A tuneable minimal cell membrane reveals that two lipid species suffice for life

Justice, Isaac; Kiesel, Petra; Safronova, Nataliya; von Appen, Alexander; Saenz, James P.

doi:10.1101/2023.10.24.563757

Cited by 3 publications

(4 citation statements)

References 85 publications

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“…Additional applications might lie in engineering minimal cell systems to function with only two classes of lipids (35); if one of those classes were a sterol-lipid, it would be interesting to test if the cell membrane phase separated. Micron-scale, reversible, liquid-liquid phase separation has previously been observed in more complex membranes of living cells (1, 36), where it has biological importance. For example, in yeast, phase separation of vacuole membranes promotes docking of lipid droplets during periods of nutrient restriction (37–39).…”

Section: Resultsmentioning

confidence: 96%

“…The problem is so severe that no vesicles formed from 100% Sterol-lipid B. diPhyPC's conical shape is reflected in its large cross-sectional area (e.g., 80.5 ± 1.5 Å 2 at 30˚C, compared with 67.4 ± 1 Å 2 for di(18:1)PC (27,28); values for diPhyPC under different conditions are compiled elsewhere (27,29)). A conical shape of diPhyPC is consistent with shallow membrane defects and low chain order in molecular dynamics simulations of diPhyPC membranes (30,31) We mitigated, but did not entirely solve, the problem of unstable membranes due to coneshaped lipids by switching to the alternative low-Tmelt lipid, di(18:1)PC. Transition temperatures of binary membranes of Sterol-lipid B and the alternative lipid appear to lie below the lowest temperatures accessible in our experiments.…”

Section: Alternative Lipids and Negative Controlsmentioning

confidence: 88%

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Sterol-lipids enable large-scale, liquid-liquid phase separation in bilayer membranes of only 2 components

Wilson,

Nguyen,

Gervay-Hague

et al. 2024

Preprint

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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 for two reasons: (1) the Gibbs Phase Rule states that only two components are necessary, and (2) 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, we discovered a minimal bilayer of only two components (PChemsPC and diPhyPC) that demixes into micron-scale, liquid phases. In this system, the sterol-lipid behaves as a 3:1 ratio of cholesterol to phospholipid. Our system gives the computation and theory community a two-component membrane that maps directly onto simplified theories and that can be used to validate simulation force fields. It suggests a new role for sterol-lipids in nature, and it gives experimental communities a membrane in which tie-lines (and, therefore, the lipid composition of each phase) are easily determined and will be consistent across multiple laboratories.Significance StatementA wide diversity of bilayer membranes, from those with hundreds of lipids (e.g., vacuoles of living yeast cells) to those with very few (e.g., artificial vesicles) phase separate into micron-scale liquid domains. The number of components required for liquid-liquid phase separation has been perplexing: only two should be necessary, but more are required except in extraordinary circumstances. What minimal set of molecular characteristics leads to liquid-liquid phase separation in bilayer membranes? This question inspired us to search for single, joined “sterol-lipid” molecules to replace both a sterol and a phospholipid in membranes undergoing liquid-liquid phase separation. By producing phase-separating membranes with only two components, we mitigate experimental challenges in determining tie-lines and in maintaining constant chemical potentials of lipids.

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Section: Resultsmentioning

confidence: 96%

Section: Alternative Lipids and Negative Controlsmentioning

confidence: 88%

Sterol-lipids enable large-scale, liquid-liquid phase separation in bilayer membranes of only 2 components

Wilson,

Nguyen,

Gervay-Hague

et al. 2024

Preprint

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“…Nevertheless, utilization of both cholesterol and SM by Mycoplasma showcases the unique composition of their membranes, which is in many way convergent with tremendously more complicated mammalian lipidomes. And while the mammalian membranes lipid composition is challenging to manipulate, mycoplasma membranes are tuneable 3 , and therefor offer a promising choice for further investigation of the role of sphingomyelin in membrane adaptation.…”

