| Prion diseases | |
|---|---|
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| Microscopic "holes" are characteristic in prion-affected tissue sections, causing the tissue to develop a "spongy" architecture. This causes deterioration of that "spongy" tissue in the brain. | |
| Specialty | Infectious disease |
Prions are misfolded proteins with the ability to transmit their misfolded shape onto normal variants of the same protein. They characterize several fatal and transmissible neurodegenerative diseases in humans and many other animals. It is not known what causes the normal protein to misfold; the abnormal three-dimensional structure is suspected of conferring infectious properties, collapsing nearby protein molecules into the same shape. The word prion derives from "proteinaceous infectious particle". The hypothesized role of a protein as an infectious agent stands in contrast to all other known infectious agents such as viruses, bacteria, fungi and parasites, all of which contain nucleic acids (DNA, RNA or both).
Prion variants of the prion protein (PrP), whose specific function is uncertain, are hypothesized as the cause of transmissible spongiform encephalopathies (TSEs), including scrapie in sheep, chronic wasting disease (CWD) in deer, bovine spongiform encephalopathy (BSE) in cattle (commonly known as "mad cow disease") and Creutzfeldt–Jakob disease (CJD) in humans. All known prion diseases in mammals affect the structure of the brain or other neural tissue; all are progressive, have no known effective treatment and are always fatal. Until 2015, all known mammalian prion diseases were considered to be caused by the prion protein (PrP), however in 2015 multiple system atrophy (MSA) was found to be transmissible and was hypothesized to be caused by a prion form of alpha-synuclein.
Prions form abnormal aggregates of proteins called amyloids, which accumulate in infected tissue and are associated with tissue damage and cell death. Amyloids are also responsible for several other neurodegenerative diseases such as Alzheimer's disease and Parkinson's disease. Prion aggregates are stable, and this structural stability means that prions are resistant to denaturation by chemical and physical agents: they cannot be destroyed by ordinary disinfection or cooking. This makes disposal and containment of these particles difficult.
A prion disease is a type of proteopathy, or disease of structurally abnormal proteins. In humans, prions are believed to be the cause of Creutzfeldt–Jakob disease (CJD), its variant (vCJD), Gerstmann–Sträussler–Scheinker syndrome (GSS), fatal familial insomnia (FFI) and kuru.[2] There is also evidence suggesting prions may play a part in the process of Alzheimer’s disease, Parkinson's disease and amyotrophic lateral sclerosis (ALS), and these have been termed prion-like diseases. Several yeast proteins have also been identified as having prionogenic properties. Prion replication is subject to epimutation and natural selection just as for other forms of replication, and their structure varies slightly between species.
Recent scientific observations show the need to refine the prion hypothesis. Synthetic prions, created in the laboratory independent of any biological source, have little or no ability to cause infection with TSEs; however, when synthetic prions are administered in combination with cofactors, such as phosphatidylethanolamine and RNA molecules, this can transmit TSEs. It has also been shown that scrapie and Creutzfeldt–Jakob disease may require agent-specific nucleic acids for transmission of infection. Most recently, it was shown that mice with severe combined immunodeficiency do not develop scrapie following inoculation with brain tissue from animals infected with scrapie, suggesting that either the role of immunity in prion pathogenesis is incompletely understood or that there is some other flaw in current understanding of prion pathophysiology.
Prion protein
Discovery
In the 1950's, Carleton Gajdusek began research which eventually showed that kuru could be transmitted to chimpanzees by what was possibly a new infectious agent, work for which he eventually won the 1976 Nobel prize. During the 1960s, two London-based researchers, radiation biologist Tikvah Alper and biophysicist John Stanley Griffith, developed the hypothesis that the transmissible spongiform encephalopathies are caused by an infectious agent consisting solely of proteins. Earlier investigations by E.J. Field into scrapie
and kuru had found evidence for the transfer of pathologically inert
polysaccharides that only become infectious post-transfer, in the new
host. Alper and Griffith wanted to account for the discovery that the mysterious infectious agent causing the diseases scrapie and Creutzfeldt–Jakob disease resisted ionizing radiation.
(A single ionizing "hit" normally destroys an entire infectious
particle, and the dose needed to hit half the particles depends on the
size of the particles. Empirical results of ionizing doses applied to
the unknown infectious substance evidenced an infectious particle size
too small to be a viral mechanism.) In his paper, entitled
"Self-replication and Scrapie", Griffith proposed three ways in which a
protein could be a pathogen. In the first hypothesis, he suggested that if the protein is the product of a normally suppressed gene,
and introducing the protein could induce the gene's expression, that
is, wake the dormant gene up, then the result would be a process
indistinguishable from replication, as the gene's expression would
produce the protein, which would then go wake the gene up in other cells.
