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Results for “Small noncoding RNA”

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At least 19 recordsLinked to original sources

PubMed2024

N6-methyladenosine modification of a parvovirus-encoded small noncoding RNA facilitates viral DNA replication through recruiting Y-family DNA polymerases.

Human bocavirus 1 (HBoV1) is a human parvovirus that causes lower respiratory tract infections in young children. It contains a single-stranded (ss) DNA genome of ~5.5 kb that encodes a small noncoding RNA of 140 nucleotides known as bocavirus-encoded small RNA (BocaSR), in addition to viral proteins. Here, we determined the secondary structure of BocaSR in vivo by using DMS-MaPseq. Our findings reveal that BocaSR undergoes N6-methyladenosine (m6A) modification at multiple sites, which is critical for viral DNA replication in both dividing HEK293 cells and nondividing cells of the human airway epithelium. Mechanistically, we found that m6A-modified BocaSR serves as a mediator for recruiting Y-family DNA repair DNA polymerase (Pol) η and Pol κ likely through a direct interaction between BocaSR and the viral DNA replication origin at the right terminus of the viral genome. Thus, this report represents direct involvement of a viral small noncoding RNA in viral DNA replication through m6A modification.

Humans↗

PubMed2026

The Small Noncoding RNA, RsaC, Is Essential for Staphylococcus aureus Virulence.

BACKGROUND: Bacterial small noncoding RNAs (sRNAs) play critical roles in virulence, stress adaptation, and host-pathogen interactions. Transcriptomic analyses during infection can help reveal pathogen-derived sRNAs required for pathogenesis, providing valuable insights for the development of novel therapeutic strategies. However, the low abundance of pathogen biomass within the host tissues poses a significant challenge for such analyses. METHODS: We employed 2-step cell disruption to enrich Staphylococcus aureus cells from infected mouse organs and conducted RNA sequencing (RNA-seq) analysis to examine staphylococcal sRNAs expressed during infection. qRT-PCR was used to confirm the gene expression. A knockout mutant of highly expressed sRNA, RsaC, was generated, and RNA-seq under in vivo as well as in vitro aerobic and anaerobic conditions were compared between the wild-type and ΔrsaC strains. Virulence of S. aureus was assessed using both mouse and silkworm survival assays. RESULTS: We identified RsaC as one of the most highly expressed sRNAs in mouse organs with consistent increment over time postinfection. Through gene disruption and complementation, we demonstrated that RsaC is an independent virulence determinant required for full pathogenicity of S. aureus in a murine infection model. In addition, RsaC influenced gene expression in response to oxygen availability and host-associated stress. Further analysis revealed that mutation of 2 genes downregulated in ΔrsaC in vivo, NWMN_RS03420 (sodium: proton antiporter) and NWMN_RS12015 (hypothetical protein), reduced S. aureus virulence in a silkworm model. CONCLUSIONS: These findings identify RsaC as a novel independent virulence determinant that supports S. aureus adaptation within the host.

Animals↗

PubMed2025

An RNA polymerase III tissue and tumor atlas uncovers context-specific activities linked to 3D epigenome regulatory mechanisms.

RNA polymerase III (Pol III) produces a plethora of small noncoding RNA species involved in diverse cellular processes, from transcription regulation and splicing to RNA stability, translation, and proteostasis. Though Pol III activity is broadly coupled with cellular demands for protein synthesis and growth, a more precise understanding of gene-level dynamics and context-specific expression patterns remains missing, in part due to challenges related to sequencing and mapping Pol III-derived small ncRNAs. Here, we establish a predictive multi-tissue map of human Pol III activity across 19 tissues and 23 primary cancer subtypes by comprehensively profiling the chromatin accessibility of canonical Pol III-transcribed gene classes. Our framework relies on the unique relationship between gene accessibility and Pol III transcription, inferring activity through uniform binary classification of ATAC-seq enrichment at Pol III-transcribed genes. By characterizing multi-context gene uniformity, we provide a definition of the core Pol III transcriptome, broadly active across specialized tissues, and catalog genes with varied levels of context specificity. Our genomic Pol III atlas uncovers variable levels of activity across tissues, including sharp contraction of the Pol III transcriptome in heart and brain tissues and frequent expansion across diverse cancers. We show that both tissue- and tumor-specific genes are significantly enriched within lamina-associated domains (LADs), and that aberrant expression of nuclear lamin proteins is sufficient to induce Pol III-emergent patterns at tumor-specific genes. Together, these findings link Pol III dynamics to subnuclear compartmentalization and provide a resource for better understanding Pol III expansion and small RNA biogenesis in cancer.

