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ABSTRACT: Single cell RNA sequencing of nc886, a non-coding RNA transcribed by RNA polymerase III, with a primer spike-in strategy

PROVIDER: PRJNA1091369 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR28431557_1.fastq.gz	Fastqsanger.gz
	SRR28431557_2.fastq.gz	Fastqsanger.gz
	SRR28431558_1.fastq.gz	Fastqsanger.gz
	SRR28431558_2.fastq.gz	Fastqsanger.gz

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Single cell RNA sequencing of nc886, a non-coding RNA transcribed by RNA polymerase III, with a primer spike-in strategy

Project description:Single cell RNA sequencing (scRNA-seq) has emerged as a versatile tool in biology, enabling comprehensive genomic-level characterization of individual cells. Currently, most scRNA-seq methods generate barcoded cDNAs by capturing polyA tails of mRNAs, which excludes many non-coding RNAs (ncRNAs), especially those transcribed by RNA polymerase III (Pol III). Although previously thought to be expressed constitutively, Pol III-transcribed ncRNAs are expressed variably in healthy and disease states and play important roles therein, necessitating their profiling at the single cell level. In this study, we have developed a measurement protocol for nc886 as a model case, as an initial step for scRNA-seq for Pol III-transcribed ncRNAs. Specifically, we spiked in an oligo-tagged nc886-specific primer during the polyA tail capture process for the 5’-reading in scRNA-seq. We then produced sequencing libraries for standard 5’ gene expression and oligo-tagged nc886 separately, to accommodate different cDNA sizes and ensure undisturbed transcriptome analysis. We applied this protocol in three cell lines which express high, low, and zero levels of nc886, respectively. Our results show that the identification of oligo tags exhibited limited target specificity, and sequencing reads of nc886 enabled the correction of non-specific priming. These findings suggest that gene-specific primers (GSPs) can be employed to capture RNAs lacking a polyA tail, with subsequent sequence verification ensuring accurate gene expression counting. Moreover, we embarked on an analysis of differentially expressed genes in cell line sub-clusters with differential nc886 expression, demonstrating variations in gene expression phenotypes. Collectively, the primer spike-in strategy allows us for a combined analysis of ncRNAs and gene expression phenotype.

2024-07-17 | GSE269878 | GEO

Single cell RNA sequencing of nc886, a non-coding RNA transcribed by RNA polymerase III, with a primer spike-in strategy

Project description:Single cell RNA sequencing of nc886, a non-coding RNA transcribed by RNA polymerase III, with a primer spike-in strategy

| PRJNA1124055 | ENA

Roles of AtR8 lncRNA in defense responses in Arabidopsis thaliana

Project description:AtR8 is a long non-coding RNA transcribed by RNA polymerase III in Arabidopsis which is responsive to hypoxic stress and salicylic acid. To understand the role of AtR8 lncRNA, microarray was carried out in the mutant seedlings of AtR8 and compared with wild type seedlings.

2018-07-04 | GSE100800 | GEO

nc886, a noncoding RNA of anti-proliferative role, is suppressed by CpG DNA methylation in human gastric cancer

Project description:nc886 is a 101 nucleotides long non-coding RNA that is also known as a precursor microRNA or a vault RNA. nc886 has been suggested to be a tumor suppressor, mainly inferred by its expression pattern as well as its genomic location at human chromosome 5q31, a locus for a tumor suppressor gene(s). In order to understand potential roles of nc886 in gastric cancer, we performed the knockdown of nc886 in nonmalignant HFE-145 cells and identified and analyzed genes whose expression is activated by nc886 knockdown in gastric cancer.

