Search Results (713)

The prion-forming regions (PFRs) of yeast prion proteins are usually located at either the N- or C-terminus of a protein. In the Sup35 prion, the main prion structure contains 71 N-terminal residues. Here, we investigated the importance of the terminal PFR location for prion properties. Two prionogenic sequences of 29 and 30 residues and two random sequences of 23 and 15 residues were added to the Sup35 N-terminus, making the original PFR internal. These proteins were overproduced in yeast with two variants of the Sup35 prion. Mapping of the prion-like structures of these proteins by partial proteinase K digestion showed that in most cases, the extensions acquired an amyloid fold, and, strikingly, the prion structure was no longer present or was substantially altered at its original location. The addition of two to five residues to the Sup35 N-terminus often resulted in prion instability and loss when the respective genes were used to replace chromosomal SUP35. The structures of yeast prions Mot3, Swi1, Lsb2, candidate prions Asm4, Nsp1, Cbk1, Cpp1, and prions based on scrambled Sup35 PFRs were mapped. The mapping showed that the N-terminal location of a QN-rich sequence predisposes to, but does not guarantee, the formation of a prion structure by it and that the prion structure located near a terminus does not always include the actual terminus, as in the cases of Sup35 and Rnq1. Full article

Located in the Rocky Mountains within the Arapahoe and Roosevelt National Forests, Colorado State University’s Mountain Campus in Pingree Park hosted the 24th Annual Rocky Mountain Virology Association’s meeting in 2024. A total of 165 participants, both regional and international, participated in the 3-day event, which consisted of 48 talks and 42 posters. These presentations discussed developments in prion research, current affairs, and novel tools in virology; investigated arboviruses and their vectors, as well as molecular foundations of viral interactions; and provided increased understanding of viral immunology and vaccines. This year’s Randall Jay Cohrs keynote presentation unveiled how viral infections disrupt intestinal homeostasis via Sting-dependent NK-kB signaling. This novel research demonstrated the importance of immunological pathways in the virus-induced disruption of homeostasis. Nested in the valley of the Rocky Mountains, participants could enjoy the fall colors and partake in hiking and fishing all while discussing science and networking amongst a variety of scientists. This report encapsulates selected presentations from the 24th Annual Rocky Mountain Virology Association meeting. Full article

The formation of self-perpetuating protein aggregates such as amyloids is associated with various diseases and provides a basis for transmissible (infectious or heritable) protein isoforms (prions). Many human proteins involved in the regulation of transcription contain potentially amyloidogenic regions. Here, it is shown that short N-terminal isoforms of the human protein PHC3, a component of the chromatin-modifying complex PRC1 (Polycomb repressive complex 1), can form prion-like aggregates in yeast assays, exhibit amyloid properties in the E. coli-based C-DAG assay, and produce detergent-resistant aggregates when ectopically expressed in cultured human cells. Moreover, aggregates of short isoforms can sequester the full-length PHC3 protein, causing its accumulation in the cytosol instead of the nucleus. The introduction of an aggregating short PHC3 isoform alters the transcriptional profile of cultured human cells. It is proposed that the aggregation of short isoforms is involved in the feedback downregulation of PRC1 activity, leading to more open chromatin configuration. Full article

Prion diseases are fatal neurodegenerative disorders affecting humans and animals, and the central pathogenic event is the conversion of normal prion protein (PrP^C) into the pathogenic PrP^Sc isoform. Previous studies have identified nanobodies that specifically recognize PrP^C and inhibit the PrP^C to PrP^Sc conversion in vitro. In this study, we investigated the potential for in vivo expression of anti-PrP^C nanobodies and evaluated their impact on prion disease. The coding sequences of three nanobodies were packaged into recombinant adeno-associated virus (rAAV) and were administered via intracerebroventricular (ICV) injection in newborn mice. We found that the expression of these nanobodies remained robust for over 180 days, with no observed detrimental effects. To assess their therapeutic potential, we performed ICV injections of nanobody-expressing rAAVs in newborn mice, followed by intracerebral prion inoculation at 5–6 weeks of age. One nanobody exhibited a small yet statistically significant therapeutic effect, extending survival time from 176 days to 184 days. Analyses of diseased brains revealed that the nanobodies did not alter the pathological changes. Our findings suggest that high levels of anti-PrP^C nanobodies are necessary to delay disease progression. Further optimization of the nanobodies, AAV vectors, or delivery methods is essential to achieve a significant therapeutic effect. Full article

