Phosphoproteomic Changes Induced by Cell-Derived Matrix and Their Effect on Tumor Cell Migration and Cytoskeleton Remodeling

Increased fibrotic extracellular matrix (ECM) deposition promotes tumor invasion, the first step of the metastatic cascade. Yet, the underlying mechanisms are poorly understood as conventional studies of tumor cell migration are often performed in 2D culture lacking the compositional and structural complexity of native ECM. Moreover, these studies frequently focus on select candidate pathways potentially overlooking other relevant changes in cell signaling. Here, we combine a cell-derived matrix (CDM) model with phosphotyrosine phosphoproteomic analysis to investigate tumor cell migration on fibrotic ECM relative to standard tissue culture plastic (TCP). Our results suggest that tumor cells cultured on CDMs migrate faster and in a more directional manner than their counterparts on TCP. These changes in migration correlated with decreased cell spreading and increased cell elongation. While the formation of phosphorylated focal adhesion kinase (pFAK)+ adhesion complexes did not vary between TCP and CDMs, time-dependent phosphoproteomic analysis identified that the SRC family kinase LYN may be differentially regulated. Pharmacological inhibition of LYN decreased tumor cell migration and cytoskeletal rearrangement on CDMs but also on TCP suggesting that LYN regulates tumor cell migration on CDMs in combination with other mechanisms. These data highlight how the combination of physicochemically complex in vitro systems with phosphoproteomics can help identify signaling mechanisms by which fibrotic ECM regulates tumor cell migration.