Fig. 7. CTLA-4-deficient B-1a cells and their IgM^neg decedents induce Tfh cells and GCs in congenic recipients.

a Live CD4⁺ T cells and CD19⁺ B cells from spleens of indicated mice were gated to reveal Tfh (PD-1^hi ICOS⁺) and GC B (CD95^hi CD38^neg) cells, respectively. b Data summarizing the numbers of Tfh and GC B cells in spleens of indicated mouse groups is shown, n = 7–10 mice per group. Each dot represents data for an individual mouse, *p < 0.003, **p < 0.0002, ***p < 0.002, ****p < 0.0003, *****p < 0.004, ns, not significant (p < 0.2), nonparametric Wilcoxon one-way test. About 5 × 10⁶ of peritoneal B-1a (pB-1a) and 2 × 10⁵ of IgM^neg splenic B-1a (sB-1a) cells were transferred. c TCRβ CDR3 tree-map plots (upper) illustrate the CDR3 nucleotide sequences expressed by non-Tfh T and Tfh cells from indicated mice. One representative plot from four or six independent T-cell samples is shown. Each rectangle in a given tree-map represents a unique CDR3 nucleotide sequence and the size of each rectangle denotes the relative frequency of an individual CDR3 sequence. The box plot summarizes the D50 metric analysis that quantifies TCRβ CDR3 nucleotide sequence diversity for each T-cell sample from indicated mouse group. Each dot represents data for a T-cell sample from an individual mouse, n = 4–6 T-cell samples per group, *p < 0.006, ns, not significant (p < 0.2), nonparametric Wilcoxon one-way test. Low D50 values are associated with less TCRβ CDR3 diversity. Sequence information for each T-cell sample is summarized in supplementary Table 3. d Live myeloid^neg CD3^neg CD19⁺ splenic B cells from indicated CB.17 mice were gated to reveal GC B cells (CD19⁺CD95⁺GL7⁺), which were further gated to show IgG1, IgG1b and IgG1a surface expression. Statistical summary for five independent experiments is shown in Supplementary Table 5. Box plots in b, c: box draws 75% (upper), 50% (center line) and 25% (down) quartile, the maxima and minima outliers are shown as top and bottom line, respectively.