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Ku counteracts mobilization of PARP1 and MRN in chromatin damaged with DNA double-strand breaks.
- Source :
-
Nucleic acids research [Nucleic Acids Res] 2011 Dec; Vol. 39 (22), pp. 9605-19. Date of Electronic Publication: 2011 Aug 31. - Publication Year :
- 2011
-
Abstract
- In mammalian cells, the main pathway for DNA double-strand breaks (DSBs) repair is classical non-homologous end joining (C-NHEJ). An alternative or back-up NHEJ (B-NHEJ) pathway has emerged which operates preferentially under C-NHEJ defective conditions. Although B-NHEJ appears particularly relevant to genomic instability associated with cancer, its components and regulation are still largely unknown. To get insights into this pathway, we have knocked-down Ku, the main contributor to C-NHEJ. Thus, models of human cell lines have been engineered in which the expression of Ku70/80 heterodimer can be significantly lowered by the conditional induction of a shRNA against Ku70. On Ku reduction in cells, resulting NHEJ competent protein extracts showed a shift from C- to B-NHEJ that could be reversed by addition of purified Ku protein. Using a cellular fractionation protocol after treatment with a strong DSBs inducer followed by western blotting or immunostaining, we established that, among C-NHEJ factors, Ku is the main counteracting factor against mobilization of PARP1 and the MRN complex to damaged chromatin. In addition, Ku limits PAR synthesis and single-stranded DNA production in response to DSBs. These data support the involvement of PARP1 and the MRN proteins in the B-NHEJ route for the repair of DNA DSBs.<br /> (© The Author(s) 2011. Published by Oxford University Press.)
- Subjects :
- Antigens, Nuclear genetics
Cell Fractionation
Cell Line
Chromatin chemistry
Chromatin metabolism
DNA-Binding Proteins antagonists & inhibitors
DNA-Binding Proteins genetics
Gene Knockdown Techniques
Humans
Ku Autoantigen
Poly (ADP-Ribose) Polymerase-1
Poly(ADP-ribose) Polymerase Inhibitors
Antigens, Nuclear metabolism
DNA Breaks, Double-Stranded
DNA End-Joining Repair
DNA-Binding Proteins metabolism
Poly(ADP-ribose) Polymerases metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1362-4962
- Volume :
- 39
- Issue :
- 22
- Database :
- MEDLINE
- Journal :
- Nucleic acids research
- Publication Type :
- Academic Journal
- Accession number :
- 21880593
- Full Text :
- https://doi.org/10.1093/nar/gkr656