Supplementary Figure 7: BRWD1 regulates accessibility and nucleosome positioning.

(a) Nucleosome positioning at RSS and J_κs in WT and Brwd1^mut (Mut) small pre-B cells. Data are representative of two independent experiments. (b) Accessibility (open chromatin) and nucleosome positioning at 3ʹE_κ. Data are representative of two independent experiments. (c) DNA footprinting analysis of ±1 kb region for WT (red) and Brwd1^mut (blue) small pre-B cells at H3K9Ac, BRWD1+H3K9Ac, H3S10pK14Ac and BRWD1+H3S10pK14Ac peaks centered at 0. Nucleosome differential (upper panel) and accessibility (lower panel) were demonstrated. For nucleosome differential (Y-axis), values greater than 0 indicate the presence of a nucleosome, whereas values less than 0 tend to be nucleosome free. The X-axis is the distance in bp from the indicated peak or motif. (d) Alignment of BRWD1, H3K9Ac and H3S10pK14Ac enrichment at GAGA motifs. The Y-axis represents the normalized immunoprecipitation signal distribution for GAGA motifs centered at 0. (e) Position of short GAGAG (CTCTC) motifs in J_κ and E_κi regions. (f) Quantitative RT-PCR analysis of the expression of Rag1 and Rag2 in flow-sorted small pre-B cells from BM of WT and Brwd1^mut mice (n = 2). (g) Quantitative RT-PCR analysis of the expression of Tcfe2a (encodes E2A), Pax5, Ikzf1 (IKAROS), Irf4, Irf8, Smarca4 and Myc in flow-sorted small pre-B cells from BM of WT and Brwd1^mut mice (n = 3).