Studies on signal transduction pathways of jasmonic acid in suspension-cultured rice cells

本研究试图分离茉莉酸（JA）反应基因，并对其进行鉴定，以阐明JA在水稻悬浮细胞中的信号转导途径。此外，还合成了JA受体的分子探针，并发现水稻JA反应基因RRJ 1和RRJ 2分别编码一种推测的水稻胱硫醚γ-裂解酶和一种丙酮酸脱羧酶。RRJ 1可能参与含硫氨基酸的生物合成和/或代谢及相关的生化过程，RRJ 2可能参与激发子诱导的水稻细胞防御反应的能量代谢。这是首次从单子叶植物中克隆OPR基因。由于OsOPR 1比JA的天然前体（+）-cis-OPDA更优先催化（-）-cis-OPDA的还原，OsOPR 1可能有助于JA的其他生理活性。 ...更多信息 RERJ 1是一个早期JA应答基因，编码一个具有碱性区/螺旋-环-螺旋基序的转录调节因子。通过对正义和反义RERJ 1基因过表达转基因水稻植株的表型分析，推测RERJ 1基因可能参与了茉莉酸诱导的水稻芽生长抑制。RERJ 1也很可能参与其他生理现象，包括防御反应。JA在几丁质诱导的水稻悬浮培养细胞中作为一个关键的信号转导子参与了二萜类植物抗毒素的合成。我们从几丁质诱导的悬浮培养的水稻细胞中分离了编码二萜环化酶的cDNA，并证明该基因产物具有内消旋-卡萨-12，15-二烯合酶的功能。这是第一次报道分离的cDNA编码的二萜环化酶参与生物合成的二萜烃，这是一个推定的前体二萜类植物抗毒素在水稻。本研究以水稻二萜类植物抗毒素（-）-phytocassanes（-）-Phytocassanes）为研究对象，合成了几种JA衍生物，包括JA-氨基酸偶联物、JA-生物素偶联物、JA-地塞米松异二聚体、和JA-荧光素缀合物作为JA-结合蛋白的分子探针的候选物。除JA-荧光素偶联物外，其他化合物在水稻幼苗试验、大豆PAL诱导试验和/或水稻乳内酯A诱导试验中均表现出明显的JA样活性，因此它们可能是分离和鉴定JA结合蛋白的有用探针。少

In this project, we have tried to isolate and characterize jasmonic acid (JA)-responsive genes to elucidate the signal transduction pathways of JA in suspension-cultured rice cells. In addition, molecular probes for JA receptors were also synthesized.The JA-responsive genes RRJ1 and RRJ2 were found to encode a putative rice cystathionine γ-lyase and a pyruvate decarboxylase, respectively. RRJ1 is possibly involved in biosynthesis and/or metabolism of sulfur-containing amino acids and related biochemical processes, and RRJ2, in energy metabolism for elicitor-induced defense reactions in suspension-cultured rice cells.We succeeded in cloning of the JA-responsive gene OsOPR1 encoding 12-oxophytodienoic acid (OPDA) reductase (OPR) that is one of key biosynthetic enzymes of JA. This is the first cloning of an OPR gene from a monocot plant. Since OsOPR1 catalyze reduction of (-)-cis-OPDA preferentially over (+)-cis-OPDA, a natural precursor of JA, OsOPR1 might contribute to other physiologic … More al phenomena than JA biosynthesis.RERJ1 was isolated as an early JA-responsive gene which encodes a transcriptional regulator possessing a basic region/helix-loop-helix motif. Based on analysis of phenotypes of sense- or antisense-RERJ1 mRNA overexpressing transgenic rice plants, RERJ1 is suggested to be involved in JA-induced inhibition of shoot growth in rice. Involvement of RERJ1 in other physiological phenomena including defense response is also quite possible. Futher analysis of biological functions of RERJ1 is in progress.JA functions as a key signal transducer in a chitin elicitor-induced diterpenoid phytoalexin production in suspension-cultured rice cells. We isolated a cDNA encoding a diterpene cyclase from the chitin-elicited suspension-cultured rice cells, and demonstrated that the gene product functioned as ent-cassa-12, 15-diene synthase. This is the first report of isolation of a cDNA encoding a diterpene cyclase involved in biosynthesis of a diterpene hydrocarbon which is a putative precursor of diterpenoid phytoalexins in rice. ent-Cassa-12, 15-diene synthase cDNA isolated here will be a useful tool to investigate the regulatory mechanisms of biosynthesis of the rice diterpenoid phytoalexins (-)-phytocassanes during plant development and by change of environmental conditions.We synthesized several types of JA derivatives including JA-amino acid conjugates, JA-biotin conjugate, JA-dexamethasone heterodimer, and JA-fluoresceine conjugate as candidates for molecular probes for JA-binding proteins. Since these synthesized compounds except JA-fluoresceine conjugate exhibited significant JA-like activities in rice seedling assay, soybean PAL-inducing assay, and/or rice momilactone A-inducing assay, they might be useful probes to isolate and characterize JA-binding proteins. Less