title: Pyk2 and Syk participate in functional activation of granulocytic HL-60 cells in a different manner.
abstract: The roles of the protein tyrosine kinases Pyk2 (also called RAFTK or CAK beta) 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 beta2 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 beta2 integrin was detected. In contrast, Syk was not tyrosine phosphorylated by fMLP stimulation but constitutively associated with beta2 integrin. Stimulation with fMLP also caused the alteration of beta2 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-beta2 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-beta2 integrin antibody. In vitro kinase assays revealed that Pyk2 and Syk represented kinase activities to induce tyrosine phosphorylation of several molecules in the anti-beta2 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 beta2 integrin activation and a subsequent beta2 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 beta2 integrin before fMLP stimulation. (Blood. 2000;96:1733-1739)
authors.affiliation: Department of Hematology and Oncology, Graduate School of Medicine, Kyoto University, Kyoto, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Miura
authors.forename: Y
authors.initials: Y
authors.lastname: Tohyama
authors.forename: T
authors.initials: T
authors.lastname: Hishita
authors.forename: A
authors.initials: A
authors.lastname: Lala
authors.forename: E
authors.initials: E
authors.lastname: De Nardin
authors.forename: Y
authors.initials: Y
authors.lastname: Yoshida
authors.forename: H
authors.initials: H
authors.lastname: Yamamura
authors.forename: T
authors.initials: T
authors.lastname: Uchiyama
authors.forename: K
authors.initials: K
authors.lastname: Tohyama
mesh_terms: D018821:CD18 Antigens / Q000378:metabolism; D002448:Cell Adhesion / Q000187:drug effects; D002454:Cell Differentiation; D004792:Enzyme Precursors / Q000378:metabolism*; D005340:Fibrinogen / Q000378:metabolism; D051418:Focal Adhesion Kinase 2; D006098:Granulocytes / Q000166:cytology; D018922:HL-60 Cells / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D010766:Phosphorylation / Q000187:drug effects; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D000072377:Syk Kinase; D014443:Tyrosine / Q000378:metabolism
chemical_list: D018821:CD18 Antigens; D004792:Enzyme Precursors; D047908:Intracellular Signaling Peptides and Proteins; D014443:Tyrosine; D009240:N-Formylmethionine Leucyl-Phenylalanine; D005340:Fibrinogen; D011505:Protein-Tyrosine Kinases; D051418:Focal Adhesion Kinase 2; C000606896:SYK protein, human; D000072377:Syk Kinase