Publications by authors named "Lisa F Boyd"

20 Publications

  • Page 1 of 1

A local regulatory T cell feedback circuit maintains immune homeostasis by pruning self-activated T cells.

Cell 2021 Jul 21;184(15):3981-3997.e22. Epub 2021 Jun 21.

Lymphocyte Biology Section, Laboratory of Immune System Biology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892-1892, USA. Electronic address:

A fraction of mature T cells can be activated by peripheral self-antigens, potentially eliciting host autoimmunity. We investigated homeostatic control of self-activated T cells within unperturbed tissue environments by combining high-resolution multiplexed and volumetric imaging with computational modeling. In lymph nodes, self-activated T cells produced interleukin (IL)-2, which enhanced local regulatory T cell (Treg) proliferation and inhibitory functionality. The resulting micro-domains reciprocally constrained inputs required for damaging effector responses, including CD28 co-stimulation and IL-2 signaling, constituting a negative feedback circuit. Due to these local constraints, self-activated T cells underwent transient clonal expansion, followed by rapid death ("pruning"). Computational simulations and experimental manipulations revealed the feedback machinery's quantitative limits: modest reductions in Treg micro-domain density or functionality produced non-linear breakdowns in control, enabling self-activated T cells to subvert pruning. This fine-tuned, paracrine feedback process not only enforces immune homeostasis but also establishes a sharp boundary between autoimmune and host-protective T cell responses.
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http://dx.doi.org/10.1016/j.cell.2021.05.028DOI Listing
July 2021

Alterations in the HLA-B*57:01 Immunopeptidome by Flucloxacillin and Immunogenicity of Drug-Haptenated Peptides.

Front Immunol 2020 9;11:629399. Epub 2021 Feb 9.

Laboratory of Immunology, Office of Biotechnology Products, Center for Drugs Evaluation and Research, Food and Drug Administration, Silver Spring, MD, United States.

Neoantigen formation due to the interaction of drug molecules with human leukocyte antigen (HLA)-peptide complexes can lead to severe hypersensitivity reactions. Flucloxacillin (FLX), a β-lactam antibiotic for narrow-spectrum gram-positive bacterial infections, has been associated with severe immune-mediated drug-induced liver injury caused by an influx of T-lymphocytes targeting liver cells potentially recognizing drug-haptenated peptides in the context of HLA-B*57:01. To identify immunopeptidome changes that could lead to drug-driven immunogenicity, we used mass spectrometry to characterize the proteome and immunopeptidome of B-lymphoblastoid cells solely expressing HLA-B*57:01 as MHC-I molecules. Selected drug-conjugated peptides identified in these cells were synthesized and tested for their immunogenicity in HLA-B*57:01-transgenic mice. T cell responses were evaluated by immune assays. The immunopeptidome of FLX-treated cells was more diverse than that of untreated cells, enriched with peptides containing carboxy-terminal tryptophan and FLX-haptenated lysine residues on peptides. Selected FLX-modified peptides with drug on P4 and P6 induced drug-specific CD8 T cells . FLX was also found directly linked to the HLA K146 that could interfere with KIR-3DL or peptide interactions. These studies identify a novel effect of antibiotics to alter anchor residue frequencies in HLA-presented peptides which may impact drug-induced inflammation. Covalent FLX-modified lysines on peptides mapped drug-specific immunogenicity primarily at P4 and P6 suggesting these peptide sites as drivers of off-target adverse reactions mediated by FLX. FLX modifications on HLA-B*57:01-exposed lysines may also impact interactions with KIR or TCR and subsequent NK and T cell function.
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http://dx.doi.org/10.3389/fimmu.2020.629399DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7900192PMC
June 2021

Synthetic nanobody-SARS-CoV-2 receptor-binding domain structures identify distinct epitopes.

bioRxiv 2021 Jan 27. Epub 2021 Jan 27.

