Non-Coding RNA

21 pages, 2376 KiB

Open AccessArticle

Anti-HIV-1 Effect of the Fluoroquinolone Enoxacin and Modulation of Pro-Viral hsa-miR-132 Processing in CEM-SS Cells

by Verena Schlösser, Helen Louise Lightfoot, Christine Leemann, Aathma Merin Bejoy, Shashank Tiwari, Jeffrey L. Schloßhauer, Valentina Vongrad, Andreas Brunschweiger, Jonathan Hall, Karin J. Metzner and Jochen Imig

Non-Coding RNA 2025, 11(1), 8; https://doi.org/10.3390/ncrna11010008 - 20 Jan 2025

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Abstract

Background: Despite tremendous advances in antiretroviral therapy (ART) against HIV-1 infections, no cure or vaccination is available. Therefore, discovering novel therapeutic strategies remains an urgent need. In that sense, miRNAs and miRNA therapeutics have moved intensively into the focus of recent HIV-1-related investigations. [...] Read more.

Background: Despite tremendous advances in antiretroviral therapy (ART) against HIV-1 infections, no cure or vaccination is available. Therefore, discovering novel therapeutic strategies remains an urgent need. In that sense, miRNAs and miRNA therapeutics have moved intensively into the focus of recent HIV-1-related investigations. A strong reciprocal interdependence has been demonstrated between HIV-1 infection and changes of the intrinsic cellular miRNA milieu. This interrelationship may direct potential alterations of the host cells’ environment beneficial for the virus or its suppression of replication. Whether this tightly balanced and controlled battle can be exploited therapeutically remains to be further addressed. In this context, the fluoroquinolone antibiotic Enoxacin has been demonstrated as a potent modulator of miRNA processing. Here, we test the hypothesis that this applies also to selected HIV-1-related miRNAs. Methods: We studied the effect of Enoxacin on HIV-1 replication coupled with miRNA qRT-PCR analysis of HIV-1-related miRNAs in CEM-SS and MT-4 T-cells. The effects of miRNA mimic transfections combined with Enoxacin treatment on HIV-1 replication were assessed. Finally, we employed an in vitro DICER1 cleavage assay to study the effects of Enoxacin on a pro-HIV-1 miRNA hsa-miR-132 processing. Results: We established that Enoxacin, but not the structurally similar compound nalidixic acid, exhibits strong anti-HIV-1 effects in the T-cell line CEM-SS, but not MT-4. We provide experimental data that this effect of Enoxacin is partly attributed to the specific downregulation of mature hsa-miR-132-3p, but not other tested pro- or anti-HIV-1 miRNAs, which is likely due to affecting DICER1 processing. Conclusions: Our findings show an anti-retroviral activity of Enoxacin at least in part by downregulation of hsa-miR-132-3p, which may be relevant for future antiviral therapeutic applications by modulation of the RNA interference pathway. Full article

(This article belongs to the Section Small Non-Coding RNA)

