p53 in SV40-transformed DNA-damaged human cells binds to its cognate sequence but fails to transactivate target genes

Human SV40-transformed cells contain high levels of stabilized p53 of which only a fraction is complexed with the SV40 large tumor antigen (T-antigen) . This raises the question whether the p53 which is not complexed with T-an tigen retains some biological activity. Two human SV40-transformed cell lin es, BEAS and SV80, were investigated. A significant level of constitutive c ognate-sequence-specific DNA-binding of p53 was detected by electrophoretic mobility shift assay (EMSA.) of cell extracts. Upon DNA damage by treatmen t with mitomycin C the DNA-binding activity was increased, as known for cel ls with wild-type p53. However, in both cell lines, before and after DNA da mage, p53 was not able to transactivate a target gene as shown by reporter gene assay. Hence, the capability of p53 to bind its cognate sequence is a prerequisite but no proof of p53 transactivating activity. Nuclear p53 leve ls were not further increased after mitomycin C treatment, occasionally rat her slightly decreased, often accompanied by an even larger decrease in amo unt of T-antigen. In conclusion, SV40-transformation of human cells has cau sed a loss of essential features of wild-type p53 activity, even in that fr action of p53 not in physical complex with SV40 T-antigen.