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Activation-induced cytidine deaminase turns on somatic hypermutation in hybridomas

Abstract

The production of high-affinity protective antibodies requires somatic hypermutation (SHM) of the antibody variable (V)-region genes. SHM is characterized by a high frequency of point mutations that occur only during the centroblast stage of B-cell differentiation. Activation-induced cytidine deaminase (AID), which is expressed specifically in germinal-centre centroblasts1, is required for this process, but its exact role is unknown2. Here we show that AID is required for SHM in the centroblast-like Ramos cells, and that expression of AID is sufficient to induce SHM in hybridoma cells, which represent a later stage of B-cell differentiation that does not normally undergo SHM. In one hybridoma, mutations were exclusively in G·C base pairs that were mostly within RGYW or WRCY motifs, suggesting that AID has primary responsibility for mutations at these nucleotides. The activation of SHM in hybridomas indicates that AID does not require other centroblast-specific cofactors to induce SHM, suggesting either that it functions alone or that the factors it requires are expressed at other stages of B-cell differentiation.

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Figure 1: hAID expression induces SHM in non-mutating Ramos cells.
Figure 2: AID induces SHM in hybridomas.
Figure 3: Mutations observed in hybridoma clones.

References

  1. Muramatsu, M. et al. Specific expression of activation-induced cytidine deaminase (AID), a novel member of the RNA-editing deaminase family in germinal centre B cells. J. Biol. Chem. 274, 18470–18476 (1999)

    Article  CAS  Google Scholar 

  2. Kinoshita, K. & Honjo, T. Linking class-switch recombination with somatic hypermutation. Nature Rev. Mol. Cell Biol. 2, 493–503 (2001)

    Article  CAS  Google Scholar 

  3. Sale, J. E. & Neuberger, M. S. TdT-accessible breaks are scattered over the immunoglobulin V domain in a constitutively hypermutating B cell line. Immunity 9, 859–869 (1998)

    Article  CAS  Google Scholar 

  4. Denépoux, S. et al. Induction of somatic mutation in a human B cell line in vitro. Immunity 6, 35–46 (1997)

    Article  Google Scholar 

  5. Zan, H. et al. Induction of Ig somatic hypermutation and class switching in a human monoclonal IgM+ IgD+ B cell line in vitro: definition of the requirements and modalities of hypermutation.. J. Immunol. 162, 3437–3447 (1999)

    CAS  Google Scholar 

  6. Zhang, W. et al. Clonal instability of V region hypermutation in the Ramos Burkitt's lymphoma cell line. Int. Immunol. 13, 1175–1184 (2001)

    Article  CAS  Google Scholar 

  7. Wagner, S. D., Milstein, C. & Neuberger, M. S. Codon bias targets mutation. Nature 376, 732 (1995)

    Article  ADS  CAS  Google Scholar 

  8. Rogozin, I. B. & Kolchanov, N. A. Somatic hypermutagenesis in immunoglobulin genes. II. Influence of neighbouring base sequences on mutagenesis. Biochim. Biophys. Acta 1171, 11–18 (1992)

    Article  CAS  Google Scholar 

  9. Connor, A., Wiersma, E. & Shulman, M. J. On the linkage between RNA processing and RNA translatability. J. Biol. Chem. 269, 25178–25184 (1994)

    CAS  Google Scholar 

  10. Lin, M. M., Zhu, M. & Scharff, M. D. Sequence dependent hypermutation of the immunoglobulin heavy chain in cultured B cells. Proc. Natl Acad. Sci. USA 94, 5284–5289 (1997)

    Article  ADS  CAS  Google Scholar 

  11. Luria, S. E. & Delbruck, M. Mutation of bacteria from virus sensitivity to virus resistance. Genetics 28, 491–511 (1943)

    CAS  Google Scholar 

  12. Rada, C. & Milstein, C. The intrinsic hypermutability of antibody heavy and light chain genes decays exponentially. EMBO J. 20, 4570–4576 (2001)

    Article  CAS  Google Scholar 

  13. Rada, C., Yelamos, J., Dean, W. & Milstein, C. The 5′ hypermutation boundary of κ chains is independent of local and neighbouring sequences and related to the distance from the initiation of transcription. Eur. J. Immunol. 27, 3115–3120 (1997)

