Eric Long | National Institutes of Health (original) (raw)

Papers by Eric Long

Research paper thumbnail of Interleukin-2 activity can be fine tuned with engineered receptor signaling clamps

Immunity, Jan 19, 2015

Interleukin-2 (IL-2) regulates lymphocyte function by signaling through heterodimerization of the... more Interleukin-2 (IL-2) regulates lymphocyte function by signaling through heterodimerization of the IL-2Rβ and γc receptor subunits. IL-2 is of considerable therapeutic interest, but harnessing its actions in a controllable manner remains a challenge. Previously, we have engineered an IL-2 "superkine" with enhanced affinity for IL-2Rβ. Here, we describe next-generation IL-2 variants that function as "receptor signaling…

Research paper thumbnail of Natural killer cell receptors

The specificity in the recognition of hematopoietic target cells by natural killer cells is prima... more 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.

Research paper thumbnail of Mitochondrial fusion proteins in revascularized hibernating hearts

Journal of Surgical Research, 2015

Hibernating myocardium is characterized by viable yet dysfunctional myocardium secondary to chron... more Hibernating myocardium is characterized by viable yet dysfunctional myocardium secondary to chronic ischemia, with studies demonstrating incomplete early recovery after coronary artery bypass graft (CABG). We tested whether mitochondrial fusion proteins, an indicator of mitochondrial biogenesis, are increased in hibernating myocardium post-CABG. A constrictor was placed on the left anterior descending (LAD) artery of nine pigs. Four of these pigs additionally underwent CABG 12 wk later with a left internal mammary artery graft to the LAD distal to the constrictor. Five pigs had a constrictor placed but did not undergo CABG (Hib). Five pigs did not have a constrictor placed (control). Computerized tomography angiography was used to confirm stenosis at the site of constrictor placement and patency of left internal mammary artery grafts. Regional blood flows were determined at baseline and during 40 μg/kg/min dobutamine infusion. Mitochondrial proteins were quantified by Western blot. Blood flow in the LAD region after CABG was lower than remote regions during dobutamine infusion (2.54 ± 0.24 versus 3.46 ± 0.33 mL/min/g; P < 0.05). Electron transport chain proteins were ∼70% lower in Hib compared with those in control and failed to normalize after CABG. Post-CABG, PGC1α nuclear-bound content was increased compared with Hib (9.02 ± 0.48 versus 5.54 ± 0.98 arbitrary units, respectively; P < 0.05), and expression of mitofusins-1 and 2 and optic atrophy-1 more than doubled. PGC1α and mitochondrial fusion proteins are increased 4 wk post-CABG in hibernating hearts, indicating mitochondrial fusion has begun to occur and signaling early mitochondrial recovery. Future studies should address changes in maximal myocardial oxygen consumption relative to mitochondrial protein expression.

Research paper thumbnail of Vav1 Dephosphorylation by the Tyrosine Phosphatase SHP1 as a Mechanism for Inhibition of Cellular Cytotoxicity

Molecular and Cellular Biology, 2003

Here, we present data suggesting a novel mechanism for regulation of natural killer (NK) cell cyt... more Here, we present data suggesting a novel mechanism for regulation of natural killer (NK) cell cytotoxicity through inhibitory receptors. Interaction of activation receptors with their ligands on target cells induces cytotoxicity by NK cells. This activation is under negative control by inhibitory receptors that recruit tyrosine phosphatase SHP-1 upon binding major histocompatibility class I on target cells. How SHP-1 blocks

Research paper thumbnail of 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 majo... more 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 KIIK 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 (F1KYNGLIHR 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 ofpeptides bound to HLA-B*2705.