The worldwide spread of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) demands unprecedented attention. We report four X-ray crystal structures of three synthetic nanobodies (sybodies) (Sb16, Sb45 and Sb68) bind to the receptor-binding domain (RBD) of SARS-CoV-2: binary complexes of Sb16-RBD and Sb45-RBD; a ternary complex of Sb45-RBD-Sb68; and Sb16 unliganded. Sb16 and Sb45 bind the RBD at the ACE2 interface, positioning their CDR2 and CDR3 loops diametrically. Sb16 reveals a large CDR2 shift when binding the RBD. Sb68 interacts peripherally at the ACE2 interface; steric clashes with glycans explain its mechanism of viral neutralization. Superposing these structures onto trimeric spike (S) protein models indicates these sybodies bind conformations of the mature S protein differently, which may aid therapeutic design.

One Sentence Summary: X-ray structures of synthetic nanobodies complexed with the receptor-binding domain of the spike protein of SARS-CoV-2 reveal details of CDR loop interactions in recognition of distinct epitopic sites.
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http://dx.doi.org/10.1101/2021.01.27.428466DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7852268PMC
January 2021

Cutting Edge: Inhibition of the Interaction of NK Inhibitory Receptors with MHC Class I Augments Antiviral and Antitumor Immunity.

J Immunol 2020 08 29;205(3):567-572. Epub 2020 Jun 29.

Cellular Immunology Section, Laboratory of Immune System Biology, National Institute of Allergy and Infectious Diseases, Bethesda, MD 20892;

NK cells recognize MHC class I (MHC-I) Ags via stochastically expressed MHC-I-specific inhibitory receptors that prevent NK cell activation via cytoplasmic ITIM. We have identified a pan anti-MHC-I mAb that blocks NK cell inhibitory receptor binding at a site distinct from the TCR binding site. Treatment of unmanipulated mice with this mAb disrupted immune homeostasis, markedly activated NK and memory phenotype T cells, enhanced immune responses against transplanted tumors, and augmented responses to acute and chronic viral infection. mAbs of this type represent novel checkpoint inhibitors in tumor immunity, potent tools for the eradication of chronic infection, and may function as adjuvants for the augmentation of the immune response to weak vaccines.
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http://dx.doi.org/10.4049/jimmunol.2000412DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7369225PMC
August 2020

MHC-restricted Ag85B-specific CD8 T cells are enhanced by recombinant BCG prime and DNA boost immunization in mice.

Eur J Immunol 2019 09 19;49(9):1399-1414. Epub 2019 Jun 19.

Division of Microbiology, Department of Pathology and Microbiology, Nihon University School of Medicine, Nihon University, Tokyo, Japan.

Despite efforts to develop effective treatments and vaccines, Mycobacterium tuberculosis (Mtb), particularly pulmonary Mtb, continues to provide major health challenges worldwide. To improve immunization against the persistent health challenge of Mtb infection, we have studied the CD8 T cell response to Bacillus Calmette-Guérin (BCG) and recombinant BCG (rBCG) in mice. Here, we generated CD8 T cells with an rBCG-based vaccine encoding the Ag85B protein of M. kansasii, termed rBCG-Mkan85B, followed by boosting with plasmid DNA expressing the Ag85B gene (DNA-Mkan85B). We identified two MHC-I (H2-K )-restricted epitopes that induce cross-reactive responses to Mtb and other related mycobacteria in both BALB/c (H2 ) and CB6F1 (H2 ) mice. The H2-K -restricted peptide epitopes elicited polyfunctional CD8 T cell responses that were also highly cross-reactive with those of other proteins of the Ag85 complex. Tetramer staining indicated that the two H2-K -restricted epitopes elicit distinct CD8 T cell populations, a result explained by the X-ray structure of the two peptide/H2-K complexes. These results suggest that rBCG-Mkan85B vector-based immunization and DNA-Mkan85B boost may enhance CD8 T cell response to Mtb, and might help to overcome the limited effectiveness of the current BCG in eliciting tuberculosis immunity.
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http://dx.doi.org/10.1002/eji.201847988DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6722017PMC
September 2019

The Role of Molecular Flexibility in Antigen Presentation and T Cell Receptor-Mediated Signaling.

Front Immunol 2018 17;9:1657. Epub 2018 Jul 17.

Molecular Biology Section, Laboratory of Immune System Biology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, United States.

