European Journal of Biochemistry 1981-12-01

The effect of inhibition of (ADP-ribose)n biosynthesis on DNA repair assayed by the nucleoid technique.

B W Durkacz, S Shall, J Irwin

Index: Eur. J. Biochem. 121(1) , 65-9, (1981)

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Abstract

DNA damage and repair was assayed by the loss and restoration of DNA supercoiling in nucleoids. This technique was used to assess the effects of inhibition of (ADP-ribose)n biosynthesis by 3-aminobenzamide on the capacity to repair DNA of mouse leukaemia L1210 cells following damage by gamma-irradiation and by the monofunctional alkylating agent, dimethyl sulphate. 3-Aminobenzamide partially inhibits restoration of supercoiling following gamma-irradiation and dimethyl sulphate treatment, but inhibits neither the enzymic incision events leading to breaks in the DNA nor the repair synthesis. This inhibition of repair can be observed at very low doses of damaging agents. These observations confirm and extend the evidence that (ADP-ribose)n biosynthesis is required for efficient cellular recovery from DNA damage. In particular, the nucleoid technique permits the demonstration that 3-aminobenzamide inhibits DNA repair after gamma-radiation; it is not possible to draw this unequivocal conclusion with the data from alkaline sucrose gradients because this technique is too insensitive. 1-beta-D-Arabinofuranosylcytosine, which inhibits semiconservative DNA replication, also retards repair, and this effect can be reversed by the addition of deoxycytidine. The inhibitors of DNA excision repair, arabinosylcytosine, hydroxyurea or 3-aminobenzoamide increase the steady-state number of DNA breaks. Thus, they can be used to enhance even further the sensitivity of the nucleoid assay of repair.