2015-09-30 | GSE51067 | GEO

Analysis of the interactome of B2-SINE RNA transcripts

Project description:B2-SINEs are noncoding RNAs which are transcribed by RNA polymerase III (Pol III) from short interspersed nuclear elements (SINEs), which are high copy number transposable elements in the mouse genome. Unexpectedly have observed significant induction of non-coding RNAs from the B2-SINE subclass of repeat element RNAs in DRG neurons following sciatic nerve injury. We next sought to identify intracellular targets of GI-SINEs, by protein pull-down from sciatic nerve axoplasm using biotinylated B2-SINE RNA as bait. We generated two constructs from the B2 consensus sequence, comprising 5’ (1-75nt) and 3’ (75-166nt) sequences, both predicted to retain their respective structures, and used the constructs to pull down axoplasm proteins for mass spectrometry analysis. Further confirmatory experiments were performed using the mentioned -175nt sequence vs a control U1 SL3+4 RNA

2025-05-05 | PXD060976 | Pride

Pol II and its associated epigenetic marks are present at pol III-transcribed non-coding RNA genes (I)

Project description:Epigenetic control is an important aspect of gene regulation. Despite detailed understanding of many examples, the transcription of non-coding RNA genes by RNA polymerase (pol) III is less well characterized. Here we profile the epigenetic features of pol III target genes throughout the human genome. This reveals that the chromatin landscape of pol III-transcribed genes resembles that of pol II templates in many ways, although there are also clear differences. Our analysis also discovered an entirely unexpected phenomenon, namely that pol II co-localizes with the majority of genomic loci that are bound by pol III.

2010-04-25 | GSE20040 | GEO

Pol II and its associated epigenetic marks are present at pol III-transcribed non-coding RNA genes (II)

2010-04-25 | GSE20071 | GEO

Epigenetic silencing of noncoding RNA nc886 provokes oncogenes during esophageal tumorigenesis

2013-10-31 | GSE51732 | GEO

Epigenetic silencing of noncoding RNA nc886 provokes oncogenes during esophageal tumorigenesis

Project description:nc886 is a 101 nucleotides long non-coding RNA that is also known as a precursor microRNA or a vault RNA. nc886 has been suggested to be a tumor suppressor, mainly inferred by its expression pattern as well as its genomic location at human chromosome 5q31, a locus for a tumor suppressor gene(s). Pre-clinical study In order to understand potential roles of nc886 in esophageal squamous cell carcinoma (ESCC), we performed the knockdown of nc886 in nonmalignant HET-1A cells and ESCC cell lines (TE-1 and TE-8) and identified and analyzed genes whose expression is activated by nc886 knockdown in ESCC.

2013-10-31 | E-GEOD-51732 | biostudies-arrayexpress

Transcriptome-scale analysis for RNAs of medium length, designated as melRNA-seq [RppH-treated]

Project description:Small interspersed elements (SINEs) is transcribed by RNA polymerase III (Pol III) to produce RNAs of typically 100 to 500 nucleotides in length. Although the abundance of SINE RNAs can be analyzed by Northern blotting and primer extension, the nature (sequence, exact length, and genomic origin) of these RNAs cannot be revealed by these methods. Moreover, mRNA sequencing (mRNA-seq) is not able to distinguish bona fide SINE RNAs and SINE sequences present in longer transcripts. To elucidate the abundance, source loci, and sequence nature of SINE RNAs, we have established a deep sequencing method, designated as melRNA-seq (medium length RNA-seq), which can determine whole-length sequences of RNAs. The total RNA samples were treated with 5' Pyrophosphohydrolase (RppH), which allowed ligation of an RNA adaptor to the 5’ end of intact SINE RNAs. Another adaptor was ligated to the 3’ end, followed by reverse transcription, PCR amplification, size selection, and single-end deep sequencing. Analysis of two biological replicates of RNAs from mouse spermatogonia showed high reproducibility of the SINE expression data both at family level and locus level. Therefore, this new method can be used for the quantification and detailed sequence analysis of medium length non-coding RNAs, such as rRNA, snRNA, tRNAs, and SINE RNAs. The dynamic range is much wider than Northern blotting and primer extension.

2021-07-01 | GSE171593 | GEO

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