Explore The Activation Efficiency Of Different Ligand Carriers On Synnotch-Based Contact-Dependent Activation System

Objectives: synNotch receptors can get different inputs about the outside environment into cells; however, synNotch system doesn't work for soluble ligands. This study aims to explore the activation efficiency of different ligand carriers on synNotch system.Methods: SynNotch-based contact-dependent activation system was constructed in 293T cells (293T-synNotch), then the synNotch receptor ligands eGFP proteins were provided by three different carriers: 400 nm Ni magnetic beads (Mag Ni-eGFP), 300 nm carboxyl modified Fe3O4 magnetic beads (Mag COOH-deGFP), and 4T1 cell (4T1-mGFP). Three eGFP carriers were incubated with 293T-synNotch, the activation efficiencies of these three eGFP carriers were evaluated by fluorescence microscopy.Results: The 293T-synNotch cell activated by Mag Ni-eGFP shows no observable red fluorescence; when Mag COOH-deGFP are incubated with 293T-synNotch, about 50%o cells with red fluorescence appeared, also eGFP proteins have not dropped off from the Mag COOH-deGFP beads; 4T1-mGFP cells show the highest activation efficiency, about 21% synNotch cells are activated, when 4T1-mGFP and 293T-synNotch are incubated at high cell density.Conclusion: The 4T1-mGFP cells are the most effective ligands for synNotch receptor activation.