The Prion Hypothesis at Forty: Enlightening or Deceptive?

Spoiler: The prion hypothesis was developed in the context of misleading premises, skewed interpretations of experimental data and observations, and omission of previous findings and knowledge.

When it comes to experimental work, science has built-in mechanisms for self-correction [1], but they are weak [2-4]. These mechanisms are even weaker in the case of broad paradigms and theories, which often have a long life of their own, even when faced with conflicting data and observations [5]. This certainly seems to be the case with the Amyloid Hypothesis and related concepts [6, 7], which are imbedded in two other questionable major paradigms: the protein misfolding dogma and the prion hypothesis that have dominated the thinking on the etiology of Alzheimer’s diseases (AD), Parkinson’s disease (PD), amyotrophic lateral sclerosis (ALS), Creutzfeldt-Jakob disease (CJD), and other neurodegenerative diseases for decades. Unless the validity of these broad paradigms is addressed in a timely and comprehensive manner, the understanding of neurodegenerative diseases and their associated pathology is likely to remain entrapped in confusion for years to come.

After the discovery that the amyloidogenic proteins implicated in neurodegenerative diseases, including prion protein (PrP), amyloid-β (Aβ), tau, α-synuclein (α-Syn), and TDP-43, acquire β-sheet-rich folds, a structure that was not thought to be compatible with physiological functions, these proteins were classified as misfolded proteins, and the associated disorders as protein misfolding diseases (reviewed in [8, 9]). The classification of the amyloidogenic process as a protein misfolding event uncoupled the folding process, its products (i.e., the protein aggregates), and the associated pathology from the biological functions of these proteins and the evolution of their genes. In other words, the aggregation and the presumed toxicity of these proteins has been regarded as an accidental event, unrelated to their evolution and biological function.

Nevertheless, the prion hypothesis has been the critical conceptual construct that reinforced the protein misfolding paradigm and set a strong conceptual barrier between the physiological function of these proteins and the disease mechanisms. Because prions were defined and promoted as novel, protein-only infectious pathogens that self-replicate as misfolded proteins, they have been viewed as completely independent of the physiological function and the evolution of the PrP and its gene [10].

The prion hypothesis, suggesting that the pathogens causing scrapie, kuru, CJD, and other transmissible spongiform encephalopathies (TSEs) are non-viral, self-replicating protein-only pathogens lacking a nucleic acid-based genome, has been one of the most fascinating albeit controversial ideas in biology. Additionally, in conjunction with the protein misfolding dogma, the prion paradigm has been one of the leading working hypotheses explaining the pathogenic mechanisms leading to AD, PD, ALS, and other neurodegenerative disorders, as addressed and discussed in hundreds of studies and publications (reviewed in [11, 12]).

Here I present evidence and arguments that the prion hypothesis was developed on the basis of confusing and misleading premises, skewed interpretations of experimental data and observations, and the omission of previous findings and knowledge. It is important to clarify that this presentation does not question the validity or the importance of the findings and observations in the TSE field, but only their interpretation.

The Scientific Premises, Evidence, and Rationale Behind the Prion Hypothesis Are Questionable
Forty years ago, in 1982, Stanley Prusiner introduced a new term—“prion”—to designate the mysterious TSE pathogens [13], which previously had been referred to as “unconventional viruses” [14]. Interestingly, the term “prion” was first publicly announced in a newspaper article entitled “Tiny new form of life” [15] and later defined in a scientific article published in the journal Science: “In place of such terms as ‘unconventional virus’ or ‘unusual slow virus-like agent,’ the term ‘prion’ (pronounced pree-on) is suggested. Prions are small proteinaceous infectious particles which are resistant to inactivation by most procedures that modify nucleic acids.” [13].

Defining prions as “proteinaceous infectious particles” was confusing, as many other infectious agents, especially viruses, could also be characterized as “proteinaceous.” In addition, several hypotheses about the putative non-viral nature of the TSE pathogens, including the paradigm that they might be protein-only infectious agents, had already been proposed two decades earlier, in the 1960s, when it was shown that the TSE pathogens (more precisely their transmissible forms—see below) were indeed resistant to inactivation by most procedures that modify nucleic acids [16, 17].

However, the most remarkable, albeit obscured scientific and historical fact about the prion hypothesis is that, as Prusiner has explained, the term “prion” was NOT introduced to denote a non-viral, protein-only scrapie pathogen, as incorrectly reported in thousands of scientific articles and textbooks over the last four decades. Instead, the term “prion” was coined as a lexical novelty to replace the traditional “dumb” names for the scrapie pathogen: “They had their own words, that’s true, but they were really dumb. ‘Unconventional virus’ is as dumb as it gets.” [18].

As Prusiner pointed out, the idea that the term “prion” was introduced as a scientific and conceptual novelty to indicate the protein-only nature of the TSE pathogens (to the exclusion of a viral etiology) was erroneously advanced by other investigators in the field, who, for reasons difficult to decipher, have “redefined prions as infectious proteins” [19]. Prusiner was very clear on this error: “Some investigators have misinterpreted the term prion. They have used prion to signify infectious proteins (23, 24) or even as a synonym for scrapie associated fibrils (25). This misuse of the term prion has led to confusion and should be avoided.” [19].

According to Prusiner, he introduced the term “prion” following the advice of a fellow scientist: “‘When you have a better idea about the agent’s composition, you’ll need to spend some time thinking of a name for it. The name is very important. If you choose a bad name, someone will come along and rename it, and if that happens, your contributions to this discovery may become obscured. But if you give it a good name, it will stick.’” ([20], pg. 86). It seems inconceivable, though, that the outstanding contributions of Prusiner’s laboratory to the TSE field would have been obscured, absent the introduction of the term “prion.” However, this might be a naïve assumption on my part, as several articles and books have discussed at length the critical role that the word "prion" played in promoting the work performed by Prusiner and his laboratory [21-23].

As mentioned above, multiple hypotheses about the structure and biochemical composition of the pathogens causing scrapie and other TSEs were advanced in the 1960s, including that of a protein-only composition [16, 17]. How did all these broadly circulating and highly debated ideas about the nature of the TSE pathogens, including the well-publicized work on kuru which was rewarded with a Nobel Prize in the mid-1970s [14], end up being re-incarnated in the 1980s as the ill-defined prion hypothesis, which has been regarded ever since as one of the greatest novelties in biology?

The philosophers of science and the scholars of epistemology might derive more complex answers to this intriguing question and to the bizarre turn of events regarding the decades-long dispute over the etiology TSEs that led to the introduction of the novel term “prion” and of the confusing prion hypothesis. Here, I offer a rather simple explanation rooted in the way science is usually conducted in the laboratory or in the field, in which reductionist questions and research objectives dominate. When embedded in misleading premises, this approach could lead to misinterpretations and conceptual artifacts [24]. In the case of the TSE field and the prion hypothesis, the culprit has been the century-old misleading dogma of viruses as virus particles, in accordance with which viruses have been erroneously defined based on the physical and biochemically properties of their virus particles (see below).

For the last half of century, the central question and primary experimental objective in the TSE field has focused on the properties and the biochemical composition of the TSE pathogen as present in the material transmitting the disease, usually brain homogenates from diseased animals: Does the TSE pathogen in the transmissible material (i.e., the TSE inoculum) contain a nucleic acid-based genome, or not? In other words, is this TSE pathogen a virus, or a novel self-replicating protein-only pathogen?

From the early 1960s through the 1990s, hundreds of studies searched in vain for viral nucleic acids in the material used for TSE transmission. By the late 1990s, it had become abundantly clear that the TSE inoculum does not contain a TSE-specific nucleic acid. In the midst of the ‘mad-cow disease’ public health scare, the prion hypothesis and Prusiner were celebrated with the Nobel Prize: “While no single experiment can refute the existence of the “scrapie virus”, all of the data taken together from numerous experimental studies present an impressive edifice which argues that the 50-year quest for a virus has failed because it does not exist!” [10].

