification, manipulation, and use of methanogens in animal and human medicines and in industry has become a priority, promising significant results. The data in this Chapter on the immunochemical and molecular characteristics of the cell walls of methanogens, encompassing a range of evolutionary branches, will be useful to develop tools with direct applications in environmental, biotechnological, and medical research and practical endeavors. For example, the identification of distinct immunotypes in Methanobrevibacter smithii, which is by far the most frequent methanogen inhabiting the human body and plays a role in pathology by enhancing the pathogenesis of associated microbes, is the first step for determining which are the immunotypes most relevant for disease development. The same principle is valid for research directed toward the elucidation of which methanogenic immunotype is the most efficient for bioconversion of wastes with the highest yield of useful byproducts in waste-treatment plants.

HSP and molecular chaperones, both referred to in this chapter as chaperones, are key players in development and senescence. With regard to senescence, several issues are critical: the role of normal chaperones in the process of ageing itself and in preventing and controlling age-associated diseases, the role of defective chaperones (chaperonopathies) in the onset and progression of senescence and in the etiology of old-age diseases, the interaction of chaperones with the immune system, and the potential of chaperones as therapeutic agents for counteracting the deleterious effects of ageing on molecules and cells and for treating proteinopathies of the elderly (chaperonotherapy). All these issues are discussed in this chapter, focusing on Hsp60 and Hsp10 (referred to as chaperonins for the purposes of this article) by examining a sample of publications dealing with pathological conditions prevalent in aged individuals, such as atherosclerosis and coronary syndromes, neurodegenerative and degenerative joint disorders, diabetes, chronic occlusive pulmonary disease, and glaucoma (cancer is omitted because it has recently been extensively discussed elsewhere). The data show that Hsp60 and Hsp10 undergo changes in levels and distribution inside cells and tissues, including invasion of the extracellular space and plasma, with age and health status and in relation with specific pathologies. The physiological and/or pathological significance of these changes is not yet fully understood but is being actively investigated. The role of chaperonopathies, particularly those due to aberrant post-translational modifications related to stressors such as ROS, on the aggravation of senescence ought to be examined in detail in the near future. Studies on the mechanisms by which defective chaperones (molecular chaperonopathies) accelerate senescence and contribute to pathogenesis and, thus, to the development of disease (clinical chaperonopathies) should provide key information useful for developing diagnostic and therapeutic means based on chaperone genes and proteins

This article is about Hsp10 and its intracellular and extracellular forms focusing on the relationship of the latter with Early Pregnancy Factor and on their roles in cancer and immunity. Cellular physiology and survival are finely regulated and depend on the correct functioning of the entire set of proteins. Misfolded or unfolded proteins can cause deleterious effects and even cell death. The chaperonins Hsp10 and Hsp60 act together inside the mitochondria to assist protein folding. Recent studies demonstrated that these proteins have other roles inside and outside the cell, either together or independently of each other. For example, Hsp10 was found increased in the cytosol of different tumors (although in other tumors it was found decreased). Moreover, Hsp10 localizes extracellularly during pregnancy and is often indicated as Early Pregnancy Factor (EPF), which is released during the first stages of gestation and is involved in the establishment of pregnancy. Various reports show that extracellular Hsp10 and EPF modulate certain aspects of the immune response with anti-inflammatory effects in patients with autoimmune conditions improving clinically after treatment with recombinant Hsp10. Moreover, Hsp10 and EPF are involved in embryonic development, acting as a growth factor, and in cell proliferation/differentiation mechanisms. Therefore, it becomes evident that Hsp10 is not only a co-chaperonin, but an active player in its own right in various cellular functions. In this article, we present an overview of various aspects of Hsp10 and EPF as they participate in physiological and pathological processes such as the antitumor response and autoimmune diseases.

