In Vitro Membrane Remodeling by ESCRT is Regulated by Negative Feedback from Membrane Tension

Booth, Andrew; Marklew, Christopher J.; Ciani, Barbara; Beales, Paul A.

doi:10.1016/j.isci.2019.04.021

Cited by 34 publications

(48 citation statements)

References 49 publications

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“…These results demonstrate that changes in membrane permeability, fluidity and mechanical properties are not caused by any release of Ag + ions from the AgNPs, hence we confidently conclude that all the membrane perturbations observed in this investigation are produced directly by the weakly aggregating AgNPs. without the need of cellular endocytic mechanisms or external sources of energy 25,52,60,[74][75][76][77] . Generally the interaction of single proteins or particles is not strong enough to induce these large deformations of the membrane and thus many molecules or particles must cooperate to bend the membrane 54,75 .…”

Section: Flicker Spectroscopymentioning

confidence: 99%

Mechanomodulation of Lipid Membranes by Weakly Aggregating Silver Nanoparticles

et al. 2019

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Silver nanoparticles (AgNPs) have wide-ranging applications, including as additives in consumer products and in medical diagnostics and therapy. Therefore, understanding how AgNPs interact with biological systems is important for ascertaining any potential health risks due to the likelihood of high levels of human exposure. Besides any severe, acute effects, it is desirable to understand more subtle interactions that could lead to milder, chronic health impacts. Nanoparticles are small enough to be able to enter biological cells and interfere with their internal biochemistry. The initial contact between the nanoparticle and cell is at the plasma membrane. To gain fundamental mechanistic insight into AgNP–membrane interactions, we investigate these phenomena in minimal model systems using a wide range of biophysical approaches applied to lipid vesicles. We find a strong dependence on the medium composition, where colloidally stable AgNPs in a glucose buffer have a negligible effect on the membrane. However, at physiological salt concentrations, the AgNPs start to weakly aggregate and sporadic but significant membrane perturbation events are observed. Under these latter conditions, transient poration and structural remodeling of some vesicle membranes are observed. We observe that the fluidity of giant vesicle membranes universally decreases by an average of 16% across all vesicles. However, we observe a small population of vesicles that display a significant change in their mechanical properties with lower bending rigidity and higher membrane tension. Therefore, we argue that the isolated occurrences of membrane perturbation by AgNPs are due to low-probability mechanomodulation by AgNP aggregation at the membrane.

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Section: Flicker Spectroscopymentioning

confidence: 99%

Mechanomodulation of Lipid Membranes by Weakly Aggregating Silver Nanoparticles

et al. 2019

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“…2,31 ESCRT activity is negatively regulated by membrane tension, in a manner that is likely analogous to suppression of activity by higher bending rigidity. 15 We conclude that ESCRT activity must be higher at PS-containing Ld domains where the membrane is more susceptible to deformation.…”

Section: Ps In the Ld Phase Enhances Efficiency Of Ilv Formationmentioning

confidence: 75%

“…We and others 14,15,29 have successfully used GUVs composed of homogeneous PC:Cholesterol:PS mixtures to reconstitute the membrane remodelling activity of the ESCRT-II/ESCRT-III complex. Based on phase diagrams available in the literature for DOPC:DPPC:chol mixtures, 30 we selected a baseline membrane composition of 35:35:30 as it is fairly central within the liquid ordered (Lo) -liquid disordered (Ld) coexistence region of the phase diagram.…”

Section: Strategymentioning

confidence: 99%

“…The presence of fluorescent dye in the lumens of free-floating ILVs, after incubation with ESCRT proteins, being a strong indication that those ILVs were formed as a result of ESCRT activity. 15 The number of dye-containing ILVs was counted and then expressed as a number of ILVs per GUV volume equivalent, by taking the luminal volume observed during large area tile-scan imaging of GUV samples (cross sectional area x section thickness), divided by the volume of a typical 20 m diameter GUV Figure 3. Automation of ILV counting was made possible by using the calcein-Co 2+ fluorescent dye-quencher pair in this assay, as fluorescence in the bulk medium could be quenched after the ESCRT incubation period by addition of the membrane-impermeable quencher (CoCl2).…”