Antigen presentation is a cellular process that involves a number of steps, beginning with the production of peptides by proteolysis or aberrant synthesis and the delivery of peptides to cellular compartments where they are loaded on MHC class I (MHC-I) or MHC class II (MHC-II) molecules. The selective loading and editing of high-affinity immunodominant antigens is orchestrated by molecular chaperones: tapasin/TAP-binding protein, related for MHC-I and HLA-DM for MHC-II. Once peptide/MHC (pMHC) complexes are assembled, following various steps of quality control, they are delivered to the cell surface, where they are available for identification by αβ receptors on CD8 or CD4 T lymphocytes. In addition, recognition of cell surface peptide/MHC-I complexes by natural killer cell receptors plays a regulatory role in some aspects of the innate immune response. Many of the components of the pathways of antigen processing and presentation and of T cell receptor (TCR)-mediated signaling have been studied extensively by biochemical, genetic, immunological, and structural approaches over the past several decades. Until recently, however, dynamic aspects of the interactions of peptide with MHC, MHC with molecular chaperones, or of pMHC with TCR have been difficult to address experimentally, although computational approaches such as molecular dynamics (MD) simulations have been illuminating. Studies exploiting X-ray crystallography, cryo-electron microscopy, and multidimensional nuclear magnetic resonance (NMR) spectroscopy are beginning to reveal the importance of molecular flexibility as it pertains to peptide loading onto MHC molecules, the interactions between pMHC and TCR, and subsequent TCR-mediated signals. In addition, recent structural and dynamic insights into how molecular chaperones define peptide selection and fine-tune the MHC displayed antigen repertoire are discussed. Here, we offer a review of current knowledge that highlights experimental data obtained by X-ray crystallography and multidimensional NMR methodologies. Collectively, these findings strongly support a multifaceted role for protein plasticity and conformational dynamics throughout the antigen processing and presentation pathway in dictating antigen selection and recognition.
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http://dx.doi.org/10.3389/fimmu.2018.01657DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6056622PMC
July 2018

A transgenic mouse model for HLA-B*57:01-linked abacavir drug tolerance and reactivity.

J Clin Invest 2018 07 21;128(7):2819-2832. Epub 2018 May 21.

Laboratory of Immunology, Division of Biotechnology Review and Research III (DBRR III), Office of Biotechnology Products (OBP), Office of Pharmaceutical Quality (OPQ), Center for Drug Evaluation and Research (CDER), FDA, Silver Spring, Maryland, USA.

Adverse drug reactions (ADRs) are a major obstacle to drug development, and some of these, including hypersensitivity reactions to the HIV reverse transcriptase inhibitor abacavir (ABC), are associated with HLA alleles, particularly HLA-B*57:01. However, not all HLA-B*57:01+ patients develop ADRs, suggesting that in addition to the HLA genetic risk, other factors may influence the outcome of the response to the drug. To study HLA-linked ADRs in vivo, we generated HLA-B*57:01-Tg mice and show that, although ABC activated Tg mouse CD8+ T cells in vitro in a HLA-B*57:01-dependent manner, the drug was tolerated in vivo. In immunocompetent Tg animals, ABC induced CD8+ T cells with an anergy-like phenotype that did not lead to ADRs. In contrast, in vivo depletion of CD4+ T cells prior to ABC administration enhanced DC maturation to induce systemic ABC-reactive CD8+ T cells with an effector-like and skin-homing phenotype along with CD8+ infiltration and inflammation in drug-sensitized skin. B7 costimulatory molecule blockade prevented CD8+ T cell activation. These Tg mice provide a model for ABC tolerance and for the generation of HLA-B*57:01-restricted, ABC-reactive CD8+ T cells dependent on both HLA genetic risk and immunoregulatory host factors.
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http://dx.doi.org/10.1172/JCI99321DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6025983PMC
July 2018

Effects of Cross-Presentation, Antigen Processing, and Peptide Binding in HIV Evasion of T Cell Immunity.

J Immunol 2018 03 26;200(5):1853-1864. Epub 2018 Jan 26.

Vaccine Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892.