Research paper thumbnail of Antigen presentation mediated by recycling of surface HLA-DR molecules

Nature, 1995

CLASS II histocompatibility molecules associate with peptides derived from antigens that are proc... more 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 molecules1, associated invariant chain2,3, and HLA-DM4,5. Exceptions to these rules have been reported6-10, but without description of an underlying mechanism. Here we show that presentation of immunodominant epitopes in the haemagglutinin

Research paper thumbnail of Expression of uncoupling protein-2 remains increased within hibernating myocardium despite successful coronary artery bypass grafting at 4 wk post-revascularization

Journal of Surgical Research, 2015

Background-We have previously shown that mitochondrial uncoupling protein-2 (UCP-2) is increased ... more Background-We have previously shown that mitochondrial uncoupling protein-2 (UCP-2) is increased in a swine model of hibernating myocardium (HM). Although UCP-2 reduces oxidant stress, it can promote inefficiency of the electron transport chain. In this study, we tested whether UCP-2 remains increased in revascularized HM (RHM) following coronary artery bypass grafting (CABG). Methods-Seven swine underwent thoracotomy with placement of a constrictor on the left anterior descending artery (LAD). Twelve weeks later, a left internal mammary artery (LIMA) graft was placed on the distal LAD. Four weeks post-CABG, CT angiography documented patent grafts and function. At the terminal study, blood flow to the LAD and remote territories were assessed during high-dose dobutamine and mitochondria isolated from both regions for analysis. Comparisons were made to a group of swine with HM who underwent constrictor placement without bypass grafting (n=4). Results-During dobutamine infusion, RHM demonstrated lower blood flows (2.44±0.23 versus 3.43±0.30 ml/min/g; P<0.05) and reduced wall thickening (33±9% versus 52±13%; P<0.05)

Research paper thumbnail of Negative Signaling Pathways of the Killer Cell Inhibitory Receptor and Fc~/RIIbl Require Distinct Phosphatases

Summary Inhibition of natural killer (NK) cells by the killer cell inhibitory receptor (KIR) invo... more Summary Inhibition of natural killer (NK) cells by the killer cell inhibitory receptor (KIR) involves re- cruitment of the tyrosine phosphatase SHP-1 by KIR and is prevented by expression of a dom- inant negative SHP-1 mutant. Another inhibitory receptor, the low affinity Fc receptor for im- munoglobulin G (IgG) (Fc',/Rllbl), has been shown to bind SHP-1 when cocross-hnked with the

Research paper thumbnail of Isoforms of the invariant chain regulate transport of MHC class II molecules to antigen processing compartments

The Journal of Cell Biology, 1996

Newly synthesized class II molecules of the major histocompatibility complex must be transported ... more 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 appa...

Research paper thumbnail of HLA class I recognition by killer cell Ig-like receptors

Seminars in Immunology, 2000

Natural killer NK cell activation is regulated by a combination of positive and negative signals ... more Natural killer NK cell activation is regulated by a combination of positive and negative signals coming from repertoires of activating and inhibitory receptors expressed on the surface of each NK cell. Receptors specific for major histo-() compatibility complex MHC class I molecules on target cells inhibit NK effector functions such as cytotoxicity and cytokine production. As reviewed here, the killer cell Ig-like () receptor KIR family on human NK cells includes receptors with activating as well as inhibitory potential, receptors that have no known ligand, and inhibitory receptors with welldefined specificities for MHC class I such as HLA-B and HLA-C.

Research paper thumbnail of Cell surface HLA-DR-invariant chain complexes are targeted to endosomes by rapid internalization

Proceedings of the National Academy of Sciences, 1993

Clas U molecules of the major histocompatibilty complex (MHIC) bind peptides derived from protein... more Clas U molecules of the major histocompatibilty complex (MHIC) bind peptides derived from protein antigens delivered Into enldocytic compartments and present these peptides to CD4+ T cells. The prcursors to fbnctional MHIC clas U molecules loaded with peptides are complexes of the invariant chain asociated with dass U ap heterodimers.

Research paper thumbnail of Antigen presentation mediated by recycling of surface HLA-DR molecules

Nature, 1995

Class II histocompatibility molecules associate with peptides derived from antigens that are proc... more 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.

