Processive AID-catalysed cytosine deamination on single-stranded DNA simulates somatic hypermutation

Pham, Phuong
Bransteitter, Ronda
Petruska, John
Goodman, Myron F.

Departments of Biological Sciences and Chemistry, Hedco Molecular Biology Laboratories, University of Southern California, University Park, Los Angeles, California 90089-1340, USA

Received 10 April; accepted 28 May 2003; doiL10.1038/nature01760.

Published online 18 June 2003.

Supplementary Information accompanies the paper on www.nature.com/nature.

Correspondence and requests for materials should be addressed to M.F.G. ([email protected]).

Nature 424(6944):p 103-107, July 3, 2003.

Activation-induced cytidine deaminase (AID) is a protein required for B cells to undergo class switch recombination and somatic hypermutation (SHM)-two processes essential for producing high-affinity antibodies. Purified AID catalyses the deamination of C to U on single-stranded (ss)DNA. Here, we showin vitrothat AID-catalysed C deaminations occur preferentially on 5′ WRCsequences in accord with SHM spectra observedin vivo.Although about 98% of DNA clones suffer no mutations, most of the remaining mutated clones have 10-70 C to T transitions per clone. Therefore, AID carries out multiple C deaminations on individual DNA strands, rather than jumping from one strand to another. The avid binding of AID to ssDNA could result from its large net positive charge (+11) at pH 7.0, owing to a basic amino-terminal domain enriched in arginine and lysine. Furthermore, AID exhibits a 15-fold preference for C deamination on the non-transcribed DNA strand exposed by RNA polymerase than the transcribed strand protected as a RNA-DNA hybrid. These deamination results on ssDNA bear relevance to three characteristic features of SHM: preferential mutation at C sites within WRChotspot sequences, the broad clonal mutagenic heterogeneity of antibody variable regions targeted for mutation, and the requirement for active transcription to obtain mutagenesis.