Fig. 5
From: Initiation of HIV neutralizing B cell lineages with sequential envelope immunizations
Analysis of receptor editing in CH103 germ line knock-in mice. a–c Env reactivity in mature B cell compartments of CH103 unmutated common ancestor (UCA) double knock-in (dKI) mice. a Representative pseudocolored dot plots showing typical patterns of reduced TF Env+ mature (follicular) B cells in naive CH103 UCA dKI mice. Shown as controls for background TF Env staining are WT (B6) mice. b Graphical summaries of percentages of differential binding in transitional and mature compartments in naive CH103 UCA dKI mice. Each dot represents an individual animal. ***P < 0.001, two-tailed Student’s t test. c Regression analysis of developmental blockade severity and residual mature B cell fraction retaining Env specificity. d Loss of CH103 KI light chain (LC) rescues splenic mature B cell development and reverses anergic phenotype in CH103 UCA VHDJH ++ (“heavy-chain (HC)-only”) KI mice. **P < 0.005, two-tailed Student’s t test. e Progressively decreased differential binding of splenic transitional and mature B cells (means + SEM) in CH103 UCA dKI (n = 6), heterozygous (het) dKI (n = 5), and VHDJH +/+ KI (n = 4) mice. CH01 het dKI mice (n = 2), a model that undergoes no negative selection, and WT B6 mice (n = 4) are shown as positive and negative controls for Env binding, respectively. f, g Extensive and highly-restricted receptor editing of the CH103 λ3-20 LC by “master editor” Vκ5-39, as revealed in CH103 UCA het dKI mice. f Receptor editing in CH103 UCA het dKI mice. Shown are pie charts of HC/LC usage and breakdown of Vκ family and Jκ usage, among LCs paired to the KI VH4-59-bearing HC. Individual mature B cells from naive CH103 het dKI mice were obtained by single-cell sorting on the total (unselected for TF Env binding) mature follicular B cell repertoire, and VHDJH/VλJλ Ig gene pairs from single cells were recovered by RT-PCR for sequencing and immunogenetic analysis. g Polyreactivity/autoreactivity profiles of the CH103 UCA and Vκ5-39 editor M5808 monoclonal antibodies (mAbs) on human protoarrays. The >500-fold binding compared to control mAb, that is, the official autoreactivity “cutoff”, is indicated by dashed lines. Also shown are candidate autoantigens bound by the CH103 UCA mAb (circled in blue, in the red shaded box) and to which binding by M5808 is eliminated (green-shaded box)
