GROWTH-INHIBITION AND CELL-KILLING BY N-METHYL-N-NITROSOUREA - METABOLIC ALTERATIONS THAT ACCOMPANY POLY(ADP-RIBOSYL)ATION

The effect of inhibition of poly(ADP-ribose) polymerase (PARP) on the growth arrest and cell killing induced by N-methyl-N-nitrosourea (MNU) was studied in L929 fibroblasts. Depletion of NAD and ATP preceded th e cell killing by a 1-h exposure to 10 or 15 mM MNU. 3-Aminobenzamide (ABA), an inhibitor of PARP, spared the depletion of NAD and ATP and p revented the cell killing. With 5 mM MNU, a depletion of NAD was promp tly reversed, and there was no loss of ATP and no cell death. Aphidico lin, a DNA polymerase inhibitor, prevented the restoration of NAD, wit h resulting depletion of ATP and death of the cells, effects that were prevented by ABA, Azide together with 2-deoxyglucose depleted ATP, fo llowed by a loss of NAD and cell death, changes that occurred in the a bsence of DNA single strand breaks (DNA SSB). ABA prevented the deplet ion of NAD, but not that of ATP, nor the cell killing. MNU (2.5 mM) in hibited cell growth without effect on the viability of the cells. ABA potentiated the cell growth inhibition. Thus, inhibition of PARP poten tiates cell growth inhibition by limiting DNA repair mechanisms. Alter natively inhibition of the DNA repair response to more extensive DNA d amage prevents cell killing. The ATP depletion caused by poly(ADP-ribo syl)ation, rather than DNA SSB and the loss of NAD, is the more critic al event in the cell killing. (C) 1995 Academic Press, Inc.