Journal Article↗

PubMed2025

Identification and Analysis of Small Nucleolar RNAs by Real-Time Quantitative PCR.

One of the greatest scientific achievements of the twenty-first century is the completion of The Human Genome Project (HGP). Thereafter, we came to know that the human genome codes nearly 2% for making proteins and thus named as coding genes, suggesting the rest of the genome as noncoding or junk. However, research in the past two decades has shown and established that noncoding RNAs are major contributors of regulating and modulating the various function of cells as well as tissues. Noncoding RNAs can be classified as basis of their sizes in two categories, long noncoding RNAs (>200 nt) and small noncoding RNAs (<200 nt). Small nucleolar RNAs (snoRNAs) are part of the small noncoding RNA family and primarily reside inside the nucleus of eukaryotes. Sno RNAs can be divided into two major categories based on their distinguished structure and function; these are C/D box and HACA box snoRNAs. They participate in the posttranscriptional modifications on ribosomal RNAs (r-RNAs), transfer RNAs (t-RNAs), messenger RNAs (m-RNAs), and small nuclear RNAs (snRNAs). Sno RNAs act as guide RNAs to modify other noncoding RNAs by pseudouridylation or 2'O ribomethylation. We discussed in this protocol about one of the widely used techniques for detection and analysis of snoRNAs, i.e., real-time quantitative PCR (RT-qPCR).

RNA, Small Nucleolar↗

PubMed2026

Rop-Mediated Suppression of RpoS Production Increases Resistance to Nitric Oxide.

We identified the RNA-binding protein Rop, encoded on the pOSAK1 plasmid of enterohaemorrhagic Escherichia coli (EHEC), as a novel factor that enhances nitric oxide (NO) resistance, although it has previously been reported to regulate plasmid copy number. The Rop-induced increase in NO resistance was significantly reduced in several small noncoding RNA (sRNA) gene-deficient EHEC mutants. Among these sRNAs, DsrA, ArcZ, and RprA were directly involved in the translational regulation of rpoS expression, suggesting that Rop modulates rpoS expression through sRNAs. To examine this mechanism, we generated sRNA gene-deficient mutants with an additional deletion of the 5' untranslated region (5' UTR) of rpoS, which is required for translational regulation. The increase in NO resistance by Rop was restored in the double mutant, suggesting that this phenotype is mediated by Rop-dependent interactions between sRNAs and the 5' UTR of rpoS mRNA. Furthermore, Rop promoted rpoS mRNA degradation, an effect that likely suppresses RpoS production and may thereby enhance NO resistance. Finally, an hfq-deficient EHEC mutant exhibited no increase in NO resistance in the presence of Rop, indicating that Hfq is essential for Rop-mediated NO resistance.

Nitric Oxide↗

PubMed2024

Small noncoding RNAs and sperm nuclear basic proteins reflect the environmental impact on germ cells.

BACKGROUND: Molecular techniques can complement conventional spermiogram analyses to provide new information on the fertilizing potential of spermatozoa and to identify early alterations due to environmental pollution. METHODS: Here, we present a multilevel molecular profiling by small RNA sequencing and sperm nuclear basic protein analysis of male germ cells from 33 healthy young subjects residing in low and high-polluted areas. RESULTS: Although sperm motility and sperm concentration were comparable between samples from the two sites, those from the high-pollution area had a higher concentration of immature/immune cells, a lower protamine/histone ratio, a reduced ability of sperm nuclear basic proteins to protect DNA from oxidative damage, and an altered copper/zinc ratio in sperm. Sperm levels of 32 microRNAs involved in intraflagellar transport, oxidative stress response, and spermatogenesis were different between the two areas. In parallel, a decrease of Piwi-interacting RNA levels was observed in samples from the high-polluted area. CONCLUSIONS: This comprehensive analysis provides new insights into pollution-driven epigenetic alterations in sperm not detectable by spermiogram.

