Regulation and lectin activity of the human neutrophil peripheral lymph node homing receptor

We characterize the nature and regulation of a human neutrophil cell surface antigen recognized by monoclonal antibodies (the DREG series) against a human lymphocyte peripheral lymph node homing receptor. Human neutrophils express high levels of the DREG antigen, whose expression is downregulated after treatment with phorbol myristate acetate, or the chemotactic factors C5a and FMLP. Interestingly, C5a treatment also downregulated the monocyte DREG antigen, but had no effect on expression of the lymphocyte molecule. Within 3 minutes after treatment with C5a, greater than 80% of neutrophil DREG antigen expression is lost, and essentially the molecule is completely removed from the cell surface by 5 minutes. The human neutrophil DREG antigen is 10 Kd larger than the lymphocyte molecule. These features are similar to those of the mouse neutrophil MEL-14 antigen (murine peripheral lymph node homing receptor). The mannose-6-phosphate rich phosphomannan (PPME) binds human lymphocytes via the DREG antigen. PPME also binds neutrophils, but little difference in binding is seen between unactivated and activated cells. We show that PPME binding to unactivated neutrophils is mediated primarily by a cation- and DREG antigen-dependent mechanism, whereas activated neutrophil-PPME binding is DREG antigen- and cation-independent, and may be due to the translocation of lysosomal mannose-6-phosphate receptors to the cell surface. The DREG antibodies offer powerful tools for analyzing the role of homing receptors in human neutrophil-endothelial cell interactions, and also may prove valuable in the clinical assessment of neutrophil activation.

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Regulation and lectin activity of the human neutrophil peripheral lymph node homing receptor

Blood ◽

10.1182/blood.v76.1.178.178 ◽

1990 ◽

Vol 76 (1) ◽

pp. 178-183 ◽

Cited By ~ 42

Author(s):

MA Jutila ◽

TK Kishimoto ◽

EC Butcher

Keyword(s):

Lymph Node ◽

Cell Surface ◽

Human Neutrophil ◽

Human Lymphocytes ◽

Human Neutrophils ◽

Peripheral Lymph Node ◽

Homing Receptor ◽

Lymph Node Homing ◽

Peripheral Lymph ◽

Mannose 6 Phosphate

Abstract We characterize the nature and regulation of a human neutrophil cell surface antigen recognized by monoclonal antibodies (the DREG series) against a human lymphocyte peripheral lymph node homing receptor. Human neutrophils express high levels of the DREG antigen, whose expression is downregulated after treatment with phorbol myristate acetate, or the chemotactic factors C5a and FMLP. Interestingly, C5a treatment also downregulated the monocyte DREG antigen, but had no effect on expression of the lymphocyte molecule. Within 3 minutes after treatment with C5a, greater than 80% of neutrophil DREG antigen expression is lost, and essentially the molecule is completely removed from the cell surface by 5 minutes. The human neutrophil DREG antigen is 10 Kd larger than the lymphocyte molecule. These features are similar to those of the mouse neutrophil MEL-14 antigen (murine peripheral lymph node homing receptor). The mannose-6-phosphate rich phosphomannan (PPME) binds human lymphocytes via the DREG antigen. PPME also binds neutrophils, but little difference in binding is seen between unactivated and activated cells. We show that PPME binding to unactivated neutrophils is mediated primarily by a cation- and DREG antigen-dependent mechanism, whereas activated neutrophil-PPME binding is DREG antigen- and cation-independent, and may be due to the translocation of lysosomal mannose-6-phosphate receptors to the cell surface. The DREG antibodies offer powerful tools for analyzing the role of homing receptors in human neutrophil-endothelial cell interactions, and also may prove valuable in the clinical assessment of neutrophil activation.

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The peripheral lymph node homing receptor, LECAM-1, is involved in CD18-independent adhesion of human neutrophils to endothelium

Biochemical and Biophysical Research Communications ◽

10.1016/0006-291x(91)90511-5 ◽

1991 ◽

Vol 174 (1) ◽

pp. 236-243 ◽

Cited By ~ 52

Author(s):

R. Hallmann ◽

M.A. Jutila ◽

C.W. Smith ◽

D.C. Anderson ◽

T.K. Kishimoto ◽

...

Keyword(s):

Lymph Node ◽

Human Neutrophils ◽

Peripheral Lymph Node ◽

Homing Receptor ◽

Lymph Node Homing ◽

Peripheral Lymph

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Characterization of a human homologue of the murine peripheral lymph node homing receptor.