Research paper thumbnail of Processing pathways for presentation of cytosolic antigen to MHC class II-restricted T cells

Nature, 1992

Antigens presented to CD4+ T cells derive primarily from exogenous proteins that are processed in... more Antigens presented to CD4+ T cells derive primarily from exogenous proteins that are processed into peptides capable of binding to class II major histocompatibility complex (MHC) molecules in an endocytic compartment. In contrast, antigens presented to CD8+ T cells derive mostly from proteins processed in the cytosol, and peptide loading onto class I MHC molecules in an early exocytic compartment is dependent on a transporter for antigen presentation encoded in the class II MHC region. Endogenous cytosolic antigen can also be presented by class II molecules. Here we show that, unlike class I-restricted recognition of antigen, HLA-DR1-restricted recognition of cytosolic antigen occurs in mutant cells without a transporter for antigen presentation. In contrast, DR1-restricted recognition of a short cytosolic peptide is dependent on such a transporter. Thus helper T-cell epitopes can be generated from cytosolic antigens by several mechanisms, one of which is distinct from the classical class I pathway.

Research paper thumbnail of Vav1 Dephosphorylation by the Tyrosine Phosphatase SHP-1 as a Mechanism for Inhibition of Cellular Cytotoxicity

Molecular and Cellular Biology, 2003

Here, we present data suggesting a novel mechanism for regulation of natural killer (NK) cell cyt... more Here, we present data suggesting a novel mechanism for regulation of natural killer (NK) cell cytotoxicity through inhibitory receptors. Interaction of activation receptors with their ligands on target cells induces cytotoxicity by NK cells. This activation is under negative control by inhibitory receptors that recruit tyrosine phosphatase SHP-1 upon binding major histocompatibility class I on target cells. How SHP-1 blocks the activation pathway is not known. To identify SHP-1 substrates, an HLA-C-specific inhibitory receptor fused to a substrate-trapping mutant of SHP-1 was expressed in NK cells. Phosphorylated Vav1, a regulator of actin cytoskeleton, was the only protein detectably associated with the catalytic site of SHP-1 during NK cell contact with target cells expressing HLA-C. Vav1 trapping was independent of actin polymerization, suggesting that inhibition of cellular cytotoxicity occurs through an early dephosphorylation of Vav1 by SHP-1, which blocks actin-dependent acti...

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

Journal of Experimental Medicine, 1996

Natural killer (NK) cells express a repertoire of killer cell inhibitory receptors (KIR) for majo... more 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 RRSKEI...

Research paper thumbnail of The Direct Binding of a p58 Killer Cell Inhibitory Receptor to Human Histocompatibility Leukocyte Antigen (HLA)-Cw4 Exhibits Peptide Selectivity

Journal of Experimental Medicine, 1997

Natural killer (NK) cells in mice and humans express a number of structurally diverse receptors t... more Natural killer (NK) cells in mice and humans express a number of structurally diverse receptors that inhibit target cell lysis upon recognition of major histocompatibility complex (MHC) class I molecules expressed on targets. The contribution of peptide to the structural features of class I required for NK cell inhibition appears to vary depending on the type of receptor engaged. Thus, while there is no peptide specificity in NK inhibition mediated by Ly-49A in the mouse, human histocompatibility antigen (HLA)-B*2705–specific NK clones displayed selectivity for peptides. In this report, we examine the role of peptide in the recognition of HLA-C by the defined killer cell inhibitory receptor (KIR) cl42 with established specificity for HLA-Cw4. Binding of soluble KIR cl42 molecules to HLA-Cw4 expressed on transporter associated with antigen presentation (TAP)-deficient RMA-S cells occurred only upon exogenous peptide loading. Moreover, there was peptide selectivity in that certain sub...

Research paper thumbnail of Type I transmembrane receptor with inhibitory function in mouse mast cells and NK cells

Research paper thumbnail of Killer cell inhibitory receptors: diversity, specificity, and function

Immunological Reviews, 1997

NK cells selectively kill target cells that fail to express self-MHC class I molecules. This sele... more 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.

Research paper thumbnail of Recruitment of Tyrosine Phosphatase HCP by the Killer Cell Inhibitory Receptor

Immunity, 1996

Cytolysis of target cells by natural killer (NK) cells and by some cytotoxic T cells occurs unles... more 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.

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

Immunity, 1995

Recognition of major histocompatibility class 1 molecules on target ce Ils by natural killer (NK)... more Recognition of major histocompatibility class 1 molecules on target ce Ils by natural killer (NK) cells confers selective protection from NK-mediated Iysis. Crosslinking of the p58 NK receptor, involved in the recognition of HLA-C alleles, delivers a negative signal that prevents target cell Iysis. 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. Ali 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.

