Human immunoglobulin selection associated with class switch and possible tolerogenic origins for Cδ class-switched B cells
J. Clin. Invest. Nai-Ying Zheng, et al. 113:1188 doi:10.1172/JCI20255 [
Go to this article.]
Figure 5Analysis of the VJ
κ repertoire and configuration of the V
κ loci for evidence of receptor editing. (A) Mean ± SD of V
κ and J
κ gene segments used at significantly different frequencies between naive (four donors), Cδ-CS (five donors), and combined total memory (two donors) plus IgG
+ memory (two donors). (B) Semi-quantitative PCR was used to determine if increased V
κ4-1 usage provides evidence of receptor editing for Cδ-CS cells. V
κ4-1 is the most J
κ-proximal V
κ gene and is inverted. Thus, primary V
κ4-1 rearrangements retain the RSS junction (left), but following any other primary inversional rearrangement, a secondary (editing) V
κ4-1 rearrangement will delete the RSS junction (right). Thus, analysis of the genetic configuration of V
κ4-1–to-J
κ2 rearrangements provides a measure of receptor editing for all V
κ genes. The relative proportion of primary versus editing rearrangements was determined by PCR using the primer sets indicated to get the ratio of V
κ4-1–to-J
κ2 rearrangements (primers: V
κ4J
κ2-P1, blue arrows; and V
κ4J
κ2-P2, green arrows) compared to RSS junctions (RSS-P1, red arrows; and RSS-P2, orange arrows). (C) Genomic DNA template from naive, Cδ-CS, and memory B cells were normalized for V
κ4-1–to-J
κ2 rearrangements (Rearr.); then the relative proportion of RSS junctions were determined by PCR at three dilutions of template (wedges). There was ten-fold less V
κ4-1–to-J
κ2 RSS junctions for the Cδ-CS population, indicating that the increased V
κ4-1 rearrangements for this population are due to receptor editing. (–), buffer controls.
