You will be presented with the abstract, introduction, and discussion section from a biomedical research article. Your task is to create a summary that a layperson can understand, capturing the essence of the research article. Keep essential scientific terms, but ensure that the language remains clear and the concepts are explained in an uncomplicated manner.

Title: Binding of Superantigen Toxins into the CD28 Homodimer Interface Is Essential for Induction of Cytokine Genes That Mediate Lethal Shock}
Article:
Abstract:
Bacterial superantigens, a diverse family of toxins, induce an inflammatory cytokine storm that can lead to lethal shock. CD28 is a homodimer expressed on T cells that functions as the principal costimulatory ligand in the immune response through an interaction with its B7 coligands, yet we show here that to elicit inflammatory cytokine gene expression and toxicity, superantigens must bind directly into the dimer interface of CD28. Preventing access of the superantigen to CD28 suffices to block its lethality. Mice were protected from lethal superantigen challenge by short peptide mimetics of the CD28 dimer interface and by peptides selected to compete with the superantigen for its binding site in CD28. Superantigens use a conserved β-strand/hinge/α-helix domain of hitherto unknown function to engage CD28. Mutation of this superantigen domain abolished inflammatory cytokine gene induction and lethality. Structural analysis showed that when a superantigen binds to the T cell receptor on the T cell and major histocompatibility class II molecule on the antigen-presenting cell, CD28 can be accommodated readily as third superantigen receptor in the quaternary complex, with the CD28 dimer interface oriented towards the β-strand/hinge/α-helix domain in the superantigen. Our findings identify the CD28 homodimer interface as a critical receptor target for superantigens. The novel role of CD28 as receptor for a class of microbial pathogens, the superantigen toxins, broadens the scope of pathogen recognition mechanisms.
Introduction:
As the principal costimulatory receptor, CD28 is a critical regulator of the immune response[1]–[3]. Expressed constitutively on T cells, CD28 is a homodimer that interacts with its B7 coligands to transduce the signal essential for an immediate T cell response[2]–[5]. B7-2 is expressed constitutively and induced immediately, whereas B7-1 is expressed only later[5],[6]; hence, interaction of B7-2 with CD28 regulates signaling by antigens early in the immune response[7],[8]. Apart from its B7 coreceptors, CD28 has no other known ligands. Bacterial superantigens are a family of proteins secreted by Staphylococcus aureus and Streptococcus pyogenes that induce toxic shock by activating an immune response, orders of magnitude higher in intensity than that elicited by regular antigens. Bypassing restricted presentation of conventional antigens which typically activate <0. 01% of T cells, superantigens bind directly as intact proteins to most major histocompatibility class II( MHC-II) and T cell receptor( TCR) molecules, activating up to 20% of T cells[9]–[11]. This results in massive induction of the T helper 1( Th1) cytokines interleukin-2( IL2), interferon-γ( IFN-γ), and tumor necrosis factor( TNF) that mediate toxic shock. Nevertheless, superantigen action also requires CD28 costimulation[12]–[14]. Induction of human T helper 1( Th1) cytokine gene expression by divergent superantigens is inhibited by a dodecamer peptide that protects mice from their lethal effect[15]. The peptide shows homology to the β-strand( 8) /hinge/α-helix( 4) domain in staphylococcal enterotoxin( SE) B, structurally conserved in superantigens yet remote from MHC-II and TCR binding sites. Because this domain is essential for superantigen action[15], we considered that it may engage a third receptor. We show here that this receptor is CD28. This costimulatory molecule is co-opted by superantigens as receptor. Superantigens must bind directly into the homodimer interface of CD28 to elicit toxicity. Blocking access of a superantigen to CD28 is sufficient to block lethal toxic shock. Peptide mimetics of the CD28 dimer interface inhibited the induction of Th1 cytokines and protected mice from lethal superantigen exposure. These experiments identify the CD28 homodimer interface as a receptor target for superantigen toxins and reveal a novel role for CD28 in pathogen recognition.
Discussion:
These experiments identify the CD28 dimer interface as a receptor target for superantigens. We show that to deliver the signal for Th1 cytokine induction, a superantigen must co-opt CD28 as third receptor, in addition to its full reliance on costimulatory signaling through B7-2/CD28. Thus, superantigens make unconventional use not only of MHC-II and TCR through direct binding but also of CD28. Independent lines of evidence show that the binding site for superantigens in CD28 is the crystallographic homodimer interface. The superantigen domain that engages CD28 contains the conserved β-strand/hinge/α-helix motif[15], remote from MHC-II and TCR binding sites, leaving it accessible to CD28( Figure 7). Blocking access of a superantigen to CD28 is sufficient to block lethal toxic shock. Mice were protected from lethal SEB challenge by peptide mimetics of the CD28 dimer interface and of the β-strand/hinge/α-helix domain in superantigens, as well as by random peptides selected to compete with SEB for its binding site in CD28. The finding that CD28 is a direct sensor of a class of microbial pathogens, superantigen toxins, broadens the scope of pathogen recognition mechanisms. Independent lines of evidence support direct binding of superantigen to CD28. SEB bound to cell-surface CD28 even in the absence of MHC-II or TCR. Novel peptide antagonists of SEB, protective against lethal toxic shock, were selected by phage display solely for their affinity for the SEB binding site in CD28. Surface plasmon resonance showed that CD28 binds directly to SEB, whether one or the other was immobilized and even when both were free in solution, and specifically to peptide mimetics of the SEB β-strand( 8) /hinge/α-helix( 4) domain but not, or poorly, to mutant peptides in which 1–2 amino acids were changed to alanine. Conversely, another mutation in such a mimetic peptide, leading to higher SEB antagonist activity[15], enhanced its affinity for CD28. This peptide blocked binding of SEB to cell-surface CD28. Thus, SEB uses its β-strand( 