Identification of a vinyl reductase gene for chlorophyll synthesis in Arabidopsis thaliana and implications for the evolution of Prochlorococcus species
Identification of a vinyl reductase gene for chlorophyll synthesis in Arabidopsis thaliana and implications for the evolution of Prochlorococcus species
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DOI：
10.1105/tpc.104.027276
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发表时间：
2005-01-01
期刊：
影响因子：
11.6
通讯作者：
Tanaka, A
中科院分区：
文献类型：
--
作者：
Nagata, N;Tanaka, R;Tanaka, A
各种生物对叶绿素代谢进行了广泛的研究，在高等植物中几乎所有的叶绿素生物合成基因都已被鉴定出来。然而，目前只有3，8-二乙烯基原叶绿素内酯-8-乙烯基还原酶(DVR)的基因还没有被鉴定，DVR是单乙烯基叶绿素合成所必需的。在本研究中，我们通过甲烷磺酸乙酯诱变，分离到了一个能积累二乙烯基叶绿素而不是单乙烯基叶绿素的拟南芥突变体。对该突变体进行图位克隆，获得了一个与异黄酮还原酶基因序列相似的基因(AT5G18660)。野生型基因的转化补充了突变的表型。在大肠杆菌中表达了由AT5G18660编码的重组蛋白，并发现该蛋白能催化二乙烯基叶绿素转化为单乙烯基叶绿素，从而证明该基因编码一个功能性的DVR。DVR是由拟南芥基因组中的单拷贝基因编码的。随着DVR的鉴定，最终在高等植物中鉴定出了叶绿素生物合成所需的所有基因。对拟南芥全基因组的分析表明，它有15种酶，由27个基因编码，参与从谷氨酰-tRNA(Glu)到叶绿素b的生物合成。此外，DVR基因的鉴定有助于了解海洋原氯球菌的进化。海洋原氯球菌是一种海洋蓝藻，在公海中占主导地位，在利用二乙烯基叶绿素方面很少见。在海洋假单胞菌基因组中未发现DVR同源基因，而在聚球藻WH8102基因组中发现了DVR同源物，这与原氯球藻属和聚球藻属海洋蓝藻中二乙烯基叶绿素的分布一致。
Chlorophyll metabolism has been extensively studied with various organisms, and almost all of the chlorophyll biosynthetic genes have been identified in higher plants. However, only the gene for 3,8-divinyl protochlorophyllide a 8-vinyl reductase (DVR), which is indispensable for monovinyl chlorophyll synthesis, has not been identified yet. In this study, we isolated an Arabidopsis thaliana mutant that accumulated divinyl chlorophyll instead of monovinyl chlorophyll by ethyl methanesulfonate mutagenesis. Map-based cloning of this mutant resulted in the identification of a gene (AT5G18660) that shows sequence similarity with isoflavone reductase genes. The mutant phenotype was complemented by the transformation with the wild-type gene. A recombinant protein encoded by AT5G18660 was expressed in Escherichia coli and found to catalyze the conversion of divinyl chlorophyllide to monovinyl chlorophyllide, thereby demonstrating that the gene encodes a functional DVR. DVR is encoded by a single copy gene in the A. thaliana genome. With the identification of DVR, finally all genes required for chlorophyll biosynthesis have been identified in higher plants. Analysis of the complete genome of A. thaliana showed that it has 15 enzymes encoded by 27 genes for chlorophyll biosynthesis from glutamyl-tRNA(glu) to chlorophyll b. Furthermore, identification of the DVR gene helped understanding the evolution of Prochlorococcus marinus, a marine cyanobacterium that is dominant in the open ocean and is uncommon in using divinyl chlorophylls. A DVR homolog was not found in the genome of P. marinus but found in the Synechococcus sp WH8102 genome, which is consistent with the distribution of divinyl chlorophyll in marine cyanobacteria of the genera Prochlorococcus and Synechococcus.