Cholesterol Promotes Interaction of the Protein CLIC1 with Phospholipid Monolayers at the Air-Water Interface.

Khondker Hossain,Stephen Holt,Stella Valenzuela,Heba Al Khamici

doi:10.3390/membranes6010015

Khondker Hossain, Stephen Holt + Show 2 more

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https://doi.org/10.3390/membranes6010015

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Abstract

CLIC1 is a Chloride Intracellular Ion Channel protein that exists either in a soluble state in the cytoplasm or as a membrane bound protein. Members of the CLIC family are largely soluble proteins that possess the intriguing property of spontaneous insertion into phospholipid bilayers to form integral membrane ion channels. The regulatory role of cholesterol in the ion-channel activity of CLIC1 in tethered lipid bilayers was previously assessed using impedance spectroscopy. Here we extend this investigation by evaluating the influence of cholesterol on the spontaneous membrane insertion of CLIC1 into Langmuir film monolayers prepared using 1-palmitoyl-2-oleoylphosphatidylcholine, 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-ethanolamine and 1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-L-serine alone or in combination with cholesterol. The spontaneous membrane insertion of CLIC1 was shown to be dependent on the presence of cholesterol in the membrane. Furthermore, pre-incubation of CLIC1 with cholesterol prior to its addition to the Langmuir film, showed no membrane insertion even in monolayers containing cholesterol, suggesting the formation of a CLIC1-cholesterol pre-complex. Our results therefore suggest that CLIC1 membrane interaction involves CLIC1 binding to cholesterol located in the membrane for its initial docking followed by insertion. Subsequent structural rearrangements of the protein would likely also be required along with oligomerisation to form functional ion channels.

Highlights

Cholesterol is an essential constituent of the plasma membrane of most eukaryotic cells, where it is distributed non-randomly and plays a crucial role in membrane organization, dynamics, function and sorting [1,2,3]
Our results provide insight into the regulatory role that phospholipids and cholesterol play in the spontaneous membrane insertion of the protein chloride intracellular channel protein 1 (CLIC1)
There are variations in CLIC1 interaction with different phospholipid monolayers, it is clear that CLIC1 has a strong preference for associating with or intercalating into pure lipid or mixed lipid monolayers containing cholesterol

Summary

Introduction

Cholesterol is an essential constituent of the plasma membrane of most eukaryotic cells, where it is distributed non-randomly and plays a crucial role in membrane organization, dynamics, function and sorting [1,2,3]. The crucial role of cholesterol in facilitating structural rearrangements of proteins upon association with the lipid bilayer, resulting in the spontaneous conversion of the protein from water-soluble to membrane-bound form, has recently been well documented. This cholesterol-dependent membrane insertion is a widely observed phenomenon and occurs in many bacterial pore-forming toxins (PFTs), the cholesterol-dependent cytolysins (CDCs), as well as the human protein perforin, members of the complement membrane attack complex [4]. Membranes 2016, 6, 15 bilayer membranes (tBLMs) have revealed that cholesterol regulates the ion channel activity of the eukaryotic chloride intracellular channel protein 1 (CLIC1) into model membranes [8]. In addition to ion channel activity [8,12,13,14], CLIC proteins are involved in cell cycle regulation [15]; neurotransmission [16]; and recently CLICs

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