EAGER: Overcoming transformation and germline expression barriers

EAGER：克服转化和种系表达障碍

• 批准号: 2300557
• 负责人: Lisa Timmons
• 金额: $ 30万
• 依托单位: University of Kansas Center for Research Inc
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-02-01 至 2025-01-31
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2300557&HistoricalAwards=false
• 关键词: EAGER; Overcoming; transformation; germline; expression

A cell’s genome, comprised of DNA, enables cells to function and respond to changing environments. Seminal experiments by Frederick Griffith in the 1920 revealed that some bacterial cells capture DNA from the environment (released from dying cells). The cells incorporated the captured DNA into their genomes where it functioned normally. Historically, this easy ability to acquire DNA has not been observed in plant and animal cells. From this, a common view is that plant and animal cells actively protect their genomes from invasion of foreign, or virus-derived, DNA encountered from the environment, and many genome-protection strategies are known. These protective mechanisms prevent the efficient introduction of DNA into cells in the laboratory setting where scientists might aim to correct a genetic defect or perform other genetic manipulations in cells. The goal of this project is to investigate delivery and intracellular trafficking routes for DNA introduced to cells from environmental sources using the genetically tractable, multicellular organism Caenorhabditis elegans. In addition to acquiring knowledge, new strategies for genetic manipulation and DNA delivery are predicted outcomes from this project. Undergraduate students will derive broader benefits from the project as experiments will be reserved for them. Such students will include those from groups underrepresented in STEM and from small colleges looking for summer research opportunities.Small amounts of various DNA fragments can be detected in the circulatory system of animals and plants. Detection and analysis of circulating DNA (circDNA) for the purpose of diagnosing disease, and the impact of circDNA on cell function, is an emerging and expanding field of research. circDNA activates inflammation responses in higher organisms and has been observed to incorporate into distant cells. How frequently circDNA manages to gain entry into distant cells and incorporate into that cell’s genome is a relatively unexplored question. Using Caenorhabditis elegans, we will investigate protein-mediated delivery of circDNA to distal cells. C. elegans lacks many inflammatory pathways found in higher eukaryotes, such as cGAS-STING and NF-kappaB nuclear effector responses. Thus, we can more easily and directly investigate the consequences of exposure to circulating, extracellular DNA and the consequences of its acquisition to cells. If successful, this EAGER project will provide new strategies for genetic manipulation through purposeful introduction of DNA. We may also learn of additional cell or developmental consequences to circDNA, consequences that may be avoided due to circDNA-related inflammatory responses in higher organisms.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

细胞的基因组由DNA组成，使细胞能够起作用并响应不断变化的环境。弗雷德里克·格里菲斯（Frederick Griffith）在1920年进行的精确实验表明，一些细菌细胞从环境中捕获了DNA（从垂死细胞释放）。细胞将捕获的DNA掺入其正常功能的基因组中。从历史上看，在动植物细胞中尚未观察到这种易于获得DNA的能力。从中，一个普遍的观点是，动植物细胞积极保护其基因组免受来自环境遇到的外国或病毒衍生的DNA的侵袭，并且已知许多基因组保护策略。这些受保护的机制阻止了在实验室环境中有效地将DNA引入细胞，科学家可能旨在纠正遗传缺陷或在细胞中执行其他遗传操作。该项目的目的是研究使用遗传性遗传性的多细胞生物Caenorhabditis秀中的环境来源引入细胞的DNA的传递和细胞内运输途径。除了获得知识外，该项目的结果预测了基因操纵和DNA传递的新策略。本科生将为他们保留实验，从该项目中获得更大的好处。这样的学生将包括来自STEM中代表性不足的小组和寻找夏季研究机会的小组的成员。在动物和植物的循环系统中可以检测到各种DNA碎片。出于诊断疾病的目的，循环DNA（CircDNA）的检测和分析是一个新兴和扩展的研究领域。 CircDNA激活较高生物体的炎症反应，并已观察到将其掺入远处的细胞中。 CircDNA的频率设法进入遥远的细胞并将其纳入该细胞的基因组是一个相对意外的问题。使用秀丽隐杆线虫，我们将研究蛋白质介导的CircDNA递送以区分细胞。秀丽隐杆线虫缺乏在较高的真核生物中发现的许多炎症途径，例如CGAS刺和NF-kappab核效应子反应。这是，我们可以更容易，直接研究暴露于循环，细胞外DNA及其对细胞的后果的后果。如果成功的话，这个渴望的项目将通过有目的地引入DNA为基因操作提供新的策略。我们还可能会了解CircDNA的其他细胞或发育后果，由于较高生物体中与CircDNA相关的炎症反应可能会避免的后果。该奖项反映了NSF的法定任务，并且被认为是通过基金会的知识分子优点和更广泛的影响审查标准来评估的值得支持的。