The Journal of Cell Biology ◽

10.1083/jcb.109.1.421 ◽

1989 ◽

Vol 109 (1) ◽

pp. 421-427 ◽

Cited By ~ 60

Author(s):

B R Bowen ◽

T Nguyen ◽

L A Lasky

Keyword(s):

Cell Adhesion ◽

Lymph Node ◽

Adhesion Molecule ◽

Carbohydrate Binding ◽

Human Homologue ◽

Peripheral Lymph Node ◽

Homing Receptor ◽

Lymph Node Homing ◽

Peripheral Lymph ◽

Human Receptor

Lymphocyte trafficking is a fundamental aspect of the immune system that allows B and T lymphocytes with diverse antigen recognition specificities to be exposed to various antigenic stimuli in spatially distinct regions of an organism. A lymphocyte adhesion molecule that is involved with this trafficking phenomenon has been termed the homing receptor. Previous work (Lasky, L., T. Yednock, M. Singer, D. Dowbenko, C. Fennie, H. Rodriguez, T. Nguyen, S. Stachel, and S. Rosen. 1989. Cell. 56:1045-1055) has characterized a cDNA clone encoding a murine homing receptor that is involved in trafficking of lymphocytes to peripheral lymph nodes. This molecule was found to contain a number of protein motifs, the most intriguing of which was a carbohydrate binding domain, or lectin, that is apparently involved in the adhesive interaction between murine lymphocytes and peripheral lymph node endothelium. In this study, we have used the murine cDNA clone to isolate a human homologue of this peripheral lymph node-specific adhesion molecule. The human receptor was found to be highly homologous to the murine receptor in overall sequence, but showed no sequence similarity to another surface protein that may be involved with human lymphocyte homing, the Hermes glycoprotein. The extracellular region of the human receptor contained an NH2 terminally located carbohydrate binding domain followed by an EGF-like domain and a domain containing two repeats of a complement binding motif. Transient cell transfection assays using the human receptor cDNA showed that it encoded a surface glycoprotein that cross reacted with a polyclonal antibody directed against the murine peripheral lymph node homing receptor. Interestingly, the human receptor showed a high degree of sequence homology to another human cell adhesion glycoprotein, the endothelial cell adhesion molecule ELAM.

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The Peripheral Lymph Node Homing Receptor: Tandem Structural and Functional Domains in a Novel Class of Adhesion Molecule

Cellular and Molecular Mechanisms of Inflammation ◽

10.1016/b978-0-12-150402-1.50007-x ◽

1991 ◽

pp. 61-89

Author(s):

Mark H. Siegelman ◽

Irving L. Weissman

Keyword(s):

Lymph Node ◽

Adhesion Molecule ◽

Functional Domains ◽

Peripheral Lymph Node ◽

Homing Receptor ◽

Lymph Node Homing ◽

Peripheral Lymph

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Human neutrophils release the Leu-8 lymph node homing receptor during cell activation

Blood ◽

10.1182/blood.v76.11.2381.bloodjournal76112381 ◽

1990 ◽

Vol 76 (11) ◽

pp. 2381-2388 ◽

Cited By ~ 1

Author(s):

M Berg ◽

SP James

Keyword(s):

Lymph Node ◽

Cell Activation ◽

Human Lymphocytes ◽

Sodium Dodecyl ◽

Lymphoid Cells ◽

Human Neutrophils ◽

Cdna Clones ◽

Homing Receptor ◽

Cell Extracts ◽

Lymph Node Homing

The Leu-8 molecule, the human homologue of the murine MEL-14 peripheral lymph node homing receptor, is expressed on neutrophils in both species and may be important in localization of cells to sites of inflammation. Most circulating human neutrophils express the Leu-8 molecule, and activation of neutrophils with phorbol myristate acetate causes a rapid decline in Leu-8 membrane fluorescence staining within 15 minutes. Northern blot analysis of total cellular RNA from neutrophils demonstrated two species of Leu-8 messenger RNA, a major one of 2.4 kb and a minor one of 1.9 kb. Because two different Leu-8 cDNA clones were obtained from human lymphocytes that were predicted to encode both transmembrane and phosphatidylinositol (PI)-anchored forms of the molecule, experiments were conducted to determine whether Leu-8 is anchored to neutrophils by a PI-anchor. There was a slight decrease in expression of Leu-8 on neutrophils when they were treated with PI- specific phospholipase C (PI-PLC). However, Leu-8 was abundant on neutrophils obtained from a patient with paroxysmal nocturnal hemoglobinuria. To determine the fate of the Leu-8 molecule during cell activation, neutrophils were labeled with 125I-anti-Leu-8. During activation antibody was rapidly lost from the cell surface and was not internalized, suggesting that Leu-8 is released from the cell membrane during cell activation. When cell extracts of neutrophils were compared with extracts of lymphoid cells by sodium dodecyl sulfate- polyacrylamide gel electrophoresis and immunoblotting, the Leu-8 species expressed on neutrophils had a significantly higher and more variable relative mobility (70 to 120 Kd for neutrophils v 70 Kd for Jurkat T cells). In addition, Leu-8 molecules were detected in the supernatants of activated neutrophils. These results indicate that human neutrophils express a high-molecular-weight form of the Leu-8 molecule that has a conventional transmembrane anchor and is rapidly released from the membrane during activation. The loss of the Leu-8 membrane glycoprotein during activation may be a mechanism for rapid alteration of neutrophil adhesion characteristics.

