Notch signaling is highly conserved in most animals and plays critical roles during neurogenesis as well as embryonic development. Synthetic Notch-based systems, modeled from Notch receptors, have been developed to sense and respond to a specific extracellular signal. Recent advancement of synNotch has shown promise for future use in cellular engineering to treat cancers. However, synNotch from Morsut et al. (2016) has a high level of ligand-independent activation, which limits its application. Here we show that adding an intracellular hydrophobic sequence (QHGQLWF, named as RAM7) present in native Notch, significantly reduced ligand-independent activation. Our enhanced synthetic Notch receptor (esNotch) demonstrates up to a 14.6-fold reduction in ligand-independent activation, without affecting its antigen-induced activation efficiency. Our work improves a previously reported transmembrane receptor and provides a powerful tool to develop better transmembrane signaling transduction modules for further advancement of eukaryotic synthetic biology.

Notch信号在大多数动物中高度保守，在神经发生和胚胎发育过程中发挥着关键作用。以Notch受体为模型的基于Notch的合成系统已被开发用于感知和响应特定的细胞外信号。 synNotch 的最新进展显示出未来在细胞工程中用于治疗癌症的前景。然而，Morsut 等人的 synNotch。 (2016)具有高水平的配体依赖性激活，这限制了其应用。在这里，我们表明，添加存在于天然Notch中的细胞内疏水序列（QHGQLWF，命名为RAM7），可以显着减少配体依赖性激活。我们的增强型合成 Notch 受体 (esNotch) 表现出配体依赖性激活最多减少 14.6 倍，而不影响其抗原诱导的激活效率。我们的工作改进了先前报道的跨膜受体，并为开发更好的跨膜信号转导模块提供了强大的工具，以进一步推进真核合成生物学。