An official website of the United States government
Micron-scale, liquid-liquid phase separation occurs in membranes of living cells, with physiological consequences. To discover which lipids might support phase separation in cell membranes and how lipids might partition between phases, miscibility phase diagrams have been mapped for model membranes. Typically, model membranes are composed of ternary mixtures of a lipid with a high melting temperature, a lipid with a low melting temperature, and cholesterol. Phospholipids in ternary mixtures are chosen primarily to favor stable membranes (phosphatidylcholines and sphingomyelins) or add charge (phosphatidylglycerols and phosphatidylserines). A major class of phospholipids missing from experimental ternary diagrams has been the phosphatidylethanolamines (PEs). PE-lipids constitute up to 20 mol% of common biological membranes, where they influence protein function and facilitate membrane fusion. These biological effects are often attributed to PE’s smaller headgroup, which leads to higher monolayer spontaneous curvatures and higher melting temperatures. Taken alone, the higher melting points of saturated PE-lipids imply that liquid-liquid phase separation should persist to higher temperatures in membranes containing PE-lipids. Here, we tested that hypothesis by substituting a saturated PE-lipid (DPPE) for its corresponding PC-lipid (DPPC) in two well-studied ternary membranes (DOPC/DPPC/cholesterol and DiphyPC/DPPC/cholesterol). We used fluorescence microscopy to map full ternary phase diagrams for giant vesicles over a range of temperatures. Surprisingly, we found no micron-scale, liquid-liquid phase separation in vesicles of the first mixture (DOPC/DPPE/cholesterol), and only a small region of liquid-liquid phase separation in the second mixture (DiphyPC/DPPE/cholesterol). Instead, coexisting solid and liquid phases were widespread, with the solid phase enriched in DPPE. An unusual feature of these ternary membranes is that solid and liquid-ordered phases can be distinguished by fluorescence microscopy, so tie-line directions can be estimated throughout the phase diagram, and transition temperatures to the 3-phase region (containing a liquid-disordered phase, a liquid-ordered phase, and a solid phase) can be accurately measured.
more »
« less
Sterol–lipids enable large-scale, liquid–liquid phase separation in bilayer membranes of only two components
Despite longstanding excitement and progress toward understanding liquid–liquid phase separation in natural and artificial membranes, fundamental questions have persisted about which molecules are required for this phenomenon. Except in extraordinary circumstances, the smallest number of components that has produced large-scale, liquid–liquid phase separation in bilayers has stubbornly remained at three: a sterol, a phospholipid with ordered chains, and a phospholipid with disordered chains. This requirement of three components is puzzling because only two components are required for liquid–liquid phase separation in lipid monolayers, which resemble half of a bilayer. Inspired by reports that sterols interact closely with lipids with ordered chains, we tested whether phase separation would occur in bilayers in which a sterol and lipid were replaced by a single, joined sterol–lipid. By evaluating a panel of sterol–lipids, some of which are present in bacteria, we found a minimal bilayer of only two components (PChemsPC and diPhyPC) that robustly demixes into micron-scale, liquid phases. It suggests an additional role for sterol–lipids in nature, and it reveals a membrane in which tie-lines (and, therefore, the lipid composition of each phase) are straightforward to determine and will be consistent across multiple laboratories.
