Altering the pathway of immunoglobulin hypermutation by inhibiting uracil-DNA glycosylase

Abstract

A functional immune system depends on the production of a wide range of immunoglobulin molecules. Immunoglobulin variable region (IgV) genes are diversified after gene rearrangement by hypermutation. In the DNA deamination model, we have proposed that deamination of dC residues to dU by activation-induced deaminase (AID) triggers this diversification. In hypermutating chicken DT40 B cells, most IgV mutations are dC → dG/dA or dG → dC/dT transversions, which are proposed to result from replication over sites of base loss produced by the excision activity of uracil-DNA glycosylase. Blocking the activity of uracil-DNA glycosylase should instead lead to replication over the dU lesion, resulting in dC → dT (and dG → dA) transitions. Here we show that expression in DT40 cells of a bacteriophage-encoded protein that inhibits uracil-DNA glycosylase shifts the pattern of IgV gene mutations from transversion dominance to transition dominance. This is good evidence that antibody diversification involves dC → dU deamination within the immunoglobulin locus itself.

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Figure 1: Uracil-DNA glycosylase activity in the DT40 B-cell line.
Figure 2: Analysis of Vλ mutations in surface IgM-loss variants sorted from pEF-Ugi and control transfectants.
Figure 3: Analysis of Vλ mutations in unsorted populations of pEF-Ugi and control transfectants after 3–8 weeks of clonal expansion.

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Acknowledgements

We thank R. Savva for the Ugi cassette, S. Takeda for XRCC2-deficient DT40 cells, R. Grenfell for help with cell sorting, and R. Harris, S. Petersen-Mahrt, C. Rada and J. Sale for discussions. J.D.N. was supported by a César Milstein fellowship and the Fundación Antorchas, Argentina.

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Correspondence to Michael S. Neuberger.

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Di Noia, J., Neuberger, M. Altering the pathway of immunoglobulin hypermutation by inhibiting uracil-DNA glycosylase. Nature 419, 43–48 (2002). https://doi.org/10.1038/nature00981

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