N⁶-methyladenosine (m⁶A), the most common internal RNA modification in eukaryotes, plays a vital role in post-transcriptional regulation. The YT521-B homology (YTH) domain plays a pivotal role in the methylation-dependent recognition of m⁶A. In this study, we performed an in-depth analysis of the YTH domain-containing RNA-binding protein family in Taxus chinensis (T. chinensis), a species renowned for its rich content of taxol, a significant compound in cancer therapy. We identified and analyzed six YTH domain-containing proteins in T. chinensis, elucidating their phylogenetic relationships, conserved domain, gene structures, conserved motifs, and chromosomal locations. The prion-like domain analyses provided insights into their potential functions in liquid–liquid phase separation and mRNA metabolism. Quantitative tissue analysis revealed TcYTH1 as the most highly expressed gene among the six TcYTH members. Additionally, we investigated the expression profiles of TcYTH genes under various stress conditions, such as high light, ABA, and PEG treatments. The expression levels of all TcYTH genes changed significantly under stress, revealing their involvement in stress response mechanisms. Our research provides novel insights into the YTH genes family in T. chinensis, emphasizing their potential roles in growth regulation and stress tolerance. The identification and analysis of these genes lay the groundwork for future studies on their functional roles in plant biology. Full article

(1) Background: Parkinson’s disease (PD) is characterized by the pathological accumulation of α-synuclein (α-syn) containing Lewy bodies (LBs) and Lewy neurites (LNs) within neurons. Growing evidence indicates that α-syn may propagate throughout the nervous system in a manner similar to prion-like transmission. Extracellular vesicles (EVs) may contribute to this pathway. We and others have reported that ATP13A2/PARK9 deficiency results in decreased EVs while its overexpression leads to increased EV generation. For analyzing EV-mediated α-syn secretion in neighboring neurons, we planned to alter Atp13a2 levels in vivo. (2) Methods: Three months after inoculating mouse α-syn fibrils into the striatum of Atp13a2-null and wild-type mice, we stained brain sections with anti-phosphorylated α-syn antibodies and then quantified LBs/LNs. We also examined the effect of increased levels of ATP13A2 by injecting lentivirus carrying human ATP13A2. Finally, we used cultured astrocytes and microglia for α-syn uptake and release, which were mediated by EVs. (3) Results: While LBs/LNs were formed in the entire brains, no significant difference was observed in LB/LN formation between Atp13a2-deficient and wild-type mice. Interestingly, the overexpression of ATP13A2 led to decreased LB/LN formation in the entire brains. Microglia and astrocytes released EVs more than neurons. EVs released from microglia and astrocytes contained more α-syn PFFs than those from neurons. (4) Conclusions: These results suggest that enhanced EV secretion by increased ATP13A2 levels attenuate the spreading of α-syn in brains, suggesting a protective role of ATP13A2 in α-synucleinopathies Full article

Prion diseases are fatal neurodegenerative diseases that can be transmitted by infectious protein particles, PrP^Scs, encoded by the endogenous prion protein gene (PRNP). The origin of prion seeds is unclear, especially in non-human hosts, and this identification is pivotal to preventing the spread of prion diseases from host animals. Recently, an abnormally high amyloid propensity in prion proteins (PrPs) was found in a frog, of which the genetic variations in the PRNP gene have not been investigated. In this study, genetic polymorphisms in the PRNP gene were investigated in 194 Dybowski’s frogs using polymerase chain reaction (PCR) and amplicon sequencing. We carried out in silico analyses to predict functional alterations according to non-synonymous single nucleotide polymorphisms (SNPs) using PolyPhen-2, PANTHER, SIFT, and MutPred2. We used ClustalW2 and MEGA X to compare frog PRNP and PrP sequences with those of prion-related animals. To evaluate the impact of the SNPs on protein aggregation propensity and 3D structure, we utilized AMYCO and ColabFold. We identified 34 novel genetic polymorphisms including 6 non-synonymous SNPs in the frog PRNP gene. The hydrogen bond length varied at codons 143 and 207 according to non-synonymous SNPs, even if the electrostatic potential was not changed. In silico analysis predicted S143N to increase the aggregation propensity, and W6L, C8Y, R211W, and L241F had damaging effects on frog PrPs. The PRNP and PrP sequences of frogs showed low homology with those of prion-related mammals. To the best of our knowledge, this study was the first to discover genetic polymorphisms in the PRNP gene in amphibians. Full article

Parkinson’s disease (PD) is the second most common neurodegenerative disease in the world. Currently, PD is incurable, and the diagnosis of PD mainly relies on clinical manifestations. The central pathological event in PD is the abnormal aggregation and deposition of misfolded α-synuclein (α-Syn) protein aggregates in the Lewy body (LB) in affected brain areas. Behaving as a prion-like seeding, the misfolded α-syn protein can induce and facilitate the aggregation of native unfolded α-Syn protein to aggravate α-Syn protein aggregation, leading to PD progression. Recently, in a blood-based α-Syn seeding amplification assay (SAA), Kluge et al. identified pathological α-Syn seeding activity in PD patients with Parkin (PRKN) gene variants. Additionally, pathological α-syn seeding activity was also identified in sporadic PD and PD patients with Leucine-rich repeat kinase 2 (LRRK2) or glucocerebrosidase (GBA) gene variants. Principally, the α-Syn SAA can be used to detect pathological α-Syn seeding activity, which will significantly enhance PD diagnosis, progression monitoring, prognosis prediction, and anti-PD therapy. The significance and future strategies of α-Syn SAA protocol are highlighted and proposed, whereas challenges and limitations of the assay are discussed. Full article