more »
« less
- PAR ID:
- 10584442
- Publisher / Repository:
- National Academy of Sciences
- Date Published:
- Journal Name:
- Proceedings of the National Academy of Sciences
- Volume:
- 121
- Issue:
- 38
- ISSN:
- 0027-8424
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
More Like this
-
-
We show, via molecular simulations, that not only does cholesterol induce a lipid order, but the lipid order also enhances cholesterol localization within the lipid leaflets. Therefore, there is a strong interdependence between these two phenomena. In the ordered phase, cholesterol molecules are predominantly present in the bilayer leaflets and orient themselves parallel to the bilayer normal. In the disordered phase, cholesterol molecules are mainly present near the center of the bilayer at the midplane region and are oriented orthogonal to the bilayer normal. At the melting temperature of the lipid bilayers, cholesterol concentration in the leaflets and the bilayer midplane is equal. This result suggests that the localization of cholesterol in the lipid bilayers is mainly dictated by the degree of ordering of the lipid bilayer. We validate our findings on 18 different lipid bilayer systems, obtained from three different phospholipid bilayers with varying concentrations of cholesterol. To cover a large temperature range in simulations, we employ the Dry Martini force field. We demonstrate that the Dry and the Wet Martini (with polarizable water) force fields produce comparable results.more » « less
-
Many intrinsically disordered peptides have been shown to undergo liquid–liquid phase separation and form complex coacervates, which play various regulatory roles in the cell. Recent experimental studies found that such phase separation processes may also occur at the lipid membrane surface and help organize biomolecules during signaling events; in some cases, phase separation of proteins at the membrane surface was also observed to lead to significant remodeling of the membrane morphology. The molecular mechanisms that govern the interactions between complex coacervates and lipid membranes and the impacts of such interactions on their structure and morphology, however, remain unclear. Here we study the coacervation of poly-glutamate (E 30 ) and poly-lysine (K 30 ) in the presence of lipid bilayers of different compositions. We carry out explicit-solvent coarse-grained molecular dynamics simulations by using the MARTINI (v3.0) force-field. We find that more than 20% anionic lipids are required for the coacervate to form stable contact with the bilayer. Upon wetting, the coacervate induces negative curvature to the bilayer and facilitates local lipid demixing, without any peptide insertion. The magnitude of negative curvature, extent of lipid demixing, and asphericity of the coacervate increase with the concentration of anionic lipids. Overall, we observe a decrease in the number of contacts among the polyelectrolytes as the droplet spreads over the bilayer. Therefore, unlike previous suggestions, interactions among polyelectrolytes do not constitute a driving force for the membrane bending upon wetting by the coacervate. Rather, analysis of interaction energy components suggests that bending of the membrane is favored by enhanced interactions between polyelectrolytes with lipids as well as with counterions. Kinetic studies reveal that, at the studied polyelectrolyte concentrations, the coacervate formation precedes bilayer wetting.more » « less
-
Biological supramolecular assemblies, such as phospholipid bilayer membranes, have been used to demonstrate signal processing via short-term synaptic plasticity (STP) in the form of paired pulse facilitation and depression, emulating the brain’s efficiency and flexible cognitive capabilities. However, STP memory in lipid bilayers is volatile and cannot be stored or accessed over relevant periods of time, a key requirement for learning. Using droplet interface bilayers (DIBs) composed of lipids, water and hexadecane, and an electrical stimulation training protocol featuring repetitive sinusoidal voltage cycling, we show that DIBs displaying memcapacitive properties can also exhibit persistent synaptic plasticity in the form of long-term potentiation (LTP) associated with capacitive energy storage in the phospholipid bilayer. The time scales for the physical changes associated with the LTP range between minutes and hours, and are substantially longer than previous STP studies, where stored energy dissipated after only a few seconds. STP behavior is the result of reversible changes in bilayer area and thickness. On the other hand, LTP is the result of additional molecular and structural changes to the zwitterionic lipid headgroups and the dielectric properties of the lipid bilayer that result from the buildup of an increasingly asymmetric charge distribution at the bilayer interfaces.more » « less
-
null (Ed.)The plasma membrane of eukaryotic cells is known to be compositionally asymmetric. Certain phospholipids, such as sphingomyelin and phosphatidylcholine species, are predominantly localized in the outer leaflet, while phosphatidylethanolamine and phosphatidylserine species primarily reside in the inner leaflet. While phospholipid asymmetry between the membrane leaflets is well established, there is no consensus about cholesterol distribution between the two leaflets. We have performed a systematic study, via molecular simulations, of how the spatial distribution of cholesterol molecules in different “asymmetric” lipid bilayers are affected by the lipids’ backbone, head-type, unsaturation, and chain-length by considering an asymmetric bilayer mimicking the plasma membrane lipids of red blood cells, as well as seventeen other asymmetric bilayers comprising of different lipid types. Our results reveal that the distribution of cholesterol in the leaflets is solely a function of the extent of ordering of the lipids within the leaflets. The ratio of the amount of cholesterol matches the ratio of lipid order in the two leaflets, thus providing a quantitative relationship between the two. These results are understood by the observation that asymmetric bilayers with equimolar amount of lipids in the two leaflets develop tensile and compressive stresses due to differences in the extent of lipid order. These stresses are alleviated by the transfer of cholesterol from the leaflet in compressive stress to the one in tensile stress. These findings are important in understanding the biology of the cell membrane, especially with regard to the composition of the membrane leaflets.more » « less
- Free Publicly Accessible Full Text
-
Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1073/pnas.2401241121
