PROJECT SUMMARY/ABSTRACT ? PROJECT 4 Migration is an essential component of a functional immune system. Responding to both chemical and mechanical signaling cues, cells need to move in complex environments including both within and between tissues. To accomplish this, cells need to generate internal forces which can be coupled to their surrounding matrix through adhesions. Both of these physical interactions must be delicately balanced in time and space to ensure motility. While this balance has been extensively explored in the context of mesenchymal cell migration, these relationships remain ill-defined in the amoeboid migration modes used by T cells and other immune cells. Our overall goal is to define the mechanical interactions that enable and guide immune cell migration. Chemokines, which stimulate the protrusion machinery, and integrins, which mediate adhesion, are thought to be the primary biological effectors of T cell migration. Mechanically, cytoskeletal dynamics consisting of actin polymerization and myosin contractility are the predominant mechanisms for generating forces in cells. The interplay of these components defines the motility of the cell. We hypothesize that the migration behaviors of different immune cells lie along a single continuum, differing only in their relative contributions of adhesion and force generation. We further speculate that effector programming leads to differences in the activation thresholds for these physical interactions that may modify the way distinct effector subsets respond to their physical microenvironment. To test this hypothesis, we propose to make precise mechanical measurements of the migration machinery and determine how they affect the migration efficiency of T cells in vitro. We will then use our in vitro findings as a basis to interpret similar morphological behaviors and interactions in the more complex in vivo inflamed tissue environment. Aim 1: To determine the relation between actin polymerization and traction stress in T cells. Aim 2: To determine how ECM composition, organization and material properties regulate adhesion in T cells. Aim 3: Do T cells adapt their migration behavior to the microenvironment in vivo? Our findings will elucidate the underlying mechanical mechanisms regulating T cell migration and can be used to develop new targets for future therapies.