PLANT GENE REGULATION DURING N-ASSIMILATION/FIXATION

编码谷氨酰胺不同多肽的基因 合成酶（GS）在高等植物中提供了一个有吸引力的系统， 研究调节核基因的分子机制 参与植物特有的两个功能过程，即 叶绿体发育和固氮。 我们建议 检查编码不同形式GS的基因 （叶绿体和胞质），探索分子基础， 它们在叶、根和氮中的差异表达， 固定豆科植物的根瘤。 本研究的具体目的 将解决以下问题：1）mRNA编码 叶绿体或胞质GS多肽是不同的， 唯一受调控的GS基因，或者它们是 差异转录起始产物还是RNA加工产物？ 2)哪些因素导致了特定的 叶片中叶绿体GS mRNA的表达（即光，组织- 类型）？ 3)参与的分子机制是什么？ 氮素对豆科植物根系GS mRNA的诱导作用 修复根瘤菌 编码不同GS多肽的cDNA克隆将被克隆。 每个GS的特征和特异性寡核苷酸探针 mRNA种类将用于检查其在体内的表达， 叶（即亮/暗），根，以及在根瘤发育过程中， 豆类。 将分离同源核GS基因， 表征，并用作分子探针，以确定 分子事件，围绕其调节表达， vivo. 特别是，我们将通过以下方式研究转录调控： 光，组织型，或根瘤菌感染，在孤立的核， 体外 推定的反式作用转录起始因子 将在体外检测对GS基因转录具有特异性。 野生 型和启动子缺失突变体的GS将被引入到 外源植物（通过Ti载体）及其在植物中的表达 监测转基因植物对光照、组织类型或 确定所需顺式作用DNA元件的根瘤菌感染 基因表达的调控。 除了定义 分子机制，影响差异调节的一个 相关基因家族，从长远来看，这一信息可能会B 应用于现有基因的修饰（即在非豆科植物中） 与根瘤菌成功结合。

The genes encoding the distinct polypeptides of glutamine synthetase (GS) in higher plants present an attractive system to study the molecular mechanisms which regulate nuclear genes involved in two functional processes unique to plants, namely chloroplast development and nitrogen fixation. We propose to examine the genes encoding the distinct forms of GS (chloroplastic and cytosolic), to explore the molecular basis for their differential expression in leaves, roots, and in nitrogen- fixing root nodules of legumes. The specific aims of this study will address the following questions: 1) Are the mRNAs encoding chloroplast or cytosolic GS polypeptides the products of distinct, uniquely regulated GS genes, or alternatively are they the products of differential transcript initiation or RNA processing? 2) What are the factors which contribute to the specific expression of chloroplast GS mRNA in leaves (i.e. light, tissue- type)? 3) What are the molecular mechanisms involved in the induction of GS mRNA in legume roots infected with nitrogen- fixing Rhizobia? cDNA clones encoding the distinct GS polypeptides will be characterized and oligonucleotide probes specific for each GS mRNA species will be used to examine its expression in vivo in leaves (i.e. light/dark), roots, and during nodule development in legumes. The cognate nuclear GS genes will be isolated, characterized, and used as molecular probes, to determine the molecular events which surround their regulated expression in vivo. In particular, we will examine transcriptional regulation by: light, tissue-type, or Rhizobial infection, in isolated nuclei in vitro. Putative trans-acting transcription initiation factors specific for GS gene transcription will be examined in vitro. Wild type and promoter deletion mutants of GS will be introduced into foreign plants (via Ti vectors) and their expression in the transgenic plants monitored in response to light, tissue-type, or Rhizobial infection to define cis-acting DNA elements required for regulated gene expression. In addition to defining the molecular mechanisms which effect a differential regulation of a family of related genes, in the long term, this information may b applied to the modification of existing genes (i.e. in non-legumes) to effect a successful association with Rhizobium.