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Figure 1
TaqMan qRT-PCR analysis of selected anti- and pro-HIV miRNAs in CEM-SS cells. Cells were treated for 4 and 7 days post-infection with 50 µM final concentration of (a) Enoxacin, (b) nalidixic acid relative to equivalent volume of DMSO control. * p < 0.05 or ** p < 0.01 two-tailed Student’s t-test. Error bars indicate ± 1 s.d. Full article ">Figure 2
Effect of Enoxacin and nalidixic acid on HIV-1 replication in (a) CEM-SS cells over two to ten days p.i. (red arrows indicate time points of qPCR in (c)) measured by p24-ELISA. (b) Fold inhibition of Enoxacin at 50 µM final concentration on HIV-1 replication compared to controls in CEM-SS and (c) MT-4 cells were spin-infected with HIV-1HXB2 at MOI 0.01 and monitored for p24 expression in cell culture supernatant. Untreated cells infected with HIV-1 served as positive, DMSO treatment as negative and no virus as background controls, respectively. (n = 4, technical duplicates, DMSO control = nominator, fold-inhibition = 1), ** p < 0.01, two-tailed Mann Whitney U test. Error bars indicate ± 1 s.d. (d) Pro-HIV-1 miR-132-3p relative expression mirrors the anti-HIV-1 effect of Enoxacin at days 4 and 7 but not control miR-223 (anti-HIV-1) or miR-23-a (T-cell miRNA) compared to DMSO by qRT-PCR. ** p < 0.01 two-tailed Student’s t-test. Error bars indicate ± 1 s.d., red arrows indicate time points of significance for p24-ELISA and corresponding qPCR. Full article ">Figure 3
Hsa-miR-132-3p enhances HIV-1 replication in CEM-SS cells. Cells were spin-infected with HIV-1HXB2 at MOI 0.01 and monitored for p24 expression in supernatant at days 0 and 10 post-infection (p.i.) reverse transfected with miR-mimic or scrambled control at 100 nM plus DMSO. DMSO/mock transfection served as negative and cells and virus alone as positive control. Grey arrow indicates time of transfection. n = 2, * p < 0.05 or ** p < 0.01, two-tailed Student’s t-test. Error bars indicate ± 1 s.d. Full article ">Figure 4
Hsa-miR-132-3p partially rescues Enoxacin-dependent anti-HIV-1 effect in CEM-SS cells. Cells were spin-infected with HIV-1HXB2 at MOI 0.01 and monitored for p24 expression in supernatant over 4 to 10 days post-infection (p.i.) in the presence of 50 µM final concentration of Enoxacin or respective equivalent volume of DMSO control as shown as fold HIV-1 inhibition by p24 ELISA. Untreated cells infected with HIV-1 served as positive control. Cells were reverse transfected in DMSO or Enoxacin treatment with 100 nM concentration of hsa-miR-132-3p mimic or scrambled control. DMSO and Enoxacin treatment with mock transfection served as additional controls. n = 2, two-tailed Mann-Whitney U-test, error bars indicate ± 1 s.d. Full article ">Figure 5
In vitro DICER1 cleavage assay in CEM-SS lysates (a) or recombinant DICER1 protein alone (b). miR-132 hairpin was 5′- (a) or 3′ (b) γ-phosphate labeled and incubated in 100 µM, 300 µM and 1 mM Enoxacin or DMSO control at 37 °C for between 0 and 60 min and PAGE-gel separated. Pre-miR and miR bands were densitometrically quantified and DICER1 processing effect was plotted as normalized ratio to DMSO (a) or % cleavage (b). Note: See full autoradiograms and other replicates <a href="#app1-ncrna-11-00008" class="html-app">Figure S5</a>. (c) In vitro SHAPE-MaP reactivity scores plotted on RNAstructure prediction of pre-miR-132 in the presence of 150 and 300 µM Enoxacin and negative control (0 µM Enoxacin). Red triangles indicate DICER1 and blue triangles DROSHA cleavage sites. Light red sequences indicate significant structural rearrangements (marked by asterisks) determined by SHAPE scores of Enoxacin treatment vs. control. Full article ">

12 pages, 2348 KiB

Open AccessReview

The Role of Long Non-Coding RNA in the Pathogenesis of Psoriasis

by Kajetan Kiełbowski, Anna Jędrasiak, Estera Bakinowska and Andrzej Pawlik

Non-Coding RNA 2025, 11(1), 7; https://doi.org/10.3390/ncrna11010007 - 17 Jan 2025

Viewed by 389

Abstract

Psoriasis is a chronic immune-mediated disease with complex pathogenesis. The altered proliferation and differentiation of keratinocytes, together with the activity of dendritic cells and T cells, are crucial drivers of psoriasis progression. Long non-coding RNAs (lncRNAs) are composed of over 200 nucleotides and [...] Read more.

