Mechanosensitive signaling and cell adhesion during migration

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DESCRIPTION (provided by applicant): Cell migration is a complex process that involves both physical adhesive mechanics and intracellular signaling, which are profoundly interdependent on one another. Mechanisms of cell migration have been studied intensively using single cells cultured in vitro, but our understanding of cell migration in vivo remains limited. Cell migration in vivo often occurs in multicellular arrays that produce bulk tissue movements critical to morphogenesis, normal adult physiology, and various pathologies. While the function of mechanical signaling generated through cell-matrix traction forces has frequently been the subject of investigation, mechanosensitive responses to tension in cell-cell adhesions and the subsequent effects of these adhesive contacts on cell migration have not been elucidated. I will test the hypothesis that mechanical signaling, initiated by cadherin-mediated cell-cell adhesions in intact migratory tissues, has a central role in coordinating cell polarization and directional migration of individual cells to direct intact tissue movements. Mechanisms of cell migration are widely conserved between different species, tissues, and cell types. The migratory mesendoderm that traverses the fibronectin matrix of the blastocoel roof during Xenopus gastrulation will be used as a model system for these studies. This model provides an ideal system in which to study normal in vivo migratory processes because the motile behaviors of the intact tissue are retained when cultured in vitro and thus, the system is accessible to a variety of experimental manipulations. There are four specific aims proposed to address the significance of cell-cell adhesion in the propagation of mechanical signals that regulate mesendoderm cell polarity and migration. In specific aim 1, mesendoderm explants and a magnetic bead "pull" assay will be used to establish the role of biomechanical tension on C-cadherin dependent adhesions as an important regulator of polarization and directional migration in this tissue. Cadherins and integrins provide a physical link between extracellular ligands and internal cytoskeletal elements. GFP fusion proteins will be used to track cytoskeletal dynamics as a consequence of mechanical signaling initiated by cadherin adhesions in specific aim 2. Actin cytoskeletal dynamics involved in migration is signaled by spatiotemporal activation of the small Rho family GTPases. In specific aim 3, FRET analysis will be used to examine the local subcellular activation of Rho family GTPases as a result of C-cadherin and a5 (31 integrin-dependent mechanosensitive cell signaling. Finally, traction force microscopy and reflection contrast microscopy approaches will be used in specific aim 4 to determine the specific force relationship between cell-matrix adhesions and C-cadherin cell-cell contacts in migrating mesendoderm.