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CAREER: A Synthetic Biology Platform to Map and Engineer the Diverse Epigenetic Space

职业：绘制和设计多样化表观遗传空间的合成生物学平台

基本信息

批准号：
2144539
负责人：
Albert Keung
金额：
$ 82.58万
依托单位：
North Carolina State University
依托单位国家：
美国
项目类别：
Continuing Grant
财政年份：
2022
资助国家：
美国
起止时间：
2022-02-01 至 2027-01-31
项目状态：
未结题

来源：
https://www.nsf.gov/awardsearch/showAward?AWD_ID=2144539&HistoricalAwards=false
关键词：
CAREER Synthetic Biology Platform Map

项目摘要

This award is funded in whole or in part under the American Rescue Plan Act of 2021 (Public Law 117-2).This project will develop new technological platforms and educational innovations that will unlock our understanding of and ability to control the expression of genes in higher order organisms from yeast and plants to humans, with broad applications in biotechnology. Chromatin is the layer of hundreds of proteins bound on top of the genome. It confers regulatory handles over diverse cellular and organismal processes including aging and cancer, and it could be leveraged in biotechnological applications including bioremediation, environmental sensing, and bio-based production of commodity chemicals. Chromatin also provides a strong topic for pedagogical innovation and enrichment. This project will develop hands-on experiments for middle and high school students that connect molecular changes in chromatin to effects observable by eye, in this case the binding of yeast cells to the surface of an agar plate. Furthermore, the numerical and biochemical diversity of chromatin advocates for new pedagogical approaches at the collegiate and PhD levels. Specifically, this project will develop new coursework modules that train the next generation of scientists in how to handle and analyze large data sets. American Rescue Plan funding of this project will support the investigator at a critical stage in his career.Chromatin is incredibly diverse at the molecular level. There are more than 80 different amino acid residues on five histone proteins known to undergo over 20 distinct post-translational modifications. Hundreds of proteins, often containing domains referred to as writers and readers, both regulate and interact with this complex biochemical palette. Through the development of high content technologies paired with biochemical and genetic approaches it has become clear that much of this diversity is functional. However, unlike many methods that can characterize chromatin at high throughput, functional assays remain substantially more limited in scale. In particular, characterizing and engineering the enzymatic activities and specificities of epigenetic writers still rely largely on low throughput biochemical assays that require recombinant protein production and purification that are challenging to scale even with robotic liquid handling. This proposal seeks to build upon recent synthetic biology platforms the investigator and his team have developed using yeast surface display and epigenome editors to unlock access to the diversity of chromatin. With a focus on histone acetyltransferases (HAT), they will establish proof-of-principle demonstrations that these systems can 1) map the residue specificities of all known human HATs; 2) connect sequence to function by accessing the very large enzyme and substrate sequence space by directed evolution approaches; and 3) probe the impact of molecular context on function. While the investigations focus on HATs, the investigator and his team seek to motivate the future expansion and application of this experimental platform to the diverse biochemistry of chromatin.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.

该奖项的全部或部分资金均根据《2021 年美国救援计划法案》（公法 117-2）提供。该项目将开发新技术平台和教育创新，以解锁我们对从酵母和植物到人类的高级生物体中控制基因表达的理解和能力，并在生物技术领域具有广泛的应用。染色质是结合在基因组顶部的数百种蛋白质的层。它赋予对包括衰老和癌症在内的多种细胞和有机过程的监管能力，并且可以在生物技术应用中利用，包括生物修复、环境传感和商品化学品的生物基生产。染色质还为教学创新和丰富提供了一个强有力的主题。该项目将为中学生和高中生开发动手实验，将染色质的分子变化与肉眼可观察到的效应联系起来，在本例中是酵母细胞与琼脂板表面的结合。此外，染色质的数量和生化多样性提倡在大学和博士水平上采用新的教学方法。具体来说，该项目将开发新的课程模块，培训下一代科学家如何处理和分析大型数据集。美国救援计划对该项目的资助将在研究人员职业生涯的关键阶段提供支持。染色质在分子水平上具有令人难以置信的多样性。已知五种组蛋白上有超过 80 个不同的氨基酸残基，可经历 20 多种不同的翻译后修饰。数百种蛋白质通常包含被称为写入器和读取器的域，它们调节这种复杂的生化调色板并与之相互作用。通过高内涵技术的发展与生化和遗传方法相结合，我们已经清楚地认识到这种多样性在很大程度上是具有功能性的。然而，与许多可以高通量表征染色质的方法不同，功能测定在规模上仍然受到很大限制。特别是，表征和设计表观遗传写入器的酶活性和特异性仍然很大程度上依赖于低通量生化测定，这些测定需要重组蛋白生产和纯化，即使使用机器人液体处理，也难以规模化。该提案旨在建立在研究者和他的团队使用酵母表面展示和表观基因组编辑器开发的最新合成生物学平台的基础上，以解锁染色质多样性。他们将重点关注组蛋白乙酰转移酶 (HAT)，建立原理验证演示，证明这些系统可以 1) 绘制所有已知人类 HAT 的残基特异性； 2) 通过定向进化方法访问非常大的酶和底物序列空间，将序列与功能连接起来； 3）探讨分子背景对功能的影响。虽然研究重点是 HAT，但研究人员和他的团队力求推动该实验平台在染色质多样化生物化学方面的未来扩展和应用。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。