Unlike cytosolic processing and presentation of viral Ags by virus-infected cells, Ags first expressed in infected nonprofessional APCs, such as CD4 T cells in the case of HIV, are taken up by dendritic cells and cross-presented. This generally requires entry through the endocytic pathway, where endosomal proteases have first access for processing. Thus, understanding virus escape during cross-presentation requires an understanding of resistance to endosomal proteases, such as cathepsin S (CatS). We have modified HIV-1 gp120 by mutating a key CatS cleavage site (ThrThr) in the V3 loop of the immunodominant epitope IGPGRAFY to IGPGRAFY to prevent digestion. We found this mutation to facilitate cross-presentation and provide evidence from MHC binding and X-ray crystallographic structural studies that this results from preservation of the epitope rather than an increased epitope affinity for the MHC class I molecule. In contrast, when the protein is expressed by a vaccinia virus in the cytosol, the wild-type protein is immunogenic without this mutation. These proof-of-concept results show that a virus like HIV, infecting predominantly nonprofessional presenting cells, can escape T cell recognition by incorporating a CatS cleavage site that leads to destruction of an immunodominant epitope when the Ag undergoes endosomal cross-presentation.
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http://dx.doi.org/10.4049/jimmunol.1701523DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5890335PMC
March 2018

Crystal structure of a TAPBPR-MHC I complex reveals the mechanism of peptide editing in antigen presentation.

Science 2017 11 12;358(6366):1064-1068. Epub 2017 Oct 12.

Molecular Biology Section, Laboratory of Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.

Central to CD8 T cell-mediated immunity is the recognition of peptide-major histocompatibility complex class I (p-MHC I) proteins displayed by antigen-presenting cells. Chaperone-mediated loading of high-affinity peptides onto MHC I is a key step in the MHC I antigen presentation pathway. However, the structure of MHC I with a chaperone that facilitates peptide loading has not been determined. We report the crystal structure of MHC I in complex with the peptide editor TAPBPR (TAP-binding protein-related), a tapasin homolog. TAPBPR remodels the peptide-binding groove of MHC I, resulting in the release of low-affinity peptide. Changes include groove relaxation, modifications of key binding pockets, and domain adjustments. This structure captures a peptide-receptive state of MHC I and provides insights into the mechanism of peptide editing by TAPBPR and, by analogy, tapasin.
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http://dx.doi.org/10.1126/science.aao5154DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6320693PMC
November 2017

An allosteric site in the T-cell receptor Cβ domain plays a critical signalling role.

Nat Commun 2017 05 16;8:15260. Epub 2017 May 16.

Department of Chemistry and Biochemistry, University of California Santa Cruz, Santa Cruz, California 95064, USA.

The molecular mechanism through which the interaction of a clonotypic αβ T-cell receptor (TCR) with a peptide-loaded major histocompatibility complex (p/MHC) leads to T-cell activation is not yet fully understood. Here we exploit a high-affinity TCR (B4.2.3) to examine the structural changes that accompany binding to its p/MHC ligand (P18-I10/H2-D). In addition to conformational changes in complementarity-determining regions (CDRs) of the TCR seen in comparison of unliganded and bound X-ray structures, NMR characterization of the TCR β-chain dynamics reveals significant chemical shift effects in sites removed from the MHC-binding site. Remodelling of electrostatic interactions near the Cβ H3 helix at the membrane-proximal face of the TCR, a region implicated in interactions with the CD3 co-receptor, suggests a possible role for an allosteric mechanism in TCR signalling. The contribution of these TCR residues to signal transduction is supported by mutagenesis and T-cell functional assays.
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http://dx.doi.org/10.1038/ncomms15260DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5440810PMC
May 2017

Measuring Protein Interactions by Optical Biosensors.

Curr Protoc Protein Sci 2017 04 3;88:20.2.1-20.2.25. Epub 2017 Apr 3.

National Institutes of Health, Bethesda, Maryland.

This unit gives an introduction to the basic techniques of optical biosensing for measuring equilibrium and kinetics of reversible protein interactions. Emphasis is placed on description of robust approaches that will provide reliable results with few assumptions. How to avoid the most commonly encountered problems and artifacts is also discussed. © 2017 by John Wiley & Sons, Inc.
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http://dx.doi.org/10.1002/cpps.31DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5776739PMC
April 2017

Interaction of TAPBPR, a tapasin homolog, with MHC-I molecules promotes peptide editing.

Proc Natl Acad Sci U S A 2016 Feb 11;113(8):E1006-15. Epub 2016 Feb 11.

