Effects of increased UV-B radiation on the lichen Cladonia arbuscula spp. mitis: UV-absorbing pigments and DNA damage

Abstract: Lichens, a form of symbiosis between a fungal and a photosynthetic partner, are believed to be a group that evolved early in the history of terrestrial life. As a consequence, they probably experienced higher ultraviolet (UV) radiation fluxes than at present. However, in the last two decades, due to the depletion of the stratospheric ozone layer, increasing levels of ultraviolet-B radiation (280-315 nm) are reaching the earth€ surface. In this thesis the effects of enhanced UV-B radiation were studied on the apical (€tips€) and central (€stems€) regions of the podetial thallus of the lichen Cladonia arbuscula ssp. mitis. Effects of UV-B radiation on lichen DNA were investigated by measuring the accumulation of cyclobutane pyrimidine dimers (CPDs) by means of the enzyme-linked immunosorbant assay technique (ELISA). ELISA was also used for assessing the capacity of lichens to repair DNA damage by measuring the decrease in CPD content. UV-absorbing pigment content and changes in penetration of light within the tissue were investigated by means of spectrophotometric measurements of lichen acetone-ethanolic extracts and fibre optic measurements of internal radiation gradients, respectively. After 7 days of exposure to high light (HL; 800 _mol m-2 s-1) with or without enhanced UV-B radiation (13 kJ m-2 day-1) UV-absorbing pigments and CPDs accumulated in both tips and stems of air-dry lichens. However, stems showed a larger increase in total phenolic content and lower CPD accumulation than tips. In an outdoor experiment, after three months exposure to natural sunlight, where the UV-B component had been removed or where supplemental UV-B radiation had been added (simulating 15% ozone depletion), the highest increases in pigment content were found in stems of those lichens grown under supplemental UV-B radiation. Fibre optic measurements showed a decreased light penetration (280 nm) within the thallus of lichens grown under enhanced UV-B radiation. Short-term exposure to UV-B radiation (7.4 kJ m-2 day-1) of hydrated lichen thalli kept at either 25oC or at 2oC resulted in the accumulation of CPDs. Subsequent exposure to photosynthetically active radiation (PAR, 300 _mol m-2 s-1, 400-700 nm) decreased the CPD content only in hydrated lichen thalli kept at 25oC, whereas there was no change in thalli kept at 2oC. Exposure to UV-B radiation in combination with PAR resulted in the absence of accumulation of CPDs in hydrated lichen thalli kept at 25 oC, whereas in hydrated thalli kept at 2oC CPD levels were as high as after exposure to UV-B only. Throughout all experiments stems showed consistently lower levels of CPDs and a higher degree of repair than tips. Monitoring of photolyase repairing activity also showed that a higher percentage of pyrimidine dimers was repaired in stems than in tips. From the results it was evident that water content and temperature represent two important factors in the response of Cladonia arbuscula ssp. mitis to UV-B radiation, since air-dry thalli were more susceptible to UV-induced DNA damage and less capable of repairing CPDs than hydrated thalli. Further, hydrated thalli kept at low temperatures were not able to repair the accumulated DNA damage. An apparent protective mechanism against UV radiation in air-dry lichens was the accumulation of UV-absorbing pigments, which consequently reduced the penetration of light within the thallus. Tips have shown to be those thallus parts most susceptible to damage caused by UV-B radiation as they accumulated the highest amounts of DNA damage, exhibited the least repair capacity and the lowest relative increase in phenolic pigment content.