Sign in to use this feature.

Years

Between: -

Subjects

remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline

Journals

remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline

Article Types

Countries / Regions

remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline

Search Results (579)

Search Parameters:
Keywords = Rnase2

Order results
Result details
Results per page
Select all
Export citation of selected articles as:
24 pages, 2655 KiB  
Article
Ribosomal RNA-Specific Antisense DNA and Double-Stranded DNA Trigger rRNA Biogenesis and Insecticidal Effects on the Insect Pest Coccus hesperidum
by Vol Oberemok, Nikita Gal’chinsky, Ilya Novikov, Alexander Sharmagiy, Ekaterina Yatskova, Ekaterina Laikova and Yuri Plugatar
Int. J. Mol. Sci. 2025, 26(15), 7530; https://doi.org/10.3390/ijms26157530 - 4 Aug 2025
Abstract
Contact unmodified antisense DNA biotechnology (CUADb), developed in 2008, employs short antisense DNA oligonucleotides (oligos) as a novel approach to insect pest control. These oligonucleotide-based insecticides target pest mature rRNAs and/or pre-rRNAs and have demonstrated high insecticidal efficacy, particularly against sap-feeding insect pests, [...] Read more.
Contact unmodified antisense DNA biotechnology (CUADb), developed in 2008, employs short antisense DNA oligonucleotides (oligos) as a novel approach to insect pest control. These oligonucleotide-based insecticides target pest mature rRNAs and/or pre-rRNAs and have demonstrated high insecticidal efficacy, particularly against sap-feeding insect pests, which are key vectors of plant DNA viruses and among the most economically damaging herbivorous insects. To further explore the potential of CUADb, this study evaluated the insecticidal efficacy of short 11-mer antisense DNA oligos against Coccus hesperidum, in comparison with long 56-mer single-stranded and double-stranded DNA sequences. The short oligos exhibited higher insecticidal activity. By day 9, the highest mortality rate (97.66 ± 4.04%) was recorded in the Coccus-11 group, while the most effective long sequence was the double-stranded DNA in the dsCoccus-56 group (77.09 ± 6.24%). This study also describes the architecture of the DNA containment (DNAc) mechanism, highlighting the intricate interactions between rRNAs and various types of DNA oligos. During DNAc, the Coccus-11 treatment induced enhanced ribosome biogenesis and ATP production through a metabolic shift from carbohydrates to lipid-based energy synthesis. However, this ultimately led to a ‘kinase disaster’ due to widespread kinase downregulation resulting from insufficient ATP levels. All DNA oligos with high or moderate complementarity to target rRNA initiated hypercompensation, but subsequent substantial rRNA degradation and insect mortality occurred only when the oligo sequence perfectly matched the rRNA. Both short and long oligonucleotide insecticide treatments led to a 3.75–4.25-fold decrease in rRNA levels following hypercompensation, which was likely mediated by a DNA-guided rRNase, such as RNase H1, while crucial enzymes of RNAi (DICER1, Argonaute 2, and DROSHA) were downregulated, indicating fundamental difference in molecular mechanisms of DNAc and RNAi. Consistently, significant upregulation of RNase H1 was detected in the Coccus-11 treatment group. In contrast, treatment with random DNA oligos resulted in only a 2–3-fold rRNA decrease, consistent with the normal rRNA half-life maintained by general ribonucleases. These findings reveal a fundamental new mechanism of rRNA regulation via complementary binding between exogenous unmodified antisense DNA and cellular rRNA. From a practical perspective, this minimalist approach, applying short antisense DNA dissolved in water, offers an effective, eco-friendly and innovative solution for managing sternorrhynchans and other insect pests. The results introduce a promising new concept in crop protection: DNA-programmable insect pest control. Full article
(This article belongs to the Special Issue New Insights into Plant and Insect Interactions (Second Edition))
Show Figures