Background: Creutzfeldt–Jakob disease (CJD) is a rare, fatal, and transmissible neurodegenerative disorder caused by prion proteins. Handling specimens from individuals with suspected or confirmed cases presents a safety challenge to hospital workers including clinical laboratory staff. As no national guidelines exist, the clinical pathology laboratory must establish protocols for handling these specimens to ensure sufficient protective measures. This study aims to explore how various medical institutions manage CJD specimens, as a first step toward developing standardized preanalytical protocols for safe specimen handling by health care professionals. Methods: An electronic survey was generated and disseminated to diplomats of the American Board of Clinical Chemistry and was posted on the Listserv platform of the American Society for Microbiology and the Artery forum of the Association for Diagnostics and Laboratory Medicine. The survey evaluated various procedures and precautions implemented, the nature of the specimens processed, and whether they are processed in-house or sent to reference laboratories. Results: A total of 49 responses were collected. Most respondents (64%) noted their laboratories process specimens with a clinical suspicion of CJD regardless of the level of suspicion, 13% handled specimens only if the degree of suspicion was low, and 16% did not process specimens in-house at all. Among those who process CJD specimens, practices varied greatly, including different levels of precautions, use of biological safety cabinets, aliquoting, disposal, and disinfection procedures. Conclusions: A lack of standardization across laboratories exists for the handling of specimens of patients with suspected CJD. This study summarizes the approaches reported by survey respondents, providing a rationale for developing protocols for the safe handling of these specimens and highlighting the need to develop uniform universal standardized processing procedures. Full article

Prion diseases, including Creutzfeldt–Jakob disease (CJD), are deadly neurodegenerative disorders characterized by the buildup of abnormal prion proteins in the brain. This accumulation disrupts neuronal functions, leading to the rapid onset of psychiatric symptoms, ataxia, and cognitive decline. The urgency of timely diagnosis for effective treatment necessitates the identification of strongly correlated biomarkers in bodily fluids, which makes our research crucial. In this study, we employed a fully automated multiplex ELISA (Ella^®) to measure the concentrations of 14-3-3 protein, total tau protein, and neurofilament light chain (NF-L) in cerebrospinal fluid (CSF) and serum samples from patients with prion disease and analyzed their link to disease prognosis. However, in North American and European cases, we did not confirm a correlation between NF-L levels and survival time. This discrepancy is believed to stem from differences in treatment policies and measurement methods between Japan and the United States. Nonetheless, our findings suggest that NF-L concentrations could be an early diagnostic marker for CJD patients with further enhancements. The potential impact of our findings on the early diagnosis of CJD patients is significant. Future research should focus on increasing the number of sCJD cases studied in Japan and gathering additional evidence using next-generation measurement techniques. Full article

Prion diseases, or transmissible spongiform encephalopathies (TSEs), are a group of invariably fatal neurodegenerative disorders. One of the candidate genes involved in prion diseases is the shadow of the prion protein (SPRN) gene. Raccoon dogs, a canid, are considered to be a prion disease-resistant species. To date, the genetic polymorphisms of the SPRN gene and the predicted protein structure of the shadow of prion protein (Sho) have not been explored in raccoon dogs. SPRN was amplified using polymerase chain reaction (PCR). We also investigated the genetic polymorphisms of SPRN by analyzing the frequencies of genotypes, alleles, and haplotypes, as well as the linkage disequilibrium among the identified genetic variations. In addition, in silico analysis with MutPred-Indel was performed to predict the pathogenicity of insertion/deletion polymorphisms. Predicted 3D structures were analyzed by the Alphafold2. We found a total of two novel synonymous single nucleotide polymorphisms and three insertion/deletion polymorphisms. In addition, the 3D structure of the Sho protein in raccoon dogs was predicted to resemble that of the Sho protein in dogs. This is the first study regarding the genetic and structural characteristics of the raccoon dog SPRN gene. Full article

