Dinitrophenol inhibits the rejoining of radiation-induced DNA breaks by L-cells

The production and rejoining of X-ray-induced single-stranded DNA breaks was studied using the alkaline sucrose density gradient technique and by measuring the disappearance of both 5' termini and 3'-OH termini using polynucleotide kinase and DNA polymerase, respectively. All studies were conducted using L-cell suspensions irradiated both in the presence and absence of 2,4-dinitrophenol (DNP), an uncoupler of oxidative phosphorylation. Results show that the induction of single-stranded DNA breaks probably includes a nucleolytic component in addition to indirect free radical effects. A greater number of breaks were produced in the absence of DNP, suggesting that depressed adenosine triphosphate (ATP) levels reduce endogenous nucleolytic activity. The rejoining mechanism is enzymatic and requires an available ATP supply for operation. In the presence of DNP no DNA rejoining was observed following 30 min incubation after 10,000 rad. These results suggest that DNA breaks produced may be characterized by 5'-PO(4)-3'-OH termini and are rejoined by DNA ligase.