Microbes that produce methane gas, methanogens, were identified as Archaea in the 1970s but their possible role in disease is only emerging now, after they were found in the large intestine, mouth, and vagina. Significant associations were observed, for instance, between levels of methanogens in periodontal pockets and severity of periodontitis, and between quantities of methanogens in the large intestine and diseases such as colon cancer and diverticulosis. Recently, a role for intestinal methanogens in obesity was proposed. The lesson learned is that for methanogens we have to look at their pathogenicity from a different angle in comparison to classic pathogens that invade tissues and release toxins. This type of pathogenicity has not yet been described for methanogens. Instead, methanogens seem to participate in pathogenicity indirectly, favoring the growth of other microbes, which are directly involved in pathogenesis. This indirect role should not be minimized. On the contrary, it has become clear that a fundamental change of approach to the understanding and control of microbial diseases must be implemented. A comprehensive strategy is needed to elucidate the syntrophic associations that are essential for a healthy relation among microbes (including methanogens) and between them and the host organism, and to unveil those associations that lead to disease.

The stress response, stress proteins, heat-shock genes and proteins, molecular chaperone genes and proteins, and a number of closely related molecules and cellular processes have been studied over the last few decades. A huge amount of information has accumulated that is scattered in printed and electronic literature and databases. Most of this information constitutes the subject matter of the science of chaperonology. More recently, the concept of chaperone pathology, sick chaperones, has evolved since various pathological conditions have been identified in which defective chaperones play an etiologic role. These conditions are the chaperonopathies. Recent findings on chaperonopathies are briefly discussed in this article. Chaperonopathies occur at all ages; as a rule the genetic cases have an early clinical onset while the acquired chaperonopathies become manifest in the elderly and/or in association with other diseases. Other fields of chaperonology, which will most likely be expanded in the near future, are the study of extracellular chaperones, chaperone networks, the therapeutic use of chaperones (i.e., chaperonotherapy) to manage chaperonopathies and to improve cell performance in the face of stress, the evaluation of chaperones as diagnostic markers and as prognostic indicators, and the development of antichaperone agents to suppress chaperone-gene expression or inhibit chaperone function when chaperones contribute to disease rather than the opposite.

The molecular chaperones Hsp60 and Hsp10 are, according to recent reports, involved in cancer development and progression. We, for instance, have found that their expression varies with distinctive patterns in different malignancies: they are overexpressed in colorectal, exocervical and prostate carcinogenesis, and colorectal cancer progression, but they are downregulated during bronchial carcinogenesis. There is also evidence showing that Hsp60 and Hsp10 can be used as therapeutic agents, for example in rheumatoid arthritis. In view of these findings we want now to call attention to the potential of Hsp60 and Hsp10 in cancer therapy.

The study of molecular chaperones (genetics, structure, location, physiology, pathology, and therapeutics) has developed into a science with specific objectives, methods, and hypotheses, a discipline we called chaperonology. Subdisciplines of chaperonology include the study of pathological chaperones (chaperonopathies) and the analysis of their genes in sequenced genomes (chaperonomics). Chaperonopathies are pathological conditions in which one type of chaperone is deficient due to a genetic or acquired defect that modifies the chaperone’s structure and/or makes the chaperone unavailable for functioning when needed. Experimental and clinical data show that chaperones and their genes can be used for treating various pathological conditions, thus justifying the development of chaperonotherapy. We discuss recent work showing that chaperonotherapy is on solid foundations: the data demonstrate that molecular chaperones counteract pathogenetic mechanisms in disease and during stress.

The participation of molecular chaperones in the process of senescence and in the mechanisms of age-related diseases is currently under investigation in many laboratories. However, accurate, complete information about the number and diversity of chaperone genes in any given genome is scarce. Consequently, the results of efforts aimed at elucidating the role of chaperones in ageing and disease are often confusing and contradictory. To remedy this situation, we have developed chaperonomics, including means to identify and characterize chaperone genes and their families applicable to humans and model organisms. The problem is difficult because in eukaryotic organisms chaperones have evolved into complex multi-gene families. For instance, the occurrence of multiple paralogs in a single genome makes it difficult to interpret results if consideration is not given to the fact that similar but distinct chaperone genes can be differentially expressed in separate cellular compartments, tissues, and developmental stages. The availability of complete genome sequences allows implementation of chaperonomics with the purpose of understanding the composition of chaperone families in all cell compartments, their evolutionary and functional relations and, ultimately, their role in pathogenesis. Here, we present a series of concatenated, complementary procedures for identifying, characterizing, and classifying chaperone genes in genomes and for elucidating evolutionary relations and structural features useful in predicting functional properties. We illustrate the procedures with applications to the complex family of hsp70 genes and show that the kind of data obtained can provide a solid basis for future research.

