Microbiology & Immunology - Research Publications
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ItemNatural micropolymorphism in human leukocyte antigens provides a basis for genetic control of antigen recognition(ROCKEFELLER UNIV PRESS, 2009-01-16)Human leukocyte antigen (HLA) gene polymorphism plays a critical role in protective immunity, disease susceptibility, autoimmunity, and drug hypersensitivity, yet the basis of how HLA polymorphism influences T cell receptor (TCR) recognition is unclear. We examined how a natural micropolymorphism in HLA-B44, an important and large HLA allelic family, affected antigen recognition. T cell-mediated immunity to an Epstein-Barr virus determinant (EENLLDFVRF) is enhanced when HLA-B*4405 was the presenting allotype compared with HLA-B*4402 or HLA-B*4403, each of which differ by just one amino acid. The micropolymorphism in these HLA-B44 allotypes altered the mode of binding and dynamics of the bound viral epitope. The structure of the TCR-HLA-B*4405(EENLLDFVRF) complex revealed that peptide flexibility was a critical parameter in enabling preferential engagement with HLA-B*4405 in comparison to HLA-B*4402/03. Accordingly, major histocompatibility complex (MHC) polymorphism can alter the dynamics of the peptide-MHC landscape, resulting in fine-tuning of T cell responses between closely related allotypes.
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ItemA minimal binding footprint on CD1d-glycolipid is a basis for selection of the unique human NKT TCR(ROCKEFELLER UNIV PRESS, 2008-04-14)Although it has been established how CD1 binds a variety of lipid antigens (Ag), data are only now emerging that show how alphabeta T cell receptors (TCRs) interact with CD1-Ag. Using the structure of the human semiinvariant NKT TCR-CD1d-alpha-galactosylceramide (alpha-GalCer) complex as a guide, we undertook an alanine scanning mutagenesis approach to define the energetic basis of this interaction between the NKT TCR and CD1d. Moreover, we explored how analogues of alpha-GalCer affected this interaction. The data revealed that an identical energetic footprint underpinned the human and mouse NKT TCR-CD1d-alpha-GalCer cross-reactivity. Some, but not all, of the contact residues within the Jalpha18-encoded invariant CDR3alpha loop and Vbeta11-encoded CDR2beta loop were critical for recognizing CD1d. The residues within the Valpha24-encoded CDR1alpha and CDR3alpha loops that contacted the glycolipid Ag played a smaller energetic role compared with the NKT TCR residues that contacted CD1d. Collectively, our data reveal that the region distant to the protruding Ag and directly above the F' pocket of CD1d was the principal factor in the interaction with the NKT TCR. Accordingly, although the structural footprint at the NKT TCR-CD1d-alpha-GalCer is small, the energetic footprint is smaller still, and reveals the minimal requirements for CD1d restriction.
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ItemThe immunogenicity of a viral cytotoxic T cell epitope is controlled by its MHC-bound conformation(ROCKEFELLER UNIV PRESS, 2005-11-07)Thousands of potentially antigenic peptides are encoded by an infecting pathogen; however, only a small proportion induce measurable CD8(+) T cell responses. To investigate the factors that control peptide immunogenicity, we have examined the cytotoxic T lymphocyte (CTL) response to a previously undefined epitope ((77)APQPAPENAY(86)) from the BZLF1 protein of Epstein-Barr virus (EBV). This peptide binds well to two human histocompatibility leukocyte antigen (HLA) allotypes, HLA-B*3501 and HLA-B*3508, which differ by a single amino acid at position 156 ((156)Leucine vs. (156)Arginine, respectively). Surprisingly, only individuals expressing HLA-B*3508 show evidence of a CTL response to the (77)APQPAPENAY(86) epitope even though EBV-infected cells expressing HLA-B*3501 process and present similar amounts of peptide for CTL recognition, suggesting that factors other than peptide presentation levels are influencing immunogenicity. Functional and structural analysis revealed marked conformational differences in the peptide, when bound to each HLA-B35 allotype, that are dictated by the polymorphic HLA residue 156 and that directly affected T cell receptor recognition. These data indicate that the immunogenicity of an antigenic peptide is influenced not only by how well the peptide binds to major histocompatibility complex (MHC) molecules but also by its bound conformation. It also illustrates a novel mechanism through which MHC polymorphism can further diversify the immune response to infecting pathogens.
