Institute of Neuropathology, University Hospital of Zurich, Zurich, Switzerland
Transmissible spongiform encephalopathies (TSEs) are inevitably lethal neurodegenerative diseases that affect humans ...and a large variety of animals. The infectious agent responsible for TSEs is the prion, an abnormally folded and aggregated protein that propagates itself by imposing its conformation onto the cellular prion protein (PrP C ) of the host. PrP C is necessary for prion replication and for prion-induced neurodegeneration, yet the proximal causes of neuronal injury and death are still poorly understood. Prion toxicity may arise from the interference with the normal function of PrP C , and therefore, understanding the physiological role of PrP C may help to clarify the mechanism underlying prion diseases. Here we discuss the evolution of the prion concept and how prion-like mechanisms may apply to other protein aggregation diseases. We describe the clinical and the pathological features of the prion diseases in human and animals, the events occurring during neuroinvasion, and the possible scenarios underlying brain damage. Finally, we discuss potential antiprion therapies and current developments in the realm of prion diagnostics.
Reduction of native prion protein (PrP) levels in the brain is an attractive strategy for the treatment or prevention of human prion disease. Clinical development of any PrP-reducing therapeutic will ...require an appropriate pharmacodynamic biomarker: a practical and robust method for quantifying PrP, and reliably demonstrating its reduction in the central nervous system (CNS) of a living patient. Here we evaluate the potential of ELISA-based quantification of human PrP in human cerebrospinal fluid (CSF) to serve as a biomarker for PrP-reducing therapeutics. We show that CSF PrP is highly sensitive to plastic adsorption during handling and storage, but its loss can be minimized by the addition of detergent. We find that blood contamination does not affect CSF PrP levels, and that CSF PrP and hemoglobin are uncorrelated, together suggesting that CSF PrP is CNS derived, supporting its relevance for monitoring the tissue of interest and in keeping with high PrP abundance in brain relative to blood. In a cohort with controlled sample handling, CSF PrP exhibits good within-subject test–retest reliability (mean coefficient of variation, 13% in samples collected 8–11 wk apart), a sufficiently stable baseline to allow therapeutically meaningful reductions in brain PrP to be readily detected in CSF. Together, these findings supply a method for monitoring the effect of a PrP-reducing drug in the CNS, and will facilitate development of prion disease therapeutics with this mechanism of action.
The cellular prion protein (PrPC) has a C-terminal globular domain and a disordered N-terminal region encompassing five octarepeats (ORs). Encounters between Cu(II) ions and four OR sites produce ...interchangeable binding geometries; however, the significance of Cu(II) binding to ORs in different combinations is unclear. To understand the impact of specific binding geometries, OR variants were designed that interact with multiple or single Cu(II) ions in specific locked coordinations. Unexpectedly, we found that one mutant produced detergent-insoluble, protease-resistant species in cells in the absence of exposure to the infectious prion protein isoform, scrapie-associated prion protein (PrPSc). Formation of these assemblies, visible as puncta, was reversible and dependent upon medium formulation. Cobalamin (Cbl), a dietary cofactor containing a corrin ring that coordinates a Co3+ ion, was identified as a key medium component, and its effect was validated by reconstitution experiments. Although we failed to find evidence that Cbl interacts with Cu-binding OR regions, we instead noted interactions of Cbl with the PrPC C-terminal domain. We found that some interactions occurred at a binding site of planar tetrapyrrole compounds on the isolated globular domain, but others did not, and N-terminal sequences additionally had a marked effect on their presence and position. Our studies define a conditional effect of Cbl wherein a mutant OR region can act in cis to destabilize a globular domain with a wild type sequence. The unexpected intersection between the properties of PrPSc's disordered region, Cbl, and conformational remodeling events may have implications for understanding sporadic prion disease that does not involve exposure to PrPSc.
The misfolding and aggregation of specific proteins is a common hallmark of many neurodegenerative disorders, including highly prevalent illnesses such as Alzheimer's and Parkinson's diseases, as ...well as rarer disorders such as Huntington's and prion diseases. Among these, only prion diseases are 'infectious'. By seeding misfolding of the PrP(C) (normal conformer prion protein) into PrP(Sc) (abnormal disease-specific conformation of prion protein), prions spread from the periphery of the body to the central nervous system and can also be transmitted between individuals of the same or different species. However, recent exciting data suggest that the transmissibility of misfolded proteins within the brain is a property that goes way beyond the rare prion diseases. Evidence indicates that non-prion aggregates tau, α-syn (α-synuclein), Aβ (amyloid-β) and Htt (huntingtin) aggregates can also move between cells and seed the misfolding of their normal conformers. These findings have enormous implications. On the one hand they question the therapeutical use of transplants, and on the other they indicate that it may be possible to bring these diseases to an early arrest by preventing cell-to-cell transmission. To better understand the prion-like spread of these protein aggregates it is essential to identify the underlying cellular and molecular factors. In the present review we analyse and discuss the evidence supporting prion-like spreading of amyloidogenic proteins, especially focusing on the cellular and molecular mechanisms and their significance.
