Superantigens: Molecular Basis for Their Role in Human Diseases

Streptococcal toxic shock syndrome (STSS) is caused by a sudden release of one or more streptococcal superantigens (SAgs) into the blood. The conditions that result in this sudden production of toxin remain one of the major questions in the study and treatment of invasive streptococcal disease. The availability of streptococcal genomes has simplified the effort of identifying new SAgs. Minor allelic variation has been found for streptococcal pyrogenic exotoxin (SPE)-A and SPE-C. Two cocrystal structures of streptococcal SAgs bound to soluble TcR reveal how SPE-A interacts with murineVβ8.2 and SPE-C interacts with human Vβ2.1 complexes. Streptococcal SAgs when combined with small doses of endotoxin such as lipopolysaccharide (LPS) are lethal in rabbits. The focus on speA without knowledge of other more potent streptococcal SAgs such as streptococcal mitogenic exotoxin Z (SMEZ) and SPE-J tends to reduce the significance of these earlier studies by adding the potential for two or more SAgs to contribute to toxicity. Kawasaki disease (KD) is an acute multisystem vasculitis of unknown etiology that affects mostly young children and is now recognized as the leading cause of acquired heart disease in children in the developed world. The streptococcal SAgs are a family of secreted toxins similar to their staphylococcal cousins, but with some unique features, in particular with regard to expression and potency. It is clear that the SAg genes originated in staphylococci, and the fact that most are encoded in mobile phage suggests that they confer a significant advantage to the organism.

This chapter provides an introduction to bacterial superantigen (SAg) and presents the current understanding of disease association and causation by SAgs, with the intent of identifying needed future research studies of SAgs. Bacterial SAgs include those produced by β-hemolytic streptococci (including Streptococcus pyogenes), Streptococcus aureus and Streptococcus intermedius, Mycoplasma arthritidis, and certain species of Yersinia. In the chapter the authors consider only T-cell SAgs; definitions listed may be easily adapted to define B-cell SAgs. The pyrogenic toxin SAgs include the staphylococcal enterotoxins (SEs), which are known to cause emesis when administered orally to monkeys, and thus are the causes of staphylococcal food poisoning. Streptococcal pyrogenic exotoxins (SPEs) (erythrogenic toxins, scarlet fever toxins) now include the classic SPEs A and C, but also serotypes G, H, I, J, K, L, M, streptococcal superantigen (SSA), and streptococcal mitogenic exotoxin Zn (SMEZ). The last aspect of the author's presentation of new horizons for SAg research is based on the premise that human diseases caused by SAgs begin with their interaction with epithelial cells. Both group A streptococci and S. aureus cause infections that begin on skin and mucous membranes and, most of these organisms make SAgs. Clearly, additional studies are necessary to clarify all aspects of the interaction with mucosal and skin surfaces. These studies are likely to more fully define the extent of SAg involvement in human illnesses, and at the same time characterize the mechanism of SAg penetration of these surfaces.

This chapter begins with an outline of the original observations that led to the discovery of Mycoplasma arthritidis-derived superantigen (MAM) and discusses the structural basis of its interaction with class II major histocompatibility complex (MHC) molecules and T-cell receptors (TCRs). It reviews recent work on the interaction of MAM with Toll-like receptors and their importance in control of innate and adaptive immunity and in disease expression. The chapter discusses the potential role of MAM-like superantigens (SAgs) in autoimmune disease and how this might relate to recent findings on Toll-like receptor (TLR) control of adaptive immunity. Antigen-induced activation of T cells leading to cytokine production and proliferation requires signals delivered by antigen-presenting cells (APC), i.e., macrophages or dendritic cells (DCs), through costimulatory molecules such as B7-1 (CD80) and B7-2 (CD86) that are related membrane-bound molecules. In view of the demonstrated allelic specificity of MAM for MHC molecules it is also tempting to project that these polymorphisms may influence the outcome of the MAM/TLR interactions. A recent observation that has relevance to the potential role of MAM-like SAgs in human rheumatoid arthritis (RA) is that in preliminary studies transgenic mice bearing the HLA-DR and HLA-DQ alleles that predispose to RA develop a type 1 adaptive cytokine profile in response to MAM, whereas cells from MAM-injected mice bearing those alleles that protect against RA develop a type 2 cytokine profile.