What if “the 50-year quest for a virus has failed,” not because the virus did not exist, but because the rationale and experimental approaches adopted in the search for a viral nucleic acid in the TSE transmissible material as a means to identify the putative virus were fundamentally flawed? Indeed, what if defining the nature of the TSE pathogens, or that of any infectious agent or organism, based on their properties and biochemical composition in a particular stage of their life cycle misrepresents their true nature?

Consider, for example, the original prions, which are seabirds, types of petrel that acquired their name long before the scrapie agent ([25]). One might try to identify and describe them on the basis of their properties and features during the egg stage of their life cycle: no wings, no feathers, no squealing, basically no prions, just eggs, as fascinating as they might be. Or consider Chlamydia trachomatis, an infectious bacterial pathogen frequently found on college campuses [26], which, until the 1960s, was classified as a virus. Certainly, the physical, biochemical, and biological properties of the transmissible extracellular stage in the life cycle of this pathogen do not represent the properties of C. trachomatis as an obligate intracellular pathogen. And then, there is a huge, over a century-old Elephant in the Grand Room of Biology—the misleading dogma of viruses as virus particles, or virions. Ever since viruses were identified at the end of the 19th century as infectious agents that passed through porcelain filters thought to retain all other pathogens, such as bacteria, viruses have been conceptually misidentified with virus particles and erroneously defined on the basis of the physical, biochemical, and biological properties of those particles [27-32], as described and illustrated in virtually all scientific literature ever since.

For example, in his seminal book, The Molecular Biology of the Gene, published half-a-century ago, James Watson, who surely was highly familiar with nucleic acids, wrote: “All viruses differ fundamentally from cells, which have both DNA and RNA, in that viruses contain only one type of nucleic acid, which may be either DNA or RNA” [33]. A decade later, in A Dictionary of Virology, viruses were defined as “Infectious units consisting of either RNA or DNA enclosed in a protective coat” [34], and in the 1990s, a classical microbiology textbook stated that viruses “consist of a genome, either RNA or DNA, that is surrounded by a protective protein shell” [35].

Certainly, the authors of these scientific texts were fully aware that, during the intracellular stage of their life cycle, thousands of diverse viruses have both type of nuclei acids, DNA as well as RNA, and that many of them are much more complex than a nucleic acid wrapped in a protein coat. Yet, all these renowned scientists have fallen victims to the misleading dogma of viruses as virus particles, as they identified viruses conceptually with virus particles and defined them based in the physical and biochemical properties of these particles. This is a strong example of the power of concepts in science: a concept that blatantly misrepresents experimental findings, observations, and knowledge can still be viable for decades, or, as in the case of viruses, for more than a century. In the context of the historical conception of viruses as virus particles and of the main dispute whether the TSE pathogens are viruses or different type of pathogens, it is not surprising, therefore, that, similar to viruses, prions were defined as “small proteinaceous infectious particles” [13].

In context of a new perspective on the evolution and nature of viruses that questioned the dogma of viruses as virus particles [27], soon after the gene for the PrP was shown to be presumably encoded by a host chromosomal gene [36, 37], I proposed that the transmissible forms of the TSE pathogens were protein-only particles produced by an endogenous virus [38]. To expand on the lexical novelty already introduced by Prusiner, I called these endogenous viruses "prionic viruses," while reserving the term “prion” for their transmissible protein-only particle [38].

Shortly after the publication of this hypothesis, William Haseltine and Roberto Patarca showed that the PrP and its gene share sequence domains that are similar and collinear to those that occur in the retroviral reverse transcriptase gene and protein [39]. Later, it was also shown that PrP shares structural and functional properties with HIV-1 fusion peptide [40] and RNA binding and chaperoning properties with nucleocapsid protein NCP7 of HIV-1 [41]. Moreover, as I previously discussed [42], one of the most striking features of the PrP gene is the lack of introns within the protein coding region of the gene a rare phenomenon among vertebrate genes, but one that is highly predictable for viral genes.

It is well known that most viruses produce defective virus particles that do not contain the viral genome. Although these virus particles can be transmitted to other host cells or individuals, in the absence of the viral genome, they cannot establish a productive infection [43]. However, this is not the case with endogenous viruses, which have their genome integrated into the host genome and are therefore present in all cells [44]. For example, the human genome contains millions of endogenous viral sequences, which surpass the number of human genes many times, and some of these endogenous virus genes encode for proteins that self-assemble into virus particles that do not contain the viral genome (see below). If an endogenous virus produces protein-only particles that enter other cells of the same or, if inadvertently transmitted, different individuals, where they induce the assembly of similar particles, then, these ‘small proteinaceous infectious particles’ would appear to be non-viral, self-replicating protein-only infectious agents. However, this only occurs if they are mistakenly taken out of the context of their endogenous virus etiology [42]. In this case, it would be nonsensical to search for a viral nucleic acid in the transmissible particles in order to establish their viral or non-viral nature. Certainly, in this case, the creed of the prion hypothesis that the “scrapie virus…does not exist” (see Prusiner’s quote above) would collapse.

Remarkably, it was recently discovered that the Arc protein, a master regulator of synaptic plasticity, memory formation, and cognition, is encoded by an endogenous virus gene evolutionary related to those encoding for the Gag polyproteins found in some retrotransposons and retroviruses [45]. Surprisingly, it was found that, like some other proteins produced by human endogenous viruses, Arc protein assembles into virus-like particles, which exit their host cell and enter other cells through a process that resembles viral transmission [46, 47].

The idea of an endogenous viral origin of PrP has also been advanced by Charles Weissmann, whose laboratory cloned and sequenced the PrP gene and generated the first PrP gene knockout animal model [36, 48], both of which represented major breakthroughs in the TSE field: “Another possibility is that PrP/PrPSc is derived from an ancient pathogen, the genetic material of which was integrated into the genome of its host and harnessed to fulfil a useful function while its pathogenic potential was minimized” [49]. However, by the time this idea was proposed, the prion hypothesis was considered one of the most amazing conceptual novelties in biology, and it had already been engraved in textbooks and in the minds of researchers, scholars, and students as a bona fide scientific fact, despite lingering questions regarding its validity. To quote a reflection by Rudy Castellani and Mark Smith in their quest to address parallel problems with the Amyloid Hypothesis [50], the prion hypothesis was also “too big to fail.”

By the time Prusiner received the Nobel Prize in 1997, fifteen years after he introduced the term “prion,” he had changed the definition of prions from “small proteinaceous infectious particles which are resistant to inactivation by most procedures that modify nucleic acids” [13] to one stating unambiguously that “a prion is a proteinaceous infectious particle that lacks nucleic acid” (emphasis added; [10]). Yet, more than a decade later, Prusiner abandoned the definition of prions that focused on the “infectious particles” in favor of one focusing on the PrP’s isomeric conformational changes and their self-propagating property: “Prions are proteins that acquire alternative conformations that become self-propagating” [51].

This radical change was prompted by the fact that the “new biological principle of infection,” the novelty for which Prusiner received the Nobel prize, lost its universal aura, when it was established that many proteins defined as prions or ‘prion-like’ were not infectious. Significantly, the new definition expanded the prion paradigm to a group of human diseases of extraordinary medical and public health relevance, in which the protein misfolding dogma was well established [8, 11]. Indeed, as illustrated in hundreds of publications (reviewed in [11, 12, 52], the prion paradigm has been increasingly used to explain the conformational changes and misfolding of Aβ, tau, α-Syn, and TDP-43, the primary proteins implicated in AD, PD ALS, and other neurodegenerative diseases.

However, as discussed in the next section, the conformational changes, the cyclic assembly of PrP into various aggregates, which were interpreted as prion replication, and the associated pathogenic mechanisms can be explained in context of PrP’s physiological function. Similarly, the biological functions of Aβ, tau, α-Syn, and TDP-43 are foundational in understanding their aggregation, which currently is interpreted as a protein misfolding event, as well as their associated pathogenic mechanisms that lead to neurodegeneration.

