Improving our picture of the plasma membrane: Rafts induce ordered domains in a simplified model cytoplasmic leaflet

Enoki, Thais A.; Feigenson, Gerald W.

doi:10.1016/j.bbamem.2022.183995

Cited by 18 publications

(34 citation statements)

References 34 publications

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“…Fetal Bovine Serum (FBS) was purchased from Atlanta Biologicals (Atlanta, GA). Alexa Fluor 488 (AF488) NHS ester (Invitrogen) was used to fluorescently label monoclonal anti-DNP (2,4-dinitrophenyl) immunoglobulin E (IgE) yielding AF488-IgE as described previously (27). The multivalent antigen (Ag), DNP-BSA, was prepared by conjugating DNP sulfonate (Sigma-Aldrich) to bovine serum albumin (BSA) (36).…”

Section: Methodsmentioning

confidence: 99%

“…Of particular interest is the series of recent experiments in lipid bilayers demonstrating that lipid compositional asymmetry, which is commonly maintained in live cells (24,25), may drive phase separation in the inner leaflet (26)(27)(28)(29). Simply, ordered/disordered phase-like separation in outer leaflet lipids can induce phase-like separation in the inner leaflet.…”

Section: Introductionmentioning

confidence: 99%

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Lipid Driven Inter-leaflet Coupling of Plasma Membrane Order Regulates FcεRI Signaling in Mast Cells

Yang

Wagenknecht-Wiesner

Yin

et al. 2022

Preprint

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Engagement of high affinity immunoglobulin E (IgE) receptor FcεRI with extracellular, multivalent antigen (Ag) stabilizes co-existing ordered and disordered phases in the inner leaflet of the plasma membrane. This optimally controls biochemical interactions between signaling components required for transmembrane (TM) signaling in mast cells. The biophysical organization of the resting inner leaflet is poised to respond appropriately to this extracellular stimulation. The resting inner leaflet is generally less ordered than the outer leaflet, with a lipid composition that does not spontaneously phase separate in model membranes. We proposed that coupling between the two leaflets mediates separation into different phase-like domains in the inner leaflet. To test this hypothesis in live cells, we first established a straightforward approach to evaluate changes in membrane order due to inter-leaflet coupling by measuring inner leaflet diffusion of phase-specific lipid probes with Imaging Fluorescence Correlation Spectroscopy (ImFCS) before and after methyl-α-cyclodextrin (mαCD)-catalyzed exchange of outer leaflet lipids (LEX) with exogenous order- or disorder-promoting phospholipids. We examined the functional impact of LEX by monitoring two Ag-stimulated cellular responses, namely early-stage recruitment of Syk kinase to the inner leaflet and late-stage exocytosis of secretory granules (degranulation). Based on changes in probe diffusion, we observed global increase or decrease of inner leaflet order when outer leaflet is exchanged with order or disorder promoting lipids, respectively, in unstimulated cells. Furthermore, the degree of stimulated Syk recruitment and degranulation correlates with the inner leaflet order of the resting cells, which was varied using LEX. Overall, combined LEX and ImFCS platform provides strong evidence of lipid-based control of stimulated TM signaling in live mast cells. In addition, our functional results imply that resting-state lipid composition and ordering of the outer leaflet sets the ordering of the inner leaflet, likely via interleaflet coupling, and correspondingly modulates TM signaling initiated by antigen-activated IgE-FcεRI.

