RESUMEN
Central to adaptive immunity is the interaction between the αß T cell receptor (TCR) and peptide presented by the major histocompatibility complex (MHC) molecule. Presumably reflecting TCR-MHC bias and T cell signaling constraints, the TCR universally adopts a canonical polarity atop the MHC. We report the structures of two TCRs, derived from human induced T regulatory (iT(reg)) cells, complexed to an MHC class II molecule presenting a proinsulin-derived peptide. The ternary complexes revealed a 180° polarity reversal compared to all other TCR-peptide-MHC complex structures. Namely, the iT(reg) TCR α-chain and ß-chain are overlaid with the α-chain and ß-chain of MHC class II, respectively. Nevertheless, this TCR interaction elicited a peptide-reactive, MHC-restricted T cell signal. Thus TCRs are not 'hardwired' to interact with MHC molecules in a stereotypic manner to elicit a T cell signal, a finding that fundamentally challenges our understanding of TCR recognition.
Asunto(s)
Autoantígenos/metabolismo , Complejo Mayor de Histocompatibilidad/inmunología , Receptores de Antígenos de Linfocitos T/metabolismo , Inmunidad Adaptativa , Presentación de Antígeno , Autoantígenos/química , Autoantígenos/genética , Células Cultivadas , Antígeno HLA-DR4/química , Antígeno HLA-DR4/genética , Antígeno HLA-DR4/metabolismo , Antígenos de Histocompatibilidad Clase II/química , Antígenos de Histocompatibilidad Clase II/genética , Antígenos de Histocompatibilidad Clase II/metabolismo , Humanos , Complejo Mayor de Histocompatibilidad/genética , Modelos Moleculares , Mutagénesis Sitio-Dirigida , Proinsulina/química , Proinsulina/genética , Proinsulina/inmunología , Dominios y Motivos de Interacción de Proteínas , Receptores de Antígenos de Linfocitos T/química , Receptores de Antígenos de Linfocitos T/genética , Linfocitos T Reguladores/inmunologíaRESUMEN
The central immunological role of HLA class I (HLA-I) in presenting peptide Ags to cellular components of the immune system has been the focus of intense study for >60 y. A confounding factor in the study of HLA-I has been the extreme polymorphism of these molecules. The mAb W6/32 has been a fundamental reagent bypassing the issue of polymorphism by recognizing an epitope that is conserved across diverse HLA-I allotypes. However, despite the widespread use of W6/32, the epitope of this Ab has not been definitively mapped. In this study, we present the crystal structure of the Fab fragment of W6/32 in complex with peptide-HLA-B*27:05. W6/32 bound to HLA-B*27:05 beneath the Ag-binding groove, recognizing a discontinuous epitope comprised of the α1, α2, and α3 domains of HLA-I and ß2-microglobulin. The epitope comprises a region of low polymorphism reflecting the pan-HLA-I nature of the binding. Notably, the W6/32 epitope neither overlaps the HLA-I binding sites of either T cell Ag receptors or killer cell Ig-like receptors. However, it does coincide with the binding sites for leukocyte Ig-like receptors and CD8 coreceptors. Consistent with this, the use of W6/32 to block the interaction of NK cells with HLA-I only weakly impaired inhibition mediated by KIR3DL1, but impacted HLA-LILR recognition.
RESUMEN
Type I interferons are important in regulating immune responses to pathogens and tumors. All interferons are considered to signal via the heterodimeric IFNAR1-IFNAR2 complex, yet some subtypes such as interferon-ß (IFN-ß) can exhibit distinct functional properties, although the molecular basis of this is unclear. Here we demonstrate IFN-ß can uniquely and specifically ligate to IFNAR1 in an IFNAR2-independent manner, and we provide the structural basis of the IFNAR1-IFN-ß interaction. The IFNAR1-IFN-ß complex transduced signals that modulated expression of a distinct set of genes independently of Jak-STAT pathways. Lipopolysaccharide-induced sepsis was ameliorated in Ifnar1(-/-) mice but not Ifnar2(-/-) mice, suggesting that IFNAR1-IFN-ß signaling is pathologically relevant. Thus, we provide a molecular basis for understanding specific functions of IFN-ß.
