Biochemical and biophysical features of both oligomer/fibril and cell membrane in amyloid cytotoxicity

Stefani, Massimo

doi:10.1111/j.1742-4658.2010.07889.x

Cited by 173 publications

(165 citation statements)

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“…There is considerable evidence that amyloid assemblies interact with membranes, supporting the notion that these interactions may play a role in amyloid-associated cellular dysfunction (11,13,34). However, the physical basis for these interactions and the mechanisms of amyloid-induced dysfunction remain unresolved.…”

Section: Discussionmentioning

confidence: 92%

“…In some systems, there is evidence suggesting that prefibrillar oligomers, rather than the fully formed fibrils, are the source of toxicity (8,9). In these cases, cytotoxicity is thought to result from the formation of specific membrane pores (10,11) although alternative models including membrane destabilization or membrane thinning have also been proposed (12)(13)(14)(15). In other cases, toxicity may reside with the amyloid fibrils themselves.…”

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confidence: 99%

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Direct three-dimensional visualization of membrane disruption by amyloid fibrils

Milanesi

Sheynis

Xue

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

172

177

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Protein misfolding and aggregation cause serious degenerative conditions such as Alzheimer's, Parkinson, and prion diseases. Damage to membranes is thought to be one of the mechanisms underlying cellular toxicity of a range of amyloid assemblies. Previous studies have indicated that amyloid fibrils can cause membrane leakage and elicit cellular damage, and these effects are enhanced by fragmentation of the fibrils. Here we report direct 3D visualization of membrane damage by specific interactions of a lipid bilayer with amyloid-like fibrils formed in vitro from β 2 -microglobulin (β 2 m). Using cryoelectron tomography, we demonstrate that fragmented β 2 m amyloid fibrils interact strongly with liposomes and cause distortions to the membranes. The normally spherical liposomes form pointed teardrop-like shapes with the fibril ends seen in proximity to the pointed regions on the membranes. Moreover, the tomograms indicated that the fibrils extract lipid from the membranes at these points of distortion by removal or blebbing of the outer membrane leaflet. Tiny (15-25 nm) vesicles, presumably formed from the extracted lipids, were observed to be decorating the fibrils. The findings highlight a potential role of fibrils, and particularly fibril ends, in amyloid pathology, and report a previously undescribed class of lipid-protein interactions in membrane remodelling.T he failure of molecular chaperones to prevent the accumulation of misfolded proteins results in protein aggregation and amyloid formation, processes associated with severe human degenerative diseases (1, 2). Despite the attention focused on these problems during the century since these disorders were first identified (3-5) and advances in understanding the structure of the cross-β conformation of amyloid fibrils in atomic detail (6, 7), the basic pathological mechanisms of amyloidosis remain poorly understood and therapeutic intervention is lacking. The identity of the toxic species and the mechanisms of cytotoxicity remain major unsolved problems. In some systems, there is evidence suggesting that prefibrillar oligomers, rather than the fully formed fibrils, are the source of toxicity (8, 9). In these cases, cytotoxicity is thought to result from the formation of specific membrane pores (10, 11) although alternative models including membrane destabilization or membrane thinning have also been proposed (12-15). In other cases, toxicity may reside with the amyloid fibrils themselves. Evidence that toxicity correlates with fibrillar assemblies has been reported for yeast and mammalian prion proteins (16, 17), human lysozyme (18), Huntingtin exon 1, α-synuclein (19), and Amyloid-β (Aβ) (20, 21). Furthermore, Aβ plaques have been shown to form rapidly in vivo and to precede neuropathological changes in a mouse model (22). The end surfaces of fibrils (herein termed "fibril ends") are unusually reactive entities: they play a key role in catalyzing recruitment and conformational conversion of amyloid-forming proteins (23, 24) and provide the sites for templated...

