Kinetic modeling of the CRISPR-Cas9 genome editing system

密苏里大学哥伦比亚分校的陈世杰教授获得了化学系化学理论、模型和计算方法项目的奖励。Chen博士开发了计算和理论方法来研究集群规则间隔短回文重复序列(CRISPR)- CRISPR相关蛋白9 （Cas9）基因组编辑过程的物理机制。CRISPR-Cas9基因组编辑是一种越来越强大的工具，其广泛的应用超出了我们的预期。然而，控制CRISPR基因编辑过程的物理原理尚不清楚，因此，基因编辑的合理设计和定量预测仍然是一个重大挑战。陈博士及其同事将运用物理和化学理论揭示CRISPR基因编辑的动力学机制，建立系统动力学特性与基因编辑效率之间的定量关系。该研究成果将通过免费访问的软件和web服务器部署，将导致新的、准确的CRISPR- cas9基因组编辑设计，并影响CRISPR的应用，从更好的作物物种遗传操作到更精确的各种疾病基因治疗。在教育和推广方面，这项研究将为课程开发提供新的材料，为学生和博士后学者创造多学科培训机会，并从长远来看，培养我们在理论和计算化学和生物学方面的下一代领导者。这项研究的核心是开发CRISPR-Cas9基因编辑系统的动力学模型。在项目的第一部分，为了找到基因组编辑的关键动力学步骤，陈博士和同事将使用随机森林方法从各种实验收集的大型数据集中提取关键成分。在项目的第二部分，通过关注关键的动力学步骤，他们将采用统计力学理论来模拟r环形成的动力学途径，这是基因编辑动力学的关键步骤。在项目的第三部分，陈博士和同事们正在使用实验数据测试、完善和验证模型。本研究的结果将是一个新的模型，能够预测CRISPR-Cas9系统的动力学机制，如动力学途径、过渡态、动力学中间体和速率常数，并对于给定的CRISPR-Cas9系统，估计靶向和脱靶编辑的基因组编辑效率。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。

Professor Shi-Jie Chen of the University of Missouri-Columbia is supported by an award from the Chemical Theory, Models and Computational Methods program in the Division of Chemistry. Dr. Chen develops computational and theoretical methods to investigate the physical mechanisms for the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)- CRISPR-associated protein 9 (Cas9) genome editing process. CRISPR-Cas9 genome editing is an increasingly powerful tool with widespread applications beyond all our expectations. However, the physical principles governing the CRISPR gene editing process are unclear and as a result, the rational design and quantitative prediction for gene editing remain a significant challenge. Dr. Chen and coworkers will employ physical and chemical theories to uncover the kinetic mechanism of CRISPR gene editing and establish the quantitative relationship between the kinetic properties of the system and the gene editing efficiency. The research result, deployed through freely accessible software and web servers, will lead to new, accurate designs of CRISPR-Cas9 genome editing and impact CRISPR applications from better genetic manipulation of crop species to more precise gene therapy for various diseases. On the educational and outreach front, this research will lead to new materials for curriculum development, create multi-disciplinary training opportunities for students and postdoctoral scholars, and in the long term, train our next generation leaders in theoretical and computational chemistry and biology. At the center of this research is the development of a kinetic model for CRISPR-Cas9 gene editing system. In the first part of the project, to find the critical kinetic step in genome editing, Dr. Chen and coworkers will use a random forest method to extract the key ingredients from the large data sets collected from various experiments. In the second part of the project, by focusing on the critical kinetic steps, they will employ statistical mechanical theory to model kinetic pathways of the R-loop formation, the critical step in gene editing kinetics. In the third part of the project, Dr. Chen and coworkers are testing, refining, and validating the model using experimental data. The result of this research will be a new model capable of predicting the kinetic mechanism of the CRISPR-Cas9 system, such as kinetic pathways, transition states, kinetics intermediates, and rate constants, and for a given CRISPR-Cas9 system, estimating the genome editing efficiency for both on- and off-target 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.