Section: Ps In the Ld Phase Enhances Efficiency Of Ilv Formationmentioning

confidence: 99%

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The influence of phosphatidylserine localisation and lipid phase on membrane remodelling by the ESCRT-II/ESCRT-III complex

Booth

Marklew

Ciani

et al. 2020

Preprint

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The endosomal sorting complex required for transport (ESCRT) organises in supramolecular structures on the surface of lipid bilayers to drive membrane invagination and scission of intraluminal vesicles (ILVs), a process also controlled by membrane mechanics. However, ESCRT association with the membrane is also mediated by electrostatic interactions with anionic phospholipids. Phospholipid distribution within natural biomembranes is inhomogeneous due to, for example, the formation of lipid rafts and curvature-driven lipid sorting. Here, we have used phase-separated giant unilamellar vesicles (GUVs) to investigate the link between phosphatidylserine (PS)-rich lipid domains and ESCRT activity. We employ GUVs composed of phase separating lipid mixtures, where unsaturated DOPS and saturated DPPS lipids are incorporated individually or simultaneously to enhance PS localisation in liquid disordered (Ld) and/or liquid ordered (Lo) domains, respectively. PS partitioning between the coexisting phases is confirmed by a fluorescent Annexin V probe. Ultimately, we find that ILV generation promoted by ESCRTs is significantly enhanced when PS lipids localise within Ld domains. However, the ILVs that form are rich in Lo lipids. We interpret this surprising observation as preferential recruitment of the Lo phase beneath the ESCRT complex due to its increased rigidity, where the Ld phase is favoured in the neck of the resultant buds to facilitate the high membrane curvature in these regions of the membrane during the ILV formation process. Ld domains offer lower resistance to membrane bending, demonstrating a mechanism by which the composition and mechanics of membranes can be coupled to regulate the location and efficiency of ESCRT activity.

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“…4 D); this may have the added benefit of promoting the ultimate annular fusion event necessary for nuclear envelope sealing. Similarly, membrane remodeling by ESCRTs may itself increase local membrane tension (Booth et al, 2019) and trigger a local lipid synthesis response, which may contribute membrane to help seal nuclear ruptures (Kinugasa et al, 2019) and the nuclear envelope at the end of mitosis (Lee et al, 2020; Penfield et al, 2020).…”

Section: Resultsmentioning

confidence: 99%

Direct PA-binding by Chm7 is required for nuclear envelope surveillance at herniations

Thaller

Tong

Marklew

et al. 2020

Preprint

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19Mechanisms that control nuclear membrane remodeling are essential to maintain the integrity of the 20 nucleus but remain to be fully defined. Here, we identify a phosphatidic acid (PA)-binding activity in the 21 nuclear envelope-specific ESCRT, Chm7, in budding yeast. PA-binding is mediated through a conserved 22 hydrophobic stretch of amino acids, which confers specific binding to the inner nuclear membrane (INM). 23This INM-binding is independent but nonetheless required for interaction with the LAP2-emerin-MAN1 24 (LEM) domain protein, Heh1 (LEM2). Consistent with the functional importance of PA-binding, mutation of 25 this region inhibits recruitment of Chm7 to the INM and abolishes Chm7 function in the context of nuclear 26 envelope herniations or "blebs" that form during defective nuclear pore complex (NPC) biogenesis. In 27 fact, we show that PA accumulates at nuclear envelope herniations. We suggest that local control of PA 28 metabolism is important for ensuring productive nuclear envelope remodeling and that its dysregulation 29 may contribute to pathologies associated with defective NPC assembly. 30 31 109 nucleus or at the nuclear envelope. Thus, NES1 did not appreciably alter Chm7 distribution, although we 110 note that fewer cytosolic structures were visible compared to wildtype Chm7-GFP.

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In Vitro Membrane Remodeling by ESCRT is Regulated by Negative Feedback from Membrane Tension

Cited by 34 publications

References 49 publications

Mechanomodulation of Lipid Membranes by Weakly Aggregating Silver Nanoparticles

Mechanomodulation of Lipid Membranes by Weakly Aggregating Silver Nanoparticles

The influence of phosphatidylserine localisation and lipid phase on membrane remodelling by the ESCRT-II/ESCRT-III complex

Direct PA-binding by Chm7 is required for nuclear envelope surveillance at herniations

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