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Ceramide channels and mitochondrial outer membrane permeability

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Abstract

Among the permeability pathways in the mitochondrial outer membrane (MOM), whose elucidation was pioneered by Kathleen Kinnally, there is one formed by the lipid, ceramide. Electron microscopic visualization shows that ceramide channels are large cylindrical structures of varying pore size, with a most frequent size of 10 nm in diameter, large enough to allow all soluble proteins to translocate between the cytosol and the mitochondrial intermembrane space. Similar results were obtained with electrophysiological measurements. Studies of the dynamics of the channels are consistent with a right cylinder. Ceramide channels form at mole fractions of ceramide that are found in the MOM early in the apoptotic process, before or at the time of protein release from mitochondria. That these channels are good candidates for the protein release pathway is supported by the fact that channel formation is inhibited by anti-apoptotic proteins and favored by Bax. Bcl-xL inhibits ceramide channel formation by binding to the apolar ceramide tails using its hydrophobic grove. Bax interaction with the polar regions of ceramide results in MOM permeabilization through synergy with ceramide. Evidence that ceramide channels actually function to favor apoptosis in vivo is supported by the expression of Bcl-xL containing point mutations in cells induced to undergo apoptosis. The Bcl-xL mutants inhibit differentially Bax and ceramide channels and thus tease apart, to some extent, these two modes of MOM permeabilization. Ceramide channels have the right properties and appropriate regulation to be key players in the induction of apoptosis.

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Acknowledgments

This work was supported by funding from the National Science Foundation (MCB-1023008).

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Correspondence to Marco Colombini.

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Supplemental Figure 1

Panel A. The working model of a ceramide channel consisting of 48 columns of ceramide arranged in an anti-parallel fashion. The phospholipid bilayer surrounding the channel is not illustrated. This forms an aqueous pore that is 10 nm in diameter, the size most commonly observed experimentally. The pore is lined by the polar ends of the ceramide molecules with the oxygens in red. The crystal structure of Bcl-xL is superimposed on one portion of the channel at the bottom of the figure. It is positioned to overlap the apolar tails of a ceramide at the end of one of the columns. Panel B. A pair of ceramide columns in an anti-parallel orientation illustrates the channel structure in detail. Each column consists of 6 ceramide molecules connected by hydrogen bonding (yellow) both through the amide linkages and the twin hydroxyls. Some water molecules are also involved in the hydrogen-bonded network. (Reprinted with permission from Chang et al., 2015) (GIF 205 kb)

High Resolution Image (TIFF 3188 kb)

Supplemental Figure 2

Metabolic pathways that generate or consume ceramide. (GIF 10 kb)

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Supplemental Figure 3

Model of the Bcl-xL/ceramide interaction from in-silico docking experiments. Structure of C16-ceramide, in green, bound to Bcl-xL, shown in ribbon mode. Panel A some of the amino acid side chains: F97, F105 and F146 are shown in full space filling structure and labeled as “F″ so as to illustrate their interaction with the acyl chain of ceramide. Panel B illustrates the polar interactions between the ceramide hydroxyls (shown in space filling mode) and two side chains if Bcl-xL: R100 and Y195. R100 has 2 hydrogen bonds with the oxygen of the hydroxyl on carbon 3 of ceramide (labeled as 3-OH). The hydroxyl of Y195 (labeled as “Y″) is aligned next to the C1 hydroxyl (labeled as 1-OH) of ceramide in such a way that their dipoles are antiparallel. (Reprinted with permission from Chang et al., 2015) (GIF 132 kb)

High Resolution Image (TIFF 3858 kb)

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Colombini, M. Ceramide channels and mitochondrial outer membrane permeability. J Bioenerg Biomembr 49, 57–64 (2017). https://doi.org/10.1007/s10863-016-9646-z

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