Analysis of Promoter Activity for the Gene Encoding Pyruvate Orthophosphate Dikinase in Stably Transformed C(4) Flaveria Species

The C(4) enzyme pyruvate orthophosphate dikinase is encoded by a single gene, Pdk, in the C(4) plant Flaveria trinervia. This gene also encodes enzyme isoforms located in the chloroplast and in the cytosol that do not have a function in C(4) photosynthesis. Our goal is to identify cis-acting DNA sequences that regulate the expression of the gene that is active in the C(4) cycle. We fused 1.5 kb of a 5′ flanking region from the Pdk gene, including the entire 5′ untranslated region, to the uidA reporter gene and stably transformed the closely related C(4) species Flaveria bidentis. β-Glucuronidase (GUS) activity was detected at high levels in leaf mesophyll cells. GUS activity was detected at lower levels in bundle-sheath cells and stems and at very low levels in roots. This lower-level GUS expression was similar to the distribution of mRNA encoding the nonphotosynthetic form of the enzyme. We conclude that cis-acting DNA sequences controlling the expression of the C(4) form in mesophyll cells and the chloroplast form in other cells and organs are co-located within the same 5′ region of the Pdk gene.

C4酶丙酮酸磷酸二激酶（pyruvate orthophosphate dikinase）由C4植物三脉黄花菊（Flaveria trinervia）中的单个Pdk基因编码。该基因同时编码定位于叶绿体与细胞质的酶同工型，这类同工型并不参与C4光合作用过程。本研究旨在鉴定可调控该基因在C4循环中活跃表达的顺式作用DNA序列。我们将Pdk基因长1.5kb的5'侧翼区（包含完整的5'非翻译区）与uidA报告基因进行融合，并对近缘C4物种双花黄花菊（Flaveria bidentis）开展稳定转化。在叶片叶肉细胞中可检测到高水平的β-葡萄糖苷酸酶（β-Glucuronidase, GUS）活性；在维管束鞘细胞与茎组织中可检测到较低水平的GUS活性，而在根组织中仅能检测到极低水平的GUS活性。这种低水平的GUS表达模式，与编码该酶非光合形式的mRNA的分布特征相符。综上，我们认为调控叶肉细胞中C4形式酶表达、以及其他细胞与器官中叶绿体形式酶表达的顺式作用DNA序列，共同定位于Pdk基因的同一5'区域内。