V-region mutation in vitro, in vivo, and in silico reveal the importance of the enzymatic properties of AID and the sequence environment

Thomas MacCarthy, Susan L. Kalis, Sergio Roa, Phuong Pham, Myron F. Goodman, Matthew D. Scharff, Aviv Bergman

Research output: Contribution to journalArticle

27 Scopus citations

Abstract

The somatic hypermutation of Ig variable regions requires the activity of activation-induced cytidine deaminase (AID) which has previously been shown to preferentially deaminate WRC (W = A/T, R = A/G) motif hot spots in in vivo and in vitro assays. We compared mutation profiles of in vitro assays for the 3′ flanking intron of VhJ558-Jh4 region to previously reported in vivo profiles for the same region in the Msh2-/-Ung-/- mice that lack base excision and mismatch repair.Wefound that the in vitro and in vivo mutation profiles were highly correlated for the top (nontranscribed) strand, while for the bottom (transcribed) strand the correlation is far lower. We used an in silico model of AID activity to elucidate the relative importance of motif targeting in vivo. We found that the mutation process entails substantial complexity beyond motif targeting, a large part of which is captured in vitro. To elucidate the contribution of the sequence environment to the observed differences between the top and bottom strands, we analyzed intermutational distances. The bottom strand shows an approximately exponential distribution of distances in vivo and in vitro, as expected from a null model. However, the top strand deviates strongly from this distribution in that mutations approximately 50 nucleotides apart are greatly reduced, again both in vivo and in vitro, illustrating an important strand asymmetry. While we have confirmed that AID targeting of hot and cold spots is a key part of the mutation process, our results suggest that the sequence environment plays an equally important role.

Original languageEnglish (US)
Pages (from-to)8629-8634
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume106
Issue number21
DOIs
StatePublished - May 26 2009

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Keywords

  • Simulation
  • Somatic hypermutation
  • Variable-region

ASJC Scopus subject areas

  • General

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