Involvement of Shc and Cbl-PI 3-kinase in Lyn-dependent proliferative signaling pathways for G-CSF

Granulocyte colony-stimulating factor (G-CSF) is the major hematopoietic fa ctor which controls the production and differentiation of granulocytes. The G-CSF receptor (G-CSFR) belongs to the superfamily of the cytokine recepto rs, which transduce signals via the activation of cytosolic protein tyrosin e kinases (PTK), To determine the role of specific PTK in G-CSF signaling w e expressed the human G-CSFR in cell lines derived from DT40 B cells, which lack either the Src-related Lyn or Syk, Wild-type (wt) and syk-deficient c ells underwent increased DNA synthesis in response to G-CSF; lyn-deficient cells did not. The purpose of these studies is to identify Lyn's downstream effectors in mediating DNA synthesis, While G-CSF stimulated Pas activity in all cell lines, G-CSF failed to induce the tyrosine phosphorylation of S hc in lyn-deficient cells. G-CSF induced a statistically significant activa tion of Erk1/Erk2 Kinase or p90Rsk only in the wt cells. G-CSF induced the tyrosine phosphorylation of Cbl and increased activity of PI 3-kinase in wi ld-type and syk-deficient, but non in lyn-deficient, cells. Inhibition of S hc by over-expression of its SH2 or PTB domains or PI 3-kinase by either tr eatment with wortmannin or expression of the CblY731F mutant decreased G-CS F-induced DNA synthesis, Thus, the Lyn, Cbl-PI 3-kinase, and Shc/non-Pas-de pendent pathways correlate with the ability of cells to respond to G-CSF wi th increased DNA synthesis.