Yang Xiaoyu, Rocks Jason W, Jiang Kaiyi, Walters Andrew J, Rai Kshitij, Liu Jing, Nguyen Jason, Olson Scott D, Mehta Pankaj, Collins James J, Daringer Nichole M, Bashor Caleb J
Department of Bioengineering, Rice University; Houston, TX 77030, USA.
Graduate Program in Systems, Synthetic and Physical Biology, Rice University; Houston, TX 77030, USA.
bioRxiv. 2023 Nov 14:2023.09.11.557100. doi: 10.1101/2023.09.11.557100.
Protein phosphorylation signaling networks play a central role in how cells sense and respond to their environment. Here, we describe the engineering of artificial phosphorylation networks in which "push-pull" motifs-reversible enzymatic phosphorylation cycles consisting of opposing kinase and phosphatase activities-are assembled from modular protein domain parts and then wired together to create synthetic phosphorylation circuits in human cells. We demonstrate that the composability of our design scheme enables model-guided tuning of circuit function and the ability to make diverse network connections; synthetic phosphorylation circuits can be coupled to upstream cell surface receptors to enable fast-timescale sensing of extracellular ligands, while downstream connections can regulate gene expression. We leverage these capabilities to engineer cell-based cytokine controllers that dynamically sense and suppress activated T cells. Our work introduces a generalizable approach for designing and building phosphorylation signaling circuits that enable user-defined sense-and-respond function for diverse biosensing and therapeutic applications.
蛋白质磷酸化信号网络在细胞感知和响应环境的过程中起着核心作用。在此，我们描述了人工磷酸化网络的构建，其中“推挽”基序——由相反的激酶和磷酸酶活性组成的可逆酶促磷酸化循环——由模块化蛋白质结构域部分组装而成，然后连接在一起，在人类细胞中创建合成磷酸化电路。我们证明，我们的设计方案的可组合性能够实现对电路功能的模型引导调整以及建立多样化网络连接的能力；合成磷酸化电路可以与上游细胞表面受体偶联，以实现对细胞外配体的快速时间尺度感知，而下游连接可以调节基因表达。我们利用这些能力设计基于细胞的细胞因子控制器，以动态感知和抑制活化的T细胞。我们的工作引入了一种可推广的方法，用于设计和构建磷酸化信号电路，从而为各种生物传感和治疗应用实现用户定义的感知和响应功能。
