Arahidopsis CHLOROPHYLLASE 1 protects young leaves from long-term photodamage by facilitating FtsH-mediated D1 degradation in photosystemⅡrepair

The proteolytic degradation of the photodamaged D1 core subunit during the photosystemⅡ(PSⅡ)repair cycle is well understood,but chlorophyll turnover during D1 degradation remains unclear.Here,we report that Arabidopsis thaliana CHLOROPHYLLASE 1(CLH1)plays important roles in the PSII repair process.The abundance of CLH1 and CLH2 peaks in young leaves and is induced by high-light exposure.Seedlings of clh1 single and clh1-1/2-2 double mutants display increased photoinhibition after long-term high-light exposure,whereas seedlings overexpressing CLH1 have enhanced light tolerance compared with the wild type.CLH1 is localized in the developing chloroplasts of young leaves and associates with the PSⅡ-dismantling complexes RCC1 and RC47,with a preference for the latter upon exposure to high light.Furthermore,degradation of damaged D1 protein is retarded in young clh1-1/2-2 leaves after 18-h highlight exposure but is rescued by the addition of recombinant CLH1 in vitro.Moreover,overexpression of CLH1 in a variegated mutant(var2~2)that lacks thylakoid protease FtsH2,with which CLH1 interacts,suppresses the variegation and restores D1 degradation.A var2-2 clh1-1/2-2triple mutant shows more severe variegation and seedling death.Taken together,these results establish CLH1 as a long-sought chlorophyll dephytylation enzyme that is involved in PSⅡrepair and functions in long-term adaptation of young leaves to high-light exposure by facilitating FtsH-mediated D1 degradation.

AtCLH2, a Typical but Possibly Distinctive Chlorophyllase Gene in Arabidopsis

Chlorophyllase （EC 3.1.1.14） is involved in the first step of chlorophyll degradation. Isolation of chlorophyllase genes greatly facilitates characterization of chlorophyllase properties and elucidation of molecular regulation of their in vivo activities. There are two chlorophyllase genes, AtCLH1 and AtCLH2, in Arabidopsis thaliana. The in vivo roles of AtCLH1 have been reported previously. However, few studies have been carried out on AtCLH2. Here, we show that purified recombinant Chlase2, encoded by AtCLH2, exhibits in vitro chlorophyllase activity. Interestingly, ＂activation＂ of in vitro activity of the recombinant Chlase2 required higher concentrations of a detergent or a polar solvent. To determine its activity in vivo, the expression of AtCLH2 was inhibited by RNA interference. RNAi plants showed decreased contents of chlorophyllide without a substantial change in the total amount of the extractable chlorophyll and consequently presented lower chlorophyllide to chlorophyll ratios in their leaves. In addition, the two AtCLHs exhibited differential expression patterns. Our results suggest that AtCLH2 might play a distinctive role in chlorophyll catabolism in vivo.