Adaptive immunity relies on the antigen-specific B cell (BCR) and T cell (TCR) receptors to survey for pathogenic events. Upon recognition of a microbe by these receptors, an immune response is initiated, resulting in the differentiation of the antigen-specific lymphocytes into effector cells, which eliminate the pathogen. According to the IMGT database, there are 23 and 18 functional T cell (TCR) receptors Vβ-families in humans and mice, respectively. The majority of known superantigens (SAgs) are of either bacterial or viral origin, the prototypes being the bacterial pathogens Staphylococcus aureus and Streptococcus pyogenes and the murine mammary tumor virus (MMTV). While the exogenous virus encoded SAg is critical for the viral infection, the expression of endogenous MMTV SAgs leads to thymic deletion of T cells with the corresponding Vβ-chains, hence rendering the mice resistant to infection by exogenous MMTV encoding SAgs with the same Vβ specificity. During thymic maturation, only T cells that receive signals above a certain threshold survive by undergoing positive selection. Expression of a strong endogenous SAg during the negative selection process often results in thymic deletion of the entire population of T-cell expression TCR Vβ-chains that are reactive to the SAg, while a weak SAg activity leads to partial deletion or anergy of the reactive T cells.

Yersinia pseudotuberculosis-derived mitogens (YPMs) a, b, and c, have been identified as superantigen (SAg) from gram-negative bacteria. The T-cell activation by SAgs triggers the pathological changes that are based on multiple organ failure, seen in several infectious diseases, such as acute and systemic Y. pseudotuberculosis infection, toxic shock syndrome (TSS) in children and adults, and TSS in neonates, neonatal TSS-like exanthematous disease (NTED). This chapter reviews progress in research on YPMs and the diseases caused by them. It also discusses research findings in TSS and NTED, because these findings provide clues to the pathogenic mechanism of systemic Y. pseudotuberculosis infection. Genes encoding SAgs are frequently carried within mobile genetic elements, especially bacteriophages, and several features of YPMs strongly suggest involvement of mobile genetic elements in carrying the SAg genes. First, YPM genes have not been present in all Y. pseudotuberculosis strains examined. Second, their guanine and cytosine (GC) content is significantly lower (34.6 to 35.3%) than in the genomic core (46.5%). The authors reproduced the response patterns seen in patients with TSS and systemic Y. pseudotuberculosis infection in mouse experiments. They hypothesize that the biased expansion is based on different binding affinities of TCR Vβ-elements for the complex of SAg/MHC class II molecules. Patients with TSS and systemic Y. pseudotuberculosis infection exhibit overlapping clinical manifestations. An NTED patient with exceptionally severe clinical manifestations was found to have an adult-type massive, protracted expansion of Vβ-2+ T cells.

This chapter outlines the details of the core three-dimensional structure of the superantigens (SAgs) and discusses precisely what is conserved among the family along with the evolutionary reasons for conservation. It then discusses the variation that has been grafted onto the conserved structure, allowing different members of the family to bind to major histocompatibility class II molecules (MHC-II) and the T-cell receptors (TCRs) in many different configurations, with different affinities and, in the case of TCR binding, different specificities. The interactions between SAgs and their target MHC-II molecules and TCRs cover virtually all permutations of the following binding modes: MHC-II α-chain binding; MHC-II β-chain binding; TCR Vα binding and restriction; TCR Vβ binding and restriction; SAg oligomerization; MHC-II cross-linking. For the purposes of discussion these variations are loosely divide into MHC-II α-chain binding and its associated TCR interactions, MHC-II α-chain binding and its associated TCR interactions, and SAg oligomerization and MHC-II cross-linking. As secreted proteins from two highly adapted human pathogens, S. aureus and S. pyogenes, SAgs and the related staphylococcal superantigen-like proteins (SSLs) must be subject to severe immune pressure due to their potent effects. Despite their notoriety, it is likely that, for the most part, the effects of SAg and SSL secretion are relatively benign, and that only when local concentrations rise, or synergistic relationships with other factors apply, do they trigger the severe invasive disease with which they are associated.