Asunto(s)
Interferón beta/química , Interferón beta/metabolismo , Receptor de Interferón alfa y beta/química , Receptor de Interferón alfa y beta/metabolismo , Transducción de Señal , Animales , Modelos Animales de Enfermedad , Femenino , Lipopolisacáridos/efectos adversos , Ratones , Ratones Noqueados , Modelos Moleculares , Complejos Multiproteicos/química , Complejos Multiproteicos/metabolismo , Unión Proteica , Conformación Proteica , Estabilidad Proteica , Receptor de Interferón alfa y beta/genética , Choque Séptico/inducido químicamente , Choque Séptico/genética , Choque Séptico/metabolismo , Choque Séptico/mortalidadRESUMEN
Activating and inhibitory receptors on natural killer (NK) cells have a crucial role in innate immunity, although the basis of the engagement of activating NK cell receptors is unclear. The activating receptor Ly49H confers resistance to infection with murine cytomegalovirus by binding to the 'immunoevasin' m157. We found that m157 bound to the helical stalk of Ly49H, whereby two m157 monomers engaged the Ly49H dimer. The helical stalks of Ly49H lay centrally across the m157 platform, whereas its lectin domain was not required for recognition. Instead, m157 targeted an 'aromatic peg motif' present in stalks of both activating and inhibitory receptors of the Ly49 family, and substitution of this motif abrogated binding. Furthermore, ligation of m157 to Ly49H or Ly49C resulted in intracellular signaling. Accordingly, m157 has evolved to 'tackle the legs' of a family of NK cell receptors.
Asunto(s)
Infecciones por Herpesviridae/inmunología , Antígenos de Histocompatibilidad Clase I/inmunología , Inmunidad Innata/inmunología , Células Asesinas Naturales/inmunología , Muromegalovirus/inmunología , Subfamilia A de Receptores Similares a Lectina de Células NK/inmunología , Secuencias de Aminoácidos/inmunología , Secuencia de Aminoácidos , Animales , Cristalografía por Rayos X , Femenino , Ratones , Ratones Endogámicos BALB C , Modelos Moleculares , Datos de Secuencia Molecular , Transducción de Señal/inmunología , Organismos Libres de Patógenos Específicos , Resonancia por Plasmón de SuperficieRESUMEN
Psoriasis is a chronic skin disease characterized by hyperproliferative epidermal lesions infiltrated by autoreactive T cells. Individuals expressing the human leukocyte antigen (HLA) C∗06:02 allele are at highest risk for developing psoriasis. An autoreactive T cell clone (termed Vα3S1/Vß13S1) isolated from psoriatic plaques is selective for HLA-C∗06:02, presenting a peptide derived from the melanocyte-specific autoantigen ADAMTSL5 (VRSRRCLRL). Here we determine the crystal structure of this psoriatic TCR-HLA-C∗06:02 ADAMTSL5 complex with a stabilized peptide. Docking of the TCR involves an extensive complementary charge network formed between negatively charged TCR residues interleaving with exposed arginine residues from the self-peptide and the HLA-C∗06:02 α1 helix. We probed these interactions through mutagenesis and activation assays. The charged interface spans the polymorphic region of the C1/C2 HLA group. Notably the peptide-binding groove of HLA-C∗06:02 appears exquisitely suited for presenting highly charged Arg-rich epitopes recognized by this acidic psoriatic TCR. Overall, we provide a structural basis for understanding the engagement of melanocyte antigen-presenting cells by a TCR implicated in psoriasis while simultaneously expanding our knowledge of how TCRs engage HLA-C.
Asunto(s)
Antígenos HLA-C , Psoriasis , Humanos , Electricidad Estática , Péptidos/química , Psoriasis/patología , Receptores de Antígenos de Linfocitos T/genética , Proteínas ADAMTSRESUMEN
Type I natural killer T cells (NKT cells) are characterized by an invariant variable region 14-joining region 18 (V(α)14-J(α)18) T cell antigen receptor (TCR) α-chain and recognition of the glycolipid α-galactosylceramide (α-GalCer) restricted to the antigen-presenting molecule CD1d. Here we describe a population of α-GalCer-reactive NKT cells that expressed a canonical V(α)10-J(α)50 TCR α-chain, which showed a preference for α-glucosylceramide (α-GlcCer) and bacterial α-glucuronic acid-containing glycolipid antigens. Structurally, despite very limited TCRα sequence identity, the V(α)10 TCR-CD1d-α-GlcCer complex had a docking mode similar to that of type I TCR-CD1d-α-GalCer complexes, although differences at the antigen-binding interface accounted for the altered antigen specificity. Our findings provide new insight into the structural basis and evolution of glycolipid antigen recognition and have notable implications for the scope and immunological role of glycolipid-specific T cell responses.
