Tobacco plants expressing T4 endonuclease V show enhanced sensitivity to ultraviolet light and DNA alkylating agents

Abstract

DNA repair processes and UV-filtering pigments protect organisms from the cytotoxicity of UV light and endow plants with a high degree of natural UV resistance. In an attempt to further enhance this UV resistance we have constructed transgenic tobacco lines that express a DNA repair enzyme encoded by the bacteriophage T4 denV gene. The denV gene encodes endonuclease V, an enzyme which initiates base excision repair of cyclobutane pyrimidine dimers. Its presence is expected to provide transgenotes with a repair pathway complementary to, but likely distict from, the repair pathways found in tobacco. The denV gene, flanked by a CaMV 35S promoter and poly(A) addition site, was introduced into tobacco and mature plants regenerated. The transgenotes expressed high levels of a UV-specific endonuclease and no such activity was found in control plants. Curiously, assays which detected several different biological endpoints showed that the denV + transgenotes were also hypersensitive to UV-C light. This hypersensitivity segregated with the denV gene and was not caused by altered concentrations of UV-filtering pigments. Moreover, the denV + transgenotes were also hypersensitive to alkylating agents. This odd phenotype is most readily explained if endogenous plant DNA repair systems cannot tolerate the high levels of baseless lesions that would be generated by a transgenically expressed β-eliminating lyase such as endonuclease V.