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Genes May 2024PIWI-interacting RNAs (piRNAs), a class of small non-coding RNAs (sncRNAs) with 24-32 nucleotides (nt), were initially identified in the reproductive system. Unlike... (Review)
Review
PIWI-interacting RNAs (piRNAs), a class of small non-coding RNAs (sncRNAs) with 24-32 nucleotides (nt), were initially identified in the reproductive system. Unlike microRNAs (miRNAs) or small interfering RNAs (siRNAs), piRNAs normally guide P-element-induced wimpy testis protein (PIWI) families to slice extensively complementary transposon transcripts without the seed pairing. Numerous studies have shown that piRNAs are abundantly expressed in the brain, and many of them are aberrantly regulated in central neural system (CNS) disorders. However, the role of piRNAs in the related developmental and pathological processes is unclear. The elucidation of piRNAs/PIWI would greatly improve the understanding of CNS development and ultimately lead to novel strategies to treat neural diseases. In this review, we summarized the relevant structure, properties, and databases of piRNAs and their functional roles in neural development and degenerative disorders. We hope that future studies of these piRNAs will facilitate the development of RNA-based therapeutics for CNS disorders.
Topics: Humans; RNA, Small Interfering; Animals; Argonaute Proteins; Nervous System Diseases; Neurogenesis
PubMed: 38927589
DOI: 10.3390/genes15060653 -
Antibiotics (Basel, Switzerland) Jun 2024Leptospirosis is a major zoonotic disease caused by pathogenic spirochetes in the genus Leptospira, affecting over a million people annually and causing approximately...
Leptospirosis is a major zoonotic disease caused by pathogenic spirochetes in the genus Leptospira, affecting over a million people annually and causing approximately 60,000 deaths. , a key causative agent, likely possesses defense systems against bacteriophages (leptophages), yet these systems are not well understood. We analyzed 402 genomes of using the DefenseFinder tool to identify and characterize the antiphage defense systems. We detected 24 unique systems, with CRISPR-Cas (Clustered Regularly Interspaced Short Palindromic Repeats and CRISPR-associated proteins), PrrC, Borvo, and Restriction-Modification (R-M) being the most prevalent. Notably, Cas were identified in all strains, indicating their central role in phage defense. Furthermore, there were variations in the antiphage system distribution across different serovars, suggesting unique evolutionary adaptations. For instance, Retron was found exclusively in the Canicola serovar, while prokaryotic Argonaute proteins (pAgo) were only detected in the Grippotyphosa serovar. These findings significantly enhance our understanding of 's antiphage defense mechanisms. They reveal the potential for the development of serovar-specific phage-based therapies and underscore the importance of further exploring these defense systems.
PubMed: 38927188
DOI: 10.3390/antibiotics13060522 -
Cell Reports Jun 2024Inhibition of nucleic acid targets is mediated by Argonaute (Ago) proteins guided by RNA or DNA. Although the mechanisms underpinning the functions of eukaryotic and...
Inhibition of nucleic acid targets is mediated by Argonaute (Ago) proteins guided by RNA or DNA. Although the mechanisms underpinning the functions of eukaryotic and "long" prokaryotic Ago proteins (pAgos) are well understood, those for short pAgos remain enigmatic. Here, we determine two cryoelectron microscopy structures of short pAgos in association with the NADase-domain-containing protein Sir2-APAZ from Geobacter sulfurreducens (GsSir2/Ago): the guide RNA-target DNA-loaded GsSir2/Ago quaternary complex (2.58 Å) and the dimer of the quaternary complex (2.93Å). These structures show that the nucleic acid binding causes profound conformational changes that result in disorder or partial dissociation of the Sir2 domain, suggesting that it adopts a NADase-active conformation. Subsequently, two RNA-/DNA-loaded GsSir2/Ago complexes form a dimer through their MID domains, further enhancing NADase activity through synergistic effects. The findings provide a structural basis for short-pAgo-mediated defense against invading nucleic acids.
