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Biomedical subjects

E O Long

Publications and source records attributed to E O Long.

At least 55 records · Page 3Linked to original sources
PubMed1997

T cell response to myelin basic protein in the context of the multiple sclerosis-associated HLA-DR15 haplotype: peptide binding, immunodominance and effector functions of T cells.

In this study, we evaluated the role of the two functional HLA-DR heterodimers, DR2a (DR alpha paired with the beta chain encoded by DRB5*0101) and DR2b (DR alpha paired with the beta chain encoded by DRB1*1501), that are coexpressed in the multiple sclerosis (MS)-associated haplotype HLA-DR15 Dw2, in presenting myelin basic protein (MBP) peptides to MBP-specific T cell lines (TCL). Our results show that both HLA-DR molecules serve as restriction elements for HLA-DR15-restricted TCL. Slightly higher numbers of TCL use DR2a as restriction element, and the epitopes contained in the immunodominant C-terminal region (131-159) are uniquely restricted by DR2a. The immunodominant middle epitope (81-99) is recognized in the context of both DR2a and DR2b, but this specificity strongly dominates the DR2b-restricted T cell response. Overall, immunodominance in the MBP-specific T cell response correlated well with peptide binding to DR2a or DR2b, demonstrating that the affinity of MHC-peptide interactions is important for shaping the T cell response to this autoantigen. Furthermore, we show that binding of the middle MBP peptide to HLA-DR15 molecules prevents cleavage by cathepsin D, a protease abundantly found in endosomal processing compartments, and thus contributes to its immunodominance. Surprisingly, the restriction element employed by MBP-specific T cell clones influenced the effector function (i.e., cytotoxic activity) of T cells irrespective of their peptide fine specificity.

Alleles↗
PubMed1997

Natural killer cell receptors.

The specificity in the recognition of hematopoietic target cells by natural killer cells is primarily provided by inhibitory receptors and several such receptors have been identified in the past year. Surprisingly, the recognition of MHC class I molecules by inhibitory receptors on human natural killer cells involves two different types of receptors, one with Ig domains (killer cell inhibitory receptor), and another with C-type lectin domains (CD94-NKG2). Mouse natural killer cells recognize MHC class I molecules through the C-type lectin Ly49 receptors but also express a receptor, of unknown ligand specificity, that is related to the killer cell inhibitory receptor.

Animals↗
PubMed1997

Killer cell inhibitory receptors: diversity, specificity, and function.

NK cells selectively kill target cells that fail to express self-MHC class I molecules. This selective killing results from a balance between inhibitory NK receptors specific for MHC class I molecules and activating receptors that are still largely unknown. Isolation of molecular clones for the human killer cell inhibitory receptors (KIR) revealed that KIR consist of a family of molecules with Ig ectodomains and cytoplasmic tails of varying length. Soluble complexes of KIR and HLA-C molecules established that KIR recognizes and binds to its ligand as an autonomous receptor. A functional expression system in human NK clones demonstrated that a single KIR can provide both recognition of MHC class I and delivery of a dominant negative signal to the NK cell. Functional evidence has been obtained for a role of the tyrosine phosphatase SHP-1 in KIR-mediated inhibition. The presence of a conserved motif used to recruit and activate SHP-1 in the cytoplasmic tail of KIR and of the mouse Ly-49 inhibitory receptor (otherwise structurally unrelated to KIR) represents an interesting case of evolutionary convergence. Furthermore, the motif led to the identification of other receptors with inhibitory potential, including a type I Ig-like receptor shared by mouse mast cells and NK cells.

Amino Acid Sequence↗
PubMed1996

Presentation of a cytosolic antigen by major histocompatibility complex class II molecules requires a long-lived form of the antigen.

Class I and II molecules of the major histocompatibility complex present peptides to T cells. Class I molecules bind peptides that have been generated in the cytosol by proteasomes and delivered into the endoplasmic reticulum by the transporter associated with antigen presentation. In contrast, class II molecules are very efficient in the presentation of antigens that have been internalized and processed in endosomal/lysosomal compartments. In addition, class II molecules can present some cytosolic antigens by a TAP-independent pathway. To test whether this endogenous class II presentation pathway was linked to proteasome-mediated degradation of antigen in the cytosol, the N-end rule was utilized to produce two forms of the influenza virus matrix protein with different in vivo half-lives (10 min vs. 5 h) when expressed in human B cells. Whereas class I molecules presented both the short- and the long-lived matrix proteins, class II molecules presented exclusively the long-lived form of antigen. Thus, rapid degradation of matrix protein in the cytosol precluded its presentation by class II molecules. These data suggest that the turnover of long-lived cytosolic proteins, some of which is mediated by delivery into endosomal/ lysosomal compartments, provides a mechanism for immune surveillance by CD4+ T cells.

