Poly(ADP-ribose) Glycohydrolase Deficiency Sensitizes Mouse ES Cells to DNA Damaging Agents

H.      Fujihara; H.      Ogino; D.      Maeda; H.      Shirai; T.      Nozaki; N.      Kamada; K.      Jishage; S.      Tanuma; T.      Takato; T.      Ochiya; T.      Sugimura; M.      Masutani

Abstract

Poly(ADP-ribose) glycohydrolase (Parg) is the main enzyme for degradation of poly(ADP-ribose) by splitting ribose-ribose bonds. Parg-deficient (Parg+/- and Parg-/-) mouse ES cell lines have been established by disrupting both alleles of Parg exon 1 through gene-targeting. A transcript encoding a full length isoform of Parg was eliminated and only low amounts of Parg isoforms were detected in Parg-/- embryonic stem (ES) cells. Poly(ADP-ribose) degradation activity was decreased to one-tenth of that in Parg+/+ ES cells. Parg-/- ES cells exhibited the same growth rate as Parg+/+ ES cells in culture. Sensitivity of Parg-/- ES cells to various DNA damaging agents, including an alkylating agent dimethyl sulfate, cisplatin, gemcitabine, 5-fluorouracil, camptothecin, and γ-irradiation was examined by clonogenic survival assay. Parg-/- ES cells showed enhanced lethality after treatment with dimethyl sulfate, cisplatin and γ-irradiation compared with wildtype (Parg+/+) ES cells (p < 0.05, respectively). In contrast, a sensitization effect by Parg-deficiency was not observed with gemcitabine and camptothecin. These results suggest the possibility that functional inhibition of Parg leads to sensitization of tumor cells to some chemo- and radiation therapies.

Keywords: Poly(ADP-ribose) glycohydrolase, Knockout, ES cell, DNA damaging agent, Alkylating agent, γ-irradiation, cisplatin, 5-fluorouracil