Functional Analyses of an Epicuticular Wax Gene from Arabidopsis

所有植物的角质层表面都含有表皮蜡（EW）。扫描电镜显示，在许多植物物种中，电子束由一个无定形的由棒状结构和不规则的具有圆形边缘的板状结构覆盖的基底层组成。蜡是由长链脂肪族化合物的混合物组成的，这种化合物可以通过将植物的空中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、酯类、-二酮、醛、伯醇和仲醇、酮、游离酸和三萜。在植物物种之间，无论是在蜡中存在的化学类别的数量还是它们的相对丰度，都存在着巨大的多样性。即使在单个植物内，各种器官也可以通过EW组成的数量和质量差异来表征。人们对蜡的生化途径了解甚少，对这些分子通过细胞壁和角质层向表面出口的了解就更少了。然而，大量的EW突变体已经在拟南芥中分离出来，这使得研究人员能够采用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其中的两个基因，CER4和CER21。CER4位点的突变消除了伯醇的积累，而CER21位点的突变消除了几乎所有的醛类。这两个基因都将被测序。它们在叶、茎和角质部的表达水平将在发育过程中得到表征。此外，CER4和CER21基因的表达将在几种环境下生长的植物中进行研究。这两个基因的细胞型特异性表达将被研究。亚细胞定位将通过针对CER4和CER21蛋白的抗体进行原位分析。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。有大量的证据表明，氨基酸的组成在食草昆虫选择寄主植物中起着重要的作用。通过基因工程方法对EW进行修饰，将使我们能够在一些重要的农用作物中提供对各种害虫的抗性。拟南芥修饰的结果可用于农艺植物蜡质修饰。

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.