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ItemHierarchical self-tolerance to T cell determinants within the ubiquitous nuclear self-antigen La (SS-B) permits induction of systemic autoimmunity in normal mice(ROCKEFELLER UNIV PRESS, 1996-11-01)Systemic autoimmune diseases are frequently associated with clustering of high titer autoantibody responses towards nuclear self-antigens. Little is known, however, about the extent of immune tolerance to the target nuclear antigens or the events leading to the complex autoantibody responses that are characteristic of systemic autoimmunity. To address these issues, we have examined the mouse immune response to La autoantigen (mLa) and the homologous human La antigen (hLa), which are components of the La(SS-B)/Ro(SS-A) ribonucleoprotein (RNP) complex targeted in systemic lupus erythematosus and primary Sjögren's syndrome. The findings reveal the presence of hierarchical T cell tolerance involving multiple autodeterminants within the La autoantigen expressed by normal H-2k and H-2a mice. At one end of this spectrum, there was no detectable T or B cell autoimmunity observed in mice that were immunized with the immunodominant mLa287-301 determinant, which differed by a single residue in its core sequence from the homologous but highly immunogenic human La288-302 determinant. Interestingly, the mLa287-301 peptide acted as an altered peptide ligand that specifically antagonized the activation of an hLa288-302-specific T cell hybridoma. In contrast to the tolerogenic mLa287-301 determinant, a range of autoimmune potential was identified among poorly tolerizing, subdominant self-peptides present within mouse La autoantigen. Notably, immunization of normal mice with the autologous subdominant La25-44 and La106-129 determinants resulted in limited or no detectable autoantibody response. In contrast, immunization with the subdominant mouse La13-30 determinant induced a proliferative T cell response associated with the appearance of specific autoantibodies recognizing multiple intrastructural (La) and intermolecular components (Ro) of the murine La/Ro RNP. The findings suggest how diversified autoimmunity might follow initiation of immunity to simple peptide mimics of poorly tolerogenic determinants that are present within ubiquitous self-antigens.
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ItemA 320-kilobase artificial chromosome encoding the human HLA DR3-DQ2 MHC haplotype confers HLA restriction in transgenic mice(AMER ASSOC IMMUNOLOGISTS, 2002-03-15)MHC class II haplotypes control the specificity of Th immune responses and susceptibility to many autoimmune diseases. Understanding the role of HLA class II haplotypes in immunity is hampered by the lack of animal models expressing these genes as authentic cis-haplotypes. In this study we describe transgenic expression of the autoimmune prone HLA DR3-DQ2 haplotype from a yeast artificial chromosome (YAC) containing an intact similar320-kb region from HLA DRA to DQB2. In YAC-transgenic mice HLA DR and DQ gene products were expressed on B cells, macrophages, and dendritic cells, but not on T cells indicating cell-specific regulation. Positive selection of the CD4 compartment by human class II molecules was 67% efficient in YAC-homozygous mice lacking endogenous class II molecules (Abeta(null/null)) and expressing only murine CD4. A broad range of TCR Vbeta families was used in the peripheral T cell repertoire, which was also purged of Vbeta5-, Vbeta11-, and Vbeta12-bearing T cells by endogenous mouse mammary tumor virus-encoded superantigens. Expression of the HLA DR3-DQ2 haplotype on the Abeta(null/null) background was associated with normal CD8-dependent clearance of virus from influenza-infected mice and development of CD4-dependent protection from otherwise lethal infection with Salmonella typhimurium. HLA DR- and DQ-restricted T cell responses were also elicited following immunization with known T cell determinants presented by these molecules. These findings demonstrate the potential for human MHC class II haplotypes to function efficiently in transgenic mice and should provide valuable tools for developing humanized models of MHC-associated autoimmune diseases.
