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PMID: 8666673 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Deficiency of Src family kinases p59/61hck and p58c-fgr results in defective adhesion-dependent neutrophil functions.

The Journal of cell biology ·Vol. 133 ·No. 4 ·1996-05-00 ·Pages 895-910

Lowell CA, Fumagalli L, Berton G

Abstract

Cross-linking of the neutrophil-beta 2- or beta 3-related leukocyte response integrins by extracellular matrix (ECM) proteins or monoclonal antibodies (mAb) stimulates cytoskeletal rearrangement leading to cell spreading and respiratory burst. Tyrosin phosphorylation of a variety of proteins and activation of the Src family kinases within polymorphonuclear leukocytes (PMN) have recently been implicated in the intracellular signaling pathways generated by leukocyte integrins (Yan, S.R., L. Fumagalli, and G Berton. 1995. J. Inflammation. 45:217-311.) To directly test whether these functional responses are dependent on the Src family kinases p59/61hck and p58c-fgr, we examined adhesion-dependent respiratory burst in PMNs isolated from hck -/-, fgr -/-, and hck -/- fgr -/- knockout mice. Purified bone marrow PMNS from wild-type mice released significant amounts of O2- when adherent to fibrinogen-, fibronectin-, or collagen-coated surfaces, in the presence of activating agents such as tumor necrosis factor (TNF) or formyl-methionyl-leucyl-phenylalanine, as described for human PMNs. PMNs from hck-/-fgr-/- double-mutant mic, however, failed to respond. This defect was specific for integrin signaling, since respiratory burst was normal in hck-/-fgr-/-PMNs stimulated by immune complexes or PMA. Stimulation of respiratory burst was observed in TNF-primed wild-type PMN plated on surfaces coated with murine intracellular adhesion molecule-1 (ICAM-1), while hck-/-fgr-/- PMNs, failed to respond. Direct cross-linking of the subunits of beta 2 and beta 2 integrins by surface-bound mAbs was elicited O2- production by wild-type PMNs, while the double-mutant hck-/-fgr-/- cells failed to respond. Photomicroscopy and cell adhesion assays revealed that the impaired functional responses of hck-/-fgr-/- PMNs were caused by defective spreading and tight adhesion on either ECM protein- or mAb-coated surfaces. In contrast, hck-/-or fgr-/-single mutant cells produced O2- at levels equivalent to wild-type cells on ECM protein, murine ICAM-1, and antiintegrin mAb-coated surfaces. Hence, either p59/61 hck and p 58c-fgr is required for signaling through leukocyte beta 2 and beta 3 integrins leading to PMN spreading and respiratory burst. This is the first direct genetic evidence of the importance of Src family kinases in integrin signaling within leukocytes, and it is also the best example of overlapping function between members of this gene family within a defined signal transduction pathway.

MeSH Terms
Animals Antibodies, Monoclonal/immunology Antigens, CD/immunology Bone Marrow/immunology Bone Marrow Cells CD11 Antigens/immunology CD18 Antigens/immunology,pharmacology Cell Adhesion Cells, Cultured Fibrinogen Humans Integrin beta3 Integrins/immunology Intercellular Adhesion Molecule-1/immunology Mice Mice, Knockout Mice, Mutant Strains N-Formylmethionine Leucyl-Phenylalanine/pharmacology Neutrophils/immunology Platelet Membrane Glycoproteins/immunology Protein-Tyrosine Kinases/deficiency,physiology Proto-Oncogene Proteins/deficiency,physiology Proto-Oncogene Proteins c-hck Respiratory Burst Signal Transduction Superoxides/metabolism Tumor Necrosis Factor-alpha/pharmacology src-Family Kinases/physiology
Chemicals
Antibodies, Monoclonal Antigens, CD CD11 Antigens CD18 Antigens Integrin beta3 Integrins Platelet Membrane Glycoproteins Proto-Oncogene Proteins Tumor Necrosis Factor-alpha Superoxides Intercellular Adhesion Molecule-1 N-Formylmethionine Leucyl-Phenylalanine Fibrinogen Protein-Tyrosine Kinases HCK protein, human Hck protein, mouse Proto-Oncogene Proteins c-hck proto-oncogene proteins c-fgr src-Family Kinases
Authors & Affiliations
3 authors, click to expand affiliations / ORCID
Lowell C A
Department of Laboratory Medicine, University of California, San Francisco 94143, USA.
Fumagalli L
Berton G
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Article Info
Journal
The Journal of cell biology
Abbr.
J Cell Biol
ISSN
0021-9525
DOI
10.1083/jcb.133.4.895
Published
1996-05-00
Pages
895-910
Language
English
Region
United States
NLM ID
0375356
PMCID
PMC2120842
Subset
IM
Grants
NIDDK NIH HHS · DK50267 · United States
External Links
PubMed source DOI: 10.1083/jcb.133.4.895 PMC Full Text View journal details
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