title: Fine mapping of V(D)J recombinase mediated rearrangements in human lymphoid malignancies.
abstract: BACKGROUND
  Lymphocytes achieve diversity in antigen recognition in part by rearranging genomic DNA at loci encoding antibodies and cell surface receptors. The process, termed V(D)J recombination, juxtaposes modular coding sequences for antigen binding. Erroneous recombination events causing chromosomal translocations are recognized causes of lymphoid malignancies. Here we show a hybridization based method for sequence enrichment can be used to efficiently and selectively capture genomic DNA adjacent to V(D)J recombination breakpoints for massively parallel sequencing. The approach obviates the need for PCR amplification of recombined sequences.
  
  
  RESULTS
  Using tailored informatics analyses to resolve alignment and assembly issues in these repetitive regions, we were able to detect numerous recombination events across a panel of cancer cell lines and primary lymphoid tumors, and an EBV transformed lymphoblast line. With reassembly, breakpoints could be defined to single base pair resolution. The observed events consist of canonical V(D)J or V-J rearrangements, non-canonical rearrangements, and putatively oncogenic reciprocal chromosome translocations. We validated non-canonical and chromosome translocation junctions by PCR and Sanger sequencing. The translocations involved the MYC and BCL-2 loci, and activation of these was consistent with histopathologic features of the respective B-cell tumors. We also show an impressive prevalence of novel erroneous V-V recombination events at sites not incorporated with other downstream coding segments.
  
  
  CONCLUSIONS
  Our results demonstrate the ability of next generation sequencing to describe human V(D)J recombinase activity and provide a scalable means to chronicle off-target, unexpressed, and non-amplifiable recombinations occurring in the development of lymphoid cancers.
authors: McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University, Baltimore, MD, USA. kburns9@jhmi.edu.
authors: Eitan
authors: E
authors: Halper-Stromberg
authors: Jared
authors: J
authors: Steranka
authors: Nicolas
authors: N
authors: Giraldo-Castillo
authors: Timothy
authors: T
authors: Fuller
authors: Stephen
authors: S
authors: Desiderio
authors: Kathleen H
authors: KH
authors: Burns
mesh_terms: D001402:B-Lymphocytes / Q000201:enzymology; D001483:Base Sequence; D063646:Carcinogenesis / Q000235:genetics; D045744:Cell Line, Tumor; D056905:Chromosome Breakpoints; D002874:Chromosome Mapping; D015321:Gene Rearrangement; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D007945:Leukemia, Lymphoid / Q000201:enzymology / Q000235:genetics*; D017422:Sequence Analysis, DNA; D014178:Translocation, Genetic; D045567:VDJ Recombinases / Q000502:physiology*
chemical_list: D045567:VDJ Recombinases
references: Nature. 1998 Aug 20;394(6695):744-51
references: 9723614
references: Brief Bioinform. 2010 Sep;11(5):524-34
references: 20427421
references: Blood. 1987 Dec;70(6):1816-23
references: 2823937
references: Blood. 1996 Nov 15;88(10):3937-44
references: 8916960
references: Methods Mol Biol. 2012;882:569-604
references: 22665256
references: Nucleic Acids Res. 1998 Jan 1;26(1):297-303
references: 9399859
references: Nature. 1986 Jan 2-8;319(6048):28-33
references: 3484541
references: Genes Immun. 2012 Jul;13(5):363-73
references: 22551722
references: Genome Res. 2002 Apr;12(4):656-64
references: 11932250
references: Nat Biotechnol. 2011 Jan;29(1):24-6
references: 21221095
references: Proc Natl Acad Sci U S A. 1987 Apr;84(8):2396-400
references: 3104914
references: Genes Chromosomes Cancer. 2012 Mar;51(3):290-9
references: 22120970
references: Adv Cancer Res. 1982;37:319-80
references: 6305160
references: Int Immunol. 1998 Feb;10(2):241-6
references: 9533453
references: Lancet. 1999 Oct 30;354(9189):1499-503
references: 10551495
references: Eur J Immunol. 1992 Jun;22(6):1561-5
references: 1601042
references: Cell. 2008 Dec 12;135(6):1130-42
references: 19070581
references: Proc Natl Acad Sci U S A. 1985 May;82(9):2925-9
references: 3873070
references: Nucleic Acids Res. 2006 Jan 1;34(Database issue):D781-4
references: 16381979
references: Cell. 1998 Aug 21;94(4):463-70
references: 9727489
references: Bioinformatics. 2009 May 1;25(9):1189-91
references: 19151095
references: Blood. 1999 Jun 15;93(12):4079-85
references: 10361104
references: Nucleic Acids Res. 2011 Jan;39(Database issue):D895-900
references: 21051346
references: MAbs. 2011 Jan-Feb;3(1):17-20
references: 21099370
references: J Immunol. 1992 Jul 15;149(2):511-6
references: 1624797
references: Genome Res. 2010 May;20(5):623-35
references: 20308636