Figure 1

12 pages, 757 KiB  
Brief Report
DNA-Programmable Oligonucleotide Insecticide Eriola-11 Targets Mitochondrial 16S rRNA and Exhibits Strong Insecticidal Activity Against Woolly Apple Aphid (Eriosoma lanigerum) Hausmann
by Vol Oberemok, Kate Laikova, Oksana Andreeva, Anastasia Dmitrienko, Tatiana Rybareva, Jamin Ali and Nikita Gal’chinsky
Int. J. Mol. Sci. 2025, 26(15), 7486; https://doi.org/10.3390/ijms26157486 - 2 Aug 2025
Viewed by 190
Abstract
The potent and selective ‘genetic zipper’ method for insect pest control consists of three essential components: an antisense DNA (the finder), its complementary mature rRNA or pre-rRNA of the pest (the target), and the host’s endogenous DNA-guided rRNase (the degrader). Although this approach [...] Read more.
The potent and selective ‘genetic zipper’ method for insect pest control consists of three essential components: an antisense DNA (the finder), its complementary mature rRNA or pre-rRNA of the pest (the target), and the host’s endogenous DNA-guided rRNase (the degrader). Although this approach has been validated, the spectrum of effective rRNA targets remains insufficiently explored. In this study, we report for the first time the insecticidal efficacy of a novel oligonucleotide insecticide, Eriola-11, which targets the mitochondrial 16S rRNA of the woolly apple aphid Eriosoma lanigerum Hausmann. We hypothesized that the antisense-mediated silencing of mitochondrial rRNA would impair aphid viability and lead to physiological disruptions associated with mitochondrial energy metabolism. Eriola-11 was applied either once or twice (with a 24 h interval) to aphid-infested plants, and aphid mortality was recorded over 14 days. Mitochondrial 16S rRNA expression levels were quantified using molecular assays, and the degradation kinetics of Eriola-11 were assessed in aphid tissue homogenates. Results showed significant insecticidal activity, with 67.55% mortality after a single treatment and 83.35% after two treatments. Treated aphids exhibited the loss of their characteristic white woolly wax covering, and mitochondrial 16S rRNA expression was reduced 0.66-fold relative to the control. Additionally, Eriola-11 was fully degraded by aphid DNases from tissue homogenates within 3 h, highlighting its rapid biodegradability. These findings establish mitochondrial 16S rRNA as a viable target for antisense insecticides and expand the catalogue of potential rRNA-based targets, offering a promising avenue for environmentally sustainable pest control strategies. Full article
(This article belongs to the Special Issue Antisense Oligonucleotides: Versatile Tools with Broad Applications)
Show Figures

Figure 1

15 pages, 1506 KiB  
Review
Dilated Cardiomyopathy and Sensorimotor Polyneuropathy Associated with a Homozygous ELAC2 Variant: A Case Report and Literature Review
by Francesco Ravera, Filippo Angelini, Pier Paolo Bocchino, Gianluca Marcelli, Giulia Gobello, Giuseppe Giannino, Guglielmo Merlino, Benedetta De Guidi, Andrea Destefanis, Giulia Margherita Brach Del Prever, Carla Giustetto, Guglielmo Gallone, Stefano Pidello, Antonella Barreca, Silvia Deaglio, Gaetano Maria De Ferrari, Claudia Raineri and Veronica Dusi
Cardiogenetics 2025, 15(3), 20; https://doi.org/10.3390/cardiogenetics15030020 - 31 Jul 2025
Viewed by 125
Abstract
Variants in ELAC2, a gene encoding the mitochondrial RNase Z enzyme essential for mitochondrial tRNA processing, have been associated with severe pediatric-onset mitochondrial dysfunction, primarily presenting with developmental delay, hypertrophic cardiomyopathy (HCM), and lactic-acidosis. We hereby report the case of a 25-year-old [...] Read more.
Variants in ELAC2, a gene encoding the mitochondrial RNase Z enzyme essential for mitochondrial tRNA processing, have been associated with severe pediatric-onset mitochondrial dysfunction, primarily presenting with developmental delay, hypertrophic cardiomyopathy (HCM), and lactic-acidosis. We hereby report the case of a 25-year-old young woman presenting with dilated cardiomyopathy (DCM) and peripheral sensorimotor polyneuropathy, harboring a homozygous variant in ELAC2. The same variant has been reported only once so far in a case of severe infantile-onset form of HCM and mitochondrial respiratory chain dysfunction, with in vitro data showing a moderate reduction in the RNase Z activity and supporting the current classification as C4 according to the American College of Medical Genetics (ACMG) criteria (PS3, PM2, PM3, PP4). Our extensive clinical, imaging, histological, and genetic investigations support a causal link between the identified variant and the patient’s phenotype, despite the fact that the latter might be considered atypical according to the current state of knowledge. A detailed review of the existing literature on ELAC2-related disease is also provided, highlighting the molecular mechanisms underlying tRNA maturation, mitochondrial dysfunction, and the variable phenotypic expression. Our case further expands the clinical spectrum of ELAC2-related cardiomyopathies to include a relatively late onset in young adulthood and underscores the importance of comprehensive genetic testing in unexplained cardiomyopathies with multisystem involvement. Full article
(This article belongs to the Section Rare Disease-Genetic Syndromes)
Show Figures

