CAREER: Develop PROTAC-CID, a Novel Small Molecule Inducible Platform for Controllable Gene Expression in Mammalian Cells

职业：开发 PROTAC-CID，一种新型小分子诱导平台，用于哺乳动物细胞中可控基因表达

• 批准号: 2143626
• 负责人: Xue Gao
• 金额: $ 50.21万
• 依托单位: William Marsh Rice 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: 2022-07-01 至 2027-06-30
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2143626&HistoricalAwards=false
• 关键词: CAREER; Develop; PROTAC; CID; Novel

It is difficult to control gene expression in mammalian cells. Accomplishing this is key to developing effective cell therapies. Small molecules can exert that control. Unfortunately, small molecules are often toxic, and tend to have a limited, on-off capability. The aim of this project is to develop an improved small molecule system for mammalian cells. This system will be tunable instead of on/off. Highly sensitive, it will interact with multiple genes to direct more complex behaviors. Undergraduates from underrepresented groups and high school teachers will learn techniques of synthetic biology and CRISPR technology.Proteolysis Targeting Chimeric (PROTAC)-Chemically Induced Dimerization (CID) systems will be developed as sensitive, multiplex, tunable, and safe toolkits to control gene expression in mammalian cells. Combining PROTAC-CID with genome editors will enable precise genomic modification. This will greatly increase the accuracy and safety of using genome editors for basic and therapeutic applications. The specific research goals are to 1) establish a novel PROTAC-CID inducible gene expression system; 2) engineer multiplex inducible gene regulations; 3) design high-induction and low-basal level PROTAC-CID systems for Cre recombinase expression, and 4) develop PROTAC-CID inducible CRISPR base editors. Successful completion should result in novel PROTAC-CID inducible gene expression platforms for multiplex gene regulations in living eukaryotic cells. These systems will provide deep insights into the approaches to achieve high-induction and low basal-level gene expressions. Finally, applying PROTAC-CID systems to CRISPR base editors will permit inducible genomic DNA modification at a defined time to reduce the off-target effect and increase the accuracy and safety in genome editing.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.

在哺乳动物细胞中控制基因表达是困难的。实现这一点是开发有效细胞疗法的关键。小分子可以发挥这种控制作用。不幸的是，小分子通常是有毒的，并且往往具有有限的开关能力。该项目的目的是为哺乳动物细胞开发一种改进的小分子系统。该系统将是可调的，而不是开/关。 高度敏感，它将与多个基因相互作用，以指导更复杂的行为。来自弱势群体的本科生和高中教师将学习合成生物学和CRISPR技术。蛋白质水解靶向嵌合（PROTAC）-化学诱导二聚化（CID）系统将被开发为敏感、多元、可调和安全的工具包，以控制哺乳动物细胞中的基因表达。将PROTAC-CID与基因组编辑器相结合将实现精确的基因组修饰。这将大大提高使用基因组编辑器进行基础和治疗应用的准确性和安全性。具体研究目标为：1）建立新型PROTAC-CID诱导型基因表达系统; 2）设计多重诱导型基因调控; 3）设计高诱导低基础水平的PROTAC-CID Cre重组酶表达系统; 4）开发PROTAC-CID诱导型CRISPR碱基编辑器。成功完成应导致新的PROTAC-CID诱导型基因表达平台，用于活真核细胞中的多重基因调控。这些系统将为实现高诱导和低基础水平基因表达的方法提供深入的见解。最后，将PROTAC-CID系统应用于CRISPR碱基编辑器将允许在规定的时间进行诱导型基因组DNA修饰，以减少脱靶效应，提高基因组编辑的准确性和安全性。该奖项反映了NSF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。