N⁶-methyladenosine (m⁶A) is a widespread post-transcriptional modification of RNA in eukaryotes. The conserved YTH-domain-containing RNA binding protein has been widely reported to serve as a typical m⁶A reader in various species. However, no studies have reported the m⁶A readers in Ginkgo biloba (G. biloba). In this study, a systematic analysis of the m⁶A reader (YTH) gene family was performed on G. biloba, identifying 10 YTH genes in its genome. Phylogenetic analysis of protein-coding sequences revealed that YTH genes from G. biloba could be classified into two subgroups: GbDC1 and GbDC2 in GbDC and GbDF1-8 in GbDF, each with similar motifs and gene structures. In G. biloba, the predicated aromatic cage pocket of the YTH domains in the YTH gene family is uniformly composed of tryptophan residues (WWW). Subcellular localization experiments verified that GbDC1 is indeed localized in the nucleus, while GbDF1 is localized in both the nucleus and the cytoplasm. The expression patterns of the identified m⁶A reader genes showed a wide distribution but were tissue-specific. Most genes were highly expressed in leaves, followed by the stem, while the lowest expression tendency was found in the roots. Cis-regulatory element analysis predicted the possible functions of YTH genes in G. biloba, which were mainly responsive to plant hormones such as ABA and MeJA, as well as stress responses. Furthermore, the expression levels of YTH genes indeed changed significantly after ABA, MeJA, and NaCl treatments, suggesting that they can be affected by these abiotic factors. In addition, the PLAAC prediction results indicate that prion domains exist in GbDF1, GbDF2, GbDF3, GbDF4, GbDF6, GbDF7, GbDF8, and GbDC1, and phase separation is possible. This study provides a foundation for further investigation of the effects of m⁶A methylation on gene expression regulation in G. biloba and other forest trees. Full article

The brain contains many interconnected and complex cellular and molecular mechanisms. Injury to the brain causes permanent dysfunctions in these mechanisms. So, it continues to be an area where surgical intervention cannot be performed except for the removal of tumors and the repair of some aneurysms. Some agents that can cross the blood–brain barrier and reach neurons show neuroprotective effects in the brain due to their anti-apoptotic, anti-inflammatory and antioxidant properties. In particular, some agents act by reducing or modulating the accumulation of protein aggregates in neurodegenerative diseases (Alzheimer’s disease, Parkinson’s disease, Huntington’s disease, Amyotrophic lateral sclerosis, and prion disease) caused by protein accumulation. Substrate accumulation causes increased oxidative stress and stimulates the brain’s immune cells, microglia, and astrocytes, to secrete proinflammatory cytokines. Long-term or chronic neuroinflammatory response triggers apoptosis. Brain damage is observed with neuronal apoptosis and brain functions are impaired. This situation negatively affects processes such as motor movements, memory, perception, and learning. Neuroprotective agents prevent apoptosis by modulating molecules that play a role in apoptosis. In addition, they can improve impaired brain functions by supporting neuroplasticity and neurogenesis. Due to the important roles that these agents play in central nervous system damage or neurodegenerative diseases, it is important to elucidate many mechanisms. This review provides an overview of the mechanisms of flavonoids, which constitute a large part of the agents with neuroprotective effects, as well as vitamins, neurotransmitters, hormones, amino acids, and their derivatives. It is thought that understanding these mechanisms will enable the development of new therapeutic agents and different treatment strategies. Full article

This study evaluated influenza A virus (IAV) detection and genetic diversity over time, specifically at the human–swine interface in breeding and nursery farms. Active surveillance was performed monthly in five swine farms in the Midwest United States targeting the employees, the prewean piglets at sow farms, and the same cohort of piglets in downstream nurseries. In addition, information was collected at enrollment for each employee and farm to assess production management practices, IAV vaccination status, diagnostic procedures, and biosecurity. Farm employee and swine samples were screened by IAV reverse transcription real-time polymerase chain reaction (RT-rtPCR), followed by IAV subtyping RT-rtPCR and whole genome sequencing on PCR-positive samples. This study showed higher positivity of IAV RNA detection in nursery pigs compared to prewean pigs, and more whole genome sequences were also obtained in the nursery phase. Surveillance of farm employees revealed two detections of H3N2 representing the 2022–2023 human IAV season, confirming the presence of influenza in farm employees while present at work, and thus highlighting the importance of biosecurity measures at the human–swine interface. This study highlights the importance of routine active surveillance to understand the dynamics of IAV at the farm level in both farm employees and swine. Full article

In this review, we focus on the biophysical and structural aspects of the oligomeric states of physiologically intrinsically disordered proteins and peptides tau, amyloid-β and α-synuclein and partly disordered prion protein and their isolations from animal models and human brains. These protein states may be the most toxic agents in the pathogenesis of Alzheimer’s and Parkinson’s disease. It was shown that oligomers are important players in the aggregation cascade of these proteins. The structural information about these structural states has been provided by methods such as solution and solid-state NMR, cryo-EM, crosslinking mass spectrometry, AFM, TEM, etc., as well as from hybrid structural biology approaches combining experiments with computational modelling and simulations. The reliable structural models of these protein states may provide valuable information for future drug design and therapies. Full article