HB4C5 is a human antibody producing plasma B cell line that expresses the recombination activating gene-1 (RAG-1) and RAG-2 constitutively, but undergoes few secondary immunoglobulin gene rearrangements when cultured in fetal bovine serum-containing medium. Here, we found that depletion of serum from the culture media induces secondary V(λ)J(λ) rearrangement in this cell line. To investigate the induction mechanism of secondary V(λ)J(λ) rearrangement, we assessed the expression levels of RAG-1 and RAG-2 products, V(λ) germline transcription level and the amount of V(λ) signal broken ends (SBE) in HB4C5 cells cultured in serum-supplemented or serum-free medium. Western-blot analysis showed that the expression level for the RAG-1 and RAG- 2 proteins was not affected by the serum depletion. V(λ) germline transcript was found to be constitutively expressed in HB4C5 cell line and this transcription level was not affected by the lack of serum. On the other hand, the amount of V(λ) SBE was shown to be increased in HB4C5 cells cultured in serum-free medium, suggesting that this increased formation of V(λ) SBE at least partly contributed to the enhanced occurrence of secondary V(λ)J(λ) rearrangement in HB4C5 cells cultured in the serum-free condition. These results indicate that expression of RAG proteins and V(λ) germline transcription is not enough to undergo secondary V(λ)J(λ) rearrangement in this cell line.
All Science Journal Classification (ASJC) codes
- Molecular Biology