Sialic acid–binding immunoglobulin-like lectin 8 (Siglec-8) is an activating receptor mediating β2-integrin–dependent function in human eosinophils

Daniela J. Carroll, Jeremy Alastair O'Sullivan, David B. Nix, Yun Cao, Michael Tiemeyer, Bruce Scott Bochner*

*Corresponding author for this work

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

Background: Siglec-8 is a CD33 subfamily cell-surface receptor selectively expressed on human eosinophils. After cytokine priming, Siglec-8 mAb or glycan ligand binding causes eosinophil apoptosis associated with reactive oxygen species (ROS) production. Most CD33-related Siglecs function as inhibitory receptors, but the ability of Siglec-8 to stimulate eosinophil ROS production and apoptosis suggests that Siglec-8 might instead function as an activating receptor. Objective: We sought to determine the role of IL-5 priming and identify the signaling molecules involved in Siglec-8 function for human eosinophils. Methods: We used an mAb and/or a multimeric synthetic sulfated sialoglycan ligand recognizing Siglec-8 in combination with integrin blocking antibodies, pharmacologic inhibitors, phosphoproteomics, and Western blot analysis to define the necessity of various proteins involved in Siglec-8 function for human eosinophils. Results: Cytokine priming was required to elicit the unanticipated finding that Siglec-8 engagement promotes rapid β2-integrin–dependent eosinophil adhesion. Also novel was the finding that this adhesion was necessary for subsequent ROS production and apoptosis. Siglec-8–mediated ROS was generated through reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activation because pretreatment of eosinophils with catalase (an extracellular superoxide scavenger) or NSC 23766 (a Rac GTPase inhibitor) completely inhibited Siglec-8–mediated eosinophil apoptosis. Finally, engagement of Siglec-8 on IL-5–primed eosinophils resulted in increased phosphorylation of Akt, p38, and c-Jun N-terminal kinase 1 that was also β2-integrin dependent; pharmacologic inhibition of these kinases completely prevented Siglec-8–mediated eosinophil apoptosis. Conclusions: These data demonstrate that Siglec-8 functions uniquely as an activating receptor on IL-5–primed eosinophils through a novel pathway involving regulation of β2-integrin–dependent adhesion, NADPH oxidase, and a subset of protein kinases.

Original languageEnglish (US)
Pages (from-to)2196-2207
Number of pages12
JournalJournal of Allergy and Clinical Immunology
Volume141
Issue number6
DOIs
StatePublished - Jun 1 2018

Fingerprint

Saliva
Eosinophils
Lectins
Immunoglobulins
Sialic Acid Binding Immunoglobulin-like Lectins
Apoptosis
Reactive Oxygen Species
NADP
Integrins
Oxidoreductases
Mitogen-Activated Protein Kinase 8
Cytokines
Ligands
Blocking Antibodies
GTP Phosphohydrolases
Interleukin-5
Cell Surface Receptors
Interleukin-8
Superoxides
Catalase

Keywords

  • Akt
  • Eosinophil
  • Siglec-8
  • apoptosis
  • c-Jun N-terminal kinase (JNK)
  • p38
  • phosphoproteomics
  • reduced nicotinamide adenine dinucleotide phosphate oxidase (NADPH)
  • β-integrin

ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology

Cite this

@article{84bfab1757364d45b9ce56cd1c9b2297,
title = "Sialic acid–binding immunoglobulin-like lectin 8 (Siglec-8) is an activating receptor mediating β2-integrin–dependent function in human eosinophils",
abstract = "Background: Siglec-8 is a CD33 subfamily cell-surface receptor selectively expressed on human eosinophils. After cytokine priming, Siglec-8 mAb or glycan ligand binding causes eosinophil apoptosis associated with reactive oxygen species (ROS) production. Most CD33-related Siglecs function as inhibitory receptors, but the ability of Siglec-8 to stimulate eosinophil ROS production and apoptosis suggests that Siglec-8 might instead function as an activating receptor. Objective: We sought to determine the role of IL-5 priming and identify the signaling molecules involved in Siglec-8 function for human eosinophils. Methods: We used an mAb and/or a multimeric synthetic sulfated sialoglycan ligand recognizing Siglec-8 in combination with integrin blocking antibodies, pharmacologic inhibitors, phosphoproteomics, and Western blot analysis to define the necessity of various proteins involved in Siglec-8 function for human eosinophils. Results: Cytokine priming was required to elicit the unanticipated finding that Siglec-8 engagement promotes rapid β2-integrin–dependent eosinophil adhesion. Also novel was the finding that this adhesion was necessary for subsequent ROS production and apoptosis. Siglec-8–mediated ROS was generated through reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activation because pretreatment of eosinophils with catalase (an extracellular superoxide scavenger) or NSC 23766 (a Rac GTPase inhibitor) completely inhibited Siglec-8–mediated eosinophil apoptosis. Finally, engagement of Siglec-8 on IL-5–primed eosinophils resulted in increased phosphorylation of Akt, p38, and c-Jun N-terminal kinase 1 that was also β2-integrin dependent; pharmacologic inhibition of these kinases completely prevented Siglec-8–mediated eosinophil apoptosis. Conclusions: These data demonstrate that Siglec-8 functions uniquely as an activating receptor on IL-5–primed eosinophils through a novel pathway involving regulation of β2-integrin–dependent adhesion, NADPH oxidase, and a subset of protein kinases.",
keywords = "Akt, Eosinophil, Siglec-8, apoptosis, c-Jun N-terminal kinase (JNK), p38, phosphoproteomics, reduced nicotinamide adenine dinucleotide phosphate oxidase (NADPH), β-integrin",
author = "Carroll, {Daniela J.} and O'Sullivan, {Jeremy Alastair} and Nix, {David B.} and Yun Cao and Michael Tiemeyer and Bochner, {Bruce Scott}",
year = "2018",
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day = "1",
doi = "10.1016/j.jaci.2017.08.013",
language = "English (US)",
volume = "141",
pages = "2196--2207",
journal = "Journal of Allergy and Clinical Immunology",
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}

Sialic acid–binding immunoglobulin-like lectin 8 (Siglec-8) is an activating receptor mediating β2-integrin–dependent function in human eosinophils. / Carroll, Daniela J.; O'Sullivan, Jeremy Alastair; Nix, David B.; Cao, Yun; Tiemeyer, Michael; Bochner, Bruce Scott.