Psoriasis is a chronic immune-mediated disease with complex pathogenesis. The altered proliferation and differentiation of keratinocytes, together with the activity of dendritic cells and T cells, are crucial drivers of psoriasis progression. Long non-coding RNAs (lncRNAs) are composed of over 200 nucleotides and exert a large variety of functions, including the regulation of gene expression. Under pathological conditions, the expression of lncRNAs is frequently dysregulated. Recent studies demonstrated that lncRNAs significantly affect major cellular processes, and their aberrant expression is likely involved in the pathogenesis of various disorders. In this review, we will discuss the role of lncRNAs in the pathophysiology of psoriasis. We will summarize recent studies that investigated the relationships between lncRNAs and keratinocyte proliferation and pro-inflammatory responses. Full article

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13 pages, 3151 KiB

Open AccessArticle

In Silico Prediction of Maize microRNA as a Xanthine Oxidase Inhibitor: A New Approach to Treating Hyperuricemia Patients

by Manas Joshi and Mohd Mabood Khan

Non-Coding RNA 2025, 11(1), 6; https://doi.org/10.3390/ncrna11010006 - 15 Jan 2025

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Abstract

Introduction: Hyperuricemia is characterized by increased uric acid (UA) in the body. The ability to block xanthine oxidase (XO) is a useful way to check how different bioactive molecules affect hyperuricemia. Previous reports showed the significant effect of corn against hyperuricemia disorder with [...] Read more.

Introduction: Hyperuricemia is characterized by increased uric acid (UA) in the body. The ability to block xanthine oxidase (XO) is a useful way to check how different bioactive molecules affect hyperuricemia. Previous reports showed the significant effect of corn against hyperuricemia disorder with its anti-XO activity. The identification of stable Zea mays miRNA (zma-miR) in humans has opened up a new avenue for speculation about its part in regulating novel human gene targets. Aims: The aim of this study was to investigate the prospects of zma-miRs in XO gene regulation, the possible mechanism, and the interaction analysis of the zma-miR-XO mRNA transcript. Method: Significant features of miRNA-mRNA interaction were revealed using two popular miRNA target prediction software—intaRNA (version 3.3.1) and RNA hybrid (version 2.2.1) Results: Only 12 zma-miR-156 variants, out of the 325 zma-miR’s sequences reported in the miRNA database, efficiently interact with the 3′UTR of the XO gene. Characteristics of miRNA-mRNA interaction were as follows: the positioning of zma-miR-156 variants shows that they all have the same 11-mer binding sites, guanine (G), and uracil (U) loops at the 13th and 14th positions from the 5′ end, and no G: U wobble pairing. These factors are related to the inhibition of functional mRNA expression. Additionally, the zma-miR-156 variants exhibit a single-base variation (SBV), which leads to distinct yet highly effective alterations in their interaction pattern with the XO mRNA transcript and the corresponding free energy values. Conclusion: Therefore, we propose that zma-miR-156 variants may be a promising new bioactive compound against hyperuricemia and related diseases. Full article

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15 pages, 1409 KiB

Open AccessArticle

Plasma Humanin and Non-Coding RNAs as Biomarkers of Endothelial Dysfunction in Rheumatoid Arthritis: A Pilot Study

by Donatella Coradduzza, Sara Cruciani, Biagio Di Lorenzo, Maria Rosaria De Miglio, Angelo Zinellu, Margherita Maioli, Serenella Medici, Gian Luca Erre and Ciriaco Carru

Non-Coding RNA 2025, 11(1), 5; https://doi.org/10.3390/ncrna11010005 - 14 Jan 2025

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Abstract

Background: Rheumatoid arthritis (RA) is a chronic autoimmune disorder associated with an increased risk of cardiovascular disease (CVD), largely driven by peripheral endothelial dysfunction (ED). Humanin, a mitochondrial-derived peptide, has been suggested to play a protective role in endothelial function. However, the relationship [...] Read more.

