MECHANISTIC INSIGHTS INTO THE ?INSIDE-OUT? SIGNALING OF INTEGRINS

Carnegie-Mellon University, Pittsburgh PA

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Abstract

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. The overall goal of our research activities is to achieve a detailed mechanistic understanding of signal transduction processes triggered by molecular recognition in membrane protein complex systems. One of the funded research programs in our lab focuses on platelet integrin ?IIb?3, a membrane heterodimeric protein that mediates platelet aggregation by binding the adhesive plasma proteins fibrinogen and von Willebrand factor. This protein plays an essential role in initiating arrest of bleeding at sites of vascular injury, but also in thrombus formation, which may lead to heart attacks and stroke. Like other integrins, ?IIb?3 integrin can signal bidirectionally through long-range allosteric changes that are induced by the interactions of their short cytoplasmic tails with intracellular proteins such as talin (inside-out signaling), or by interactions of their extracellular domains with extracellular matrix components or soluble ligands (outside-in signaling). Interactions between the transmembrane (TM) helices of integrin ? and ? subunits have been suggested to play an important role in integrin activation and clustering. Notably, the TM helices are believed to interact in the integrin resting state, but to separate upon talin-induced integrin activation and cellular adhesion. However, the specificity and affinity between integrin TM helices have been difficult to demonstrate, and to relate to the activation process. Recent models derived from solution NMR and X-ray scattering experiments have clarified some details of the integrin TM interaction, but also raised questions about the atomic-level mechanism of the integrin signal processing. We propose to study the dynamical details of the interaction of the TM regions of integrin ?IIb and ?3 subunits in the presence or absence of talin, with the ultimate goal of understanding the molecular mechanism by which talin causes TM dimerization disruption. From a computational perspective, understanding the structural and thermodynamic features of integrin TM dimerization is not a trivial problem. To a large extent, this is due to the stochastic nature of this biological process, and the consequent need to perform multiple long-timescale molecular dynamics (MD) simulations of fully atomistic representations of protein complexes in an explicit lipid-water environment. Anton is uniquely positioned to provide detailed structural and dynamical information about the dimerization of integrin TM regions. Thus, we propose to carry out on Anton multiple microsecond-scale MD simulations, whose results are expected to provide breakthrough insights into the "inside-out" signaling of ?IIb?3 integrins.