Guilt by Association?—Aβ, α-Synuclein Make Mixed Oligomers
The presence of small oligomeric protein assemblies with neurotoxic properties has become a common thread in several neurodegenerative diseases—in Alzheimer disease, there are amyloid-β (Aβ) and tau, and in Parkinson disease, there is α-synuclein. Now that thread may stitch the different diseases together even more tightly, with a new study suggesting that under some conditions, Aβ and α-synuclein may form mixed oligomers. The study, published September 4 in PLoS ONE by Eliezer Masliah and colleagues at the University of California at San Diego, combines molecular modeling, biophysical techniques, and cell culture experiments to support the idea that Aβ/α-synuclein co-complexes assume a ring-like conformation on membranes. Further, the data suggest that the mixed oligomers may act as cation channels and perturb cellular calcium levels.
Aβ and α-synuclein pathologies frequently overlap. In dementia with Lewy body disease (DLB), α-synuclein inclusions and amyloid plaques coexist. Also, 15-25 percent of Alzheimer's cases develop Lewy bodies and motor deficits similar to Parkinson disease, while some Parkinson's patients develop dementia, often in association with amyloid plaques. These overlapping disorders suggest there may be an association between Aβ and α-synuclein, and indeed years ago Masliah and colleagues found a fragment of α-synuclein in amyloid plaques (Ueda et al., 1993).
The two proteins clearly cooperate in animal models: APP-expressing mice crossed with α-synuclein transgenics show enhanced accumulation and toxicity of α-synuclein (Masliah et al., 2001; Mandal et al., 2006), while α-synuclein mice display elevated Aβ aggregation (see ARF related news story).
In the new study, first author Igor Tsigelny and colleagues initially looked to see if the two proteins were found together in vivo. As they had shown before, the presence of Aβ increased the levels of aggregated α-synuclein in APP/α-synuclein transgenic mice. Moreover, in brain extracts from patients with LBD or the transgenic mice, the two proteins immunoprecipitated together, suggesting a direct interaction.
Tsigelny then used computer modeling to simulate what happens when the proteins meet in the presence of a lipid membrane. The molecular dynamics modeling suggest that interactions occurred between the N-terminus of Aβ and both N- and C-termini of α-synuclein. Importantly, the simulation suggested that docking of Aβ onto an α-synuclein dimer stabilized the complex on the membrane, and promoted the addition of α-synuclein monomers. Progressive recruitment of more α-synuclein resulted in ring-like hybrid oligomers of five or six α-synuclein molecules with the Aβ on the membrane. Over time, energy-minimizing simulations suggested that the oligomers became embedded in the membrane. Even when the two proteins started on opposite sides of the lipid bilayer, as they do in their natural habitat of the brain, the simulations suggested they could penetrate and interact in the membrane.
For real-life evidence of such interactions, the investigators mixed monomeric or aggregated Aβ and α-synuclein, and demonstrated that Aβ promoted the aggregation of α-synuclein. The proteins directly interacted, as evidenced by co-immunoprecipitation, and a mutational analysis showed that complex formation depended on residues in N-terminal 18 amino acids of Aβ, which were the same as the contact residues identified in the modeling experiments.
Both α-synuclein and Aβ on their own can form pore-like oligomers (see Lashuel et al., 2002; Quist et al., 2005; ARF related news story), but in the in vitro conditions of the current study, only the mixture assumed well-defined ring-like structures as visualized by electron microscopy. Either protein alone adopted globular structures 5-10 nm across. At longer incubation, the mixed oligomers formed fibrils. Adding lipids enhanced formation of rings by either protein alone or together.
Several studies have suggested that pore formation by synuclein could contribute to neurodegeneration, so the researchers measured ion currents in cells overexpressing α-synuclein. They found elevated whole-cell cation currents in α-synuclein expressing cells, which were further enhanced by treatment with soluble Aβ. Cation pores might contribute to calcium dysregulation by allowing influx from external stores. In agreement with this idea, the synuclein -expressing cells had a twofold elevation in calcium levels, which was further increased by treatment with Aβ.
“This is the first paper that shows co-oligomers,” Masliah told ARF. “We know that amyloid interaction with α-synuclein can lead to α-synuclein aggregation, and that α-synuclein pathology could be enhanced by Aβ in vivo, but we didn’t know how. This work shows how the two proteins might come from different compartments to interact and form unusual hybrid aggregates that lodge into the membrane and cause this pore type of pathology.” In addition, he said, there are probably indirect mechanisms by which the two proteins could enhance each other’s toxicity.
The mixed oligomers could be a target for therapeutics, Masliah also noted. “If our model is correct, the next logical hypothesis is that if you block the interactions, you should disaggregate these complexes and ameliorate the disease. From the combination of modeling and biological experiments, we have detailed molecular information about the interaction that can be used to design drugs to block the interaction, and that is what we are trying to do.”—Pat McCaffrey