The roles of the protein tyrosine kinases Pyk2 (also called RAFTK or CAK β) and Syk in the process of functional activation of human myeloid cells were examined. During granulocytic differentiation of HL-60 cells with dimethyl sulfoxide (DMSO), the amounts of Pyk2 and β2 integrin increased, whereas the amount of Syk was abundant before differentiation and did not change during differentiation. When the granulocytic cells were stimulated with N-formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP), tyrosine phosphorylation of Pyk2 occurred promptly and subsequent association of Pyk2 with β2 integrin was detected. In contrast, Syk was not tyrosine phosphorylated by fMLP stimulation but constitutively associated with β2 integrin. Stimulation with fMLP also caused the alteration of β2 integrin to an activated form, a finding that was confirmed by the observation of fMLP-induced cell attachment on fibrinogen-coated dishes and inhibition of this attachment by pretreatment with anti-β2 integrin antibody. Cell attachment to fibrinogen caused the enhanced tyrosine phosphorylation of Pyk2 and the initial tyrosine phosphorylation of Syk, which was also inhibited by pretreatment with anti-β2 integrin antibody. In vitro kinase assays revealed that Pyk2 and Syk represented kinase activities to induce tyrosine phosphorylation of several molecules in the anti-β2 integrin immunoprecipitates of the attached cells. These results showed that Pyk2 is involved in the functional activation of granulocytic cells in 2 signaling pathways: an fMLP receptor-mediated 'inside-out' signaling pathway that might cause β2 integrin activation and a subsequent β2 integrin-mediated 'outside-in' signaling pathway. Syk was activated in relation to cell attachment to fibrinogen as a result of 'outside-in' signaling, although it was already associated with β2 integrin before fMLP stimulation. (C) 2000 by The American Society of Hematology.
|Number of pages||7|
|Publication status||Published - 01-09-2000|
All Science Journal Classification (ASJC) codes
- Cell Biology