Regulation of Diterpene Biosynthesis in Higher Plants

这项研究的总体目标是加深对高等植物中调节二萜代谢物生物合成的机制的理解，这些二萜代谢物有助于植物在环境和生物胁迫下生存的能力。重点放在环化酶的调节上，该酶催化产生生理活性二萜代谢物(S)的途径的第一步，因此是一个合理的调节部位。一个靶酶是酪烯合成酶，它催化蓖麻籽幼苗中可能的植物保鲜蛋白酪宾的合成。果胶片段激发子处理幼苗后，酪蛋白合成酶基因在转录水平上被激活。含有酪蛋白合成酶编码序列的基因组克隆已被分离。这项研究的一个主要目的是鉴定酪氨酸合成酶基因以及参与果胶片段激发子对该基因转录激活的顺式和反式作用因子。这些研究对于植物抗病具有重要意义，因为植物保卫素是植物抗病反应中的防御成分之一。另一个实验目标是Kaurene合成酶，它是赤霉素调节蛋白家族途径中的关键调节酶，有助于调节环境对植物生长和发育的影响。一个主要的目的是将考拉烯合成酶纯化为均一，并用纯化的蛋白制备特异性抗体和考拉烯合成酶基因探针。这些探针将被用来测量植物组织中Kaurene合成酶mRNA的水平，以及它们在不同环境条件下的水平变化，作为它们合成赤霉素的相对潜力的指标。

The general objective of this research is to develop understanding of the mechanisms operating in higher plants to regulate the biosynthesis of diterpenoid metabolites that contribute to a plant's ability to survive environmental and biological stresses. The focus is on the regulation of cyclization enzymes that catalyze the first committed step of the pathway leading to a physiologically active diterpenoid metabolite(s), and thus is a logical site for regulation. One target enzyme is casbene synthetase, which catalyzes the synthesis of the putative phytoalexin, casbene, in castor bean seedlings. The casbene synthetase gene is transcriptionally activated in response to treatment of the seedlings with pectic fragment elicitors. Genomic clones containing casbene synthetase coding sequences have been isolated. A major goal of the research is to characterize the casbene synthetase gene and the cis- and trans-acting factors that participate in the transcriptional activation of the gene in response to pectic fragment elicitors. These investigations are of importance in connection with plant disease resistance since phytoalexins are among the defensive components that appear in a resistance response. Another experimental target is kaurene synthetase, a key regulatory enzyme in the pathway leading to the gibberellin family of regulators that help mediate environmental influences on plant growth and development. A major objective is to purify kaurene synthetase to homogeneity, and to use the purified protein to prepare specific antibodies and kaurene synthetase cDNA probes. These probes will be used to measure kaurene synthetase mRNA levels in plant tissues, and changes in their levels under different environmental conditions, as an indicator of their relative potentials for gibberellin biosynthesis.