In: Journal of Allergy and Clinical Immunology, Vol. 141, No. 6, 01.06.2018, p. 2196-2207.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Sialic acid–binding immunoglobulin-like lectin 8 (Siglec-8) is an activating receptor mediating β2-integrin–dependent function in human eosinophils

AU - Carroll, Daniela J.

AU - O'Sullivan, Jeremy Alastair

AU - Nix, David B.

AU - Cao, Yun

AU - Tiemeyer, Michael

AU - Bochner, Bruce Scott

PY - 2018/6/1

Y1 - 2018/6/1

N2 - Background: Siglec-8 is a CD33 subfamily cell-surface receptor selectively expressed on human eosinophils. After cytokine priming, Siglec-8 mAb or glycan ligand binding causes eosinophil apoptosis associated with reactive oxygen species (ROS) production. Most CD33-related Siglecs function as inhibitory receptors, but the ability of Siglec-8 to stimulate eosinophil ROS production and apoptosis suggests that Siglec-8 might instead function as an activating receptor. Objective: We sought to determine the role of IL-5 priming and identify the signaling molecules involved in Siglec-8 function for human eosinophils. Methods: We used an mAb and/or a multimeric synthetic sulfated sialoglycan ligand recognizing Siglec-8 in combination with integrin blocking antibodies, pharmacologic inhibitors, phosphoproteomics, and Western blot analysis to define the necessity of various proteins involved in Siglec-8 function for human eosinophils. Results: Cytokine priming was required to elicit the unanticipated finding that Siglec-8 engagement promotes rapid β2-integrin–dependent eosinophil adhesion. Also novel was the finding that this adhesion was necessary for subsequent ROS production and apoptosis. Siglec-8–mediated ROS was generated through reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activation because pretreatment of eosinophils with catalase (an extracellular superoxide scavenger) or NSC 23766 (a Rac GTPase inhibitor) completely inhibited Siglec-8–mediated eosinophil apoptosis. Finally, engagement of Siglec-8 on IL-5–primed eosinophils resulted in increased phosphorylation of Akt, p38, and c-Jun N-terminal kinase 1 that was also β2-integrin dependent; pharmacologic inhibition of these kinases completely prevented Siglec-8–mediated eosinophil apoptosis. Conclusions: These data demonstrate that Siglec-8 functions uniquely as an activating receptor on IL-5–primed eosinophils through a novel pathway involving regulation of β2-integrin–dependent adhesion, NADPH oxidase, and a subset of protein kinases.

AB - Background: Siglec-8 is a CD33 subfamily cell-surface receptor selectively expressed on human eosinophils. After cytokine priming, Siglec-8 mAb or glycan ligand binding causes eosinophil apoptosis associated with reactive oxygen species (ROS) production. Most CD33-related Siglecs function as inhibitory receptors, but the ability of Siglec-8 to stimulate eosinophil ROS production and apoptosis suggests that Siglec-8 might instead function as an activating receptor. Objective: We sought to determine the role of IL-5 priming and identify the signaling molecules involved in Siglec-8 function for human eosinophils. Methods: We used an mAb and/or a multimeric synthetic sulfated sialoglycan ligand recognizing Siglec-8 in combination with integrin blocking antibodies, pharmacologic inhibitors, phosphoproteomics, and Western blot analysis to define the necessity of various proteins involved in Siglec-8 function for human eosinophils. Results: Cytokine priming was required to elicit the unanticipated finding that Siglec-8 engagement promotes rapid β2-integrin–dependent eosinophil adhesion. Also novel was the finding that this adhesion was necessary for subsequent ROS production and apoptosis. Siglec-8–mediated ROS was generated through reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activation because pretreatment of eosinophils with catalase (an extracellular superoxide scavenger) or NSC 23766 (a Rac GTPase inhibitor) completely inhibited Siglec-8–mediated eosinophil apoptosis. Finally, engagement of Siglec-8 on IL-5–primed eosinophils resulted in increased phosphorylation of Akt, p38, and c-Jun N-terminal kinase 1 that was also β2-integrin dependent; pharmacologic inhibition of these kinases completely prevented Siglec-8–mediated eosinophil apoptosis. Conclusions: These data demonstrate that Siglec-8 functions uniquely as an activating receptor on IL-5–primed eosinophils through a novel pathway involving regulation of β2-integrin–dependent adhesion, NADPH oxidase, and a subset of protein kinases.

KW - Akt

KW - Eosinophil

KW - Siglec-8

KW - apoptosis

KW - c-Jun N-terminal kinase (JNK)

KW - p38

KW - phosphoproteomics

KW - reduced nicotinamide adenine dinucleotide phosphate oxidase (NADPH)

KW - β-integrin

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U2 - 10.1016/j.jaci.2017.08.013

DO - 10.1016/j.jaci.2017.08.013

M3 - Article

VL - 141

SP - 2196

EP - 2207

JO - Journal of Allergy and Clinical Immunology

JF - Journal of Allergy and Clinical Immunology

SN - 0091-6749

IS - 6

ER -