Extended Data Fig. 2: DNA damage induces interactions between shieldin and CST.
a) IgM-to-IgA CSR frequencies in Shld3 knockout CH12-F3 cell lines, n = 3 independent experiments, where each data point is the average of 3 technical replicates from a single experiment. Mean ± SEM. Representative flow cytometry plots for IgM-to-IgA CSR. Representative data, n > 3 independent experiments. b) IgM-to-IgA CSR frequencies in Shld1 knockout CH12-F3 cell lines, n = 3 independent experiments, where each data point is the average of 3 technical replicates from a single experiment. Mean ± SEM. Representative flow cytometry plots for IgM-to-IgA CSR. Representative data, n > 3 independent experiments. c) B cell Shld1 interactome as defined by LC–MS/MS and LFQ. Scatter plot depicts log2 fold-enrichment of indicated TwinStep–immunocomplexes across 2 independent experiments. d) B cell Shld1 IR-dependent interactome as defined by LC–MS/MS and LFQ. Scatter plot depicts log2 fold-enrichment of indicated TwinStep–immunocomplexes across 2 independent experiments. e) Western blot analysis of CH12-F3 cell extracts from pulldown experiments. Cells were pre-treated with ATM, ATR inhibitor or both for 1 hr prior to irradiation. Representative of n = 2 independent experiments. f) Western blot analysis of CH12-F3 cell extracts from pulldown experiments. Representative of n = 2 independent experiments. g) Splenic B cells cultured with the indicated stimuli (96 h) and stained for surface IgG1, IgE, IgG2b or IgG3. n = 3-5 mice per genotype. CSR 100%, mean immunoglobulin isotype switch frequency of 2 control animals in each experiment. Significance was determined by two-way ANOVA with Tukey’s correction. Mean ± SEM. h) CTV-labelled purified B cells were stimulated as indicated and stained for surface IgG on day 4. Representative of n = 2 experiments.
