Related Experiment Videos
Functional interaction between DNA-PK and c-Abl in response to DNA damage
1Division of Cancer Pharmacology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, Massachusetts 02115, USA.
Abstract:
How DNA damage is converted into intracellular signals that can control cell behaviour is unknown. The c-Abl protein tyrosine kinase is activated by ionizing radiation and certain other DNA-damaging agents, whereas the DNA-dependent protein kinase (DNA-PK), consisting of a serine/threonine kinase and Ku DNA-binding subunits, requires DNA double-strand breaks or other DNA lesions for activation. Here we demonstrate that c-Abl interacts constitutively with DNA-PK. Ionizing radiation stimulates binding of c-Abl to DNA-PK and induces an association of c-Abl with Ku antigen. We show that DNA-PK phosphorylates and activates c-Abl in vitro. Cells deficient in DNA-PK are defective in c-Abl activation induced by ionizing radiation. In a potential feedback mechanism, c-Abl phosphorylates DNA-PK, but not Ku, in vitro. Phosphorylation of DNA-PK by c-Abl inhibits the ability of DNA-PK to form a complex with DNA. We also show that treatment of cells with ionizing radiation results in phosphorylation of DNA-PK that is dependent on c-Abl. Our results support the hypothesis that there are functional interactions between c-Abl and DNA-PK in the response to DNA damage.
Insights
The study reveals how DNA damage signals are transmitted within cells. DNA-dependent protein kinase (DNA-PK) activates c-Abl protein tyrosine kinase following DNA damage, forming a crucial signaling pathway.
Area of Science:
- Molecular Biology
- Cellular Signaling
- DNA Repair Mechanisms
Background:
- The precise mechanisms converting DNA damage into cellular signals remain unclear.
- Protein tyrosine kinases like c-Abl and DNA-dependent protein kinase (DNA-PK) are activated by DNA-damaging agents.
- DNA-PK, comprising kinase and Ku subunits, requires DNA lesions for activation.
Purpose of the Study:
- To elucidate the functional interactions between c-Abl and DNA-PK in response to DNA damage.
- To investigate how DNA damage signaling is mediated through these protein kinases.
- To understand the role of these interactions in cellular behavior control.
Main Methods:
- Demonstration of constitutive and radiation-stimulated interactions between c-Abl and DNA-PK/Ku complex.
- In vitro kinase assays to assess phosphorylation and activation of c-Abl by DNA-PK.
- Analysis of c-Abl activation in DNA-PK deficient cells following ionizing radiation.
- In vitro phosphorylation of DNA-PK by c-Abl and assessment of its effect on DNA binding.
Main Results:
- c-Abl constitutively interacts with DNA-PK, with binding enhanced by ionizing radiation.
- DNA-PK phosphorylates and activates c-Abl in vitro; this activation is impaired in DNA-PK deficient cells.
- c-Abl phosphorylates DNA-PK (but not Ku) in vitro, inhibiting DNA-PK's DNA-binding ability.
- Ionizing radiation induces c-Abl-dependent phosphorylation of DNA-PK in cells.
Conclusions:
- Functional interactions between c-Abl and DNA-PK are critical for the cellular response to DNA damage.
- DNA-PK-mediated activation of c-Abl and c-Abl-mediated regulation of DNA-PK form a signaling network.
- These interactions likely play a role in controlling cell behavior following DNA damage.