Functional Analyses of an Epicuticular Wax Gene from Arabidopsis

9419093 Feldmann所有植物的角质层表面都含有表皮蜡（EW）。 扫描电子显微镜显示，在许多植物物种的EW由一个无定形的地面层覆盖的杆状结构和不规则形状的板状结构与圆形边缘。 蜡由长链脂肪族化合物的混合物组成，其可以通过将植物的地上部分浸入氯仿或己烷中几秒钟，然后蒸发溶剂来分离。 脂肪族化合物包括烷烃、酯、β-二酮、醛、伯醇和仲醇、酮、游离酸和三萜。 在植物物种中存在巨大的多样性，无论是在蜡中存在的化学类别的数量，还是在它们的相对丰度上。 甚至在单个植物内，不同的器官可以通过EW组成的定量和定性差异来表征。 蜡的生化途径知之甚少，甚至更少的是知道这些分子通过细胞壁和角质层的出口到表面。 然而，大量的EW突变体已经在拟南芥中分离，这使得研究人员可以采取遗传和分子方法来解剖表皮蜡质途径。 本提案的目的是克隆和表征这些基因中的两个，CER 4和CER 21。 CER 4基因座的突变消除了伯醇的积累，而CER 21基因座的突变消除了几乎所有的醛。 这两个基因都将被测序。 其在叶、茎和长角果中的表达水平将在发育期间表征。 此外，将在几种环境条件下生长的植物中研究CER 4和CER 21基因的表达。 将研究这两个基因的细胞类型特异性表达。 将通过使用针对CER 4和CER 21蛋白产生的抗体进行原位分析进行亚细胞定位。 还将研究这些基因在植物中的功能，即，它们在生物合成、运输或调节中是否重要。 大量证据表明，EW的组成在植食性昆虫对寄主植物的选择中是重要的。 通过基因工程方法对EW进行修饰，应使我们能够在一些重要的农作物中提供对各种害虫的抗性。 从拟南芥的修饰获得的结果可用于农业植物上的蜡修饰。

9419093 Feldmann All plants contain epicuticular wax (EW) on the surface of the cuticle. Scanning electron microscopy has revealed that in many plant species the EW consists of an amorphous ground layer covered by rod-shaped structures and irregular shaped plate-like structures with rounded edges. The wax is composed of a mixture of long chain aliphatic compounds which can be isolated by dipping an aerial portion of the plant into chloroform or hexane for a few seconds and then evaporating the solvent. The aliphatic compounds include alkanes, esters, beta-diketones, aldehydes, primary and secondary alcohols, ketones, free acids, and triterpenes. There is tremendous diversity among plant species both in the number of chemical classes present in a wax and in their relative abundance. Even within an individual plant the vairous organs can be characterized by quantitative and qualitative differences in the EW composition. The wax biochemical pathway is poorly understood and even less is known about the export of these molecules through the cell wall and cuticle to the surface. However, a large number of EW mutants have been isolated in Arabidopsis, which allows the investigator to take a genetic and molecular approach to dissecting the epicuticular wax pathway. The objective of this proposal is to clone and characterize two of these genes, CER4 and CER21. The mutation at the CER4 locus eliminates the accumulation of primary alcohols whicle a mutation at the CER21 locus eliminates nearly all of the aldehydes. Both genes will be sequenced. Their expression level in leaves, stems and siliques will be characterized during development. Also, the expression of the CER4 and CER21 genes will be studied in plants grown under several encironmental regimes. The cell-type specific expression of these two genes will be studied. Subcellular localization will be performed via in situ analyses with antibodies raised against the CER4 and CER21 proteins. Also s tudied will be the function of these genes in the plant, i.e., are they important in biosynthesis, transport, or regulation. There is substantial evidence to show that the composition of the EW is important in host plant selection by herbivorous insects. Modification of the EW via genetic engineering methods should enable us to provide resistance to various insect pests in a number of important agronomic crops. The results obtained from the modification of Arabidopsis can be used to modify the waxes on agronomic plants.