Bacterial superantigens (SAgs) are small, highly mitogenic proteins that cross-link antigen-presenting cells (APCs) and T cells by binding simultaneously to the immunoreceptors major histocompatibility complex (MHC) class II and the T-cell receptors (TCRs) leading to the stimulation of large numbers of T cells. The advantages for microbes of secreting SAgs are not very well understood, but apart from disturbing the normal immune response they might prolong survival by promoting local inflammation, thereby increasing the blood and nutrient supply. The ability of SAgs to activate the immune system systemically became clear when evidence was presented that the interaction took place outside the antigen groove. SEA is an interesting SAg because it can interact with both the generic site on the α-chain of MHC class II and the zinc-dependent site on the β-chain of major histocompatibility complex (MHC) class II. The crystal structure shows that SED forms homodimers where two carboxy-terminal β-sheets from two SED molecules are packed against each other burying a large solvent-inaccessible area. At high concentrations the streptococcal SAg Streptococcus pyogenes pyrogenic exotoxins (SPE-C) has the ability to cross-link two MHC molecules through a high-affinity, zinc-dependent interaction with the β-chain due to homodimer formation using residues in its amino-terminal domain (SPE-C probably binds MHC as a monomer under physiological conditions). Approximately two-thirds of the buried surface area is contributed by interaction with the β1-helix on MHC and one-third by the antigenic peptide, strongly implying that the peptide plays an important role in binding the SAg.

Bacterial superantigens (SAgs) are powerful T-cell stimulatory molecules produced primarily by Staphylococcus aureus and Streptococcus pyogenes. Bacterial SAgs possess the unique ability to cross-link major histocompatibility complex (MHC) class II molecules and T-cell receptors, which in turn is responsible for their ability to illicit an immune response several orders of magnitude greater than that of conventional peptide antigens. The division of staphylococcal and streptococcal SAgs into subfamilies based on amino acid sequence, structure, and physiological information has caused some disagreement in the scientific community. Conventional antigens are processed internally by antigen-presenting cells (APCs) and displayed as discrete peptides on the cell surface by MHC class II molecules. These peptide antigens are then recognized by T-cell receptors (TCRs) specific to that peptide. Crystal structures of SAgs in complex with MHC class II molecules via both the generic site (SEB and TSST-1 in complex with HLA-DR1) and the high-affinity site SpeC in complex with HLA-DR2 and SEH in complex with HLA-DR1 have allowed a detailed examination of these interactions. A zinc ion plays a critical role in the binding of SME-Z2, SpeG, and SpeH to MHC class II molecules, as the binding of all three of these toxins to LG-2 cells is significantly reduced by chelating the zinc. The chapter talks about binding to the t-cell receptor, formation of the trimeric complex for signal transduction, and other structural features and idiosyncrasies.

The skin is an important target for microbial infection. One key strategy by which microbes exacerbate skin disease is via the production of microbial toxins. This chapter focuses on the role of staphylococcal and streptococcal superantigens in human skin diseases, beginning with a review of the properties of superantigens and the mechanisms by which they cause cutaneous immunologic responses. In addition to activated keratinocytes, several other skin cell types constitutively express HLA-DR on their cell surface and are therefore targets for superantigen action. Superantigen-mediated stimulation of monocytes is a consequence of binding and transducing a positive signal through major histocompatibility complex (MHC) class II molecules. The type of antigen-presenting cell (APC) used to present superantigen may influence T-cell development. Staphylococcal superantigens have also been shown to induce human T-cell resistance to corticosteroids and may thereby complicate the treatment of atopic dermatitis (AD) and other inflammatory diseases treated with corticosteroids. The structure of ETB is similar to ETA except that the oxyanion hole, which forms part of the catalytic site, is in the closed or inactive conformation for ETA, but in the open or active conformation for ETB. Both Staphylococcus aureus and Staphylococcus pyogenes are commensal organisms in humans, so the fact that they have the potential to activate the immune response in such a dramatic fashion means that their expression must be tightly controlled and that the immune system must deal with their continuous presence.

