The Molecular Basis for Integrin-Mediated Bidirectional Signaling

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Leukocytes are essential immune components protecting the body against foreign invaders. Adhesion, migration, extravasation, and cell-cell communication are mediated though the bidirectional signaling of β2 integrins, which are integral membrane proteins found on the leukocyte surface. Due to their complex and multifunctional roles, dysregulation of β2 integrins is linked to autoimmune, cardiac and pulmonary pathologies as well as infectious diseases and several cancers. Although integrins are prime therapeutic targets, drug development has been hindered due to unanticipated side effects that arise from large gaps in our understanding of the mechanisms that drive specificity and integrin activation. I will decipher the molecular basis for β2 integrin activation, ligand recognition, and bidirectional signaling using an integrative approach. I'll build on my expertise in cryoEM method development to capture high- resolution conformational snapshots of isolated β2 integrin and ligand-bound complexes to reveal the dynamic structural rearrangements associated with signal transduction and identify key residues mediating ligand specificity. Using cell-surface expressed integrins, I'll assess the functional consequences of mutating these residues on binding of ligands and conformation- specific antibodies and on adhesion, phagocytosis, and cell motility. To gain broad insight into integrin allostery in a near-native context, I'll develop a membrane mimetic system using next- generation styrene maleic acid copolymers. Membrane lipids influence integrin activation and ligand binding and are key to forming stable complexes. These polymers will afford a stream- lined method to extract and purify nanodiscs embedded with pre- formed integrin-ligand complexes in their native environment. I will use this system to study integrin in complex with talin, the central integrin- activator protein that binds a conserved motif on the cytoplasmic region on integrin. This will reveal in molecular detail how integrins are activated to relay signals allosterically across the plasma membrane and define a molecular basis for bidirectional signaling as well as provide a framework for designing biochemical, biophysical, and mechano- sensitive experiments to study larger complexes and gain comprehensive insight into integrin function. Ultimately, this work will provide a structural blueprint for the rational design of therapeutics for autoimmune diseases, which is a long-term goal of the lab.