INTERACTIONS BETWEEN DNA AND THE LYSR-TYPE PROTEIN OCCR

DNA 与 LYSR 型蛋白 OCCR 之间的相互作用

• 批准号: 3046590
• 负责人: WILLIAM C FUQUA
• 金额: $ 2.27万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-09-14 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3046590
• 关键词: DNA binding protein; DNA footprinting; gel mobility shift assay; gene induction /repression; gene interaction; genetic promoter element; genetic transcription; regulatory gene

The process of Agrobacterium tumefaciens of plants is a well studied phenomenon, and is the only bonafide example of natural genetic engineering. The infecting bacterium transfers a fragment of its DNA from a tumor inducing (Ti) plasmid, into the host plant genome. Genes encoded on this fragment direct tumor formation and the synthesis of unusual sugars and amino acids termed opines, which serve as Agrobacterium-specific carbon/nitrogen sources. In turn, the opines stimulate the expression of A. tumefaciens genes involved in their uptake and utilization, conjugal transfer of the Ti plasmid, and a number of other functions that may be involved in tumor colonization. In octopine-type plasmids this induction is mediated at least in part by the LysR-type transcriptional activator OccR, which induces the transcription of these genes in response to the opine octopine and which also autoregulates its own synthesis. One of the targets for OccR regulation is the overlapping, divergently transcribed promoter region between the occ (octopine catabolism) operon and the occR gene itself. This unusual genetic organization is typical of LysR-type systems and probably allows simultaneous control of both operons. In the proposed study, we shall examine the interaction of OccR with this transcriptional control region. Evidence is presented that OccR binds to this region in the presence or absence of octopine, and that octopine induces conformational changes in Occr and/or its target site. We hypothesize that; i) OccR bends its target site, that octopine incites modulation of this bending, and that these changes result in the induction of transcription, or ii) that octopine-induced conformational changes in OccR itself result in transcriptional activation. The proposed investigation will employ biochemical, molecular biological, and genetic techniques to test these hypotheses.

根癌农杆菌对植物的作用过程已得到充分研究 现象，并且是自然遗传的唯一真实例子 工程。感染细菌转移其 DNA 片段 从肿瘤诱导（Ti）质粒进入宿主植物基因组。 基因 该片段编码直接肿瘤形成和合成 不寻常的糖和氨基酸，称为opines，其作用是 农杆菌特异性碳/氮源。 反过来，意见 刺激根癌农杆菌基因的表达，这些基因参与其 Ti 质粒的摄取和利用、接合转移以及 可能参与肿瘤定植的许多其他功能。 在章鱼碱型质粒中，这种诱导至少部分是由 LysR 型转录激活因子 OccR，可诱导 这些基因响应奥品章鱼碱而转录，并且 还自动调节其自身的合成。 OccR 的目标之一 调控是重叠的、不同转录的启动子区域 occ（章鱼碱分解代谢）操纵子和 occR 基因本身之间。 这种不寻常的遗传组织是 LysR 型系统的典型特征 可能允许同时控制两个操纵子。 在提议的 研究中，我们将研究 OccR 与此的相互作用 转录控制区。 有证据表明 OccR 结合 在存在或不存在章鱼碱的情况下到达该区域，并且章鱼碱 诱导 Occr 和/或其靶位点的构象变化。 我们 假设； i) OccR 弯曲其目标位点，章鱼碱会激发 这种弯曲的调制，并且这些变化导致 转录的诱导，或 ii) 章鱼碱诱导的构象 OccR 本身的变化会导致转录激活。 这 拟议的研究将采用生物化学、分子生物学、 和遗传技术来检验这些假设。