Figure 1

16 pages, 1177 KiB  
Article
Genetic Differentiation of Ornamental and Fruit-Bearing Prunus laurocerasus Revealed by SSR and S-Locus Markers
by Attila Hegedűs, Péter Honfi, Sezai Ercisli, Gulce Ilhan, Endre György Tóth and Júlia Halász
Horticulturae 2025, 11(7), 854; https://doi.org/10.3390/horticulturae11070854 - 19 Jul 2025
Viewed by 357
Abstract
Cherry laurel (Prunus laurocerasus) is an understudied, highly polyploid (22×) species that is widely used as an ornamental shrub and as a fruit-bearing plant in Türkiye. We analyzed 43 accessions—33 ornamental cultivars and 10 fruit-bearing selections—by examining size variations in 10 [...] Read more.
Cherry laurel (Prunus laurocerasus) is an understudied, highly polyploid (22×) species that is widely used as an ornamental shrub and as a fruit-bearing plant in Türkiye. We analyzed 43 accessions—33 ornamental cultivars and 10 fruit-bearing selections—by examining size variations in 10 simple sequence repeat (SSR) markers and the first intron region of the self-incompatibility ribonuclease (S-RNase) gene. A total of 498 alleles were detected across 11 loci, with the highest number of alleles observed at the S-locus. The SSR loci amplified between 4 (ASSR63) and 17 (BPPCT039) alleles per accession, with eight of the 11 primers generating more than 12 alleles per accession. Two markers, BPPCT040 and CPSCT021, uniquely distinguished all tested accessions. Of the alleles, only 178 (36%) were shared between the ornamental and fruit-bearing groups, reflecting significant genetic differentiation. A dendrogram and principal coordinate analysis revealed three distinct groups. Group 1 included most Hungarian and some European cultivars. Groups 2 (Western European cultivars) and 3 (Turkish selections) exhibited higher average allele numbers, suggesting greater genetic diversity in these groups. Our results indicate that cultivated cherry laurels originate from a broad genetic base and show clear genetic divergence between ornamental and fruit-bearing selections, likely due to differing long-term selection pressures. The observed genetic variability is consistent with the polyploid nature of the species and supports the presumed self-incompatible phenotype. This is the first study to report SSR fingerprints for ornamental cultivars and fruit-bearing selections, providing a potential tool for use in breeding programs. Full article
Show Figures

Graphical abstract

28 pages, 2988 KiB  
Review
Circular RNAs as Targets for Developing Anticancer Therapeutics
by Jaewhoon Jeoung, Wonho Kim, Hyein Jo and Dooil Jeoung
Cells 2025, 14(14), 1106; https://doi.org/10.3390/cells14141106 - 18 Jul 2025
Viewed by 539
Abstract
Circular RNA (CircRNA) is a single-stranded RNA arising from back splicing. CircRNAs interact with mRNA, miRNA, and proteins. These interactions regulate various life processes, including transcription, translation, cancer progression, anticancer drug resistance, and metabolism. Due to a lack of cap and poly(A) tails, [...] Read more.
Circular RNA (CircRNA) is a single-stranded RNA arising from back splicing. CircRNAs interact with mRNA, miRNA, and proteins. These interactions regulate various life processes, including transcription, translation, cancer progression, anticancer drug resistance, and metabolism. Due to a lack of cap and poly(A) tails, circRNAs show exceptional stability and resistance to RNase degradation. CircRNAs exhibit dysregulated expression patterns in various cancers and influence cancer progression. Stability and regulatory roles in cancer progression make circRNAs reliable biomarkers and targets for the development of anticancer therapeutics. The dysregulated expression of circRNAs is associated with resistance to anticancer drugs. Enhanced glycolysis by circRNAs leads to resistance to anticancer drugs. CircRNAs have been known to regulate the response to chemotherapy drugs and immune checkpoint inhibitors. Exogenous circRNAs can encode antigens that can induce both innate and adaptive immunity. CircRNA vaccines on lipid nanoparticles have been shown to enhance the sensitivity of cancer patients to immune checkpoint inhibitors. In this review, we summarize the roles and mechanisms of circRNAs in anticancer drug resistance and glycolysis. This review discusses clinical applications of circRNA vaccines to overcome anticancer drug resistance and enhance the efficacy of immune checkpoint inhibitors. The advantages and disadvantages of circRNA vaccines are also discussed. Overall, this review stresses the potential value of circRNAs as new therapeutic targets and diagnostic/prognostic biomarkers for cancer Full article
Show Figures

