STIM1 activates CRAC channels through rotation of the pore helix to open a hydrophobic gate

Megumi Yamashita,Christopher E. Ing,Priscilla S.-W. Yeung,Murali Prakriya,Beth A. McNally,Régis Pomès

doi:10.1038/ncomms14512

Abstract

Store-operated Ca2+ release-activated Ca2+ (CRAC) channels constitute a major pathway for Ca2+ influx and mediate many essential signalling functions in animal cells, yet how they open remains elusive. Here, we investigate the gating mechanism of the human CRAC channel Orai1 by its activator, stromal interacting molecule 1 (STIM1). We find that two rings of pore-lining residues, V102 and F99, work together to form a hydrophobic gate. Mutations of these residues to polar amino acids produce channels with leaky gates that conduct ions in the resting state. STIM1-mediated channel activation occurs through rotation of the pore helix, which displaces the F99 residues away from the pore axis to increase pore hydration, allowing ions to flow through the V102-F99 hydrophobic band. Pore helix rotation by STIM1 also explains the dynamic coupling between CRAC channel gating and ion selectivity. This hydrophobic gating mechanism has implications for CRAC channel function, pharmacology and disease-causing mutations.

Highlights

Store-operated Ca2 þ release-activated Ca2 þ (CRAC) channels constitute a major pathway for Ca2 þ influx and mediate many essential signalling functions in animal cells, yet how they open remains elusive
Their clinical relevance is underscored by loss- and gain-of-function mutations that lead to devastating immunodeficiencies, muscle weakness and bleeding disorders[4,5], and a growing list of diseases from asthma to pain for which CRAC channels are being explored for the development of new therapeutics[6,7]
This conclusion is supported by molecular dynamics simulations (MD) simulations, which showed that the V102A mutation increases water and monovalent ion occupancy in the pore[23], and lanthanide resonance energy transfer (LRET) measurements that imply that the V102A pore is similar in conformation to closed channels[19]

Summary

Introduction

Store-operated Ca2 þ release-activated Ca2 þ (CRAC) channels constitute a major pathway for Ca2 þ influx and mediate many essential signalling functions in animal cells, yet how they open remains elusive. The Ca2 þ selectivity filter resides towards the extracellular end of the pore and is formed by a ring of six glutamate residues (E106 in hOrai1) that bind Ca2 þ ions[11,12,13,14,15] This overall structure, of a narrow central pore flanked by TM1 helices, agrees well with previous electrophysiological and biochemical studies that have probed the functional pore architecture of Orai[1] channels[16,17]. Based on the constitutive ion permeation evoked by mutations of V102 (V102C/A/S/T)[21,22], this residue was proposed to function as a hydrophobic gate by presenting a free energy barrier for water and cations[21,23] Consistent with this overall concept, lanthanide resonance energy transfer (LRET) measurements implicate V102 in STIM1-mediated gating[19] and molecular simulations show that pore hydration is markedly increased in the constitutively permeant V102A mutant[23]. STIM1 binding leads to a modest rotation of the pore helix, which displaces F99 residues away from the central pore axis to disrupt the V102-F99 hydrophobic band, thereby increasing pore hydration and permitting ion conduction

Methods

Results

Conclusion