Research paper thumbnail of Interleukin-2 activity can be fine tuned with engineered receptor signaling clamps

Immunity, Jan 19, 2015

Interleukin-2 (IL-2) regulates lymphocyte function by signaling through heterodimerization of the... more Interleukin-2 (IL-2) regulates lymphocyte function by signaling through heterodimerization of the IL-2Rβ and γc receptor subunits. IL-2 is of considerable therapeutic interest, but harnessing its actions in a controllable manner remains a challenge. Previously, we have engineered an IL-2 "superkine" with enhanced affinity for IL-2Rβ. Here, we describe next-generation IL-2 variants that function as "receptor signaling…

Research paper thumbnail of Natural killer cell receptors

The specificity in the recognition of hematopoietic target cells by natural killer cells is prima... more 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.

Research paper thumbnail of Mitochondrial fusion proteins in revascularized hibernating hearts

Journal of Surgical Research, 2015

Hibernating myocardium is characterized by viable yet dysfunctional myocardium secondary to chron... more Hibernating myocardium is characterized by viable yet dysfunctional myocardium secondary to chronic ischemia, with studies demonstrating incomplete early recovery after coronary artery bypass graft (CABG). We tested whether mitochondrial fusion proteins, an indicator of mitochondrial biogenesis, are increased in hibernating myocardium post-CABG. A constrictor was placed on the left anterior descending (LAD) artery of nine pigs. Four of these pigs additionally underwent CABG 12 wk later with a left internal mammary artery graft to the LAD distal to the constrictor. Five pigs had a constrictor placed but did not undergo CABG (Hib). Five pigs did not have a constrictor placed (control). Computerized tomography angiography was used to confirm stenosis at the site of constrictor placement and patency of left internal mammary artery grafts. Regional blood flows were determined at baseline and during 40 μg/kg/min dobutamine infusion. Mitochondrial proteins were quantified by Western blot. Blood flow in the LAD region after CABG was lower than remote regions during dobutamine infusion (2.54 ± 0.24 versus 3.46 ± 0.33 mL/min/g; P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.05). Electron transport chain proteins were ∼70% lower in Hib compared with those in control and failed to normalize after CABG. Post-CABG, PGC1α nuclear-bound content was increased compared with Hib (9.02 ± 0.48 versus 5.54 ± 0.98 arbitrary units, respectively; P &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt; 0.05), and expression of mitofusins-1 and 2 and optic atrophy-1 more than doubled. PGC1α and mitochondrial fusion proteins are increased 4 wk post-CABG in hibernating hearts, indicating mitochondrial fusion has begun to occur and signaling early mitochondrial recovery. Future studies should address changes in maximal myocardial oxygen consumption relative to mitochondrial protein expression.

Research paper thumbnail of Vav1 Dephosphorylation by the Tyrosine Phosphatase SHP1 as a Mechanism for Inhibition of Cellular Cytotoxicity

Molecular and Cellular Biology, 2003

Here, we present data suggesting a novel mechanism for regulation of natural killer (NK) cell cyt... more Here, we present data suggesting a novel mechanism for regulation of natural killer (NK) cell cytotoxicity through inhibitory receptors. Interaction of activation receptors with their ligands on target cells induces cytotoxicity by NK cells. This activation is under negative control by inhibitory receptors that recruit tyrosine phosphatase SHP-1 upon binding major histocompatibility class I on target cells. How SHP-1 blocks

Research paper thumbnail of 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 majo... more 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 KIIK 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 (F1KYNGLIHR 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 ofpeptides bound to HLA-B*2705.

