Optogenetic control of NOTCH1 signalling

The Notch signalling pathway is a crucial regulator of cell differentiation as well as tissue organisation. Dysregulation of Notch signalling has been linked to the pathogenesis of different diseases. Notch plays a key role in breast cancer progression by controlling the interaction between the tumour cells and the microenvironment as well as by increasing cell motility and invasion. NOTCH1 is a mechanosensitive receptor, where mechanical force is required to activate the proteolytic cleavage and release of the Notch intracellular domain (NICD). Here, we circumvent this step by regulating Notch activity by light. To achieve this, we have engineered a membrane-bound optogenetic NOTCH1 receptor (optoNotch) to control the activation of NOTCH1 intracellular domain (N1ICD) and its downstream transcriptional activities. Using optoNotch we confirm that NOTCH1 activation increases cell proliferation in MCF7 and MDA-MB-468 breast cancer cells in 2D and spheroid 3D cultures. OptoNotch allows fine-tuning ligand-independent regulation of N1ICD to understand the spatiotemporal complexity of Notch signalling.