Figure 1

SB1518 effectively blocks signaling pathways in JAK2^WT-expressing cell lines. (a) 293T (b) Jurkat and (c) Karpas 1106P cells were pretreated with SB1518 for 3 h and 293T, and Jurkat cells were treated for 10 min with 100 ng/ml interferon-γ as indicated. After lysis, pJAK2 (Y1007/8) was detected by immunoblotting (IB) with anti-p(1007/8)JAK2 antibody. pSTAT1, pSTAT3 and pSTAT5 were detected by IB with anti-p(Y701)STAT1, anti-p(Y705)STAT3 or anti-p(694)STAT5 antibody, respectively. As a loading control, the same membranes were re-probed with anti-JAK2, anti-STAT1 or anti-actin antibody. (d) Jurkat cells were pre-treated with SB1518 for 3 h and were treated with 5 ng/ml pervanadate for 10 min as indicated. After lysis, JAK1 and JAK2 were immunoprecipitated and their phosphorylation status was determined by immunoblotting using phospho-specific antibodies. The same membranes were re-probed with anti-JAK1/or JAK2 antibody to detect the total protein levels.