Local nascent protein deposition and remodeling guide mesenchymal stromal cell mechanosensing and fate in three-dimensional hydrogels

Hydrogels serve as valuable tools for studying cell-extracellular matrix (ECM) interactions in three-dimensional (3D) environments that recapitulate aspects of native ECM. However, the impact of early protein deposition on cell behavior within hydrogels has largely been overlooked. Using a bio-orthogonal labeling technique, we visualized nascent proteins within a day of culture across a range of hydrogels. In two engineered hydrogels of interest in 3D mechanobiology studies – proteolytically degradable covalently crosslinked hyaluronic acid (HA) and dynamic viscoelastic HA hydrogels – mesenchymal stromal cell (MSC) spreading, YAP/TAZ nuclear translocation, and osteogenic differentiation were observed with culture. However, inhibition of cellular adhesion to nascent proteins or reduction in nascent protein remodeling reduced MSC spreading and nuclear translocation of YAP/TAZ, resulting in a shift towards adipogenic differentiation. Our findings emphasize the role of nascent proteins in the cellular perception of engineered materials and have implications for in vitro cell signaling studies and application to tissue repair.