Figure 1

17 pages, 12102 KiB  
Article
Multiomics Integration of Parkinson’s Disease Datasets Reveals Unexpected Roles of IRE1 in Its Pathology
by Bianka Alexandra Pasat, Matthieu Moncan, Eleftherios Pilalis, Afshin Samali, Aristotelis Chatziioannou and Adrienne M. Gorman
Int. J. Mol. Sci. 2025, 26(14), 6711; https://doi.org/10.3390/ijms26146711 - 12 Jul 2025
Viewed by 343
Abstract
Parkinson’s disease (PD) is the second most common neurodegenerative disease. It primarily affects the motor system but is also associated with a range of cognitive impairments that can manifest early in disease progression, indicating its multifaceted nature. In this paper, we performed a [...] Read more.
Parkinson’s disease (PD) is the second most common neurodegenerative disease. It primarily affects the motor system but is also associated with a range of cognitive impairments that can manifest early in disease progression, indicating its multifaceted nature. In this paper, we performed a meta-analysis of transcriptomics and proteomics data using MultiOmicsIntegrator to gain insights into the post-transcriptional modifications and deregulated pathways associated with this disease. Our results reveal differential isoform usage between control and PD patient brain samples that result in enriched alternative splicing events, including an extended UTR length, domain loss, and the upregulation of non-coding isoforms. We found that Inositol-Requiring Enzyme 1 (IRE1) is active in PD samples and examined the role of its downstream signaling through X-box binding mRNA 1 (XBP1) and regulated IRE1-dependent decay (RIDD). We identified several RIDD candidates and showed that the enriched alternative splicing events observed are associated with RIDD. Moreover, in vitro mRNA cleavage assays demonstrated that OSBPL3, C16orf74, and SLC6A1 mRNAs are targets of IRE1 RNAse activity. Finally, a pathway enrichment analysis of both XBP1s and RIDD targets in the PD samples uncovered associations with processes such as immune response, oxidative stress, signal transduction, and cell–cell communication that have previously been linked to PD. These findings highlight a potential regulatory role of IRE in PD. Full article
Show Figures

Figure 1

39 pages, 10640 KiB  
Review
Endogenous Ribonucleases: Therapeutic Targeting of the Transcriptome Through Oligonucleotide-Triggered RNA Inactivation
by Daria A. Chiglintseva, Olga A. Patutina and Marina A. Zenkova
Biomolecules 2025, 15(7), 965; https://doi.org/10.3390/biom15070965 - 4 Jul 2025
Viewed by 430
Abstract
The selective regulation of gene expression at the RNA level represents a rapidly evolving field offering substantial clinical potential. This review examines the molecular mechanisms of intracellular enzymatic systems that utilize single-stranded nucleic acids to downregulate specific RNA targets. The analysis encompasses antisense [...] Read more.
The selective regulation of gene expression at the RNA level represents a rapidly evolving field offering substantial clinical potential. This review examines the molecular mechanisms of intracellular enzymatic systems that utilize single-stranded nucleic acids to downregulate specific RNA targets. The analysis encompasses antisense oligonucleotides and synthetic mimics of small interfering RNA (siRNA), microRNA (miRNA), transfer RNA-derived small RNA (tsRNA), and PIWI-interacting RNA (piRNA), elucidating their intricate interactions with crucial cellular machinery, specifically RNase H1, RNase P, AGO, and PIWI proteins, mediating their biological effects. The functional and structural characteristics of these endonucleases are examined in relation to their mechanisms of action and resultant therapeutic outcomes. This comprehensive analysis illuminates the interactions between single-stranded nucleic acids and their endonuclease partners, covering antisense inhibition pathways as well as RNA interference processes. This field of research has important implications for advancing targeted RNA modulation strategies across various disease contexts. Full article
(This article belongs to the Section Biomacromolecules: Proteins, Nucleic Acids and Carbohydrates)
Show Figures

