TRANSCRIPTION ABNORMALITIES POTENTIATE APOPTOSIS OF NORMAL HUMAN FIBROBLASTS

Background: Apoptosis is a natural process by which damaged and potent ially tumorigenic cells are removed. Induction of apoptosis is importa nt in chemotherapy aimed at eliminating cancer cells. We address the m echanisms by which this process can be triggered in cells that are rec alcitrant to cell death induced by DNA-damaging agents. Materials and Methods: Normal human fibroblasts and lymphoblasts, and fibroblasts wi th defined genetic changes, were treated with DNA-damaging agents and inhibitors of transcription. Western blotting was used to study the ex pression of some of the key factors involved in the response to DNA da mage and the induction of apoptosis, namely, p53, p21(WAF1,Cip1) Mdm2, Bax, and CD95 (Fas/APO1). Apoptosis was followed by various criteria, including DNA fragmentation, specific proteolysis, cell morphology, v iability, and FAGS scan for sub-G1 cells. Results: Normal human fibrob lasts were more resistant than lymphoblasts to DNA damage-induced apop tosis. The DNA-damaging agents mitomycin C and cisplatin induced rapid apoptosis of fibroblasts with defects in the repair of transcribed DN A, compared with wild-type cells or those with defects in overall geno me repair. Shortterm treatment with inhibitors of RNA polymerase IT tr anscription, actinomycin D, and alpha-amanitin induced rapid cell deat h of normal fibroblasts. These results show that there is a link betwe en defective transcription and apoptosis. Treatments and genetic backg rounds that favored apoptosis were associated with efficient and prolo nged induction of p53 and often altered or imbalanced expression of it s downstream effectors p21(WAF1,Cip1) and Mdm2, whereas there were no changes in Bax or CD95 (Fas/APO1). Conclusion: Transcription inhibitor s increase p53 levels and are better inducers of apoptosis than DNA-da maging agents in some cell types;Apoptosis might be triggered by block ed polymerases and/or faulty expression of downstream effecters.