Tanaka-Ueguchi M, Itoh H, Oyama N, Koshioka M, Matsuoka M
Nagoya University, BioScience Center, Japan.
Plant J. 1998 Aug;15(3):391-400. doi: 10.1046/j.1365-313x.1998.00217.x.
Ectopic expression of the homeobox gene, NTH15 (Nicotiana tabacum homeobox 15) in transgenic tobacco leads to abnormal leaf and flower morphology, accompanied by a decrease in the content of the active gibberellin, GA1. Quantitative analysis of intermediates in the GA biosynthetic pathway revealed that the step from GA19 to GA20 was blocked in transgenic tobacco plants overexpressing NTH15. To investigate the relationship between the expression of NTH15 and genes involved in GA biosynthesis, we isolated three cDNA clones from tobacco encoding two types of GA 20-oxidase and a 3 beta-hydroxylase. RNA gel blot analysis revealed that the expression of one gene (Ntc12, encoding GA 20-oxidase), which in wild-type tobacco plants was abundantly expressed in leaves, was strongly suppressed in the transformants. The expression level of Ntc12 decreased with increasing severity of phenotype of transgenic tobacco leaves. The abnormal leaf morphology was largely overcome by treatment with GA20 or GA1 but not by GA19. These data strongly suggest that overexpression of NTH15 inhibits the expression of Ntc12, resulting in reduced levels of active GA and abnormal leaf morphology in transgenic tobacco plants. In situ hybridization in wild-type tobacco revealed that expression of Ntc12 occurred mainly in the rib meristem, cells surrounding the procambium and in leaf primordia. Expression was not seen in the tunica, corpus and procambium, tissues in which NTH15 was predominantly expressed. The contrasting expression patterns of these genes may reflect their antagonistic functions in the formation of lateral organs from the shoot apical meristem.
同源异型框基因NTH15（烟草同源异型框15）在转基因烟草中的异位表达导致叶片和花的形态异常，同时活性赤霉素GA1的含量降低。对赤霉素生物合成途径中间产物的定量分析表明，在过表达NTH15的转基因烟草植株中，从GA19到GA20的步骤被阻断。为了研究NTH15的表达与参与赤霉素生物合成的基因之间的关系，我们从烟草中分离出三个cDNA克隆，它们编码两种类型的GA 20-氧化酶和一种3β-羟化酶。RNA凝胶印迹分析表明，一个基因（Ntc12，编码GA 20-氧化酶）在野生型烟草植株的叶片中大量表达，而在转化体中受到强烈抑制。Ntc12的表达水平随着转基因烟草叶片表型严重程度的增加而降低。用GA20或GA1处理可在很大程度上克服异常的叶片形态，而用GA19处理则不能。这些数据有力地表明，NTH15的过表达抑制了Ntc12的表达，导致转基因烟草植株中活性GA水平降低和叶片形态异常。在野生型烟草中的原位杂交显示，Ntc12的表达主要发生在肋分生组织、原形成层周围的细胞和叶原基中。在NTH15主要表达的组织，即表皮、皮层和原形成层中未观察到表达。这些基因相反的表达模式可能反映了它们在茎尖分生组织形成侧生器官过程中的拮抗作用。
