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DNA-dependent protein kinase is not required for accumulation of p53 or cell cycle arrest after DNA damage
W K Rathmell1, W K Kaufmann, J C Hurt
1Department of Medicine, Stanford University School of Medicine, California 94305, USA.
Abstract:
In response to DNA damage, cells transduce a signal that leads to accumulation and activation of p53 protein, transcriptional induction of several genes, including p21, gadd45, and gadd153, and cell cycle arrest. One hypothesis is that the signal is mediated by DNA-dependent protein kinase (DNA-PK), which consists of a catalytic subunit (DNA-PKcs) and a regulatory subunit (Ku). DNA-PK has several characteristics that support this hypothesis: Ku binds to DNA damaged by nicks or double-strand breaks, DNA-PKcs is activated when Ku binds to DNA, DNA-PK will phosphorylate p53 and other cell cycle regulatory proteins in vitro, and DNA-PKcs shares homology with ATM, which is mutated in ataxia telangiectasia and involved in signaling the p53 response to ionizing radiation. The hypothesis was tested by analyzing early passage fibroblasts from severe combined immunodeficient mice, which are deficient in DNA-PK. After exposure to ionizing radiation, UV radiation, or methyl methane-sulfonate, severe combined immunodeficient and wild-type cells were indistinguishable in their response. The accumulation of p53, induction of p21, gadd45, and gadd153, and arrest of the cell cycle in G1 and G2 occurred normally. Therefore, DNA-PK is not required for the p53 response or cell cycle arrest after DNA damage.
Insights
DNA-dependent protein kinase (DNA-PK) was hypothesized to mediate DNA damage signaling. However, studies show DNA-PK is not required for p53 activation or cell cycle arrest following DNA damage.
Area of Science:
- Molecular Biology
- Cell Biology
- Genetics
Background:
- DNA damage triggers cellular responses, including p53 protein accumulation, gene induction (p21, gadd45, gadd153), and cell cycle arrest.
- DNA-dependent protein kinase (DNA-PK), comprising DNA-PKcs and Ku subunits, was hypothesized to mediate this signaling pathway.
- DNA-PK's characteristics, such as Ku binding to damaged DNA and DNA-PKcs activation, supported its proposed role in DNA damage response.
Purpose of the Study:
- To investigate the role of DNA-dependent protein kinase (DNA-PK) in the cellular response to DNA damage.
- To determine if DNA-PK is essential for p53 protein accumulation, gene induction, and cell cycle arrest after exposure to various DNA-damaging agents.
Main Methods:
- Utilized fibroblasts from severe combined immunodeficient (SCID) mice, which lack functional DNA-PK.
- Exposed both SCID and wild-type cells to DNA-damaging agents: ionizing radiation, UV radiation, and methyl methanesulfonate.
- Analyzed and compared the cellular responses, including p53 accumulation, p21, gadd45, and gadd153 induction, and cell cycle progression (G1 and G2 arrest).
Main Results:
- SCID and wild-type cells exhibited indistinguishable responses to DNA damage.
- Normal accumulation of p53 protein was observed in both cell types.
- Induction of p21, gadd45, and gadd153, as well as cell cycle arrest in G1 and G2 phases, occurred similarly in DNA-PK deficient and sufficient cells.
Conclusions:
- DNA-dependent protein kinase (DNA-PK) is not required for the p53-mediated response to DNA damage.
- The signaling pathway leading to p53 activation and cell cycle arrest after DNA damage does not depend on DNA-PK activity.