Nakagawara Eiki, Sakuraba Yasuhito, Yamasato Akihiro, Tanaka Ryouichi, Tanaka Ayumi
Institute of Low Temperature Science, Hokkaido University, N19 W8 Kita-ku, Sapporo, Japan.
Plant J. 2007 Mar;49(5):800-9. doi: 10.1111/j.1365-313X.2006.02996.x. Epub 2007 Feb 9.
Chlorophyll b is one of the major light-harvesting pigments in green plants and it is essential for optimal light harvesting. Chlorophyll b is synthesized from chlorophyll a by chlorophyllide a oxygenase (CAO) which consists of A, B and C domains. Previously, we demonstrated that the C domain alone has a catalytic function, while the A and B domains control the level of CAO protein in response to chlorophyll b accumulation. We hypothesized that the accumulation of chlorophyll b triggers the proteolytic degradation of CAO. In this study, in order to gain further insight into this regulatory mechanism we screened for mutants that have defects in the control of CAO accumulation. Seeds from a transgenic line of Arabidopsis which overexpressed a CAO-GFP fusion were mutagenized and their progenies were screened by laser-scanning confocal microscopy for mutants showing an elevated level of GFP fluorescence. One particular mutant (dca1) exhibited stronger GFP fluorescence and accumulated a GFP-CAO fusion protein at a higher level. Concomitantly, the chlorophyll a to b ratio decreased in this mutant. The mutation in the dca1 mutant was mapped to the ClpC1 gene, thereby indicating that a chloroplast Clp protease is involved in regulating chlorophyll b biosynthesis through the destabilization of CAO protein in response to the accumulation of chlorophyll b.
叶绿素b是绿色植物中主要的捕光色素之一，对于最佳的光捕获至关重要。叶绿素b由叶绿素a通过叶绿素酸酯a加氧酶（CAO）合成，该酶由A、B和C结构域组成。此前，我们证明单独的C结构域具有催化功能，而A和B结构域则根据叶绿素b的积累来控制CAO蛋白的水平。我们推测叶绿素b的积累会触发CAO的蛋白水解降解。在本研究中，为了进一步深入了解这种调控机制，我们筛选了在CAO积累控制方面存在缺陷的突变体。对过表达CAO-GFP融合蛋白的拟南芥转基因系的种子进行诱变，并通过激光扫描共聚焦显微镜对其后代进行筛选，以寻找GFP荧光水平升高的突变体。一个特定的突变体（dca1）表现出更强的GFP荧光，并积累了更高水平的GFP-CAO融合蛋白。与此同时，该突变体中叶绿素a与b的比例降低。dca1突变体中的突变被定位到ClpC1基因，从而表明叶绿体Clp蛋白酶通过响应叶绿素b的积累使CAO蛋白不稳定来参与调节叶绿素b的生物合成。
