Protrusive and contractile cell-matrix adhesions have important roles in cell adhesion, movement and signaling. Until recently, there has been a paucity of systems with which to research cell protrusions at molecular level. This laboratory previously discovered that thrombospondin-1, a cell-adhesion molecule, specifically induces protrusive lamellipodia, spikes and filopodia that contain the actin cross-linking protein, fascin, in core F-actin bundles. These fascin-containing structures are necessary for cell adhesion and migration on TSP-1 substrata, contribute to adhesion and migration on fibronectin substrata and are also formed in response to certain peptide growth factors. We have identified a novel molecular pathway by which thrombospondin-1 acts through clustering of the cell-surface proteoglycan syndecan-1 to stimulate the assembly of fascin-containing protrusions. The central hypothesis of the proposed research is that molecular components necessary for assembly of fascin-containing cell protrusions participate in protein complexes that contribute to the regulation of cell-matrix adhesion and cell movement. The proposed research focuses on the elucidatation of the molecular processes downstream of syndecan- 1 that mediate the organization of fascin cytoskeletal structures, the initial integration of these molecular processes into cell signaling networks, and the identification of mechanisms by which fascin-containing protrusions contribute to cell movements in three-dimensional matrix. These goals will be addressed experimentally by, 1), elucidation of the molecular requirements for regions within the syndecan-1 extracellular and cytoplasmic domains in transducing the organization of fascin protrusions; 2), the discovery of key binding partners for the V region of the syndecan-1 cytoplasmic domain and how these are integrated into regulation of fascin organization through the activation of Cdc42 and Rac small GTPases ; 3), the identification of molecular processes by which fascin structures contribute to migration in 3-dimensional matrix. Success in these complementary aims will advance new knowledge and understanding of the roles of fascin-based cytoskeletal structures in the important fundamental cellular activities of matrix-adhesion and cell migration. By opening up a new understanding of the molecular processes downstream of a specific extracellular inducer of fascin-containing protrusions, the proposed research will develop knowledge that is an essential basis for further investigation of fascin protrusions and associated regulatory molecules as therapeutic targets.