Research paper thumbnail of Antigen presentation mediated by recycling of surface HLA-DR molecules

Nature, 1995

CLASS II histocompatibility molecules associate with peptides derived from antigens that are proc... more 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 molecules1, associated invariant chain2,3, and HLA-DM4,5. Exceptions to these rules have been reported6-10, but without description of an underlying mechanism. Here we show that presentation of immunodominant epitopes in the haemagglutinin

Research paper thumbnail of Expression of uncoupling protein-2 remains increased within hibernating myocardium despite successful coronary artery bypass grafting at 4 wk post-revascularization

Journal of Surgical Research, 2015

Background-We have previously shown that mitochondrial uncoupling protein-2 (UCP-2) is increased ... more Background-We have previously shown that mitochondrial uncoupling protein-2 (UCP-2) is increased in a swine model of hibernating myocardium (HM). Although UCP-2 reduces oxidant stress, it can promote inefficiency of the electron transport chain. In this study, we tested whether UCP-2 remains increased in revascularized HM (RHM) following coronary artery bypass grafting (CABG). Methods-Seven swine underwent thoracotomy with placement of a constrictor on the left anterior descending artery (LAD). Twelve weeks later, a left internal mammary artery (LIMA) graft was placed on the distal LAD. Four weeks post-CABG, CT angiography documented patent grafts and function. At the terminal study, blood flow to the LAD and remote territories were assessed during high-dose dobutamine and mitochondria isolated from both regions for analysis. Comparisons were made to a group of swine with HM who underwent constrictor placement without bypass grafting (n=4). Results-During dobutamine infusion, RHM demonstrated lower blood flows (2.44±0.23 versus 3.43±0.30 ml/min/g; P<0.05) and reduced wall thickening (33±9% versus 52±13%; P<0.05)

Research paper thumbnail of Negative Signaling Pathways of the Killer Cell Inhibitory Receptor and Fc~/RIIbl Require Distinct Phosphatases

Summary Inhibition of natural killer (NK) cells by the killer cell inhibitory receptor (KIR) invo... more Summary Inhibition of natural killer (NK) cells by the killer cell inhibitory receptor (KIR) involves re- cruitment of the tyrosine phosphatase SHP-1 by KIR and is prevented by expression of a dom- inant negative SHP-1 mutant. Another inhibitory receptor, the low affinity Fc receptor for im- munoglobulin G (IgG) (Fc',/Rllbl), has been shown to bind SHP-1 when cocross-hnked with the

Research paper thumbnail of Isoforms of the invariant chain regulate transport of MHC class II molecules to antigen processing compartments

The Journal of Cell Biology, 1996

Newly synthesized class II molecules of the major histocompatibility complex must be transported ... more 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 appa...

Research paper thumbnail of HLA class I recognition by killer cell Ig-like receptors

Seminars in Immunology, 2000

Natural killer NK cell activation is regulated by a combination of positive and negative signals ... more Natural killer NK cell activation is regulated by a combination of positive and negative signals coming from repertoires of activating and inhibitory receptors expressed on the surface of each NK cell. Receptors specific for major histo-() compatibility complex MHC class I molecules on target cells inhibit NK effector functions such as cytotoxicity and cytokine production. As reviewed here, the killer cell Ig-like () receptor KIR family on human NK cells includes receptors with activating as well as inhibitory potential, receptors that have no known ligand, and inhibitory receptors with welldefined specificities for MHC class I such as HLA-B and HLA-C.

Research paper thumbnail of Cell surface HLA-DR-invariant chain complexes are targeted to endosomes by rapid internalization

Proceedings of the National Academy of Sciences, 1993

Clas U molecules of the major histocompatibilty complex (MHIC) bind peptides derived from protein... more Clas U molecules of the major histocompatibilty complex (MHIC) bind peptides derived from protein antigens delivered Into enldocytic compartments and present these peptides to CD4+ T cells. The prcursors to fbnctional MHIC clas U molecules loaded with peptides are complexes of the invariant chain asociated with dass U ap heterodimers.

Research paper thumbnail of Antigen presentation mediated by recycling of surface HLA-DR molecules

Nature, 1995

Class II histocompatibility molecules associate with peptides derived from antigens that are proc... more 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.

Research paper thumbnail of Processing pathways for presentation of cytosolic antigen to MHC class II-restricted T cells

Nature, 1992

Antigens presented to CD4+ T cells derive primarily from exogenous proteins that are processed in... more Antigens presented to CD4+ T cells derive primarily from exogenous proteins that are processed into peptides capable of binding to class II major histocompatibility complex (MHC) molecules in an endocytic compartment. In contrast, antigens presented to CD8+ T cells derive mostly from proteins processed in the cytosol, and peptide loading onto class I MHC molecules in an early exocytic compartment is dependent on a transporter for antigen presentation encoded in the class II MHC region. Endogenous cytosolic antigen can also be presented by class II molecules. Here we show that, unlike class I-restricted recognition of antigen, HLA-DR1-restricted recognition of cytosolic antigen occurs in mutant cells without a transporter for antigen presentation. In contrast, DR1-restricted recognition of a short cytosolic peptide is dependent on such a transporter. Thus helper T-cell epitopes can be generated from cytosolic antigens by several mechanisms, one of which is distinct from the classical class I pathway.

