Down-regulation of DNA repair synthesis at DNA single-strand interruptions in poly(ADP-ribose) polymerase-1 deficient murine cell extracts.
The functional involvement of poly(ADP-ribose) polymerase-1 (PARP-1) in the repair of DNA single- and double-strand breaks, DNA base damage, and related repair substrate intermediates remains unclear. Using an in vitro DNA repair assay and cell extracts derived from PARP-1 deficient or wild-type murine embryonic fibroblasts, we investigated the DNA synthesis and ligation steps associated with the rejoining of DNA single-strand interruptions containing 3'-OH, and either 5'-OH or 5'-P termini. Complete repair leading to DNA rejoining was similar between PARP-1 deficient cells and wild-type controls and poly(ADP-ribose) synthesis was, as expected, greatly reduced in PARP-1 deficient cell extracts. The incorporation of [32P]dCMP into repaired DNA at the site of a lesion was reduced two-three-fold in PARP-1 deficient cell extracts, demonstrating a decrease in repair patch size. Addition of purified PARP-1 to levels approximating those present in wild-type extracts did not stimulate DNA repair synthesis. We conclude that PARP-1 is not required for the efficient processing and rejoining of single-strand interruptions with defined 3'-OH and 5'-OH or 5'-P termini. Decreased DNA repair synthesis observed in PARP-1 deficient cell extracts is associated with reduced cellular expression of several factors required for long-patch base excision repair (BER), including FEN-1 and DNA ligase I.
Sanderson
Russell J
RJ
Cancer Research UK London Research Institute, Clare Hall Laboratories, South Mimms, Hertfordshire EN6 3LD, UK.
Lindahl
Tomas
T
D000818:Animals; D004249:DNA Damage / Q000235:genetics; D004260:DNA Repair / Q000235:genetics* / Q000502:physiology; D015536:Down-Regulation / Q000235:genetics* / Q000502:physiology; D005347:Fibroblasts; D051379:Mice; D018345:Mice, Knockout; D011065:Poly(ADP-ribose) Polymerases / Q000172:deficiency / Q000235:genetics*
D011065:Poly(ADP-ribose) Polymerases