Many agents of physical, chemical, or biological nature, have the potential for causing cell stress. These agents are called stressors and their effects on cells are due to protein denaturation. Cells, microbes, for instance, perform their physiological functions and survive stress only if they have their proteins in the necessary concentrations and shapes. To be functional a protein shape must conform to a specific three-dimensional arrangement, named the native configuration. When a stressor (e.g., temperature elevation or heat shock, decrease in pH, hypersalinity, heavy metals) hits a microbe, it causes proteins to lose their native configuration, which is to say that stressors cause protein denaturation. The cell mounts an anti-stress response: house-keeping genes are down-regulated and stress genes are activated. Among the latter are the genes that produce the Hsp70(DnaK), Hsp60, and small heat protein (sHsp) families of stress proteins. Hsp70(DnaK) is part of the molecular chaperone machine together with Hsp40(DnaJ) and GrpE, and Hsp60 is a component of the chaperonin complex. Both the chaperone machine and the chaperonins play a crucial role in assisting microbial proteins to reach their native, functional configuration and to regain it when it is partially lost due to stress. Proteins that are denatured beyond repair are degraded by proteases so they do not accumulate and become a burden to the cell. All Archaea studied to date possess chaperonins but only some methanogens have the chaperone machine. A recent genome survey indicates that Archaea do not harbor well conserved equivalents of the co-chaperones trigger factor, Hip, Hop, BAG-1, and NAC, although the data suggest that Archaea have proteins related to Hop and to the NAC alpha subunit whose functions remain to be elucidated. Other anti-stress means involve osmolytes, ion traffic, and formation of multicellular structures. All cellular anti-stress mechanisms depend on genes whose products are directly involved in counteracting the effects of stressors, or are regulators. The latter proteins monitor and modulate gene activity. Biomethanation depends on the concerted action of at least three groups of microbes, the methanogens being one of them. Their anti-stress mechanisms are briefly discussed in this Chapter from the standpoint of their role in biomethanation with emphasis on their potential for optimizing bioreactor performance. Bioreactors usually contain stressors that come with the influent, or are produced during the digestion process. If the stressors reach levels above those that can be dealt with by the anti-stress mechanisms of the microbes in the bioreactor, the microbes will die or at least cease to function. The bioreactor will malfunction and crash. Manipulation of genes involved in the anti-stress response, particularly those pertinent to the synthesis and regulation of the Hsp70(DnaK) and Hsp60 molecular machines, is a promising avenue for improving the capacity of microbes to withstand stress, and thus to continue biomethanation even when the bioreactor is loaded with harsh waste. The engineering of methanogenic consortia with stress-resistant microbes, made on demand for efficient bioprocessing of stressor-containing effluents and wastes, is a tangible possibility for the near future. This promising biotechnological development will soon become a reality due to the advances in the study of the stress response and anti-stress mechanisms at the molecular and genetic levels.

