DBP6: Integrin
Integrins are major adhesion receptors that transmit signals bidirectionally across the plasma membrane, playing significant roles in diverse biological processes including cell migration, morphogenesis, immune response, and vascular hemostasis. Subversion of integrin function and signaling pathways contributes to the progression of a wide variety of diseases, including cancer, inflammatory disease, thrombosis and infection. For example, leukocyte adhesion deficiency, a genetic disorder that affects the body's immune system, is caused by inherited defects of integrins; Neurofibromatosis type II, characterized by the formation of multiple nervous system tumors, is another disease found to correlate with integrin mutation. Since integrins are involved in the pathogenesis of a diverse array of acquired and hereditary diseases, they are major targets for therapeutic intervention.
Integrins contain two non-covalently associated type I transmembrane glycoprotein α and β subunits; each subunit contains large extracellular domains, a single-spanning transmembrane domain, and a short cytoplasmic domain. The ability of integrin extracellular domains to bind ligands ("activation") regulates cell adhesion and signal transduction. Prior studies have demonstrated that integrin activation correlates with rearrangements in the extracellular, transmembrane and cytoplasmic domains. The overall atomic-level structures of integrin ectodomains, stemming from the crystal structures of αVβ3, αIIbβ3 and the recently resolved αXβ2, all show a compact bent conformation with a low affinity for ligands. Electron microscopy and biochemical studies have demonstrated that integrins also adopt extended conformations, either with a closed headpiece or with an open headpiece. Extension of bent integrins exposes the ligand-binding sites; swing-out of the hybrid domain results in an open conformation with a higher affinity for ligands.
Integrin domain organization and cartoon models of bidirectional signaling. B: bent conformation; E: extended, closed conformation; O: extended, open conformation.
A key feature of integrins is that through binding to both extracellular and intracellular ligands, integrins provide a transmembrane link for bidirectional transmission, namely outside-in and inside-out signaling, of mechanical force and biochemical signals across the plasma membrane. During outside-in signaling, changes in the extracellular environment like stress or ion/ligand concentration, induce integrin to transition from a bent conformation to an extended one with closed headpiece (B→E transition); ligand binding further induces the closed headpiece of integrin to open (E→O transition), resulting in conformational changes in the integrin transmembrane and cytoplasmic domains that mediate outside signals into the cell. During inside-out signaling, intracellular ligands, e.g., talin, bind to the β- tail, causing conformational changes of extracellular domains, thereby increasing integrin affinity for extracellular ligands. Inside-out signaling allows cells to regulate cell adhesion and migration by controlling the activity of their integrins. Current experimental techniques have very limited potential to capture the details of integrin conformational transition during signal transduction, therefore, the molecular basis of integrin bidirectional transmembrane signaling, e.g., the relationship between activation and conformational change, remains unclear.
The Center's parallel molecular dynamics (MD) simulation program NAMD has proven to be successful in studying the mechanical response of biological macromolecules to environmen- tal signals. Through MD simulations, the Center has discovered many mechanistic details of biomolecular systems, such as the muscle protein titin, the neural cell adhesion molecule complex, the blood clot component fibrinogen, and the ubiquitous cellular mechanics proteins cadherin and ankyrin, in many cases predicting biomolecular behavior and guiding experimental measurement.
In the case of integrin, MD simulation provides a promising tool to investigate, at the atomic level, the response of integrin to ligand binding during outside-in and inside-out signaling. Pioneering MD simulations of integrins have revealed some key elements of signal transduction. The Center's collaborator, Timothy A. Springer (Harvard U.), has successfully crystallized αIIbβ3 integrins adopting a closed conformation in the absence of ligand and an open conformation with ligand bound. The Springer lab has also crystallized five intermediate conformations of integrin during ligand-induced E→O transition (see letter of support). These new structures shed light on how the conformational signals are generated and transmitted by integrins, but the dynamical detail needs to be investigated through MD simulations. Another collaborator, Taekjip Ha (UIUC), performed fluorescence measurements to monitor the pulling force that integrins exert on the Arg-Gly-Asp (RGD) tripeptide (see letter of support), a cell adhesion motif on the extracellular matrix protein fibronectin studied extensively by the Center. MD simulations are called for in order to reveal the molecular origin of adhesion strength between integrin and the RGD motif, continuing our study in.
To determine the molecular mechanism of integrin signaling and, therefore, to assist development of new therapeutical strategies to integrin-related diseases, the Center will adapt its program NAMD to machines with over 100,000 cores achieving petascale power and develop advanced computational methods, such as hybrid QM/MM (quantum mechanics/molecular mechanics) and enhanced sampling techniques, to accurately describe the process of integrin signal transduction. Based on new integrin structures and new data of integrin adhesion strength provided by the collaborators, as well as new computational technology being developed in the Center's programs NAMD and VMD, we will study systematically the bidirectional signal transduction of integrin, namely the B to E (Aim 1) and the E to O (Aim 2) transitions of αIIbβ3 integrin during outside-in signaling, and talin-dependent inside-out signaling (Aim 3).