The Biological Functions of PrP, Aβ, tau, α-Syn, and TDP-43 Are Critical in Assessing the Validity of the Prion Hypothesis and Protein Misfolding Dogma
Six years after Prusiner’s Nobel Prize for the prion hypothesis, Kurt Wüthrich, another Nobel laureate for his work on the three-dimensional structure of biological macromolecules, including PrP aggregates, submitted that: “we must understand the function of the normal prion protein before we can understand prion diseases” (cited in [53]). Now, almost twenty years later, the biological function of PrP is as elusive as ever, which begs the question: do we still not understand prion diseases?

Like the other amyloidogenic proteins implicated in neurodegenerative diseases, PrP has been one of the most studied proteins ever. Although defining function in biology is still a work in progress [54, 55], it is surprising that the physiological functions of PrP, Aβ, tau, α-Syn, and TDP-43 remain enigmatic decades after their discovery. Given their evolutionary conservation, which suggests important biological functions, the absence of strong phenotypic effects in gene-knockout animal models is puzzling. As I previously suggested [42, 56, 57], the lack of progress in defining the biological function of these proteins reflects the fundamental scientific confusion induced by the misleading working paradigms in the field, the protein misfolding dogma and the prion hypothesis.

In the context of the proposed model for the evolutionary origin of the PrP gene from an ancestral endogenous virus, I proposed that PrP, which is expressed at high levels not only in the brain but also in the immune cells, is an innate immunity protein that performs its protective functions by participating in two major overlapping mechanisms or pathways: (i) directly, by blocking the life cycle of various microbial and viral pathogens, for example, by damaging the microbial cellular membrane or the host cell membranes required for viral replication, or (ii) indirectly, by inducing the death of host cells through various inflammatory and non-inflammatory mechanisms, which limits the spread of infection [42, 57].

Additionally, drawing on comparative data with the other primary proteins implicated in neurodegenerative disorders, and on broad new interpretations of the existing data and observations in context of an evolutionary framework, I proposed that:

(i) Like PrP, Aβ, tau, α-Syn, and TDP-43 are members of the innate immune system;
(ii) The conformational changes of these proteins and their assembly into various oligomers and amyloid aggregates are not protein misfolding events, as they have been defined for decades, nor are they prion-replication activities, but rather they are part of these proteins’ evolutionarily selected innate immune repertoire;
(iii) The reactions and activities associated with the function of these proteins in innate immunity leads to pathological and neurodegenerative events that define AD, PD, ALS, and CJD as innate (auto)immune disorders [42, 57].

The protective role of PrP and the other proteins in this group has been addressed and reviewed in dozens of publications [58-66]. To perform their innate immunity functions, these proteins have been evolutionarily selected to acquire multiple functional conformations and to assemble into various oligomer structures, or Innate Immunity Complexes (IICs), which mimic pathogen- or danger-associated molecular patterns (PAMPs & DAMPs). In their native, unengaged, ligand-like conformation, these proteins can exist as monomers or small oligomers that contain primarily α-helix protein folds. Upon contact with microbial or viral components, or after detecting ‘danger signals’ associated with diverse microbial and viral infections or other types of injuries, including physical, biochemical, immunological, and age- related injuries, they assume new isomeric conformations that are rich in β-sheet folds, a defining characteristic of amyloids. By mimicking PAMS and DAMS, these IICs amplify and transduce these signals to downstream innate immunity pathways (for reviews of putative PrP signaling properties and pathogenic pathways, see [67-76]).

Although, like other members of the immune system, PrP, Aβ, tau, α-Syn, and TDP-4 have been strongly selected against extended pathogenic reactions that would lead to autoimmune diseases, they run a fine line between ‘protection’ and ‘pathogenicity.’ Indeed, in the context of their complex immune activities, which are exercised within the narrow gap between protection and injury, these putative innate immunity proteins perform reactions that could be narrowly defined as a gain of (toxic) function (GOF), or a loss of (physiological) function (LOF) [77, 78]. However, as is the case with most immune system activities, including autoimmune reactions, these events are part and parcel of their evolutionarily selected biological functions. It is important to point out that the innate immunity paradigm addresses one of the main disputes in the field of neurodegenerative disease, the conflicting views over the question whether GOF or LOF are the primary pathogenic mechanisms in this groups of diseases [79, 80].

A model for the cycling molecular mechanisms of PrP, Aβ, Tau, PrP, α-Syn, and TDP-43, which lead to their functional aggregation into various IICs, as well as their various roles in the immune system, has been described earlier in greater detail [57]. One of the most puzzling phenomena associated with PrP, as well as with Aβ, Tau, PrP, α-Syn, and TDP-43, has been the apparent existence of ‘strains,’ a topic at the center of the decades-long dispute over the viral or non-viral nature of TSE pathogens. Currently, the existence of ‘strains’ is one of the most contentious propositions in the effort to understand the apparent variable toxicity associated with the protein aggregates implicated in AD, PD, ALS, and other neurodegenerative diseases.

In the context of the innate immunity model, the ‘strain’ phenomenon is expected. Briefly, during their cycling innate immunity reactions, PrP, Aβ, Tau, α-Syn, and TDP-43 assemble into various IICs, which are recognized by the native parental protein molecules with differential kinetics, leading to a selection process and the preferential formation of certain IICs. This selection process, which is sustained by the conformational flexibility of these proteins—a remarkable process that could be envisioned as an adaptive immune feature—opens the door to pathogenic (auto)immune reactions. A second selection process occurs in association with the propensity of various IICs to circulate among neighboring cells and tissues, whereby they become the targets of the resident parental proteins, leading to an expanded continuous cycle, which can be defined fundamentally as an ‘autoimmune’ phenomenon. Although the cycling process can be resolved, at least partially, by the formation of large benign assemblies, such as amyloid plaques and tangles, the neuronal damage remains a permanent feature of the brain’s tissues, which contain primarily post-mitotic cells. Although the amino acid sequence of the proteins within the IICs can be a strong determinant of the IICs’ specific structural features and reactivity, many other factors, including, for example, pH or the presence of metal ions, can influence their ‘strain’ properties.

How does the biological function of all these proteins in innate immunity challenge the fundamental tenets of the protein misfolding dogma and the prion hypothesis and inform our understanding of neurodegenerative diseases and the associated pathogenic mechanisms?

As relevant as the prion hypothesis might have been, or not, in addressing and explaining TSEs from a medical and public health perspective, in context of biology, the holy grail of the prion hypothesis has been the idea of protein self-replication. The following are three recent quotations that reflect the current, state-of-the-art understanding of prion replication:

(i) “Prion diseases are neurodegenerative disorders caused by conformational conversion of the cellular prion protein (PrPC) into scrapie prion protein (PrPSc). As the main component of prion, PrPSc acts as an infectious template that recruits and converts normal cellular PrPC into its pathogenic, misfolded isoform” [81];
(ii) "According to the widely accepted ‘protein-only’ hypothesis, an abnormal PrP isoform is the infectious agent acting to replicate itself with high fidelity by recruiting endogenous PrPC” [82]; and,
(iii) “PrPSc multimers propagate by binding and refolding PrPC as they elongate” [83] (emphasis added).

As I previously proposed, a prion (whatever it is), or PrPSc in its monomeric or multimeric form, does NOT “recruit,” “bind,” “convert,” and “refold” PrPC and, surely, it is NOT “acting to replicate itself with high fidelity”[42, 57]. That is just a conceptual mirage rooted in the misleading protein misfolding dogma and the prion hypothesis. Instead, according to the innate immunity model, it is the PrPC molecule that acts to recognize and bind the PrPSc, and it is PrPC which changes its conformation and assembles into IICs to perform its innate immunity functions (for a discussion on the innate property of PrPC to acquire multiple isomeric conformation, see [84]). By analogy, it is the antibody that recognizes, binds to, and changes its conformation to form an antibody-antigen complex [85], NOT vice versa, as an agnostic unaware of the biological functions of antibodies and the evolution of their genes could argue. To expand on this analogy, the newly formed IICs can be envisioned as ‘antigenic targets’ for the ‘antibody- like’ parental innate immunity proteins, which have innate flexibility in their binding and assembling conformations.

