Lipka B, Steinmüller K, Rosche E, Börsch D, Westhoff P
Institut für Entwicklungs- und Molekularbiologie der Pflanzen, Heinrich-Heine-Universität, Düsseldorf, Germany.
Plant Mol Biol. 1994 Dec;26(6):1775-83. doi: 10.1007/BF00019491.
To study the molecular evolution of NADP-dependent malic enzyme (NADP-ME) in the genus Flaveria a leaf-specific cDNA library of the C3 plant F. pringlei was screened for the presence of sequences homologous to the C4 isoform gene (named modA) of the C4 plant F. trinervia. The cDNAs isolated contained varying numbers of identical restriction fragments suggesting that they were derived from a single gene. This was supported by Southern hybridisation experiments with genomic DNA from F. trinervia and F. pringlei. Nucleotide sequence analysis of a full-size clone identified the presence of a typical plastidic transit peptide and revealed that the mature modA proteins of F. trinervia (C4) and F. pringlei (C3) are 90% similar. These findings indicate that C3 plants, like C4 species, possess a plastidic isoform of NADP-ME and that the modA genes of the two species represent orthologous genes. Northern analyses showed that modA transcripts accumulate to similar levels in leaves, stems and roots of F. pringlei. The expression of this gene in F. pringlei thus appears to be rather constitutive. In contrast, the modA gene of F. trinervia is abundantly expressed in leaves, but maintains its expression in stems and roots. It has to be concluded from these data that the leaf-specific increase in the expression level was a key step which was taken during the evolution of the C4 isoform modA gene starting from a C3 ancestral gene.
为了研究黄顶菊属中依赖于NADP的苹果酸酶（NADP-ME）的分子进化，对C3植物普氏黄顶菊的叶特异性cDNA文库进行筛选，寻找与C4植物三脉黄顶菊的C4同工型基因（命名为modA）同源的序列。分离得到的cDNA含有不同数量的相同限制性片段，这表明它们来源于单个基因。三脉黄顶菊和普氏黄顶菊基因组DNA的Southern杂交实验支持了这一点。对一个全长克隆进行核苷酸序列分析，确定存在典型的质体转运肽，并显示三脉黄顶菊（C4）和普氏黄顶菊（C3）的成熟modA蛋白相似度为90%。这些发现表明，C3植物与C4植物一样，拥有一种质体同工型的NADP-ME，并且这两个物种的modA基因代表直系同源基因。Northern分析表明，modA转录本在普氏黄顶菊的叶、茎和根中积累到相似水平。因此，该基因在普氏黄顶菊中的表达似乎相当组成型。相比之下，三脉黄顶菊的modA基因在叶中大量表达，但在茎和根中也保持表达。从这些数据可以得出结论，表达水平的叶特异性增加是从C3祖先基因开始的C4同工型modA基因进化过程中的关键一步。