His second hypothesis forms the basis of the modern prion theory, and
proposed that an abnormal form of a cellular protein can convert normal
proteins of the same type into its abnormal form, thus leading to
replication. His third hypothesis proposed that the agent could be an antibody if the antibody was its own target antigen,
as such an antibody would result in more and more antibody being
produced against itself. However, Griffith acknowledged that this third
hypothesis was unlikely to be true due to the lack of a detectable immune response.
Francis Crick
recognized the potential significance of the Griffith protein-only
hypothesis for scrapie propagation in the second edition of his "Central dogma of molecular biology"
(1970): While asserting that the flow of sequence information from
protein to protein, or from protein to RNA and DNA was "precluded", he
noted that Griffith's hypothesis was a potential contradiction (although
it was not so promoted by Griffith). The revised hypothesis was later formulated, in part, to accommodate reverse transcription (which both Howard Temin and David Baltimore discovered in 1970).
In 1982, Stanley B. Prusiner of the University of California, San Francisco
announced that his team had purified the hypothetical infectious
protein, which did not appear to be present in healthy hosts, though
they did not manage to isolate the protein until two years after
Prusiner's announcement.[28][29] The protein was named a prion, for "Proteinacious infectious particle", taken from the words protein and infection.
When the prion was discovered, Griffith's first hypothesis, that the
protein was the product of a normally silent gene was favored by many.
It was subsequently discovered, however, that the same protein
exists in normal hosts but in different form.
Following the discovery of the same protein in different form in
uninfected individuals, the specific protein that the prion was composed
of was named the Prion Protein (PrP), and Griffith's second hypothesis
that an abnormal form of a host protein can convert other proteins of
the same type into its abnormal form, became the dominant theory. Prusiner won the Nobel Prize in Physiology or Medicine in 1997 for his research into prions.
Structure
The protein that prions are made of (PrP) is found throughout the
body, even in healthy people and animals. However, PrP found in
infectious material has a different structure and is resistant to proteases, the enzymes in the body that can normally break down proteins. The normal form of the protein is called PrPC, while the infectious form is called PrPSc – the C refers to 'cellular' PrP, while the Sc refers to 'scrapie', the prototypic prion disease, occurring in sheep. While PrPC is structurally well-defined, PrPSc is certainly polydisperse
and defined at a relatively poor level. PrP can be induced to fold into
other more-or-less well-defined isoforms in vitro, and their
relationship to the form(s) that are pathogenic in vivo is not yet
clear.
PrPC
PrPC is a normal protein found on the membranes of cells. It has 209 amino acids (in humans), one disulfide bond, a molecular mass of 35–36 kDa and a mainly alpha-helical structure. Several topological forms exist; one cell surface form anchored via glycolipid and two transmembrane forms. The normal protein is not sedimentable; meaning that it cannot be separated by centrifuging techniques. Its function is a complex issue that continues to be investigated. PrPC binds copper (II) ions with high affinity. The significance of this finding is not clear, but it is presumed to relate to PrP structure or function. PrPC is readily digested by proteinase K and can be liberated from the cell surface in vitro by the enzyme phosphoinositide phospholipase C (PI-PLC), which cleaves the glycophosphatidylinositol (GPI) glycolipid anchor. PrP has been reported to play important roles in cell-cell adhesion and intracellular signaling in vivo, and may therefore be involved in cell-cell communication in the brain.
PrPres
Protease-resistant PrPSc-like protein (PrPres) is the name given to any isoform of PrPc which is structurally altered and converted into a misfolded proteinase K-resistant form in vitro. To model conversion of PrPC to PrPSc in vitro, Saborio et al. rapidly converted PrPC into a PrPres by a procedure involving cyclic amplification of protein misfolding. The term "PrPres" has been made to distinguish between PrPSc, which is isolated from infectious tissue and associated with the transmissible spongiform encephalopathy agent. For example, unlike PrPSc, PrPres may not necessarily be infectious.
PrPSc
The infectious isoform of PrP, known as PrPSc, or simply the prion, is able to convert normal PrPC proteins into the infectious isoform by changing their conformation, or shape; this, in turn, alters the way the proteins interconnect. PrPSc always causes prion disease. Although the exact 3D structure of PrPSc is not known, it has a higher proportion of β-sheet structure in place of the normal α-helix structure. Aggregations of these abnormal isoforms form highly structured amyloid
fibers, which accumulate to form plaques. It is unclear as to whether
these aggregates are the cause of cell damage or are simply a
side-effect of the underlying disease process.
The end of each fiber acts as a template onto which free protein
molecules may attach, allowing the fiber to grow. Under most
circumstances, only PrP molecules with an identical amino acid sequence
to the infectious PrPSc are incorporated into the growing fiber. However, rare cross-species transmission is also possible.
