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。它们已经被定位到七个互补组（dwf 1-dwf 7）;在其中三个（dwf 1、dwf 3和dwf 4）中，存在至少一个与T-DNA共分离的等位基因。最近，三个矮秆，dwf 1，det 2（dwf 6）和cpd（dwf 3）被证明是通过外源施用芸苔素（BL）补充。除dwf 2（bri）外，其它所有矮秆材料的矮秆表型均得到BL的补充。克隆的矮秆基因（DEI 2，CPD和在较小程度上DWFI）的序列表明，它们参与BL生物合成。假设bri（油菜素内酯不敏感）是BL的受体。这留下了三个BL-互补的基因座，DWF 4，DWFS和DWF 7，在任何水平上都没有得到很大的表征。这里所描述的矮秆植物的集合很可能代表了拟南芥基因组对这种类型的矮秆表型的接近饱和。 本研究的主要目的是：1）对拟南芥中油菜素甾醇相关矮秆基因组进行饱和分析，确定其互作位点; 2）进一步研究dwfS、dwf 6和dwp基因（形态学、解剖学和生理学）; 3）通过检查所有六个BL-响应性矮化的双重和多重突变体来测试关于BL生物合成的假设; 4）利用来自T-DNA标记的突变体的3'基因组克隆鉴定和表征全长DWF 4 cDNA和基因组克隆;这包括测序几个等位基因和分子互补; 6）分析在dwis和其它细胞扩增突变体中BL生物合成和BL调节基因的表达，和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。 该分析将允许开发更精确的BL生物合成模型。此外，DWF 4的克隆将为研究BL途径增加另一种分子工具。这些目标的完成将提供实质性的信息，油菜素类固醇在细胞伸长和植物生长发育的其他过程中的作用。

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.