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Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Lipid Driven Inter-leaflet Coupling of Plasma Membrane Order Regulates FcεRI Signaling in Mast Cells

Yang

Wagenknecht-Wiesner

Yin

et al. 2022

Preprint

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“…Users aiming at aGUVs who did not exit to the emulsion-based methods (C1, C2) before may go for hemifusion-based exchange (D1) with an excess area of solid supported membranes (see also [ 45 , 46 , 47 ]). This approach works without a lipid carrier and the elimination of the donor reservoir after exchange is straightforward.…”

Section: Navigating the Preparation Of Asymmetric Model Membranesmentioning

confidence: 99%

“…Preparation of aGUVs and data collection needs less than five hours. The resulting vesicles show high asymmetries approaching 100%, given the large excess of the SLB donor area over the GUV area [ 45 , 46 , 47 ]. However, during the hemifusion process or when aGUVs are sheared off the SLB, transient pores are formed so that lipid flip-flop may occur.…”

Section: Navigating the Preparation Of Asymmetric Model Membranesmentioning

confidence: 99%

A Guide to Your Desired Lipid-Asymmetric Vesicles

Krompers

Heerklotz

2023

Membranes

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Liposomes are prevalent model systems for studies on biological membranes. Recently, increasing attention has been paid to models also representing the lipid asymmetry of biological membranes. Here, we review in-vitro methods that have been established to prepare free-floating vesicles containing different compositions of the classic two-chain glycero- or sphingolipids in their outer and inner leaflet. In total, 72 reports are listed and assigned to four general strategies that are (A) enzymatic conversion of outer leaflet lipids, (B) re-sorting of lipids between leaflets, (C) assembly from different monolayers and (D) exchange of outer leaflet lipids. To guide the reader through this broad field of available techniques, we attempt to draw a road map that leads to the lipid-asymmetric vesicles that suit a given purpose. Of each method, we discuss advantages and limitations. In addition, various verification strategies of asymmetry as well as the role of cholesterol are briefly discussed. The ability to specifically induce lipid asymmetry in model membranes offers insights into the biological functions of asymmetry and may also benefit the technical applications of liposomes.

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“…Transleaflet coupling has been reported to induce domains in non-domain forming lipid mixtures (9, 10), affect lateral lipid diffusion (11), or cross-link the melting transitions of individual leaflets (7). Recently, we reported that the extent of hydrocarbon interdigitation allows to tweak the lateral packing of lipids in the apposing leaflet through competing cohesive and entropic interactions of the acyl chains (12).…”

Section: Introductionmentioning

confidence: 99%

Distributing Aminophospholipids Asymmetrically Across Leaflets Causes Anomalous Membrane Stiffening

Frewein

Piller

Semeraro

et al. 2022

Preprint

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We studied the mechanical leaflet coupling of prototypic mammalian plasma membranes using neutron spin-echo spectroscopy. In particular, we examined a series of asymmetric phospholipid vesicles with phosphatidylcholine and sphingomyelin enriched in the outer leaflet and inner leaflets composed of phosphatidylethanolamine / phosphatidylserine mixtures. The bending rigidities of most asymmetric membranes were anomalously high, exceeding even those of symmetric membranes formed from their cognate leaflets. Only asymmetric vesicles with outer leaflets enriched in sphingolipid displayed bending rigidities in conformity with these symmetric controls. We performed complementary small-angle neutron and X-ray experiments on the same vesicles to examine possible links to structural coupling mechanisms, which would show up in corresponding changes in membrane thickness. Additionally, we estimated differential stress between leaflets originating either from a mismatch of their lateral areas or spontaneous curvatures. However, no correlation with asymmetry-induced membrane stiffening was observed. To reconcile our findings, we speculate that an asymmetric distribution charged or H-bond forming lipids may induce an intraleaflet coupling which increases the weight of hard undulatory modes of membrane fluctuations and hence the overall membrane stiffness.

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Improving our picture of the plasma membrane: Rafts induce ordered domains in a simplified model cytoplasmic leaflet

Cited by 18 publications

References 34 publications

Lipid Driven Inter-leaflet Coupling of Plasma Membrane Order Regulates FcεRI Signaling in Mast Cells

Lipid Driven Inter-leaflet Coupling of Plasma Membrane Order Regulates FcεRI Signaling in Mast Cells

A Guide to Your Desired Lipid-Asymmetric Vesicles

Distributing Aminophospholipids Asymmetrically Across Leaflets Causes Anomalous Membrane Stiffening

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