Mutagenesis dependent upon the combination of activation-induced deaminase expression and a double-strand break

Laboratory of Structural Biology, National Institutes of Health, NIEHS, 111 T.W. Alexander Drive, PO Box 12233, MD F1-12, Research Triangle Park, NC 27709, USA.
Molecular Immunology (Impact Factor: 2.97). 11/2010; 48(1-3):164-70. DOI: 10.1016/j.molimm.2010.08.013
Source: PubMed


We explored DNA metabolic events potentially relevant to somatic hypermutation (SHM) of immunoglobulin genes using a yeast model system. Double-strand break (DSB) formation has been discussed as a possible component of the SHM process during immunoglobulin gene maturation. Yet, possible mechanisms linking DSB formation with mutagenesis have not been well understood. In the present study, a linkage between mutagenesis in a reporter gene and a double-strand break at a distal site was examined as a function of activation-induced deaminase (AID) expression. Induction of the DSB was found to be associated with mutagenesis in a genomic marker gene located 7 kb upstream of the break site: mutagenesis was strongest with the combination of AID expression and DSB induction. The mutation spectrum of this DSB and AID-mediated mutagenesis was characteristic of replicative bypass of uracil in one strand and was dependent on expression of DNA polymerase delta (Polδ). These results in a yeast model system illustrate that the combination of DSB induction and AID expression could be associated with mutagenesis observed in SHM. Implications of these findings for SHM of immunoglobulin genes in human B cells are discussed.

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    • "We chose to use APOBEC3G* for this experiment since it gave a good mutation load but a lower proportion of kataegic mutations than AID* (Supplementary file 1B): any enhancement of kataegis would therefore be more readily detectable. Consistent with previous findings (Poltoratsky et al., 2010), induction of I-SceI expression resulted in an increased frequency canonical APOBEC3G CC context. Two-thirds of the CAN1 region mutations in the KanMX-ISceI RS cells were transversions. "
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    • "Regions of ssDNA are recognized as a prerequisite for kataegis-like events induced by an alkylation agent in yeast, and by extrapolation, have been proposed to be a prerequisite for the kataegistic action of deaminases in humans [3]. Double-strand DNA breaks in the vicinity of a reporter gene synergistically stimulate mutagenesis by AID, and in yeast this behavior might be related to the generation of ssDNA during homologous recombination [8]. "
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