Antigen Presentation↗
PubMed1996

Peptide sequence requirements for the recognition of HLA-B*2705 by specific natural killer cells.

NK clones that were inhibited by target cell expression of HLA-B*2705 displayed peptide-specific recognition of HLA-B*2705. To evaluate the specificity of this recognition, synthetic versions of 14 endogenous ligands of HLA-B*2705 were tested for their ability to provide protection from NK-mediated lysis by binding to surface HLA-B*2705 molecules on RMA-S cells deficient in the transporter for Ag presentation. Several unrelated peptides inhibited lysis by the same NK clones. Despite similar capacities to stabilize HLA-B*2705 molecules on RMA-S cells, the 14 peptides differed widely in their abilities to provide protection. Single amino acid substitutions in both a protective and a nonprotective peptide revealed the importance of residues 7 and 8 in the peptide for recognition by NK clones, thus localizing the peptide influence to a polymorphic region of the alpha-helix of HLA class I molecules known to control discrimination among allelic variants of HLA-B and HLA-C by NK cells.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

A p70 killer cell inhibitory receptor specific for several HLA-B allotypes discriminates among peptides bound to HLA-B*2705.

Natural killer (NK) cells express a repertoire of killer cell inhibitory receptors (KIR) for major histocompatibility complex (MHC) class I molecules. KIR specificity for MHC class I can be broad, as in the case of a single p70 KIR that can recognize several HLA-B allotypes, including HLA-B*2705. On the other hand, recognition of MHC class I can also be highly specific, as in the case of NK clones that recognize HLA-B*2705 in a peptide-specific manner. Most NK cells express multiple KIR sequences. To determine whether the broad and specific types of HLA-B recognition by NK cells reflect the use of different receptors or a property of a single KIR we analyzed the recognition of HLA-B*2705 by the p70 KIR-11, known to recognize several HLA-B allotypes. Vaccinia virus-mediated expression of KIR-11 in NK clones resulted in inhibition by HLA-B*2705 molecules on wild type but not on target cells deficient in the transporter for antigen presentation (TAP). Two peptides (FRYNGLIHR and RRSKEITVR) loaded onto HLA-B*2705 molecules on TAP-deficient cells provided protection from lysis by NK cells expressing KIR-11 but three other B27-specific peptides did not. As the five peptides bound to HLA-B*2705 with similar stability, these data demonstrate that a single KIR specific for several HLA-B allotypes recognizes a subset of peptides bound to HLA-B*2705.

Clone Cells↗
PubMed1996

Direct binding of a soluble natural killer cell inhibitory receptor to a soluble human leukocyte antigen-Cw4 class I major histocompatibility complex molecule.

Natural killer (NK) cells expressing specific p58 NK receptors are inhibited from lysing target cells that express human leukocyte antigen (HLA)-C class I major histocompatibility complex molecules. To investigate the interaction between p58 NK receptors and HLA-Cw4, the extracellular domain of the p58 NK receptor specific for HLA-Cw4 was overexpressed in Escherichia coli and refolded from purified inclusion bodies. The refolded NK receptor is a monomer in solution. It interacts specifically with HLA-Cw4, blocking the binding of a p58-Ig fusion protein to HLA-Cw4-expressing cells, but does not block the binding of a p58-Ig fusion protein specific for HLA-Cw3 to HLA-Cw3-expressing cells. The bacterially expressed extracellular domain of HLA-Cw4 heavy chain and beta2-microglobulin were refolded in the presence of a HLA-Cw4-specific peptide. Direct binding between the soluble p58 NK receptor and the soluble HLA-Cw4-peptide complex was observed by native gel electrophoresis. Titration binding assays show that soluble monomeric receptor forms a 1:1 complex with HLA-Cw4, independent of the presence of Zn2+. The formation of complexes between soluble, recombinant molecules indicates that HLA-Cw4 is sufficient for specific ligation by the NK receptor and that neither glycoprotein requires carbohydrate for the interaction.

Amino Acid Sequence↗
PubMed1996

Mutations in human CD4 impair the functional interaction with different human and mouse class II isotypes and alleles.