Molecular Biology Section, Laboratory of Immunology, National Institute of Allergy and Infectious Diseases/National Institutes of Health, Bethesda, MD 20892;

Peptide loading of major histocompatibility complex class I (MHC-I) molecules is central to antigen presentation, self-tolerance, and CD8(+) T-cell activation. TAP binding protein, related (TAPBPR), a widely expressed tapasin homolog, is not part of the classical MHC-I peptide-loading complex (PLC). Using recombinant MHC-I molecules, we show that TAPBPR binds HLA-A*02:01 and several other MHC-I molecules that are either peptide-free or loaded with low-affinity peptides. Fluorescence polarization experiments establish that TAPBPR augments peptide binding by MHC-I. The TAPBPR/MHC-I interaction is reversed by specific peptides, related to their affinity. Mutational and small-angle X-ray scattering (SAXS) studies confirm the structural similarities of TAPBPR with tapasin. These results support a role of TAPBPR in stabilizing peptide-receptive conformation(s) of MHC-I, permitting peptide editing.
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http://dx.doi.org/10.1073/pnas.1519894113DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4776512PMC
February 2016

A Novel MHC-I Surface Targeted for Binding by the MCMV m06 Immunoevasin Revealed by Solution NMR.

J Biol Chem 2015 Nov 13;290(48):28857-68. Epub 2015 Oct 13.

the Molecular Biology Section, Laboratory of Immunology, NIAID, National Institutes of Health, Bethesda, Maryland 20892

As part of its strategy to evade detection by the host immune system, murine cytomegalovirus (MCMV) encodes three proteins that modulate cell surface expression of major histocompatibility complex class I (MHC-I) molecules: the MHC-I homolog m152/gp40 as well as the m02-m16 family members m04/gp34 and m06/gp48. Previous studies of the m04 protein revealed a divergent Ig-like fold that is unique to immunoevasins of the m02-m16 family. Here, we engineer and characterize recombinant m06 and investigate its interactions with full-length and truncated forms of the MHC-I molecule H2-L(d) by several techniques. Furthermore, we employ solution NMR to map the interaction footprint of the m06 protein on MHC-I, taking advantage of a truncated H2-L(d), "mini-H2-L(d)," consisting of only the α1α2 platform domain. Mini-H2-L(d) refolded in vitro with a high affinity peptide yields a molecule that shows outstanding NMR spectral features, permitting complete backbone assignments. These NMR-based studies reveal that m06 binds tightly to a discrete site located under the peptide-binding platform that partially overlaps with the β2-microglobulin interface on the MHC-I heavy chain, consistent with in vitro binding experiments showing significantly reduced complex formation between m06 and β2-microglobulin-associated MHC-I. Moreover, we carry out NMR relaxation experiments to characterize the picosecond-nanosecond dynamics of the free mini-H2-L(d) MHC-I molecule, revealing that the site of interaction is highly ordered. This study provides insight into the mechanism of the interaction of m06 with MHC-I, suggesting a structural manipulation of the target MHC-I molecule at an early stage of the peptide-loading pathway.
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http://dx.doi.org/10.1074/jbc.M115.689661DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4661401PMC
November 2015

The structure of mouse cytomegalovirus m04 protein obtained from sparse NMR data reveals a conserved fold of the m02-m06 viral immune modulator family.

Structure 2014 Sep 7;22(9):1263-1273. Epub 2014 Aug 7.

Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD 20892, USA. Electronic address:

Immunoevasins are key proteins used by viruses to subvert host immune responses. Determining their high-resolution structures is key to understanding virus-host interactions toward the design of vaccines and other antiviral therapies. Mouse cytomegalovirus encodes a unique set of immunoevasins, the m02-m06 family, that modulates major histocompatibility complex class I (MHC-I) antigen presentation to CD8+ T cells and natural killer cells. Notwithstanding the large number of genetic and functional studies, the structural biology of immunoevasins remains incompletely understood, largely because of crystallization bottlenecks. Here we implement a technology using sparse nuclear magnetic resonance data and integrative Rosetta modeling to determine the structure of the m04/gp34 immunoevasin extracellular domain. The structure reveals a β fold that is representative of the m02-m06 family of viral proteins, several of which are known to bind MHC-I molecules and interfere with antigen presentation, suggesting its role as a diversified immune regulation module.
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http://dx.doi.org/10.1016/j.str.2014.05.018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4156524PMC
September 2014

A structural and molecular dynamics approach to understanding the peptide-receptive transition state of MHC-I molecules.

