Figure 6: Inactivation of GSK3β by p38 MAPK-mediated phosphorylation on Ser³⁸⁹ is required for B cells to survive the DNA damage response triggered by CSR.

(a) Purified B cells from WT and GSK3β-KI mice (n=3) were activated for four days with LPS and IL-4 and the surface level of IgG1 determined by flow cytometry. (b) B cells from WT and GSK3β-KI mice (n=3) were activated for 4 days either in the presence or absence of the GSK3 inhibitor X (GSK-Inh) and the percent of IgG1 positive B cells was determined as in a. (c) B cells from WT and GSK3β-KI mice (n=3) were activated as described above either in the presence or absence of the p38 MAPK inhibitor SB203580 (SB) and the percent of IgG1 positive B cells was determined as in a. (d) B cells from WT (solid black line) and GSK3β-KI (grey filled) mice (n=3) were loaded with CFSE and activated for four days and CFSE staining was examined flow cytometry analysis. (e) B cells from WT and GSK3β-KI mice (n=4) were activated for three days and viability determined by trypan blue staining. Percentage of cell survival relative to activated WT B cells is provided (% relative to WT cells). (f) B cells from WT and GSK3β-KI mice (n=4) were activated for three days and cell death was measured by AnnexinV staining and flow cytometry analysis. Percentage of AnnexinV positive cells for each cell type is shown. (g) B cells from WT and GSK3β-KI mice (n=3) were activated for three days and stained for AnnexinV, B220 and IgG1. The percentage of AnnexinV+ cells in the B220+IgG+ and B220+IgG- populations are shown. *P value<0.05 as determined by t- test. Data are shown as mean±s.e.m. and are representative of three or more independent experiments.