Abstract
CRAC channel is ubiquitous and its importance in the regulation of the immune system is testified by the severe immunodeficiencies caused by its mutations. In this work we took advantage of the availability of open and closed structures of this channel to run for the first time simulations of the whole gating process reaching the relevant time-scale with an enhanced sampling technique, Targeted Molecular Dynamics. Our simulations highlighted a complex allosteric propagation of the conformational change from peripheral helices, where the activator STIM1 binds, to the central pore helices. In agreement with mutagenesis data, our simulations revealed the key role of residue H206 whose displacement creates an empty space behind the hydrophobic region of the pore, thus releasing a steric brake and allowing the opening of the channel. Conversely, the process of pore closing culminates with the formation of a bubble that occludes the pore even in the absence of steric block. This mechanism, known as “hydrophobic gating”, has been observed in an increasing number of biological ion channels and also in artificial nanopores. Our study therefore shows promise not only to better understand the molecular origin of diseases caused by disrupted calcium signaling, but also to clarify the mode of action of hydrophobically gated ion channels, possibly even suggesting strategies for the biomimetic design of synthetic nanopores.
Highlights
The ubiquitous Calcium Release Activated Calcium channel (CRAC) mediates Ca2+-influx through the plasma membrane of non-excitable cells in metazoans activating communication cascades that elicit a wide range of functions (Prakriya and Lewis, 2015) like gene expression, cell proliferation, secretion of inflammatory mediators, and cell migration
Our equilibrium simulations show a peak of the hydrophobicity profile in correspondence of the hydrophobic region, where the Potential of Mean Force of water has its maximum
This suggests a scenario of functional occlusion of the pore even in the absence of steric block, which is the hallmark of hydrophobic gating (Aryal et al, 2015)
Summary
The ubiquitous Calcium Release Activated Calcium channel (CRAC) mediates Ca2+-influx through the plasma membrane of non-excitable cells in metazoans activating communication cascades that elicit a wide range of functions (Prakriya and Lewis, 2015) like gene expression, cell proliferation, secretion of inflammatory mediators, and cell migration. STIM1 migrates to the junctions between ER and plasma membranes where it oligomerizes and undergoes a conformational transition that exposes the CRAC activation domain (CAD) (Park et al, 2009). The interaction of STIM1 CAD domain with CRAC outermost helices (TM4) induces the opening of the CRAC channel that results in an influx of calcium that refills the stores of the ER and activates a number of signalling pathways including those necessary for the activation of immune response genes in T cells (Feske et al, 2005)
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