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Section: Discussionmentioning

confidence: 92%

mentioning

confidence: 99%

Direct three-dimensional visualization of membrane disruption by amyloid fibrils

Milanesi

Sheynis

Xue

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

172

177

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“…The natural amyloidogenic peptides, such as Aβ, undergo spontaneous intermolecular rearrangement in solution to generate miscellaneous misfolding species (27). However, in fact, most polypeptides can adopt a native structure or form amyloid fibrils by transitioning through a precursor state (16,28).…”

Section: Resultsmentioning

confidence: 99%

Nucleic acid-containing amyloid fibrils potently induce type I interferon and stimulate systemic autoimmunity

Domizio¹,

Dorta‐Estremera²,

Gagea

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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The immunopathophysiologic development of systemic autoimmunity involves numerous factors through complex mechanisms that are not fully understood. In systemic lupus erythematosus, type I IFN (IFN-I) produced by plasmacytoid dendritic cells (pDCs) critically promotes the autoimmunity through its pleiotropic effects on immune cells. However, the host-derived factors that enable abnormal IFN-I production and initial immune tolerance breakdown are largely unknown. Previously, we found that amyloid precursor proteins form amyloid fibrils in the presence of nucleic acids. Here we report that nucleic acid-containing amyloid fibrils can potently activate pDCs and enable IFN-I production in response to self-DNA, self-RNA, and dead cell debris. pDCs can take up DNA-containing amyloid fibrils, which are retained in the early endosomes to activate TLR9, leading to high IFNα/β production. In mice treated with DNA-containing amyloid fibrils, a rapid IFN response correlated with pDC infiltration and activation. Immunization of nonautoimmune mice with DNA-containing amyloid fibrils induced antinuclear serology against a panel of selfantigens. The mice exhibited positive proteinuria and deposited antibodies in their kidneys. Intriguingly, pDC depletion obstructed IFN-I response and selectively abolished autoantibody generation. Our study reveals an innate immune function of nucleic acid-containing amyloid fibrils and provides a potential link between compromised protein homeostasis and autoimmunity via a pDC-IFN axis.autoimmune disease | innate immune response | disease model

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“…Therefore, a gel shift assay can be performed to visualize the direct interaction between the stabilized soluble protein oligomers and DNA, which would result in the retarded migration of DNA on an agarose gel (Figure 2). Cytotoxicity is an archetypical character of soluble protein oligomers 14 . We have shown before that the stabilized soluble protein oligomers dose-dependently induce cell death that is detectible by FACS, trypan blue or CellTiter Blue Subsequently the samples were separated on 1% agarose gel.…”

Section: Representative Resultsmentioning

confidence: 99%

“…These peptides usually lack well defined structure and spontaneously form amyloid in solution over time, a process mediated by a partially misfolded intermediate that serves as the precursor of insoluble amyloid. The precursor of amyloid is also called soluble protein oligomer, which is rare and unstable when generated from natural peptides 14,15 . However, as described earlier, almost any protein in principle can adopt amyloid conformation under the appropriate conditions.…”

Section: Introductionmentioning

confidence: 99%

Rapid Generation of Amyloid from Native Proteins <em>In vitro</em>

Dorta‐Estremera

Cao

2013

JoVE

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Proteins carry out crucial tasks in organisms by exerting functions elicited from their specific three dimensional folds. Although the native structures of polypeptides fulfill many purposes, it is now recognized that most proteins can adopt an alternative assembly of beta-sheet rich amyloid. Insoluble amyloid fibrils are initially associated with multiple human ailments, but they are increasingly shown as functional players participating in various important cellular processes. In addition, amyloid deposited in patient tissues contains nonproteinaceous components, such as nucleic acids and glycosaminoglycans (GAGs). These cofactors can facilitate the formation of amyloid, resulting in the generation of different types of insoluble precipitates. By taking advantage of our understanding how proteins misfold via an intermediate stage of soluble amyloid precursor, we have devised a method to convert native proteins to amyloid fibrils in vitro. This approach allows one to prepare amyloid in large quantities, examine the properties of amyloid generated from specific proteins, and evaluate the structural changes accompanying the conversion. Video LinkThe video component of this article can be found at

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Biochemical and biophysical features of both oligomer/fibril and cell membrane in amyloid cytotoxicity

Cited by 173 publications

References 93 publications

Direct three-dimensional visualization of membrane disruption by amyloid fibrils

Direct three-dimensional visualization of membrane disruption by amyloid fibrils

Nucleic acid-containing amyloid fibrils potently induce type I interferon and stimulate systemic autoimmunity

Rapid Generation of Amyloid from Native Proteins <em>In vitro</em>

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