The structure-function of the CD4-class II MHC interaction was investigated. Two functional assays were used to assess the responses of the 3DT52.5.8 murine T cell hybridoma expressing human CD4 (h-CD4) or murine CD4 (m-CD4). First, we determined the responses of the CD4+ and CD4-effector cells toward DAP-3 cells co-expressing the cognate alloantigen H-2Dd together with several human (DRw52b, DR4-Dw4, DR2A, and DPw2) and murine (I-Ab, I-Ak, IA alpha b I-A beta k and I-Ek) class II alleles and isotypes. We found that h-CD4 and m-CD4 strongly enhance the T cell response to H-2Dd, demonstrating that interspecies CD4/class II interactions occur efficiently. Furthermore, mutations in h-CD4 at positions 19, 89, and 165 markedly reduced the interaction with both human class II and mouse class II, indicating that the structural features of this cross-species interaction are strongly conserved. This was further supported by the finding that a h-CD4 deletion mutant (deletion F43-S49) interacted with both human and murine class II. Moreover, as 3DT cells express the responsive V beta element for the bacterial superantigen staphylococcal enterotoxin B, a co-receptor assay was conducted. DAP-3 cells expressing only class II molecules were used as APCs to present staphylococcal enterotoxin B to h-CD4+ and m-CD4+ T cells. h-CD4 and m-CD4 were able to enhance the T cell response to staphylococcal enterotoxin B, further demonstrating the conservation of the CD4-class II MHC interaction.

Alleles↗
PubMed1996

Recruitment of tyrosine phosphatase HCP by the killer cell inhibitor receptor.

Cytolysis of target cells by natural killer (NK) cells and by some cytotoxic T cells occurs unless prevented by inhibitory receptors that recognize MHC class I on target cells. Human NK cells express a p58 inhibitory receptor specific for HLA-C. We report association of the tyrosine phosphatase HCP with the p58 receptor in NK cells. HCP association was dependent on tyrosine phosphorylation of p58. Phosphotyrosyl peptides corresponding to the p58 tail bound and activated HCP in vitro. Furthermore, introduction of an inactive mutant HCP into an NK cell line prevented the p58-mediated inhibition of target cell lysis. These data imply that the inhibitory function of p58 is dependent on its tyrosine phosphorylation and on recruitment and activation of HCP.

Amino Acid Sequence↗
PubMed1996

Isoforms of the invariant chain regulate transport of MHC class II molecules to antigen processing compartments.

Newly synthesized class II molecules of the major histocompatibility complex must be transported to endosomal compartments where antigens are processed for presentation to class II-restricted T cells. The invariant chain (Ii), which assembles with newly synthesized class II alpha- and beta-chains in the endoplasmic reticulum, carries one or more targeting signals for transport to endosomal compartments where Ii dissociates from alpha beta Ii complexes. Here we show that the transport route of alpha beta Ii complexes is regulated selectively by two forms of Ii (p33 and p35) that are generated by the use of alternative translation initiation sites. Using a novel quantitative surface arrival assay based on labeling with [6-3H]-D-galactose combined with biochemical modification at the cell surface with neuraminidase, we demonstrate that newly synthesized alpha beta Ii molecules containing the Ii-p33 isoform can be detected on the cell surface shortly after passage through the Golgi apparatus/trans-Golgi network. A substantial amount of these alpha beta Ii complexes are targeted to early endosomes either directly from the trans-Golgi network or after internalization from the cell surface before their delivery to antigen processing compartments. The fraction of alpha beta Ii complexes containing the p35 isoform of Ii with a longer cytosolic domain was not detected at the cell surface as determined by iodination of intact cells and the lack of susceptibility to neuraminidase trimming on ice. However, treatment with neuraminidase at 37 degrees C did reveal that some of the alpha beta Ii-p35 complexes traversed early endosomes. These results demonstrate that a fraction of newly synthesized class II molecules arrive at the cell surface as alpha beta Ii complexes before delivery to antigen processing compartments and that class II alpha beta Ii complexes associated with the two isoforms of Ii are sorted to these compartments by different transport routes.

Antigen Presentation↗
PubMed1995

The Ig-related killer cell inhibitory receptor binds zinc and requires zinc for recognition of HLA-C on target cells.

Members of the Ig superfamily are predominantly receptors that mediate interactions between cells or provide signals to cells when binding specific ligands. Here we describe an Ig-related receptor that requires zinc for its function. Killer cell inhibitory receptors (KIR) belonging to the Ig superfamily mediate inhibition of NK cells upon recognition of HLA-C molecules on target cells. An abundance of histidine residues in the first extracellular domain of KIR, including the signature zinc binding motif HEXXH, suggested that this receptor may bind zinc. Two distinct KIR molecules that mediate recognition of HLA-Cw4 and -Cw8, respectively, bound specifically to zinc affinity columns. Furthermore, addition of the zinc chelator 1,10-phenanthroline during chromium release assays reversed the inhibition of killing by NK clones specific for HLA-Cw4 or HLA-Cw8, demonstrating that zinc is necessary for the inhibitory function of KIR. Such functionally relevant zinc binding has not been described for other members of the Ig superfamily and may represent a novel regulatory mechanism for Ag receptor-ligand interactions.

Amino Acid Sequence↗
PubMed1995

HLA-DR polymorphism affects the interaction with CD4.