Asunto(s)
Galactosilceramidas/inmunología , Células T Asesinas Naturales/inmunología , Receptores de Antígenos de Linfocitos T alfa-beta/inmunología , Adyuvantes Inmunológicos/farmacología , Secuencia de Aminoácidos , Animales , Antígenos Bacterianos/inmunología , Antígenos CD1d/inmunología , Línea Celular , Galactosilceramidas/farmacología , Glucuronatos/inmunología , Humanos , Ratones , Ratones Mutantes , Datos de Secuencia Molecular , Receptores de Antígenos de Linfocitos T alfa-beta/genéticaRESUMEN
HLA class I molecules that represent ligands for the inhibitory killer cell Ig-like receptor (KIR) 3DL1 found on NK cells are categorically defined as those HLA-A and HLA-B allotypes containing the Bw4 motif, yet KIR3DL1 demonstrates hierarchical recognition of these HLA-Bw4 ligands. To better understand the molecular basis underpinning differential KIR3DL1 recognition, the HLA-ABw4 family of allotypes were investigated. Transfected human 721.221 cells expressing HLA-A*32:01 strongly inhibited primary human KIR3DL1+ NK cells, whereas HLA-A*24:02 and HLA-A*23:01 displayed intermediate potency and HLA-A*25:01 failed to inhibit activation of KIR3DL1+ NK cells. Structural studies demonstrated that recognition of HLA-A*24:02 by KIR3DL1 used identical contacts as the potent HLA-B*57:01 ligand. Namely, the D1-D2 domains of KIR3DL1 were placed over the α1 helix and α2 helix of the HLA-A*24:02 binding cleft, respectively, whereas the D0 domain contacted the side of the HLA-A*24:02 molecule. Nevertheless, functional analyses showed KIR3DL1 recognition of HLA-A*24:02 was more sensitive to substitutions within the α2 helix of HLA-A*24:02, including residues Ile142 and Lys144 Furthermore, the presence of Thr149 in the α2 helix of HLA-A*25:01 abrogated KIR3DL1+ NK inhibition. Together, these data demonstrate a role for the HLA class I α2 helix in determining the hierarchy of KIR3DL1 ligands. Thus, recognition of HLA class I is dependent on a complex interplay between the peptide repertoire, polymorphisms within and proximal to the Bw4 motif, and the α2 helix. Collectively, the data furthers our understanding of KIR3DL1 ligands and will inform genetic association and immunogenetics studies examining the role of KIR3DL1 in disease settings.
Asunto(s)
Antígenos HLA-A , Células Asesinas Naturales , Receptores KIR3DL1 , Antígenos HLA-A/química , Antígenos HLA-A/inmunología , Humanos , Células Asesinas Naturales/química , Células Asesinas Naturales/inmunología , Conformación Proteica en Hélice alfa , Dominios Proteicos , Receptores KIR3DL1/química , Receptores KIR3DL1/inmunologíaRESUMEN
Micropolymorphisms within human leukocyte antigen (HLA) class I molecules can change the architecture of the peptide-binding cleft, leading to differences in peptide presentation and T cell recognition. The impact of such HLA variation on natural killer (NK) cell recognition remains unclear. Given the differential association of HLA-B*57:01 and HLA-B*57:03 with the control of HIV, recognition of these HLA-B57 allomorphs by the killer cell immunoglobulin-like receptor (KIR) 3DL1 was compared. Despite differing by only two polymorphic residues, both buried within the peptide-binding cleft, HLA-B*57:01 more potently inhibited NK cell activation. Direct-binding studies showed KIR3DL1 to preferentially recognize HLA-B*57:01, particularly when presenting peptides with positively charged position (P)Ω-2 residues. In HLA-B*57:01, charged PΩ-2 residues were oriented toward the peptide-binding cleft and away from KIR3DL1. In HLA-B*57:03, the charged PΩ-2 residues protruded out from the cleft and directly impacted KIR3DL1 engagement. Accordingly, KIR3DL1 recognition of HLA class I ligands is modulated by both the peptide sequence and conformation, as determined by the HLA polymorphic framework, providing a rationale for understanding differences in clinical associations.
