NIAID Scientists Detail First Structure of a Natural Mammalian Prion

The near-atomic structure of a chronic wasting disease (CWD) prion should help scientists explain how CWD prions spread and become the most naturally infectious of the many mammalian protein aggregation diseases. NIAID scientists revealed the structure in a new study in Acta Neuropathologica. Such detailed knowledge could guide the rational design of vaccines and therapeutics, as well as identify mechanisms that protect humans from CWD pathogens in deer, elk, moose, and reindeer.

Many brain diseases of humans and other mammals involve specific proteins (e.g., prion protein or PrP) gathering into abnormal thread-like structures that grow by sticking to normal versions of the same protein. These threads can also fragment and spread throughout the nervous system and accumulate to deadly levels. For unknown reasons, CWD prions are more naturally contagious than most other protein aggregates and are spreading rampantly among cervid species in North America, Korea and northern Europe. Recalling the bovine spongiform encephalopathy (BSE) or “mad cow disease” epidemic of the mid-1980s and mid-1990s, there are concerns that CWD might similarly be transmissible to humans.

To date, no CWD transmission to humans has been substantiated, and the new CWD structure suggests preliminarily why we might be protected. The structure also reveals multiple differences between CWD and previously determined structures of highly infectious, but experimentally rodent-adapted, PrP-based prions. Differences are even more profound when compared to largely non-transmissible PrP filaments isolated from humans with Gerstmann-Sträussler-Scheinker syndrome, a genetic prion disorder.

PrP-based prion diseases are degenerative, untreatable, and fatal diseases of the central nervous system that occur in people and other mammals. These diseases primarily involve the brain, but also can affect the eyes and other organs. CWD-infected animals shed infectious prions in their feces, urine, and other fluids and body components while alive, and from their carcasses after dying. The prions can remain infectious in the environment for years. 

Scientists at NIAID’s Rocky Mountain Laboratories in Hamilton, Montana, determined the CWD structure from the brain tissue from a naturally infected white-tailed deer. They isolated the prions and froze them in glass-like ice. Then, using electron microscopy techniques, they developed a 3-D electron density map that indicated the detailed shapes of the protein molecules within the prion structure. This involved taking nearly 80,000 video clips of the sample, magnified 105,000 times the original size, at various orientations. They marked prion filaments in the video clips and collected more than 500,000 overlapping sub-images. They isolated about 7,300 of the highest quality sub-images and then used supercomputers to generate a 3-D density map and a molecular model to fit the map.

Vaccine development is among the many research areas where scientists could use high-resolution prion structures to advance their work. The study authors note that previous attempts to develop vaccines against CWD in cervids failed to be protective, and, at least in one case, had the opposite effect. They speculate that one explanation for adverse vaccine effects could be that antibody binding to the sides, rather than the ends of prion fibril surfaces, promotes fragmentation – creating infectious particles rather destroying them. Thus, a strategy to explore with vaccines and small-molecule inhibitors, they say, is to target the tips of prion structures where binding and conversion of prion protein molecules occurs.

The research team is planning to solve other naturally occurring prion structures, hoping to advance its understanding of the molecular basis of prion transmission and disease.

References:

P Alam, F Hoyt, E Artikis, et al. Cryo-EM structure of a natural prion: chronic wasting disease fibrils from deer. Acta Neuropathologica DOI: 10.1007/s00401-024-02813-y (2024).

A Kraus et al. High-resolution structure and strain comparison of infectious mammalian prions. Molecular Cell. DOI: 10.1016/j.molcel.2021.08.011. (2021).

In the early 1980s, scientists identified clumps of abnormal, misfolded prion protein in mammals as the cause of brain-wasting diseases, now called prion diseases. Since that time, they have struggled to answer: What does a normal prion protein do?

The answer, they believe, could help lead them to develop treatments and disease-prevention measures against human prion diseases, such as Creutzfeldt-Jakob disease, fatal familial insomnia and kuru, as well as animal prion diseases, such as scrapie in sheep and chronic wasting disease in cervids.