In principle, three distinct mechanisms mediate superantigenic impact on the onset/relapse of autoimmune disease. First, superantigens may, following presentation on antigen-presenting cells, directly activate autoreactive T and B cells, which will migrate to the appropriate organ and once there contribute to tissue destruction by production of chemokines, proinflammatory cytokines, and tissue destructive proteinases. Second, innocent (i.e., nonautoreactive) bystander lymphocytes will be activated, thereby triggering nonspecific inflammatory response that might lead to disease relapse in the chronic phase of autoimmune disease. Finally, superantigens are able to activate the antigen-presenting cells, such as macrophages. Microbial superantigens encompass viral and bacterial proteins that share the ability to interact with major histocompatibility complex (MHC) class II and the T-cell receptor, thereby bypassing the conventional antigen-processing pathway. Superantigens have been implicated in several inflammatory diseases with and without autoimmune background. With these conditions one can include Kawasaki syndrome, psoriasis, rheumatoid arthritis, autoimmune myositis, and diabetes mellitus. Superantigens are able to interact with synoviocytes thereby triggering their production of chemokines (RANTES, monocyte chemoattractant protein-1 [MCP-1], and interleukin-8 [IL-8]). It has been clear for a decade that microbial superantigens have a significant impact on the expression of autoimmune and immune-mediated diseases. Indeed, depending on the timing and dose of superantigen, injecting of superantigen may affect the incidence and course of an autoimmune disease. Further studies on the therapeutic uses of superantigens as well as specific silencing of superantigen-mediated aberrant immune responses are definitely merited.

There have been different attempts with superantigen to modify experimental allergic encephalomyelitis (EAE), which is taken as an animal model for multiple sclerosis (MS). MS is not simply an inflammatory disease like EAE. It is well established that in MS chronic degeneration runs in parallel to outbreaks of inflammation. The portions of inflammation and degeneration may vary so that different types of disease course can be distinguished. Recent studies indicate that the degenerative process MS is not always secondary to but may also precede neuroinflammation. The chapter presents experiments which demonstrate that superantigen expressed in the brain or spinal cord is capable of inducing neuroinflammation as in MS. Horizontal sections of the cerebral hemispheres at the corpus callosum and at the level of the lateral ventricles were stained with hematoxylin/eosin. It is well known that relapsing-remitting MS patients with frequent relapses progress faster than patients with less frequent relapses. HLA associations identified in patients with a relapsing-remitting course of MS may be due to major histocompatibility complex (MHC)-specific binding properties of T-cell superantigens. Different so-called autoimmune diseases involve nerves or muscles. Pathogens like bacteria or viruses are involved in part of them. Although it is likely that true autoimmune phenomena following molecular mimicry are of pathogenetic relevance in some of these diseases, it seems possible that superantigens are involved in parallel or alone in a subgroup of patients with autoimmune neuritis or myositis.

The uncontrolled immunological response triggered by microbial superantigens has been implicated in the etiology of numerous human disorders. The observations that superantigens can differentially activate T cells from different individuals suggested that genetic heterogeneity contributes to the clinical phenotype following exposure to superantigens. More recently, the findings that differences in the avidity of HLA molecules to bind superantigens and present them to T cells could dictate the strength and quality of the cytokine response have provided perhaps the strongest evidence about the contribution of the HLA haplotype to susceptibility or resistance to superantigen-induced disorders. The development of HLA class II transgenic murine models that are superantigen sensitive has been a critical step toward understanding the effect of HLA polymorphism on superantigen-mediated disease expression. Mycoplasma arthritidis, which can cause a chronic inflammatory polyarthritis in genetically susceptible strains of rodents, produces a soluble factor designated M. arthritidis mitogen (MAM) with superantigenic properties. The generation of transgenic mice expressing human HLA molecules has been an important step toward the creation of in vivo models for investigating the function of disease-associated HLA class II. Genetic susceptibility to autoimmunity in humans and experimental animal models is due to the presence of multiple disease loci. For many common autoimmune diseases little is known about the potential role of superantigens.

Toxic shock syndrome (TSS) is a serious acute bacterial disease characterized by fever, diffuse erythematous rash, hypotension, multiorgan involvement, and desquamation of the skin one to two weeks after onset. Various immunomodulatory agents and antisuperantigen therapeutic strategies have been proposed. One such strategy includes the administration of intravenous polyspecific immunoglobulin (IVIG). This chapter reviews the mechanistic actions and use of IVIG as adjunctive therapy for TSS. Patients were considered to have streptococcal TSS if they had hypotension in combination with two or more of the following: acute renal failure, coagulation abnormalities, liver abnormalities, adult respiratory distress syndrome (ARDS), generalized rash, and necrotizing fasciitis. The observation that staphylococcal TSS patients normally do not have detectable bacteremia, yet the patients demonstrate significant systemic features, suggested that TSS was the result of a toxemia. Most Streptococcus pyogenes strains express several different superantigens, and strains harbor in general genes encoding three to five of the superantigens, but the repertoire of genes varies between strains. S. pyogenes and Staphylococcus aureus are major human pathogens largely due to their ability to modulate and exploit the host defense mechanisms. Conventional therapy of invasive S. pyogenes infections has consisted of antimicrobials and, when necessary in severe invasive disease, support of vital functions for those patients with streptococcal TSS and surgery for those patients with necrotizing fasciitis.

