Prions are a totally unique disease-causing agent. They are known to be responsible for bovine spongiform encephalopathy (BSE or mad cow disease), Creutzfelt Jacob Disease (CJD), scrapie, and Kuru. They are so unusual because they contain no genetic material in the form of DNA or RNA like a virus or bacteria. They are composed entirely of protein, which was previously thought to be incapable of causing an infectious disease.

It seems that the proteins responsible for these diseases are normal proteins present in the brains of sheep, cattle, and humans. In their normal state, they are folded in a certain conformation, consisting of mainly alpha-helical domains. In the variant, disease causing form, they seem to be folded in a different conformation, mostly beta-pleated-sheets. It seems that this switch in structure can be transmitted, as though the variant (disease causing form) catalyzes the change in otherwise normal proteins.

The disease has been proven transmissible through cannibalism (eating brains in particular for Kuru), inoculation, and through genetic predisposition (most common in CJD, due to a mutation in the protein chain that makes it more likely to switch conformation). It has been shown that brain extract from BSE-infected cattle is capable of inducing CJD in chimpanzees, suggesting that the same could happen for humans.

These diseases all affect the brain, causing ataxia, progessive dementia, and ultimately, death in their respective species.
Prion-mediated diseases are an interesting direct application of the protein folding problem to pathology. It highlights the difference between kinetic and thermodynamic stability. The innocuous alpha helical form folds very quickly and is a metastable conformation. The disease causing beta sheet conformation, is energetically slightly more stable, however it is kinetically inaccessible (i.e. it takes a very, very long time to find this conformation randomly). This process is sped up only by catalysis, where one protein in the disease conformation helps another reach the same shape. Upon doing so, these proteins assemble and form plaques. These plaques eventually build up in the cells, disrupting normal cell processes, causing disease.

Because prions are proteins and not viruses or bacteria, they are small enough to penetrate the blood brain barrier, which normally keeps disease out of the brain. As a result, these plaques build up in neurons, causing mental illness and eventually death.

This write-up is an attempt to give an overview of the current status of the intriguing puzzle of prion research. Some things are known more or less for sure, more is suspected and speculated, and I’ll end with some remaining questions. The aspects per section are mentioned in pretty much of a random order.

What they think they know
1. A prion is a proteinaceous infectious particle and can induce spongiform encephalopathies in a lot of animals, not only sheep (scrapie), cows (BSE) and humans (CJD and Kuru), but also rodents, minks (TME), elks (CWD) and probably pigs, chicken and some farmed fish.
2. BSE was first recognized in 1986, whereas Kuru had already been identified in 1957 (not the disease-causing agent itself, but a source where it’s residing in the harmful conformation. More about that below), scrapie is known for at least 200 years.
3. The harmless variant of the prion is called PrPc (Prion Particle cellular), the harmful one PrPsc (from scrapie).
4. There are no chemical differences found between the two variants. However, a difference in enzyme susceptibility has been found: plasminogen is the first endogenous factor discriminating between the normal and pathological prions. On the nerve cells, the molecules that help turn plasminogen into plasmin are localized on the same specialized patches of membrane as normal PrP. Now, isn’t that convenient? (FYI: plasminogen is a pro-protease that has a function related to neuronal exitotoxicity, it is the inactive form of plasmin (and plasmin is thought to allows synapses to remodel themselves, a crucial process for thought and memory). See also points II and A.). Further, there’s this kinetic difficult accessible but thermodynamic stable PrPsc resulting from PrPc (see write-up Halcyon&on).
5. There are two ways to get the prion disease:
  • Inheritable: about 10-15% of the CJD cases because one (or a few) out of the 750 codons is different as a result of a point mutation in the DNA: Leucine is substituted for Proline. They’ve found 18 of those mutations up till now. Another important codon appears to be nr 129, building in Methionine into the PrP-chain instead of Valine. About 37% is so-called homozygote for this codon 129, meaning that both helices need to have this defect (“MM”). At first they thought homozygosity was required for developing the disease, but recent results show that heterozygotes will die too, the only difference is the longer incubation period. 51% of the population is heterozygote (MV, one strand coding for the Methionine and one for Valine).
    Reduced PrPc-gene expression delays the onset of the disease, whereas absence of the PrP gene makes the tested knockout mice fully resistant to infection. So production of PrPc is required for the PrPc to turn into PrPsc. On the other hand, excessive PrP production results in destruction of muscles and peripheral nerves.
  • Communicable: iatrogenic (e.g. via corneal transplantation, human growth hormone medicines), animal feed, food, and in some species like sheep also via blood.
6. The infections affect different species in different ways. The result can be no symptoms, but being able to pass on the prions to other animals who in turn get sick (might be the case with pigs and chicken, not tested yet). When sheep eat BSE-contaminated feed, they develop a disease clinically similar to scrapie. But when their meat is fed to mice, the mice develop BSE symptoms. Further, BSE tends to affect the brain and neurons primarily, whereas the PrPsc in sheep can be found all over the body.

