UV-B-Inducible and Constitutive Genes that Mediate Repair and Toleration Of UV-Damaged DNA in the Plant Arabidopsis Thaliana

Abstract

Plant responses to UV-induced DNA damage are of particular interest because plants are exposed almost continuously to the sunlight on which they depend for energy and developmental signals. The small crucifer Arabidopsis thaliana is an ideal model plant for genetic and molecular biological studies of DNA-damage-repair/toleration (DRT) responses. We previously showed that Arabidopsis plants primarily used photoreaction to remove cyclobutane pyrimidine dimers (CPDs), the major UV photoproducts in DNA, from their genomes; excision repair of CPDs was about 5% as efficient. Arabidopsis photoreactivating enzyme (photolyase) levels are developmentally regulated (more activity in midlife than young plants), and inducible by UV treatment of plants.

To gain information about the range of DRT activities in Arabidopsis, and to obtain DRT gene probes for regulatory studies, we recently selected for Arabidopsis cDNAs, in an expression library, that partially corrected the phenotypes of E. coli mutants lacking repair/toleration activities. Of the six cDNAs isolated, the products of three—DRT 100, DRT111 and DRT112—appear to catalyze either strand-exchange or resolution steps of homologous recombination; in plants, these activities may mediate recombinational toleration of unrepaired damage. DRT101 and DRT102 may encode UV-specific excision repair activities. Four of the DNA sequences—DRT100, 101, 111 and 112—predict chloroplast-targeted proteins.

We now find levels of DRT100 and DRT101 mRNA to be increased three- to four-fold by UV irradiation of whole plants. DRT 100 and DRT 101 induction patterns differ from one another and from the pattern for the UV-inducible CHS gene (encodes the flavonoid-biosynthesis enzyme chalcone synthase): 1) all three genes are induced by UV light, only DRT 100 and DRT 101 by the DNA crosslinking agent Mitomycin C, and only DRT 100 by the alkylating agent methylmethane sulfonate; 2) CHS and DRT 101 mRNA levels are maximal immediately after a 2-h UV-B pulse, whereas DRT100 mRNA peaks 3-h later; 3) a UV-B pulse induces CAS mRNA maximally in 7-day-old plants, and much less in older plants, whereas DRT100 and DRT101 show higher inducibility at 14 days and 21 days. This last result, and the developmental pattern of photolyase expression, suggest that older Arabidopsis plants may rely more on DRT activities, and perhaps less on shielding by flavonoids, than younger ones.