Now, a new study published in iScience from NIAID scientists at Rocky Mountain Laboratories in Hamilton, Montana, and colleagues provides details of how prion protein functions in the retina of mouse eyes, helping them respond to light.

The scientists used mice specially bred without prion protein to compare to wild mice with natural prion protein. Prior studies have suggested that prion protein may have a role in how nerves transmit signals to other nerves at specialized junctions, known as neural synapses. So, knowing that prion protein exists naturally in the eye, the researchers examined mouse retina for a specific neural synapse role.

A key tool the researchers used involved measuring the electroretinographic (ERG) responses – the amount of time it took for the retina in mice to respond to a flash of light. Remember as a kid in school learning about rods and cones in the eye and how they convert light signals to help the brain understand vision? The same is true in mice.

Compared to the wild mice with prion protein, the scientists observed deficiencies in ERG responses for mice without prion protein. The deficiencies affected the normal function of the rods and cones. And – using the ERG data and neural synapse information – they found that the deficiencies originated in the portion of the retina where natural prion protein was most highly concentrated.

Though additional study is needed, the researchers believe the prion protein may act like scaffolding to help cells and elements of the eye, such as rods and cones, to stabilize neural synapses. And they believe prion protein must be present for rods and cones to function normally.

The research team hopes these findings help colleagues who study prion diseases better understand what might occur in humans when natural forms of prion protein are therapeutically removed. New treatment strategies for prion diseases focus on using drugs that remove natural prion protein to eliminate the potential for misfolding and clumping. But researchers do not know whether that could result in unwanted outcomes, such as possibly affecting vision. These findings also could extend to other protein-related neurodegenerative diseases, such as Alzheimer’s (amyloid beta protein) and Parkinson’s diseases (alpha synuclein protein).

Scientists from Duke University and the McLaughlin Research Institute in Great Falls, Montana, collaborated on the study.

Reference: J Striebel, et al. The prion protein is required for normal responses to light stimuli by photoreceptors and bipolar cells. iScience DOI: 10.1016/j.isci.2024.110954 (2024).

Novel Study Model Reveals New Understanding of Fatal Familial Insomnia
Cerebral Organoids Show NIAID Investigators Disease Characteristics

Fatal familial insomnia (FFI) is a little-known yet horrific disease in which people die from lack of sleep. A protein mutation in the brain prevents sleep, and the body gradually deteriorates. Fortunately, the disease is extremely rare. Fewer than 1,000 people in the United States are estimated to have FFI, according to the NIH’s Genetic and Rare Diseases Information Center. Unfortunately for those with the disease, it can be hereditary – thus the “familial” aspect in the name and importance of developing diagnostic tests and treatments. 

FFI is among a group of unusual neurologic conditions known as prion diseases – those caused by normally harmless prion protein that can malfunction and kill brain cells. Because FFI is found in the brain (suspected origin is the thalamus) studying its spread is not possible until a patient has died. But at that point, valuable disease information is not available because the brain no longer functions. Likewise, studies in rodents and laboratory glassware only have provided limited information.

In a new study published in PLOS Genetics, scientists from the National Institute of Allergy and Infectious Diseases (NIAID) developed a cerebral organoid model to study the exact protein mutation that causes FFI. Human cerebral organoids are small balls of brain cells ranging in size from a poppy seed to a pea; scientists use human skin cells to create organoids. Cerebral organoids have organization, structure, and electrical signaling systems similar to human brain tissue. Because they can survive in a controlled environment for months to years, cerebral organoids also are ideal for studying nervous system diseases over lengthy periods of time.

In the new study using the FFI organoids, NIAID scientists working at Rocky Mountain Laboratories in Hamilton, Mont., compared cell functions – primarily in neurons – between the FFI model and organoids without the FFI protein mutation, making several important observations about the mutation’s effect on brain cells.

They believe the abnormalities likely are features of asymptomatic FFI that may lead to disease.

They surprisingly did not observe spontaneous change in shape or spread of the FFI protein to additional prion protein. Such spread typically is a trademark of prion disease – changing prion protein throughout the brain from the normal shape to the malfunctioning folded shape.

