Lipids in Mitochondrial Macroautophagy: Phase Behavior of Bilayers Containing Cardiolipin and Ceramide

Abstract

Cardiolipin (CL) is a key lipid for damaged mitochondrial recognition by the LC3/GABARAP human autophagy proteins. The role of ceramide (Cer) in this process is unclear, but CL and Cer have been proposed to coexist in mitochondria under certain conditions. Varela et al. showed that in model membranes composed of egg sphingomyelin (eSM), dioleoyl phosphatidylethanolamine (DOPE), and CL, the addition of Cer enhanced the binding of LC3/GABARAP proteins to bilayers. Cer gave rise to lateral phase separation of Cer-rich rigid domains but protein binding took place mainly in the fluid continuous phase. In the present study, a biophysical analysis of bilayers composed of eSM, DOPE, CL, and/or Cer was attempted to understand the relevance of this lipid coexistence. Bilayers were studied by differential scanning calorimetry, confocal fluorescence microscopy, and atomic force microscopy. Upon the addition of CL and Cer, one continuous phase and two segregated ones were formed. In bilayers with egg phosphatidylcholine instead of eSM, in which the binding of LC3/GABARAP proteins hardly increased with Cer in the former study, a single segregated phase was formed. Assuming that phase separation at the nanoscale is ruled by the same principles acting at the micrometer scale, it is proposed that Cer-enriched rigid nanodomains, stabilized by eSM:Cer interactions formed within the DOPE- and CL-enriched fluid phase, result in structural defects at the rigid/fluid nanointerfaces, thus hypothetically facilitatingLC3/GABARAP protein interaction.

Effects of a N-Maleimide-derivatized Phosphatidylethanolamine on the Architecture and Properties of Lipid Bilayers

“…Finally, GUVs were transferred to the visualizing chamber, where 150 mM NaCl, 50 mM Tris-HCl and pH 7.5 buffer was added. Images were acquired on a Leica SP5 confocal microscope with a 63× Water Planar Apochromat 1.2 NA objective at 23 • C. The excitation and emission wavelengths used for Rho-PE were 514 nm and 580-600 nm, respectively [27].…”

Biophysics in Membrane of Cells

“…Delving into the microcosm of lipid rafts, the studies “Lipids in Mitochondrial Macroautophagy: Phase Behavior of Bilayers Containing Cardiolipin and Ceramide” [ 8 ] and “Effect of Lipid Raft Disruptors on Cell Membrane Fluidity Studied by Fluorescence Spectroscopy” [ 9 ] examined the roles of specific lipids in the context of membrane function. From the molecular interactions between cardiolipin, ceramide, and autophagy proteins to the effects of raft disruptors on membrane fluidity, these studies illuminated the delicate balance within lipid rafts.…”