title: Overexpression of human poly(ADP-ribose) polymerase in transfected hamster cells leads to increased poly(ADP-ribosyl)ation and cellular sensitization to gamma irradiation.
abstract: Poly(ADP-ribosyl)ation is a posttranslational modification of nuclear proteins catalyzed by poly(ADP-ribose) polymerase (PARP), an enzyme which uses NAD+ as substrate. Binding of PARP to DNA single-strand or double-strand breaks leads to enzyme activation. Inhibition of poly(ADP-ribose) formation impairs the cellular recovery from DNA damage. Here we describe stable transfectants of the Chinese hamster cell line CO60 that constitutively overexpress human PARP (COCF clones). Immunofluorescence analysis of gamma-irradiation-stimulated poly(ADP-ribose) synthesis revealed consistently larger fractions of cells positive for this polymer in the COCF clones than in control clones, which failed to express human PARP. HPLC-based quantitative determination of in vivo levels of poly(ADP-ribose) confirmed this result and revealed that the basal polymer levels of undamaged cells were significantly higher in the COCF clones. The COCF clones were sensitized to the cytotoxic effects of gamma irradiation compared with control transfectants and parental cells. This effect could not be explained by depletion of cellular NAD+ or ATP pools. Together with the well-known cellular sensitization by inhibition of poly(ADP-ribosyl)ation, our data lead us to hypothesize that an optimal level of cellular poly(ADP-ribose) accumulation exists for the cellular recovery from DNA damage.
authors.affiliation: Deutsches Krebsforschungszentrum, Forschungsschwerpunkt Angewandte Tumorvirologie, Unité INSERM U375, Heidelberg, Germany.
authors.forename: L
authors.initials: L
authors.lastname: Van Gool
authors.forename: R
authors.initials: R
authors.lastname: Meyer
authors.forename: E
authors.initials: E
authors.lastname: Tobiasch
authors.forename: C
authors.initials: C
authors.lastname: Cziepluch
authors.forename: J C
authors.initials: JC
authors.lastname: Jauniaux
authors.forename: A
authors.initials: A
authors.lastname: Mincheva
authors.forename: P
authors.initials: P
authors.lastname: Lichter
authors.forename: G G
authors.initials: GG
authors.lastname: Poirier
authors.forename: A
authors.initials: A
authors.lastname: Bürkle
authors.forename: J H
authors.initials: JH
authors.lastname: Küpper
mesh_terms: D000246:Adenosine Diphosphate Ribose / Q000378:metabolism* / Q000528:radiation effects*; D000255:Adenosine Triphosphate / Q000528:radiation effects; D000818:Animals; D016466:CHO Cells / Q000201:enzymology* / Q000378:metabolism / Q000528:radiation effects; D002470:Cell Survival / Q000528:radiation effects; D006224:Cricetinae; D005720:Gamma Rays; D006801:Humans; D009243:NAD / Q000528:radiation effects; D011065:Poly(ADP-ribose) Polymerases / Q000096:biosynthesis* / Q000235:genetics / Q000528:radiation effects; D014162:Transfection
chemical_list: D009243:NAD; D000246:Adenosine Diphosphate Ribose; D000255:Adenosine Triphosphate; D011065:Poly(ADP-ribose) Polymerases