Many agents of physical, chemical, or biological nature, have the potential for causing cell stress. These agents are called stressors and their effects on cells are due to protein denaturation. Cells, microbes, for instance, perform their physiological functions and survive stress only if they have their proteins in the necessary concentrations and shapes. To be functional a protein shape must conform to a specific three-dimensional arrangement, named the native configuration. When a stressor (e.g., temperature elevation or heat shock, decrease in pH, hypersalinity, heavy metals) hits a microbe, it causes proteins to lose their native configuration, which is to say that stressors cause protein denaturation. The cell mounts an anti-stress response: house-keeping genes are down-regulated and stress genes are activated. Among the latter are the genes that produce the Hsp70(DnaK), Hsp60, and small heat protein (sHsp) families of stress proteins. Hsp70(DnaK) is part of the molecular chaperone machine together with Hsp40(DnaJ) and GrpE, and Hsp60 is a component of the chaperonin complex. Both the chaperone machine and the chaperonins play a crucial role in assisting microbial proteins to reach their native, functional configuration and to regain it when it is partially lost due to stress. Proteins that are denatured beyond repair are degraded by proteases so they do not accumulate and become a burden to the cell. All Archaea studied to date possess chaperonins but only some methanogens have the chaperone machine. A recent genome survey indicates that Archaea do not harbor well conserved equivalents of the co-chaperones trigger factor, Hip, Hop, BAG-1, and NAC, although the data suggest that Archaea have proteins related to Hop and to the NAC alpha subunit whose functions remain to be elucidated. Other anti-stress means involve osmolytes, ion traffic, and formation of multicellular structures. All cellular anti-stress mechanisms depend on genes whose products are directly involved in counteracting the effects of stressors, or are regulators. The latter proteins monitor and modulate gene activity. Biomethanation depends on the concerted action of at least three groups of microbes, the methanogens being one of them. Their anti-stress mechanisms are briefly discussed in this Chapter from the standpoint of their role in biomethanation with emphasis on their potential for optimizing bioreactor performance. Bioreactors usually contain stressors that come with the influent, or are produced during the digestion process. If the stressors reach levels above those that can be dealt with by the anti-stress mechanisms of the microbes in the bioreactor, the microbes will die or at least cease to function. The bioreactor will malfunction and crash. Manipulation of genes involved in the anti-stress response, particularly those pertinent to the synthesis and regulation of the Hsp70(DnaK) and Hsp60 molecular machines, is a promising avenue for improving the capacity of microbes to withstand stress, and thus to continue biomethanation even when the bioreactor is loaded with harsh waste. The engineering of methanogenic consortia with stress-resistant microbes, made on demand for efficient bioprocessing of stressor-containing effluents and wastes, is a tangible possibility for the near future. This promising biotechnological development will soon become a reality due to the advances in the study of the stress response and anti-stress mechanisms at the molecular and genetic levels.

A brief survey is presented of salient findings on transcription in the Archaea, focussing on stress genes of the hsp70(dnaK) locus, which code for the molecular chaperones Hsp70(DnaK), Hsp40(DnaJ), and GrpE. Archaeal basal factors and some recently characterized regulators pertinent to non-stress genes are presented first to show their similarities and differences with equivalents in organisms of the other two phylogenetic domains, Bacteria and Eucarya, and to reveal clues on how these or similar factors might transcribe and regulate the archaeal stress genes. The second part of the article deals with the hsp70(dnaK)-locus genes, particularly those from Methanosarcina mazeii, because they are virtually the only ones within the methanogenic Archaea whose patterns of constitutive and stress-induced expressions have been studied. Therefore, these genes, provide a standardized model system to elucidate transcription initiation and regulation at the molecular level in this phylogenetic group. Promoters, and other cis-acting sites that are, or might be, involved in stress-gene expression are described. Conformational changes of basal transcription factors after interaction with stress-gene promoters are discussed that suggest ways for generating a large diversity of initiation complexes using a few factors and DNA sites in different combinations. Likewise, the effects of stress on DNA topology and on TBP-TFB-promoter complex formation and tightness are described, which might also contribute to the generation of transcription-initiation complex diversity. This diversity would be key to differential gene expression, namely, which genes are transcribed, when (basal, steady expression vs. sporadic stress-induced expression), and to what level. Future research should investigate this diversity, and the mechanism of complex formation and action at the atomic, molecular, and supramolecular levels to elucidate the dynamics of transcription initiation in real time.