Designing and implementing synthetic biological pattern formation remains challenging due to underlying theoretical complexity as well as the difficulty of engineering multicellular networks biochemically. Here, we introduce a cell-in-the-loop approach where living cells interact through in silico signaling, establishing a new testbed to interrogate theoretical principles when internal cell dynamics are incorporated rather than modeled. We present an easy-to-use theoretical test to predict the emergence of contrasting patterns in gene expression among laterally inhibiting cells. Guided by the theory, we experimentally demonstrate spontaneous checkerboard patterning in an optogenetic setup, where cell-to-cell signaling is emulated with light inputs calculated in silico from real-time gene expression measurements. The scheme successfully produces spontaneous, persistent checkerboard patterns for systems of sixteen patches, in quantitative agreement with theoretical predictions. Our research highlights how tools from dynamical systems theory may inform our understanding of patterning, and illustrates the potential of cell-in-the-loop for engineering synthetic multicellular systems.

由于潜在的理论复杂性以及生物化学工程多细胞网络的难度，设计和实现合成生物模式的形成仍然具有挑战性。在这里，我们引入了一种细胞在环方法，其中活细胞通过计算机信号进行相互作用，建立了一个新的测试平台，以在合并内部细胞动力学而不是建模时质疑理论原理。我们提出了一种易于使用的理论测试来预测横向抑制细胞中基因表达对比模式的出现。在该理论的指导下，我们通过实验证明了光遗传学装置中的自发棋盘图案，其中细胞间信号传导是通过实时基因表达测量在计算机中计算出的光输入来模拟的。该方案成功地为十六个斑块的系统产生了自发的、持久的棋盘图案，与理论预测在数量上一致。我们的研究强调了动力系统理论的工具如何帮助我们理解模式，并说明了细胞在环技术在工程合成多细胞系统方面的潜力。