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Mechanisms of hybrid oligomer formation in the pathogenesis of combined Alzheimer's and Parkinson's diseases.

Tsigelny IF, Crews L, Desplats P, Shaked GM, Sharikov Y, Mizuno H, Spencer B, Rockenstein E, Trejo M, Platoshyn O, Yuan JX, Masliah E - PLoS ONE (2008)

Bottom Line: While progressive accumulation of amyloid beta protein (Abeta) oligomers has been identified as one of the central toxic events in AD, accumulation of alpha-synuclein (alpha-syn) resulting in the formation of oligomers and protofibrils has been linked to PD and Lewy body Disease (LBD).These results support the contention that Abeta directly interacts with alpha-syn and stabilized the formation of hybrid nanopores that alter neuronal activity and might contribute to the mechanisms of neurodegeneration in AD and PD.The broader implications of such hybrid interactions might be important to the pathogenesis of other disorders of protein misfolding.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, University of California San Diego, La Jolla, California, United States of America.

ABSTRACT

Background: Misfolding and pathological aggregation of neuronal proteins has been proposed to play a critical role in the pathogenesis of neurodegenerative disorders. Alzheimer's disease (AD) and Parkinson's disease (PD) are frequent neurodegenerative diseases of the aging population. While progressive accumulation of amyloid beta protein (Abeta) oligomers has been identified as one of the central toxic events in AD, accumulation of alpha-synuclein (alpha-syn) resulting in the formation of oligomers and protofibrils has been linked to PD and Lewy body Disease (LBD). We have recently shown that Abeta promotes alpha-syn aggregation and toxic conversion in vivo, suggesting that abnormal interactions between misfolded proteins might contribute to disease pathogenesis. However the molecular characteristics and consequences of these interactions are not completely clear.

Methodology/principal findings: In order to understand the molecular mechanisms involved in potential Abeta/alpha-syn interactions, immunoblot, molecular modeling, and in vitro studies with alpha-syn and Abeta were performed. We showed in vivo in the brains of patients with AD/PD and in transgenic mice, Abeta and alpha-synuclein co-immunoprecipitate and form complexes. Molecular modeling and simulations showed that Abeta binds alpha-syn monomers, homodimers, and trimers, forming hybrid ring-like pentamers. Interactions occurred between the N-terminus of Abeta and the N-terminus and C-terminus of alpha-syn. Interacting alpha-syn and Abeta dimers that dock on the membrane incorporated additional alpha-syn molecules, leading to the formation of more stable pentamers and hexamers that adopt a ring-like structure. Consistent with the simulations, under in vitro cell-free conditions, Abeta interacted with alpha-syn, forming hybrid pore-like oligomers. Moreover, cells expressing alpha-syn and treated with Abeta displayed increased current amplitudes and calcium influx consistent with the formation of cation channels.

Conclusion/significance: These results support the contention that Abeta directly interacts with alpha-syn and stabilized the formation of hybrid nanopores that alter neuronal activity and might contribute to the mechanisms of neurodegeneration in AD and PD. The broader implications of such hybrid interactions might be important to the pathogenesis of other disorders of protein misfolding.

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Electrophysiological analysis of the cellular effects of hybrid Aβ/α-syn multimers.293T cells were infected with lenti-α-syn and lenti-GFP and treated with Aβ and analyzed by electrophysiology. (A) Immunoblot analysis showing expression levels of α-syn and GFP in treated cultures. (B) Immunocytochemical analysis demonstrating high efficiency of infection and co-localization between α-syn and GFP. (C, D) Representative currents (C) elicited by depolarizing the cells from a holding potential of −50 mV to a series of test potentials ranging from −80 mV to +80 mV, and corresponding current voltage-relationship curves (D), in α-syn-expressing cells treated with vehicle (α-syn, n = 5), vector-transfected cells treated with Aβ (Aβ, n = 5), and α-syn-expressing cells treated with Aβ (α-syn+Aβ, n = 6).
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pone-0003135-g007: Electrophysiological analysis of the cellular effects of hybrid Aβ/α-syn multimers.293T cells were infected with lenti-α-syn and lenti-GFP and treated with Aβ and analyzed by electrophysiology. (A) Immunoblot analysis showing expression levels of α-syn and GFP in treated cultures. (B) Immunocytochemical analysis demonstrating high efficiency of infection and co-localization between α-syn and GFP. (C, D) Representative currents (C) elicited by depolarizing the cells from a holding potential of −50 mV to a series of test potentials ranging from −80 mV to +80 mV, and corresponding current voltage-relationship curves (D), in α-syn-expressing cells treated with vehicle (α-syn, n = 5), vector-transfected cells treated with Aβ (Aβ, n = 5), and α-syn-expressing cells treated with Aβ (α-syn+Aβ, n = 6).

Mentions: Since previous studies have suggested that the α-syn ring-like structures might form pores in the membrane that could be responsible for the neurotoxic effects of α-syn oligomers [25], [26], [28], [30], we investigated whether abnormally high levels of ion currents are detected in cells over-expressing α-syn and if this process might be enhanced by Aβ. For this purpose, we recorded and compared whole-cell cation currents in human embryonic kidney (HEK) 293T cells transduced with lentiviral (lenti) vectors expressing α-syn or green fluorescent protein (GFP) as a control in the presence or absence of Aβ (Figure 7). Immunoblot analysis confirmed that cells expressed comparable levels of α-syn (Figure 7A). Double-labeling verified that in co-transduced cells, GFP was also expressed with α-syn (Figure 7B). The target cells (displaying green fluorescence) for electrophysiological measurements were identified by co-transduction with the lenti-GFP vector (Figure 7B).


