Extended Data Fig. 3: Iα deletion promotes CSR to upstream S regions.
From: Fundamental roles of chromatin loop extrusion in antibody class switching
a, Illustration of dominant, deletional CSR between Sμ and Sα in CH12F3 cells. b, Illustration of Cas9–gRNA targeting (lightning bolts) used to generate the CH12F3NCΔ line. c, Southern blot confirmation (using BamHI digestion and a JH4 probe) of the CH12F3NCΔ lines (done twice independently with similar results). d, Western blot confirmation of AID expression or lack of expression, respectively, in AID sufficient and deficient (via targeted deletion) CH12F3NCΔ and IαΔ lines following stimulation with anti-CD40–IL-4–TGFβ for 72 h (done twice independently with similar results). e, FACS analysis for surface IgA expression in CH12F3NCΔ AID−/− cells stimulated with anti-CD40–IL-4–TGFβ for 72 h (done three times independently with similar results). f, Three repeats of CSR-HTGTS-seq data shown in Fig. 2b for anti-CD40–IL-4–TGFβ-stimulated CH12F3NCΔ and IαΔ cells (three biologically independent repeats). Junctions are plotted at 2.5-kb bin size. The blue lines indicate deletional joining and the red lines indicate inversional joining. Bar graph shows percentages of junctions located in different regions from CH12F3NCΔ and IαΔ cells. Data are mean ± s.d. from three biologically independent repeats. P values were calculated via an unpaired two-tailed t-test. g, FACS analysis of IgA, IgG3 and IgG2b surface expression in CH12F3NCΔ and IαΔ cells stimulated with anti-CD40–IL-4–TGFβ for 72 h (four biologically independent repeats). Bar graph shows percentages of IgA, IgG3 and IgG2b expression on CH12F3NCΔ and IαΔ cells. Data are mean ± s.d. from four biologically independent repeats. P values were calculated via an unpaired two-tailed t-test.