    Article  CAS  Google Scholar 

  14. Tao, W. & Bothwell, A. L. Development of B cell lineages during a primary anti-hapten immune response. J. Immunol. 145, 3216–3222 (1990)

    CAS  Google Scholar 

  15. Rada, C., Ehrenstein, M. R., Neuberger, M. S. & Milstein, C. Hot spot focusing of somatic hypermutation in MSH2-deficient mice suggests two stages of mutational targeting. Immunity 9, 135–141 (1998)

    Article  CAS  Google Scholar 

  16. Spencer, J., Dunn, M. & Dunn-Walters, D. K. Characteristics of sequences around individual nucleotide substitutions in IgVH genes suggest different GC and AT mutators. J. Immunol. 162, 6596–6601 (1999)

    CAS  Google Scholar 

  17. Bachl, J., Carlson, C., Gray-Schopfer, V., Dessing, M. & Olsson, C. Increased transcription levels induce higher mutation rates in a hypermutating cell line. J. Immunol. 166, 5051–5017 (2001)

    Article  CAS  Google Scholar 

  18. Phung, Q. H. et al. Increased hypermutation at G and C nucleotides in immunoglobulin variable genes from mice deficient in the MSH2 mismatch repair protein. J. Exp. Med. 187, 1745–1751 (1998)

    Article  CAS  Google Scholar 

  19. Reynaud, C. A. et al. Mismatch repair and immunoglobulin gene hypermutation: did we learn something? Immunol. Today 20, 522–527 (1999)

    Article  CAS  Google Scholar 

  20. Vora, K. A. et al. Severe attenuation of the B cell immune response in Msh2-deficient mice. J. Exp. Med. 189, 471–481 (1999)

    Article  CAS  Google Scholar 

  21. Wiesendanger, M., Kneitz, B., Edelmann, W. & Scharff, M. D. Somatic mutation in MSH3, MSH6, and MSH3/MSH6-deficient mice reveals a role for the MSH2–MSH6 heterodimer in modulating the base substitution pattern. J. Exp. Med. 191, 579–584 (2000)

    Article  CAS  Google Scholar 

  22. Zan, H. et al. The translesional polymerase ζ plays a major role in Ig and Bcl-6 somatic mutation. Immunity 14, 643–653 (2001)

    Article  CAS  Google Scholar 

  23. Zeng, X. et al. DNA polymerase η is an A-T mutator in somatic hypermutation of immunoglobulin variable genes. Nature Immunol. 2, 537–541 (2001)

    Article  CAS  Google Scholar 

  24. Rogozin, I. B., Pavlov, Y. I., Bebenek, K., Matsuda, T. & Kunkel, T. A. Somatic mutation hotspots correlate with DNA polymerase η error spectrum. Nature Immunol. 2, 530–536 (2001)

    Article  CAS  Google Scholar 

  25. Kobrin, B. J., Casadevall, A., Pirofski, L.-A., Schiff, C. & Scharff, M. D. in Somatic Hypermutation in V Regions (ed. Steele, E. J.) 11–28 (CRC, Boca Raton, 1990)

    Google Scholar 

  26. Kohler, G. & Milstein, C. Continuous cultures of fused cells secreting antibody of predefined specificity. Nature 256, 495–497 (1975)

    Article  ADS  CAS  Google Scholar 

  27. Sack, S. Z., Bardwell, P. D. & Scharff, M. D. Testing the reverse transcriptase model of somatic mutation. Mol. Immunol. 38, 303–311 (2001)

    Article  CAS  Google Scholar 

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Acknowledgements

We thank B. Diamond, J. Warner, M. Goodman and R. Laskov for critical review of the manuscript, and A. Bothwell for providing the P1-5 hybridoma. This work was supported by grants from the National Institutes of Health to P.D.B., to C.J.W. and to M.D.S., who is also supported by the Harry Eagle chair provided by the National Women's Division of the Albert Einstein College of Medicine. A.M. is a recipient of Cancer Research Institute and Harry Eagle fellowships.

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Correspondence to Matthew D. Scharff.

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Martin, A., Bardwell, P., Woo, C. et al. Activation-induced cytidine deaminase turns on somatic hypermutation in hybridomas. Nature 415, 802–806 (2002). https://doi.org/10.1038/nature714

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