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Nature Communications Dec 2023Despite the importance of Nitric Oxide (NO) as signaling molecule in both plant and animal development, the regulatory mechanisms downstream of NO remain largely...
Despite the importance of Nitric Oxide (NO) as signaling molecule in both plant and animal development, the regulatory mechanisms downstream of NO remain largely unclear. Here, we show that NO is involved in Arabidopsis shoot stem cell control via modifying expression and activity of ARGONAUTE 4 (AGO4), a core component of the RNA-directed DNA Methylation (RdDM) pathway. Mutations in components of the RdDM pathway cause meristematic defects, and reduce responses of the stem cell system to NO signaling. Importantly, we find that the stem cell inducing WUSCHEL transcription factor directly interacts with AGO4 in a NO dependent manner, explaining how these two signaling systems may converge to modify DNA methylation patterns. Taken together, our results reveal that NO signaling plays an important role in controlling plant stem cell homeostasis via the regulation of de novo DNA methylation.
Topics: DNA Methylation; Arabidopsis Proteins; Nitric Oxide; Meristem; Arabidopsis; RNA; Gene Expression Regulation, Plant
PubMed: 38049411
DOI: 10.1038/s41467-023-43705-1 -
Nucleic Acids Research Jan 2024Prokaryotic Argonautes (pAgos) play a vital role in host defense by utilizing short nucleic acid guides to recognize and target complementary nucleic acids. Despite...
Prokaryotic Argonautes (pAgos) play a vital role in host defense by utilizing short nucleic acid guides to recognize and target complementary nucleic acids. Despite being the majority of pAgos, short pAgos have only recently received attention. Short pAgos are often associated with proteins containing an APAZ domain and a nuclease domain including DUF4365, SMEK, or HNH domain. In contrast to long pAgos that specifically cleave the target DNA, our study demonstrates that the short pAgo from Thermocrispum municipal, along with its associated DUF4365-APAZ protein, forms a heterodimeric complex. Upon RNA-guided target DNA recognition, this complex is activated to nonspecifically cleave DNA. Additionally, we found that the TmuRE-Ago complex shows a preference for 5'-OH guide RNA, specifically requires a uridine nucleotide at the 5' end of the guide RNA, and is sensitive to single-nucleotide mismatches between the guide RNA and target DNA. Based on its catalytic properties, our study has established a novel nucleic acid detection method and demonstrated its feasibility. This study not only expands our understanding of the defense mechanism employed by short pAgo systems but also suggests their potential applications in nucleic acid detection.
Topics: Argonaute Proteins; DNA; Endonucleases; Nucleic Acids; Prokaryotic Cells; Actinobacteria; RNA, Bacterial
PubMed: 38048327
DOI: 10.1093/nar/gkad1145 -
BioMed Research International 2023In different regions of the world, cowpea (Vigna unguiculata (L.) Walp.) is an important vegetable and an excellent source of protein. It lessens the malnutrition of the...
In different regions of the world, cowpea (Vigna unguiculata (L.) Walp.) is an important vegetable and an excellent source of protein. It lessens the malnutrition of the underprivileged in developing nations and has some positive effects on health, such as a reduction in the prevalence of cancer and cardiovascular disease. However, occasionally, certain biotic and abiotic stresses caused a sharp fall in cowpea yield. Major RNA interference (RNAi) genes like Dicer-like (DCL), Argonaute (AGO), and RNA-dependent RNA polymerase (RDR) are essential for the synthesis of their associated factors like domain, small RNAs (sRNAs), transcription factors, micro-RNAs, and acting factors that shield plants from biotic and abiotic stresses. In this study, applying BLASTP search and phylogenetic tree analysis with reference to the RNAi (AtRNAi) genes, we discovered 28 VuRNAi genes, including 7 , 14 , and 7 genes in cowpea. We looked at the domains, motifs, gene structures, chromosomal locations, subcellular locations, gene ontology (GO) terms, and regulatory factors (transcription factors, micro-RNAs, and -acting elements (CAEs)) to characterize the VuRNAi genes and proteins in cowpea in response to stresses. Predicted , (a, b), , , and genes might have an impact on cowpea growth, development of the vegetative and flowering stages, and antiviral defense. The VuRNAi gene regulatory features miR395 and miR396 might contribute to grain quality improvement, immunity boosting, and pathogen infection resistance under salinity and drought conditions. Predicted CAEs from the VuRNAi genes might play a role in plant growth and development, improving grain quality and production and protecting plants from biotic and abiotic stresses. Therefore, our study provides crucial information about the functional roles of VuRNAi genes and their associated components, which would aid in the development of future cowpeas that are more resilient to biotic and abiotic stress. The manuscript is available as a preprint at this link: doi:10.1101/2023.02.15.528631v1.