Section: Discussionmentioning

confidence: 99%

“…Consequently, cells are continuously engaged in a balancing act to ensure that the composition of their membrane lipidomes is properly tuned to optimize both bioactivity and mechanical properties, such as bending rigidity 1 and fluidity 2 . Although a tuneable membrane can be built from as few as two lipid structures 3 , cellular lipidomes are far more complex, ranging from 10s of lipids in bacteria 4 to 100s in eukaryotic organisms 5678 . Despite the fundamental role of lipids in shaping membrane function and cellular fitness, how cells employ the collective properties of lipids to build responsive interfaces is still largely undefined.…”

Section: Introductionmentioning

confidence: 99%

From hot to cold: dissecting lipidome adaptation inMycoplasma mycoidesand the Minimal Cell JCVI-Syn3B

Safronova,

Junghans,

Saenz

2023

Preprint

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Cell membranes insulate and mediate interactions between life and its environment, with lipids determining their properties and functions. However, the intricacies of how cells adjust their lipidome compositions to tune membrane properties remain relatively undefined. The complexity of most model organisms has made it challenging to characterize lipidomic adaptation. An ideal model system would be a relatively simple organism with a single membrane that can adapt to environmental changes, particularly temperature, which is known to affect membrane properties. To this end, we used quantitative shotgun lipidomics to analyze temperature adaptation inMycoplasma mycoidesand its minimal synthetic counterpart, JCVI-Syn3B. Comparing with lipidomes from eukaryotes and bacteria, we observed a universal logarithmic distribution of lipid abundances. Additionally, the extent of lipid remodeling needed for temperature adaptation appears relatively constrained, irrespective of lipidomic or organismal complexity. Through lipid features analysis, we demonstrate head group-specific acyl chain remodeling as characteristic of temperature-induced lipidome adaptation and its deficiency in Syn3B is associated with impaired homeoviscous adaptation. Temporal analysis uncovers a two-stage cold adaptation process: swift cholesterol and cardiolipin shifts followed by gradual acyl chain modifications. This work provides an in-depth analysis of lipidome adaptation in minimal cells, laying a foundation to probe the fundamental design principles of living membranes.

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Chemically defined lipid diets reveal the versatility of lipidome remodeling in genomically minimal cells

Safronova,

Junghans,

Oertel

et al. 2024

Preprint

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All cells are encapsulated in a lipid membrane that provides a responsive interface between life and its environment. Although simple membranes can be built from a single type of lipid, cellular membranes contain 10s to 100s of unique lipid species. Deciphering the significance of lipidome complexity is a central challenge in understanding the design principles of living membranes. While functions of individual lipids have been extensively studied, understanding how lipidomes collectively contribute to membrane function and cell phenotypes is experimentally challenging in most organisms. To address this challenge, we turned to the simple pathogenic organismMycoplasma mycoidesand its genomically derived Minimal Cell JCVI-syn3B, to establish a living minimal membrane model system in which lipidome complexity can be experimentally manipulated. By complexing lipids with cyclodextrins, we introduce a chemically defined approach to deliver lipid 'diets' with different chemistries to cells, resulting in cellular lipidomes with as few as seven to nearly 30 lipids species. We explored how lipidome size and composition influences cell growth, osmotic sensitivity, and membrane adaptability to changes in growth temperature. Our findings indicate that lipidome composition dictates membrane adaptation to temperature change. Moreover, we show that lipidome diversity enhances cellular robustness to hypoosmotic shock. We further show that impaired acyl chain remodeling in the minimal cell is associated with impaired membrane temperature adaptation. Finally, we demonstrate as a proof of principle, how cells with tuneable lipidomes can be used as experimental chassis for screening membrane active antimicrobial peptides. Our study introduces an experimental resource and foundation for deciphering the role of lipidome complexity in membrane function and cellular fitness.

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A tuneable minimal cell membrane reveals that two lipid species suffice for life

Cited by 3 publications

References 85 publications

Sterol-lipids enable large-scale, liquid-liquid phase separation in bilayer membranes of only 2 components

Sterol-lipids enable large-scale, liquid-liquid phase separation in bilayer membranes of only 2 components

From hot to cold: dissecting lipidome adaptation inMycoplasma mycoidesand the Minimal Cell JCVI-Syn3B

Chemically defined lipid diets reveal the versatility of lipidome remodeling in genomically minimal cells

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