Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Article
  • Published:

Initiation of allelic exclusion by stochastic interaction of Tcrb alleles with repressive nuclear compartments

Nature Immunology volume 9, pages 802–809 (2008)Cite this article

Abstract

Studies of antigen-receptor loci have linked directed monoallelic association with pericentromeric heterochromatin to the initiation or maintenance of allelic exclusion. Here we provide evidence for a fundamentally different basis for T cell antigen receptor-β (Tcrb) allelic exclusion. Using three-dimensional immunofluorescence in situ hybridization, we found that germline Tcrb alleles associated stochastically and at high frequency with the nuclear lamina or with pericentromeric heterochromatin in developing thymocytes and that such interactions inhibited variable–to–diversity-joining (Vβ-to-DβJβ) recombination before β-selection. The introduction of an ectopic enhancer into Tcrb resulted in fewer such interactions and impaired allelic exclusion. We propose that initial Vβ-to-DβJβ recombination events are generally monoallelic in developing thymocytes because of frequent stochastic, rather than directed, interactions of Tcrb alleles with repressive nuclear compartments. Such interactions may be essential for Tcrb allelic exclusion.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Subnuclear localization of Tcrb alleles in Rag2−/− pro–B cells, Rag2−/− DN thymocytes and sorted wild-type DP thymocytes.
Figure 2: Subnuclear localization of Tcra alleles in Rag2−/− pro–B cells, Rag2−/− DN thymocytes and sorted wild-type DP thymocytes.
Figure 3: Colocalization of Tcrb, Tcra and Actb alleles with γ-satellite or lamin B1.
Figure 4: Distribution of nuclei with zero, one or two Tcrb alleles localized together with γ-satellite or lamin B1.
Figure 5: Subnuclear localization of rearranged and unrearranged Tcrb alleles.
Figure 6: Influence of Eα on subnuclear localization and allelic exclusion.

Similar content being viewed by others

References

  1. Bassing, C.H., Swat, W. & Alt, F.W. The mechanism and regulation of chromosomal V(D)J recombination. Cell 109, Suppl S45–S55 (2002).

    Article  CAS  PubMed  Google Scholar 

  2. Michie, A.M. & Zuniga-Pflucker, J.C. Regulation of thymocyte differentiation: pre-TCR signals and β-selection. Semin. Immunol. 14, 311–323 (2002).

    Article  CAS  PubMed  Google Scholar 

  3. Khor, B. & Sleckman, B.P. Allelic exclusion at the TCRβ locus. Curr. Opin. Immunol. 14, 230–234 (2002).

    Article  CAS  PubMed  Google Scholar 

  4. Mostoslavsky, R., Alt, F.W. & Rajewsky, K. The lingering enigma of the allelic exclusion mechanism. Cell 118, 539–544 (2004).

    Article  CAS  PubMed  Google Scholar 

  5. Jackson, A.M. & Krangel, M.S. Turning T-cell receptor β recombination on and off: more questions than answers. Immunol. Rev. 209, 129–141 (2006).

    Article  PubMed  Google Scholar 

  6. Jung, D., Giallourakis, C., Mostoslavsky, R. & Alt, F.W. Mechanism and control of V(D)J recombination at the immunoglobulin heavy chain locus. Annu. Rev. Immunol. 24, 541–570 (2006).

    Article  CAS  PubMed  Google Scholar 

  7. Tripathi, R., Jackson, A. & Krangel, M.S. A change in the structure of Vβ chromatin associated with TCR β allelic exclusion. J. Immunol. 168, 2316–2324 (2002).

    Article  CAS  PubMed  Google Scholar 

  8. Jackson, A., Kondilis, H.D., Khor, B., Sleckman, B.P. & Krangel, M.S. Regulation of T cell receptor-β allelic exclusion at a level beyond accessibility. Nat. Immunol. 6, 189–197 (2005).

    Article  CAS  PubMed  Google Scholar 

  9. Agata, Y. et al. Regulation of T cell receptor β gene rearrangements and allelic exclusion by the helix-loop-helix protein, E47. Immunity 27, 871–884 (2007).

    Article  CAS  PubMed  Google Scholar 

  10. Skok, J.A. et al. Reversible contraction by looping of the Tcra and Tcrb loci in rearranging thymocytes. Nat. Immunol. 8, 378–387 (2007).

    Article  CAS  PubMed  Google Scholar 

  11. Mostoslavsky, R. κ chain monoallelic demethylation and the establishment of allelic exclusion. Genes Dev. 12, 1801–1811 (1998).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  12. Mostoslavsky, R. et al. Asynchronous replication and allelic exclusion in the immune system. Nature 414, 221–225 (2001).