Asunto(s)
Antígenos de Histocompatibilidad Clase I/genética , Células Asesinas Naturales/fisiología , Polimorfismo Genético/genética , Antígenos de Histocompatibilidad Clase I/química , Antígenos de Histocompatibilidad Clase I/fisiología , Humanos , Activación de Linfocitos/genética , Modelos Moleculares , Polimorfismo Genético/fisiología , Receptores KIR/genéticaRESUMEN
BACKGROUND AND AIMS: Growth hormone (GH) is important for liver regeneration after partial hepatectomy (PHx). We investigated this process in C57BL/6 mice that express different forms of the GH receptor (GHR) with deletions in key signaling domains. APPROACH AND RESULTS: PHx was performed on C57BL/6 mice lacking GHR (Ghr-/- ), disabled for all GH-dependent Janus kinase 2 signaling (Box1-/- ), or lacking only GH-dependent signal transducer and activator of transcription 5 (STAT5) signaling (Ghr391-/- ), and wild-type littermates. C57BL/6 Ghr-/- mice showed striking mortality within 48 hours after PHx, whereas Box1-/- or Ghr391-/- mice survived with normal liver regeneration. Ghr-/- mortality was associated with increased apoptosis and elevated natural killer/natural killer T cell and macrophage cell markers. We identified H2-Bl, a key immunotolerance protein, which is up-regulated by PHx through a GH-mediated, Janus kinase 2-independent, SRC family kinase-dependent pathway. GH treatment was confirmed to up-regulate expression of the human homolog of H2-Bl (human leukocyte antigen G [HLA-G]) in primary human hepatocytes and in the serum of GH-deficient patients. We find that injury-associated innate immune attack by natural killer/natural killer T cell and macrophage cells are instrumental in the failure of liver regeneration, and this can be overcome in Ghr-/- mice by adenoviral delivery of H2-Bl or by infusion of HLA-G protein. Further, H2-Bl knockdown in wild-type C57BL/6 mice showed elevated markers of inflammation after PHx, whereas Ghr-/- backcrossed on a strain with high endogenous H2-Bl expression showed a high rate of survival following PHx. CONCLUSIONS: GH induction of H2-Bl expression is crucial for reducing innate immune-mediated apoptosis and promoting survival after PHx in C57BL/6 mice. Treatment with HLA-G may lead to improved clinical outcomes following liver surgery or transplantation.
Asunto(s)
Hormona del Crecimiento/deficiencia , Antígenos H-2/metabolismo , Antígenos HLA-G/metabolismo , Regeneración Hepática/inmunología , Hígado/fisiología , Animales , Apoptosis/inmunología , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Células Cultivadas , Técnicas de Cocultivo , Técnicas de Silenciamiento del Gen , Antígenos H-2/genética , Antígenos HLA-G/genética , Antígenos HLA-G/aislamiento & purificación , Hepatectomía , Hepatocitos , Humanos , Inmunidad Innata , Células Asesinas Naturales/inmunología , Células Asesinas Naturales/metabolismo , Hígado/cirugía , Macrófagos/inmunología , Macrófagos/metabolismo , Ratones , Células T Asesinas Naturales/inmunología , Células T Asesinas Naturales/metabolismo , Cultivo Primario de Células , Proteínas Recombinantes/genética , Proteínas Recombinantes/inmunología , Proteínas Recombinantes/metabolismo , Transducción de Señal/genética , Transducción de Señal/inmunologíaRESUMEN
The race to identify a successful treatment for COVID19 will be defined by fundamental research into the replication cycle of the SARS-CoV-2 virus. This has identified five distinct stages from which numerous vaccination and clinical trials have emerged alongside an innumerable number of drug discovery studies currently in development for disease intervention. Informing every step of the viral replication cycle has been an unprecedented 'call-to-arms' by the global structural biology community. Of the 20 main SARS-CoV-2 proteins, 13 have been resolved structurally for SARS-CoV-2 with most having a related SARS-CoV and MERS-CoV structural homologue totalling some 300 structures currently available in public repositories. Herein, we review the contribution of structural studies to our understanding of the virus and their role in structure-based development of therapeutics.