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ItemQuantitative and qualitative influences of tapasin on the class I peptide repertoire(AMER ASSOC IMMUNOLOGISTS, 2001-01-15)Tapasin is critical for efficient loading and surface expression of most HLA class I molecules. The high level surface expression of HLA-B*2705 on tapasin-deficient 721.220 cells allowed the influence of this chaperone on peptide repertoire to be examined. Comparison of peptides bound to HLA-B*2705 expressed on tapasin-deficient and -proficient cells by mass spectrometry revealed an overall reduction in the recovery of B*2705-bound peptides isolated from tapasin-deficient cells despite similar yields of B27 heavy chain and beta(2)-microglobulin. This indicated that a proportion of suboptimal ligands were associated with B27, and they were lost during the purification process. Notwithstanding this failure to recover these suboptimal peptides, there was substantial overlap in the repertoire and biochemical properties of peptides recovered from B27 complexes derived from tapasin-positive and -negative cells. Although many peptides were preferentially or uniquely isolated from B*2705 in tapasin-positive cells, a number of species were preferentially recovered in the absence of tapasin, and some of these peptide ligands have been sequenced. In general, these ligands did not exhibit exceptional binding affinity, and we invoke an argument based on lumenal availability and affinity to explain their tapasin independence. The differential display of peptides in tapasin-negative and -positive cells was also apparent in the reactivity of peptide-sensitive alloreactive CTL raised against tapasin-positive and -negative targets, demonstrating the functional relevance of the biochemical observation of changes in peptide repertoire in the tapasin-deficient APC. Overall, the data reveal that tapasin quantitatively and qualitatively influences ligand selection by class I molecules.
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ItemT cell receptor recognition of a 'super-bulged' major histocompatibility complex class I-bound peptide(NATURE PUBLISHING GROUP, 2005-11)Unusually long major histocompatibility complex (MHC) class I-restricted epitopes are important in immunity, but their 'bulged' conformation represents a potential obstacle to alphabeta T cell receptor (TCR)-MHC class I docking. To elucidate how such recognition is achieved while still preserving MHC restriction, we have determined here the structure of a TCR in complex with HLA-B(*)3508 presenting a peptide 13 amino acids in length. This complex was atypical of TCR-peptide-MHC class I interactions, being dominated at the interface by peptide-mediated interactions. The TCR assumed two distinct orientations, swiveling on top of the centrally bulged, rigid peptide such that only limited contacts were made with MHC class I. Although the TCR-peptide recognition resembled an antibody-antigen interaction, the TCR-MHC class I contacts defined a minimal 'generic footprint' of MHC-restriction. Thus our findings simultaneously demonstrate the considerable adaptability of the TCR and the 'shape' of MHC restriction.
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ItemThe CDR3 regions of an immunodominant T cell receptor dictate the 'energetic landscape' of peptide-MHC recognition(NATURE PUBLISHING GROUP, 2005-02)The energetic bases of T cell recognition are unclear. Here, we studied the 'energetic landscape' of peptide-major histocompatibility complex (pMHC) recognition by an immunodominant alphabeta T cell receptor (TCR). We quantified and evaluated the effect of natural and systematic substitutions in the complementarity-determining region (CDR) loops on ligand binding in the context of the structural detail of each component of the immunodominant TCR-pMHC complex. The CDR1 and CDR2 loops contributed minimal energy through direct recognition of the antigen and instead had a chief function in stabilizing the ligated CDR3 loops. The underlying energetic basis for recognition lay in the CDR3 loops. Therefore the energetic burden of the CDR loops in the TCR-pMHC interaction is variable among TCRs, reflecting the inherent adaptability of the TCR in ligating different ligands.
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ItemStructural basis for a major histocompatibility complex class Ib-restricted T cell response(NATURE PUBLISHING GROUP, 2006-03)In contrast to antigen-specific immunity orchestrated by major histocompatibility complex (MHC) class Ia molecules, the ancestrally related nonclassical MHC class Ib molecules generally mediate innate immune responses. Here we have demonstrated the structural basis by which the MHC class Ib molecule HLA-E mediates an adaptive MHC-restricted cytotoxic T lymphocyte response to human cytomegalovirus. Highly constrained by host genetics, the response showed notable fine specificity for position 8 of the viral peptide, which is the sole discriminator of self versus nonself. Despite the evolutionary divergence of MHC class Ia and class Ib molecules, the structure of the T cell receptor-MHC class Ib complex was very similar to that of conventional T cell receptor-MHC class Ia complexes. These results emphasize the evolutionary 'ambiguity' of HLA-E, which not only interacts with innate immune receptors but also has the functional capacity to mediate virus-specific cytotoxic T lymphocyte responses during adaptive immunity.