Tyrosine phosphorylation in human neutrophil

Area of Science:

• Immunology
• Cell Signaling
• Biochemistry

Background:

• Protein tyrosine phosphorylation is a key regulatory mechanism in cellular signaling.
• Human neutrophils play critical roles in immune responses.
• Granulocyte-macrophage colony-stimulating factor (GM-CSF) and chemotactic factors modulate neutrophil functions.

Purpose of the Study:

• To investigate the role of protein tyrosine phosphorylation in human neutrophil activation by GM-CSF and fMet-Leu-Phe.
• To identify specific proteins involved in these signaling pathways.
• To explore the involvement of G-proteins and potential receptor tyrosine kinase activity.

Main Methods:

• Immunoblotting using phosphotyrosine-specific antibodies.
• Treatment of neutrophils with GM-CSF, fMet-Leu-Phe, and phorbol esters.
• Use of specific tyrosine kinase inhibitors (ST 638) and pertussis toxin.
• Measurement of superoxide production and actin polymerization.

Main Results:

• GM-CSF and fMet-Leu-Phe rapidly increased tyrosine phosphorylation of several neutrophil proteins, including p40 and p54.
• These increases were inhibited by the tyrosine kinase inhibitor ST 638 and, for some proteins, by pertussis toxin.
• The p40 protein was identified as likely Gi alpha 2, and p78/p92 as potential GM-CSF receptor subunits.
• ST 638 also inhibited fMet-Leu-Phe-induced superoxide production.

Conclusions:

• GM-CSF and fMet-Leu-Phe responses in neutrophils are partly mediated by tyrosine phosphorylation.
• The GM-CSF receptor appears coupled to a pertussis toxin-sensitive G-protein.
• The findings suggest the GM-CSF receptor may possess intrinsic tyrosine kinase activity.