Recombinant C-terminally truncated prion protein PrP23-144 (which corresponds to the Y145Stop PrP variant associated with a Gerstmann–Sträussler–Scheinker-like prion disease) spontaneously forms ...amyloid fibrils with a parallel in-register β-sheet architecture and β-sheet core mapping to residues ∼112–139. Here we report that mice (both tga20 and wild type) inoculated with a murine (moPrP23-144) version of these fibrils develop clinical prion disease with a 100% attack rate. Remarkably, even though fibrils in the inoculum lack the entire C-terminal domain of PrP, brains of clinically sick mice accumulate longer proteinase K-resistant (PrPres) fragments of ∼17–32 kDa, similar to those observed in classical scrapie strains. Shorter, Gerstmann–Sträussler–Scheinker-like PrPres fragments are also present. The evidence that moPrP23-144 amyloid fibrils generated in the absence of any cofactors are bona fide prions provides a strong support for the protein-only hypothesis of prion diseases in its pure form, arguing against the notion that nonproteinaceous cofactors are obligatory structural components of all infectious prions. Furthermore, our finding that a relatively short β-sheet core of PrP23-144 fibrils (residues ∼112–139) with a parallel in-register organization of β-strands is capable of seeding the conversion of full-length prion protein to the infectious form has important implications for the ongoing debate regarding structural aspects of prion protein conversion and molecular architecture of mammalian prions.
The normal cellular isoform of prion protein, designated PrP
, is constitutively converted to the abnormally folded, amyloidogenic isoform, PrP
, in prion diseases, which include Creutzfeldt-Jakob ...disease in humans and scrapie and bovine spongiform encephalopathy in animals. PrP
is a membrane glycoprotein consisting of the non-structural
-terminal domain and the globular C-terminal domain. During conversion of PrP
to PrP
, its 2/3 C-terminal region undergoes marked structural changes, forming a protease-resistant structure. In contrast, the N-terminal region remains protease-sensitive in PrP
. Reverse genetic studies using reconstituted PrP
-knockout mice with various mutant PrP molecules have revealed that the N-terminal domain has an important role in the normal function of PrP
and the conversion of PrP
to PrP
. The N-terminal domain includes various characteristic regions, such as the positively charged residue-rich polybasic region, the octapeptide repeat (OR) region consisting of five repeats of an octapeptide sequence, and the post-OR region with another positively charged residue-rich polybasic region followed by a stretch of hydrophobic residues. We discuss the normal functions of PrP
, the conversion of PrP
to PrP
, and the neurotoxicity of PrP
by focusing on the roles of the N-terminal regions in these topics.
Recent reports indicate that a growing number of intracellular proteins are not only prone to pathological aggregation but can also be released and “infect” neighboring cells. Therefore, many complex ...diseases may obey a simple model of propagation where the penetration of seeds into hosts determines spatial spread and disease progression. We term these proteins prionoids, as they appear to infect their neighbors just like prions—but how can bulky protein aggregates be released from cells and how do they access other cells? The widespread existence of such prionoids raises unexpected issues that question our understanding of basic cell biology.
Human prion diseases (HPDs) are fatal neurodegenerative disorders characterized by abnormal prion proteins (PrPSc). However, the detection of prion seeding activity in patients with high sensitivity ...remains challenging. Even though real-time quaking-induced conversion (RT-QuIC) assay is suitable for detecting prion seeding activity in a variety of specimens, it shows lower accuracy when whole blood, blood plasma, and blood-contaminated tissue samples are used. In this study, we developed a novel technology for the in vitro amplification of abnormal prion proteins in HPD to the end of enabling their detection with high sensitivity known as the enhanced quaking-induced conversion (eQuIC) assay.
Three antibodies were used to develop the novel eQUIC method. Thereafter, SD50 seed activity was analyzed using brain tissue samples from patients with prion disease using the conventional RT-QUIC assay and the novel eQUIC assay. In addition, blood samples from six patients with solitary prion disease were analyzed using the novel eQuIC assay.
The eQuIC assay, involving the use of three types of human monoclonal antibodies, showed approximately 1000-fold higher sensitivity than the original RT-QuIC assay. However, when this assay was used to analyze blood samples from six patients with sporadic human prion disease, no prion activity was detected.
The detection of prion seeding activity in blood samples from patients with sporadic prion disease remains challenging. Thus, the development of alternative methods other than RT-QuIC and eQuIC will be necessary for future research.
Foiling deadly prions Wadman, Meredith
Science (American Association for the Advancement of Science),
2024-Mar-22, Letnik:
383, Številka:
6689
Journal Article
Recenzirano
Can the course of fatal prion diseases be changed by removing the protein before it goes bad?
On passaging synthetic prions, two isolates emerged with incubation times differing by nearly 100 days. Using conformational-stability assays, we determined the guanidine hydrochloride (Gdn-HCI) ...concentration required to denature 50% of disease-causing prion protein ($PrP^{Sc}$) molecules, denoted as the $Gdn\cdotHCl_{1/2}$ value. For the two prion isolates enciphering shorter and longer incubation times, $Gdn\cdotHCl_{1/2}$ values of 2.9 and 3.7 M, respectively, were found. Intrigued by this result, we measured the conformational stabilities of 30 prion isolates from synthetic and naturally occurring sources that had been passaged in mice. When the incubation times were plotted as a function of the $Gdn\cdotHCl_{1/2}$ values, a linear relationship was found with a correlation coefficient of 0.93. These findings demonstrate that (i) less stable prions replicate more rapidly than do stable prions, and (ii) a continuum of $PrP^{Sc}$ structural states enciphers a multitude of incubation-time phenotypes. Our data argue that cellular machinery must exist for propagating a large number of different $PrP^{Sc}$ conformers, each of which enciphers a distinct biological phenotype as reflected by a specific incubation time. The biophysical explanation for the unprecedented plasticity of $PrP^{Sc}$ remains to be determined.
PDF