Male↗

PubMed2020

LINC00922 regulates epithelial-mesenchymal transition, invasive and migratory capacities in breast cancer through promoting NKD2 methylation.

Breast cancer ranks as the major reason for mortality in women populations, accounting for 23% of all cancer deaths. One in every three Asian women encounters the risk of this cancer in their lifetime. Long intergenic non-coding RNAs (lincRNAs) have emerged as tumor promoters and suppressors. The molecular mechanism of breast cancer remains elusive. Therefore, the current study aimed to explore the role lincRNA LINC00922 plays in the development of breast cancer. Breast cancer tissues and adjacent tissues were obtained from 109 patients with breast cancer. The RNA extraction and quantification and immunohistochemical staining characterized the high expression of LINC00922 and low expression of NKD2 in breast cancer tissues in comparison to its adjacent counterparts. Furthermore, the ectopic expression and knockdown experiments were conducted to figure out the in vivo and in vitro effects of LINC00922 on breast cancer progression. The ectopically expressed LINC00922 activated the Wnt signaling pathway, promoted epithelial-mesenchymal transition, cell proliferative, invasive and migratory capacities, tumor growth and metastasis. Additionally, the RIP and ChIP assay identified that LINC00922 recruited DNMT1, DNMT3A and DNMT3B proteins in the promoter region of NKD2 to promote NKD2 promoter methylation, thus reducing the NKD2 expression. Moreover, the Wnt signaling pathway was activated subsequent to NKD2 silencing, which was reversed by LINC00922 silencing. Lastly, the anti-oncogenic effects of LINC00922 inhibition was antagonized after NKD2 knocked down. The current study provides evidence that LINC00922 acts as a tumor promoter by promoting NKD2 methylation. Hopefully, it provides a novel potential gene target for the treatment of breast cancer.

Adaptor Proteins, Signal Transducing↗

PubMed2025

A Glimpse of Noncoding RNAs: Secondary Structure, Emerging Trends, and Potential Applications in Human Diseases.

An appealing strategy for the treatment of several diseases is the therapeutic targeting of noncoding RNAs (ncRNAs), such as microRNAs (miRNAs) and long noncoding RNAs (lncRNAs). Many antisense oligonucleotides and small interfering RNAs have been tested in clinical studies over the past 10 years, and several of these have received FDA approval. However, trial results have thus far been mixed, with some studies reporting strong effects and others showing low effectiveness or side effects, including toxicity. Clinical trials for alternative entities like antimiRNAs are underway, and interest in lncRNA-based therapies is constantly growing. From this perspective, we discuss the basic overview of ncRNAs, their significant role as therapeutic biomarkers against different diseases, and the role of secondary structure in noncoding RNAs.

Humans↗

PubMed2026

Long noncoding RNA GAS5 disrupts intestinal epithelial barrier function by increasing small vault RNA levels.