Graphical abstract

12 pages, 4906 KiB  
Review
Therapeutic Approaches for C9ORF72-Related ALS: Current Strategies and Future Horizons
by Marco Cattaneo, Eleonora Giagnorio, Giuseppe Lauria and Stefania Marcuzzo
Int. J. Mol. Sci. 2025, 26(13), 6268; https://doi.org/10.3390/ijms26136268 - 28 Jun 2025
Viewed by 780
Abstract
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the loss of upper and lower motor neurons. One of its major genetic causes is C9ORF72, where mutations lead to hexanucleotide repeat expansions in the C9ORF72 gene. These expansions drive disease progression [...] Read more.
Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease characterized by the loss of upper and lower motor neurons. One of its major genetic causes is C9ORF72, where mutations lead to hexanucleotide repeat expansions in the C9ORF72 gene. These expansions drive disease progression through mechanisms, including the formation of toxic RNAs and the accumulation of damaged proteins such as dipeptide repeats (DPRs). This review highlights these pathogenic mechanisms, focusing on RNA foci formation and the accumulation of toxic DPRs, which contribute to neuronal damage. It also discusses promising targeted therapies, including small molecules and biological drugs, designed to counteract these specific molecular events. Small molecules such as G-quadruplex stabilizers, proteasome and autophagy modulators, and RNase-targeting chimeras show potential in reducing RNA foci and DPR accumulation. Furthermore, targeting enzymes involved in repeat-associated non-AUG (RAN) translation and nucleocytoplasmic transport, which are crucial for disease pathogenesis, opens new therapeutic avenues. Even some anti-viral drugs show encouraging results in preclinical studies. Biological drugs, such as antisense oligonucleotides and gene-editing technologies like CRISPR-Cas, were explored for their potential to specifically target C9ORF72 mutations and modify the disease’s molecular foundations. While preclinical and early clinical data show promise, challenges remain in optimizing delivery methods, ensuring long-term safety, and improving efficacy. This review concludes by emphasizing the importance of continued research and the potential for these therapies to alter the disease trajectory and improve patient outcomes. Full article
(This article belongs to the Section Molecular Neurobiology)
Show Figures

Figure 1

17 pages, 962 KiB  
Article
Specific Heat-Killed Lactic Acid Bacteria Enhance Mucosal Aminopeptidase N Activity in the Small Intestine of Aged Mice
by Takeshi Tsuruta, Mami Wakisaka, Takumi Watanabe, Aoi Nishijima, Akihito Ikeda, Mao Teraoka, Tianyang Wang, Kuiyi Chen and Naoki Nishino
Int. J. Mol. Sci. 2025, 26(12), 5742; https://doi.org/10.3390/ijms26125742 - 15 Jun 2025
Viewed by 575
Abstract
Aminopeptidase N (APN), an enzyme expressed in the small intestinal mucosa, is involved in dietary protein digestion. Previous studies have shown that oral administration of fermented milk containing lactic acid bacteria (LAB) enhances mucosal APN activity in young mice. This study aimed to [...] Read more.
Aminopeptidase N (APN), an enzyme expressed in the small intestinal mucosa, is involved in dietary protein digestion. Previous studies have shown that oral administration of fermented milk containing lactic acid bacteria (LAB) enhances mucosal APN activity in young mice. This study aimed to investigate whether LAB strains stimulate mucosal APN activity in aged mice and to evaluate its relevance to age-related changes in body composition. The underlying molecular mechanisms were also explored in vitro. Experiment 1: Aged C57BL/6J mice were fed diets supplemented with heat-killed LAB strains—Enterococcus faecalis OU-23 (EF), Leuconostoc mesenteroides OU-03 (LM), or Lactiplantibacillus plantarum SNK12 (LP). Compared to the aged Control group, the ileal APN activity was significantly higher in the LP group. LP administration also elevated serum Gla-osteocalcin levels and decreased serum CTX-1 levels. Experiment 2: IEC-6 cells were co-cultured with LP that had been treated with RNase, DNase, or lysozyme. APN activity was significantly lower in cells co-cultured with DNase- or lysozyme-treated LP compared to those co-cultured with untreated LP. A specific LAB strain may enhance mucosal APN activity in the aged intestine, potentially contributing to improved bone metabolism. This effect may be mediated by bacterial DNA and peptidoglycan. Full article
Show Figures

