Fig 2: Adoptively transferred engineered B cells enable memory retention upon immunization.a ELISA of sera collected 14 days following either prime or boost immunization, quantified using an anti-idiotypic antibody to 3BNC117. ####pv < 0.0001, #pv = 0.0465 for two-way ANOVA and **pv = 0.0024, ***pv = 0.0003 for Tukey’s multiple comparison (n = 3, each dot represents a mouse). b Analysis by flow cytometry of CD45.1 expression and gp120 binding in the GCs of mice following prime or boost immunizations, gating on live, singlets, B220+, GL-7+. c Quantitation of b. ####pv < 0.0001, ###pv = 0.0003 for two-way ANOVA and ***pv = 0.0008 and **upper = pv = 0.0095 and **lower = pv = 0.0067 for Tukey’s multiple comparison (n = 6, each dot represents a mouse). d, e Analysis by flow cytometry of CD38 or CD138 expression among donor derived cells in the spleens of recipient mice after prime or boost immunizations by the gp120 antigens from either the THRO4156.18 (THRO, Red) or the YU2.DG (YU2, Blue) HIV strains, gated on live, singlets, CD45.1+. ###pv = 0.0003, ##pv = 0.0044, #(D) = pv = 0.0338, #(E) = pv = 0.0125, for two-way ANOVA and **pv = 0.0012, *(D) = pv = 0.0222, *(E) = pv = 0.0143, Tukey’s multiple comparison (n = 3, each dot represents a mouse). For gating strategy see Supplementary Fig. 12.

Fig 3: LSD1 loss promotes intratumoral CD8+ T cell persistence.a Experimental design of administering FTY720 to block T cell egress. b Representative flow plots showing frequencies of CD4+ and CD8+ T cells in PBMCs of mice receiving FTY720 or vehicle. c Cell numbers of CD8+ TILs on day 22. d Tumor growth curves of WT and Cd4-Cre+Lsd1f/f (cKO) mice implanted with MC38 tumors and treated with FTY720 or vehicle. e Experimental design of CD8+ TILs co-transfer and assessment of intratumoral persistence. f Frequencies of CD45.1+ wildtype and CD45.2+ LSD1-deficient cells among transferred TCRß+CD8+ TILs (n = 3 per group). Data are pooled from two experiments and presented as mean ± SEM (c, d, f). Sample sizes are as indicated. Statistical significance was determined by two-sided unpaired t test (c, f) or two-way ANOVA (d). Source data are provided as a Source data file.

Fig 4: Cellular and molecular characterization of MMc in the offspring’s brain.a Experimental set-up of the mating strategies developed to identify MMc in brain based on surface expression of CD45.2 and H-2Db/b in CD45.2/1 and H-2Db/d offspring (left) and based on tdTomato-fluorescent reporter-expression (right). b Number of CD45.2 and H-2Db/b MMc among fetal brain cells on E18.5 (n = 43) and P8 (n = 17), using the gating strategy shown in Supplementary Fig. 1a. c Number of tdTomato-expressing MMc among fetal brain cells on E18.5 (n = 7). d Representative contour plots used to identify tdTomato+ MMc in fetal brain by flow cytometry (left) and tdTomato negative control (right). e UMAP dimensionality reduction plot of scRNA-seq data set from 4305 MMc isolated from fetal brains (n = 21, pooled from 4 litters). The phenotypes of the nine transcriptionally distinct MMc clusters are marked by the color keys. f Dot plots revealing the expression of cluster-specific genes in the nine MMc clusters identified in (e), using the same color keys. The dot size indicates the percentage of expression, the shade represents the average expression within the respective cluster. Data are displayed as mean ± SEM in (b) and (c), each circle corresponds to one pup.

Fig 5: Following the clonal composition of individual GC responding to foreign antigen over days to weeks.(A) Schematic overview of experimental approach for NP-CGG immunization (imm), tamoxifen treatment, surgical implantation of the CLNW, and multiphoton intravital microscopy in CD45.2 Aicda-Cre-Confetti mice. Data represent one independent experiment with three mice each. (B) Frequency of most dominant color for each individual GC as observed from 7 to 19 days post-imm in 3 mice. (C) Divergence Index for each individual GC as observed from 7 to 19 days post-imm in 3 mice. (D) Meander Index for each individual GC as observed from 7 to 19 days post-imm in 3 mice. (E) GC labeled E as in (B), (C), and (D) with pie chart to elaborate quantification below at each observation from days 7–19. (F) GC labeled F as in (B), (C), and (D) with pie chart to elaborate quantification below at each observation from days 7–19. Prominent clonal inversion observed from days 11–15. (G) GC labeled G as in (B), (C), and (D) with pie chart to elaborate quantification below at each observation from days 7–19. (H) GC labeled H as in (B), (C), and (D) with pie chart to elaborate quantification below at each observation from days 7–19. Double clonal inversion observed from days 11–13 and days 17–19. Please note that in E-H, representative images and quantification charts do not display 1:1 correspondence, since the latter are based on multiple z-planes throughout the volume of each of the GCs.10.7554/eLife.33051.015Figure 5—source data 1.NPCGG tracking experimental data for each observed GC to calculate relative frequency, dominance, relative frequency, and meander score.