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Recombination signal sequences restrict chromosomal V(D)J recombination beyond the 12/23 rule

Nature volume 405, pages 583–586 (2000)Cite this article

Abstract

The genes encoding the variable regions of lymphocyte antigen receptors are assembled from variable (V), diversity (D) and joining (J) gene segments1. V(D)J recombination is initiated by the recombinase activating gene (RAG)-1 and -2 proteins, which introduce DNA double-strand breaks between the V, D and J segments and their flanking recombination signal sequences (RSSs). Generally expressed DNA repair proteins then carry out the joining reaction2,3. The conserved heptamer and nonamer sequences of the RSSs are separated by non-conserved spacers of 12 or 23 base pairs (forming 12-RSSs and 23-RSSs). The 12/23 rule, which is mediated at the level of RAG-1/2 recognition and cutting4,5, specifies that V(D)J recombination occurs only between a gene segment flanked by a 12-RSS and one flanked by a 23-RSS1. Vβ segments are appended to DJβ rearrangements, with little or no direct Vβ to Jβ joining, despite 12/23 compatibility of Vβ 23-RSSs and Jβ12-RSSs6,7. Here we use embryonic stem cells and mice with a modified T-cell receptor (TCR)β locus containing only one Dβ (Dβ1) gene segment and one Jβ (Jβ1) gene cluster to show that the 5′ Dβ1 12-RSS, but not the Jβ1 12-RSSs, targets rearrangement of a diverse Vβ repertoire. This targeting is precise and position-independent. This additional restriction on V(D)J recombination has important implications for the regulation of variable region gene assembly and repertoire development.

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Figure 1: Generation of Jβ1ω/ω ES cells.
Figure 2: Vβ and Jβ repertoire in WT, Jβ1ω/ω and Jβ1M5/M5 mice.
Figure 3: Rearrangement of the Jβ1M4 allele.
Figure 4: Rearrangement analysis of Jβ1M3 and Jβ1M5 alleles.

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Acknowledgements

We thank O. Kanagawa and K. Murphy for advice and discussions and D. Fenoglio for assistance on cell sorting. This work is supported in part by the National Institutes of Health (F.W.A.). B.P.S. is a recipient of a Career Development Award from the Burroughs Wellcome Fund. C.H.B. was a fellow of the Irvington Institute for Immunological Research. F.G. is an associate of the Howard Hughes Medical Institute. F.W.A. is an investigator of the Howard Hughes Medical Institute.

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  1. Howard Hughes Medical Institute Children's Hospital and Department of Genetics, Harvard Medical School and The Center for Blood Research, Boston, 02115, Massachusetts , USA

    Craig H. Bassing, Frederick W. Alt, Margaux D'Auteuil, Barbara B. Woodman, Frank Gärtner & Laurie Davidson

  2. Washington University School of Medicine Department of Pathology and Immunology, 660 S. Euclid Ave., Campus Box 8118, St. Louis, 63110 , Missouri, USA

    Maureen M. Hughes, Tara D. Wehrly, J. Michael White & Barry P. Sleckman

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  1. Craig H. Bassing
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Correspondence to Barry P. Sleckman.

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Bassing, C., Alt, F., Hughes, M. et al. Recombination signal sequences restrict chromosomal V(D)J recombination beyond the 12/23 rule. Nature 405, 583–586 (2000). https://doi.org/10.1038/35014635

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