Asunto(s)
Antivirales/química , Antivirales/uso terapéutico , COVID-19/terapia , Descubrimiento de Drogas/métodos , SARS-CoV-2 , Antivirales/síntesis química , COVID-19/inmunología , Desarrollo de Medicamentos/métodos , Genoma Viral , Humanos , Modelos Moleculares , Elementos Estructurales de las Proteínas , SARS-CoV-2/química , SARS-CoV-2/efectos de los fármacos , SARS-CoV-2/inmunología , SARS-CoV-2/fisiología , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/fisiología , Relación Estructura-Actividad , Proteínas Estructurales Virales/química , Proteínas Estructurales Virales/fisiología , Replicación Viral/efectos de los fármacos , Replicación Viral/fisiología , Tratamiento Farmacológico de COVID-19RESUMEN
The antigen receptor for natural killer T cells (NKT TCR) binds CD1d-restricted microbial and self-lipid antigens, although the molecular basis of self-CD1d recognition is unclear. Here, we have characterized NKT TCR recognition of CD1d molecules loaded with natural self-antigens (Ags) and report the 2.3 Å resolution structure of an autoreactive NKT TCR-phosphatidylinositol-CD1d complex. NKT TCR recognition of self- and foreign antigens was underpinned by a similar mode of germline-encoded recognition of CD1d. However, NKT TCR autoreactivity is mediated by unique sequences within the non-germline-encoded CDR3ß loop encoding for a hydrophobic motif that promotes self-association with CD1d. Accordingly, NKT cell autoreactivity may arise from the inherent affinity of the interaction between CD1d and the NKT TCR, resulting in the recognition of a broad range of CD1d-restricted self-antigens. This demonstrates that multiple self-antigens can be recognized in a similar manner by autoreactive NKT TCRs.
Asunto(s)
Antígenos CD1d/inmunología , Autoantígenos , Células T Asesinas Naturales/inmunología , Animales , Cristalografía por Rayos X , Ratones , Ratones Endogámicos C57BL , Modelos Moleculares , Complejos Multiproteicos , Receptores de Células Asesinas Naturales/inmunologíaRESUMEN
The surveillance of target cells by natural killer (NK) cells utilizes an ensemble of inhibitory and activating receptors, many of which interact with major histocompatibility complex (MHC) class I molecules. NK cell recognition of MHC class I proteins is important developmentally for the acquisition of full NK cell effector capacity and during target cell recognition, where the engagement of inhibitory receptors and MHC class I molecules attenuates NK cell activation. Human NK cells have evolved two broad strategies for recognition of human leukocyte antigen (HLA) class I molecules: (i) direct recognition of polymorphic classical HLA class I proteins by diverse receptor families such as the killer cell immunoglobulin-like receptors (KIRs), and (ii) indirect recognition of conserved sets of HLA class I-derived peptides displayed on the non-classical HLA-E for recognition by CD94-NKG2 receptors. In this review, we assess the structural basis for the interaction between these NK receptors and their HLA class I ligands and, using the suite of published KIR and CD94-NKG2 ternary complexes, highlight the features that allow NK cells to orchestrate the recognition of a range of different HLA class I proteins.
Asunto(s)
Antígenos HLA/inmunología , Antígenos de Histocompatibilidad Clase I/inmunología , Células Asesinas Naturales/inmunología , Receptores de Células Asesinas Naturales/inmunología , Animales , Antígenos HLA/química , Antígenos HLA/metabolismo , Antígenos de Histocompatibilidad Clase I/química , Antígenos de Histocompatibilidad Clase I/metabolismo , Humanos , Células Asesinas Naturales/metabolismo , Ligandos , Modelos Moleculares , Unión Proteica , Estructura Terciaria de Proteína , Receptores de Células Asesinas Naturales/química , Receptores de Células Asesinas Naturales/metabolismoRESUMEN
The interaction of IFN-ß with its receptor IFNAR1 (interferon α/ß receptor subunit 1) is vital for host-protective anti-viral and anti-proliferative responses, but signaling via this interaction can be detrimental if dysregulated. Whereas it is established that IFNAR1 is an essential component of the IFNAR signaling complex, the key residues underpinning the IFN-ß-IFNAR1 interaction are unknown. Guided by the crystal structure of the IFN-ß-IFNAR1 complex, we used truncation variants and site-directed mutagenesis to investigate domains and residues enabling complexation of IFN-ß to IFNAR1. We have identified an interface on IFNAR1-subdomain-3 that is differentially utilized by IFN-ß and IFN-α for signal transduction. We used surface plasmon resonance and cell-based assays to investigate this important IFN-ß binding interface that is centered on IFNAR1 residues Tyr240 and Tyr274 binding the C and N termini of the B and C helices of IFN-ß, respectively. Using IFNAR1 and IFN-ß variants, we show that this interface contributes significantly to the affinity of IFN-ß for IFNAR1, its ability to activate STAT1, the expression of interferon stimulated genes, and ultimately to the anti-viral and anti-proliferative properties of IFN-ß. These results identify a key interface created by IFNAR1 residues Tyr240 and Tyr274 interacting with IFN-ß residues Phe63, Leu64, Glu77, Thr78, Val81, and Arg82 that underlie IFN-ß-IFNAR1-mediated signaling and biological processes.