PubMed: 38923459
DOI: 10.1016/j.celrep.2024.114391 -
Asian Pacific Journal of Cancer... Jun 2024The aim of this study was to evaluate the expression profiles of PIWI-like protein- 2 (PIWIL2), and HepPar1 and their immunohistochemical (IHC) characteristics in...
OBJECTIVE
The aim of this study was to evaluate the expression profiles of PIWI-like protein- 2 (PIWIL2), and HepPar1 and their immunohistochemical (IHC) characteristics in Hepatocellular Carcinoma (HCC), and determine their correlation with clinicopathological parameters of this type of cancer to determine their diagnostic value in combination.
METHODS
Seventy-five patients with HCC were assessed for the expression of PIWIL2 in serum and tissue using real-time polymerase chain reaction (RT-PCR) and IHC was performed for PIWIL2 and HepPar1 was performed on all patients.
RESULTS
A statistically significantly higher level of PIWIL2 was found in HCC compared to controls (p≤0.001). Both HepPar1 and PIWIL2 were detected in 84% of HCC cases, the diagnostic and prognostic factors for PIWIL2 were found to be significant in liver tumour tissue samples and non-tumorous sections p<0.001, and the same was observed for serum samples and results of healthy serum controls (p<0.001) when compared to AFP.
CONCLUSION
Our results affirm the hypothesis that reactivation of PIWI expression in various caner types is crucial for cancer development, and that a possible panel maybe used for these markers HCC diagnosis.
Topics: Humans; Carcinoma, Hepatocellular; Liver Neoplasms; Argonaute Proteins; Biomarkers, Tumor; Male; Female; Middle Aged; Prognosis; Case-Control Studies; Follow-Up Studies; Adult; alpha-Fetoproteins; Aged
PubMed: 38918675
DOI: 10.31557/APJCP.2024.25.6.2123 -
Frontiers in Bioengineering and... 2024Boolean gates, the fundamental components of digital circuits, have been widely investigated in synthetic biology because they permit the fabrication of biosensors and...
Boolean gates, the fundamental components of digital circuits, have been widely investigated in synthetic biology because they permit the fabrication of biosensors and facilitate biocomputing. This study was conducted to design and construct Boolean gates in the yeast , the main component of which was the RNA interference pathway (RNAi) that is naturally absent from the budding yeast cells. We tested different expression cassettes for the siRNA precursor (a giant hairpin sequence, a DNA fragment-flanked by one or two introns-between convergent promoters or transcribed separately in the sense and antisense directions) and placed different components under the control of the circuit inputs (i.e., the siRNA precursor or proteins such as the Dicer and the Argonaute). We found that RNAi-based logic gates are highly sensitive to promoter leakage and, for this reason, challenging to implement . Convergent-promoter architecture turned out to be the most reliable solution, even though the overall best performance was achieved with the most difficult design based on the siRNA precursor as a giant hairpin.
PubMed: 38895554
DOI: 10.3389/fbioe.2024.1392967 -
International Journal of Molecular... May 2024DNA methylation is an important way to regulate gene expression in eukaryotes. In order to reveal the role of DNA methylation in the regulation of germ cell-specific...
DNA methylation is an important way to regulate gene expression in eukaryotes. In order to reveal the role of DNA methylation in the regulation of germ cell-specific gene expression during spermatogenesis of Japanese flounder (), the expression profiles of () and () genes in the gonads of female, male, and sex-reversed pseudo-male were analyzed, and the dynamic of DNA methylation was investigated. As a result, and genes were highly expressed in the testis of both male and pseudo-male , with significant variation among male individuals. The DNA methylation levels in the promoter regions of both and were negatively correlated with their expression levels, which may contribute to the transcriptional regulation of genes during spermatogenesis. There was also sperm quality variation among male , and the sperm curvilinear velocity was positively correlated with the expression of both and genes. These results indicated that the DNA methylation in and promoter regions may affect the initiation of gene transcription, thereby regulating gene expression and further affecting the spermatogenesis process and gamete quality in .
