Mechanisms promoting translocations in editing and switching peripheral B cells

Abstract

Variable, diversity and joining gene segment (V(D)J) recombination assembles immunoglobulin heavy or light chain (IgH or IgL) variable region exons in developing bone marrow B cells, whereas class switch recombination (CSR) exchanges IgH constant region exons in peripheral B cells. Both processes use directed DNA double-strand breaks (DSBs) repaired by non-homologous end-joining (NHEJ). Errors in either V(D)J recombination or CSR can initiate chromosomal translocations, including oncogenic IgH locus (Igh) to c-myc (also known as Myc) translocations of peripheral B cell lymphomas. Collaboration between these processes has also been proposed to initiate translocations. However, the occurrence of V(D)J recombination in peripheral B cells is controversial. Here we show that activated NHEJ-deficient splenic B cells accumulate V(D)J-recombination-associated breaks at the lambda IgL locus (Igl), as well as CSR-associated Igh breaks, often in the same cell. Moreover, Igl and Igh breaks are frequently joined to form translocations, a phenomenon associated with specific Igh–Igl co-localization. Igh and c-myc also co-localize in these cells; correspondingly, the introduction of frequent c-myc DSBs robustly promotes Igh–c-myc translocations. Our studies show peripheral B cells that attempt secondary V(D)J recombination, and determine a role for mechanistic factors in promoting recurrent translocations in tumours.

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Figure 1: Role of AID and RAG in generating Igh, Igk and Igl breaks in CD21-cre, Xrcc4c/- splenic B cells.
Figure 2: Frequent Igh–Igl translocations in activated XRCC4-deficient splenic B cells.
Figure 3: Frequent cell-type and Igλ locus-specific Igh and Igl co-localization.
Figure 4: DSBs in c-myc are rate-limiting for Igh–c-myc translocations in activated splenic B cells.

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Acknowledgements

We thank Alt laboratory members for discussions, and Y. L. Chen, J. M. Bianco and M. Moghimi for technical assistance. This work was supported by the National Insitutes of Health (NIH) grant 5P01CA92625 and a Leukemia and Lymphoma Society of America (LLS) SCORE grant to F.W.A. and K.R. M.G. is and J.H.W. was a Special Fellow of the LLS. J.H.W. and D.R.W. are supported by an NIH training grant and C.T.Y. was supported by an NCI training grant. A.N. is supported by the Intramural Research program of the NIH, NCI, Center for Cancer Research. F.W.A. is an Investigator of the Howard Hughes Medical Institute.

Author Contributions F.W.A., J.H.W., M.G. and C.T.Y. planned studies and interpreted data. J.H.W. performed most experiments, including mouse breeding, B cell studies, FISH, and Igh and Igl PCR studies. C.T.Y. bred mice and performed B cell analyses. M.G. generated and analysed c-myc25IsceI/WT mice and performed FISH and Igh–c-myc translocation studies. P.G., T.H., and E.H. provided technical assistance. S.D. and A.N. provided expertise in 3D interphase FISH. A.A.Z. generated the 25 IsceI array. D.R.W. performed RAG expression studies and mesenteric lymph node B cell analyses. K.R. provided RAG conditional knockout mice and helped interpret data. F.W.A., J.H.W. and M.G. wrote the paper.

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Correspondence to Frederick W. Alt.

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Wang, J., Gostissa, M., Yan, C. et al. Mechanisms promoting translocations in editing and switching peripheral B cells. Nature 460, 231–236 (2009). https://doi.org/10.1038/nature08159

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