Mol Immunol 2013 Sep 28;55(2):123-5. Epub 2012 Nov 28.

Molecular Biology Section, Laboratory of Immunology, National Institute of Allergy and Infectious Diseases NIAID, NIH, Bethesda, MD 20892-1892, USA.

The mature conformation of major histocompatibility complex class I (MHC-I) proteins depends on the presence of bound peptides, permitting recognition at the cell surface by CD8(+) T lymphocytes. Newly synthesized MHC-I molecules in the endoplasmic reticulum are maintained in a peptide-receptive (PR) transition state by several chaperones until they are released concomitant with the loading of peptides. By determining the crystallographic structure of a region of an MHC-I molecule that is recognized by a unique monoclonal antibody and comparing this with docking and molecular dynamics simulations with the whole molecule, we demonstrate the movement of a hinged unit supporting the part of the binding groove that interacts with the amino terminal residues of the bound peptide. This unit contains a conserved 310 helix that flips from an exposed "open" position in the PR form to a "closed" position in the peptide-loaded (PL) mature molecule. These analyses indicate how this segment of the MHC-I molecule moves to help establish the A and B pockets critical for tight peptide binding and the stable structure required for antigen presentation and T cell recognition at the cell surface.
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http://dx.doi.org/10.1016/j.molimm.2012.10.021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3632263PMC
September 2013

Structural basis of mouse cytomegalovirus m152/gp40 interaction with RAE1γ reveals a paradigm for MHC/MHC interaction in immune evasion.

Proc Natl Acad Sci U S A 2012 Dec 19;109(51):E3578-87. Epub 2012 Nov 19.

Molecular Biology Section, Laboratory of Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.

Natural killer (NK) cells are activated by engagement of the NKG2D receptor with ligands on target cells stressed by infection or tumorigenesis. Several human and rodent cytomegalovirus (CMV) immunoevasins down-regulate surface expression of NKG2D ligands. The mouse CMV MHC class I (MHC-I)-like m152/gp40 glycoprotein down-regulates retinoic acid early inducible-1 (RAE1) NKG2D ligands as well as host MHC-I. Here we describe the crystal structure of an m152/RAE1γ complex and confirm the intermolecular contacts by mutagenesis. m152 interacts in a pincer-like manner with two sites on the α1 and α2 helices of RAE1 reminiscent of the NKG2D interaction with RAE1. This structure of an MHC-I-like immunoevasin/MHC-I-like ligand complex explains the binding specificity of m152 for RAE1 and allows modeling of the interaction of m152 with classical MHC-I and of related viral immunoevasins.
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http://dx.doi.org/10.1073/pnas.1214088109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3529088PMC
December 2012

The peptide-receptive transition state of MHC class I molecules: insight from structure and molecular dynamics.

J Immunol 2012 Aug 29;189(3):1391-9. Epub 2012 Jun 29.

Molecular Biology Section, Laboratory of Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.

MHC class I (MHC-I) proteins of the adaptive immune system require antigenic peptides for maintenance of mature conformation and immune function via specific recognition by MHC-I-restricted CD8(+) T lymphocytes. New MHC-I molecules in the endoplasmic reticulum are held by chaperones in a peptide-receptive (PR) transition state pending release by tightly binding peptides. In this study, we show, by crystallographic, docking, and molecular dynamics methods, dramatic movement of a hinged unit containing a conserved 3(10) helix that flips from an exposed "open" position in the PR transition state to a "closed" position with buried hydrophobic side chains in the peptide-loaded mature molecule. Crystallography of hinged unit residues 46-53 of murine H-2L(d) MHC-I H chain, complexed with mAb 64-3-7, demonstrates solvent exposure of these residues in the PR conformation. Docking and molecular dynamics predict how this segment moves to help form the A and B pockets crucial for the tight peptide binding needed for stability of the mature peptide-loaded conformation, chaperone dissociation, and Ag presentation.
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http://dx.doi.org/10.4049/jimmunol.1200831DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3422668PMC
August 2012

Availability of autoantigenic epitopes controls phenotype, severity, and penetrance in TCR Tg autoimmune gastritis.

