DNA-damage-repair/toleration (DRT) activities defend cells against killing and mutagenesis by UV light and other agents. The mechanisms by which expression of mammalian DRT genes is stimulated by DNA-damaging agents, and modulated in various tissues, remain poorly understood. The plant Arabidopsis thaliana, in which four DNA-damage-induced DRT genes have recently been described, provides a genetically amenable and experimentally convenient model system for analysis of DRT-gene expression in whole differentiated organisms. The UVB-induced, tissue- specific and growth-stage-specific expression of genes encoding such plant flavonoid-biosynthesis enzymes as chalcone synthase (CHS) and chalcone isomerase (CHI) provides a well-studied paradigm of regulated gene expression. The applicant will, in the sponsor's laboratory, analyze UV-induced CHS and CHI gene expression, using techniques that he will subsequently employ to study DRT gene regulation in his own laboratory. Specifically, the applicant will construct mutant Arabidopsis CHS and CHI promoters, using previous analysis of parsley CHA promoter elements as a guide, fuse the mutant promoters to a reporter gene (GUS), measure transient expression in Arabidopsis protoplasts, and construct transgenic plants encoding promoter::GUS for analysis of tissue-specific and growth-stage-modulated expression.
