The prion protein as a receptor for amyloid-beta

I’m currently – 03/05/13 – in the process of updating this. I’ve added the section “Update (May 2013)”, which is still a work in process (i.e. otherwise I would delete this note here).

The title of this node comes from a Nature communication in which recent research linking the prion protein (PrP) and the amyloid-beta (Abeta) protein was discussed. This is a new area of investigation which has seen some intense research in recent years. The obvious reason for this interest is that it suggests a link between different diseases: misfolded PrP is responsible for prion diseases, which include CJD and BSE (aka Mad Cows Disease), while aggregates of Abeta are very prominent in Alzheimer’s disease.

Who is this for?

Everyone. But given that this is a fairly specialized topic I have written this in sections of increasing technicality. For the non-specialist, I’d recommend reading the sections “Introduction” and “Lauren et al. (2009)”. You might find the section called “Continuing research” a bit unforgiving, so please feel comfortable then skipping to the final section. Even if you don’t have any science background please give the next few paragraphs a go, they were written for you. 🙂

Introduction:

Neurons and proteins

The human body is made up of cells – something like 10 to 100 trillion cells. Each cell can be imagined as a small ball containing a copy of our DNA in 23 pairs of chromosomes. The DNA sequence encodes for all the different proteins the cells need in order to make a functional organism (like you). Our DNA encodes on the order of one hundred thousand unique protein forms, which all interact to form signalling cascades and modify their cellular environments.

There are different types of cells. The reason they differ is that even though they all have the same DNA, they don’t read the DNA the same way. At certain points during development, cells are given fates – some cells are told to read their DNA one way and so become neurons while others might read it another way and become muscle cells. Neurons are cells which are specialized to process and transmit information and form the major component of our brain and nervous systems, (the nervous system is what we call the information networks made of neurons).

When a cell’s fate (i.e. the way it reads its DNA) leads it to make a certain protein which is encoded in its DNA we say that that protein has been expressed. Among the myriad proteins expressed by neurons are two that are of interest here: the prion protein (abbreviated PrP) and the Amyloid-beta Precursor Protein (abbreviated APP).

PrP and the protein-only hypothesis

Research into PrP can be divided into two fields. One field is concerned with what the physiological function of PrP is, that is, why millions of years of evolution have produced and maintained the existence of this protein. The second field is concerned with PrP as the substrate of a group of related diseases known as prion diseases¹.

At the present time, there is strong evidence supporting roles for PrP in neuronal resilience to stress (e.g. following strokes), regulation of cell metabolism (i.e. energy usage and production), regulation of copper homoeostasis, immune cells’ responses, and more. If this list looks haphazard that is because in biology everything connects to everything².

The second question about PrP and disease stems from the protein-only hypothesis. This can be understood in the following way: there are many protein misfolding diseases. These are diseases that involve a protein misfolding to form protein aggregates, and includes diseases like Alzheimer’s disease and Huntington’s disease. Prion diseases also involve protein misfolding, but unlike those other diseases, the diseases only require PrP. Whereas Alzheimer’s requires some sort of hereditary mutation or toxic environment to manifest protein misfolding, in prion diseases all you need is a misfolded prion. In prion diseases misfolding prions (called PrP^sc) interact with healthy PrP (called PrP^c), teaching the healthy PrP to become misfolded too. That means that unlike diseases like Alzheimer’s, prion diseases can be spread by exposure of healthy PrP to misfolded PrP.

APP processing and amyloid plaques

APP is another protein which is expressed significantly by neurons and has been a major interest due to the amyloid hypothesis in Alzheimer’s disease. Simply stated, the hypothesis suggests that the toxic plaques (protein aggregates) which form during Alzheimer’s disease form part of the disease mechanism (i.e. how the disease makes you sick). The way APP contributes to plaques is by being processed to form amyloid-beta (Abeta) – the protein itself gets cut to release a small protein called amyloid-beta which can aggregate.

Now here’s a good question: so APP processing into Abeta forms plaques, but why does this prominently affect neurons? Why is Alzheimer’s primarily a brain disease? Why doesn’t it cause muscle dystrophy or your skin to fall off? What I’m getting at is that perhaps if we know why Abeta affects neurons preferentially we could understand what the mechanism of Abeta toxicity is.

Based on what we know from above, how could we answer this question? Above I talked about cells having different fates and expressing different proteins. If we knew every single protein expressed by neurons, and then we put those proteins one by one into non-neuronal cells, then we could check which protein is responsible for Abeta’s affinity and toxicity to neurons.

Lauren et al. (2009)

In February 2009 a group published a paper in the prestigious science journal Nature. The paper was entitled “Cellular prion protein mediates impairment of synaptic plasticity by amyloid-beta oligomers” and was authored by Juha Lauren, David A. Gimbel, Haakon B. Nygaard, John W. Gilbert and Stephen M. Strittmatter.

