Millisecond UV-B irradiation evokes prolonged elevation of cytosolic-free Ca2+ and stimulates gene expression in transgenic parsley cell cultures

Chalcone synthase (CHS) is a key enzyme leading to the generation of protec tive flavonoids in plants under environmental stress. Expression of the CHS gene is strongly upregulated by exposures to UV light, a response also obs erved in heterotrophic parsley cell cultures. Although there are hints that the stimulus for CHS expression may be coupled to UV-B irradiation through a rise in cytosolic-free Ca2+ ([Ca2+](i)), the temporal relationship of th ese events has never been investigated critically. To explore this question , we have used a CHS promoter/luciferase (CHS/LUC) reporter gene fusion and recorded its expression and [Ca2+](i) elevation in a transgenic parsley ce ll culture following millisecond light pulses. Luciferase expression was en hanced maximally seven- (+/- 2) fold by 30 10ms flashes of UV-B light. The response was specific to wavelengths of 300-330 nm and could be inhibited i n the presence of the Ca2+ channel blocker nifedipine. In parallel measurem ents, using Fura-2 fluorescence ratio microphotometry, we found that 10ms U V-B flashes also evoked it gradual and prolonged rise of [Ca2+](i) in the p arsley cells which was irreversible within the timescale of these experimen ts, but could be prevented by prior treatment with nifedipine. These, and a dditional results, indicate a remarkably high temporal sensitivity to, and specificity for, UV-B light in CHS gene expression independent of UV-mediat ed DNA damage by thymine dimerization. The ability of transient UV-B stimul ation to evoke prolonged elevations of [Ca2+](i) suggests a functional coup ling between the initial light stimulus and subsequent gene expression that takes place many tons of minutes later.