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，使细胞能够发挥功能并对不断变化的环境做出反应。 1920年，弗雷德里克·格里菲斯（Frederick Griffith）的精子实验揭示了一些细菌细胞从环境中捕获DNA（从垂死的细胞中释放）。 这些细胞将捕获的DNA整合到它们的基因组中，使其正常发挥功能。 从历史上看，这种容易获得DNA的能力在植物和动物细胞中还没有观察到。 由此，一个普遍的观点是，植物和动物细胞主动保护其基因组免受来自环境的外来或病毒衍生的DNA的入侵，并且许多基因组保护策略是已知的。 这些保护机制阻止了DNA在实验室环境中有效地引入细胞，科学家可能旨在纠正遗传缺陷或在细胞中进行其他遗传操作。 该项目的目标是调查使用遗传上易处理的多细胞生物秀丽隐杆线虫从环境来源引入细胞的DNA的递送和细胞内运输途径。 除了获取知识，该项目还预测了基因操作和DNA递送的新策略。 本科生将从该项目中获得更广泛的利益，因为实验将为他们保留。 这些学生将包括那些来自STEM中代表性不足的群体和寻找夏季研究机会的小型学院的学生。在动物和植物的循环系统中可以检测到少量的各种DNA片段。 检测和分析循环DNA（circDNA）以诊断疾病，以及circDNA对细胞功能的影响，是一个新兴和不断扩大的研究领域。 circDNA激活高等生物体中的炎症反应，并已观察到掺入远距离细胞。 circDNA进入远端细胞并整合到该细胞基因组中的频率是一个相对未探索的问题。 使用秀丽隐杆线虫，我们将研究蛋白质介导的circDNA传递到远端细胞。 C.线虫缺乏许多在高等真核生物中发现的炎症途径，如cGAS-STING和NF-κ B核效应子反应。 因此，我们可以更容易和直接地研究暴露于循环的细胞外DNA的后果以及其获得细胞的后果。 如果成功，EAGER项目将通过有目的地引入DNA为遗传操作提供新的策略。 我们还可以了解到额外的细胞或发育后果circcDNA，后果，可以避免由于circDNA相关的炎症反应在高等organism.This奖项反映了NSF的法定使命，并已被认为是值得通过使用基金会的智力价值和更广泛的影响审查标准进行评估的支持。