Search

Integrin αIIbβ3 is a predominant type of integrin abundantly expressed on the surface of platelets and its activation regulates the process of thrombosis. Talin and kindlin are cytoplasmic proteins that bind to integrin and modulate its affinity for extracellular ligands. While the molecular details of talin-mediated integrin activation are known, the mechanism of kindlin involvement in this process remains elusive. Here, we demonstrate that the interplay between talin and kindlin promotes integrin activation. Our all-atomic molecular dynamics simulations on complete transmembrane and cytoplasmic domains of integrin αIIbβ3, talin1 F2/F3 subdomains, and kindlin2 FERM domain in an explicit lipid-water environment over microsecond timescale, unraveled the role of kindlin as an enhancer of the talin interaction with the membrane proximal region of β–integrin. The cooperation of kindlin with talin results in a complete disruption of salt bridges between R995 on αIIb and D723/E726 on β3. Furthermore, kindlin modifies the molecular mechanisms of inside-out activation by decreasing the crossing angle between transmembrane helices of integrin αIIb-β3, which eventually results in parallelization of integrin dimer. In addition, our control simulation featuring integrin in complex with kindlin reveals that kindlin binding is not sufficient for unclasping the inner membrane and outer membrane interactions of integrin dimer, thus ruling out the possibility of solitary action of kindlin in integrin activation.Statement of SignificanceUsing the newly solved crystal structure of kindlin, we investigated, for the first time, the molecular mechanism of kindlin-mediated integrin activation through simultaneous binding of talin and kindlin. We demonstrate in atomist details how kindlin cooperates with talin to promote the activation of integrin αIIb-β3.

Mechanical stresses directly regulate the function of several proteins of the integrin-mediated focal adhesion complex as they experience intra- and extracellular forces. Kindlin is a largely overlooked member of the focal adhesion complex whose roles in cellular mechanotransduction are only recently being identified. Recent crystallographic experiments have revealed that kindlins can form dimers that bind simultaneously to two integrins, providing a mechanistic explanation of how kindlins may promote integrin activation and clustering. In this study, using the newly identified molecular structure, we modeled the response of the kindlin2 dimer in complex with integrin β1 to mechanical cytoskeletal forces on integrins. Using molecular dynamics simulations, we show that forces on integrins are directly transmitted to the kindlin2 dimerization site, resulting in a shift in an R577-S550/E553 interaction network at this site. Under force, R577 on one protomer switches from interacting with S550 to forming new hydrogen bonds with E553 on the neighboring protomer, resulting in the strengthening of the kindlin2 dimer in complex with integrin β1. This force-induced strengthening is similar to the catch-bond mechanisms that have previously been observed in other adhesion molecules. Based on our results, we propose that the kindlin2 dimer is mechanosensitive and can strengthen integrin-mediated focal adhesions under force by shifting the interactions at its dimerization sites.

Immune response mechanisms at a cellular level are mostly triggered by a decrease in the concentration of Ca2+ within the endoplasmic reticulum (ER), followed by the opening of the calcium release-activated calcium (CRAC) channels that leads to increased intracellular Ca2+ concentrations. Recently, the stromal interaction molecule (STIM) was determined to be the ER Ca2+ sensor that activates the channel in response to the depletion of intracellular calcium content. However, the molecular details of STIM interaction with Orai that causes the channel opening are not yet known. Understanding these molecular processes would shed light on this signal transduction pathway and opens up new doors for tapping into such a mechanism with profound biological and clinical implications.CRAC channels in the plasma membrane are integral membrane proteins that play a central role in cellular signaling by generating the calcium influx. Orai protein is a pore subunit of the channel, which has 3 homologs (Orai-1, Orai-2, and Orai-3).We develop all-atomic molecular dynamics models of the STIM1/Orai-1 complex with the aim to demonstrate how the binding of STIM1 to C-terminus of Orai-1 will result in a conformational change in Orai-1 protein complex leading to the activation of CRAC channels. Different mutations of Orai-1 C-terminus are considered and the influence of factors such as ER tension on the enhancement of this conformational change is explored in accordance with experimental evidences.