Background: Rheumatoid arthritis (RA) is a chronic autoimmune disorder associated with an increased risk of cardiovascular disease (CVD), largely driven by peripheral endothelial dysfunction (ED). Humanin, a mitochondrial-derived peptide, has been suggested to play a protective role in endothelial function. However, the relationship between Humanin levels and ED in RA, as well as the interaction between Humanin and non-coding RNAs such as Long Non-Coding RNA GAS5, microRNA-21 (miR-21), and microRNA-103 (miR-103), remains unclear. Objective: This study aimed to investigate the relationship between circulating Humanin levels, non-coding RNAs (GAS5, miR-21, miR-103), and endothelial dysfunction (ED) in patients with RA. Additionally, we explored the correlation between Humanin expression and specific non-coding RNAs (GAS5, miR-21, and miR-103) to better understand their potential role in vascular health. Methods: Peripheral ED was assessed using flow-mediated pulse amplitude tonometry, with Ln-RHI values <0.51 indicating dysfunction. Humanin levels, GAS5, miR-21, and miR-103 were measured in RA patients. Univariate and multivariate analyses were conducted to determine the relationship between these biomarkers and ED. Kaplan–Meier survival analysis and ROC curve analysis were used to assess the prognostic value of Humanin. Results: Higher Humanin levels were significantly associated with better endothelial function (OR = 0.9774, p = 0.0196). Kaplan–Meier analysis demonstrated that higher Humanin levels correlated with improved survival (p < 0.0001). The non-coding RNAs (GAS5, miR-21, and miR-103) did not show significant associations with ED. Conclusions: Humanin is a potential protective biomarker for endothelial dysfunction and survival in RA patients. Further research is needed to explore the interaction between Humanin and non-coding RNAs in the context of vascular health. Full article

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miRNA expression in plasma and exosomes. The expression of miR-21 Panels (A) and (B) and miR-103 Panels (C) and (D) was evaluated in plasma and exosomes. mRNA levels were normalized to U6snRNA. The data are presented as the mean ± SD relative to the control (* p ≤ 0.05, ** p ≤ 0.01, and **** p ≤ 0.0001). Full article ">Figure 2
Lnc-RNA GAS5 expression in plasma (A) and (B). Expression levels were normalized to Glyceraldehyde-3-Phosphate-Dehydrogenase (GAPDH). The data are presented as the mean ± SD relative to the control (**** p ≤ 0.0001). Full article ">Figure 3
Humanin levels in plasma were determined by ELISA. Data are presented as mean ± SD relative to control (**** p ≤ 0.0001). Full article ">Figure 4
ROC curve analysis for Humanin’s predictive value for ED. Full article ">Figure 5
Kaplan–Meier survival analysis to evaluate the prognostic value of Humanin levels for survival outcomes. 0 = patients with serum Humanin concentration < 124.44pg/mL; 1 = patients with serum Humanin concentration ≥ 124.44pg/mL. Full article ">Figure 6
The Kaplan–Meier survival curve showing the survival probabilities for the two groups based on the ROC-determined Humanin levels. Group 0 (blue): Higher survival probability. Group 1 (red dashed): Lower survival probability. The number at risk at different time points is detailed in <a href="#ncrna-11-00005-t005" class="html-table">Table 5</a>. Full article ">

23 pages, 1698 KiB

Open AccessArticle

Integrative Analysis of Whole-Genome and Transcriptomic Data Reveals Novel Variants in Differentially Expressed Long Noncoding RNAs Associated with Asthenozoospermia

by Maria-Anna Kyrgiafini, Maria Katsigianni, Themistoklis Giannoulis, Theologia Sarafidou, Alexia Chatziparasidou and Zissis Mamuris

Non-Coding RNA 2025, 11(1), 4; https://doi.org/10.3390/ncrna11010004 - 14 Jan 2025

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Abstract

Background/Objectives: Asthenozoospermia, characterized by reduced sperm motility, is a common cause of male infertility. Emerging evidence suggests that noncoding RNAs, particularly long noncoding RNAs (lncRNAs), play a critical role in the regulation of spermatogenesis and sperm function. Coding regions have a well-characterized [...] Read more.