What they suspect
I. Some prions cause disease quickly, while other prions (in the same species) have a longer incubation period. There is a tendency to believe that this may be dependent on the amino acid sequence: with one or a few other amino acids built in into the chain, one (or some) crucial parts of the prion may, or may not, be less stable hence more susceptible to changing the á-helix into the ß-sheet (i.e. PrPc into the harmful PrPsc). Tests were conducted with a PrP hybrid built of mice PrP-gene code and flanking human PrP code, built into and expressed in mice. A hybrid protein was produced. Then brain tissue of patients who died of CJD was introduced into the mice. Weird stuff was, that the transgenic mice became ill much more frequently and faster than did mice carrying a full human PrP gene (diverges from mice PrP at 28 position). This suggest that similarity in the supposed active region is important, as well as a “chaperonemolecule normally involved in folding nascent protein chains, recognized one or both mouse-derived regions of the PrPc.
II. PrPc is predominantly found on the surface of neurons, attached by a glycoinositol phospholipid anchor. What is its purpose there? (See also 4)
III. Ingested prions may be absorbed across the gut wall at Peyers patches. The resident bacteria may be facilitating a protective immune response (but others note that the white blood cells don’t respond to prions at all). The lymphoid cells take up the prions by phagocytose, so that the prions can travel to lymphoid nodes, spleen and tonsils and probably replicate there. “Eventually they gain access to a nerve” (How?) and can propagate or travel via the axon and spinal cord up to the brain. Either glial cells or neural cells can propagate the disease independently. And the fact that intercerebral injection of PrPsc alone doesn’t cause pathology means that “cells must be making PrP for a pathological result”. I don’t think that’s a good conclusion, instead of “must” it should be at least “may” (and see point II of this section): the fact that there’s no known physiological function doesn’t mean that it doesn’t exist. If you don’t know where, what and how exactly to test (as is the situation trying to solve the mystery around prions; “standard” infectious particles are normally micro-organisms or viruses, at least some DNA or RNA involved, never a parasiting protein) it is not likely you’re going to find it easily.
IV. Results tend to be in the direction that the conformation change apparently happens on a membrane in the cell interior. Apparently in neurons. They accumulate in the lysosomes in the cell, which will burst and then infect other cells. The plaques mentioned in the previous write-up are not always observed, nor that it disturbs intracellular processes per se. See also next point V.
V. There is a suspicion of the existence of prion “strains”, multiple possible conformations of prions. This is based on the idea: hey, we’ve found two conformations, if there are two, it makes sense if there are more, and that would explain the different results seen (disease symptoms, location of the prions in the body.)

What they really don’t know
(or I just couldn’t find the answer)
A. All research seems to be directed towards the harm the prions can do, but the PrPc variant is produced in a lot of animals, which indicates that the gene sequence is evolutionary old. Does this mean that PrPc is also beneficial to the body? Why is it produced in the first place? Involved in synaptic functions? (See points II and 4)
B. After transcription and translation of the PrPc, there are extensive post-translational modification (mainly conformational changes) made to the protein, meaning that the protein is “advanced and evolved”. How does that happen? What exactly is changed, and what is the sequence of these events?
C. Pigs, chicken, sheep and farmed fish all have been exposed to BSE-contaminated feed. But what is the difference in the DNA and tertiary structure of the prions? And I don’t mean the answer that there are 30 different positions between bovine and human prions, or the 7 between sheep and cows, I know that. But some parts of the protein are less important for functioning than others. Are those differences in supposed active sites of the prion? The disease pattern in humans tend to be more similar to sheep, so do sheep infect humans more easily? (Side note: currently there doesn’t seem to be a “consistent correlation” between countries herding and eating a lot of sheep and CJD, but see also next point) And if the transmission pattern thought to be similar, what about blood transfusions? (This is being investigated now)
D. If the sheep fed with BSE-contaminated meat can pass on their prions to mice, can they do that to humans too? And if yes, are the symptoms similar to scrapie, or BSE, or something else we don’t know yet? I don’t want to be highly suggestive, but take a disease like Alzheimer: physiologically the brain gets porous in a relatively similar way like CJD, leading to the question: Are they both related to prions or prion-like molecules?
E. PrPsc catalyses the conformation (only? And alone?) change from the á-helix in PrPc into the ß-sheet in PrPsc. But initially only “harmless” PrP is produced. There must have been a first flipping prion. Produced by the body itself, e.g. that the post-translational machinery doesn’t work properly anymore? Was the protein PrPc getting “old” and changed conformation? To which extend is it possible for a PrPsc from say a cow to induce the flipping of PrPc in humans (or whatever other test animal) (see point I)?
F. They don’t know the process during the conformation change.