Research paper thumbnail of Vav1 Dephosphorylation by the Tyrosine Phosphatase SHP-1 as a Mechanism for Inhibition of Cellular Cytotoxicity

Molecular and Cellular Biology, 2003

Here, we present data suggesting a novel mechanism for regulation of natural killer (NK) cell cyt... more Here, we present data suggesting a novel mechanism for regulation of natural killer (NK) cell cytotoxicity through inhibitory receptors. Interaction of activation receptors with their ligands on target cells induces cytotoxicity by NK cells. This activation is under negative control by inhibitory receptors that recruit tyrosine phosphatase SHP-1 upon binding major histocompatibility class I on target cells. How SHP-1 blocks the activation pathway is not known. To identify SHP-1 substrates, an HLA-C-specific inhibitory receptor fused to a substrate-trapping mutant of SHP-1 was expressed in NK cells. Phosphorylated Vav1, a regulator of actin cytoskeleton, was the only protein detectably associated with the catalytic site of SHP-1 during NK cell contact with target cells expressing HLA-C. Vav1 trapping was independent of actin polymerization, suggesting that inhibition of cellular cytotoxicity occurs through an early dephosphorylation of Vav1 by SHP-1, which blocks actin-dependent acti...

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

Journal of Experimental Medicine, 1996

Natural killer (NK) cells express a repertoire of killer cell inhibitory receptors (KIR) for majo... more 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 RRSKEI...

Research paper thumbnail of The Direct Binding of a p58 Killer Cell Inhibitory Receptor to Human Histocompatibility Leukocyte Antigen (HLA)-Cw4 Exhibits Peptide Selectivity

Journal of Experimental Medicine, 1997

Natural killer (NK) cells in mice and humans express a number of structurally diverse receptors t... more Natural killer (NK) cells in mice and humans express a number of structurally diverse receptors that inhibit target cell lysis upon recognition of major histocompatibility complex (MHC) class I molecules expressed on targets. The contribution of peptide to the structural features of class I required for NK cell inhibition appears to vary depending on the type of receptor engaged. Thus, while there is no peptide specificity in NK inhibition mediated by Ly-49A in the mouse, human histocompatibility antigen (HLA)-B*2705–specific NK clones displayed selectivity for peptides. In this report, we examine the role of peptide in the recognition of HLA-C by the defined killer cell inhibitory receptor (KIR) cl42 with established specificity for HLA-Cw4. Binding of soluble KIR cl42 molecules to HLA-Cw4 expressed on transporter associated with antigen presentation (TAP)-deficient RMA-S cells occurred only upon exogenous peptide loading. Moreover, there was peptide selectivity in that certain sub...

Research paper thumbnail of Type I transmembrane receptor with inhibitory function in mouse mast cells and NK cells

Research paper thumbnail of Killer cell inhibitory receptors: diversity, specificity, and function

Immunological Reviews, 1997

NK cells selectively kill target cells that fail to express self-MHC class I molecules. This sele... more 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.

Research paper thumbnail of Recruitment of Tyrosine Phosphatase HCP by the Killer Cell Inhibitory Receptor

Immunity, 1996

Cytolysis of target cells by natural killer (NK) cells and by some cytotoxic T cells occurs unles... more 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.

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

Immunity, 1995

Recognition of major histocompatibility class 1 molecules on target ce Ils by natural killer (NK)... more Recognition of major histocompatibility class 1 molecules on target ce Ils by natural killer (NK) cells confers selective protection from NK-mediated Iysis. Crosslinking of the p58 NK receptor, involved in the recognition of HLA-C alleles, delivers a negative signal that prevents target cell Iysis. 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. Ali 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.