Normal function PrP
The
physiological function of the prion protein remains poorly understood.
While data from in vitro experiments suggest many dissimilar roles,
studies on PrP knockout mice
have provided only limited information because these animals exhibit
only minor abnormalities. In research done in mice, it was found that
the cleavage of PrP proteins in peripheral nerves causes the activation
of myelin repair in Schwann cells and that the lack of PrP proteins caused demyelination in those cells.
PrP and regulated cell death
MAVS,
RIP1, and RIP3 are prion-like proteins found in other parts of the
body. They also polymerise into filamentous amyloid fibers which
initiate regulated cell death in the case of a viral infection to
prevent the spread of virions to other, surrounding cells.
PrP and long-term memory
A review of evidence in 2005 suggested that PrP may have a normal function in maintenance of long-term memory. As well, a 2004 study found that mice lacking genes for normal cellular PrP protein show altered hippocampal long-term potentiation.
A recent study that might explain why this is found that neuronal
protein CPEB has a similar genetic sequence to yeast prion proteins. The
prion-like formation of CPEB is essential for maintaining long-term
synaptic changes associated with long term memory formation.
PrP and stem cell renewal
A
2006 article from the Whitehead Institute for Biomedical Research
indicates that PrP expression on stem cells is necessary for an
organism's self-renewal of bone marrow. The study showed that all long-term hematopoietic stem cells
express PrP on their cell membrane and that hematopoietic tissues with
PrP-null stem cells exhibit increased sensitivity to cell depletion.
PrP and innate immunity
There is some evidence that PrP may play a role in innate immunity,
as the expression of PRNP, the PrP gene, is upregulated in many viral
infections and PrP has antiviral properties against many viruses,
including HIV.
Prion replication mechanism
Heterodimer model of prion propagation
Fibril model of prion propagation.
The first hypothesis that tried to explain how prions replicate in a protein-only manner was the heterodimer model. This model assumed that a single PrPSc molecule binds to a single PrPC molecule and catalyzes its conversion into PrPSc. The two PrPSc molecules then come apart and can go on to convert more PrPC.
However, a model of prion replication must explain both how prions
propagate, and why their spontaneous appearance is so rare. Manfred Eigen showed that the heterodimer model requires PrPSc to be an extraordinarily effective catalyst, increasing the rate of the conversion reaction by a factor of around 1015. This problem does not arise if PrPSc exists only in aggregated forms such as amyloid, where cooperativity may act as a barrier to spontaneous conversion. What is more, despite considerable effort, infectious monomeric PrPSc has never been isolated.
An alternative model assumes that PrPSc exists only as fibrils, and that fibril ends bind PrPC and convert it into PrPSc. If this were all, then the quantity of prions would increase linearly, forming ever longer fibrils. But exponential growth of both PrPSc and of the quantity of infectious particles is observed during prion disease. This can be explained by taking into account fibril breakage.
A mathematical solution for the exponential growth rate resulting from
the combination of fibril growth and fibril breakage has been found. The exponential growth rate depends largely on the square root of the PrPC concentration. The incubation period is determined by the exponential growth rate, and in vivo data on prion diseases in transgenic mice match this prediction. The same square root dependence is also seen in vitro in experiments with a variety of different amyloid proteins.
The mechanism of prion replication has implications for designing
drugs. Since the incubation period of prion diseases is so long, an
effective drug does not need to eliminate all prions, but simply needs
to slow down the rate of exponential growth. Models predict that the
most effective way to achieve this, using a drug with the lowest
possible dose, is to find a drug that binds to fibril ends and blocks
them from growing any further.
Prion diseases and their transmission properties
Until 2015 all known mammalian prion diseases were considered to be caused by the prion protein, PrP; in 2015 multiple system atrophy was found to be transmissible and was hypothesized to be caused by a new prion, the misfolded form of a protein called alpha-synuclein. The endogenous, properly folded form of the prion protein is denoted PrPC (for Common or Cellular), whereas the disease-linked, misfolded form is denoted PrPSc (for Scrapie), after one of the diseases first linked to prions and neurodegeneration. The precise structure of the prion is not known, though they can be formed by combining PrPC, polyadenylic acid, and lipids in a protein misfolding cyclic amplification (PMCA) reaction. Proteins showing prion-type behavior are also found in some fungi, which has been useful in helping to understand mammalian prions. Fungal prions do not appear to cause disease in their hosts.
Prions cause neurodegenerative disease by aggregating extracellularly within the central nervous system to form plaques known as amyloid, which disrupt the normal tissue structure. This disruption is characterized by "holes" in the tissue with resultant spongy architecture due to the vacuole formation in the neurons. Other histological changes include astrogliosis and the absence of an inflammatory reaction. While the incubation period
for prion diseases is relatively long (5 to 20 years), once symptoms
appear the disease progresses rapidly, leading to brain damage and
death. Neurodegenerative symptoms can include convulsions, dementia, ataxia (balance and coordination dysfunction), and behavioural or personality changes.
