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Nucleic Acids Research Advance Access originally published online on November 16, 2006
Nucleic Acids Research 2006 34(21):6345-6351; doi:10.1093/nar/gkl830
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Nucleic Acids Research, 2006, Vol. 34, No. 21 6345-6351
© 2006 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Molecular Biology

NHEJ-deficient DT40 cells have increased levels of immunoglobulin gene conversion: evidence for a double strand break intermediate

Ephraim S. Tang and Alberto Martin*

Department of Immunology, 5265 Medical Sciences Building, University of Toronto Ontario, Toronto, Canada M5S 1A8

*To whom correspondence should be addressed. Tel: +416 978 4235; Fax +416 978 1938; Email: alberto.martin{at}utoronto.ca

Received June 20, 2006. Revised October 6, 2006. Accepted October 6, 2006.

Activation-induced cytidine deaminase (AID) likely initiates immunoglobulin gene-conversion (GC) by deaminating cytidines within the V-region of chicken B-cells. However, the intervening DNA lesion required to initiate GC remains elusive. GC could be initiated by a single strand break or a double strand break (DSB). To distinguish between these possibilities, we examined GC in the chicken DT40 B cell line deficient in non-homologous end joining (NHEJ). It is known that the NHEJ and homologous recombination DNA repair pathways compete for DSBs. In light of this, if a DSB is the major intermediate, deficiency in NHEJ should result in increased levels of GC. Here we show that DNA–PKcs–/–/– and Ku70–/– DT40 cells had 5- to 10-fold higher levels of GC relative to wildtype DT40 as measured by surface IgM reversion and sequencing of the V-region. These data suggest that a DSB is the major DNA lesion that initiates GC.


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