Characterization of Brassinosteroid Dwarfs of Arabidopsis: Cloning of DWF4

K.A.费尔德曼9604439从T-DNA和EMS诱变的群体中分离出一系列矮化(细胞伸长)突变体。它们被映射到七个互补组(Dwf1-Dwf7)；在其中三个组(Dwf1、DwF3和Dwf4)中，至少有一个等位基因与T-DNA共聚集。最近，外源油菜素内酯(BL)对Dwf1、Det2(Dwf6)和Cpd(Dwf3)这三个矮秆品种起到了补充作用。除Dwf2(Bri)外，其余矮秆品种的矮秆表型也得到了BL型的补充。克隆的矮秆基因(DEI2、CPD和较小程度的DWFI)序列表明它们参与了BL的生物合成。BRI(油菜素内酯不敏感)被认为是BL的受体。这就留下了三个BL互补的基因座，即DWF4、DWFS和DWF7，它们在任何水平上都基本上没有特征。这里描述的矮秆的收集很可能代表了这种类型的矮秆表型的拟南芥基因组接近饱和。这一建议的目标包括：1)油菜素类固醇相关矮秆的拟南芥基因组饱和；确定相互作用的基因座；2)进一步表征DWFS、DWF6和DWP(形态、解剖和生理)；3)通过检测所有6个对油菜素类固醇相关的矮秆的双突变体和多突变体来检验关于BL生物合成的假说；4)利用来自T-DNA标记突变体的3‘基因组克隆来鉴定和表征全长的DWF4 cDNA和基因组克隆；这包括对几个等位基因进行测序和分子互补；5)确定表达和定位DWF4的组织；6)分析BL生物合成基因和BL调控基因在DWI和其他细胞扩增突变体中的表达；7)研究BL生物合成基因的异位表达对细胞大小和形态的影响。这一分析将使发展更精确的生物合成模型成为可能。此外，DWF4基因的克隆还将为研究BL途径提供另一种分子工具。这些目标的完成将为油菜素类固醇在细胞伸长和植物生长发育的其他过程中起什么作用提供大量信息。

K.A. Feldmann 9604439 A collection of dwarf (cell elongation) mutants have been isolated from T-DNA and EMS mutagenized populations. They have been mapped to seven complementation groups (dwfl -dwf7); in three of these (dwfl, dwf3, and dwf4) there is at least one allele that cosegregates with a T-DNA. Recently, three dwarfs, dwfl, det2 (dwf6) and cpd (dwf3) were shown to be complemented by the exogenous application of brassinolide (BL). The dwarf phenotype in all the other dwarfs except dwf2 (bri) was also complemented by BL. Sequences of cloned dwarf genes (DEI2, CPD and to a lesser extent DWFI) indicate that they are involved in BL biosynthesis. bri (brassinolide-insensitive) is hypothesized to be a receptor for BL. This leaves three BL-complemented loci, DWF4, DWFS and DWF7, that remain largely uncharacterized at any level. The collection of dwarfs described here very likely represents near saturation of the Arabidopsis genome for this type of dwarf phenotype. The objectives of this proposal include: 1) saturation of the Arabidopsis genome for brassinosteroid-related dwarfs; identify interacting loci; 2) further characterization of dwfS, dwf6 and dwp (morphology, anatomy, and physiology); 3) test hypotheses about BL biosynthesis by examining double and multiple mutants of all six BL-responsive dwarfs; 4) identify and characterize full length DWF4 cDNA and genomic clones utilizing a 3' genomic clone from a T-DNA tagged mutant; this includes sequencing several alleles and molecular complementation; 5) define the tissues in which DWF4 is expressed and localized; 6) analyze the expression of BL biosynthetic and BL-regulated genes in dwis and other cell expansion mutants, and 7) study the effect of ectopic expression of BL biosynthesis genes on cell size and morphology. This analysis will allow the development of a more precise biosynthetic model for BL. In addition, the cloning of DWF4 will add another molecular tool for studying the BL pathway. Completion of these objectives will pro vide substantial information on what role brassinosteroids have in cell elongation and other processes in plant growth and development.