Topics: Vigna; RNA Interference; Phylogeny; Gene Expression Regulation, Plant; Plants, Genetically Modified; MicroRNAs; Transcription Factors
PubMed: 38046903
DOI: 10.1155/2023/8832406 -
Cancer Biomarkers : Section a of... 2024P-Element-induced wimpy testis (PIWI) proteins, when in combination with PIWI-interacting RNA (piRNA), are engaged in the epigenetic regulation of gene expression in...
BACKGROUND
P-Element-induced wimpy testis (PIWI) proteins, when in combination with PIWI-interacting RNA (piRNA), are engaged in the epigenetic regulation of gene expression in germline cells. Different types of tumour cells have been found to exhibit abnormal expression of piRNA, PIWIL-mRNAs, and proteins. We aimed to determine the mRNA expression profiles of PIWIL1, PIWIL2, PIWIL3, & PIWIL4, in hepatocellular carcinoma patients, and to associate their expression patterns with clinicopathological features.
METHODS
The expression patterns of PIWIL1, PIWIL2, PIWIL3, PIWIL4 mRNA, was assessed via real-time quantitative polymerase chain reaction (RT-QPCR), on tissue and serum samples from HCC patients, their impact for diagnosis was evaluated by ROC curves, prognostic utility was determined, and In Silico analysis was conducted for predicted variant detection, association with HCC microRNAs and Network Analysis.
RESULTS
Expression levels were significantly higher in both HCC tissue and serum samples than in their respective controls (p< 0.001). Additionally, the diagnostic performance was assessed, Risk determination was found to be statistically significant.
CONCLUSION
PIWIL mRNAs are overexpressed in HCC tissue and serum samples, the expression patterns could be valuable molecular markers for HCC, due to their association with age, tumour grade and pattern. To the best of our knowledge, our study is the first to report the expression levels of all PIWIL mRNA and to suggest their remarkable values as diagnostic and prognostic biomarkers, in addition to their correlation to HCC development. Additionally, a therapeutic opportunity might be also suggested through in silico miRNA prediction for HCC and PIWIL genes through DDX4 and miR-124-3p.
Topics: Male; Humans; Carcinoma, Hepatocellular; Epigenesis, Genetic; Piwi-Interacting RNA; Testis; Liver Neoplasms; RNA, Messenger; Argonaute Proteins
PubMed: 38043006
DOI: 10.3233/CBM-230134 -
The Journal of Biological Chemistry Jan 2024Argonaute (AGO) proteins in all three domains of life form ribonucleoprotein or deoxyribonucleoprotein complexes by loading a guide RNA or DNA, respectively. Since all... (Review)
Review
Argonaute (AGO) proteins in all three domains of life form ribonucleoprotein or deoxyribonucleoprotein complexes by loading a guide RNA or DNA, respectively. Since all AGOs retain a PIWI domain that takes an RNase H fold, the ancestor was likely an endoribonuclease (i.e., a slicer). In animals, most miRNA-mediated gene silencing occurs slicer independently. However, the slicer activity of AGO is indispensable in specific events, such as development and differentiation, which are critical for vertebrates and thus cannot be replaced by the slicer-independent regulation. This review highlights the distinctions in catalytic activation mechanisms among slicing-competent AGOs, shedding light on the roles of two metal ions in target recognition and cleavage. The precision of the target specificity by the RNA-induced silencing complexes is reevaluated and redefined. The possible coevolutionary relationship between slicer-independent gene regulation and AGO-binding protein, GW182, is also explored. These discussions reveal that numerous captivating questions remain unanswered regarding the timing and manner in which AGOs employ their slicing activity.
Topics: Animals; Argonaute Proteins; Ribonucleases; RNA, Guide, CRISPR-Cas Systems; RNA, Small Interfering; RNA-Induced Silencing Complex
PubMed: 38029964
DOI: 10.1016/j.jbc.2023.105499 -
BMC Cancer Nov 2023To investigate the potential correlation between piwi-like RNA-mediated gene silencing 1 (PIWIL1) polymorphisms and susceptibility to epithelial ovarian cancer (EOC).
OBJECTIVE
To investigate the potential correlation between piwi-like RNA-mediated gene silencing 1 (PIWIL1) polymorphisms and susceptibility to epithelial ovarian cancer (EOC).
METHODS
A case-control study was conducted to evaluate the susceptibility of EOC using multinomial logistic regression analysis. The study analyzed the relationship between five functional single nucleotide polymorphisms (SNPs) in the PIWIL1 gene and EOC risk. Genotyping of 288 cases and 361 healthy samples from South China was identified using a TaqMan assay. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated to estimate the relationship between the five selected SNPs and EOC susceptibility.
