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Generation of Gross Chromosomal Rearrangements by a Single Engineered DNA Double Strand Break
Zhijun Qiu1, Zhenhua Zhang1, Anna Roschke1
1Genetics Branch National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA.
Abstract:
Gross chromosomal rearrangements (GCRs), including translocations, inversions amplifications, and deletions, can be causal events leading to malignant transformation. GCRs are thought to be triggered by DNA double strand breaks (DSBs), which in turn can be spontaneous or induced by external agents (eg. cytotoxic chemotherapy, ionizing radiation). It has been shown that induction of DNA DSBs at two defined loci can produce stable balanced chromosomal translocations, however, a single engineered DNA DSB could not. Herein, we report that although a single engineered DNA DSB in H2AX "knockdown" cells did not generate GCRs, repair of a single engineered DNA DSB in fibroblasts that had ablated H2ax did produce clonal, stable GCRs, including balanced translocations and megabase-pair inversions. Upon correction of the H2ax deficiency, cells no longer generated GCRs following a single engineered DNA DSB. These findings demonstrate that clonal, stable GCRs can be produced by a single engineered DNA DSB in H2ax knockout cells, and that the production of these GCRs is ameliorated by H2ax expression.
Insights
Gross chromosomal rearrangements (GCRs) can cause cancer. Researchers found that repairing a single DNA double-strand break (DSB) in cells lacking H2AX protein can lead to GCRs, a process reduced by H2AX expression.
Area of Science:
- Genetics
- Molecular Biology
- Cancer Research
Background:
- Gross chromosomal rearrangements (GCRs) are key events in malignant transformation.
- GCRs are often initiated by DNA double-strand breaks (DSBs).
- Previous studies indicated that a single engineered DSB could not induce GCRs.
Purpose of the Study:
- To investigate if a single engineered DNA double-strand break (DSB) can induce GCRs in the absence of H2AX.
- To determine the role of H2AX in the repair of DSBs and subsequent GCR formation.
Main Methods:
- Engineered single DNA double-strand breaks (DSBs) in H2AX-deficient (knockout) and H2AX-knockdown cells.
- Analyzed repair outcomes, including chromosomal translocations and inversions.
- Restored H2AX expression to assess its effect on GCR production.
Main Results:
- A single engineered DSB did not induce GCRs in H2AX-knockdown cells.
- Repair of a single engineered DSB in H2AX-ablated fibroblasts produced stable, clonal GCRs, including translocations and megabase-pair inversions.
- Restoring H2AX expression prevented GCR formation after a single engineered DSB.
Conclusions:
- A single engineered DSB can generate clonal, stable GCRs in H2AX-deficient cells.
- H2AX plays a critical role in suppressing GCR formation following DSB repair.
- These findings highlight H2AX's function in maintaining genome stability.
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