Background/Objectives: Asthenozoospermia, characterized by reduced sperm motility, is a common cause of male infertility. Emerging evidence suggests that noncoding RNAs, particularly long noncoding RNAs (lncRNAs), play a critical role in the regulation of spermatogenesis and sperm function. Coding regions have a well-characterized role and established predictive value in asthenozoospermia. However, this study was designed to complement previous findings and provide a more holistic understanding of asthenozoospermia, this time focusing on noncoding regions. This study aimed to identify and prioritize variants in differentially expressed (DE) lncRNAs found exclusively in asthenozoospermic men, focusing on their impact on lncRNA structure and lncRNA–miRNA–mRNA interactions. Methods: Whole-genome sequencing (WGS) was performed on samples from asthenozoospermic and normozoospermic men. Additionally, an RNA-seq dataset from normozoospermic and asthenozoospermic individuals was analyzed to identify DE lncRNAs. Bioinformatics analyses were conducted to map unique variants on DE lncRNAs, followed by prioritization based on predicted functional impact. The structural impact of the variants and their effects on lncRNA–miRNA interactions were assessed using computational tools. Gene ontology (GO) and KEGG pathway analyses were employed to investigate the affected biological processes and pathways. Results: We identified 4173 unique variants mapped to 258 DE lncRNAs. After prioritization, 5 unique variants in 5 lncRNAs were found to affect lncRNA structure, while 20 variants in 17 lncRNAs were predicted to disrupt miRNA–lncRNA interactions. Enriched pathways included Wnt signaling, phosphatase binding, and cell proliferation, all previously implicated in reproductive health. Conclusions: This study identifies specific variants in DE lncRNAs that may play a role in asthenozoospermia. Given the limited research utilizing WGS to explore the role of noncoding RNAs in male infertility, our findings provide valuable insights and a foundation for future studies. Full article

(This article belongs to the Special Issue Exploring Non-coding RNAs: Insights into Male Infertility)

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21 pages, 1908 KiB

Open AccessReview

Perspectives in MicroRNA Therapeutics for Cystic Fibrosis

by Alessia Finotti and Roberto Gambari

Non-Coding RNA 2025, 11(1), 3; https://doi.org/10.3390/ncrna11010003 - 12 Jan 2025

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Abstract

The discovery of the involvement of microRNAs (miRNAs) in cystic fibrosis (CF) has generated increasing interest in the past years, due to their possible employment as a novel class of drugs to be studied in pre-clinical settings of therapeutic protocols for cystic fibrosis. [...] Read more.

The discovery of the involvement of microRNAs (miRNAs) in cystic fibrosis (CF) has generated increasing interest in the past years, due to their possible employment as a novel class of drugs to be studied in pre-clinical settings of therapeutic protocols for cystic fibrosis. In this narrative review article, consider and comparatively evaluate published laboratory information of possible interest for the development of miRNA-based therapeutic protocols for cystic fibrosis. We consider miRNAs involved in the upregulation of CFTR, miRNAs involved in the inhibition of inflammation and, finally, miRNAs exhibiting antibacterial activity. We suggest that antago-miRNAs and ago-miRNAs (miRNA mimics) can be proposed for possible validation of therapeutic protocols in pre-clinical settings. Full article

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Figure 1
MicroRNA therapeutics. Picture created using <a href="http://Bio-Render.com" target="_blank">Bio-Render.com</a> (7 November 2024). Full article ">Figure 2
MicroRNA Therapeutics: the anti-miRNA approach. A forced inhibition of the miRNA activity can be obtained using antago-miRNA (anti-miRNA) oligonucleotides (AMOs) (e.g., DNA, RNA, and nucleic acids analogs such as LNA, PNA, 2′-MOE), delivered with vectors (A) or bioconjugated for an increased cellular uptake (e.g., R-PNAs) and/or a targeted delivery (B). MicroRNA inhibition can also be achieved using anti-miRNA sponge RNA sequences that contain multiple microRNA binding sites (C). The approach based on the use of zipper oligonucleotides is shown in panel (D) [<a href="#B31-ncrna-11-00003" class="html-bibr">31</a>]. The binding between the miRNA and the anti-miRNA molecules leads to the inactivation of the miRNA, as it can no longer bind to its molecular target, i.e., messenger RNA, thus increasing protein production. Picture created using <a href="http://Bio-Render.com" target="_blank">Bio-Render.com</a> (7 November 2024). Full article ">Figure 3
MicroRNA therapeutics: the “miRNA-masking” approach. The down-regulation of microRNA functions is obtained through miRNA masking oligonucleotides and analogs delivered to cells (A,B), which act by masking the miRNAs binding site of target mRNAs through a direct hybridization of the miRNA “mask” with the 3′UTR region of mRNA. Created using <a href="http://Bio-Render.com" target="_blank">Bio-Render.com</a> (7 November 2024). Full article ">Figure 4
MicroRNA therapeutics: the “miRNA-replacement” approach. This strategy is based on the use of molecules (mature double-strand microRNA mimics or pre-miRNA oligonucleotides) that can restore physiological levels of miRNA with consequent inhibition of mRNA translation. Created using <a href="http://Bio-Render.com" target="_blank">Bio-Render.com</a> (7 November 2024). Full article ">Figure 5
Mechanism of action and molecular targets of miR-93-5p according to the reports published by Fabbri et al. [<a href="#B106-ncrna-11-00003" class="html-bibr">106</a>], Xu et al. [<a href="#B109-ncrna-11-00003" class="html-bibr">109</a>], and Gao et al. [<a href="#B111-ncrna-11-00003" class="html-bibr">111</a>]. MicroRNA miR-93-5p directly interacts with IL-8 mRNA, thereby inhibiting IL-8 production and release [<a href="#B106-ncrna-11-00003" class="html-bibr">106</a>]; in addition, miR-93-5p inhibits IRAK1, thereby preventing NF-kB activation and expression of NF-kB-dependent genes, such as IL-8 [<a href="#B109-ncrna-11-00003" class="html-bibr">109</a>]; in addition, miR-93-5p is able to interact with TLR-4 mRNA [<a href="#B112-ncrna-11-00003" class="html-bibr">112</a>], thereby down-regulating the NF-kB pathway. Full article ">

