RoL: EAGER: DESYN-C3 Membraneless organelles by design: a biomimetic approach

RoL：EAGER：DESYN-C3 无膜细胞器设计：仿生方法

• 批准号: 1844327
• 负责人: Giovanna Ghirlanda
• 金额: $ 30万
• 依托单位: Arizona State University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2021-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1844327&HistoricalAwards=false
• 关键词: RoL; EAGER; DESYN; C3; Membraneless

The cell has to organize a large number of reactions. It accomplishes this by isolating certain reactions and molecules inside of compartments known as organelles. Sometimes these organelles exist within membranes. Sometimes they exist simply as a distinct liquid phase. Controlling organelle formation and composition is central to creating a synthetic cell. Synthetic cells could be used in the biomanufacturing of chemicals, biofuels, and therapeutics, among potential applications. This project will investigate whether synthetic organelles without membranes can house complex reaction sequences. High school students will be involved in this project directly, through hands-on training in synthetic biology and participation in the international genetically engineered machine (iGEM) competition. Their challenge will be to develop protein tags that can trigger formation of the liquid phase organelles. They will also share their experiences on a blog. This will hopefully increase their enthusiasm and interest in pursuing careers in STEM-related fields.Liquid-liquid phase separation (LLPS) of proteins is a fundamental process in living cells. It underpins the formation of functional, non-membrane bound, liquid-like compartments involved in cell function and development. Examples include the nucleolus, Cajal bodies, nuclear speckles, germ granules, processing bodies, stress granules, and cell signaling compartments. These membraneless organelles can separate and concentrate specific sets of molecules in a highly dynamic manner. They also exchange components with their microenvironments rapidly. Within the liquid droplet, proteins maintain or acquire a folded structure, and conserve functions such as specific protein recognition. Increasingly, these structures are being implicated in diseases caused by protein aggregation. In this project, we will test whether LLPS can be exploited to enhance catalysis. An organelle that brings together elements of an enzyme cascade will be constructed and evaluated. These membraneless organelles will be evaluated by expressing the modular proteins in vivo and visualizing the formation of LLPS droplets by microscopy. Further, we will extend this approach to enable enzymatic cascades that include artificial metalloenzymes. If successful, this proposal will establish an innovative, simple method to generate organelles for synthetic biology, complementing existing methods such as encapsulation by protein shells or by vesicles.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.

细胞必须组织大量的反应。它通过将某些反应和分子隔离在被称为细胞器的隔间内来实现这一点。有时这些细胞器存在于细胞膜内。有时它们只是作为一种独特的液相存在。控制细胞器的形成和组成是创造合成细胞的核心。 合成细胞可用于化学品，生物燃料和治疗剂的生物制造，以及潜在的应用。这个项目将研究没有膜的合成细胞器是否可以容纳复杂的反应序列。高中生将通过合成生物学的实践培训和参加国际遗传工程机器（iGEM）竞赛直接参与该项目。他们的挑战将是开发能够触发液相细胞器形成的蛋白质标签。他们还将在博客上分享他们的经历。这将有望提高他们在干细胞相关领域从事职业的热情和兴趣。蛋白质的液-液相分离（LLPS）是活细胞中的基本过程。它支持参与细胞功能和发育的功能性、非膜结合、液体样隔室的形成。例子包括核仁、卡哈尔体、核斑点、胚芽颗粒、加工体、应激颗粒和细胞信号室。这些无膜细胞器可以以高度动态的方式分离和浓缩特定的分子组。它们还与它们的微环境快速交换组件。在液滴内，蛋白质保持或获得折叠结构，并保留诸如特异性蛋白质识别的功能。这些结构越来越多地与蛋白质聚集引起的疾病有关。在这个项目中，我们将测试LLPS是否可以用来增强催化作用。一个细胞器，汇集了酶级联的元素将被构建和评估。这些无膜细胞器将通过在体内表达模块蛋白并通过显微镜观察LLPS液滴的形成来评估。此外，我们将扩展这种方法，使酶级联反应，包括人工金属酶。如果成功的话，该提案将建立一种创新的、简单的方法来产生用于合成生物学的细胞器，补充现有的方法，如蛋白质壳或囊泡封装。该奖项反映了NSF的法定使命，并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，被认为值得支持。