All known prion diseases are untreatable and fatal. However, a vaccine developed in mice may provide insight into providing a vaccine to resist prion infections in humans.
Additionally, in 2006 scientists announced that they had genetically
engineered cattle lacking a necessary gene for prion production – thus
theoretically making them immune to BSE,
building on research indicating that mice lacking normally occurring
prion protein are resistant to infection by scrapie prion protein.
In 2013, a study revealed that 1 in 2,000 people in the United Kingdom
might harbour the infectious prion protein that causes vCJD.
Many different mammalian species can be affected by prion diseases, as the prion protein (PrP) is very similar in all mammals.
Due to small differences in PrP between different species it is unusual
for a prion disease to transmit from one species to another. The human
prion disease variant Creutzfeldt–Jakob disease, however, is thought to
be caused by a prion that typically infects cattle, causing bovine spongiform encephalopathy and is transmitted through infected meat.
Transmission
It has been recognized that prion diseases can arise in three different ways: acquired, familial, or sporadic.
It is often assumed that the diseased form directly interacts with the
normal form to make it rearrange its structure. One idea, the "Protein
X" hypothesis, is that an as-yet unidentified cellular protein (Protein
X) enables the conversion of PrPC to PrPSc by bringing a molecule of each of the two together into a complex.
Current research suggests that the primary method of infection in
animals is through ingestion. It is thought that prions may be
deposited in the environment through the remains of dead animals and via
urine, saliva, and other body fluids. They may then linger in the soil
by binding to clay and other minerals.
A University of California research team, led by Nobel Prize winner Stanley Prusiner, has provided evidence for the theory that infection can occur from prions in manure.
And, since manure is present in many areas surrounding water
reservoirs, as well as used on many crop fields, it raises the
possibility of widespread transmission. It was reported in January 2011
that researchers had discovered prions spreading through airborne
transmission on aerosol particles, in an animal testing experiment focusing on scrapie infection in laboratory mice. Preliminary evidence supporting the notion that prions can be transmitted through use of urine-derived human menopausal gonadotropin, administered for the treatment of infertility, was published in 2011.
Prions in plants
In 2015, researchers at The University of Texas Health Science Center at Houston
found that plants can be a vector for prions. When researchers fed
hamsters grass that grew on ground where a deer that died with chronic wasting disease
(CWD) was buried, the hamsters became ill with CWD, suggesting that
prions can bind to plants, which then take them up into the leaf and
stem structure, where they can be eaten by herbivores, thus completing
the cycle. It is thus possible that there is a progressively
accumulating number of prions in the environment.
Sterilization
Infectious particles possessing nucleic acid
are dependent upon it to direct their continued replication. Prions,
however, are infectious by their effect on normal versions of the
protein. Sterilizing prions, therefore, requires the denaturation
of the protein to a state in which the molecule is no longer able to
induce the abnormal folding of normal proteins. In general, prions are
quite resistant to proteases, heat, ionizing radiation, and formaldehyde treatments, although their infectivity can be reduced by such treatments. Effective prion decontamination relies upon protein hydrolysis or reduction or destruction of protein tertiary structure. Examples include sodium hypochlorite, sodium hydroxide, and strongly acidic detergents such as LpH. 134 °C (274 °F) for 18 minutes in a pressurized steam autoclave has been found to be somewhat effective in deactivating the agent of disease. Ozone sterilization is currently being studied as a potential method for prion denaturation and deactivation.
Renaturation of a completely denatured prion to infectious status has
not yet been achieved; however, partially denatured prions can be
renatured to an infective status under certain artificial conditions.
The World Health Organization
recommends any of the following three procedures for the sterilization
of all heat-resistant surgical instruments to ensure that they are not
contaminated with prions:
- Immerse in 1N sodium hydroxide and place in a gravity-displacement autoclave at 121 °C for 30 minutes; clean; rinse in water; and then perform routine sterilization processes.
- Immerse in 1N sodium hypochlorite (20,000 parts per million available chlorine) for 1 hour; transfer instruments to water; heat in a gravity-displacement autoclave at 121 °C for 1 hour; clean; and then perform routine sterilization processes.
- Immerse in 1N sodium hydroxide or sodium hypochlorite (20,000 parts per million available chlorine) for 1 hour; remove and rinse in water, then transfer to an open pan and heat in a gravity-displacement (121 °C) or in a porous-load (134 °C) autoclave for 1 hour; clean; and then perform routine sterilization processes.