    Article  CAS  PubMed  Google Scholar 

  13. Schlissel, M. Allelic exclusion of immunoglobulin gene rearrangement and expression: why and how? Semin. Immunol. 14, 207–212 (2002).

    Article  CAS  PubMed  Google Scholar 

  14. Liang, H.E., Hsu, L.Y., Cado, D. & Schlissel, M.S. Variegated transcriptional activation of the immunoglobulin κ locus in pre-B cells contributes to the allelic exclusion of light-chain expression. Cell 118, 19–29 (2004).

    Article  CAS  PubMed  Google Scholar 

  15. Liang, H.E. et al. The “dispensable” portion of RAG2 is necessary for efficient V-to-DJ rearrangement during B and T cell development. Immunity 17, 639–651 (2002).

    Article  CAS  PubMed  Google Scholar 

  16. Brown, C.R. & Silver, P.A. Pore-ing the right dose. Nat. Cell Biol. 8, 430–431 (2006).

    Article  CAS  PubMed  Google Scholar 

  17. Kosak, S.T. & Groudine, M. Form follows function: The genomic organization of cellular differentiation. Genes Dev. 18, 1371–1384 (2004).

    Article  CAS  PubMed  Google Scholar 

  18. Pickersgill, H. et al. Characterization of the Drosophila melanogaster genome at the nuclear lamina. Nat. Genet. 38, 1005–1014 (2006).

    Article  CAS  PubMed  Google Scholar 

  19. Misteli, T. Beyond the sequence: cellular organization of genome function. Cell 128, 787–800 (2007).

    Article  CAS  PubMed  Google Scholar 

  20. Schneider, R. & Grosschedl, R. Dynamics and interplay of nuclear architecture, genome organization, and gene expression. Genes Dev. 21, 3027–3043 (2007).

    Article  CAS  PubMed  Google Scholar 

  21. Hewitt, S.L., High, F.A., Reiner, S.L., Fisher, A.G. & Merkenschlager, M. Nuclear repositioning marks the selective exclusion of lineage-inappropriate transcription factor loci during T helper cell differentiation. Eur. J. Immunol. 34, 3604–3613 (2004).

    Article  CAS  PubMed  Google Scholar 

  22. Zink, D. et al. Transcription-dependent spatial arrangements of CFTR and adjacent genes in human cell nuclei. J. Cell Biol. 166, 815–825 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Williams, R.R. et al. Neural induction promotes large-scale chromatin reorganisation of the Mash1 locus. J. Cell Sci. 119, 132–140 (2006).

    Article  CAS  PubMed  Google Scholar 

  24. Reddy, K.L., Zullo, J.M., Bertolino, E. & Singh, H. Transcriptional repression mediated by repositioning of genes to the nuclear lamina. Nature 452, 243–247 (2008).

    Article  CAS  PubMed  Google Scholar 

  25. Kosak, S.T. et al. Subnuclear compartmentalization of immunoglobulin loci during lymphocyte development. Science 296, 158–162 (2002).

    Article  CAS  PubMed  Google Scholar 

  26. Fuxa, M. et al. Pax5 induces V-to-DJ rearrangements and locus contraction of the immunoglobulin heavy-chain gene. Genes Dev. 18, 411–422 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Yang, Q., Riblet, R. & Schildkraut, C.L. Sites that direct nuclear compartmentalization are near the 5′ end of the mouse immunoglobulin heavy-chain locus. Mol. Cell. Biol. 25, 6021–6030 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Fisher, A.G. & Merkenschlager, M. Gene silencing, cell fate and nuclear organisation. Curr. Opin. Genet. Dev. 12, 193–197 (2002).

    Article  CAS  PubMed  Google Scholar 

  29. Su, R.C., Sridharan, R. & Smale, S.T. Assembly of silent chromatin during thymocyte development. Semin. Immunol. 17, 129–140 (2005).

    Article  CAS  PubMed  Google Scholar 

  30. Brown, K.E. et al. Association of transcriptionally silent genes with Ikaros complexes at centromeric heterochromatin. Cell 91, 845–854 (1997).

    Article  CAS  PubMed  Google Scholar 

  31. Brown, K.E., Baxter, J., Graf, D., Merkenschlager, M. & Fisher, A.G. Dynamic repositioning of genes in the nucleus of lymphocytes preparing for cell division. Mol. Cell 3, 207–217 (1999).

    Article  CAS  PubMed  Google Scholar 

  32. Skok, J.A. et al. Nonequivalent nuclear location of immunoglobulin alleles in B lymphocytes. Nat. Immunol. 2, 848–854 (2001).

    Article  CAS  PubMed  Google Scholar 

  33. Roldan, E. et al. Locus 'decontraction' and centromeric recruitment contribute to allelic exclusion of the immunoglobulin heavy-chain gene. Nat. Immunol. 6, 31–41 (2005).