Similarly, the pathogenic mechanisms implicated in neurodegenerative diseases do NOT represent generic activities of accidentally misfolded garden-variety proteins. According to the innate immunity model, the pathogenic mechanisms and activities of these proteins are linked to and can be explained in context of their evolutionary selected biological functions in innate immunity. Interestingly, a finding that is not well known outside the TSE field, and which increases the confusion surrounding the prion hypothesis, is the apparent dissociation of prions, as entities that transmit the disease and self-replicate, from the enigmatic toxic entities that induce pathogenicity [86, 87].

Unlike the protein misfolding dogma and the prion hypothesis, in which the aggregation and activities of PrP, Aβ, Tau, α-Syn, and TDP-43 are considered accidental events, their putative biological function in innate immunity would explain the molecular mechanisms leading to neurodegeneration. In this case, the activity of all these proteins could be explored from the same angle, both in “healthy” and in “disease” states. Additionally, the similar biological function of all these proteins would explain their interactions across multiple neurodegenerative conditions.

Oversight of Previous Studies on Transmissible Amyloids Was Critical in the Formulation of the Prion Hypothesis and Its Promotion as a Fundamental Novelty in Biology
In a seminal 1993 article on the molecular mechanism of amyloid formation in scrapie and AD, Joseph Jerrett and Peter Lansbury wrote, “By striking analogy to experimental transmission of scrapie, systemic amyloidosis can be induced in hamsters by intraperitoneal injection of a preparation derived from sonicated amyloid fibrils” and “Thus, scrapie may be a form of transmissible amyloidosis” [88]. To set this statement in its historical context, the first experimental studies on homologous and heterologous transmission of systemic amyloids were performed in the 1960s [89, 90], during the same period when the dispute over the nature of TSEs pathogens emerged as one of the most contentious issues, both in the medical field and public health and in biology (for an outline of the parallel studies on transmissible amyloids and TSEs, see [91]).

Interestingly, unlike the putative functions of PrP, Aβ, Tau, α-Syn, and TDP-43 in innate immunity, which although supported by direct evidence are still hypothetical [57], the precursor of Amyloid A (AA), the protein causing systemic AA-amyloidosis, ever since it was discovered half a century ago, has been recognized as an innate immunity acute reactive protein, which is expressed in response to various infections and other inflammatory conditions (reviewed in [91, 92]).

It is also important to emphasize that, by 1960s, it was well established that the amyloids were protein aggregates rich in beta sheet conformation (reviewed in [9]). Paradoxically, despite these early studies showing that systemic amyloidosis was transmitted by an amyloid (i.e., by a protein agent) labeled “amyloid enhancing factor” (AEF), which just like prions represented a “new biological principle of infection,” the systemic AA-amyloidosis was eventually branded as a “prion disease” and the transmissible amyloid (the AEF) as a “prion” [92]. After all, it seems that Prusiner was right about the significance of lexical novelties and drive in science [22, 93]; lexically, the term “prion” was apparently superior to the term “AEF” in promoting one of the biggest novelties in modern biology.

A Way Forward
When it comes to misleading but well-established broad concepts and paradigms, the pace of change is usually measured in generational time. In the case of the century-old misleading concept of viruses as virus particles, change is silently trickling down, under the influence of the sheer volume of knowledge regarding the life cycle of recently discovered complex viruses, which no longer can be reduced conceptually or even metaphorically to the maxim “‘A virus is piece of bad news wrapped in protein’” [94].

The historical, misleading view about viruses has delayed the discovery of a large group of complex viruses, labeled “giant viruses,” by decades. It is also clear that this view has obstructed the thinking about the origin and evolution of viruses and their role in the evolution of cellular domains [29, 32, 95, 96]. It is not surprising, therefore, that the discovery of complex viruses prompted researchers to ask radical questions—“What if we have totally missed the true nature of (at least some) viruses?”—and to provide uncomfortable answers: identifying viruses with the virus particles might “be a case of ‘when the finger points to the stars, the fool looks at the finger’” [28].

Although the conception of a virus as virus particles has led to other blunders and constraints in the interpretation and integration of data and observations in the field of virology, including some that are of medical and public health relevance (e.g., [97]), one of its most consequential effects was on the half-century dispute over the nature of TSE pathogens and the formulation of the influential and celebrated prion hypothesis. Indeed, just as in the case of viruses, the thinking about TSE pathogens and the related working hypothesis focused on the physical and biochemical properties of the infectious entities, and this led to endless disputes. Additionally, as emphasized throughout this essay, by setting a strong conceptual barrier between the pathogenic mechanisms associated with PrP, Aβ, Tau, α-Syn, TDP-43, and their biological function, the prion and protein misfolding paradigms have constrained progress in the field of neurodegenerative diseases.

Forty years after its inception, the prion hypothesis is still shifting and drifting, as some of its most ardent supporters seem to be disappointed with its prospect of moving the field forward [98]. In science, extraordinary claims require extraordinary evidence. The hypothesis that the TSEs were caused by self-replicating proteins, which was first proposed in the 1960’s and eventually reincarnated as the prion hypothesis, was nothing short of an extraordinary claim. However, even if Prusiner and the other supporters of the prion hypothesis were right in their claim that “The 50-year quest for a virus has failed because it does not exist,” this “fact” was a red herring, because it pointed to the absence of a replicating virus and not to extraordinary evidence for a replicating protein.

As discussed above, the prion hypothesis makes little biological or evolutionary sense. Apparently, it doesn’t make much sense from a pure physiochemical perspective either. Indeed, on its 40th anniversary, the prion hypothesis has been ‘celebrated’ with another refuting analysis, which, drawing on principles of thermodynamics, conveys the candid message: “Proteins Do Not Replicate, They Precipitate” [99]. Perhaps the best kept secret about the prion hypothesis is the fact that the prions, whatever they are, can arise de novo in the absence of parental prions, that is, in the absence of so-called prions’ hereditary information or templating activity [84, 100, 101].

Unlike the prion hypothesis, the claims associated with innate immunity model are only extraordinary in their radical departure from the current paradigms, but not in a biological or evolutionary sense. Nevertheless, a question that is undoubtedly on readers’ minds concerns the claim that all the primary proteins implicated in neurodegenerative diseases are members of the innate immune system. What selective pressure would drive such an evolutionary path for multiple proteins expressed at high level in the brain?

In one word - viruses. A study titled “Viruses are a dominant driver of protein adaptation in mammals” found that at least a third of the adaptive mutations in the human genome have been fixed in response to viruses [102]. Unlike cellular infectious agents, such as bacterial pathogens, which maintain a cellular structure within their host cells, viruses have a "molecular structure" [27, 32], in which their molecules are more or less dispersed within the host cell. Because of this novel biological structure, the viral molecules come into direct contact with numerous cellular proteins, and therefore, many of these proteins can acquire antiviral properties [102, 103]. This is particularly important in tissues and organs, such as the brain, that are under limited surveillance by the adaptive immune system and, therefore, represent a privileged niche for infectious agents. In these cases, it would be expected that the repertoire of innate immunity members and their activities would increase significantly.

Interestingly, other well-known proteins that are prone to aggregation, to such an extent that they have been labeled prions or prion-like entities, have recently been shown to have innate immunity functions. One of the most-studied proteins, p53, which was discovered four decades ago in association with viral antigens and has been implicated in most human cancers, has been shown to assemble into aggregates that have been labelled “prions.” However, this assertion has been questioned [104]. Remarkably, Arnold Levine, a pioneer in the p53 field, has recently suggested in a review of the field that the primary biological function of p53 is in innate immunity: “The p53 gene and protein are part of the innate immune system, and play an important role in infectious diseases, senescence, aging, and the surveillance of repetitive DNA and RNAs” [105].

