Background: The tumor suppressor protein p53 which can mediate an ionizing radiation-induced G1 arrest in mammalian cells, forms complexes with SV40 large T antigen (L-T-Ag). We have analyzed the p53 levels, the capability to undergo a G1 arrest and the radiosensitivity of two SV40 transformed fibroblast strains differing in their large T antigen expression.
Material and methods: One of the two strains (VA13F) is the commercially available form of Wi38VA13, the other (VA13E) arose spontaneously from the original one in our laboratory. Their p53 levels were measured by means of flow cytometry (FCM) and Western blot (WB) with two p53 antibodies (Ab-3, clone PAb240; Ab-6, clone DO-1; both Oncogene Science). Cell cycle distributions were determined flow cytometrically after BrdU labeling at regular time intervals after exposure to 250 kV X-rays. Radiosensitivity was assessed in a clonogenicity assay.
Results: The p53 levels of the two strains corresponded to their large T antigen expression, presumably due to complex formation between the two proteins. The strain with a high p53 level did not show a G1 arrest and had a relatively high radiosensitivity, whereas the strain with a low p53 level showed a significant G1 arrest and a lower radiosensitivity.
Conclusion: These results suggest that 1. complex formation between the large T antigen and p53 reduces the latter's functionality; 2. in these two strains the G1 arrest is one of the factors determining radiosensitivity.