RESULTS
Among the five SNPs analyzed, the rs10848087 G > A and rs7957349 G > C variants significantly increased the susceptibility of EOC, rs10773771 C > T was associated with a decreased risk of EOC, while the rs35997018 and rs1106042 variants were not in Hardy-Weinberg equilibrium (p < 0.05). The rs10848087 G > A was significantly associated with increased risk of EOC in individuals with metastasis, FIGO stage I and III, low and high pathological grade, tumor numbers ≤ 3 and > 3, tumor size > 3 cm and ≤ 3 cm, pregnant more than 3 times, pre-menopausal status, and strong positive expression of ER (estrogen receptor), PR (progesterone receptor), PAX8 (paired-box 8), wild-type p53 (tumor protein 53), WT1 (Wilm's tumor gene), P16 (cyclin-dependent kinase inhibitor 2A). In addition, rs10848087 G > A enhanced the EOC risk of cases with negative/mild positive expression of wild p53 and Ki67, and with or without mutant p53 expression. The rs7957349 G > C variant was linked to an increased risk of EOC in subgroups with certain characteristics, including age equal or less than 53 years, metastasis, clinical stage I, low pathological grade, tumor number, tumor size, pregnant times, post-menopause, pre-menopause, and strong positive expression of wild p53 and Ki67 (Antigen identified by monoclonal antibody Ki-67), as well as without mutant p53 expression. The rs10773771 CT/TT alleles were identified to have a protective effect on EOC in women aged 53 years or older, as well as in cases with metastasis, advanced clinical stage, high pathological grade, multiple tumors, tumor size equal to or less than 3 cm, history of pregnancy, post-menopausal status, and strong positive expression of ER, PR, wild-type p53, PAX8, WT1, P16, and Ki67. Furthermore, rs10773771 CT/TT also showed a protective effect in patients with negative or mildly positive expression of PR, PAX8, wild-type p53, WT1, and P16, as well as positive expression of mutant p53. Compared to the reference haplotype GCG, individuals harboring haplotypes GTG were found to have a significantly decreased susceptibility to EOC. PIWIL1 was significantly expressed in the thyroid, pituitary, and adrenal glands with rs7957349 CC alleles.
CONCLUSIONS
PIWIL1 rs10848087 and rs7957349 were associated with increased risk of EOC, while rs10773771 may have a protective effect against EOC. These genetic variants may serve as potential biomarkers for EOC susceptibility in the South China population.
Topics: Female; Humans; Argonaute Proteins; Carcinoma, Ovarian Epithelial; Case-Control Studies; East Asian People; Genetic Predisposition to Disease; Genotype; Ki-67 Antigen; Ovarian Neoplasms; Polymorphism, Single Nucleotide; Tumor Suppressor Protein p53; China; Middle Aged
PubMed: 38012622
DOI: 10.1186/s12885-023-11651-2 -
Trends in Cell Biology Nov 2023The endoplasmic reticulum (ER) is central to the processing of luminal, transmembrane, and secretory proteins, and maintaining a functional ER is essential for... (Review)
Review
The endoplasmic reticulum (ER) is central to the processing of luminal, transmembrane, and secretory proteins, and maintaining a functional ER is essential for organismal physiology and health. Increased protein-folding load on the ER causes ER stress, which activates quality control mechanisms to restore ER function and protein homeostasis. Beyond protein quality control, mRNA decay pathways have emerged as potent ER fidelity regulators, but their mechanistic roles in ER quality control and their interrelationships remain incompletely understood. Herein, we review ER-associated RNA decay pathways - including regulated inositol-requiring enzyme 1α (IRE1α)-dependent mRNA decay (RIDD), nonsense-mediated mRNA decay (NMD), and Argonaute-dependent RNA silencing - in ER homeostasis, and highlight the intricate coordination of ER-targeted RNA and protein decay mechanisms and their association with antiviral defense.
PubMed: 38008608
DOI: 10.1016/j.tcb.2023.11.003 -
Plants (Basel, Switzerland) Nov 2023Sex segregation increases the cost of production through seed-based propagation. Therefore, in vitro techniques are an attractive option for clonal propagation,...