The set-up for this experiment is incredible and is worth elaborating upon. The group accessed a neuronal cDNA (complementary DNA) library. These cDNA are small pieces of DNA which separately correspond to every single protein expressed by neurons. They took all the 5000 cDNA’s from this library and expressed them in different combinations to form 225,000 clonal cell lines – this means they took different combinations of cDNA from the library and made a non-neuronal cell express it, then grew that cell up into large numbers. After making 225,000 separate cell lines, they treated the cells with a solution containing Abeta and measured which cell lines could grab onto the toxic Abeta. From all this, they came up with one strong candidate – PrP – and a number of weaker candidates (not considered here).

So they’d identified PrP as binding toxic Abeta. What next? They continued their research by asking whether PrP was important for the actual toxicity observed with Abeta. They showed that toxic Abeta (i.e. grouped Abeta) depends on PrP expression to kill cells, and that this occurs somehow by Abeta actually interacting with PrP, and that even Abeta’s ability to reduce neuronal signalling (i.e. the way different neurons send each other messages) is dependent on PrP. This was all extraordinarily impressive. So now what?

Continuing research

When Stephen Strittmatter discovered an unexpected link between key proteins in two devastating brain maladies — Alzheimer’s disease and Creutzfeldt–Jakob disease (CJD) — researchers in the field agreed that he was on to something big. But a year on, conflicting results, including findings published in Nature this month, have clouded that rosy picture and highlight the challenge faced by researchers seeking a way to arrest Alzheimer’s disease progression.

“Key Alzheimer’s finding questioned” Nature (2010)

Since the original paper, a number of groups have pursued the question of PrP’s relationship to Abeta toxicity. Unfortunately, as indicated by the above quote, the picture has quickly become more complicated than the original results revealed. Here I intend to summarise the experiments described by those papers that have been published since Lauren’s 2009 Nature Paper. Please note that many of these are from leaders in the fields. The suggestions of contradicting results should NOT be construed as indicating incompetency or fraudulence. Biology is the most complicated system we know, and these researchers are amongst the leaders in that field³.

Lauren et al. (2009) Nature. Show that exogenously applied Abeta oligomers are specifically neurotoxic in the presence of PrP. Used both cell models and transgenic mice.
Balducci et al. (February 2010) PNAS. This was the first group to publish a response to the Lauren paper. The group injected various preparations of Abeta oligomers into mice and showed that memory impairment was Abeta but not PrP dependent. This is despite being able to repeat the first group’s conclusion that Abeta oligomers preferentially bind PrP in vitro.
Morales et al. (March 2010) J Neurosci. This is another response to the Lauren paper by another group. This group wondered whether Lauren’s paper implied that Abeta could influence toxic PrP (i.e. PrP^sc) in prion disease. They used a mouse line with mutations that cause more Abeta to form and inoculated them with misfolded PrP. They showed that the two diseases could increase each other and that Abeta and misfolded PrP can physically associate.
Gimbel et al. (May 2010) J Neurosci. This is an impressive follow up by the original group and involves crossing transgenic PrP-null and mice with mutations that cause more Abeta to form. They showed that both neurotoxicity and cognitive decline in the Alzheimer’s model mice depended on those mice also expressing PrP.
Calella et al. (July 2010) EMBO Mol Med. This follows that same premises as Gimbel’s study, however, uses different mutations for their Abeta-prone mice cohort. Whereas Gimbel used mice with the Swedish APP mutation (that produce overexpress wild-type APP) crossed with gamma-secretase mutants ([link]), this study by Calella used mice with a mutated APP (KM670/671NL) and (a different) mutant gamma-secretase. In contrast to Gimbel et al., this group did not find Alzheimer model toxicity to depend on PrP expression ([link]).
Chen et al. (August 2010) J Biol Chem. This another group investigating the physical Abeta/PrP interaction. They repeated previous results showing that PrP selectively bound Abeta oligomers. Various elements of Abeta/PrP binding were quantified, and the binding was shown to depend on PrP residues 23-27 and 92-110.
Kessels et al. (August 2010) Nature. This was a paper that was published to directly question Lauren’s original conclusions. They showed that hippocampal slices that overexpress a C-terminal section of APP (viral-based transfection), or which are treated with exogenous Abeta, display reduced LTP in a PrP-independent manner.

Lauren published a back-to-back response to Kessel’s Nature publication. Lauren emphasised that the study described by Kessel depended on viral expression of a truncated APP peptide, used greater Abeta concentrations, and did not demonstrate a dose-dependent Abeta toxicity response (i.e. observed full plasticity suppression at lowest levels). Which are all valid criticisms, but they suggest another problem: What does it mean to study and model a disease? Why should Lauren’s particular experiments be considered in any sense better than Kessel’s, or vice versa?