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Identification of a human peripheral lymph node homing receptor: a rapidly down-regulated adhesion molecule.

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.87.6.2244 ◽

1990 ◽

Vol 87 (6) ◽

pp. 2244-2248 ◽

Cited By ~ 251

Author(s):

T. K. Kishimoto ◽

M. A. Jutila ◽

E. C. Butcher

Keyword(s):

Lymph Node ◽

Adhesion Molecule ◽

Peripheral Lymph Node ◽

Homing Receptor ◽

Lymph Node Homing ◽

Peripheral Lymph

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Human neutrophils release the Leu-8 lymph node homing receptor during cell activation

Blood ◽

10.1182/blood.v76.11.2381.2381 ◽

1990 ◽

Vol 76 (11) ◽

pp. 2381-2388 ◽

Cited By ~ 52

Author(s):

M Berg ◽

SP James

Keyword(s):

Lymph Node ◽

Cell Activation ◽

Human Lymphocytes ◽

Sodium Dodecyl ◽

Lymphoid Cells ◽

Human Neutrophils ◽

Cdna Clones ◽

Homing Receptor ◽

Cell Extracts ◽

Lymph Node Homing

Abstract The Leu-8 molecule, the human homologue of the murine MEL-14 peripheral lymph node homing receptor, is expressed on neutrophils in both species and may be important in localization of cells to sites of inflammation. Most circulating human neutrophils express the Leu-8 molecule, and activation of neutrophils with phorbol myristate acetate causes a rapid decline in Leu-8 membrane fluorescence staining within 15 minutes. Northern blot analysis of total cellular RNA from neutrophils demonstrated two species of Leu-8 messenger RNA, a major one of 2.4 kb and a minor one of 1.9 kb. Because two different Leu-8 cDNA clones were obtained from human lymphocytes that were predicted to encode both transmembrane and phosphatidylinositol (PI)-anchored forms of the molecule, experiments were conducted to determine whether Leu-8 is anchored to neutrophils by a PI-anchor. There was a slight decrease in expression of Leu-8 on neutrophils when they were treated with PI- specific phospholipase C (PI-PLC). However, Leu-8 was abundant on neutrophils obtained from a patient with paroxysmal nocturnal hemoglobinuria. To determine the fate of the Leu-8 molecule during cell activation, neutrophils were labeled with 125I-anti-Leu-8. During activation antibody was rapidly lost from the cell surface and was not internalized, suggesting that Leu-8 is released from the cell membrane during cell activation. When cell extracts of neutrophils were compared with extracts of lymphoid cells by sodium dodecyl sulfate- polyacrylamide gel electrophoresis and immunoblotting, the Leu-8 species expressed on neutrophils had a significantly higher and more variable relative mobility (70 to 120 Kd for neutrophils v 70 Kd for Jurkat T cells). In addition, Leu-8 molecules were detected in the supernatants of activated neutrophils. These results indicate that human neutrophils express a high-molecular-weight form of the Leu-8 molecule that has a conventional transmembrane anchor and is rapidly released from the membrane during activation. The loss of the Leu-8 membrane glycoprotein during activation may be a mechanism for rapid alteration of neutrophil adhesion characteristics.

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Identification of a carbohydrate-based endothelial ligand for a lymphocyte homing receptor.

The Journal of Cell Biology ◽

10.1083/jcb.113.5.1213 ◽

1991 ◽

Vol 113 (5) ◽

pp. 1213-1221 ◽

Cited By ~ 183

Author(s):

Y Imai ◽

M S Singer ◽

C Fennie ◽

L A Lasky ◽

S D Rosen

Keyword(s):

Lymph Node ◽

Peripheral Lymph Node ◽

Lymphocyte Homing ◽

High Endothelial Venules ◽

Homing Receptor ◽

Calcium Dependent ◽

Lymph Node Homing ◽

Recombinant Receptor ◽

Peripheral Lymph ◽

Mouse Lymphocytes

Lymphocyte attachment to high endothelial venules within lymph nodes is mediated by the peripheral lymph node homing receptor (pnHR), originally defined on mouse lymphocytes by the MEL-14 mAb. The pnHR is a calcium-dependent lectin-like receptor, a member of the LEC-CAM family of adhesion proteins. Here, using a soluble recombinant form of the homing receptor, we have identified an endothelial ligand for the pnHR as an approximately 50-kD sulfated, fucosylated, and sialylated glycoprotein, which we designate Sgp50 (sulfated glycoprotein of 50 kD). Recombinant receptor binding to this lymph node-specific glycoprotein requires calcium and is inhibitable by specific carbohydrates and by MEL-14 mAb. Sialylation of the component is required for binding. Additionally, the glycoprotein is precipitated by MECA-79, an adhesion-blocking mAb reactive with lymph node HEV. A related glycoprotein of approximately 90 kD (designated as Sgp90) is also identified.

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