Disruptions in the integrity of the intestinal epithelium occur commonly in inflammatory bowel disease (IBD) and critical surgical disorders, but the underlying mechanisms remain largely unknown. Here we identified long noncoding RNA GAS5 as a repressor of intestinal mucosal growth and the function of the gut epithelial barrier. The levels of tissue GAS5/Gas5 increased in mouse intestinal mucosa after colitis and septic stress, as well as in human intestinal mucosa from patients with IBD. Transient and tissue-specific knockdown of Gas5 in mice using CRISPR/Cas9 enhanced the renewal of the mucosa of the small intestine, increased the levels of tight junction (TJ) proteins ZO-1, ZO-2, claudin-1, and claudin-2, and improved gut barrier function. Conversely, ectopic overexpression of GAS5 in intestinal organoids and in cultured intestinal epithelium cells decreased the levels of these TJ proteins and caused epithelial barrier dysfunction. Mechanistic studies revealed that GAS5 acted as a transcriptional enhancer of the gene (2. AUTHOR: Do you mean "genes"?) encoding small noncoding vault RNAs (vtRNAs) and that GAS5 repressed TJ expression by increasing the levels of vtRNAs. Together, our results indicate that GAS5 disrupts the integrity of the intestinal epithelium by impairing mucosal growth and epithelial barrier function and that it represses TJ expression, at least in part, via vtRNAs.

Animals↗

PubMed2025

Popcorn: prediction of short coding and noncoding genomic sequences in prokaryotes.

SUMMARY: The most challenging prokaryotic genes to identify often correspond to short ORFs (sORFs) encoding small proteins or to noncoding RNAs. RNA-seq experiments commonly evince small transcripts that do not correspond to annotated genes and are candidates for novel coding sORFs or small regulatory RNAs, but it can be difficult to accurately assess whether the numerous small transcripts are coding or not. We present Popcorn (PrOkaryotic Prediction of Coding OR Noncoding), a novel machine learning method for determining whether prokaryotic sequences are coding or noncoding. We find that Popcorn is effective in distinguishing coding from noncoding sequences, including coding sORFs and noncoding RNAs. AVAILABILITY AND IMPLEMENTATION: Freely available for use on the web at https://cs.wellesley.edu/∼btjaden/Popcorn. Source code available at https://github.com/btjaden/Popcorn and https://doi.org/10.5281/zenodo.15120075.

Open Reading Frames↗

PubMed2025

Nascent and Mature RNA Profiling by Subcellular Fractionation in Human Cells.

Transcription and RNA decay determine steady-state RNA levels in cells available for translation and RNA-mediated regulatory functions. Both processes can be assessed by various techniques, for majority, based on RNA labelling or chromatin immunoprecipitation, but require a high level of expertise. Here, we describe a cost-effective, fast, and simple protocol that enables the profiling of nascent and mature RNA in the cytoplasm, nucleoplasm, and chromatin through subcellular fractionation. The workflow can include α-amanitin inhibition of RNA Polymerase II to assess nascent RNAs as a proxy of transcriptional activity, or it can be used without this treatment to investigate distribution of partially processed or mature transcripts across distinct subcellular compartments. It is applicable for studying any of RNA biotypes, including small and long noncoding RNAs, mRNAs, and their splice variants, on both transcript-specific and transcriptome-wide scales. Nascent or mature RNAs isolated from each fraction can be further analyzed by any technique of choice (northern blot, reverse transcription, RNA sequencing).

Humans↗

PubMed2025

Quantitative Real-Time PCR for Circular RNA Detection and Analysis.

In eukaryotes, nearly 2% of the genome represented by the coding proteins. However, emerging evidence suggest more than 75% of the human genome referred to as noncoding part also plays a crucial role in governing major regulatory pathways. Noncoding RNAs can be categorized into several groups, such as microRNAs (miRNAs), small nuclear RNA (snRNAs), small nucleolar RNA (snoRNAs), transfer RNA (tRNA), and circular RNA (circRNAs), which contribute to this regulatory landscape. Circular RNAs (circRNAs) are identified as a new class of regulatory noncoding RNAs with gene regulatory roles by acting as miRNA or RNA binding protein sponges or interacting with proteins. Researchers employ quantitative real-time PCR methods to examine circular RNA expression utilizing divergent primers for identification and quantification.

RNA, Circular↗

PubMed1992

The 10 S BC-1 ribonucleoprotein particle contains identifier sequence-binding proteins that interact with an array of GCAAG/CTTGC motifs between split promoter sequences for RNA polymerase III.