24 pages, 5992 KiB

Open AccessArticle

LncRNA 3222401L13Rik Is Upregulated in Aging Astrocytes and Regulates Neuronal Support Function Through Interaction with Npas3

by Sophie Schröder, M. Sadman Sakib, Dennis M. Krüger, Tonatiuh Pena, Susanne Burkhardt, Anna-Lena Schütz, Farahnaz Sananbenesi and André Fischer

Non-Coding RNA 2025, 11(1), 2; https://doi.org/10.3390/ncrna11010002 - 9 Jan 2025

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Abstract

Aging leads to cognitive decline and increased risk of neurodegenerative diseases. While molecular changes in central nervous system (CNS) cells contribute to this decline, the mechanisms are not fully understood. Long non-coding RNAs (lncRNAs) are key regulators of cellular functions. Background/Objectives: The roles [...] Read more.

Aging leads to cognitive decline and increased risk of neurodegenerative diseases. While molecular changes in central nervous system (CNS) cells contribute to this decline, the mechanisms are not fully understood. Long non-coding RNAs (lncRNAs) are key regulators of cellular functions. Background/Objectives: The roles of lncRNAs in aging, especially in glial cells, are not well characterized. Methods: We investigated lncRNA expression in non-neuronal cells from aged mice and identified 3222401L13Rik, a previously unstudied lncRNA, as upregulated in astrocytes during aging. Results: Knockdown of 3222401L13Rik in primary astrocytes revealed its critical role in regulating genes for neuronal support and synapse organization, a function conserved in human iPSC-derived astrocytes. A 3222401L13Rik interacts with the transcription factor Neuronal PAS Domain Protein 3 (Npas3), and overexpression of Npas3 rescues deficits in astrocytes lacking 3222401L13Rik. Conclusions: These data suggest that 3222401L13Rik upregulation may help delay age-related cognitive decline. Full article

(This article belongs to the Section Clinical Applications of Non-Coding RNA)

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26 pages, 2100 KiB

Open AccessReview

RNA Metabolism and the Role of Small RNAs in Regulating Multiple Aspects of RNA Metabolism

by Pranav Dawar, Indra Adhikari, Swarupa Nanda Mandal and Bhumika Jayee

Non-Coding RNA 2025, 11(1), 1; https://doi.org/10.3390/ncrna11010001 - 24 Dec 2024

Viewed by 996

Abstract

RNA metabolism is focused on RNA molecules and encompasses all the crucial processes an RNA molecule may or will undergo throughout its life cycle. It is an essential cellular process that allows all cells to function effectively. The transcriptomic landscape of a cell [...] Read more.

