Expression of enzymes involved in chlorophyll catabolism in Arabidopsis is light controlled

Abstract

We found that the levels of mRNA of two enzymes involved in chlorophyll catabolism in Arabidopsis (Arabidopsis thaliana), products of two chlorophyllase genes, AtCLH1 and AtCLH2, dramatically increase (by almost 100- and 10-fold, respectively) upon illumination with white light. The measurements of photosystem II quantum efficiency in 3-(3,4-dichlorophenyl)-1,1-dimethylurea-inhibited leaves show that their expression is not related to photosynthesis but mediated by photoreceptors. To identify the photoreceptors involved, we used various light treatments and Arabidopsis photoreceptor mutants (cry1, cry2, cry1cry2, phot1, phot2, phot1phot2, phyA phyB, phyAphyB). In wild-type Columbia, the amount of transcripts of both genes increase after white-light irradiation but their expression profile and the extent of regulation differ considerably. Blue and red light is active in the case of AtCLH1, whereas only blue light raises the AtCLH2 mRNA level. The fundamental difference is the extent of up-regulation, higher by one order of magnitude in AtCLH1. Both blue and red light is active in the induction of AtCLH1 expression in all mutants, pointing to a complex control network and redundancy between photoreceptors. The blue-specific up-regulation of the AtCLH2 transcript is mediated by cryptochromes and modulated by phototropin1 and phytochromes. Individually darkened leaves were used to test the effects of senescence on the expression of AtCLH1 and AtCLH2. The expression profile of AtCLH1 remains similar to that found in nonsenescing leaves up to 5 d after darkening. In contrast, the light induction of AtCLH2 mRNA declines during dark treatment. These results demonstrate that the expression of enzymes involved in chlorophyll catabolism is light controlled.

Figure 1.

The effect of white light (3 h irradiation with 120 μmol m⁻² s⁻¹) on the level of the AtCLH1 and AtCLH2 transcripts in Arabidopsis leaves (4- to 6-week-old plants). A shows relative mRNA levels of AtCLH1 and AtCLH2 in the dark-adapted (black bars) and in the irradiated leaves (gray bars). B and C are the expression of AtCLH1 and AtCLH2, respectively; black bars: effects of 200 μm DCMU in 0.4% DMSO; gray bars: effects of 0.4% DMSO (control). Each bar represents an average of measurements for at least seven leaves from different plants. All qRT-PCR reactions were run in duplicate. Error bars indicate se.

Figure 2.

Light regulation of AtCLH1 and AtCLH2 transcript levels in photoreceptor-deficient Arabidopsis plants. The relative level of AtCLH1 (A) and AtCLH2 (B) mRNA in dark-adapted leaves (black bars), and after 3 h irradiation with 40 μmol m⁻² s⁻¹ of blue (white bars) or red light (gray bars). The values obtained for wild-type plants are additionally marked with stripes. The photoreceptor mutants used are specified under the respective bars. Each bar represents an average of measurements for at least 10 leaves from different plants pooled together. All qRT-PCR reactions were run in triplicate. Error bars indicate se.

Figure 3.

The expression of AtCLH1 and AtCLH2 during dark-induced leaf senescence. Individual leaves were wrapped in black paper and kept in darkness for the number of days as specified under the bars, while the plant was grown in standard photoperiod. The relative level of AtCLH1 (A) and AtCLH2 (B) in darkened leaves (black bars) and after 3 h of 120 μmol m⁻² s⁻¹ white-light irradiation (gray bars). The dark levels are indicated above the respective bars as percentage of the value for day 1. Each bar represents an average of measurements for at least six leaves coming from different plants. All qRT-PCR reactions were run in duplicate. Error bars indicate se.