Asunto(s)
Interferón beta/metabolismo , Receptor de Interferón alfa y beta/metabolismo , Transducción de Señal/fisiología , Sustitución de Aminoácidos , Animales , Línea Celular , Interferón beta/genética , Ratones , Ratones Noqueados , Mutación Missense , Dominios Proteicos , Receptor de Interferón alfa y beta/genéticaRESUMEN
Human leukocyte antigen (HLA)-C*06:02 is identified as the allele associated with the highest risk for the development of the autoimmune skin disease psoriasis. However, the diversity and mode of peptide presentation by the HLA-C*06:02 molecule remains unclear. Here, we describe the endogenous peptide repertoire of â¼3,000 sequences for HLA-C*06:02 that defines the peptide-binding motif for this HLA allomorph. We found that HLA-C*06:02 predominantly presents nonamer peptides with dominant arginine anchors at the P2 and P7 positions and a preference for small hydrophobic residues at the C terminus (PΩ). To determine the structural basis of this selectivity, we determined crystal structures of HLA-C*06:02 in complex with two self-peptides (ARTELYRSL and ARFNDLRFV) and an analogue of a melanocyte autoantigen (ADAMTSL5, VRSRR-abu-LRL) implicated in psoriasis. These structures revealed that HLA-C*06:02 possesses a deep peptide-binding groove comprising two electronegative B- and E-pockets that coincide with the preference for P2 and P7 arginine anchors. The ADAMTSL5 autoantigen possessed a P7-Leu instead of the P7-Arg residue, but nevertheless was accommodated within the HLA-C*06:02 antigen-binding cleft. Collectively, our results provide the structural basis for understanding peptide repertoire selection in HLA-C*06:02.
Asunto(s)
Proteínas ADAMTS , Presentación de Antígeno , Antígenos HLA-C , Péptidos , Proteínas ADAMTS/química , Proteínas ADAMTS/genética , Proteínas ADAMTS/inmunología , Proteínas ADAMTS/metabolismo , Secuencias de Aminoácidos , Línea Celular , Antígenos HLA-C/química , Antígenos HLA-C/genética , Antígenos HLA-C/inmunología , Antígenos HLA-C/metabolismo , Humanos , Péptidos/química , Péptidos/genética , Péptidos/inmunología , Péptidos/metabolismoRESUMEN
Ligation of the alphabeta T cell receptor (TCR) by a specific peptide-loaded major histocompatibility complex (pMHC) molecule initiates T cell signaling via the CD3 complex. However, the initial events that link antigen recognition to T cell signal transduction remain unclear. Here we show, via fluorescence-based experiments and structural analyses, that MHC-restricted antigen recognition by the alphabeta TCR results in a specific conformational change confined to the A-B loop within the alpha chain of the constant domain (Calpha). The apparent affinity constant of this A-B loop movement mirrored that of alphabeta TCR-pMHC ligation and was observed in two alphabeta TCRs with distinct pMHC specificities. The Ag-induced A-B loop conformational change could be inhibited by fixing the juxtapositioning of the constant domains and was shown to be reversible upon pMHC disassociation. Notably, the loop movement within the Calpha domain, although specific for an agonist pMHC ligand, was not observed with a pMHC antagonist. Moreover, mutagenesis of residues within the A-B loop impaired T cell signaling in an in vitro system of antigen-specific TCR stimulation. Collectively, our findings provide a basis for the earliest molecular events that underlie Ag-induced T cell triggering.