BC-1 RNA is a brain-specific small RNA transcript of identifier sequences present in the somas and dendrites of neurons. We recently reported that the RNA is complexed with a protein(s) to form a 10 S ribonucleoprotein particle (Kobayashi, S., Goto, S., and Anzai, K. (1991) J. Biol. Chem. 266, 4726-4730). We demonstrate here that this 10 S BC-1 ribonucleoprotein particle contains a DNA-binding protein(s) (Bp-1 protein) capable of interacting with a region between split promoter sequences for RNA polymerase III within the identifier sequences. The region has short inverted repeats: a perfect octanucleotide repeat (GCGCTTGCCTAGCAAGCGC) and an imperfect heptanucleotide repeat (GCCTAGCAAGCGCAAGGC), each of which contains a GCAAG/CTTGC motif. We also demonstrate that the binding of this protein either to the array of pentamer motifs or to BC-1 RNA is mutually exclusive. The molecular masses of photo-cross-linking adducts of Bp-1 protein to a 32P-labeled GCAAG/CTTGC motif-specific probe were estimated to be about 31 and 36 kDa, indicating that two species of Bp-1 proteins may be present in the brain.

Animals↗

PubMed1992

Nucleotide sequence and genetic map of cowpea severe mosaic virus RNA 2 and comparisons with RNA 2 of other comoviruses.

We report the nucleotide sequence of cowpea severe mosaic comovirus (CPSMV) genomic RNA 2. The molecule is composed of 3732 nucleotide (nt) residues, exclusive of the polyadenylate at the 3' end. Only one of the six reading frame registers has a long open reading frame, from nt 255 to nt 3260 in the polarity of encapsidated RNA and corresponding to a polyprotein of 1002 amino acid residues (aa). As has been reported for other comoviruses, a second in-frame AUG, at nt position 531, apparently also initiates translation, at least in vitro. Multiple alignments of the deduced CPSMV polyprotein aa sequence with those of bean pod mottle comovirus (BPMV), cowpea mosaic comovirus (CPMV), and red clover mottle comovirus (RCMV) were consistent with a similar size for each of the three genes: the putative movement protein, beginning at the second in-frame AUG, the large coat protein (L), and the small coat protein. Identical nucleotide sequences in the terminal noncoding regions of RNA 2 of the four viruses are limited to 9 nt at the 5' end and the 3' polyadenylate. However, extensive similarities in sequence and potential structure were found. For all three genes and the 5' untranslated region, CPSMV and BPMV are more similar to each other than either is to CPMV or RCMV, the last two being similar to each other. Observed similarities predict that both cleavage sites in the CPSMV RNA 2 polyprotein are at glutamine-serine dipeptides. A sequence of 16 aa at the amino terminus of L, determined by automated Edman degradation, matched a region of the deduced aa sequence in the polyprotein and is consistent with cleavage at the predicted glutamine-serine dipeptide.

Amino Acid Sequence↗

PubMed1992

Developmental change in subcellular location of Bp-1 protein with an ability to interact with both identifier sequence and its brain-specific transcript, BC-1 RNA.

Identifier sequences are transcribed to generate a brain-specific BC-1 RNA present as a ribonucleoprotein particle in the dendrites and somata of neurons. This ribonucleoprotein particle contains an identifier sequence-binding protein (Bp-1 protein). We report here the purification of BC-1 RNA and demonstrate that Bp-1 protein interacts directly with the RNA. We also demonstrate an accumulation of Bp-1 protein in the nucleus of brain cells from mouse fetus and newborns that precedes the postnatal increase in BC-1 RNA. Cytoplasmic Bp-1 protein present in a complex with BC-1 RNA increases postnatally with a concomitant decrease in nuclear Bp-1 protein. These observations suggest that Bp-1 protein may play a role(s) in the synthesis and nuclear export of BC-1 RNA.

Aging↗

PubMed2005

Attenuation of bunyavirus replication by rearrangement of viral coding and noncoding sequences.

