Inhibition of repair patch ligation by an inhibitor of poly(ADP-ribose) synthesis in normal human fibroblasts damaged with ultraviolet radiation.

ABSTRACT

The effect of inhibiting poly(ADP-ribose) synthesis on DNA excision repair following UV irradiation of cultured normal human fibroblasts was determined under conditions which did not perturb NAD+ concentration. Following UV irradiation, there was a transient increase in DNA strand breaks to a maximum of 800 rad eq of breaks 30 min after damage. 3-Aminobenzamide (5 mM) caused a 50% increase in the maximum number of DNA single strand breaks following damage but did not prevent the decline in strand breaks which normally occurs within the first hour after damage. Addition of 3-aminobenzamide several hours after damage, when most of the strand breaks had disappeared, caused a reaccumulation of strand breaks. 3-Aminobenzamide inhibited ligation of repair patches, as measured by exonuclease III, following damage by UV radiation and the magnitude of the inhibition was sufficient to account for the increases in strand breaks caused by 3-aminobenzamide. UV radiation alone did not lower NAD+ concentrations; however, when the repair synthesis step was inhibited by aphidicolin and hydroxyurea, the number of single strand breaks increased and the NAD+ concentration fell to 11%. 3-Aminobenzamide inhibited this depletion of NAD+ by 80%.