Eur J Immunol 2008 Dec;38(12):3339-53

Department of Biotechnology Engineering, Ort Braude College, Karmiel, Israel.

We examined TCR:MHC/peptide interactions and in vivo epitope availability to explore the Th1- or Th2-like phenotype of autoimmune disease in two TCR Tg mouse models of autoimmune gastritis (AIG). The TCR of strains A23 and A51 recognize distinct IA(d)-restricted peptides from the gastric parietal cell H/K-ATPase. Both peptides form extremely stable MHC/peptide (MHC/p) complexes. All A23 animals develop a Th1-like aggressive, inflammatory AIG early in life, while A51 mice develop indolent Th2-like AIG at 6-8 wk with incomplete penetrance. A51 T cells were more sensitive than A23 to low doses of soluble antigen and to MHC/p complexes. Staining with IA(d)/peptide tetramers was only detectable on previously activated T cells from A51. Thus, despite inducing a milder AIG, the A51 TCR displays a higher avidity for its cognate IA(d)/peptide. Nonetheless, in vivo proliferation of adoptively transferred A51 CFSE-labeled T cells in the gastric lymph node was relatively poor compared with A23 T cells. Also, DC from WT gastric lymph node, presenting processed antigen available in vivo, stimulated proliferation of A23 T cells better than A51. Thus, the autoimmune potential of these TCR in their respective Tg lines is strongly influenced by the availability of the peptide epitope, rather than by differential avidity for their respective MHC/p complexes.
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http://dx.doi.org/10.1002/eji.200838584DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2712832PMC
December 2008

Measuring protein interactions by optical biosensors.

Curr Protoc Cell Biol 2004 May;Chapter 17:Unit 17.6

National Institutes of Health, Bethesda, Maryland, USA.

In recent years, optical evanescent wave biosensors have been used to characterize protein-protein interactions, including determination of equilibrium binding constants and bimolecular rate constants. This surface binding technique can provide information about chemical on-rate constants, the lifetimes of complexes formed, and the time course of the signal. This unit provides a thorough, well-illustrated discussion of the principles of optical biosensors, experimental design, ligand immobilization, experimental data analysis, and common obstacles and possible solutions.
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http://dx.doi.org/10.1002/0471143030.cb1706s22DOI Listing
May 2004

Role of alpha3 domain of class I MHC molecules in the activation of high- and low-avidity CD8+ CTLs.

Int Immunol 2007 Dec 1;19(12):1413-20. Epub 2007 Nov 1.

Molecular Immunogenetics and Vaccine Research Section, Vaccine Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.

CD8 can serve as a co-receptor or accessory molecule on the surface of CTL. As a co-receptor, CD8 can bind to the alpha3 domain of the same MHC class I molecules as the TCR to facilitate TCR signaling. To evaluate the role of the MHC class I molecule alpha3 domain in the activation of CD8(+) CTL, we have produced a soluble 227 mutant of H-2D(d), with a point mutation in the alpha3 domain (Glu227 --> Lys). 227 mutant class I-peptide complexes were not able to effectively activate H-2D(d)-restricted CD8 T cells in vitro, as measured by IFN-gamma production by an epitope-specific CD8(+) CTL line. However, the 227 mutant class I-peptide complexes in the presence of another MHC class I molecule (H-2K(b)) (that cannot present the peptide) with a normal alpha3 domain can induce the activation of CD8(+) CTL. Therefore, in order to activate CD8(+) CTL, the alpha3 domain of MHC class I does not have to be located on the same molecule with the alpha1 and alpha2 domains of MHC class I. A low-avidity CD8(+) CTL line was significantly less sensitive to stimulation by the 227 mutant class I-peptide complexes in the presence of the H-2K(b) molecule. Thus, low-avidity CTL may not be able to take advantage of the interaction between CD8 and the alpha3 domain of non-presenting class I MHC molecules, perhaps because of a shorter dwell time for the TCR-MHC interaction.
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http://dx.doi.org/10.1093/intimm/dxm111DOI Listing
December 2007
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