Bacterial superantigens are among the most lethal of toxins. These stable proteins bind directly to most major histocompatibility (MHC) class II molecules and stimulate virtually all T cells bearing particular domains in the variable portion of the β-chain of the αβ T cell receptor (TCR), without need for processing by antigen-presenting cells. The peptides are capable of protecting mice from the lethal effects of superantigen toxins as widely different as staphylococcal enterotoxins (SE) SEB and toxic shock syndrome toxin 1 (TSST-1), and they can rescue animals already deeply into toxic shock. The superantigen antagonist peptides described in this chapter protect or rescue mice from lethal shock in a molar excess of as low as 20 fold over the toxin, implying that they bind tightly to a cellular target that is critical for superantigen action. The antagonist peptides described in the chapter provide a new molecular tool for understanding the mechanism of excessive human immune response activation by superantigens that occurs during toxic shock and for the identification of a novel target ligand that may interact with this superantigen domain. Removal of two amino acids from the dodecamer motif led to a significant decline in antagonist activity; this truncation may affect conformational stability or appropriate folding onto this putative receptor and reduce its affinity for the target.

Staphylococcal superantigens are grouped into three classes based on their primary sequence homology. In humans and monkeys, Staphylococcal enterotoxins (SEs) induce an emetic response and toxic shock at submicrogram concentrations. The mouse models commonly used today rely on the use of sensitizing agents such as D-galactosamine, actinomycin D, lipopolysaccharide (LPS), or viruses to induce toxic shock. None of the existing mouse models faithfully reproduces all of the complex events of human toxic shock syndrome. Bacterial superantigens cause toxic shock and contribute to septic complications during infection. Staphylococcal toxic shock syndrome (TSS) is characterized by fever, hypotension, desquamation of skin, fever, and dysfunction of three or more organ systems. Superantigens from both S. aureus and Streptococcus pyogenes are the causative agents of staphylococcal and streptococcal toxic shock. Given the complex pathophysiology of toxic shock, an understanding of the interaction of cellular receptors and signaling pathways used by these staphylococcal superantigens and the biological mediators they induce provides invaluable insight into selecting appropriate therapeutic targets. A list of small nonpeptide inhibitors effective in blocking the effects of superantigens is shown in this chapter. The anti-inflammatory cytokine interleukin 10 (IL-10) was used to block the production of IL-1, tumor necrosis factor alpha (TNF-α), and gamma interferon (IFN-γ), resulting in reduced lethality to superantigen-induced toxic shock. The ability to stop the inflammatory events initiated by superantigens early appears to be critical in preventing toxic shock.

Superantigens (SAgs) are a family of highly potent immunostimulatory proteins produced by bacteria or viruses. The family includes many proteins that may be unrelated by sequence or structure and yet share the ability to bypass the mechanisms of conventional antigen processing and trigger excessive activation of T cells. The massive lymphokine release by the activated cells within hours and the excessive T-cell proliferation in 2 to 3 days could ultimately lead to an immunosuppressive state and favor the microbe. Human diseases caused by microbes that utilize SAgs as their major virulence factors are characterized by fever and shock. The Staphylococcus aureus enterotoxins A-1 are thought to be the causative agents in 33% of all food-poisoning cases and are the most frequent cause of hospital-acquired infections. Presently, the treatment of superantigen-mediated infections is limited to the administration of antibiotics and handling of the state of shock. Conventional antigens are phagocytosed by antigenpresenting cells (APCs) and are processed into discrete peptides. The SAgs of S. aureus and Streptococcus pyogenes share a common architecture despite their significant difference in sequence. SAgs are globular proteins of 22 to 29 kDa, composed of two domains, amino- and carboxy-terminal, that are separated by a long, solvent-accessible α- helix spanning the center of the molecule. The T-cell receptor is a transmembrane heterodimer, composed of α- and β-chains. Each chain is composed of constant (C) and variable (V) regions.