Figure 5

PP2 and dasatinib inhibit BCR-induced LYN and JNK activation and EGR-1 upregulation. (A) Patients’ cells (UPN9) were pretreated with dasatinib (100nM) or SP600125 (10 μM) for 1 h and stimulated for 5 min or 15 min with soluble anti-IgM (10 μg/ml). Phospho-Tyr397 LYN was detected using a pan phospho-src family antibody. (B) The same experiment was done with PP2 (10 μM) on UPN 9 and UPN 13 under the same conditions of BCR stimulation for 10 min. Lines 1 and 2 have to be compared to evidence the effect of PP2 on the constitutive level of phosphorylation for Lyn. Similarly lines 3 and 4 reflect this effect upon BCR stimulation. (C) BCR-induced phospho-JNK (p54 and p46) was analyzed under treatment with dasatinib (100nM) or SP600125 (10 μM) used herein as a positive control of phospho-JNK inhibition. (D) Impact of dasatinib on BCR-induced EGR-1 expression. MCL cells were pretreated with various concentrations of dasatinib as indicated and stimulated with immobilized anti-IgM. EGR-1 mRNA and protein were analyzed by qRT-PCR at 1 h of stimulation (left panel) and western-blot at 3 h of stimulation (right panel). Relative mRNA expression was calculated compared with unstimulated cells.