8) /hinge/α-helix( 4) domain to bind CD28. We provide evidence that the superantigen binding site in CD28 is the dimer interface. CD28 mimetic peptides p1TA and p2TA corresponding to dimer interface sequences each inhibited Th1 cytokine gene induction by superantigens as widely different as SEB, SEA, and TSST-1 and protected mice from lethal SEB challenge even when present in about equimolar ratio to toxin. p1TA and p2TA directly bound SEB as well as SEA, with affinities close to that of CD28. The antagonist activity of these peptides is explained by their ability to compete with cell-surface CD28 for superantigen. p1TA and p2TA each blocked binding of SEB to cell-surface CD28. Each of five peptides overlapping distinct portions of the CD28 dimer interface was a superantigen antagonist. We did not perform mutagenesis of the CD28 dimer interface because mutations therein abrogate cell surface expression of the CD28 receptor[21], rendering interpretation problematic. The CD28 dimer interface has no known role in costimulation and is remote from the B7 binding site. SEB induces vigorous expression of Th1 and Th2 cytokines but only the Th1 response, here defined by induction of IL2, IFN-γ, and TNF-α genes, depends on CD28 engagement. By attenuating this response, antagonist peptides will reduce the synergy between these cytokines, to allow survival. Our data support the interpretation that the early Th2 cytokine response induced by a superantigen, as opposed to Th2 cell differentiation[26], is independent of CD28 signaling and hence resistant to blocking by the mimetic peptides. It is tempting to link the selective need for CD28 signaling in the superantigen-mediated induction of IL2, IFN-γ, and TNF-α genes, but not of IL4 and IL10 genes, to the distinct transcription factor dependence for expression of Th1- versus Th2-type cytokine genes, involving, respectively, T-bet[27],[28] and GATA-3[29]. CD28 monomers do not dimerize at low concentrations, indicating weak noncovalent interaction[21], facilitating displacement by superantigen. Engagement of a superantigen should displace contacts between the CD28 monomers which are linked through a disulfide bond[21] beyond the HVKGKH motif in the dimer interface. This epitope, buried in resting T cells, was rendered accessible upon exposure to SEB. Our data support the interpretation that the superantigen induces conformational change in CD28 that activates signaling, without interfering with B7-2/CD28 costimulation. More commonly, ligand binding induces receptor homodimerization needed for signaling, as for human growth hormone receptor[30] and erythropoietin receptor[31]. In contrast, the superantigen-binding domain of CD28 overlaps with its homodimer interface. In our mouse model, an accepted model for the lethality of superantigens[15], most if not all of the mice died within 48 h after a single exposure to SEB but they were protected by peptides. By contrast, HLA DR3 transgenic mice were far less sensitive to SEB challenge, requiring two toxin exposures 48 h apart before death was induced, and no protection by a β-strand( 8) /hinge/α-helix( 4) superantigen domain mimetic peptide was observed thereafter[32]. The rapid kinetics of toxin action in our mouse model reflects the rapid course of toxic shock, which was blocked by the peptides despite their limited halflife. In human PBMC, induction of Th1 cytokine gene expression, the immediate response occurring during the first hours after superantigen exposure, was sensitive to peptides. Binding of SEB to PBMC proved insensitive to β-strand( 8) /hinge/α-helix( 4) domain mimetic peptide[32] as expected, because SEB will also bind to the MHC-II and TCR, which unlike CD28 are not targeted by this peptide. The phenotype of tk2 mutant SEB shows that superantigen activity not only requires binding of CD28 via the β-strand( 8) /hinge/α-helix( 4) domain but that this binding must be with moderate affinity. The mutation led to significantly higher affinity for CD28 but this binding was nonproductive, as reflected by the complete loss of lethality of SEB. Mutations in a holoprotein can change protein conformation and thereby affect ligand binding properties. However, conformational changes in an intact protein are not necessarily reflected in short peptides derived from a limited domain; thus, in peptide p12YK, the same mutation led to loss of affinity for CD28. Given the similarly moderate affinity of SEB for CD28, TCR, and MHC-II, we propose that it is the concerted interaction of the superantigen with all three receptors that allows for stable synapse formation resulting in exceptionally strong Th1 cytokine induction and lethality. This affinity limitation will render superantigen action sensitive to regulation and allow peptide mimetics that interfere with binding of superantigen to CD28 to disrupt synapse formation, thus preventing toxic shock. Two classes of peptides block the interaction between superantigen and CD28: β-strand/hinge/α-helix domain mimetics that compete with superantigen for CD28 and CD28 homodimer interface mimetics that compete with CD28 for superantigen.
Induction of protective immunity involves the expression of inflammatory cytokines , proteins that mediate and respond to immune signals . However , excessive cytokine induction can lead to disease , including , at very high levels , lethal toxic shock . Staphylococcal and streptococcal superantigens are a broad family of bacterial protein toxins that induce such a lethal cytokine storm , orders of magnitude higher in intensity than that elicited during normal immune responses . A key participant in every immune response is the costimulatory receptor CD28 , which forms a protein dimer to mediate the immune response . Hitherto , CD28 was not known to bind microbial components . Here , we show that superantigens co-opt CD28 as their receptor and that to induce a cytokine storm , superantigens must bind directly into the dimer interface of CD28 . The interaction between CD28 and the bacterial superantigen can be blocked with peptides—short protein fragments that mimic the contact domains in the intact superantigen or in CD28 . These peptides attenuate inflammatory cytokine gene induction and thus protect animals from lethal toxic shock . Our finding that CD28 is a receptor for the superantigen toxins broadens the scope of microbial pathogen recognition mechanisms and provides a novel approach for designing therapeutics that protect against toxic shock .