    Article  CAS  PubMed  Google Scholar 

  34. Goldmit, M. et al. Epigenetic ontogeny of the Igk locus during B cell development. Nat. Immunol. 6, 198–203 (2005).

    Article  CAS  PubMed  Google Scholar 

  35. Krangel, M.S., Carabana, J., Abbarategui, I., Schlimgen, R. & Hawwari, A. Enforcing order within a complex locus: current perspectives on the control of V(D)J recombination at the murine T-cell receptor α/δ locus. Immunol. Rev. 200, 224–232 (2004).

    Article  CAS  PubMed  Google Scholar 

  36. Zhang, W. et al. Essential role of LAT in T cell development. Immunity 10, 323–332 (1999).

    Article  CAS  PubMed  Google Scholar 

  37. Jackson, A.M. & Krangel, M.S. A role for MAPK in feedback inhibition of Tcrb recombination. J. Immunol. 176, 6824–6830 (2006).

    Article  CAS  PubMed  Google Scholar 

  38. Francastel, C., Walters, M.C., Groudine, M. & Martin, D.I. A functional enhancer suppresses silencing of a transgene and prevents its localization close to centrometric heterochromatin. Cell 99, 259–269 (1999).

    Article  CAS  PubMed  Google Scholar 

  39. Ragoczy, T., Telling, A., Sawado, T., Groudine, M. & Kosak, S.T. A genetic analysis of chromosome territory looping: diverse roles for distal regulatory elements. Chromosome Res. 11, 513–525 (2003).

    Article  CAS  PubMed  Google Scholar 

  40. Ragoczy, T., Bender, M.A., Telling, A., Byron, R. & Groudine, M. The locus control region is required for association of the murine β-globin locus with engaged transcription factories during erythroid maturation. Genes Dev. 20, 1447–1457 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Khor, B. & Sleckman, B.P. Intra- and inter-allelic ordering of T cell receptor β-chain gene assembly. Eur. J. Immunol. 35, 964–970 (2005).

    Article  CAS  PubMed  Google Scholar 

  42. Shinkai, Y. et al. RAG-2-deficient mice lack mature lymphocytes owing to inability to initiate V(D)J rearrangement. Cell 68, 855–867 (1992).

    Article  CAS  PubMed  Google Scholar 

  43. Sleckman, B.P., Bardon, C.G., Ferrini, R., Davidson, L. & Alt, F.W. Function of the TCR α enhancer in αβ and γδ T cells. Immunity 7, 505–515 (1997).

    Article  CAS  PubMed  Google Scholar 

  44. Smithson, G., Medina, K., Ponting, I. & Kincade, P.W. Estrogen suppresses stromal cell-dependent lymphopoiesis in culture. J. Immunol. 155, 3409–3417 (1995).

    CAS  PubMed  Google Scholar 

  45. Solovei, I. et al. Spatial preservation of nuclear chromatin architecture during three-dimensional fluorescence in situ hybridization (3D-FISH). Exp. Cell Res. 276, 10–23 (2002).

    Article  CAS  PubMed  Google Scholar 

  46. Bertolino, E. et al. Regulation of interleukin 7-dependent immunoglobulin heavy-chain variable gene rearrangements by transcription factor STAT5. Nat. Immunol. 6, 836–843 (2005).

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We thank B. Sleckman and Y. Zhuang for comments on the manuscript. Supported by the National Institutes of Health (GM41052 and AI49934 to M.S.K.) and the Howard Hughes Medical Institute (H.S.).

Author information

Author notes

  1. Ryan J Schlimgen and Karen L Reddy: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Immunology, Duke University Medical Center, Durham, 27710, North Carolina, USA

    Ryan J Schlimgen & Michael S Krangel

  2. Department of Molecular Genetics and Cell Biology, Howard Hughes Medical Institute, The University of Chicago, Chicago, 60637, Illinois, USA

    Karen L Reddy & Harinder Singh

Authors
  1. Ryan J Schlimgen
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Karen L Reddy
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Harinder Singh
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Michael S Krangel
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

R.J.S. and K.L.R. did all experiments; M.S.K. and H.S. supervised the work; and R.J.S., K.L.R., H.S. and M.S.K. prepared the manuscript.

Corresponding authors

Correspondence to Harinder Singh or Michael S Krangel.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–3 and Supplementary Tables 1–3 (PDF 350 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Schlimgen, R., Reddy, K., Singh, H. et al. Initiation of allelic exclusion by stochastic interaction of Tcrb alleles with repressive nuclear compartments. Nat Immunol 9, 802–809 (2008). https://doi.org/10.1038/ni.1624

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ni.1624

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing