DNA double-strand breaks trigger apoptosis in p53-deficient fibroblasts

DNA double-strand breaks (DSBs) are induced by ionizing radiation (IR) and various radiomimetic agents directly, or indirectly as a consequence of DNA repair, recombination and replication of damaged DNA, They are ultimately involved in the generation of chromosomal aberrations and were reported to cause genomic instability, gene amplification and reproductive cell death. To address the question of whether DSBs act as a trigger of apoptosis, we i nduced DSBs by means of restriction enzyme electroporation and compared the effect with IR in mouse fibroblasts that differ in p53 status [wild-type ( +/+) versus p53-deficient (-/-) cells]. We show that (i) electroporation of PvuII is highly efficient in the induction of DSBs, (ii) electroporation o f PvuII increases the rate of apoptosis, but not of necrosis in p53-/- cell s, (iii) treatment with gamma -rays induces both apoptosis and necrosis in p53-/- cells, (iv) the frequency of DSBs correlates with the yield of apopt osis and (v) both PvuII and gamma -ray treatment reduce the level of anti-a poptotic Bcl-2 protein in p53-/- cells whereas the level of Bar remains una ltered. Cells expressing wild-type p53 were more resistant than p53-deficie nt cells as to the induction of apoptosis and did not show Bcl-2 decline up on treatment with PvuII and gamma -rays, The data provide evidence that blu nt-ended DSBs induced by restriction enzyme PvuII act as a highly efficient trigger of apoptosis, but not of necrosis, This process is related to Bcl- 2 decline and does not require p53.