RNA metabolism is focused on RNA molecules and encompasses all the crucial processes an RNA molecule may or will undergo throughout its life cycle. It is an essential cellular process that allows all cells to function effectively. The transcriptomic landscape of a cell is shaped by the processes such as RNA biosynthesis, maturation (RNA processing, folding, and modification), intra- and inter-cellular transport, transcriptional and post-transcriptional regulation, modification, catabolic decay, and retrograde signaling, all of which are interconnected and are essential for cellular RNA homeostasis. In eukaryotes, sRNAs, typically 20–31 nucleotides in length, are a class of ncRNAs found to function as nodes in various gene regulatory networks. sRNAs are known to play significant roles in regulating RNA population at the transcriptional, post-transcriptional, and translational levels. Along with sRNAs, such as miRNAs, siRNAs, and piRNAs, new categories of ncRNAs, i.e., lncRNAs and circRNAs, also contribute to RNA metabolism regulation in eukaryotes. In plants, various genetic screens have demonstrated that sRNA biogenesis mutants, as well as RNA metabolism pathway mutants, exhibit similar growth and development defects, misregulated primary and secondary metabolism, as well as impaired stress response. In addition, sRNAs are both the “products” and the “regulators” in broad RNA metabolism networks; gene regulatory networks involving sRNAs form autoregulatory loops that affect the expression of both sRNA and the respective target. This review examines the interconnected aspects of RNA metabolism with sRNA regulatory pathways in plants. It also explores the potential conservation of these pathways across different kingdoms, particularly in plants and animals. Additionally, the review highlights how cellular RNA homeostasis directly impacts adaptive responses to environmental changes as well as different developmental aspects in plants. Full article

(This article belongs to the Special Issue Non-Coding RNA and Their Regulatory Roles in Plant)