Fungi
In yeast, protein refolding to the prion configuration is assisted by chaperone proteins such as Hsp104. All known prions induce the formation of an amyloid fold, in which the protein polymerises into an aggregate consisting of tightly packed beta sheets.
Amyloid aggregates are fibrils, growing at their ends, and replicate
when breakage causes two growing ends to become four growing ends. The incubation period of prion diseases is determined by the exponential growth rate associated with prion replication, which is a balance between the linear growth and the breakage of aggregates.
Fungal proteins exhibiting templated conformational change were discovered in the yeast Saccharomyces cerevisiae by Reed Wickner in the early 1990s. For their mechanistic similarity to mammalian prions, they were termed yeast prions. Subsequent to this, a prion has also been found in the fungus Podospora anserina. These prions behave similarly to PrP, but, in general, are nontoxic to their hosts. Susan Lindquist's group at the Whitehead Institute
has argued some of the fungal prions are not associated with any
disease state, but may have a useful role; however, researchers at the
NIH have also provided arguments suggesting that fungal prions could be
considered a diseased state.
There is mounting evidence that fungal proteins have evolved specific
functions that are beneficial to the microorganism that enhance their
ability to adapt to their diverse environments.
As of 2012, there are eight known prion proteins in fungi, seven in Saccharomyces cerevisiae (Sup35, Rnq1, Ure2, Swi1, Mot3, Cyc8, and Mod5) and one in Podospora anserina (HET-s).[contradictory]
The article that reported the discovery of a prion form, the Mca1
protein, was retracted due to the fact that the data could not be
reproduced. Notably, most of the fungal prions are based on glutamine/asparagine-rich sequences, with the exception of HET-s and Mod5.
Research into fungal prions
has given strong support to the protein-only concept, since purified
protein extracted from cells with a prion state has been demonstrated to
convert the normal form of the protein into a misfolded form in vitro,
and in the process, preserve the information corresponding to different
strains of the prion state. It has also shed some light on prion
domains, which are regions in a protein that promote the conversion into
a prion. Fungal prions have helped to suggest mechanisms of conversion
that may apply to all prions, though fungal prions appear distinct from
infectious mammalian prions in the lack of cofactor required for
propagation. The characteristic prion domains may vary between species –
e.g., characteristic fungal prion domains are not found in mammalian
prions.
Potential treatments and diagnosis
Advancements
in computer modeling have allowed scientists to identify compounds that
can treat prion-caused diseases, such as one compound found to bind a
cavity in the PrPC and stabilize the conformation, reducing the amount of harmful PrPSc.
Antiprion antibodies capable of crossing the blood-brain-barrier and targeting cytosolic prion protein (an otherwise major obstacle in prion therapeutics) have been described.
In the last decade, some progress dealing with
ultra-high-pressure inactivation of prion infectivity in processed meat
has been reported.
In 2011, it was reported that prions could be degraded by lichens. Astemizole has been found to have anti-prion activity. Another type of chemical that may be effective against prion infection is the luminescent conjugated polythiophenes,
fluorescent compounds that are often used to stain tissue samples. In a
2015 study, it was found that that when they injected mice with a prion
disease and then with polythiophenes, the mice survived 80% longer than
the control mice that were injected only with the prion disease.
There continues to be a practical problem with diagnosis of prion
diseases, including BSE and CJD. They have an incubation period of
months to decades, during which there are no symptoms, even though the
pathway of converting the normal brain PrP protein into the toxic,
disease-related PrPSc form has started. At present, there is virtually no way to detect PrPSc
reliably except by examining the brain using neuropathological and
immunohistochemical methods after death. Accumulation of the abnormally
folded PrPSc form of the PrP protein is a characteristic of
the disease, but it is present at very low levels in easily accessible
body fluids like blood or urine. Researchers have tried to develop
methods to measure PrPSc, but there are still no fully accepted methods for use in materials such as blood.
In 2010, a team from New York described detection of PrPSc even when initially present at only one part in a hundred billion (10−11) in brain tissue. The method combines amplification with a novel technology called surround optical fiber immunoassay (SOFIA) and some specific antibodies against PrPSc. After amplifying and then concentrating any PrPSc,
the samples are labelled with a fluorescent dye using an antibody for
specificity and then finally loaded into a micro-capillary tube. This
tube is placed in a specially constructed apparatus so that it is
totally surrounded by optical fibres to capture all light emitted once
the dye is excited using a laser.
The RT-QuIC assay, a microplate reader-based
prion detection method which uses as reagents normally folded prions,
fluorescently labelled so that they "light up" when they are misfolded;
samples suspected of containing misfolded prions are added and misfolded
reagents can be detected by standard fluorescence detection methods.
The Center for Disease Control and Prevention includes a positive
RT-QuIC result in its diagnostic criteria for the probable diagnosis of
sCJD.