Like PrP and the other proteins in this group, many other well-studied innate immunity proteins, including the interferon- and TNF-families of proteins, assemble into pathogen-like aggregates in order to perform their biological functions [106-109]. Some other innate immunity proteins, such as MAVS, ASC, RIPK1, RIPK3, assemble into antiviral innate immunity complexes (i.e., IICs) that amplify and transduce pathogen-associated signals to downstream effector pathways [110- 113]. Unfortunately, consideration of these findings in the context of the confusing prion hypotheses has led to questionable interpretations and statements. For example, it makes little sense to ascertain that “unlike most prions that confer loss of function, MAVS and ASC are both gain-of-function prions” [112], when clearly these proteins self-aggregate in order to exercise their evolutionally selected function in innate immunity, which is NOT a case of “gain-of- function”; it is simply their function. Although all proteins can misfold and assemble under certain conditions into amylogenic structures [114], these are rare events under physiological conditions, and this process is likely linked to their biological function.

In summary, the innate immunity model of the biological function of Aβ, Tau, PrP, α-Syn, and TDP-43 is supported by an increasing number of studies (reviewed in [64, 115, 116]), and, to my knowledge, it is consistent with all the confirmed data and observations that are presumed to support the prion hypothesis. In addition, this model has more explanatory power than the prion hypothesis, and it unifies the current conflicting views of the pathological mechanisms in neurodegenerative diseases [57]. So, is this model the way forward?

Like many other scientific publications, this article might contain factual errors, misleading statements, or questionable arguments. Considering also the radical departure of the innate immunity model from the leading paradigms in the field of neurodegenerative diseases—the protein misfolding dogma and the prion hypothesis—it would make sense to have the claims outlined here evaluated and consequently refuted if they are not supported or consistent with the current data and observations or embraced if they are. Moreover, unless we have become numb to the dumbing down of the historical and academic aspects of science, the truth about the introduction of the term “prion” and about the use of deceptive premises for promoting the prion hypothesis should be recognized in the scientific literature, including textbooks.

However, this is unlikely to happen, because there are no strong incentives for scientists to perform these critical tasks in a timely, open, and systematic manner. The built-in mechanisms for self-correction in science [1, 117] are sustained and incentivized through the funding of new studies that confirm or correct previous findings. The process is relatively straightforward, but even in this case it might take many years for the experimental data to ‘self-correct’ (e.g., [4]). In contrast, the broad interpretation and integration of the experimental data and observations that inform working hypotheses and direct future studies are inherently more subjective, and the investigators performing such studies are inclined to protect the premises of their funded work. By analogy to the innate immunity model presented here, this leads to a chronic cycle that amplifies the existing pathogenic misconceptions, which often persist until the arrival of a new generation of scientists [5].

A sensible solution would be for the funding agencies to invest a small portion of their budget (e.g., 1%) in a post-publication peer-review system, in which the authors would be joined by independent scientists (funded through “peer-review grants”) to evaluate all the data and ideas in a field in a timely, open, and comprehensible manner. A strong, open-ended post-publication peer-review system not only would correct flawed studies and misleading interpretations expediently, but, more importantly, it would also discourage and, one hopes, prevent scientists from entering the slippery slope of deceptive science, which robs them of their peace-of-mind and can ruin their careers, sometimes with tragic consequences [118]. Retrospectively, it is likely that many controversies and flawed studies in the field of neurodegenerative diseases, such as the confusion about the prion hypothesis [19] discussed here, or the faulty studies on toxic Aβ oligomers [4], would have been resolved quickly or even prevented.

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Last comment on 26 February 2024 by Lawrence Broxmeyer, MD


Many thanks for sharing your interesting blog Dr. Bandea. I agree with many things, especially the confusing, mechanistically-unsubstantiated aspects of the prion hypothesis. Regarding the innate immune properties of amyloids, since any protein can form amyloids, I’m not sure if it is a functional property. Proteins such as insulin or myoglobin, which have very specific, non-immune functions, can still form amyloids, and so can any other peptide sequence. That’s why I subscribe to the “generic hypothesis”(coined by the late Chris Dobson), that amyloids are a generic conformation of proteins that they adopt under certain conditions. We are going to publish a book chapter soon where we postulate that a simple extension of the Anfinsen hypothesis of protein folding to supersaturated conditions can fully explain the amyloid phenomenon without the need for the assumptions of the prion hypothesis. We argue that proteins possess two thermodynamically stable conformations and possess the necessary information to adopt either of them:

1.    The native conformation, which is thermodynamically favorable under sub-supersaturation conditions and is dependent on the primary sequence and specific side-chain interactions.

2.   The cross-β (amyloid) conformation, which is thermodynamically favorable under supersaturated conditions and is dependent on generic intermolecular backbone interaction facilitated by the molecular proximity created under supersaturation.

The concentration and the availability of nucleation catalysts are the factors that decide which conformation will be adopted. But since amyloids are generic (depend on backbone interactions and the characteristic protein side chains are buried within the cross-beta architecture) and their growth is uncontrolled, I believe mechanisms (such as chaperones, and proteasomes) have evolved to prevent their formation and not the other way round. That’s probably why they are mostly associated with disease when these mechanisms fail, or when concentration is pathologically increased (gene duplication), or when nucleation catalysts invade the microenvironment (viruses, other infections). 

I find the possibility that PrPC is part of the capsid, or protein component in general of a virus, or that it is part of the toolkit of innate immunity to be fascinating. I think they are possibilities that deserve attention and research. On the other hand, I do not think that any of these possibilities necessarily contradicts or is incompatible with the prion proposal. Let me provide an example: the origin of mitochondria is aerobic bacteria that established a symbiotic relation with ancestral eukaryotic cells. There is no question about that. However, they are also organules, as much as lysosomes, peroxysomes or the Golgi apparatus are. I think that the experimental evidence supporting that PrPSc prions facilitate conversion of PrPC to PrPSc is overwhelming. And there is nothing strange about it: it happens with all seeded amyloid conversions. Incubate any protein under denaturing conditions and slightly acidic pH at 37C and, as Dobson has shown, it will eventually adopt an amyloid conformation; repeat the experiment adding a bit of your pre-formed amyloid as a seed, and the process will be much shorter. I do not see anything unusual there. 


I would like to thank Kariem Ezzat and Jesús Requena for their comments and for bringing forward a critical issue that has plagued the prion hypothesis for the last four decades: what exactly are prions? To address this question, the prions need to be clearly defined and differentiated from other protein aggregates.

When viruses were identified as a distinct group of infectious pathogens at the end of the 19th century, they were defined as novel disease agents that passed through porcelain filters assumed at the time to retain all microbial (i.e., cellular) pathogens [1]. Martinus Beijerinck labeled these ‘filterable pathogens’ with the descriptive name “contagium vivum fluidum”. Later, in the mid-1930s, Wendell Stanley isolated the tobacco mosaic virus, more exactly the virus particles produced by the virus, in a crystalline form and defined viruses as ‘self-replicating proteins’: “Tobacco-mosaic virus is regarded as an autocatalytic protein which, for the present, may be assumed to require the presence of living cells for multiplication” (all quotes in italics; [2]). This definition was abandoned after it was found that the infectious virus particles contained nucleic acids [1]. Two decades later, Andre Lwoff concluded his seminal article “The concept of virus” with the (in)famous quip: “viruses should be considered as viruses because viruses are viruses” [3]. So, should prions be considered as prions because prions are prions?

Like viruses, transmissible spongiform encephalopathies (TSEs) pathogens were conceptually identified with their transmissible forms — the “proteinaceous infectious particles” labeled “prions” by Stanley Prusiner in 1982 [4]. As emphasized by Prusiner at the time, the definition “does not prejudge the chemical composition of the scrapie prion except to state that it does contain a protein” [5]. Indeed, the new term “prion” was intended to be used without any preconceived claims about its biochemical composition and structure: “To avoid prejudging the structure of these infectious particles, three hypothetical structures for the prion were proposed: (1) proteins surrounding a nucleic acid that encodes them (a virus), (2) proteins surrounding a small noncoding polynucleotide, and (3) a proteinaceous particle devoid of nucleic acid.” [6].