Graphical abstract

12 pages, 2769 KiB  
Article
5′ DREDGE: Direct Repeat-Enabled Downregulation of Gene Expression via the 5′ UTR of Target Genes
by Sagar J. Parikh, Heather M. Terron, Luke A. Burgard, Dylan D. Butler, Frank M. LaFerla, Shelley Lane and Malcolm A. Leissring
Cells 2025, 14(12), 866; https://doi.org/10.3390/cells14120866 - 8 Jun 2025
Viewed by 680
Abstract
Despite the availability of numerous methods for controlling gene expression, there remains a strong need for technologies that maximize two key properties: selectivity and reversibility. To this end, we developed a novel approach that exploits the highly sequence-specific nature of CRISPR-associated endoribonucleases (Cas [...] Read more.
Despite the availability of numerous methods for controlling gene expression, there remains a strong need for technologies that maximize two key properties: selectivity and reversibility. To this end, we developed a novel approach that exploits the highly sequence-specific nature of CRISPR-associated endoribonucleases (Cas RNases), which recognize and cleave short RNA sequences known as direct repeats (DRs). In this approach, referred to as DREDGE (direct repeat-enabled downregulation of gene expression), selective control of gene expression is enabled by introducing one or more DRs into the untranslated regions (UTRs) of target mRNAs, which can then be cleaved upon expression of the cognate Cas RNase. We previously demonstrated that the expression of target genes with DRs in their 3′ UTRs are efficiently controlled by the DNase-dead version of Cas12a (dCas12a) with a high degree of selectivity and complete reversibility. Here, we assess the feasibility of using DREDGE to regulate the expression of genes with DRs inserted in their 5′ UTRs. Among the five different Cas RNases tested, Csy4 was found to be the most efficient in this context, yielding robust downregulation with rapid onset in doxycycline-regulatable systems targeting either a stably expressed fluorescent protein or an endogenous gene, both in a fully reversible manner. Unexpectedly, dCas12a was also found to be modestly effective despite binding essentially irreversibly to the cut mRNA on its 5′ end and thereby boosting mRNA levels. Our results expand the utility of DREDGE as an attractive method for regulating gene expression in a targeted, highly selective, and fully reversible manner. Full article
(This article belongs to the Section Cell Methods)
Show Figures

Figure 1

19 pages, 7764 KiB  
Article
Binding Specificity and Oligomerization of TSWV N Protein in the Western Flower Thrips, Frankliniella occidentalis
by Falguni Khan, Eticha Abdisa, Niayesh Shahmohammadi and Yonggyun Kim
Viruses 2025, 17(6), 826; https://doi.org/10.3390/v17060826 - 7 Jun 2025
Viewed by 538
Abstract
Tomato spotted wilt virus (TSWV) is a highly destructive plant pathogen and transmitted by several thrips including the western flower thrips, Frankliniella occidentalis. A structural N protein encoded in the viral genome represents the nucleocapsid protein by binding to the viral RNA [...] Read more.
Tomato spotted wilt virus (TSWV) is a highly destructive plant pathogen and transmitted by several thrips including the western flower thrips, Frankliniella occidentalis. A structural N protein encoded in the viral genome represents the nucleocapsid protein by binding to the viral RNA genome. However, it remains unknown how the RNA-binding protein specifically interacts with the viral RNA from host RNAs in the target cells. To study the molecular basis of N function, we produced the protein in Escherichia coli and the resulting purified recombinant protein was used to investigate the protein–RNA interactions. The recombinant N protein migrated on agarose gel to the anode in the electric field due to its high basic isoelectric point. This electrostatic property led N protein to bind to DNA as well as RNA. It also bound to both single-stranded (ssRNA) and double-stranded RNA (dsRNA). However, when the total RNA was extracted from plant tissues collected from TSWV-infected host, the RNA extract using the recombinant N protein was much richer in the TSWV genome compared to that without the protein. To investigate the specificity of N protein to ssRNA, the three-dimensional structure was predicted using the AlphaFold program and showed its trimeric oligomerization with the binding pocket for ssRNA. This was supported by the differential susceptibility of N protein with ssRNA and dsRNA against RNase attack. Furthermore, a thermal shift assay to analyze the RNA and protein interaction showed that ssRNA strongly interacted with N protein compared to dsRNA. In addition, the N gene was expressed along with the multiplication of the viral RNA genome segments from the segment-specific fluorescence in situ hybridization analysis in different tissues during different developmental stages of the virus-infected F. occidentalis. These results suggest that the functional trimeric N proteins bind to the viral RNA to form a basic nucleocapsid structure at a specific virus-replicating compartment within the host cells. Full article
(This article belongs to the Special Issue Molecular Virus–Insect Interactions, 2nd Edition)
Show Figures