A 2015 study found that a naturally occurring variant of the human prion protein in transgenic mice protected them against kuru and CJD.
SGI-1027 and related compounds were identified as a novel class
of potential anti-prion agents that preferentially function through
direct interaction with PrPC.
Role of prions in transmissible spongiform encephalopathies
The cause of the transmissible spongiform encephalopathies
(TSE) is currently unknown, but the diseases are known to be associated
with prions. Whether prions cause TSEs or are the result of infection
with another agent such as a virus is a matter of debate by a minority
of scientists. The following are some hypotheses.
Prion hypothesis
The
prion hypothesis states that the main component of the TSE agent is
composed of a misfolded protein. The prion hypothesis can be divided
into two sub-hypotheses: the protein-only hypothesis, and the
multi-component hypothesis.
Protein-only hypothesis
Prior to the discovery of prions, it was thought that all pathogens used nucleic acids
to direct their replication. The "protein-only hypothesis" states that a
protein structure can replicate without the use of nucleic acids. This
was initially controversial as it contradicts the central dogma of molecular biology, which describes nucleic acid as the central form of replicative information.
Evidence in favor of a protein-only hypothesis includes:
- Infectivity titre in TSEs roughly correlates with prion amyloid (PrPSc) titre, however, prion amyloid is undetectable in approximately 10% of CJD cases.
- No virus particles, bacteria, or fungi have been conclusively associated with prion diseases, although virus-like particles and Spiroplasma-like inclusions can be detected in some TSE cases, but not in controls (uninfected individuals).
- No nucleic acid has been conclusively associated with infectivity; agent is resistant to ultraviolet radiation and nucleases, although in 2016, studies have suggested that the agent can be destroyed by nucleases under certain situations and that part of the resistance to nucleases and radiation may be that byproducts from degenerating neurons may help protect a nucleic acid.
- No immune or inflammatory response to infection.
- PrPSc experimentally transmitted between one species and another results in PrPSc with the amino-acid sequence of the recipient species, suggesting that nucleic acid-mediated replication of the donor agent does not occur.
- Familial prion disease occurs in families with a mutation in the PrP gene, and mice with PrP mutations develop prion disease despite controlled conditions where transmission is prevented. These mice can then transmit the disease to healthy, wild type mice, suggesting that mice with PrP mutations spontaneously generate infectivity.
- Animals lacking PrPC do not contract prion disease.
Genetic factors
A gene for the normal protein has been identified: the PRNP gene. In all inherited cases of prion disease, there is a mutation in the PRNP gene. Many different PRNP mutations have been identified and these proteins are more likely to fold into abnormal prion.
Although this discovery puts a hole in the general prion hypothesis,
that prions can aggregate only proteins of identical amino acid make-up.
These mutations can occur throughout the gene. Some mutations involve
expansion of the octapeptide repeat region at the N-terminal of PrP.
Other mutations that have been identified as a cause of inherited prion
disease occur at positions 102, 117 & 198 (GSS), 200, 210 & 232
(CJD) and 178 (fatal familial insomnia, FFI). The cause of prion disease can be sporadic, genetic, or infectious, or a combination of these factors. For example, to have scrapie, both an infectious agent and a susceptible genotype must be present.
Multi-component hypothesis
Despite much effort, significant titers
of prion infectivity have never been produced by refolding pure PrP
molecules, raising doubt about the validity of the protein-only
hypothesis. In addition, the protein-only hypothesis fails to provide a
molecular explanation for the ability of prion strains to target
specific areas of the brain in distinct patterns. These shortcomings,
along with additional experimental data, have given rise to the
"multi-component" or "cofactor variation" hypothesis.
In 2007, biochemist Surachai Supattapone and his colleagues at Dartmouth College produced purified infectious prions de novo from defined components (PrPC, co-purified lipids, and a synthetic polyanionic molecule).
These researchers also showed that the polyanionic molecule required
for prion formation was selectively incorporated into high-affinity
complexes with PrP molecules, leading them to hypothesize that
infectious prions may be composed of multiple host components, including
PrP, lipid, and polyanionic molecules, rather than PrPSc alone.
In 2010, Jiyan Ma and colleagues at the Ohio State University
produced infectious prions from a recipe of bacterially expressed
recombinant PrP, POPG phospholipid, and RNA, further supporting the
multi-component hypothesis. This finding is in contrast to studies that found minimally infectious prions produced from recombinant PrP alone.
In 2012, Supattapone and colleagues purified the membrane lipid phosphatidylethanolamine as a solitary endogenous cofactor capable of facilitating the formation of high-titer recombinant prions derived from multiple prion strains. They also reported that the cofactor is essential for maintaining the infectious conformation of PrPSc, and that cofactor molecules dictate the strain properties of infectious prions.