Three decades later, prions were fundamentally redefined: “Prions are proteins that acquire alternative conformations that become self-propagating” [7]. However, as discussed below, this definition encompasses not only all amyloidogenic proteins but also many other proteins that assemble into ordered oligomeric structures.

Interestingly, the new definition of prions opened the door for perceiving them as ‘activities’ rather than as ‘physical entities.’ Indeed, as Claudia Scheckel and Adriano Aguzzi pointed out, “Although it is now generally accepted that the prion consists largely of the pathological aggregate of the prion protein, PrPSc, prions are defined as a biological activity rather than a physical entity” [8]. Considering also that the TSE ‘physical entities’ or ‘activities’ presumed to undergo self-replication are apparently distinct from ‘physical entities’ or ‘activities’ implicated in the TSE pathogenic mechanisms (yet to be clearly identified or labelled) [9-12], the definition of prions becomes even more confusing.

Nevertheless, as mentioned in the essay [13] and the comments above, under certain conditions, all proteins can acquire alternative conformations and assemble into amyloid fibers. Indeed, two decades ago, Christopher Dobson suggested that: “the ability to form amyloid is a generic property of polypeptide chains. This ability can readily be explained by the fact that the intermolecular bonds that stabilize this material involve the peptide backbone, which is common to all proteins” [14]. Therefore, if indeed prions are “proteins that acquire alternative conformations that become self-propagating” [7] then, under certain conditions, all proteins can become prions.

However, with some exceptions (e.g., secreted/surface components), natural proteins function under physiological conditions and, as Dobson suggested [14], most of them are unlikely to assemble into amyloidogenic structures, unless, as pointed out in the essay [13], they were selected for this propensity in connection with their biological function.

Interestingly, Peter Lansbury [15] and Dobson [14] proposed a protective function for the mature amyloids formed by the proteins implicated in neurodegenerative diseases: “Like the monomeric folding intermediates discussed above, a β-sheet-containing oligomeric species could bind tightly to any number of cellular targets, triggering, for example, a cytotoxic cascade (see Fig. 2). The smallest intermediate in which the binding site (possibly a β-sheet) is stable would have the greatest specific activity (moles binding site per mole protein). Thus, polymerization would compete with binding and specific activity would decrease as polymerization continued. In fact, the fibril itself actually may be protective; fibrillization would be an efficient way for the cell to sequester potentially toxic protofibrils. This proposal has ramifications for the design of screens for discovery of candidate therapeutic agents, because it suggests that some of the compounds that inhibit fibril formation actually could produce a deleterious effect by causing accumulation of a prefibrillar toxic species” (emphasis added; [15]).

This statement should resonate now stronger than ever with the field of neurodegenerative diseases, particularly Alzheimer’s disease (AD), as some of the putative therapeutic agents interfere with the formation of the plaques and tangles. Nevertheless, the hypothesis proposed by Lansbury and Dobson was advanced in the context of the protein misfolding dogma and the prion hypothesis, in which the assembly of PrP, Aβ, tau, α-Syn, and TDP-43 into various oligomeric/fibrous aggregates was considered an accidental event not related to their biological function. Therefore, it was difficult to envision the self-assembly of these proteins into mature amyloids as a selected adaptive feature. In the innate immunity hypothesis, the formation of diverse oligomeric innate immunity complexes (IICs) with putative toxic properties explains, or even entails, the evolution of regulatory and protective mechanisms, such as their sequestration into inert amyloids [13, 16]. Thus, in this hypothesis, the protective function of amyloids is rooted in a compelling evolutionary rationale.

The list of functional amyloids is growing [17-23], and there is strong evidence that PrP, Aβ, tau, α-Syn, and TDP-43 are members of the innate immune system (discussed in [13, 16]). However, as pointed out in the comments, several proteins with well-defined non-immunological functions, such as insulin, transition into amyloidogenic states under physiological, albeit stressful, conditions. Likely, the propensity of insulin to form amyloids is related to its innate property to assemble into functional ‘storage oligomers,’ akin to the genuine hormone storage amyloids (e.g., [17]), whose relatively inert amyloid status represents an evolutionary adaptive feature.

As mentioned above, even some non-amyloidogenic proteins apparently “acquire alternative conformations that become self-propagating” under physiological conditions and, therefore, fulfill the new definition of prions. Indeed, some of the most fascinating innate immunity proteins, such as MAVS and ASC, assemble into antiviral innate immunity complexes (i.e., IICs) that amplify and transduce pathogen-associated signals to downstream effector pathways. Given that the assembly of these proteins into IICs is clearly related to their biological function, their classification as “gain-of-function prions” [24] adds another confusing dimension to the prion hypothesis. Clearly, the assembly of MAVS and ASC into IICs is NOT a “gain-of-function”, but their evolutionary selected biological function, which makes them intriguing models for the putative function of PrP, Aβ, tau, α-Syn, and TDP-43 in innate immunity.

Are there other more mundane proteins that could be defined as prions under their new definition? Consider, for example, the following excerpt from a decade-old Alzforum article “Mice Tell Tale of Tau Transmission, Alzheimer’s Progression”, in which Bradley Hyman fails to see a fundamental difference between the proteins labeled prions, or prion-like, and more ordinary proteins, such as hemoglobin: “While researchers believe that misfolded, toxic forms of tau can corrupt normal tau molecules in a process called templated protein misfolding, “the notion that there is templated protein alteration is not the same as an infection,” Hyman told ARF. He noted that any protein in the body that forms an oligomer undergoes templated folding. This would include many proteins, such as, for example, hemoglobin” (quotation from [25]).

There is no stronger argument for the prion paradigm than the concept of templated protein (mis)folding, which is foundational for explaining the ‘strains’ phenomenon associated with the TSEs and other neurodegenerative diseases, including AD, PD, and ALS. The existence of TSE strains with distinct disease phenotypes was used for decades as the primary argument for the viral etiology of TSEs, and against the protein-only paradigm and prion hypothesis. Remarkably, the ‘strains’ phenomenon is now used as one of the main arguments for the existence of prions as distinct biological entities carrying ‘hereditary information,’ which presumably differentiate them from other amyloidogenic proteins.

Although the prions’ so-called hereditary information, conferring the prion strains their propagation and clinicopathological specificity, was thought for decades to be encoded in distinct misfolded PrP conformations, direct evidence supporting this presumption has only been reported recently [26-28] . Nevertheless, as exciting as the cryogenic electron microscopy (cryo-EM) studies providing this evidence are, it is not clear at this time if the conformational differences between the strains (e.g., 263K vs. RML prions) were dictated by sequence differences in the parental PrP molecules versus conformational templating. Additionally, it is not known whether non-PrP factors participated in the assembly of the ex vivo fibrils used in these cryo-EM studies; the apparent participation of essential cofactors, including RNA molecules, in the formation of infectious prions has been well documented ([29]; for putative roles of nucleic acids in the ‘life cycle’ of amyloidogenic pathogens see [30-34]). More importantly, though, all these structural findings must be evaluated in the context of the well-established but rarely articulated fact that the TSEs, as well as the entities called prions (whatever they are), can arise spontaneously in the absence of prion-based hereditary information or templating activity [35, 36]. In the context of the innate immunity model, the ‘hereditary information’ and ‘templating activity’ of prions represent a conceptual mirage rooted in the misleading protein misfolding dogma and the prion hypothesis [13, 16].

The generation of synthetic prions from recombinant PrP has been regarded as the ultimate proof for the prion hypothesis and the protein-only paradigm. The results of these studies have been mixed and often confusing, not least because of the problems with defining prions, as recently articulated by scientists from the MRC Prion Unit at UCL, Institute of Prion Diseases: “After all, to be able to determine if a synthetic prion has been made, one must first know what a prion is or, at the very least, what a prion is not” (emphasis added; [37]). Clearly, defining prions is not a frivolous academic undertaking tangential to the experimental work, but a foundational rationale for planning, conducting, and making sense of the laboratory studies. At this time, though, it is reasonable to assume that, after forty years, the definition of prions will continue to ‘shift and drift,’ to use a recent take on the ‘science of prions’ [38]. As proposed next, a reasonable solution might be to reevaluate the relevance of the prion hypothesis and prion paradigm for explaining the etiology of TSEs and other neurodegenerative diseases, and to reconsider the need for defining prions.

