Fig. 6.

Inhibition of the PLCγ pathway impairs IL-7-induced mTOR activation. a PLCγ1 and PLCγ2 are phosphorylated upon IL-7 stimulation. Pro-B cells derived from Rag1-deficient mice were stimulated with IL-7. Cell lysates were subjected to western blot analysis (upper). Cell lysates were immunoprecipitated (IP) with anti-PLCγ2 antibodies and precipitated proteins were immunoblotted with the indicated antibodies (lower). b IL-7 stimulation activates the mTOR pathway. Pro-B cells derived from Rag1-deficient mice were stimulated with IL-7 and cell lysates were subjected to western blot analysis. c PLCγ1/PLCγ2 double deficiency impairs mTOR activation. YFP⁺CD45.2⁺ pre-pro-B cells (fraction A) were sorted from the indicated BM transplantation recipients and cell lysates were subjected to western blot analysis. d PLCγ1/PLCγ2 double deficiency does not affect Stat5 activation. BM cells from the indicated recipients were stained with anti-B220 and anti-CD43 antibodies, followed by intracellular staining with anti-phosphor-Stat5 (pStat5) antibodies or isotype controls. Numbers indicate the mean fluorescent intensity (MFI) of B220⁺CD43⁺ cells of pStat5 (black) or isotype controls (gray) staining. e, f Inhibition of the PLCγ pathway or Jak3 impairs IL-7-induced activation of mTOR. Pro-B cells derived from Rag1-deficient BM were pre-treated with DMSO, U73122, edelfosine, or CP-690550; and then stimulated with IL-7. Cell lysates were subjected to western blot analysis. The number beneath each band in the western blot indicates the relative intensity of the corresponding band. Data shown are representative of 3 (a, b, d), 4 (c), 5 (e), and 3 (f) independent experiments