Double-strand break formation by the RAG complex at the bcl-2 major breakpoint region and at other non-B DNA structures in vitro.
The most common chromosomal translocation in cancer, t(14;18) at the 150-bp bcl-2 major breakpoint region (Mbr), occurs in follicular lymphomas. The bcl-2 Mbr assumes a non-B DNA conformation, thus explaining its distinctive fragility. This non-B DNA structure is a target of the RAG complex in vivo, but not because of its primary sequence. Here we report that the RAG complex generates at least two independent nicks that lead to double-strand breaks in vitro, and this requires the non-B DNA structure at the bcl-2 Mbr. A 3-bp mutation is capable of abolishing the non-B structure formation and the double-strand breaks. The observations on the bcl-2 Mbr reflect more general properties of the RAG complex, which can bind and nick at duplex-single-strand transitions of other non-B DNA structures, resulting in double-strand breaks in vitro. Hence, the present study reveals novel insight into a third mechanism of action of RAGs on DNA, besides the standard heptamer/nonamer-mediated cleavage in V(D)J recombination and the in vitro transposase activity.
Norris Comprehensive Cancer Center, Rm. 5428, University of Southern California Keck School of Medicine, 1441 Eastlake Ave., MC9176, Los Angeles, California 90033, USA.
Sathees C
SC
Raghavan
Patrick C
PC
Swanson
Yunmei
Y
Ma
Michael R
MR
Lieber
D000818:Animals; D001483:Base Sequence; D002883:Chromosomes, Human, Pair 14 / Q000235:genetics; D002887:Chromosomes, Human, Pair 18 / Q000235:genetics; D004247:DNA / Q000737:chemistry* / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D019254:Genes, bcl-2 / Q000235:genetics*; D018398:Homeodomain Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008224:Lymphoma, Follicular / Q000235:genetics*; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins; D009690:Nucleic Acid Conformation; D011995:Recombination, Genetic; D014178:Translocation, Genetic
D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D009687:Nuclear Proteins; C499311:RAG2 protein, human; C499308:Rag2 protein, mouse; C064655:V(D)J recombination activating protein 2; C064658:RAG-1 protein; D004247:DNA