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Figure 1
sRNA biogenesis models for plants and animals. (A) miRNA biogenesis pathway—RNAPII transcribes the MIR specific. Primary miRNA (pri-miRNA) is then spliced into a single hairpin structure by DICER-LIKE 1, which furthermore gets cleaved into a miRNA duplex of 21 nt with the help of nuclear dicing bodies (D-bodies) including DICER-LIKE 1/3/4 (DCL1/3/4), HYPONASTIC LEAVES 1 (HYL1), SERRATE (SE), and TOUGH (TGH). The duplex is then methylated at the 3′ ends catalyzed with HUA ENHANCER1. With the assistance of HEAT SHOCK PROTEIN70/90 (HSP70/90), the mature miRNA gets loaded into the ARGONAUTE1/10 (AGO1/10) (also known as RISC) based on the specific miRNAs. (B) Secondary siRNAs (phasiRNA, tasiRNA, and easiRNA) biogenesis model—After PHAS loci, TAS loci, and active transposons are transcribed via RNAP II, AGO1/7-loaded mature miRNA cleaves the transcribed mRNA. 5′ fragment of the cleaved mRNA degrades, while the 3′ strand is converted into a double strand with the help of RDR6. SGS3 and SDE5 help in recruiting RDR6 to the recognition site. DCL4/3 participates in ta-siRNA and phasiRNA production, whereas DCL2/4 participates in easiRNA production. The 21–24 nt mature siRNA strand then gets loaded into AGO1/7 for downstream gene regulation. (C) Plant endogenous siRNA biogenesis—The transposable elements, repetitive regions, or gene introns get transcribed with RNAP IV, which then gets converted into dsRNA with the help of RDR2/4. The dsRNA then gets cleaved into 20–24 nt fragments with the help of DCL2/3/4. HSP90 then helps to load the mature siRNA strand in the respective AGO4/6/9, based on their origin. (D) Canonical and miRNA Biogenesis in Animals—After the transcription of the MIR-specific locus with DNA-dependent RNA polymerase II, pri-miRNAs were converted to single hairpin-like structures (pre-miRNAs) with the help of Drosha. The pre-miRNA transported into the cytoplasm with Exportin1/5 proteins then gets further cleaved into miRNA duplexes by Dicer proteins. The pathways create this miRNA duplex without Dorsal/Dicer acting upon the primary miRNA. The mature miRNA strand then gets loaded in the AGO2. (E) Endo- and exogenous modes of siRNA production in Caenorhabditis elegans—siRNAs derived from ssRNA, and dsRNA are loaded into primary Argonaute proteins, ERGO-1 and RDE1, respectively. The loaded primary Argonaute protein, with the help of RRF1 and Mutator, mediates the conversion of 26 nt long 5′-guanosine siRNA into 22G siRNA. The produced siRNAs are then loaded into the secondary Argonaute proteins for downstream gene silencing. (F) Biogenesis of piRNA and the regulatory ping pong cycle of biogenesis in Drosophila melanogaster—piRNA gene sequences are marked by an upstream Ruby motif. The piRNA precursors are transcribed by RNAP II and then exported to the cytoplasm. These precursors are then processed by endonuclease Zucchini and an unknown 3′–5′ exonuclease. Via DmHen1/Pimet methyltransferase, the 3′ end of the mature piRNA gets 2′-O-methylated. The mature piRNA gets loaded into PIWI, forming piRISC to regulate methylation of TEs. Apart from PIWI protein alone, some gets loaded into Aub, which then initiates the ping pong cycle of biogenesis. Aub loaded with piRNA and AGO3 loaded with secondary piRNA repress TE activity through DNA cytosine methylation. PIWI-related Gene 1 (PRG-1) is required for primary piRNA activity, whereas HRDE-1 (Heritable RNA interference (RNAi) deficient protein-1) is the Argonaute protein that carries RdRP-amplified 22 nt, 5′-guanosine siRNA (22-GsiRNA). (Modified from [<a href="#B16-ncrna-11-00001" class="html-bibr">16</a>]). Full article ">Figure 2
PTGS and TGS Mode of Action for miRNA, siRNA, and piRNA in Plants and Animals. (A,B) Post-transcriptional gene silencing of plant and animal miRNAs through mRNA cleavage, RNA decay, and translational repression. (C) Exogenous siRNA is only able to participate in PTGS through mRNA cleavage. (D) Plant and animals endogenous siRNA and piRNAs, the loaded AGO protein, after being transported back to the nucleus, target nascent RNA Pol-V transcripts (line represented in red) through complementary siRNA and form the RdDM complex (RNA-dependent DNA methylation). GW/WG protein, associated with RNAP V, KTF1 acts as an organizer by coordinating with AGO and 5-meC (5-methylCytosine). Similarly, the AGO-associated protein RDM1 interacts with DRM2, a RdDM complex catalytically active de novo methyltransferase, and binds with single-stranded methylated DNA. DRM3, a catalytically inactive paralogue of DRM2, is also known to be involved in the RdDM complex, but its function is still unknown. After all these proteins are localized, DRM2 catalyzes methylation of cytosine in all sequence contexts. (Modified from [<a href="#B16-ncrna-11-00001" class="html-bibr">16</a>,<a href="#B18-ncrna-11-00001" class="html-bibr">18</a>]). Full article ">Figure 3
miRNA-mediated mRNA decay pathway in plants and animals. (A) miRNA binds to the complementary site in the Open Reading frame and induces endonucleolytic cleavage at the splice site (between the nucleotides 10 and 11). The 5′ fragment is uridylated by HUA Enhancer 1 Suppressor 1 (HESO 1) and is degraded by XRN4 in a 5′ to 3′ direction. Similarly, the 3′ cleaved fragments are degraded by XRN4 without uridylation. (B) miRNA, after attaching with the activated mRNA, recruits CCR4-NOT and PAN2-PAN3 deadenylase complexes to target mRNAs via the GW182 protein. These deadenylated mRNAs are then oligouridylated by TUT4/7, thus starting the general mRNA decay in mammals. Apart from deadenylation, GW182 can also promote dissociation of PAPB (poly(A) binding protein). DDX6 (the de-capping activators) are then recruited onto the CCR4-NOT complex. This helps the DCP2 enzyme in removing the 5′ 7-methylated guanine cap. Finally, XRN1 acts on the uncapped uridylated mRNA strand by performing 5′-3′ exonucleolytic decay. (Modified from [<a href="#B127-ncrna-11-00001" class="html-bibr">127</a>]). Full article ">

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Non-Coding RNA, Volume 11, Issue 1 (February 2025) – 8 articles

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