Supplementary Figure 3: BRWD1 in B cell development and immunoglobulin light chain recombination.

(a,b) LSK progenitors from WT (CD45.1) and WT (CD45.2) (a) or WT (CD45.1) and Brwd1^mut (CD45.2) (b) mice were flow sorted and mixed 1:1 and transplanted into sublethally irradiated (550 rad) Rag2^−/−Il2rg^−/− hosts. B cell development in spleen (a) and T cell development in thymi (b) were then analyzed by flow cytometry after 5 weeks of transplantation. Data are representative of four independent experiments. (c) Flow cytometric assay for apoptosis with annexin V and 7-AAD in B cell progenitors of different developmental stages isolated from WT and Brwd1^mut mice (n = 2). (d) Cell cycle analysis of B cell progenitors of different developmental stages isolated from WT and Brwd1^mut mice. Numbers above bracketed lines indicate the percentage of cells in S-G2M (n = 2). (e) Quantitative RT-PCR analysis of the expression of Ccnd2 and Ccnd3 in flow-sorted large pre-B cells from BM of WT and Brwd1^mut mice (n = 2). *P < 0.05 compared with respective controls (unpaired t-test). (f). Frequency (as a percentage) of J_κ usage in rearranged Igk in isolated small pre-B cells of WT and Brwd1^mut mice (n = 2). (g) Usage of V_κ gene family in small pre-B cells of WT and Brwd1^mut mice (n = 2). (h) Analysis of Igl-expressing immature B cells and splenic IgM-expressing B cells isolated from WT and Brwd1^mut mouse BM and spleen, respectively (n = 3).