Bunyamwera virus (BUN) is the prototype virus of the family Bunyaviridae. BUN has a tripartite negative-sense RNA genome comprising small (S), medium (M), and large (L) segments. Partially complementary untranslated regions (UTRs) flank the coding region of each segment. The terminal 11 nucleotides of these UTRs are conserved between the three segments, while the internal regions are unique. The UTRs direct replication and transcription of viral RNA and are sufficient to allow encapsidation of viral RNA into ribonucleoprotein complexes. To investigate the segment-specific functions of the UTRs, we have used reverse genetics to recover a recombinant virus (called BUN MLM) in which the L segment open reading frame (ORF) is flanked by the M segment UTRs. Compared to wild-type virus, BUN MLM virus shows growth attenuation in cultured mammalian cells and a slower disease progression in mice, produces small plaques, expresses reduced levels of L mRNA and L (RNA polymerase) protein, synthesizes less L genomic and antigenomic RNA, and has an increased particle-to-PFU ratio. Our data suggest that the packaging of BUN RNAs is not segment specific. In addition, the phenotype of BUN MLM virus supports the finding that BUN UTRs differ in their regulation of RNA synthesis but suggests that the interplay between each segment UTR and its cognate ORF may contribute to that regulation. Since BUN MLM virus is attenuated due to an essentially irreversible mutation, the rearrangement of UTRs is a feasible strategy for vaccine design for the more pathogenic members of the Bunyaviridae.

Animals↗

PubMed2024

Detection of mitochondrial tDRs in killifish embryos and other non-model organisms.

In recent years a diversity of small noncoding RNAs have been identified that originate from the mitochondrial genome. These mitosRNAs are often dominated by tRNA-derived small RNAs (mito-tDRs). Differential expression of mito-tDRs is associated with responses to stress. They also appear to be expressed differentially during development and their expression may be enriched in stress-tolerant animals. Very little is currently known about roles or modes of action of these sequences, although they are implicated in a diversity of processes such as cell cycle regulation, mRNA stability, regulation of ROS production, and import of proteins into the mitochondrion. To better understand the various roles these sequences may play, it is critical that we understand their diversity, cellular location, and the context for their expression. This protocol outlines the methodologies used to detect mitosRNAs, including mito-tDRs, in embryos and cells of the annual killifish Austrofundulus limnaeus. We highlight critical steps in the isolation of RNA, creation of sequencing libraries, bioinformatics processing of sequence data, and methods for validation of expression that support a robust discovery pipeline for mitosRNAs even from species with incomplete reference genome sequences.

Animals↗

PubMed1991

Nucleotide sequences important for translation initiation of enterovirus RNA.

An infectious cDNA clone was constructed from the genome of coxsackievirus B1 strain. A number of RNA transcripts that have mutations in the 5' noncoding region were synthesized in vitro from the modified cDNA clones and examined for their abilities to act as mRNAs in a cell-free translation system prepared from HeLa S3 cells. RNAs that lack nucleotide sequences at positions 568 to 726 and 565 to 726 were found to be less efficient and inactive mRNAs, respectively. To understand the biological significance of this region of RNA, small deletions and point mutations were introduced in the nucleotide sequence between positions 538 and 601. Except for a nucleotide substitution at 592 (U----C) within the 7-base conserved sequence, mutations introduced in the sequence downstream of position 568 did not affect much, if any, of the ability of RNA to act as mRNA. Except for a point mutation at 558 (C----U), mutations upstream of position 567 appeared to inactivate the mRNA. In the upstream region, a sequence consisting of 21 nucleotides at positions 546 to 566 is perfectly conserved in the 5' noncoding regions of enterovirus and rhinovirus genomes. These results suggest that the 7-base conserved sequence functions to maintain the efficiency of translation initiation and that the nucleotide sequence upstream of position 567, including the 21-base conserved sequence, plays essential roles in translation initiation. A deletion mutant whose genome lacks the nucleotide sequence at positions 568 to 726 showed a small-plaque phenotype and less virulence against suckling mice than the wild-type virus. Thus, reduction of the efficiency of translation initiation may result in the construction of enteroviruses with the lower-virulence phenotype.

Animals↗