Figure 1

13 pages, 1519 KiB  
Article
Multiplexed CRISPR Assay for Amplification-Free Detection of miRNAs
by P. I. Thilini De Silva, Keshani Hiniduma, Rachelle Canete, Ketki S. Bhalerao, Sherif M. Shawky, Hansana Gunathilaka, Jessica L. Rouge, Islam M. Mosa, David C. Steffens, Kevin Manning, Breno S. Diniz and James F. Rusling
Biosensors 2025, 15(6), 346; https://doi.org/10.3390/bios15060346 - 29 May 2025
Viewed by 852
Abstract
CRISPR-Cas proteins from bacteria are powerful tools for gene editing and molecular diagnostics. Expanding capacity of CRISPR to low cost, multiplexed assays of biomarkers is a key to future disease diagnostics, since multiple biomarker detection is essential for reliable diagnostics. Herein we describe [...] Read more.
CRISPR-Cas proteins from bacteria are powerful tools for gene editing and molecular diagnostics. Expanding capacity of CRISPR to low cost, multiplexed assays of biomarkers is a key to future disease diagnostics, since multiple biomarker detection is essential for reliable diagnostics. Herein we describe a multiplexed assay in a 3D-printed 96-well plate with CRISPR-Cas13a immobilized in each well to target three circulating blood biomarker microRNAs (miRNAs 34c-5p, 200c-3p, and 30e-5p) for Alzheimer’s disease (ALZ). Immobilized Cas13a is equipped with different crRNAs complementary to each miRNA target. MiRNA binding to crRNA complements activates the collateral RNase activity of Cas13a, cleaving a quenched fluorescent reporter (RNaseAlert) with fluorophore and quencher connected by an RNA oligonucleotide to enable fluorescence measurements. We achieved ultralow limits of detection (LOD) of 0.74 fg/mL for miRNA 34c-5p, 0.70 fg/mL for miRNA 30e-5p, and 7.4 fg/mL for miRNA 200c-3p, with dynamic ranges from LODs up to about 1800 pg/mL. The accuracy of the assay was validated by spike-recovery studies and good correlation of levels of patient plasma samples vs. a referee method. This new approach provides selective, sensitive multiplex miRNA biosensing, and simultaneously accommodates analysis of standards and controls. Full article
(This article belongs to the Special Issue Biosensors for Monitoring and Diagnostics)
Show Figures

Figure 1

15 pages, 2941 KiB  
Article
Mechanism of circRNA_4083 Circularization and Its Role in Regulating Cell Viability
by Wenhao Li, Ting Yang, Haojie Wang, Hao Bai, Guobin Chang and Lingling Qiu
Animals 2025, 15(11), 1527; https://doi.org/10.3390/ani15111527 - 23 May 2025
Viewed by 516
Abstract
Circular RNAs (circRNAs), a class of covalently closed non-coding RNAs, are pivotal regulators of gene expression and contributors to disease pathogenesis. This study elucidated the biogenesis, functional significance, and regulatory network of circRNA_4083, a novel exon-derived circRNA originating from exons 22 and 23 [...] Read more.
Circular RNAs (circRNAs), a class of covalently closed non-coding RNAs, are pivotal regulators of gene expression and contributors to disease pathogenesis. This study elucidated the biogenesis, functional significance, and regulatory network of circRNA_4083, a novel exon-derived circRNA originating from exons 22 and 23 of the MSH3 gene in chicken. Through comprehensive molecular characterization—including Sanger sequencing, RNase R digestion assays, and subcellular localization—we confirmed the robust stability and predominant cytoplasmic localization of circRNA_4083 across diverse chicken tissues. Mechanistic investigations revealed that reverse complementary sequences within flanking intronic regions are indispensable for its circularization, as demonstrated by overexpression plasmids (#1–#4) in DF-1 cells. Functional analyses demonstrated that circRNA_4083 significantly inhibited cell apoptosis and increased cellular viability. Integrative bioinformatics approaches predicted a competing endogenous RNA (ceRNA) network comprising 12 miRNAs and 2132 target genes (FDR < 0.05), with significant enrichment in pathways critical to genomic stability, including non-homologous end joining (NHEJ) and ubiquitin-mediated proteolysis. These findings position circRNA_4083 as a key modulator of cellular viability and genomic integrity, with potential implications for avian leukosis virus-J (ALV-J) pathogenesis and resistance breeding strategies. This work advances our understanding of circRNA-driven regulatory mechanisms in avian species and underscores their relevance in poultry health. Full article
(This article belongs to the Special Issue Livestock and Poultry Genetics and Breeding Management)
Show Figures

