Regulation of Shoot Development in Arabidopsis

植物器官再生多年来一直引起科学家的兴趣，因为在培养中再生组织的发育命运可以由两种简单的植物激素——细胞分裂素和生长素——控制。在细胞分裂素相对于生长素的高水平下，产生芽，而在生长素相对于细胞分裂素的高水平下，形成根。茎部再生不仅是一种有趣的发育现象，而且是植物繁殖和改良的重要商业过程。了解它的调控可能有助于解决茎部再生效率低下的问题，这往往阻碍了对具有商业意义的植物品种的改进。最近在该实验室进行的研究使用寡核苷酸阵列（DNA芯片）分析来描述在茎发育过程中基础的基因表达的全局模式。本研究分析了8000多个基因，其中数百个基因上调或下调，其中涉及激素信号和茎分生组织形成的基因。在拟南芥中，一些生态型（来自不同地理位置的分离物）在可比的组织培养条件下形成芽的能力不同。为了了解决定茎部发育效率的基因，研究人员对控制再生能力差异的基因（位点）进行了搜索，这些基因（位点）控制着一种生态型（哥伦比亚）和另一种生态型（Landsberg）之间再生能力的差异。在5号染色体上发现了哥伦比亚等位基因促进芽形成的主要位点。另外两个小位点在1号和4号染色体上被鉴定出来。这些位点上的兰氏等位基因提高了芽产量。该项目将研究这些基因座如何影响茎发育过程中的基因表达程序，以及5号染色体上的主要基因座是否控制一组特定基因的表达。利用含有2万多个拟南芥基因的寡核苷酸阵列（DNA芯片），鉴定5号染色体主要位点上的哥伦比亚等位基因和兰兹伯格等位基因在茎发育过程中的差异调控基因。此外，将评估拟南芥基因组中每个基因在两种生态型中的表达水平，以确定是否有基因在茎发育过程中表现出遗传的基因表达模式。如果是这样，就需要对控制基因表达可遗传变化的单个基因的位点进行搜索。这种方法被称为“基因基因组学”，将寡核苷酸阵列上任何基因的表达水平视为一种度量特征，并允许人们寻找控制这些特征的基因。这是构建拟南芥芽发育过程中基因表达因果通路的第一步。最后，我们提出了在哥伦比亚生态型中，在5号染色体上的主要位点上鉴定提高茎部发育效率的基因。这将通过一个叫做“染色体行走”的过程来完成，在这个过程中，精细的基因定位技术被用来确定候选基因。

Organ regeneration in plants has intrigued scientists for years because the developmental fate of regenerating tissue in culture can be controlled by two simple plant hormones, cytokinin and auxin. At high levels of cytokinin relative to auxin, shoots are produced, while at high levels of auxin relative to cytokinin, roots are formed. Shoot regeneration is not only an interesting developmental phenomenon but also an important commercial process for the propagation and improvement of plants. Understanding its regulation may help to solve inefficiencies in shoot regeneration that often impede efforts to improve commercially significant plant varieties.Recent studies were conducted in this laboratory using oligonucleotide array (DNA chip) analysis to describe the global pattern of gene expression that underlies the shoot development process. More than 8000 genes were analyzed in this study, and several hundred were up or downregulated among them were genes involved in hormone signaling and shoot meristem formation.In Arabidopsis, some ecotypes (isolates from different geographical locations) differ in their ability to form shoots under comparable tissue culture conditions. In the effort to understand the genes that determine the efficiency of shoot development, a search was conducted for genes (loci) that control the difference in regeneration ability between one ecotype (Columbia) that regenerates well and another ecotype (Landsberg) that does not. One major locus on chromosome 5 was identified, in which the Columbia allele enhances shoot formation. Two other minor loci were identified on chromosomes 1 and 4. Landsberg alleles at these loci enhance shoot production. The proposed project will examine how these loci impact the program of gene expression during shoot development and whether the major locus on chromosome 5 controls the expression of a specific set of genes. Oligonucleotide arrays (DNA chip) with over 20,000 Arabidopsis genes will be employed to identify the genes that are differentially regulated during shoot development by the Columbia or the Landsberg alleles at the major locus on chromosome 5.In addition, the expression level of every gene in the Arabidopsis genome will be assessed in the two ecotypes to see if any genes show heritable patterns of gene expression during shoot development. If so, searches will be undertaken for loci controlling individual genes that show heritable changes in gene expression. This approach, termed "genetical genomics," treats the expression level of any gene on an oligonucleotide array as a metric trait and permits one to seek out genes that control these traits. This represents the first step in constructing causal pathways of gene expression during shoot development in Arabidopsis.Finally, it is proposed to identify the gene(s) at the major locus on chromosome 5 that enhances the efficiency of shoot development in the Columbia ecotype. This will be done by a process called "chromosome walking," in which fine-scale gene mapping techniques are used to pinpoint candidate genes.