Difficulties associated with the prion hypothesis
The following are some of the current difficulties and challenges:
- Several different types of PrPSc occur in the brains of animals with scrapie. As PrPSc consist only of peptides, there is no known mechanism by which different prion types can occur.
- The mechanism by which the number of PrPSc molecules increases by orders-of-magnitude remains unexplained.
- There has been no satisfactory explanation as to how prion peptides with the same amino acid sequence change their 3-dimensional folding structure from an alpha helix to a beta sheet.
- The presence of damaged neurologic tissue is consistent with other hypotheses besides a prion.
- Inexplicably, mice with severe combined immunodeficiency do not develop scrapie following inoculation with brain tissue from animals infected with scrapie.
Whether prions cause disease or are merely a symptom resulting from a
different agent is still a matter of debate and research. The following
sections describe several hypotheses: some pertain to the composition
of the infectious agent (protein-only, protein with other components,
virus, or other), while others pertain to its mechanism of reproduction.
Heavy metal poisoning hypothesis
Reports suggest that imbalance of brain metal homeostasis may be a cause of PrPSc-associated
neurotoxicity, though the underlying mechanisms are difficult to
explain based on existing information. Proposed hypotheses include a
functional role for PrPC in metal metabolism, and loss of this function due to aggregation to the disease-associated PrPSc form as the cause of brain metal imbalance. Other views suggest gain of toxic function by PrPSc due to sequestration of PrPC-associated metals within the aggregates, resulting in the generation of redox-active PrPSc complexes. The physiological implications of some PrPC-metal interactions are known, while others are still unclear. The pathological implications of PrPC-metal interaction include metal-induced oxidative damage, and in some instances conversion of PrPC to a PrPSc-like form.
Viral hypothesis
The
protein-only hypothesis has been criticised by those maintaining that
the simplest explanation of the evidence to date is viral. For more than a decade, Yale University neuropathologist Laura Manuelidis has been proposing that prion diseases are caused instead by an unidentified slow virus. In January 2007, she and her colleagues published an article reporting to have found a virus in 10%, or less, of their scrapie-infected cells in culture. In 2016, Sotirios Botsios and Laura Manuelidis showed evidence that TSE specific nucleic acids may be required for infectious transmission of CJD and scrapie.
Evidence in favor of a viral hypothesis includes:
- Strain variation: differences in prion infectivity, incubation, symptomology, and progression among species resembles that seen between viruses, especially RNA viruses
- The long incubation and rapid onset of symptoms resembles lentiviruses, such as HIV-induced AIDS
- Viral-like particles that do not appear to be composed of PrP have been found in some of the cells of scrapie- or CJD-infected cell lines.
- Many viruses, including HIV which needs CD4 and CXCR4, need a receptor to attach to and enter into host cells. The host prion, PrPc may be a receptor protein for an as yet undiscovered TSE virus, explaining why animals lacking host prion do not become infected with experimental prion disease.
- A prion-like protein, called MAVS, has been shown to misfold as part of the innate immune response against pathogenic viruses, similarly the cellular prion, PrPC has been shown to have anti HIV properties, and it is hypothesized that the misfolding of the prion in TSEs may be an antiviral response against an unknown virus.
- In 2016, studies have demonstrated susceptibility to nucleases under certain situations: >99% of infectivity was destroyed, but there was no reduction of prion protein, suggesting the presence of a nucleic acid.
Studies propagating TSE infectivity in cell-free reactions and in purified component chemical reactions is thought to strongly suggest against TSE viral nature. However, some viruses, such as poliovirus, have the ability to replicate in cell-free reactions.
Virino hypothesis
The 'virino hypothesis' postulates that the TSE agent is a foreign, self replicating nucleic acid or nucleic acid fragment bound to PrP.
Spiroplasma hypothesis
Spiroplasma is a cell wall–deficient bacterium related to Mycoplasma,
which some think may be the cause of the TSEs. The lack of a cell wall
means it is not susceptible to conventional antibiotics such as penicillin, which target cell wall synthesis. Frank O. Bastian of Louisiana State University first discovered Spiroplasma-like inclusions in the brain of a CJD patient during an autopsy in 1979 and has hypothesized that this bacterium could possibly be the cause of the TSEs.
However, as of 2015, with the exception of Spiroplasma mirum
strain SMCA causing spongiform microcystic encephalitis in suckling
rats, other researchers have been unable to duplicate these findings, casting doubt on the Spiroplasma hypothesis. In defense of the Spiroplasma hypothesis, Bastian pointed out that Spiroplasma is hard to culture and that strain variation makes it hard to detect certain strains using PCR and other techniques, thus giving a false negative.
Acinetobacter-autoimmunity hypothesis
Acinetobacter is a bacterium which some think is the cause of the TSEs. Mainly because some CJD patients produce antibodies against Acinetobacter calcoaceticus.
