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，并 发育，如细胞分裂、分化、形态发生、 肿瘤发生，通常被认为是负责调节 植物细胞生长。荷尔蒙的主要作用机制是 知之甚少；然而，最近的实验证据表明 荷尔蒙有能力在转录时非常迅速地起作用 水平。生长素调控的生化和遗传学研究 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 调节它们在植物细胞中的表达和作用的机制 成长。这项建议的具体目标是： 1.分离和结构鉴定其余的生长素 豌豆中鉴定普通DNA存在的调控基因 它们的启动子区域的序列。 2.确定PSIAA4/5基因的顺式作用元件 生长素和放线菌亚胺的诱导性。元素将由以下内容定义 BAL-31启动子区域缺失分析。启动子与 适当的报告基因将通过，a)导入植物组织。 高速微弹，b)电穿孔，c)钛中间体 转型。 3.用DNA亲和层析法纯化转录因子 与PSIAA4/5启动子相互作用并分离互补DNA 通过筛选lambda gtll表达对其中一些因子进行序列测定 带有DNA特异性探针的文库。 4.分离拟南芥中反式作用基因的突变 生长素调节基因。 5.分离与可能的阻遏因子互补的DNA序列 利用互补实验研究酵母生长素调控基因 酿酒。 6.用原位RNA确定对生长素有反应的细胞类型 杂交技术和亚细胞定位蛋白质 由这些基因中的一些用合成多肽的抗体编码 通过电子显微镜观察。

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.