title: Investigating Sterol and Redox Regulation of the Ion Channel Activity of CLIC1 Using Tethered Bilayer Membranes.
abstract: The Chloride Intracellular Ion Channel (CLIC) family consists of six conserved proteins in humans. These are a group of enigmatic proteins, which adopt both a soluble and membrane bound form. CLIC1 was found to be a metamorphic protein, where under specific environmental triggers it adopts more than one stable reversible soluble structural conformation. CLIC1 was found to spontaneously insert into cell membranes and form chloride ion channels. However, factors that control the structural transition of CLIC1 from being an aqueous soluble protein into a membrane bound protein have yet to be adequately described. Using tethered bilayer lipid membranes and electrical impedance spectroscopy system, herein we demonstrate that CLIC1 ion channel activity is dependent on the type and concentration of sterols in bilayer membranes. These findings suggest that membrane sterols play an essential role in CLIC1's acrobatic switching from a globular soluble form to an integral membrane form, promoting greater ion channel conductance in membranes. What remains unclear is the precise nature of this regulation involving membrane sterols and ultimately determining CLIC1's membrane structure and function as an ion channel. Furthermore, our impedance spectroscopy results obtained using CLIC1 mutants, suggest that the residue Cys24 is not essential for CLIC1's ion channel function. However Cys24 does appear important for optimal ion channel activity. We also observe differences in conductance between CLIC1 reduced and oxidized forms when added to our tethered membranes. Therefore, we conclude that both membrane sterols and redox play a role in the ion channel activity of CLIC1.
authors.affiliation: School of Life Sciences, University of Technology Sydney, Sydney 2007, Australia. Heba.alkhamici@uts.edu.au.
authors.forename: Heba
authors.initials: H
authors.lastname: Al Khamici
authors.affiliation: School of Life Sciences, University of Technology Sydney, Sydney 2007, Australia. khondker.r.hossain@student.uts.edu.au.|Australian Centre for Neutron Scattering, Australian Nuclear Science and Technology Organisation (ANSTO), NSW 2234, Australia. khondker.r.hossain@student.uts.edu.au.
authors.forename: Khondher R
authors.initials: KR
authors.lastname: Hossain
authors.affiliation: Surgical Diagnostics Pty Ltd., Sydney 2069, Australia. BruceC.sdx@gmail.com.
authors.forename: Bruce A
authors.initials: BA
authors.lastname: Cornell
authors.affiliation: School of Life Sciences, University of Technology Sydney, Sydney 2007, Australia. stella.valenzuela@uts.edu.au.
authors.forename: Stella M
authors.initials: SM
authors.lastname: Valenzuela
keywords: CLIC; chloride intracellular ion channel proteins; cholesterol; ergosterol; tethered lipid membranes