Topics: Animals; Male; DNA Methylation; Argonaute Proteins; Flounder; Spermatozoa; Spermatogenesis; Female; Promoter Regions, Genetic; Testis; Gene Expression Regulation; Fish Proteins
PubMed: 38892123
DOI: 10.3390/ijms25115935 -
Scientific Data Jun 2024In this study we examine the impact of cell confluency on gene expression. We focused on Argonaute (AGO) protein dynamics and associated gene and protein expression in...
In this study we examine the impact of cell confluency on gene expression. We focused on Argonaute (AGO) protein dynamics and associated gene and protein expression in HEK293, A375, and SHSY5Y cell lines. As a consequence of cell confluency, AGO2 protein translocates into the nucleus. Therefore, we generated transcriptomic data using RNA sequencing to compare gene expression in subconfluent versus confluent cells, which highlighted significant alterations in gene regulation patterns directly corresponding to changes in cell density. Our study also encompasses miRNA profiling data obtained through small RNA sequencing, revealing miRNA expressional changes dependent on cellular confluency, as well as cellular localization. Finally, we derived proteomic data from mass spectrometry analyses following AGO1-4 immunoprecipitation, providing a comprehensive view of AGO interactome in both nuclear and cytoplasmic compartments under varying confluency. These datasets offer a detailed exploration of the cellular and molecular dynamics, influenced by cell confluency, presenting a valuable resource for further research in cellular biology, particularly in understanding the basic mechanisms of cell density in cancer cells.
Topics: Humans; Argonaute Proteins; Transcriptome; Proteomics; MicroRNAs; HEK293 Cells; Cell Line, Tumor; Gene Expression Profiling
PubMed: 38866801
DOI: 10.1038/s41597-024-03465-z -
Nature Communications Jun 2024A short prokaryotic Argonaute (pAgo) TIR-APAZ (SPARTA) defense system, activated by invading DNA to unleash its TIR domain for NAD(P) hydrolysis, was recently identified...
A short prokaryotic Argonaute (pAgo) TIR-APAZ (SPARTA) defense system, activated by invading DNA to unleash its TIR domain for NAD(P) hydrolysis, was recently identified in bacteria. We report the crystal structure of SPARTA heterodimer in the absence of guide-RNA/target-ssDNA (2.66 Å) and a cryo-EM structure of the SPARTA oligomer (tetramer of heterodimers) bound to guide-RNA/target-ssDNA at nominal 3.15-3.35 Å resolution. The crystal structure provides a high-resolution view of SPARTA, revealing the APAZ domain as equivalent to the N, L1, and L2 regions of long pAgos and the MID domain containing a unique insertion (insert57). Cryo-EM structure reveals regions of the PIWI (loop10-9) and APAZ (helix αN) domains that reconfigure for nucleic-acid binding and decrypts regions/residues that reorganize to expose a positively charged pocket for higher-order assembly. The TIR domains amass in a parallel-strands arrangement for catalysis. We visualize SPARTA before and after RNA/ssDNA binding and uncover the basis of its active assembly leading to abortive infection.
Topics: Argonaute Proteins; Cryoelectron Microscopy; Crystallography, X-Ray; Bacterial Proteins; Protein Domains; DNA, Single-Stranded; RNA, Guide, CRISPR-Cas Systems; Models, Molecular; Nucleic Acids; Protein Binding
PubMed: 38844755
DOI: 10.1038/s41467-024-49271-4 -
Viruses Apr 2024The cucumber mosaic virus (CMV) 2b protein is a suppressor of plant defenses and a pathogenicity determinant. Amongst the 2b protein's host targets is the RNA silencing...