Role of prions in other diseases
Prion-like
domains have been found in a variety of other mammalian proteins. Some
of these proteins have been implicated in the ontogeny of age-related
neurodegenerative disorders such as amyotrophic lateral sclerosis (ALS) a motor neuron disease, frontotemporal lobar degeneration with ubiquitin-positive inclusions (FTLD-U), Alzheimer's disease and Parkinson's disease, and Huntington's disease. They are also implicated in some forms of systemic amyloidosis including AA amyloidosis that develops in humans and animals with inflammatory and infectious diseases such as tuberculosis, Crohn's disease, rheumatoid arthritis, and HIV AIDS. AA amyloidosis, like prion disease, may be transmissible.[146]
This has given rise to the 'prion paradigm', where otherwise harmless
proteins can be converted to a pathogenic form by a small number of
misfolded, nucleating proteins.
The definition of a prion-like domain arises from the study of
fungal prions. In yeast, prionogenic proteins have a portable prion
domain that is both necessary and sufficient for self-templating and
protein aggregation. This has been shown by attaching the prion domain
to a reporter protein, which then aggregates like a known prion.
Similarly, removing the prion domain from a fungal prion protein
inhibits prionogenesis. This modular view of prion behaviour has led to
the hypothesis that similar prion domains are present in animal
proteins, in addition to PrP.
These fungal prion domains have several characteristic sequence
features. They are typically enriched in asparagine, glutamine, tyrosine
and glycine residues, with an asparagine bias being particularly
conducive to the aggregative property of prions. Historically,
prionogenesis has been seen as independent of sequence and only
dependent on relative residue content. However, this has been shown to
be false, with the spacing of prolines and charged residues having been
shown to be critical in amyloid formation.
Bioinformatic screens have predicted that over 250 human proteins
contain prion-like domains (PrLD). These domains are hypothesized to
have the same transmissible, amyloidogenic properties of PrP and known
fungal proteins. As in yeast, proteins involved in gene expression and
RNA binding seem to be particularly enriched in PrLD's, compared to
other classes of protein. In particular, 29 of the known 210 proteins
with an RNA recognition motif also have a putative prion domain.
Meanwhile, several of these RNA-binding proteins have been independently
identified as pathogenic in cases of ALS, FTLD-U, Alzheimer's disease,
and Huntington's disease.
Role in neurodegenerative disease
The
pathogenicity of prions and proteins with prion-like domains is
hypothesized to arise from their self-templating ability and the
resulting exponential growth of amyloid fibrils. The presence of amyloid
fibrils in patients with degenerative diseases has been well
documented. These amyloid fibrils are seen as the result of pathogenic
proteins that self-propagate and form highly stable, non-functional
aggregates.
While this does not necessarily imply a causal relationship between
amyloid and degenerative diseases, the toxicity of certain amyloid forms
and the overproduction of amyloid in familial cases of degenerative
disorders supports the idea that amyloid formation is generally toxic.
Specifically, aggregation of TDP-43,
an RNA-binding protein, has been found in ALS/MND patients, and
mutations in the genes coding for these proteins have been identified in
familial cases of ALS/MND. These mutations promote the misfolding of
the proteins into a prion-like conformation. The misfolded form of
TDP-43 forms cytoplasmic inclusions in afflicted neurons, and is found
depleted in the nucleus. In addition to ALS/MND and FTLD-U, TDP-43
pathology is a feature of many cases of Alzheimer's disease, Parkinson's
disease and Huntington's disease. The misfolding of TDP-43 is largely
directed by its prion-like domain. This domain is inherently prone to
misfolding, while pathological mutations in TDP-43 have been found to
increase this propensity to misfold, explaining the presence of these
mutations in familial cases of ALS/MND. As in yeast, the prion-like
domain of TDP-43 has been shown to be both necessary and sufficient for
protein misfolding and aggregation.
Similarly, pathogenic mutations have been identified in the
prion-like domains of heterogeneous nuclear riboproteins hnRNPA2B1 and
hnRNPA1 in familial cases of muscle, brain, bone and motor neuron
degeneration. The wild-type form of all of these proteins show a
tendency to self-assemble into amyloid fibrils, while the pathogenic
mutations exacerbate this behaviour and lead to excess accumulation.
Etymology and pronunciation
The word prion, coined in 1982 by Stanley B. Prusiner, is a portmanteau derived from protein and infection, hence prion, and is short for "proteinaceous infectious particle", in reference to its ability to self-propagate and transmit its conformation to other proteins. Its main pronunciation is /ˈpriːɒn/ although /ˈpraɪɒn/, as the homographic name of the bird is pronounced, is also heard. In his 1982 paper introducing the term, Prusiner specified that it be "pronounced pree-on."
