Inducible DNA breaks in Ig S regions are dependent on AID and UNG.
Class switch recombination (CSR) occurs by an intrachromosomal deletion whereby the IgM constant region gene (Cmu) is replaced by a downstream constant region gene. This unique recombination event involves formation of double-strand breaks (DSBs) in immunoglobulin switch (S) regions, and requires activation-induced cytidine deaminase (AID), which converts cytosines to uracils. Repair of the uracils is proposed to lead to DNA breaks required for recombination. Uracil DNA glycosylase (UNG) is required for most CSR activity although its role is disputed. Here we use ligation-mediated PCR to detect DSBs in S regions in splenic B cells undergoing CSR. We find that the kinetics of DSB induction corresponds with AID expression, and that DSBs are AID- and UNG-dependent and occur preferentially at G:C basepairs in WRC/GYW AID hotspots. Our results indicate that AID attacks cytosines on both DNA strands, and staggered breaks are processed to blunt DSBs at the initiating ss break sites. We propose a model to explain the types of end-processing events observed.
Schrader
Carol E
CE
Department of Molecular Genetics and Microbiology, University of Massachusetts Medical School, Worcester, MA 01655, USA. carol.schrader@umassmed.edu
Linehan
Erin K
EK
Mochegova
Sofia N
SN
Woodland
Robert T
RT
Stavnezer
Janet
J
D000818:Animals; D001402:B-Lymphocytes / Q000166:cytology / Q000276:immunology*; D003564:Cytidine Deaminase / Q000235:genetics* / Q000276:immunology; D004247:DNA / Q000235:genetics / Q000276:immunology; D045647:DNA Glycosylases / Q000235:genetics* / Q000276:immunology; D005786:Gene Expression Regulation / Q000235:genetics / Q000276:immunology; D015322:Gene Rearrangement, B-Lymphocyte / Q000235:genetics* / Q000276:immunology; D007127:Immunoglobulin Constant Regions / Q000235:genetics / Q000276:immunology; D007148:Immunoglobulin mu-Chains / Q000235:genetics* / Q000276:immunology; D051379:Mice; D008817:Mice, Mutant Strains; D011995:Recombination, Genetic / Q000235:genetics / Q000276:immunology; D027041:Somatic Hypermutation, Immunoglobulin / Q000235:genetics* / Q000276:immunology; D013154:Spleen / Q000166:cytology / Q000276:immunology; D051981:Uracil-DNA Glycosidase; D000098830:AICDA (Activation-Induced Cytidine Deaminase)
D007127:Immunoglobulin Constant Regions; D007148:Immunoglobulin mu-Chains; D004247:DNA; D045647:DNA Glycosylases; D051981:Uracil-DNA Glycosidase; D000098830:AICDA (Activation-Induced Cytidine Deaminase); D003564:Cytidine Deaminase