Asunto(s)
Antígenos/química , Receptores de Antígenos de Linfocitos T alfa-beta/química , Linfocitos T/inmunología , Animales , Antígenos/inmunología , Humanos , Complejo Mayor de Histocompatibilidad/inmunología , Mutación/genética , Péptidos/química , Péptidos/inmunología , Unión Proteica/inmunología , Estructura Terciaria de Proteína , Receptores de Antígenos de Linfocitos T alfa-beta/genética , Receptores de Antígenos de Linfocitos T alfa-beta/inmunologíaRESUMEN
Human leukocyte antigens (HLAs) are highly polymorphic proteins that initiate immunity by presenting pathogen-derived peptides to T cells. HLA polymorphisms mostly map to the antigen-binding cleft, thereby diversifying the repertoire of self-derived and pathogen-derived peptide antigens selected by different HLA allotypes. A growing number of immunologically based drug reactions, including abacavir hypersensitivity syndrome (AHS) and carbamazepine-induced Stevens-Johnson syndrome (SJS), are associated with specific HLA alleles. However, little is known about the underlying mechanisms of these associations, including AHS, a prototypical HLA-associated drug reaction occurring exclusively in individuals with the common histocompatibility allele HLA-B*57:01, and with a relative risk of more than 1,000 (refs 6, 7). We show that unmodified abacavir binds non-covalently to HLA-B*57:01, lying across the bottom of the antigen-binding cleft and reaching into the F-pocket, where a carboxy-terminal tryptophan typically anchors peptides bound to HLA-B*57:01. Abacavir binds with exquisite specificity to HLA-B*57:01, changing the shape and chemistry of the antigen-binding cleft, thereby altering the repertoire of endogenous peptides that can bind HLA-B*57:01. In this way, abacavir guides the selection of new endogenous peptides, inducing a marked alteration in 'immunological self'. The resultant peptide-centric 'altered self' activates abacavir-specific T-cells, thereby driving polyclonal CD8 T-cell activation and a systemic reaction manifesting as AHS. We also show that carbamazepine, a widely used anti-epileptic drug associated with hypersensitivity reactions in HLA-B*15:02 individuals, binds to this allotype, producing alterations in the repertoire of presented self peptides. Our findings simultaneously highlight the importance of HLA polymorphism in the evolution of pharmacogenomics and provide a general mechanism for some of the growing number of HLA-linked hypersensitivities that involve small-molecule drugs.
Asunto(s)
Presentación de Antígeno/efectos de los fármacos , Autoinmunidad/efectos de los fármacos , Autoinmunidad/inmunología , Didesoxinucleósidos/farmacología , Antígenos HLA-B/inmunología , Linfocitos T/efectos de los fármacos , Linfocitos T/inmunología , Sitios de Unión , Donantes de Sangre , Linfocitos T CD8-positivos/efectos de los fármacos , Linfocitos T CD8-positivos/inmunología , Carbamazepina/farmacología , Hipersensibilidad a las Drogas , Antígenos HLA-B/química , Humanos , Modelos Moleculares , Conformación Proteica , SíndromeRESUMEN
The killer cell Ig-like receptor 3DL1 (KIR3DL1) inhibits activation of NK cells upon interaction with HLA class I molecules such as HLA-B*57:01, which contains the Bw4 epitope spanning residues 77-83 (e.g., NLRIALR), and not with HLA allomorphs that possess the Bw6 motif (e.g., HLA-B*08:01), which differ at residues 77, 80, 81, 82, and 83. Although Bw4 residues Ile(80) and Arg(83) directly interact with KIR3DL1*001, their precise role in determining KIR3DL1-HLA-Bw4 specificity remains unclear. Recognition of HLA-B*57:01 by either KIR3DL1(+) NK cells or the NK cell line YTS transfected with KIR3DL1*001 was impaired by mutation of residues 80 and 83 of HLA-B*57:01 to the corresponding amino acids within the Bw6 motif. Conversely, the simultaneous introduction of three Bw4 residues at positions 80, 82, and 83 into HLA-B*08:01 conferred an interaction with KIR3DL1*001. Structural analysis of HLA-B*57:01, HLA-B*08:01, and mutants of each bearing substitutions at positions 80 and 83 revealed that Ile(80) and Arg(83) within the Bw4 motif constrain the conformation of Glu(76), primarily through a salt bridge between Arg(83) and Glu(76). This salt bridge was absent in HLA-Bw6 molecules as well as position 83 mutants of HLA-B*57:01. Mutation of the Bw4 residue Ile(80) also disrupted this salt bridge, providing further insight into the role that position 80 plays in mediating KIR3DL1 recognition. Thus, the strict conformation of HLA-Bw4 allotypes, held in place by the Glu(76)-Arg(83) interaction, facilitates KIR3DL1 binding, whereas Bw6 allotypes present a platform on the α1 helix that is less permissive for KIR3DL1 binding.