Still a lot of uncertainties, like with most natural science research. But that’s also its charm… well, for people who like to dig into the topic of course. If anyone knows more, or even has answers on the above questions, please node them.


Most information is from articles from New Scientist, Scientific American and Nature. A well-written resource that also provides insight in “their search for truth” is the article at http://www.nmia.com/~mdibble/prion.html. The Alchemist informed me about http://www.mad-cow.org/, that info probably will be added in the near future.

Other E2 nodes related to this topic: Kuru, mad cow disease, spongiform encephalitis, scrapie, scrapies, BSE, bovine spongiform encephalopathy, Creutzfeld-Jacob disease, CJD, Creutzfeld Jacob disease (CJD), mad pig disease.

To elaborate on Halcyon&on’s good but potentially misleading write-up, it should be pointed out that not all victims of prion diseases are found to have the plaque deposits in their brains. When these amyloid plaques were first discovered, they were thought to be the cause of death, but this theory was quickly discarded when it was established that only about 15% of all CJD victims had them to a severe degree. Subsequently, it became apparent that there was an inverse relation between the extent of the plaques and the infectiousness of the brain tissue found in post-mortem samples. Furthermore, those with the most severe spongiform encephalopathy were the least likely to have plaque deposits.

These plaques consist of PrP27-30, a fragmented form of the abnormal prion mutation (known as PrPsc). The term PrP, which refers to the abormal protein, stands for protease-resistant protein, meaning that the proteolytic enzyme Protease K has no effect on it. However, where these plaques are concerned, this apparently isn’t the case. The presence of these plaques indicate that the Protease K enzyme has had some effect on the PrPsc, reducing it to the smaller, less infectious PrP27-30, which has the tendency to accumulate in amyloid plaques. These plaques are found most commonly among victims of genetically inherited prion diseases, and are in many cases believed to be part of a sort of coping mechanism, actually enabling the patient to live longer while suffering from the disease. Post-mortem analysis of these plaques have shown that they often consist of even smaller protein fragments than PrP27-30, such as PrP11, which has a molecular weight of 11 kilodaltons.

CJD variants, such as Gerstmann-Straussler-Scheinker disease, which can take anywhere from 1 to 15 years to kill its victims, is always indicated by the presence of PrP plaques, but not always by spongiform encephalopathy. By the same token, non-variant CJD lasts around 3-12 months, and spongiform encephalopathy is always present, while PrP plaques are less common. In cases such as the former, the cause is usually famillial.

The concept of an infectious protein, or "prion", was first proposed in relationship to scrapie in sheep (Prusiner, 1982) as a "protein-dependent protein synthesis" model for the cause of the disease. That is, it was thought possible that a unique protein mediated and, in theory, performed its own peptide synthesis, independent of DNA or RNA. In order to be considered infectious, the protein would have to be capable of both transmission of itself (release) and regeneration (Wickner et al., 2002); therefore, the protein must encode itself, enhance its own replication, or affect its post-synthetic structure. Griffith (1967), mindful of the fundamentals of molecular biology, proposed that since proteins do not encode themselves, the scrapie-causing protein must either enhance its own transcription, or have an altered conformation that forces reconformation of the normal cellular protein into the infectious form. Current hypotheses follow the "altered conformation" concept, specifically that the infectious prion protein can mediate reconformation of native protein into the infectious form (Prusiner, 1998).

The term "prion", while first used to describe the proteinaceous infectious agent causing scrapie, was later used in association with other mammalian transmissible spongiform encephalopathies (TSEs), such as Mad Cow Disease (bovine spongiform encephalopathy, or BSE), Creutzfeldt-Jakob disease (CJD), and kuru (Prusiner, 1998). Later studies identified and confirmed the presence of several prions of various yeasts, including [PSI+], [URE3], and [PIN+]** of Saccharomyces cerevisiae and [Het-s] of Podospora anserina (Wickner et al., 2002).


It is worth noting here that much more is known about yeast prions than about about mammalian prions. The reason for this is somewhat obvious -- the mammalian life cycle is rather long, whereas yeasts produce mitotic progeny in a matter of hours and sexual progeny in a matter of days. Therefore, yeasts are used as model organisms for a number of mammalian disorders to understand the molecular biology and biochemistry of various phenomena; the discovery of yeast prions has led to a greater understanding of the prion mode of action.