The cucumber mosaic virus (CMV) 2b protein is a suppressor of plant defenses and a pathogenicity determinant. Amongst the 2b protein's host targets is the RNA silencing factor Argonaute 1 (AGO1), which it binds to and inhibits. In , if 2b-induced inhibition of AGO1 is too efficient, it induces reinforcement of antiviral silencing by AGO2 and triggers increased resistance against aphids, CMV's insect vectors. These effects would be deleterious to CMV replication and transmission, respectively, but are moderated by the CMV 1a protein, which sequesters sufficient 2b protein molecules into P-bodies to prevent excessive inhibition of AGO1. Mutant 2b protein variants were generated, and red and green fluorescent protein fusions were used to investigate subcellular colocalization with AGO1 and the 1a protein. The effects of mutations on complex formation with the 1a protein and AGO1 were investigated using bimolecular fluorescence complementation and co-immunoprecipitation assays. Although we found that residues 56-60 influenced the 2b protein's interactions with the 1a protein and AGO1, it appears unlikely that any single residue or sequence domain is solely responsible. In silico predictions of intrinsic disorder within the 2b protein secondary structure were supported by circular dichroism (CD) but not by nuclear magnetic resonance (NMR) spectroscopy. Intrinsic disorder provides a plausible model to explain the 2b protein's ability to interact with AGO1, the 1a protein, and other factors. However, the reasons for the conflicting conclusions provided by CD and NMR must first be resolved.
Topics: Argonaute Proteins; Cucumovirus; Arabidopsis; Arabidopsis Proteins; Protein Binding; Viral Proteins; Host-Pathogen Interactions; Viral Replicase Complex Proteins; Plant Diseases; RNA-Dependent RNA Polymerase; Methyltransferases
PubMed: 38793558
DOI: 10.3390/v16050676 -
Biosensors May 2024Since SARS-CoV-2 is a highly transmissible virus, alternative reliable, fast, and cost-effective methods are still needed to prevent virus spread that can be applied in...
Since SARS-CoV-2 is a highly transmissible virus, alternative reliable, fast, and cost-effective methods are still needed to prevent virus spread that can be applied in the laboratory and for point-of-care testing. Reverse transcription real-time fluorescence quantitative PCR (RT-qPCR) is currently the gold criteria for detecting RNA viruses, which requires reverse transcriptase to reverse transcribe viral RNA into cDNA, and fluorescence quantitative PCR detection was subsequently performed. The frequently used reverse transcriptase is thermolabile; the detection process is composed of two steps: the reverse transcription reaction at a relatively low temperature, and the qPCR performed at a relatively high temperature, moreover, the RNA to be detected needs to pretreated if they had advanced structure. Here, we develop a fast and sensitive one-tube SARS-CoV-2 detection platform based on Ultra-fast RTX-PCR and Argonaute-mediated Nucleic acid Detection (PAND) technology (URPAND). URPAND was achieved ultra-fast RTX-PCR process based on a thermostable RTX (exo-) with both reverse transcriptase and DNA polymerase activity. The URPAND can be completed RT-PCR and PAND to detect nucleic acid in one tube within 30 min. This method can specifically detect SARS-CoV-2 with a low detection limit of 100 copies/mL. The diagnostic results of clinical samples with one-tube URPAND displayed 100% consistence with RT-qPCR test. Moreover, URPAND was also applied to identify SARS-CoV-2 D614G mutant due to its single-nucleotide specificity. The URPAND platform is rapid, accurate, tube closed, one-tube, easy-to-operate and free of large instruments, which provides a new strategy to the detection of SARS-CoV-2 and other RNA viruses.
Topics: SARS-CoV-2; Pyrococcus furiosus; RNA, Viral; COVID-19; Humans; Argonaute Proteins; Real-Time Polymerase Chain Reaction; Biosensing Techniques; COVID-19 Nucleic Acid Testing
PubMed: 38785719
DOI: 10.3390/bios14050245