Like Aβ oligomers, soluble aggregates of tau and α-synuclein bind to cellular prion protein, poisoning neurons, according to a study published in Acta Neuropathologica. Using a standardized procedure to produce soluble aggregates of these three proteins, researchers led by Dominic Walsh, Brigham and Women’s Hospital, Boston, report that all three types of aggregate bind to the same sequence in cellular prion protein (PrPC). As a result, neurites shrank, and synapses became less plastic. “This work clearly suggests that inhibition of oligomer-to-PrPC binding is an important therapeutic target for multiple neurodegenerative diseases,” wrote Thomas Wisniewski, New York University School of Medicine.
- Soluble aggregates, not monomers, of Aβ, tau, and α-synuclein bind to PrP.
- Binding causes neurotoxicity.
- PrP may be a therapeutic target for several late-life neurodegenerative diseases.
To take things one step further, the researchers recapitulated the experiments with human material, isolating Aβ, tau, and α-synuclein oligomers postmortem from people who had had Alzheimer’s or Pick’s disease, or dementia with Lewy bodies. Extracts from all three diseases were toxic to induced human neurons, but not if the neurons were missing PrP. “Our findings suggest that both the synthetic and brain-derived aggregates of these proteins cause toxicity that requires the expression of PrP,” Walsh told Alzforum.
The study was published last December around the same time as two others implicating PrP in AD. Researchers led by Tara Spires-Jones at the University of Edinburgh reported that mice overexpressing amyloid precursor protein (APP) and tau upregulated PrP (Pickett et al., 2019).
Dietmar Thal and colleagues at KU Leuven, Belgium, demonstrated that soluble Aβ and phosphorylated tau (p-tau) interact with PrPC in vitro, in transgenic mice, and in Alzheimer’s disease brain (Gomes et al., 2019). And in mice that overexpressed soluble Aβ, its binding to PrP correlated with Aβ-driven acceleration of tangle pathology through the brain. “The more Aβ we have, the more likely we have interactions that exacerbate tau pathology,” Thal told Alzforum.
“The new findings by Walsh and colleagues support the role of PrP as a ‘general receptor’ of soluble protein aggregates,” wrote Thal and Luis Gomes, also from KU Leuven, in a comment to Alzforum (see below). “The binding of these aggregates also caused functional and structural deficits that were rescued when PrP was either ablated or blocked,” they noted.
Walsh’s study builds on previous work describing PrP as a critical Aβ oligomer receptor (Salazar and Strittmatter, 2017). “By and large, nearly every previous study has found that certain Aβ assemblies bind to PrP. Everyone agrees. Where people are disagreeing is on the consequences,” said Walsh. What’s more, not all forms of Aβ bind to PrP, he added, and some forms that do not bind might still be toxic. To complicate matters, while other studies indicate that PrP also binds tau and α-synuclein, no one has directly compared the interaction of all three with PrP (Ferreira et al., 2017; Hu et al., 2018; Ondrejcak et al., 2018). This is partly because, until now, there was no standardized procedure for preparing soluble protein aggregates, Walsh explained.
First author Grant Corbett and colleagues developed a reproducible procedure to generate homogenous soluble protein aggregates of Aβ, tau, and α-synuclein. First, they created insoluble, end-stage fibrillar aggregates from monomers of each, following established protocols (Buell et al., 2014; Hellstrand et al., 2010; O’Dowd et al., 2012). Next, they centrifuged the fibrils, removed the monomers, resuspended the fibrils, and then broke them into small soluble aggregates by sonication. Then they tested how each bound PrP and curtailed neurite growth of mouse primary neurons (MPN), and of human neurons derived from induced pluripotent stem-cells (iNs).
Aβ, tau, and α-synuclein monomers were not toxic to these cells, but soluble aggregates of all three proteins stunted neurites in both. This toxicity was PrP-dependent: Mouse and human neurons that lacked the prion protein showed no signs of neurite damage.
Neurotoxicity. Soluble aggregates of Aβ, tau, and α-synuclein poison mouse primary neurons in a dose-dependent manner; monomers do not. [Courtesy of Corbett et al., Acta Neuropathologica, 2019.]