Figure 1

15 pages, 657 KiB  
Article
A Comparison of Diagnostic Methods for Feline Leukemia Virus and Feline Immunodeficiency Virus: Immunochromatographic Assay and RNases Hybridization-Assisted Amplification Test Kit Compared to Reverse Transcription Quantitative Polymerase Chain Reaction
by Thanikran Suwannachote, Wisut Prasitsuwan, Thirawat Sumalai and Sakchai Ruenphet
Animals 2025, 15(10), 1484; https://doi.org/10.3390/ani15101484 - 20 May 2025
Viewed by 629
Abstract
Feline leukemia virus (FeLV) and Feline immunodeficiency virus (FIV) are globally prevalent retroviral pathogens that pose significant health risks to domestic cats. This study aimed to compare the diagnostic performance of two point-of-care—the immunochromatographic assay (ICA) and the RNase hybridization-assisted amplification (RHAM) test [...] Read more.
Feline leukemia virus (FeLV) and Feline immunodeficiency virus (FIV) are globally prevalent retroviral pathogens that pose significant health risks to domestic cats. This study aimed to compare the diagnostic performance of two point-of-care—the immunochromatographic assay (ICA) and the RNase hybridization-assisted amplification (RHAM) test kit—against reverse transcription quantitative polymerase chain reaction (RT-qPCR), the current gold standard for FeLV and FIV detection. For FeLV detection, ICA demonstrated a sensitivity of 86.89%, specificity of 96.55%, accuracy of 90.00%, and precision of 98.15%, while for FIV detection, the assay showed a sensitivity of 75.86%, specificity of 88.52%, accuracy of 84.44%, and precision of 75.86%. In contrast, the RHAM test exhibited superior performance, with FeLV detection sensitivity of 93.44%, specificity of 98.28%, accuracy of 94.44%, and precision of 98.28%. For FIV detection, RHAM demonstrated a sensitivity of 75.86%, specificity of 100%, accuracy of 92.22%, and precision of 100%. Additionally, the RHAM assay significantly reduced detection time compared to RT-qPCR, enabling expedited clinical decision-making, alleviating laboratory workload, and lowering diagnostic costs. These benefits are particularly relevant in veterinary settings with limited access to PCR-based diagnostics, where the RHAM assay represents a rapid, reliable, and resource-efficient alternative for FeLV and FIV detection. Full article
(This article belongs to the Special Issue General Epidemiology of Animal Viruses (Second Edition))
Show Figures

Figure 1

15 pages, 2367 KiB  
Article
An Engineered RNase P Ribozyme Effectively Reduces Human Coronavirus 229E Gene Expression and Growth in Human Cells
by Yujun Liu, Bin Yan, Hao Gong and Fenyong Liu
Zoonotic Dis. 2025, 5(2), 12; https://doi.org/10.3390/zoonoticdis5020012 - 12 May 2025
Viewed by 540
Abstract
The human coronavirus 229E (HCoV-229E) is a member of the human coronavirus family that includes SARS-CoV-2, the causative agent of COVID-19. Developing antiviral strategies and compounds is crucial to treat and prevent HCoV-229E infections and the associated diseases. Ribozymes derived from ribonuclease P [...] Read more.
The human coronavirus 229E (HCoV-229E) is a member of the human coronavirus family that includes SARS-CoV-2, the causative agent of COVID-19. Developing antiviral strategies and compounds is crucial to treat and prevent HCoV-229E infections and the associated diseases. Ribozymes derived from ribonuclease P (RNase P) catalytic RNA represent a novel class of promising gene-targeting agents by cleaving their target mRNA and knocking down the expression of the target mRNA. However, it has not been reported whether RNase P ribozymes block the infection and replication of HCoV-229E. We report here the engineering of an anti-HCoV-229E RNase P ribozyme to target an overlapping region of viral genomic RNA and the mRNA encoding the nucleocapsid (N) protein, which is vital for viral replication and growth. The engineered ribozyme actively hydrolyzed the viral RNA target in vitro. HCoV-229E-infected cells expressing the engineered, catalytically active ribozyme exhibited a reduction of about 85% in viral RNA levels and N protein expression, and a reduction of about 750-fold in infectious particle production, compared to cells expressing no ribozymes or a control, catalytically inactive ribozyme. Our study provides the first direct evidence of the therapeutic potential of RNase P ribozymes against human coronaviruses such as HCoV-229E. Full article
Show Figures

Figure 1

Back to TopTop