Update (May 2013):

A fair bit of work has been done to continue characterizing the link between PrP and APP between 2010-2013:

More studies have confirmed that Abeta binding to PrP induces pathological effects. There is now a greater understanding of the molecular mechanisms (viz. the nature of the cascade of molecules that interact) causing the PrP-Abeta toxicity. Evidence suggests that Abeta causes PrP to cross-link (i.e. bind itself in pairs) and recruit a molecule called cPLA2 to their lipid rafts (which are structures on the surface of cells) (Bate 2011), and cause signalling via the molecule Fyn inside the cell (Um 2012) which damages the neuron-signalling neuronal receptor NMDAR (Um 2012) as well as causing changes to the molecular-chains of Tau that are associated with Alzheimer’s disease specifically (Larson 2012). Those lipid rafts, to which one study linked cPLA2, were shown to be necessary for the toxic effect of PrP-Abeta binding, including the presence of the raft molecule LRP1 (Rushworth 2013).
PrP expression (i.e. levels) are independent of APP, regardless of which type of APP, nor whether it contains a mutation linked to Alzheimer’s (Lewis 2012).
A PrP fragment called N1, which is produced under normal conditions in cells and which has been associated with PrP’s neuroprotective function, was shown to bind Abeta in such a way as to inhibit Abeta’s neurotoxicity (Fluharty 2013).

Understanding and modelling disease

I think there are two reasonable conclusions that can be made at this point. One, Abeta seems to have some sort of selective affinity for PrP, however, it’s not clear whether this interaction has any in vivo consequences, and whether any such consequences are mechanisms of Alzheimer’s pathology.

The second conclusion is that what model you use to study your disease is very important.

The reason we model diseases is because we can’t study patients. The most obvious second-best is to study animals that display the disease. But what does it mean for a mouse or an ape to have Alzheimer’s disease? For that matter, what does it mean for a human being to have Alzheimer’s disease. There are so many different possible mutations that can increase disease rates, not to mention environmental factors – which of these are the “true” (or best) manifestations of the disease?

There’s no one disease that can be studied. Diseases can be conceptualised as states in which organisms demonstrate particular proclivities towards particular modes of toxicity. To put it another way, if you got every single Alzheimer’s patient, and you described every single one of their symptoms, you could draw up a table of percentage breakdowns. You’d include stuff like “Abeta clumps at so-and-so per cent”. You’d also include more complicated descriptions like “Abeta₄₂ fibril-dependent, nicotinic receptor-mediated toxicity at so-and-so per cent”. If we could have a perfect collection of facts similar to what I’ve described then we’d also have a perfect description of Alzheimer’s disease.

Of course, that’s not a very practical way of going about things. So instead we seek out common observations and mechanisms, and then replicate those in models. All the studies described above decided that a key element in the disease is the formation of Abeta. However, some groups favoured injections of Abeta preparations, other favoured mutants and different groups preferred different mutations and different tools for generating, expressing, and validating those mutations.

Which leaves a final question: What’s the best model for Alzheimer’s disease?

Notes.

These two fields (PrP as physiological component and as pathogenic substrate) are not mutually exclusive, and notably, overlap along the hypothesis of “pathogenic misfolding of PrP may confer toxicity by corrupting PrP’s physiological function”.
In my mind, the best way of analogising this is by considering a single person as a node which is part of a society which is a system. In biology, there are different ways of dichotomizing node/system, but in this example, the node is the protein and the system is the collection of functions undertaken by its hosting cell (i.e. person/protein as node, society/cell as system). You could ask how the person interacts with the system. I might tell you about the different properties displayed by the person, e.g. particular sorts of kindness, desires, intelligences, interests. This corresponds to a protein’s tendencies (e.g. hydrophilicity, topology, catalytic functions, etc). This is however not enough. So I tell you about what sort of person it is, e.g. consumer, religious, academic, etc. This corresponds to the categories of proteins, e.g. kinase, proteolytic protein, scaffolding protein, etc. But this still isn’t enough. What you want to know is exactly how this sort of person interacts with society, and that requires an answer which can summarise a person’s entire life! Imagine everything that a person does that interacts with society, that is the sort of understanding which we need for proteins’ interactions with a cell.
Conflict of interest: No. I have no relationship at all with any of the authors of any of the papers at the present time.

Major References.

Is PrPC a Mediator of A Toxicity in Alzheimer’s Disease? (2010) J Neurosci
Key Alzheimer’s finding questioned (2010) Nature
Balducci et al. Synthetic amyloid-beta oligomers impair long-term memory independently of cellular prion protein (2010) PNAS
Calella et al. Prion protein and Abeta-related synaptic toxicity impairment (2010) EMBO Mol Med
Chen et al. Interaction between human prion protein and amyloidbeta-(Abeta) oligomers (2010) J Biol Chem
Gimbel et al. Memory impairment in transgenic Alzheimer mice requires cellular prion protein. (2010) J Neurosci
Kessel et al. The prion protein as a receptor for amyloid-beta (2010) Nature
Lauren et al. Cellular prion protein mediates impairment of synaptic plasticity by amyloid-beta oligomers. (2009) Nature
Lauren et al. Lauren et al. reply (2010) Nature
Morales et al. Molecular crosstalk between misfolded proteins in animal models of Alzheimer’s and prion diseases. (2010) J Neurosci