Figure 6. SWAP-70 regulates IRF4 function during IL-21–induced plasma cell differentiation. (A) Purified B cells from indicated 8–10-wk-old female mice were stimulated with continuous IgM (cIgM) and CD40 in the presence or absence of 50 ng/ml IL-21. After 96 h of stimulation, prolifera- tion of B cells was measured by thymidine incorporation. The data are representative of three independent experiments. (B) Purified B cells were stimu- lated with cIgM and CD40 in presence or absence of IL-21 (5 or 50 ng/ml, as indicated). After 4 d of culture, in vitro plasma cell differentiation was measured by quantifying the percentage of CD138+ cells by FACS. The data are representative of three independent experiments. (C) Purified B cells were stimulated as in A. 72 h after culture, cells were harvested, and mRNA levels of Prdm1 were measured by real-time PCR. The data are representative of three independent experiments. (D) Purified B cells were stimulated as in A. At the times indicated, cells were harvested and mRNA levels of Prdm1 were measured by real-time PCR. The data are representative of two independent experiments. (E) Purified B cells from WT or S70/ female mice were stimu- lated as indicated. 48 h after stimulation, nuclear extracts were prepared and immunoprecipitated with an anti-IRF4 antibody (IRF4 IP) or with a control antibody (Ctrl IP). The immunoprecipitates were resolved by 8% SDS-PAGE, and then analyzed by Western blot using an anti–SWAP-70 antiserum (top). The blot was reprobed with an anti-IRF4 antibody (bottom). The data are representative of two independent experiments. (F) Purified B cells were stimu- lated for 48 h, as indicated in A. ChIP assays were performed with either an anti-IRF4 antibody or a control antibody. Real-time PCR for the IL-21 re- 590 IRF4 in autoimmunity | Biswas et al.