Major histocompatibility complex (MHC) class II molecules are highly polymorphic and bind peptides for presentation to CD4+ T cells. Functional and adhesion assays have shown that CD4 interacts with MHC class II molecules, leading to enhanced responses of CD4+ T cells after the activation of the CD4-associated tyrosine kinase p56lck. We have addressed the possible contribution of allelic polymorphism in the interaction between CD4 and MHC class II molecules. Using mouse DAP-3-transfected cells expressing different isotypes and allelic forms of the HLA-DR molecule, we have shown in a functional assay that a hierarchy exists in the ability of class II molecules to interact with CD4. Also, the study of DR4 subtypes minimized the potential contribution of polymorphic residues of the peptide-binding groove in the interaction with CD4. Chimeras between the DR4 or DR1 molecules, which interact efficiently with CD4, and DRw53, which interacts poorly, allowed the mapping of polymorphic residues between positions beta 180 and 189 that can exert a dramatic influence on the interaction with CD4.

Alleles↗
PubMed1995

Antigen presentation mediated by recycling of surface HLA-DR molecules.

Class II histocompatibility molecules associate with peptides derived from antigens that are processed in endocytic compartments. Antigen presentation to class II-restricted T cells generally requires newly synthesized class II molecules, associated invariant chain, and HLA-DM. Exceptions to these rules have been reported, but without description of an underlying mechanism. Here we show that presentation of immunodominant epitopes in the haemagglutinin protein of influenza virus and in myelin basic protein correlates with recycling of surface HLA-DR molecules. Truncation of either one of the alpha or beta cytoplasmic tails virtually eliminated internalization of HLA-DR molecules and presentation of haemagglutinin from inactive virus particles. In contrast, the invariant chain-dependent presentation of matrix antigen from the same virus particles was unaffected by these truncations. Thus HLA-DR cytoplasmic tails are not required for the conventional presentation pathway, but jointly contribute a signal for an alternative pathway involving internalization of HLA-DR molecules.

Amino Acid Sequence↗
PubMed1995

Peptide specificity in the recognition of MHC class I by natural killer cell clones.

Recognition by natural killer (NK) cells of major histocompatibility complex (MHC) class I molecules on target cells inhibits NK-mediated lysis. Here, inhibition of NK clones by HLA-B*2705 molecules mutated at single amino acids in the peptide binding site varied among HLA-B*2705-specific NK clones. In addition, a subset of such NK clones was inhibited by only one of several self peptides loaded onto HLA-B*2705 molecules expressed in peptide transporter-deficient cells, showing that recognition was peptide-specific. These data demonstrate that specific self peptides, complexed with MHC class I, provide protection from NK-mediated lysis.

Amino Acid Sequence↗
PubMed1995

Molecular clones of the p58 NK cell receptor reveal immunoglobulin-related molecules with diversity in both the extra- and intracellular domains.

Recognition of major histocompatibility class I molecules on target cells by natural killer (NK) cells confers selective protection from NK-mediated lysis. Cross-linking of the p58 NK receptor, involved in the recognition of HLA-C alleles, delivers a negative signal that prevents target cell lysis. Molecular cloning of the p58 NK receptor reported here revealed a new member of the immunoglobulin superfamily. Five distinct p58 receptors, with sequence diversity in the immunoglobulin-related domains, were identified in a single individual. All NK clones tested expressed at least one p58 member. Three different types of transmembrane and cytoplasmic domains exist, even among receptors with closely related extracellular domains. These data revealed a repertoire of NK cells with clonally distributed p58 receptors exhibiting diversity in both extracellular and intracellular domains.

Antibodies, Monoclonal↗
PubMed1994

T cell recognition of an HLA-A2-restricted epitope derived from a cleaved signal sequence.

An alternative pathway for class I-restricted antigen presentation has been suggested on the basis of peptides bound to HLA-A2 molecules in cells lacking the transporter for antigen presentation (TAP). Most of these peptides were derived from signal sequences for translocation into the endoplasmic reticulum (ER). However, it is not known whether these peptides can be presented to T cells. The hydrophobic nature of an HLA-A2-restricted T cell epitope (M1 58-66) was exploited to test whether it could be presented to T cells when derived from a signal sequence. Replacing the signal sequence of the influenza virus hemagglutinin molecule H3 with an artificial sequence containing that HLA-A2-restricted T cell epitope resulted in efficient translocation of H3 molecules into the ER and transport to the cell surface. This signal sequence-derived epitope was presented to HLA-A2-restricted T cells. Involvement of cytosolic processing for this presentation is very unlikely, because (a) presentation occurred in cells lacking TAP; (b) expression of H3 molecules with the artificial signal sequence did not produce a detectable cytosolic form of H3; and (c) presentation of the same epitope expressed in cytosolic forms of antigen required TAP. Thus, a peptide derived from a signal sequence cleaved in the ER can provide an epitope for HLA-A2-restricted T cell recognition.

ATP Binding Cassette Transporter, Subfamily B, Mem↗