Fig. 5: Conditional myogenic conversion by bioorthogonal ligation, photo-uncaging, and denatured collagen using fluorescein adapter-induced αFITC(E2)-SynNotch signaling.

a Schematic depicting αFITC(E2)-SynNotch containing a TetR-VP48 ICD. The receptor is expressed in C3H10T1/2 fibroblasts containing a TRE-regulated target gene encoding p65-MyoD-T2A-DsRed2. b Depiction of the signaling-mediated conversion of C3H10T1/2 fibroblasts in response to fluorescein-induced αFITC(E2)-SynNotch activation. Expression of p65-MyoD-T2A-DsRed2 results in the formation of red fluorescent syncytia exhibiting myogenic phenotypes. c C3H10T1/2 fibroblasts grown in TCO-BSA coated wells were left untreated (left) or treated with 2 nM Tz-5-Fluorescein (right). Cells were imaged 48 h following treatment. d C3H10T1/2 fibroblasts were grown in microwells containing immobilized and photo-patterned BSA-PC-5-fluorescein. Cells were imaged after 72 h of growth on the patterned surfaces. Expression of the myogenic marker protein myosin heavy chains (MHC) was confirmed via immunofluorescence staining. e Schematic depicting the denaturation of collagen triple helices followed by the binding of CHP-5-fluorescein to immobilized collagen strands. f Microwells containing native (left) or denatured (right) collagen-I proteins were treated with CHP-5-fluorescein (2 µM) prior to the addition of C3H10T1/2 cells. Cells were imaged after 48 h of growth. Experiments in c, d, and f were repeated independently with similar results. The schematic in (a) and cartoon components from (b) were created in BioRender⁸⁷.