Asunto(s)
Epítopos , Antígenos HLA-B , Antígeno HLA-B8 , Receptores KIR3DL1 , Secuencias de Aminoácidos , Línea Celular , Epítopos/genética , Epítopos/inmunología , Antígenos HLA-B/genética , Antígenos HLA-B/inmunología , Antígeno HLA-B8/genética , Antígeno HLA-B8/inmunología , Humanos , Células Asesinas Naturales/citología , Células Asesinas Naturales/inmunología , Mutación , Receptores KIR3DL1/genética , Receptores KIR3DL1/inmunologíaRESUMEN
Members of the killer cell immunoglobulin-like receptor (KIR) family, a large group of polymorphic receptors expressed on natural killer (NK) cells, recognize particular peptide-laden human leukocyte antigen (pHLA) class I molecules and have a pivotal role in innate immune responses. Allelic variation and extensive polymorphism within the three-domain KIR family (KIR3D, domains D0-D1-D2) affects pHLA binding specificity and is linked to the control of viral replication and the treatment outcome of certain haematological malignancies. Here we describe the structure of a human KIR3DL1 receptor bound to HLA-B*5701 complexed with a self-peptide. KIR3DL1 clamped around the carboxy-terminal end of the HLA-B*5701 antigen-binding cleft, resulting in two discontinuous footprints on the pHLA. First, the D0 domain, a distinguishing feature of the KIR3D family, extended towards ß2-microglobulin and abutted a region of the HLA molecule with limited polymorphism, thereby acting as an 'innate HLA sensor' domain. Second, whereas the D2-HLA-B*5701 interface exhibited a high degree of complementarity, the D1-pHLA-B*5701 contacts were suboptimal and accommodated a degree of sequence variation both within the peptide and the polymorphic region of the HLA molecule. Although the two-domain KIR (KIR2D) and KIR3DL1 docked similarly onto HLA-C and HLA-B respectively, the corresponding D1-mediated interactions differed markedly, thereby providing insight into the specificity of KIR3DL1 for discrete HLA-A and HLA-B allotypes. Collectively, in association with extensive mutagenesis studies at the KIR3DL1-pHLA-B*5701 interface, we provide a framework for understanding the intricate interplay between peptide variability, KIR3D and HLA polymorphism in determining the specificity requirements of this essential innate interaction that is conserved across primate species.
Asunto(s)
Antígenos HLA-B/química , Antígenos HLA-B/inmunología , Receptores KIR3DL1/química , Receptores KIR3DL1/inmunología , Secuencia de Aminoácidos , Sitios de Unión/genética , Antígenos HLA-B/genética , Humanos , Modelos Moleculares , Proteínas Mutantes/química , Proteínas Mutantes/genética , Proteínas Mutantes/inmunología , Polimorfismo Genético/genética , Estructura Terciaria de Proteína , Receptores KIR3DL1/genética , Relación Estructura-Actividad , Microglobulina beta-2/química , Microglobulina beta-2/inmunologíaRESUMEN
The engagement of natural killer cell immunoglobulin-like receptors (KIRs) with their target ligands, human leukocyte antigen (HLA) molecules, is a critical component of innate immunity. Structurally, KIRs typically have either two (D1-D2) or three (D0-D1-D2) extracellular immunoglobulin domains, with the D1 and D2 domain recognizing the α1 and α2 helices of HLA, respectively, whereas the D0 domain of the KIR3DLs binds a loop region flanking the α1 helix of the HLA molecule. KIR2DL4 is distinct from other KIRs (except KIR2DL5) in that it does not contain a D1 domain and instead has a D0-D2 arrangement. Functionally, KIR2DL4 is also atypical in that, unlike all other KIRs, KIR2DL4 has both activating and inhibitory signaling domains. Here, we determined the 2.8 Å crystal structure of the extracellular domains of KIR2DL4. Structurally, KIR2DL4 is reminiscent of other KIR2DL receptors, with the D0 and D2 adopting the C2-type immunoglobulin fold arranged with an acute elbow angle. However, KIR2DL4 self-associated via the D0 domain in a concentration-dependent manner and was observed as a tetramer in the crystal lattice by size exclusion chromatography, dynamic light scattering, analytical ultracentrifugation, and small angle x-ray scattering experiments. The assignment of residues in the D0 domain to forming the KIR2DL4 tetramer precludes an interaction with HLA akin to that observed for KIR3DL1. Accordingly, no interaction was observed to HLA by direct binding studies. Our data suggest that the unique functional properties of KIR2DL4 may be mediated by self-association of the receptor.