Collaborative Research: Designing Minimal Synthetic Cells Capable of Sensing and Self-Manipulation via Tunable Self-Assembly

合成细胞的发展对于确定细胞过程的最低要求和实现一系列新的生物医学应用具有重大意义。该项目旨在为设计合成细胞提供基础和实用的见解，这些合成细胞概括了基本的细胞功能，如传感、自我生长和生物反应。例如，感知是活细胞如何决定分子在囊泡膜上的合成、控制释放和包封的核心功能。细胞膜上的蛋白质在这些细胞功能中起关键作用，介导感知和细胞内级联反应。本项目将利用重组蛋白技术合理设计蛋白质构建块，并通过改造其组装的囊泡结构和性质，使其对外界刺激作出反应，从而展示这些功能。高分子物理学、合成生物学和蛋白质工程领域的跨学科合作将在该项目中进行，并结合外展教育努力，为佛罗里达州盖恩斯维尔和伊利诺伊州林肯郡不同级别的代表性不足的学生提供实践学习机会。该项目的主要目标是通过自下而上的方法设计球状蛋白囊泡（GPVs）的组成和物理性质，构建能够感知和响应外部信号的最小合成细胞。合成细胞的囊泡膜将由传感器头蛋白、亮氨酸拉链和弹性蛋白样多肽组成的重组融合蛋白制成。具体目标是1)通过调节传感器蛋白的横向结合来设计gpv来感知特定的化学物质，2)证明传感诱导的跨膜信号传导来激活报告转录，以及3)设计可编程的信号转导和感应时的级联扩增。该合作项目结合了对聚合物构建块的自组装，无细胞合成生物学以及蛋白质-配体相互作用和酶特异性的分子生物工程的透彻理解。最终，该项目设想了蛋白质组装合成细胞的高潜力，它可以在最小的细胞结构中操纵多种蛋白质功能。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

The development of synthetic cells is of great interest to identify the minimal requirements for cellular processes and to enable a range of novel biomedical applications. This project aims to provide both fundamental and practical insights into designing synthetic cells that recapitulate essential cellular functions, such as sensing, self-growth, and biological reactions. For example, sensing is a central function in how living cells make decisions for synthesis and controlled release and encapsulation of molecules throughout vesicle membranes. Proteins at cell membranes play a critical role in these cellular functions to mediate sensing and intracellular cascade reactions. This project will demonstrate these functions by rationally designing the protein building blocks using recombinant protein technology and by engineering their assembled vesicle structure and properties to make them respond to external stimuli. Interdisciplinary collaboration in the fields of polymer physics, synthetic biology, and protein engineering will be conducted in this project, combined with outreach educational efforts to provide hands-on learning opportunities to underrepresented students at different levels in Gainesville, Florida, and Lincolnshire, Illinois. The main objective of this project is to build minimal synthetic cells capable of sensing and responding to external signals, by engineering the compositions and physical properties of globular protein vesicles (GPVs) through the bottom-up approach. The vesicle membrane of a synthetic cell will be made from recombinant fusion proteins composed of sensor head proteins, leucine zippers, and elastin-like polypeptides. The specific objectives are 1) to engineer GPVs for sensing specific chemicals by tuning lateral association of sensor proteins, 2) to demonstrate the sensing-induced transmembrane signaling to activate transcription of reporters, and 3) to design programmable signal transduction and amplification cascades upon sensing. This collaborative project combines a thorough understanding of the self-assembly of polymeric building blocks, cell-free synthetic biology, and molecular bioengineering of protein-ligand interactions and enzyme specificity. Ultimately, this project envisions the high potential of the protein-assembled synthetic cells, which can manipulate diverse protein functions in minimal cell structure.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.