Yeast prion modes of action have been well-characterized (Wickner et al., 2002). For example, Ure2p** is a protein involved in the a pathway that signals presence of good nitrogen sources, thereby blocking the uptake of allantoic acid (a poor nitrogen source) and a similar molecule, ureidosuccinic acid (USA), by Dal5p. [URE3] is an altered form of native Ure2p; this altered protein is incapable of blocking the uptake of USA, even in the presence of good nitrogen sources. This phenotype is identical to the mutated form of the ure2 gene, but ure2** strains are incapable of forming the [URE3] prion protein.

By contrast, human and mammalian prion models have been rather poorly characterized, mostly due to lack of data surrounding the phenomenon. They were originally described (in 1954) as "slow viruses", given the lack of evidence for any bacterial transmission factor or genetic predisposition to the diseases. Inability to uncover a viral factor and advances in molecular biology later led to the posing of the prion hypothesis in the early 1980s, which is still in the process of building into a theory (covered later).

Much of the frustration found in exploration of mammalian prions stems from the lack of an effective model for most of the diseases. For obvious reasons, it is implausible to examine the steps involved in inducing prion formation in humans, at least in vivo; ethical considerations often slow progress in large animal research. The obvious answer to this conundrum would be a murine model system; after all, mice share approximately 80% or so homology with humans on a genetic basis, and mouse models are effective in studies of hundreds of other diseases, including cancers and genetic disorders. However, a mouse model strain has not yet been developed that can mimic more than small portions of prion disease progression. So far, those small pieces have presented an interesting puzzle; however, until more can be seen together, it seems there will still be questions.

Incidentally, possibly the most vexing piece of the prion puzzle is the simple fact that scientists have not yet found a function for the mammalian prion protein, or PrP, in its native form (PrPc). In fact, very little is known about the function of the gene encoding the protein, except that it is consitutively expressed in adults and heavily regulated during development. Therefore, exploration the mechanism of attack of PrPSc (the altered form) has been largely the result of guesswork.


Because of the nature of prions, Wickner et al. (2002) have outlined three criteria for the definition of a prion:
  1. Reversible curing: The prion phenomenon must respond to some form of curing; when the curing stimulus is removed, a measurable rate of reversion to prion state should occur. [URE3], [PSI+], and [PIN+] are eliminated by guanidine, and appear to depend on the presence of heat shock protein chaperones for their maintenance (reviewed in Wickner et al., 2002).
  2. Overproduction of the parent protein increases the chance of prion protein formation: Overproduction of Ure2p causes increased induction of [URE3], and overproduction of Sup35p increases incidence of [PSI+] (reviewed in Wickner et al., 2002).
  3. Prion-infected phenotype mimics the phenotype of gene mutant: Neither [URE3] nor ure2 strains block uptake of ureidosuccinic acid (USA); [PSI+] and sup35 strains both result in improper translation termination (reviewed in Wickner et al., 2002).
Prions have several other properties, including a tendency to aggregate into amyloid fibers in a heritable, self-propagating fashion (Wickner et al., 2000, 2002; Prusiner, 1998). These proteins that form these aggregates have a region of protease resistance in their amyloid forms; this region of protection is called the prion domain (Wickner et al., 2000, 2002; Prusiner, 1998). This domain occurs at residues 1-65 in [URE3]; 1-114 in [PSI+]; and residues 1-25 in [Het-s] (Wickner et al., 2002).
Stanley Prusiner (1998) compiled a similar list of characteristics of mammalian prions in the guise of a table of arguments for the proteinaceous nature of prions:

    (adapted from Prusiner, 1998)
  1. PrPSc and scrapie infectivity copurify.
  2. "The unusual properties of PrPSc mimic those of prions." Modifications of PrPSc disrupt prion activity.
  3. Levels of PrPSc are directly proportional to prion titers.
  4. No evidence for a viral or nucleic acid source for prion disease has been found.
  5. Accumulation of PrPSc results in prion disease pathology.
  6. PrP genetic mutations result in heritable prion disease and cause formation of PrPSc.
  7. Overexpression of PrPc increases incidence of PrPSc (See #2 of Wickner's list, above). Loss of the PrP gene by knockout results in an inability to produce PrPSc (see description of [URE3], above).
  8. Special variations in PrP result in lack of crossover between certain species in reference to prion disease. For example, scrapie's varied conformation relative to CJD results (possibly) in the inability of humans to get CJD from scrapie-infected sheep.
  9. PrPSc (mutant) preferentially binds PrPc (native), which yields more PrPSc (see definition of prion, above).
  10. Truncations of PrP and chimeric PrP genes alter special infectivity and result in novel prions.
  11. Diversity of prion disease is caused by various conformations of PrPSc. This variation can be generated by passage of PrPSc through different species. For example, BSE may be capable of being passed to humans and converted into vCJD, a more recently discovered version of CJD. (
  12. Strain propagation can be seen in instances of exposure of model mice to two different variants of PrPSc (the paper lists fCJD and FFI, see below), which results in different disease properties in the mice.
While some of these examples seem self-referential ("well, levels of prion protein correlate to prion titer"), the fundamental principle is sound: it is the alterations in the protein itself, not in the DNA or RNA intermediates, that results in the symptoms of prion disease.