As outlined in the essay, the prion hypothesis was formulated in context of the century-old, misleading dogma of viruses as virus particles [13, 16, 34]. Virus particles are highly specialized structures produced by some, but not all, viruses for their transmission to new host cells; therefore, based this fact alone, identifying viruses with the virus particles and defining them based on their physical and biochemical properties is flawed. Viruses pass in their life cycle through multiple stages, each with distinct physical characteristics, biochemical composition, and biological properties; thus, an integrated sum of all these stages and their characteristics define viruses [39, 40]. Similarly, identifying the TSEs pathogens physically and conceptually with their transmissible entities is misleading and, by analogy with to the new perspective on viruses, the ‘TSE pathogens’ should integrate all aspects of their ‘life cycle’ or more appropriately, their ‘amyloidogenic cycle.’

As Kariem Ezzat emphasized and as Jesús Requena pragmatically narrated in their comments: “I think that the experimental evidence supporting that PrPSc prions facilitate conversion of PrPC to PrPSc is overwhelming. And there is nothing strange about it: it happens with all seeded amyloid conversions. Incubate any protein under denaturing conditions and slightly acidic pH at 37C and, as Dobson has shown, it will eventually adopt an amyloid conformation; repeat the experiment adding a bit of your pre-formed amyloid as a seed, and the process will be much shorter. I do not see anything unusual there”.  I completely agree. There is nothing fundamentally unusual about the assembly and aggregation process of PrP, Aβ, tau, α-Syn, and TDP-43 into oligomeric structures and amyloid fibers that would differentiate them from amyloidogenic cycles that all proteins can enter, albeit very few under physiological conditions. And if that’s the case, there is no need for the prion hypothesis, for the prion paradigm, or for defining prions.


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Prions, propagative amyloids, prionoids, prion-like agents...

This opens a can of worms...I recommend the enlightening paper: "The Prion 2028 round tables (II): Abeta, tau, alpha-synuclein: are they prions, prion-like proteins or what...? (Prion, 2019, 13:41-45) in which my colleague and friend Hasier Eraña summarizes a round table he chaired during the Prion 2018 international prion meeting held in Santiago de Compostela on this subject., with participation of Stanley Prusiner, Claudio Soto, Mathias Jucker and Corinne Lasmezas among other prominent experts, not forgetting the audience. There even was an informal poll on the subject, which, contrary to the predominant opinion of the table members, showed a majority of researchers stating that Amyloid beta, tau etc. are "prion-like agents".

In my opinion, one has to stick to the original definition provided by prusiner: prions are infectious agents composed solely of protein. Now, what does "infectious" mean? Is a cancer that one passages from nude mouse to nude mouse an infectious agent? Yes, but only under very articicial circumstances...However, the oral cancer of Tasmanian devils does seem to qualify without restrictions, as it propagates between Tasmanian devils under completely natural circumstances... Is TMV infectious? Yes, but only to tobacco plants...So even the most basic part of the definition may be complicated. In my humble opinion (at least at this moment, I have changed back and forth many times...) to qualify as a prion a protein ensemble must be able to propagate under "relatively" normal conditions, meaning orally, or through an open sore or wound, or iatrogenically, and between wt individuals. This excludes Abeta and tau (unless until recent findings on apparent transmission of Abeta to recipients of contaminated growth hormone, leading to death as a consequence of Alzheimer´s disease are fully confirmed. 

About the first part of the definition "composed solely of protein", I can attest to the fact (because I do it all the time in my lab) that a mixture of recombinant PrP, dextran and detergent in an inorganic buffer, subject to a simple protocol of agitation, can become infectious. So in my experience prions (protein assemblies that are infectious by themselves) do exist.

But of course this is a fascinating subject and there are many opinions.  


Even prion advocate Kane had to admit:

Although there is little indication of chronic inflammation in the injected mice, the participation of an infectious microorganism in promoting the amyloidosis cannot yet be definitively ruled out.”



The theory surrounding neurologist Stanley Prusiner’s “prions”, a word which he himself coined for gene-less proteins that were infectious, was under a rightful cloud of suspicion from its onset. In fact, from the get go Prusiner’s prion theory was felt to be heretic, unorthodox, and contrary to accepted belief.

And the only real evidence that discoverer Stanley B. Prusiner had in his original paper was that the disease agent behind “Scrapie” was devoid of DNA or RNA - was because he couldn’t find any. Nevertheless, fueled by U.S. National Institutes of Health grants which since 1975 fed him in excess of $56 million, Prusiner began his research — working, at first on obscure diseases thought to be caused by “slow viruses”.  Prusiner would in effect rename them. “Prion” said Prusiner, “is a terrific word. It’s snappy. It’s easy to pronounce. People like it. It isn’t easy to come up with a good word in biology. One hell of a lot of bad words people introduce get thrown away.” [1]

The prion story is one that illustrates in particular how deep a role public relations can play in the scientific community. Prusiner launched the prion hypothesis in 1982, after realizing that he didn't have the necessary funding to do the meticulous and extensive experiments that might unambiguously identify the agents of these strange diseases. Eighteen years later, he still had yet to do the experiments that would offer compelling proof of the existence of prions. Perhaps Prusiner lost his motivation to do so once he started getting extraordinary amounts of press simply on the basis of his claim. (By 1986, The New York Times had seen fit to mention prions in 21 separate articles, and Reader's Digest had named prions "Killer Diseases from the Dawn of Time.") Why didn’t anyone else do the experiments? Because they remained frighteningly expensive and time-consuming—and because Prusiner got more than twice the funding of all the other U.S. researchers in the field combined.

So by April, 1982 he announced that the real culprit behind such diseases as scrapie in sheep and goats, kuru in cannibals, Creutzfeldt–Jakob disease (CJD) in humans, and  chronic wasting  disease in deer and elk was either a virus—not yet isolated—or some rogue infectious protein-only “prion”, which unlike  anything yet known could multiply, and infect—without genes. Thus, right from the beginning, it was obvious that Prusiner’s plans for the eventual dominance of prion terminology would be based, in large part, on either/or ambiguity. Such vagueness did not go unnoticed or without the criticism of several investigators [2, 3]. Carp mentions in The Journal of General Virology that “The attempt to subsume within the single term, prion, both the 'protein only' and the 'protein with nucleic acid' concepts, has made it difficult to engage in precise dialogue about the term [PRION].” [4]

Of course it did. That was exactly what was intended. The term prion was being fashioned by Prusiner to mean all things to all people. It you, for some reason, agreed with Prusiner’s protein-only hypothesis—you would be satisfied; and if you did not believe in Prusiner’s “snappy” prions—there was something in there for you as well. And if by chance you sat on the fence of uncertainty, Stanley had something for you as well. In the meantime the term’s credibility increased with each and every one of its utterances, making it harder to challenge with each passing day. Perhaps Manuelides best summed it up in Lancet as “the peculiarly American sport of betting on popular momentum.” [5]

Furthermore, from the onset, Prusiner also had Alzheimer’s and Parkinsons in his sights. In his April, 1982 announcement, he staked his sweeping territorial claim:

A knowledge of the molecular structure of prions may help identify the etiologies of some chronic degenerative diseases of humans. Development of sensitive probes for detecting prions in such diseases is needed. Diseases where prions might play an etiological role include Alzheimer's senile dementia, multiple sclerosis, Parkinson's disease, amyotrophic lateral sclerosis, diabetes mellitus, rheumatoid arthritis, and lupus erythematosus, as well as a variety of neoplastic disorders

That is if there really was such a thing as a PRION to begin with.

