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V(D)J recombination activates a p53-dependent DNA damage checkpoint in scid lymphocyte precursors
C J Guidos1, C J Williams, I Grandal
1Division of Immunology and Cancer, Hospital for Sick Children Research Institute, Toronto, Ontario, Canada.
Abstract:
Double-stranded DNA breaks (DSBs) trigger p53-mediated cell cycle arrest or apoptosis pathways that limit the oncogenic consequences of exposure to genotoxic agents, but p53-mediated responses to DSB generated by normal physiologic events have not been documented. "Broken" V(D)J coding ends accumulate in scid lymphocyte precursors as a consequence of a mutation in DNA-dependent protein kinase (DNA-PK). The ensuing failure to rearrange efficiently antigen receptors arrests lymphoid development. Here we show that scid thymocytes express high levels of p53 protein, attributable to recombinase activating gene (RAG)-dependent generation of DSB adjacent to V, D, and J gene segments. To examine the functional importance of p53 expression in vivo, we bred p53-/- scid mice. The absence of p53 facilitated production of in-frame V(D)Jbeta coding joints and developmental progression of scid thymocytes, in addition to a dramatic accumulation of pro-B cells. All mice developed disseminated pro-B or immature T cell lymphoma/leukemia by 7-12 weeks of age. We present evidence that p53 deficiency prolongs the survival of scid lymphocyte precursors harboring broken V(D)J coding ends, allowing the accumulation of aneuploid cells. These results demonstrate that a p53-mediated DNA damage checkpoint contributes to the immune deficiency characteristic of the scid mutation and limits the oncogenic potential of DSBs generated during V(D)J recombination.
Insights
The p53 protein, crucial for DNA damage response, normally prevents cancer. In severe combined immunodeficiency (scid) mice, its absence allows DNA breaks during immune receptor development, leading to lymphoma.
Area of Science:
- Immunology
- Molecular Biology
- Genetics
- Cancer Research
Background:
- Double-stranded DNA breaks (DSBs) typically activate p53-mediated pathways for cell cycle arrest or apoptosis, preventing oncogenesis from genotoxic agents.
- Physiological DSBs, such as those occurring during V(D)J recombination in lymphocyte development, have not been well-documented for p53-mediated responses.
- Severe combined immunodeficiency (scid) is characterized by a mutation in DNA-dependent protein kinase (DNA-PK), leading to accumulation of broken V(D)J coding ends and arrested lymphoid development.
Purpose of the Study:
- To investigate the role of p53 in responding to DSBs generated during physiological V(D)J recombination in lymphocyte precursors.
- To determine the functional significance of p53 in vivo within the context of the scid mutation and its impact on lymphoid development and oncogenesis.
Main Methods:
- Analysis of p53 protein levels in scid thymocytes, correlating expression with RAG-dependent DSB generation.
- Generation and study of p53 knockout (p53-/-) scid mice to assess the in vivo consequences of p53 deficiency.
- Evaluation of V(D)J recombination, lymphoid development, cell survival, aneuploidy, and lymphoma/leukemia incidence in p53-/- scid mice.
Main Results:
- Scid thymocytes exhibit high p53 protein levels due to RAG-dependent DSBs near V, D, and J gene segments.
- Absence of p53 in p53-/- scid mice facilitated in-frame V(D)Jbeta coding joint formation and thymocyte development, alongside pro-B cell accumulation.
- All p53-/- scid mice developed disseminated lymphoma/leukemia by 7-12 weeks, with evidence of prolonged survival of DSB-containing precursors and aneuploid cell accumulation.
Conclusions:
- A p53-mediated DNA damage checkpoint is integral to the immune deficiency observed in scid mutations.
- p53 deficiency in scid mice enhances the oncogenic potential of DSBs generated during V(D)J recombination, leading to lymphoma.
- These findings highlight p53's critical role in preventing cancer by safeguarding against genomic instability during normal immune system development.