Open Access Highly Accessed Research article

Fine mapping of V(D)J recombinase mediated rearrangements in human lymphoid malignancies

Eitan Halper-Stromberg1, Jared Steranka2, Nicolas Giraldo-Castillo7, Timothy Fuller38, Stephen Desiderio24 and Kathleen H Burns1356*

Author Affiliations

1 McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University, Baltimore, MD, USA

2 Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, MD, USA

3 Department of Oncology and The Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins Hospital, Baltimore, MD, USA

4 Immunology Unit of the Institute for Cell Engineering, Johns Hopkins University School of Medicine, 720 Rutland Avenue, Ross Building, Room 524, Baltimore 21205, MD, USA

5 Department of Pathology, Johns Hopkins University, Baltimore, MD, USA

6 Department of Pathology, Johns Hopkins School of Medicine, 720 Rutland Avenue, Ross Building, Room 524, Baltimore 21205, MD, USA

7 Los Andes University, Bogotá, Colombia

8 Mercer University School of Medicine, Macon, GA, USA

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BMC Genomics 2013, 14:565  doi:10.1186/1471-2164-14-565

Published: 19 August 2013

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.

Keywords:
V(D)J recombination; Oncogenic translocation; Lymphoid tumors; MYC; BCL-2; V replacement