e n i c i d e M l a t n e m i r e p x E f o l a n r u o J e h T CORRESPONDENCE Bernardo Reina-San-Martin: reinab@igbmc.fr Abbreviations used: AID, activation-induced cytidine deaminase; ChIP, chromatin immunoprecipitation; CSR, class switch recombination; DDR, DNA damage response; DSB, double-stranded DNA break; HTS, high-throughput amplicon sequencing; IgH- FISH, IgH-specific fluorescence in situ hybridization; NHEJ, nonhomologous end joining; qRT-PCR, quantitative RT-PCR; SHM, somatic hypermutation. The Rockefeller University Press $30.00 J. Exp. Med. Vol. 208 No. 8 1649-1660 www.jem.org/cgi/doi/10.1084/jem.20110118 A r t i c l e Epigenetic tether ing of AID to the donor switch region dur ing immunoglobulin class switch recombination Beena Patricia Jeevan-Raj,1 Isabelle Robert,1 Vincent Heyer,1 Adeline Page,1 Jing H. Wang,2,3,5 Florence Cammas,1 Frederick W. Alt,2,3,4,6 Régine Losson,1 and Bernardo Reina-San-Martin1 1Institut de Génétique et de Biologie Moléculaire et Cellulaire, Institut National de Santé et de Recherche Médicale Unité 964/ Centre National de Recherche Scientifique Unité Mixte de Recherche 7104, Université de Strasbourg, 67404 Illkirch, France 2Immune Disease Institute; 3Department of Medicine and 4Howard Hughes Medical Institute, Children’s Hospital Boston; 5Department of Genetics; and 6Department of Genetics and Pediatrics; Harvard Medical School, Boston, MA 02115 Immunoglobulin class switch recombination (CSR) is initiated by double-stranded DNA breaks (DSBs) in switch regions triggered by activation-induced cytidine deaminase (AID). Although CSR correlates with epigenetic modifications at the IgH locus, the relationship between these modifications and AID remains unknown. In this study, we show that during CSR, AID forms a complex with KAP1 (KRAB domain–associated protein 1) and HP1 (heterochromatin protein 1) that is tethered to the donor switch region (S) bearing H3K9me3 (trimethylated histone H3 at lysine 9) in vivo. Furthermore, in vivo disruption of this complex results in impaired AID recruit- ment to S, inefficient DSB formation, and a concomitant defect in CSR but not in somatic hypermutation. We propose that KAP1 and HP1 tether AID to H3K9me3 residues at the donor switch region, thus providing a mechanism linking AID to epigenetic modifications during CSR. During immune responses, B cells diversify their Ig genes by somatic hypermutation (SHM) and class switch recombination (CSR). SHM modifies antibody affinities by introducing mutations in the variable region of heavy (IgH) and light chain genes (Di Noia and Neuberger, 2007). CSR modulates antibody effector func- tions by replacing the antibody isotype ex- pressed through a DNA recombination reaction that joins two switch regions (Chaudhuri et al., 2007). SHM and CSR are initiated by the de- amination of cytosines to uracils in single- stranded DNA mediated by activation-induced cytidine deaminase (AID; Petersen-Mahrt et al., 2002; Chaudhuri et al., 2007; Di Noia and Neuberger, 2007). The resulting dU:dG mis- matches are differentially processed to generate mutations in variable regions during SHM or double-stranded DNA breaks (DSBs) in switch regions during CSR (Chaudhuri et al., 2007; Di Noia and Neuberger, 2007). AID appears to find its targets by its binding to Spt5 and RNA J.H. Wang’s present address is Integrated Dept. of Immunol- ogy, University of Colorado School of Medicine and National Jewish Health, Denver, CO 80206. Dr. Losson died on 2 February 2010. polymerase II (Pavri et al., 2010) and distin- guishes between variable and switch regions by its association with proteins like RPA, PKA, 14-3-3, or by the formation of higher-order DNA structures in switch regions (Chaudhuri et al., 2007;Vuong et al., 2009; Xu et al., 2010; Yamane et al., 2011).AID itself also contributes to this choice, as N-terminal mutations in AID result in normal CSR but defective SHM (Shinkura et al., 2004;Wei et al., 2011) and con- versely C-terminal truncations in AID result in normal SHM but defective CSR (Chaudhuri et al., 2007; Kracker et al., 2010). The latter phenotype is also observed in mice deficient for DNA damage response (DDR) components (Ramiro et al., 2007) and in a subset of hyper IgM patients with a CSR-specific defect (Kracker et al., 2010). It has been proposed that this domain associates with CSR-specific factors that could be required for targeting AID to switch regions (Ta et al., 2003) or for recombi- nation downstream of DSB formation either by © 2011 Jeevan-Raj et al. This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.rupress.org/terms). After six months it is available under a Creative Commons License (Attribution– Noncommercial–Share Alike 3.0 Unported license, as described at http://creative- commons.org/licenses/by-nc-sa/3.0/). 1649