Src-family kinases mediate an outside-in signal necessary for β2 integrins to achieve full activation and sustain firm adhesion of polymorphonuclear leucocytes tethered on E-selectin

Licia Totani, Antonio Piccoli, Stefano Manarini, Lorenzo Federico, Romina Pecce, Nicola Martelli, Chiara Cerletti, Paola Piccardoni, Clifford A. Lowell, Susan S. Smyth, Giorgio Berton, Virgilio Evangelista

Research output: Contribution to journalArticlepeer-review

Abstract

In cell suspensions subjected to high-shear rotatory motion, human PMN (polymorphonuclear cells) adhered to E-selectin-expressing CHO (Chinese-hamster ovary) cells (CHO-E), and formed homotypic aggregates when challenged by E-selectin-IgG fusion protein, by a mechanism that involved β2 integrins. Both heterotypic and homotypic PMN adhesion was accompanied by tyrosine phosphorylation of a 110 kDa protein (P110). This event was prevented by blocking anti-(β2 integrin) antibodies and by inhibitors of Src-family kinases, suggesting that it was part of an 'outside-in' signalling that was initiated by integrin engagement. Interestingly, Src-family kinase inhibitors prevented β2-integrin-mediated (i) homotypic PMN adhesion triggered by E-selectin-IgG, (ii) heterotypic CHO-E/PMN adhesion in mixed-cell suspensions, and (iii) firm adhesion of PMN to CHO-E monolayers under physiological flow. Similarly to PMN treated with Src-family kinase inhibitors, PMN from hck -/- fgr-/- and hck-/- fgr-/- lyn-/- mice showed significant impairment of β2-integrin- mediated adhesion to CHO-E. Moreover, the expression of β2 integrin activation epitopes at the sites of cell-cell contact in CHO-E/PMN conjugates was abolished by Src-family kinase inhibitors. One component of P110 was identified as the FAK (focal adhesion kinase) Pyk2 (proline-rich tyrosine kinase 2), which was phosphorylated in a β2 integrin- and Src-family-kinase- dependent manner. Thus, Src-family kinases, and perhaps Pyk2, mediate a signal necessary for β2 integrin function in PMN tethered by E-selectin.

Original languageEnglish
Pages (from-to)89-98
Number of pages10
JournalBiochemical Journal
Volume396
Issue number1
DOIs
Publication statusPublished - May 15 2006

Keywords

  • β2 integrin
  • Adhesion
  • Leucocyte recruitment
  • Polymorphonuclear leucocyte
  • Proline-rich tyrosine kinase 2 (Pyk2)
  • Src-family kinase

ASJC Scopus subject areas

  • Biochemistry

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