Would soluble aggregates found in diseased human brains do the same? To find out, the researchers incubated induced neurons with aqueous extracts containing Aβ, tau, or α-synuclein from two brains each of people who had died with Alzheimer’s, Pick’s, and DLB, respectively. For comparison, they used AD, Pick’s, and DLB extracts from which they had depleted these aggregates, as well as extracts from one healthy brain.
All extracts from diseased brains reduced neurite length, whereas depleted extracts did not. Strikingly, neurons that had their PrP gene knocked out by CRISPR were protected from Aβ, tau, and α-synuclein aggregates, pointing once again toward a role for PrP in toxicity.
PrP Dependence. Soluble extracts (Mock) from AD, DLB, and Pick’s brains shortened neurites of induced human neurons. Neurons lacking PrP were protected (gray shading). Extracts immunodepleted using antibodies to Aβ (AW7 ID), α-synuclein (2F112 ID), and tau (Tau5 ID) were neutralized. [Courtesy of Corbett et al., Acta Neuropathologica, 2019.]
The researchers also tested how recombinant soluble protein aggregates would affect synaptic plasticity using multielectrode recordings from hippocampal slices of both wild-type and genetically ablated PrP-deficient mice. Aβ, tau, and α-synuclein aggregates all inhibited long-term potentiation (LTP), but only when the mice expressed PrP. “It looks like the PrP receptor is the target of the protein aggregates. Then, regardless of which protein, they may lead to neurodegeneration,” Thal said. Curiously, tau aggregates blocked LTP at much lower concentrations than did Aβ or α-synuclein aggregates.
Susan Catalano, Cognition Therapeutics Inc., Pittsburgh, called this difference striking, noting that the synthetic oligomers contain β-sheet conformations. “A characteristic of β-sheet structures is promiscuous unsaturable binding, usually to negatively charged residues on a variety of extracellular matrix molecules on the cell surface, including prion and gangliosides. In this light, the dramatic difference in concentrations of tau protofibrils required to block LTP compared to Aβ and α-synuclein aggregates is noteworthy,” she wrote (Matsuzaki et al., 2018).
The paper raises questions about how Aβ and tau contribute to AD pathology. “The authors immunodepleted Aβ [from AD brain extracts] with the AW7 antibody, which seemed sufficient to rescue the toxic effects. However, those brain extracts still contained tau that apparently did not cause toxicity,” Thal and Gomes wrote.
The lack of tau toxicity in these extracts puzzled Walsh, as well. “In AD, there are two hallmark proteins, Aβ and tau, but the toxicity we saw in experiments with AD brain extracts was mediated only by Aβ, not tau,” he said. “It is not clear to me what the difference is between the tau in the AD brain extract versus the tau in the Pick’s brain extract. I can only speculate that different structures may be involved.”
Researchers have found slight differences in the structures of tau fibrils isolated from people who had AD, Pick’s, and even chronic traumatic encephalopathy (Fitzpatrick et al., 2017; Falcon et al., 2018; Falcon et al., 2019).
Tiago Outeiro, University Medical Center Göttingen in Germany, thinks that from a therapeutic standpoint targeting PrP could offer a one-stop solution. “This study supports the idea that targeting PrP may constitute a valid strategy for protecting neuronal function from a series of proteotoxic injuries,” Outeiro wrote. Walsh agreed, but cautioned that the soluble aggregates are pleomorphic, and that PrP is probably not responsible for all their toxic interactions.
And how does PrP mediate toxicity? Does it promote templated misfolding of Aβ, tau, and α-synuclein, as it does for prions? Or does it act like a cell surface receptor, transducing signals to the cytosol? “We can say at the moment that PrP is involved in the process, but whether it triggers aggregation is unclear,” Thal told Alzforum. Walsh doubts PrP seeds misfolding. “It would have to bind monomer, and it does so only when monomer concentration is high enough for spontaneous aggregation,” he said.
Researchers agree on what ought to come next. “The next big step will be to examine these interactions in animal models in more detail,” wrote Colin Masters, University of Melbourne. He wants to see how these proteins interact at a molecular level. “There is still a big gap in demonstrating a biologically relevant, direct physical interaction between these extracellular and intracellular molecular entities,” he wrote. “If PrP mediates the Aβ-tau phenomenon in Alzheimer’s disease, the most parsimonious way to demonstrate this might lie in a therapeutic strategy aimed at targeting PrP in AD, and that is doable,” Masters believes.—Sandra Blumenrath
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