Isolation and characterization of plant insulators

植物绝缘体的分离和表征

• 批准号: 2869-2011
• 负责人: Johnson, Douglas
• 金额: $ 1.75万
• 依托单位: University of Ottawa
• 依托单位国家: 加拿大
• 项目类别: Discovery Grants Program - Individual
• 财政年份: 2015
• 资助国家: 加拿大
• 起止时间: 2015-01-01 至 2016-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=568264
• 关键词: Isolation; characterization; plant; insulators

Insulators are DNA elements that are able to block inappropriate or unwanted enhancer-promoter communication (enhancer blocker activity) and/or prevent the spread of repressive chromatin into active chromatin (barrier activity). Their action is mediated by protein(s) that recognize them leading to the recruitment of many proteins and ultimately the assembly "insulator bodies" the proposed "functional unit". Thus insulators represent an additional level of transcriptional control and their study represents an opportunity to understand a basic, universal mechanism of gene regulation. Insulators from number of species have been identified including fungi, yeast, fruit fly, chicken and human but not plant. Our work indicated that in Arabidopsis two insulators, UASrpg and BEAD-1C can function as insulators-first published demonstration of insulator activity in a plant. Given the potential importance of insulators in plant gene expression and possible key roles in plant biotechnology for the control of tissue-specific transgene expression, we have embarked on a research programme to isolate and study plant insulators. Our approach is to avoid the use of candidate sequences and to tackle the problem by adapting our transgenic system for the direct selection of DNA elements with insulating activity. Sequence(s) thus isolated can be cloned into other vectors to eliminate insulators that are not functional in a variety of promoter/enhancer combinations, orientation and a range of different species. With such sequences in hand we can devise means to isolate the proteins that recognize them and test models of insulator activity proposed for animal systems. Our long term goal is to understand how these interactions regulate gene expression in plants and their evolution. In addition a function and universal plant insulator would have an impact on the design of a new generation of plant transformation vectors.

绝缘子是能够阻断不适当或不需要的增强子-启动子通讯（增强子阻断剂活性）和/或防止抑制性染色质扩散到活性染色质（屏障活性）的DNA元件。它们的作用由识别它们的蛋白质介导，导致许多蛋白质的募集，并最终组装成“绝缘体”，即所提出的“功能单位”。因此，绝缘子代表了转录控制的另一个水平，他们的研究代表了了解基因调控的基本，普遍机制的机会。 绝缘体来自许多物种，包括真菌、酵母、果蝇、鸡和人类，但不包括植物。我们的工作表明，在拟南芥中的两个绝缘子，UASrpg和BEAD-1C可以作为绝缘子的功能-第一次发表的绝缘子活动在植物中的演示。鉴于绝缘子在植物基因表达中的潜在重要性以及在植物生物技术中控制组织特异性转基因表达的可能关键作用，我们已经开始了一项研究计划，以分离和研究植物绝缘子。 我们的方法是避免使用候选序列，并通过调整我们的转基因系统直接选择具有绝缘活性的DNA元件来解决这个问题。如此分离的序列可以克隆到其他载体中以消除在多种启动子/增强子组合、取向和一系列不同物种中不起作用的绝缘子。有了这样的序列在手，我们可以设计出方法来分离识别它们的蛋白质，并测试为动物系统提出的绝缘子活动模型。我们的长期目标是了解这些相互作用如何调节植物中的基因表达及其进化。此外，功能和通用的植物绝缘子将对新一代植物转化载体的设计产生影响。