Mechanisms of hybrid oligomer formation in the pathogenesis of combined Alzheimer's and Parkinson's diseases.

Tsigelny IF, Crews L, Desplats P, Shaked GM, Sharikov Y, Mizuno H, Spencer B, Rockenstein E, Trejo M, Platoshyn O, Yuan JX, Masliah E - PLoS ONE (2008)

Electrophysiological analysis of the cellular effects of hybrid Aβ/α-syn multimers.293T cells were infected with lenti-α-syn and lenti-GFP and treated with Aβ and analyzed by electrophysiology. (A) Immunoblot analysis showing expression levels of α-syn and GFP in treated cultures. (B) Immunocytochemical analysis demonstrating high efficiency of infection and co-localization between α-syn and GFP. (C, D) Representative currents (C) elicited by depolarizing the cells from a holding potential of −50 mV to a series of test potentials ranging from −80 mV to +80 mV, and corresponding current voltage-relationship curves (D), in α-syn-expressing cells treated with vehicle (α-syn, n = 5), vector-transfected cells treated with Aβ (Aβ, n = 5), and α-syn-expressing cells treated with Aβ (α-syn+Aβ, n = 6).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2519786&req=5

pone-0003135-g007: Electrophysiological analysis of the cellular effects of hybrid Aβ/α-syn multimers.293T cells were infected with lenti-α-syn and lenti-GFP and treated with Aβ and analyzed by electrophysiology. (A) Immunoblot analysis showing expression levels of α-syn and GFP in treated cultures. (B) Immunocytochemical analysis demonstrating high efficiency of infection and co-localization between α-syn and GFP. (C, D) Representative currents (C) elicited by depolarizing the cells from a holding potential of −50 mV to a series of test potentials ranging from −80 mV to +80 mV, and corresponding current voltage-relationship curves (D), in α-syn-expressing cells treated with vehicle (α-syn, n = 5), vector-transfected cells treated with Aβ (Aβ, n = 5), and α-syn-expressing cells treated with Aβ (α-syn+Aβ, n = 6).
Mentions: Since previous studies have suggested that the α-syn ring-like structures might form pores in the membrane that could be responsible for the neurotoxic effects of α-syn oligomers [25], [26], [28], [30], we investigated whether abnormally high levels of ion currents are detected in cells over-expressing α-syn and if this process might be enhanced by Aβ. For this purpose, we recorded and compared whole-cell cation currents in human embryonic kidney (HEK) 293T cells transduced with lentiviral (lenti) vectors expressing α-syn or green fluorescent protein (GFP) as a control in the presence or absence of Aβ (Figure 7). Immunoblot analysis confirmed that cells expressed comparable levels of α-syn (Figure 7A). Double-labeling verified that in co-transduced cells, GFP was also expressed with α-syn (Figure 7B). The target cells (displaying green fluorescence) for electrophysiological measurements were identified by co-transduction with the lenti-GFP vector (Figure 7B).

Bottom Line: While progressive accumulation of amyloid beta protein (Abeta) oligomers has been identified as one of the central toxic events in AD, accumulation of alpha-synuclein (alpha-syn) resulting in the formation of oligomers and protofibrils has been linked to PD and Lewy body Disease (LBD).These results support the contention that Abeta directly interacts with alpha-syn and stabilized the formation of hybrid nanopores that alter neuronal activity and might contribute to the mechanisms of neurodegeneration in AD and PD.The broader implications of such hybrid interactions might be important to the pathogenesis of other disorders of protein misfolding.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Biochemistry, University of California San Diego, La Jolla, California, United States of America.

ABSTRACT

Background: Misfolding and pathological aggregation of neuronal proteins has been proposed to play a critical role in the pathogenesis of neurodegenerative disorders. Alzheimer's disease (AD) and Parkinson's disease (PD) are frequent neurodegenerative diseases of the aging population. While progressive accumulation of amyloid beta protein (Abeta) oligomers has been identified as one of the central toxic events in AD, accumulation of alpha-synuclein (alpha-syn) resulting in the formation of oligomers and protofibrils has been linked to PD and Lewy body Disease (LBD). We have recently shown that Abeta promotes alpha-syn aggregation and toxic conversion in vivo, suggesting that abnormal interactions between misfolded proteins might contribute to disease pathogenesis. However the molecular characteristics and consequences of these interactions are not completely clear.

Methodology/principal findings: In order to understand the molecular mechanisms involved in potential Abeta/alpha-syn interactions, immunoblot, molecular modeling, and in vitro studies with alpha-syn and Abeta were performed. We showed in vivo in the brains of patients with AD/PD and in transgenic mice, Abeta and alpha-synuclein co-immunoprecipitate and form complexes. Molecular modeling and simulations showed that Abeta binds alpha-syn monomers, homodimers, and trimers, forming hybrid ring-like pentamers. Interactions occurred between the N-terminus of Abeta and the N-terminus and C-terminus of alpha-syn. Interacting alpha-syn and Abeta dimers that dock on the membrane incorporated additional alpha-syn molecules, leading to the formation of more stable pentamers and hexamers that adopt a ring-like structure. Consistent with the simulations, under in vitro cell-free conditions, Abeta interacted with alpha-syn, forming hybrid pore-like oligomers. Moreover, cells expressing alpha-syn and treated with Abeta displayed increased current amplitudes and calcium influx consistent with the formation of cation channels.

Conclusion/significance: These results support the contention that Abeta directly interacts with alpha-syn and stabilized the formation of hybrid nanopores that alter neuronal activity and might contribute to the mechanisms of neurodegeneration in AD and PD. The broader implications of such hybrid interactions might be important to the pathogenesis of other disorders of protein misfolding.

Show MeSH
Related in: MedlinePlus