Expression of the human Bcl-2 increases resistance of Ewing's sarcoma cells to apoptosis and inhibits poly(ADP-ribose) polymerase cleavage induced by radiation

Abstract

We reported previously that Ewing's sarcoma (ES) cells respond to ionizing radiation exposure by arrest in G(2)/M phase and induction of apoptosis which occurs in conjunction with poly(ADP-ribose) polymerase (PARP) proteolytic cleavage. ES cells (A4573 cell line) do not express immunodetectable levels of Bcl-2. To determine if expression of Bcl-2 could modulate radiation-induced ES cell death, we have stably transfected A4573 cells with a full-length human bcl-2 cDNA. Expression of Bcl-2 protein rendered ES cells relatively resistant to radiation-induced apoptosis. Moreover, the anti-apoptotic activity of Bcl-2 was directly related to levels of its expression in different ES clones. Cell cycle characteristics were similar for both parental and Bcl-2 expressing ES cells following radiation treatment, although bcl-2 transfectants exhibited a more protracted G(2)/M phase arrest and lower rate of apoptosis after release from the block. Constitutive expression of Bcl-2 resulted in about two-fold inhibition of PARR cleavage in ES cells dying after ionizing radiation exposure. These data support a role for Bcl-2 protein as a negative regulatory element of PARP proteolysis at the early stages of radiation-induced apoptosis in ES cells.