Sex segregation increases the cost of production through seed-based propagation. Therefore, in vitro techniques are an attractive option for clonal propagation, especially of hermaphroditic plants. Here, we performed a temporal analysis of the proteome of calli aiming to identify the key players involved in embryogenic callus formation. Mature zygotic embryos used as explants were treated with 20 μM 2,4-dichlorophenoxyacetic acid to induce embryogenic callus. Total proteins were extracted from explants at 0 (zygotic embryo) and after 7, 14, and 21 days of induction. A total of 1407 proteins were identified using a bottom-up proteomic approach. The clustering analysis revealed four distinct patterns of protein accumulation throughout callus induction. Proteins related to seed maturation and storage are abundant in the explant before induction, decreasing as callus formation progresses. Carbohydrate and amino acid metabolisms, aerobic respiration, and protein catabolic processes were enriched throughout days of callus induction. Protein kinases associated with auxin responses, such as SKP1-like proteins 1B, accumulated in response to callus induction. Additionally, regulatory proteins, including histone deacetylase (HD2C) and argonaute 1 (AGO1), were more abundant at 7 days, suggesting their role in the acquisition of embryogenic competence. Predicted protein-protein networks revealed the regulatory role of proteins 14-3-3 accumulated during callus induction and the association of proteins involved in oxidative phosphorylation and hormone response. Our findings emphasize the modulation of the proteome during embryogenic callus initiation and identify regulatory proteins that might be involved in the activation of this process.
PubMed: 38005788
DOI: 10.3390/plants12223891 -
International Journal of Molecular... Nov 2023Heat-shock proteins (HSPs) serve as molecular chaperones in the RNA interference (RNAi) pathway of eukaryotic organisms. In model organisms, Hsp70 and Hsp90 facilitate...
Heat-shock proteins (HSPs) serve as molecular chaperones in the RNA interference (RNAi) pathway of eukaryotic organisms. In model organisms, Hsp70 and Hsp90 facilitate the folding and remodeling of the client protein Argonaute (Ago). However, the specific function of HSPs in the RNAi pathway of (L.) (Lepidoptera: Plutellidae) remains unknown. In this study, we identified and analyzed the coding sequences of and (also known as ). Both PxHsc70-4 and PxHsp83 exhibited three conserved domains that covered a massive portion of their respective regions. The knockdown or inhibition of and in vitro resulted in a significant increase in the gene expression of the dsRNA-silenced reporter gene , leading to a decrease in its RNAi efficiency. Interestingly, the overexpression of PxHsc70-4 and PxHsp83 in DBM, Sf9, and S2 cells resulted in an increase in the bioluminescent activity of dsRNA-silenced luciferase, indicating a decrease in its RNAi efficiency via the overexpression of Hsp70/Hsp90. Furthermore, the inhibition of and in vivo resulted in a significant increase in the gene expression of . These findings demonstrated the essential involvement of a specific quantity of Hsc70-4 and Hsp83 in the siRNA pathway in . Our study offers novel insights into the roles played by HSPs in the siRNA pathway in lepidopteran insects.
Topics: Humans; Animals; RNA Interference; Lepidoptera; HSP90 Heat-Shock Proteins; HSP70 Heat-Shock Proteins; Heat-Shock Proteins; RNA, Small Interfering; RNA, Double-Stranded
PubMed: 38003357
DOI: 10.3390/ijms242216167 -
International Journal of Molecular... Nov 2023The endogenous miRNAs of breast milk are the products of more than 1000 nonprotein-coding genes, giving rise to mature small regulatory molecules of 19-25 nucleotides.... (Review)
Review
The endogenous miRNAs of breast milk are the products of more than 1000 nonprotein-coding genes, giving rise to mature small regulatory molecules of 19-25 nucleotides. They are incorporated in macromolecular complexes, loaded on Argonaute proteins, sequestrated in exosomes and lipid complexes, or present in exfoliated cells of epithelial, endothelial, or immune origins. Their expression is dependent on the stage of lactation; however, their detection depends on progress in RNA sequencing and the reappraisal of the definition of small RNAs. Some miRNAs from plants are detected in breast milk, opening the possibility of the stimulation of immune cells from the allergy repertoire. Each miRNA harbors a seeding sequence, which targets mRNAs, gene promoters, or long noncoding RNAs. Their activities depend on their bioavailability. Efficient doses of miRNAs are estimated to be roughly 100 molecules in the cytoplasm of target cells from in vitro and in vivo experiments. Each miRNA is included in networks of stimulation/inhibition/sequestration, driving the expression of cellular phenotypes. Three types of stress applied during lactation to manipulate miRNA supply were explored using rodent offspring: a foster mother, a cafeteria diet, and early weaning. This review presents the main mature miRNAs described from current mothers' cohorts and their bioavailability in experimental models as well as studies assessing the potential of miR-26 or miR-320 miRNA families to alter offspring phenotypes.
Topics: Female; Humans; MicroRNAs; Milk, Human; Lactation; Nutrition Therapy; Exosomes
PubMed: 38003296
DOI: 10.3390/ijms242216106