The populations of many amphibian species, in widely scattered habitats, appear to be in severe decline; other amphibians show no such declines. There is no known single cause for the declines, but their widespread distribution suggests involvement of global agents-increased UV-B radiation, for example. We addressed the hypothesis that differential sensitivity among species to UV radiation contributes to these population declines. We focused on species-specific differences in the abilities of eggs to repair UV radiation damage to DNA and differential hatching success of embryos exposed to solar radiation at natural oviposition sites. Quantitative comparisons of activities of a key UV-damage-specific repair enzyme, photolyase, among oocytes and eggs from 10 amphibian species were reproducibly characteristic for a given species but varied >80-fold among the species. Levels of photolyase generally correlated with expected exposure of eggs to sunlight. Among the frog and toad species studied, the highest activity was shown by the Pacific treefrog (Hyla regilla), whose populations are not known to be in decline. The Western toad (Bufo boreas) and the Cascades frog (Rana cascadae), whose populations have declined markedly, showed significantly lower photolyase levels. In field experiments, the hatching success of embryos exposed to UV radiation was significantly greater in H. regkla than in R. cascadae and B. boreas. Moreover, in R. cascadae and B. boreas, hatching success was greater in regimes shielded from UV radiation compared with regimes that allowed UV radiation. These observations are thus consistent with the UV-sensitivity hypothesis.

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Removal of cyclobutane pyrimidine dimers (CBPDs) in vivo from the DNA of UV-irradiated eight-leaf seedlings of Arabidopsis thaliana was rapid in the presence of visible light (half-life about 1 hour); removal of CBPDs in the dark, presumably via excision repair, was an order of magnitude slower. Extracts of plants contained significant photolyase in vitro, as assayed by restoration of transforming activity to UV-irradiated Escherichia coii plasmids; activity was maximal from four-leaf to 12-leaf stages. UV-B treatment of seedlings for 6 hours increased photolyase specific activity in extracts twofold. Arabidopsis photolyase was markedly temperature-sensitive, both in vitro (half-life at 300C about 12 minutes) and in vivo (half-life at 300C, 30 to 45 minutes).The wavelength dependency of the photoreactivation cross-section showed a broad peak at 375 to 400 nm, and is thus similar to that for maize pollen; it overlaps bacterial and yeast photolyase action spectra.Stratospheric ozone depletion projected for the early 21st century is expected to increase the amount of DNA-damaging UV-B (290-320 nm) radiation at the earth's surface by as much as 20 to 50% (2, 10, 33). This has renewed interest in the UV-resistance mechanisms of plants, which will be exposed continually to increased UV-B fluxes.Repair of ultraviolet-light-induced DNA damage is a feature of life at all levels of complexity. Studies of bacteria (particularly Escherichia coli), yeast, and mammalian cells have elucidated four distinct mechanisms by which cells cope with UV-damaged DNA (for reviews see refs. 9, 29). In photoreactivation, a single enzyme, photolyase, uses light energy in the 300 to 500 nm range to reverse a specific class of UV photoproducts-cis, syn CBPDs.2 In excision repair, an ensemble of proteins act together to remove a wide variety of helix-distorting lesions, including at least two UV photoproducts-CBPDs and pyrimidine-(6-4)-pyrimidone photoproducts (6-4 photoproducts). Photoproducts that block DNA replication are tolerated, rather than repaired,

Photolyases are DNA repair enzymes that use energy from blue light to repair pyrimidine dimers. We report the isolation of an Arabidopsis thaliana mutant (uvr2-1) that is defective in photorepair of cyclobutylpyrimidine dimers (CPDs). Whereas uvr2-1 is indistinguishable from wild type in the absence of UV light, low UV-B levels inhibit growth and cause leaf necrosis. uvr2-1 is more sensitive to UV-B than wild type when placed under white light after UV-B treatment. In contrast, recovery in darkness or in light lacking photoreactivating blue light results in equal injury in uvr2-1 and wild type. The uvr2-1 mutant is unable to remove CPDs in vivo, and plant extracts lack detectable photolyase activity. This recessive mutation segregates as a single gene located near the top of chromosome 1, and is a structural gene mutation in the type II CPD photolyase PHR1. This mutant provides evidence that CPD photolyase is required for plant survival in the presence of UV-B light.

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