Figure 5. BCR signaling-induced V_H replacement depends on Syk and Src kinase activation.

A) Western blot analyses of BCR signaling events upon treatment with different inhibitors. EU12 μHC⁺ cells (10⁷ cells/ml) were pretreated with medium alone, DMSO, Genistein (50 μg/ml), Syk II (5 μM) or Syk III (5 μM), or PP1 (5 μM) for 30 min and stimulated with F(ab')₂ goat anti-human μHC (2 μg/ml) for 3 min. Cell lysate was analyzed by Western blot using antibodies specific for the indicated BCR signaling components. Antibodies to β-ACTIN were used to control sample loading.

B) LM-PCR detection of DSBs at the V_H3 cRSS sites in EU12 μHC⁺ cells after treatment with different inhibitors, with or without BCR stimulation. The CD19 promoter region was amplified to monitor DNA input.

C) PCR detection of V_H1→V_H3 excision circles in EU12 μHC⁺ cells after treatment with different inhibitors, with or without BCR stimulation. The CD19 promoter region was amplified to monitor DNA input.

The results shown are representatives from more than three independent experiments.