VH CDR3-dependent positive selection of murine VH12-expressing B cells in the neonate

Five to fifteen percent of peritoneal B1 (CD5⁺) cells from unmanipulated mice produce antibodies that bind bromelain-treated mouse red blood cells and the hapten phosphatidylcholine (PtC). The majority of these B cells express either of two V_H/V? gene combinations, V_H12/_V?4 or V_HH/V?9. Both the V_H11 and V_H12 genes are rearranged to J_H1 and encode third complementarity determining regions (CDR3) of restricted length and sequence. These and other observations argue strongly that PtC-specific Bl cells are antigen selected. To determine when selection of PtC-specific Bl cells begins in mice we have used the polymerase chain reaction to amplify V_H12-D-J_h1 rearrangements from livers of fetal and neonatal mice, and determined the CDR3 encoding sequences of individual clones. We find an unusually low ratio of productive (P) to non-productive (NP) rearrangements (0.4–1.0) at both developmental stages. P rearrangements in day 1 neonates are biased in D gene use and in the sequence and length of their deduced V_H CDR3. These biases are similar to those of PtC-specific B1 cells in the adult peritoneum. D gene use and CDR3 length and sequence are significantly less biased among V_H12 P rearrangements 2 to 3 days earlier in the day 18 fetal liver. We suggest that this rapid change in repertoire is due to positive ligand selection that is dependent on the sequence of V_H CDR3. We suggest further that the majority of V_H12-expressing cells are not ligand selected and consequently undergo programmed cell death. The evidence of restriction in day 1 neonatal livers and the low P/NP ratio in the fetus suggests that selection of V_H12-expressing cells begins before birth.