College of Bioscience and Biotechnology, Yangzhou University, Yangzhou 225009, China.
Joint International Research Laboratory of Agriculture and Agri-Product Safety of the Ministry of Education of China, Yangzhou University, Yangzhou 225009, China.
Plant Sci. 2020 Jan;290:110314. doi: 10.1016/j.plantsci.2019.110314. Epub 2019 Oct 17.
Chlorophyllase (CLH), which catalyzes the release of the phytol chain from chlorophyll (Chl), has been long considered to catalyze the first step of Chl degradation. Arabidopsis contains two isoforms of CLH (CLH1 and CLH2), and CLH1 was previously demonstrated to be localized in tonoplast and endoplasmic reticulum, and not be involved in Chl degradation. In contrast, CLH2 possesses a predicted signal-peptide for chloroplast localization, and phylogenetic analysis of CLHs in Arabidopsis and other species also indicate that CLH2 forms a different clade than CLH1. Therefore, the possibility remains that CLH2 is involved in the breakdown of Chl. In the current study, clh mutants lacking CLH2 or both CLH isoforms were analyzed after the induction of senescence. Results indicated that the clh knockout lines were still able to degrade Chl at the same rate as wild-type plants. Transgenic Arabidopsis plants were generated that constitutively expressed either CLH2 or CLH2 fused to a yellow fluorescent protein (YFP). Observations made using confocal microscopy indicated that CLH2-YFP was located external to chloroplasts. Additionally, in overexpression plants, CLH2 was enriched in tonoplast and endoplasmic reticulum fractions following membrane fractionation. Based on the collective data, we conclude that CLH2 is not involved in Chl breakdown during senescence in Arabidopsis.
叶绿素酶（CLH）能够催化叶绿素（Chl）中植醇链的释放，长期以来一直被认为是 Chl 降解的第一步的催化剂。拟南芥含有两种 CLH 同工型（CLH1 和 CLH2），先前的研究表明 CLH1 定位于液泡膜和内质网，不参与 Chl 降解。相比之下，CLH2 具有预测的叶绿体定位信号肽，并且拟南芥和其他物种的 CLHs 的系统发育分析也表明 CLH2 与 CLH1 形成不同的分支。因此，CLH2 参与 Chl 分解的可能性仍然存在。在本研究中，在诱导衰老后分析了缺乏 CLH2 或两种 CLH 同工型的 clh 突变体。结果表明，clh 敲除系仍能够以与野生型植物相同的速率降解 Chl。生成了组成型表达 CLH2 或与黄色荧光蛋白（YFP）融合的 CLH2 的转基因拟南芥植物。使用共聚焦显微镜观察表明，CLH2-YFP 位于叶绿体外部。此外，在过表达植物中，CLH2 在膜分离后的液泡膜和内质网部分中富集。基于这些综合数据，我们得出结论，CLH2 不参与拟南芥衰老过程中的 Chl 分解。
