STRUCTURE AND EXPRESSION OF HORMONALLY REGULATED GENES

植物激素生长素调节植物生长的各个方面， 发育，如细胞分裂、分化、形态发生， 肿瘤发生，并且通常被认为是负责调节 植物细胞生长 激素的主要作用机制是 了解甚少;然而，最近的实验证据表明， 这种激素有能力在转录水平上迅速发挥作用， 水平 植物生长素调控的生化和遗传研究 豌豆和拟南芥中的诱导基因是 在此提出，以深入了解生物化学的细节， 调节其表达及其在植物细胞中作用的机制 增长 该提案的具体目标是： 1. 分离和结构表征其余的生长素 豌豆中用于鉴定常见DNA存在的调控基因 启动子区的序列。 2. 为了确定PSIAA 4/5基因中的顺式作用元件， 生长素和放线菌酮诱导。 这些元素将被定义为 启动子区域的BAL-31缺失分析。 启动子构建体， 通过以下方式将合适的报道基因引入植物组织，a） 高速微粒，B）电穿孔，和c）Ti介导 转型 3. DNA亲和层析法纯化转录因子 与PSIAA 4/5启动子和分离的互补DNA相互作用 通过筛选λ gtll表达， 用DNA特异性探针构建文库。 4. 分离拟南芥中反式作用基因的突变， 生长素调控基因上。 5. 为了分离与假定的抑制子互补的DNA序列， 酵母中生长素调控基因的互补实验 啤酒。 6. 通过RNA原位检测确定对生长素有反应的细胞类型 杂交技术并对蛋白质进行亚细胞定位 由这些基因中的一些编码， 通过电子显微镜。

The plant hormone auxin regulates various aspects of plant growth and development such as cell division, differentiation, morphogenesis, oncogenesis, and is generally considered to be responsible for regulating plant cell growth. The primary mechanism of action of the hormone is poorly understood; however, recent experimental evidence indicates that the hormone has the capacity to act very rapidly at the transcriptional level. Biochemical and genetic studies on the regulation of the auxin inducible genes in Pisum sativum (pea) and Arabidopis thaliana are proposed herein to gain insight into the details of the biochemical machinery that regulates their expression and their role in plant cell growth. The specific aims of the proposal are: 1. To isolate and structurally characterize the rest of the auxin regulated genes in pea for identifying the presence of common DNA sequences in their promoter region. 2. To define the Cis-acting elements in the PSIAA4/5 gene responsible for auxin and cycloheximide inducibility. The elements will be defined by BAL-31 deletion analysis of the promoter region. Promoter constructs with appropriate reporter genes will be introduced into plant tissue by, a) high velocity microprojectiles, b) electroporation, and, c) Ti-mediate transformation. 3. To purify by DNA affinity chromatography transcriptional factors interacting with the PSIAA4/5 promoter and isolate complementary DNA sequences to some of these factors by screening lambda gtll expression libraries with DNA specific probes. 4. To isolate mutations in Arabidopis thaliana for genes acting in trans on the auxin regulated genes. 5. To isolate complementary DNA sequences to the putative repressor of the auxin regulated genes by complementation experiments in Saccharomyces cerevisiae. 6. To define the cell types that respond to auxin by RNA in situ hybridization techniques and to subcellularly localize the proteins encoded by some of these genes with antibodies made to synthetic peptides by means of the electron microscopy.