Department of Synthetic Biology and Immunology, National Institute of Chemistry, Ljubljana, Slovenia.
Graduate School of Biomedicine, University of Ljubljana, Ljubljana, Slovenia.
Nat Chem Biol. 2023 Jan;19(1):64-71. doi: 10.1038/s41589-022-01136-x. Epub 2022 Sep 26.
Chemically inducible systems represent valuable synthetic biology tools that enable the external control of biological processes. However, their translation to therapeutic applications has been limited because of unfavorable ligand characteristics or the immunogenicity of xenogeneic protein domains. To address these issues, we present a strategy for engineering inducible split protein regulators (INSPIRE) in which ligand-binding proteins of human origin are split into two fragments that reassemble in the presence of a cognate physiological ligand or clinically approved drug. We show that the INSPIRE platform can be used for dynamic, orthogonal and multiplex control of gene expression in mammalian cells. Furthermore, we demonstrate the functionality of a glucocorticoid-responsive INSPIRE platform in vivo and apply it for perturbing an endogenous regulatory network. INSPIRE presents a generalizable approach toward designing small-molecule responsive systems that can be implemented for the construction of new sensors, regulatory networks and therapeutic applications.
化学诱导系统是一种有价值的合成生物学工具，可实现对生物过程的外部控制。然而，由于配体特性不理想或异源蛋白结构域的免疫原性，它们在治疗应用中的转化受到了限制。为了解决这些问题，我们提出了一种工程诱导分裂蛋白调节剂（INSPIRE）的策略，其中来源于人的配体结合蛋白被分割成两个片段，在存在同源生理配体或临床批准药物的情况下重新组装。我们表明，INSPIRE 平台可用于在哺乳动物细胞中进行动态、正交和多重基因表达控制。此外，我们在体内证明了糖皮质激素响应型 INSPIRE 平台的功能，并将其应用于干扰内源性调控网络。INSPIRE 提供了一种可推广的小分子响应系统设计方法，可用于构建新型传感器、调控网络和治疗应用。