Human prions have been shown numerous times to form amyloid fibers (reviewed in Prusiner, 1998). This fiber formation takes on an apparently random shape and size, distinguishing it from viral formation (which always follows a particular pattern); however, this pattern is comparable to such disorders as Alzheimer's disease. Amyloid aggregation is also thought to be the root cause of plaque formation in the brain, which is a major defining symptom of most prion diseases. Along this line, anti-prion drugs and anti-Alzheimer's drugs are being co-examined for cross-effectiveness.


On the conformational variation of PrPSc relative to PrPc:

The native form of PrP consists of approximately 40% alpha-helices and very few beta-sheet formations. Alpha helices are smaller, fairly flexible, and usually more susceptible to proteases (enzymes that cut other proteins) because of their relatively loose shape. The infectious form of PrP, however, consists of up to 45% beta-sheets and up to 30% alpha-helices. These beta-sheets are more solid, usually larger, and less susceptible to protease activity because of the inaccessibility of the peptide backbone. The region of amyloid formation, or "prion domain", usually falls in one of the larger regions of beta-sheet formation. (Prusiner (1998) remarks on the seeming impossibility of such a huge variation in proteins of the same sequence, but alpha-helices and beta-sheets are formed in the backbone, somewhat regardless of the sequence of the R-groups.) As reiterated exhaustively already, these conformational changes are the cause of the infectivity of PrP.


Having spent all this time on the genetics and molecular biology of prions, you might have thought I'd forgotten about the actual physical manifestations of mammalian prion disease. Well, you were wrong.

Mammalian prion diseases affect the brain and neural tissue; because of their small size, they are easily able to pass the blood-brain barrier (a feat not achieved by most bacteria and viruses). Therefore, prion infections directly result in various forms of neural degeneration. Most, but not all, humans affected with prion disease develop dementia, or impairment of brain function (usually associated with cognitive functions). Some develop ataxia, or the gradual loss of movement and coordination. Most, but not all, show evidence of spongiform degeneration upon autopsy; that is, their brains appear outwardly normal, but show necrotic pockets and a softer, porous, almost squishy texture as a result of loss of brain cells. Plaque formation can also occur as a result of amyloid aggregation; this will often occur along the axons of neurons, which may make them feel more solid than they already are.

Current therapies for prion disease are elusive, and consist largely of easing pain and alleviating minor demential symptoms. Quinacrine (FDA approved for treatment of malaria, a disease caused by a pathogen which also passes the blood-brain barrier), chlorpromazine (Thorazine, FDA approved for treatment of schizophrenia), and some of their derivatives have shown promising effects in vitro against the propagation of PrPSc (Korth et al., 2001). However, further investigation is pending.


Referential Information

For what it's worth, there are no known prions of other animals, plants, fungi, protists, or bacteria. Yet.

A list of known and suspected prions, their hosts, and methods of invasion (adapted partially from Prusiner, 1998):

** -- a crash course in genetic nomenclature:
-- all names of genetic elements, whether genomic or epigenetic, are italicized.
-- gene names are always lowercase letters, followed by a number. -- proteins are named as the gene name, with the first letter capitalized. The "p" following the number indicates "protein", and the name is not italicized. -- brackets indicate genetic elements that are not encoded by the nuclear genome (epigenetic elements).

Sources:

If you never read anything else on prions, read Stanley Prusiner's Nobel Prize speech (Prusiner, 1998). This man did much of the definitive work (such as it is), and is a brilliant scientist and orator. It's really heavy reading in parts, and even I didn't bother reading everything, but if ever you needed an overview, his 20-page abridged lecture, published by PNAS, is all you could possibly want.

Much of the heavy yeast content of this node came from a section of the Introduction to my master's thesis, 'Analysis of the [KIL-d] Phenomenon of Killer Virus of Saccharomyces cerevisiae (Rutgers University, October 2002).

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