“Our findings show that the mutation causes brain cells to dysfunction without the need for misfolding,” the study states. “We could confirm that most changes were caused by the presence of the mutation” rather than interacting with prion protein lacking the mutation.

Further, the group found that neurons in the FFI organoid model were impaired because of damaged mitochondria – which typically produce energy to keep the brain cells healthy. In future studies they hope to establish a relationship between impaired mitochondria function and the mutated FFI prion protein, and whether neurons attempt to stay healthy and avoid harm from the mutated FFI prion protein by switching from mitochondria as an energy source.

They also plan to explore relationships between the mutated FFI prion protein and neurons associated with “wakefulness,” sleep and rapid eye movement in the brain.

Reference:
S Foliaki et al. Altered energy metabolism in Fatal Familial Insomnia cerebral organoids is associated with astrogliosis and neuronal dysfunction. PLOS Genetics DOI: (2023).

Prions are infectious protein pathogens that cause fatal neurodegenerative diseases in mammals. For decades, scientists have wondered how different strains of prions can propagate when they do not carry their own genes with them as they move from host to host. A new study from NIAID researchers and colleagues at Case Western Reserve University in Cleveland reveals – in near-atomic detail – how differences in the folding of the primary protein of prions (PrP) can help determine the distinct characteristics of prion strains.

The research teams are pursuing studies of prion structure to aid in the design and screening of treatments that might slow or prevent the spread of these deadly diseases. Prion diseases are caused by the corruption of the normal form of PrP that is made by all mammals. Although poorly understood, normal PrP molecules spend most of their time as individual units that play roles in multiple physiological functions.

Rarely, and for unknown reasons, normal PrP molecules can spontaneously refold into highly structured assemblies that cause brain disease. This spontaneous process seems to be responsible for the most common form of prion disease in humans, sporadic Creutzfeldt-Jakob disease. However, once formed, corrupted PrP aggregates, or prions, can be highly infectious if inadvertently transferred from one person to another by invasive medical procedures. In other mammals with poorer basic hygiene, such as livestock and wildlife, prion infections such as scrapie, chronic wasting disease, or mad cow disease can spread by more casual contact and contaminate the environment.

Multiple strains of prions have been identified in all these host species, with each giving a distinctive array of clinical presentations, molecular features, patterns of damage in the brain, and transmissibility to other species. Thus, deciphering the basis of prion strain diversity is important in understanding the risks posed by prions to which humans or animals are exposed.

Prions can reproduce in hosts a billion times or more during infection. They do so without having their own infection-specific genes because all PrP molecules are encoded by the same host gene. For this reason, scientists cannot easily track prions like they can viruses and bacteria.

The new study, published in Nature Communications, comes from the same researchers who in 2021 published the first high-resolution structure of an infectious prion protein. That work solved the structure of a specific hamster-adapted strain of scrapie, a prion disease that occurs naturally in sheep and goats. The initial study also reported lower resolution images of a mouse-adapted form of scrapie that already indicated strain-dependent differences in overall shape. 

The latest study uses cryo-electron microscopy to show the structure of this mouse-adapted second prion strain in much greater detail. The strain was depicted at a resolution of about 3 angstroms in size (1 angstrom is equal to one hundred-millionth of a centimeter). Comparing these rodent prion strains reveals that, although they share some key structural similarities, their details greatly differ overall.

Importantly, both strains are pancake-like layers of PrP molecules with a ladder-like protein substructure. Prior to the 2021 study, scientists only had educated guesses to guide their work regarding prion characteristics.

“The structural basis for how prions replicate as deadly pathogens, and to do so consistently as different strains to cause distinct diseases, has long been a major mystery in our field of research,” Dr. Byron Caughey, the senior NIAID scientist on the study, said. “Now we have a much clearer idea of how this works – at least for these first two strains that we have solved so far.”

References:
F Hoyt et al. Cryo-EM structure of anchorless RML prion reveals variations in shared motifs between distinct strains. Nature Communications. DOI: https://doi.org/10.1038/s41467-022-30458-6 (2022).

A Kraus et al. High-resolution structure and strain comparison of infectious mammalian prions. Molecular Cell. DOI: 10.1016/j.molcel.2021.08.011. (2021).