CAREER: Multicolor imaging of single mRNA translational heterogeneity in living cells

该项目的目标是了解mRNA到蛋白质的翻译是如何在活细胞的单分子水平上进行编排的。通过创建荧光显微镜的开源工具，长期存在的基因调控模型将通过直接可视化进行测试，以更好地理解并最终控制自然和工程生命系统中的蛋白质合成。该项目将为学生和初级科学家提供跨学科的国际研究经验，培养他们在先进的单分子显微镜、生物传感器工程和基因调控的生物物理建模方面的能力。该项目还将把廉价的、开源的视频显微镜纳入多管齐下的STEM推广和教育工作中，旨在通过令人兴奋的公民科学吸引不同的K-12学生和教师。虽然蛋白质合成对生命至关重要，但mRNA翻译成蛋白质的实时动态很难在体内捕获。这个问题留下了许多关于蛋白质合成如何在生命系统中被调节的悬而未决的问题，特别是在mRNA翻译起始和延伸的水平上。这个项目将通过结合单分子显微镜和多色生物传感器的创造来回答这些问题，这些传感器实际上照亮了活细胞中蛋白质合成的异质性，一次一种蛋白质。通过这种强大的技术组合，在单个mRNA翻译水平上密码子使用和开放阅读框选择的冲突模型将首次直接可视化和测试。这项工作将为体内蛋白质合成的实时分析奠定基础，在基础研究和生物技术工业中具有广泛的应用前景。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 goal of this project is to understand how translation of mRNA to protein is orchestrated at the single molecule level in living cells. Through the creation of open-source tools for fluorescence microscopy, long-standing models of gene regulation will be tested by direct visualization, to better understand and ultimately control protein synthesis in both natural and engineered living systems. This project will provide students and junior scientists with an interdisciplinary and international research experience that will train them in advanced single-molecule microscopy, biosensor engineering, and biophysical modeling of gene regulation. This project will also incorporate inexpensive, open-source video microscopes into a multi-pronged STEM outreach and education effort designed to engage a diverse population of K-12 students and teachers with exciting citizen science. Although protein synthesis is essential to life, the real-time dynamics of mRNA translation to protein have been difficult to capture in vivo. This problem has left many open questions about how protein synthesis is regulated in living systems, particularly at the level of mRNA translation initiation and elongation. This project will answer such questions by combining single-molecule microscopy with the creation of multicolor biosensors that literally light up the heterogeneity of protein synthesis in living cells, one protein at a time. With this powerful combination of technologies, conflicting models of codon usage and open reading frame selection at the level of single mRNA translation will be directly visualized and tested for the first time. This work will lay the foundation for real-time analysis of protein synthesis in vivo, with broad application to both basic research and the biotechnology industry.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.