Tsuji Yoshinori, Suzuki Iwane, Shiraiwa Yoshihiro
University of Tsukuba, Japan.
Plant Cell Physiol. 2009 Feb;50(2):318-29. doi: 10.1093/pcp/pcn200. Epub 2008 Dec 24.
The coccolithophorid Emiliania huxleyi (Haptophyta) is a representative and unique marine phytoplankton species that fixes inorganic carbon by photosynthesis and calci-fication. We examined the initial process of photosynthetic carbon assimilation by analyses of metabolites, enzymes and genes. When the cells were incubated with a radioactive substrate (2.3 mM NaH(14)CO(3)) for 10 s under illumination, 70% of the (14)C was incorporated into the 80% methanol-soluble fraction. Eighty-five and 15% of (14)C in the soluble fraction was incorporated into phosphate esters (P-esters), including the C(3) cycle intermediates and a C(4) compound, aspartate, respectively. A pulse-chase experiment showed that (14)C in P-esters was mainly transferred into lipids, while [(14)C]aspartate, [(14)C]alanine and [(14)C]glutamate levels remained almost constant. These results indicate that the C(3) cycle functions as the initial pathway of carbon assimilation and that beta-carboxylation contributes to the production of amino acids in subsequent metabolism. Transcriptional analysis of beta-carboxylases such as pyruvate carboxylase (PYC), phosphoenolpyruvate carboxylase (PEPC) and phosphoenolpyruvate carboxykinase (PEPCK) revealed that PYC and PEPC transcripts were greatly increased under illumination, whereas the PEPCK transcript decreased remarkably. PEPC activity was higher in light-grown cells than in dark-adapted cells. PYC activity was detected in isolated chloroplasts of light-grown cells. According to analysis of their deduced N-terminal sequence, PYC and PEPC are predicted to be located in the chloroplasts and mitochondria, respectively. These results suggest that E. huxleyi possesses unique carbon assimila-tion mechanisms in which beta-carboxylation by both PYC and PEPC plays important roles in different organelles.
颗石藻赫氏埃米利亚藻（定鞭藻门）是一种具有代表性且独特的海洋浮游植物物种，通过光合作用和钙化作用固定无机碳。我们通过对代谢物、酶和基因的分析，研究了光合碳同化的初始过程。当细胞在光照下与放射性底物（2.3 mM NaH(14)CO(3)）孵育10秒时，70%的(14)C被整合到80%的甲醇可溶部分中。可溶部分中85%和15%的(14)C分别被整合到磷酸酯（P-酯）中，包括C(3)循环中间体和一种C(4)化合物天冬氨酸。脉冲追踪实验表明，P-酯中的(14)C主要转移到脂质中，而[(14)C]天冬氨酸、[(14)C]丙氨酸和[(14)C]谷氨酸水平几乎保持不变。这些结果表明，C(3)循环作为碳同化的初始途径发挥作用，并且β-羧化作用在随后的代谢中有助于氨基酸的产生。对丙酮酸羧化酶（PYC）、磷酸烯醇式丙酮酸羧化酶（PEPC）和磷酸烯醇式丙酮酸羧激酶（PEPCK）等β-羧化酶的转录分析表明，PYC和PEPC转录本在光照下大幅增加，而PEPCK转录本显著下降。光照培养的细胞中PEPC活性高于暗适应细胞。在光照培养细胞的分离叶绿体中检测到了PYC活性。根据对其推导的N端序列的分析，预计PYC和PEPC分别位于叶绿体和线粒体中。这些结果表明，赫氏埃米利亚藻拥有独特的碳同化机制，其中PYC和PEPC的β-羧化作用在不同细胞器中发挥重要作用。
