Fig. 8: PDCD4 restrains MZ B cell responses to NP-Ficoll immunization.

Reconstitution of WT (filled circles) and PDCD4 KO (open squares) μMT chimeras (a) or competitive chimeras (b) was analysed in blood samples 8-9 weeks after BM transfer. Contour plots represent detection of transferred CD45.2⁺ cells by flow cytometry. Graphs display numbers of CD45.2⁺ CD19⁺ B220⁺ B cells. c Chimeric mice were immunized i.v. with NP-Ficoll and analysed after 7 days. Contour plots and graphs depict percentage of CD138⁺ TACI⁺ CD19^int/low IgD^- CD45.2⁺ PBs of μMT chimeras (top) or competitive chimeras (bottom). Full gating strategy is reported in Supplementary Fig 9b–c. a–c n = 14–15 mice for μMT chimeras; n = 8 mice for competitive chimeras. d ELISpot-analysis of splenic NP20-IgM (left) and NP20-IgG3 (right) antibody-secreting cells (ASC) 7 days after NP-Ficoll immunization of μMT chimeric mice (n = 9–10 mice per group). Unimmunized WT μMT chimeric mice were used as a control (n = 3 mice). ASC numbers were calculated as proportion of 10⁶ MZ B cells. e Endpoint titers of NP20-IgM (left) and NP20-IgG3 (right) were measured in serum of μMT chimeric mice before (d-1) and after (d4, d7) NP-Ficoll immunization (n = 9–10 mice per group). a–e Graphs display data pooled from at least two independently performed experiments (mean ± SD). Two-tailed unpaired Student’s t-test analysis was performed between WT and PDCD4 KO samples. f Puromycin incorporation 30 min following administration to μMT chimeras (top) or competitive chimeras (bottom) 9 weeks after BM transfer. Overlay histograms indicate puromycin incorporation of WT (light grey) and PDCD4 KO (dark grey) MZ B cells. g Volcano plot of proteins quantified by mass spectrometry from FACS-sorted MZ B cells that were derived from WT or PDCD4 KO competitive chimeras. Vertical dashed lines indicate fold changes of 0.5. Horizontal dashed lines indicate p values of 0.05, which were calculated using a two-tailed t-test with unequal variance (n = 3 mice per group from two independently performed BM chimeras). Source data are provided as Source Data file.