A key strategy, according to colleague Dave Bolton, who worked with Prusiner was: “If we coin a new term for it [prion], and go out and tell people of the potential link to Alzheimer’s, we’re going to draw people’s attention to this. And we’re going to get money.” [1] Yet another postdoctoral researcher in Prusiner’s lab, Paul Bendheim said that Prusiner “rammed that word [prion] down the throats of everybody in that laboratory and in the world.” [1]

All things considered, Prusiner’s idea wasn’t new. The thoughts of biologists in the 1930s who had incorrectly said that viruses were only proteins and that ‘slow viruses’ might be geneless had been proposed by and  discarded in Britain by Griffith as early as 1967 [6]. Prusiner’s rejected grant application from February 18, 1980 was for “Slow Viruses Causing Degenerative Diseases.”  Prion advocates, salvaging what remained from the experience, put forth that prions were way smaller than other viruses, without the capacity to carry genes. Yet, the biology of scrapie alone, because of its many strains, spoke for a cause with genes [7].

At every twist and turn in discovery Prusiner and colleagues, showed chameleon-like properties, camouflaging their theory with endless color combinations to fit new findings. For example, when his colleagues identified the host protein involved in the diseases under implication, they named it prion protein or PrP, and the gene that codes for it—prion protein gene. Never mind that this gene did not agree with the original hypothesis—just use “snappy” prion lexicon to self-inoculate your theory from criticism, especially if buttressed with an overarching  system of classification that included “prion science”, “prion biology” and “prion diseases’. Such classification, implied authority, suggesting an aura of inevitability for the prion hypothesis.

But in reality, “prions” were not, and never have been the only game in town.

By 1984, Rower, at the National Institute of Health, showed that despite what prion advocates were claiming,  prions were the size of small viruses, with plenty of room for genetic material [8]. In the same study Rohwer attacked the prion purists supposition that prions were immortal, citing  agents of  potential damage to them. [ibid] Early prion  workers used a test called the “incubation-time assay” to judge prion purification, utilized cautiously in England since the 1960s. A modification had cut the time to score this assay from a year to a couple of months. Rohwer commented that this test was enormously less accurate than traditional methods and that purifications using it could be off by a factor of from 100 to 1000. Such knowledge kept mad cow’s viral and bacterial theories alive.

The finding that prions were proteins normally found in the body, including the brain of healthy controls, also seemed to contradict the best evidence that they were infectious. The theory, like all questionable science, survived by finding a difference: prions from healthy animals, were quickly claimed to be “cellular” protein; those from scrapie were “scrapie” protein. Scrapie protein aggregated into rods while cellular protein did not. Another so-called “critical clue” [9]: scrapie protein survived  proteinase, while “cellular” did not. This still did not mean however that some virus or bacteria did not cause the change being attributed to prions to begin with. Cow tuberculosis [Mycobacterium bovis], for example, both by virtue of its cell-wall-deficient, virus-like forms, and that it shares  methyllysines with  other mycobacteria is also protease-resistant [10]. The amyloid proteins in Alzheimer’s, not at first linked to prions, were known at that time to be also protease resistant [11].

Healthy “cellular” prions  also remained  a mystery, but they need  not have  been. Prions  are  amyloid  and  it  was common  knowledge way before the word prion ever existed that there is a soluble serum protein component [SAP] of amyloid in healthy blood, its purpose also unknown [12]. What was clear is that the role of deposited amyloid fibrils, once formed, is to disrupt, destroy, and compromise, whether in mad cow, JCD, Scrapie, Alzheimer’s or any of the  other amyloid provoked degenerative disorders.

Prion theorists further elaborated that although proteins normally fold into three-dimensional states, protein prions sometimes ‘misfolded’, assuming an incorrect, infectious state, which somehow subsequently changed surrounding cellular protein into itself, setting off an infectious chain  reaction. And since the  damage done by prion protein seemed similar to the malfunctioning proteins in Alzheimer’s and even Parkinson’s—yes, these too  might  be caused by prions.

Critics pointed out that despite the vast expenditure [13] on research geared to verify that prions cause mad cow and the spongiform encephalopathies,—to this point prions have not been established fully as the cause of any disease. Furthermore, it remained unclear how prions destroyed brain tissue. Also, experts  pointed out, prion investigators had not proven that  protein-only-prions—even if amplified over 100-fold from an infected brain—increased infectivity. Such criticism represents perhaps the  best kept  secret of prion  researchers [14, 15].

Both CJD and scrapie can be transmitted without prions [16]. Also, brain  material from which prions and their antibodies have been removed, can still infect animals. Moreover, prions have been found in completely unrelated disease processes, such as Kawsaski syndrome and inclusion body myositis. Finally, although there were many strains of “prion”  diseases, there is no credible theory as  to  how  these strains  exist without genetic material. Aguzzi points out that abnormal prions  have  exactly the same amino acid structure as nonpathogenic prions, found  in everyone. How could  prion  proteins, then  be claimed to do what they are claimed to do [17]?

By all logical estimates, the death-knell to the prion hypothesis should have occurred with Lasmezas’s 1997 interspecies  transmission of mad cow in which more than  half of injected mice had no detectable prions [16]. If this was not enough, then there was Manuelidis’s 2002 [18] study on infectious neurons (microglia) with low prion levels in otherwise highly infectious material, which only served to support the concept that pathologic prions were the result of infection rather than  being  the  actual infectious agent. To Manuelidis  this  was  likely to be  a  virus,  although she admitted the  fundamental mystery remained. In fact, to many dissenters, some other, not as yet identified  pathogen  such as a virus or bacteria [or a mycobacteria] caused “prions” to  misfold—thus damaging the brain.

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CWD Tuberculosis Found in Spongiform Disease Formerly Attributed to Prions: Its Implication towards Mad Cow Disease, Scrapie and Alzheimer’s.

Citation: Lysenko AP, Broxmeyer L MD, Vlasenko VV, et al. CWD Tuberculosis Found in Spongiform Disease Formerly Attributed to Prions: Its Implication towards Mad Cow, Disease, Scrapie and Alzheimer’s J Mol Path Epidemol. 2017, 3:2


The TSE’S or transmissible spongiform encephalopathies, include bovine spongiform encephalopathy (also called BSE or “mad cow disease”), Creutzfeldt–Jakob disease (CJD) in humans, and “scrapie” in sheep or goats (caprine spongiform encephalopathy). They remain a mystery, their cause still hotly debated. Current mad cow diagnosis lies solely in the detection of late appearing “prions”, an acronym for hypothesized, gene-less, misfolded proteins, somehow claimed to cause the disease. Yet laboratory preparations of prions contain other things, which could include unidentified bacteria or viruses. And the only real evidence that prion originator Stanley Prusiner had in his original paper that the disease agent behind “Scrapie” in sheep and goats was devoid of DNA or RNA– was based upon the fact that he couldn’t find any. Furthermore, the rigors of prion purification alone, might, in and of themselves, have killed any causative microorganism and Heino Dringer, who did pioneer work on their nature, candidly predicts “it will turn out that the prion concept is wrong.” Roels and Walravens as well as Hartly traced Mad Cow to Mycobacterium bovis. Moreover, epidemiologic maps of the origins and peak incidence of Mad Cow in the UK, suggestively match those of England’s areas of highest bovine tuberculosis, the Southwest. The neurotoxic potential of bovine tuberculosis has for some time been well known. By 1911 Alois Alzheimer called attention to “a characteristic condition of the cortical issue which Fischer referred to as ‘spongy cortical wasting” in Alzheimer’s disease (AD). But behind AD, Fischer suspected a microbe called Streptothrix which was constantly being mistaken and confused for tuberculosis. Our present investigation of the TSEs clearly shows cell-wall-deficient (CWD) tubercular mycobacteria present, verified by molecular analysis, ELISA, PCR and microscopy to cause spongiform encephalopathy.

Keywords: Prions; Scrapie; The Spongiform Encephalopathies; Alzheimer’s disease;The eiology of Alzheimer’s Disease; Mycobacterium tuberculosis Complex