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Comprehensive map of ribosomal 2′-O-methylation and C/D box snoRNAs in Drosophila melanogaster [Fib_KD_TGIRT-seq]

ABSTRACT: The majority of Nm modifications in eukaryotes are placed by Fibrillarin, a conserved methyltransferase belonging to a ribonucleoprotein complex guided by C/D box small nucleolar RNAs (C/D box snoRNAs). We built the first comprehensive map of Nm sites in Drosophila melanogaster rRNAs using two complementary approaches (RiboMethSeq and Nanopore direct RNA sequencing) and identified their corresponding C/D box snoRNAs by whole-transcriptome sequencing (TGIRT-seq).

ORGANISM(S): Drosophila melanogaster

PROVIDER: GSE233853 | GEO | 2024/02/12

REPOSITORIES: GEO

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Comprehensive map of ribosomal 2′-O-methylation and C/D box snoRNAs in Drosophila melanogaster [Fib_KD_RiboMeth-seq]

Project description:The majority of Nm modifications in eukaryotes are placed by Fibrillarin, a conserved methyltransferase belonging to a ribonucleoprotein complex guided by C/D box small nucleolar RNAs (C/D box snoRNAs). We built the first comprehensive map of Nm sites in Drosophila melanogaster rRNAs using two complementary approaches (RiboMethSeq and Nanopore direct RNA sequencing) and identified their corresponding C/D box snoRNAs by whole-transcriptome sequencing (TGIRT-seq).

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Comprehensive map of ribosomal 2′-O-methylation and C/D box snoRNAs in Drosophila melanogaster [Tissues_TGIRT-seq]

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Next generation sequencing identifies discrete classes of box C/D snoRNAs featuring different ends and RNA binding protein dependency

Project description:The paired-end next-generation sequencing of all small RNAs of less than 200 nucleotides in length from four different human cell lines (SKOV3ip1, MCF-7, BJ-Tielf, INOF) allowed us to determine the exact sequence(s) and variations of human box C/D snoRNAs (small nucleolar RNAs), revealing processing patterns of this class of molecules. Two distinct groups of box C/D snoRNAs were identified based on the position of their ends with respect to their characteristic boxes and the terminal base pairing potential. Short box C/D snoRNAs start sharply 4 or 5 nucleotides upstream of their box C and end 2 or 3 nucleotides downstream of their box D. In contrast, long box C/D snoRNAs start 5 or 6 nucleotides upstream of their box C and end 4 or 5 nucleotides downstream of their box D, increasing the likelihood of formation of a k-turn between their boxes C and D. Sequencing of SKOV3ip1 cells following the depletions of NOP58, a core box C/D snoRNA-binding protein and of RBFOX2, a splicing factor, shows that the short box C/D snoRNA forms are significantly more affected by the depletion of RBFOX2 while the long snoRNA forms, which display more canonical box C/D snoRNA features, are significantly more affected by the depletion of NOP58. Together the data suggest that box C/D snoRNAs are divided into at least two groups of RNA with distinct maturation and functional preferences. Small RNAs (<200 nucleotides) were isolated from different human cell lines that were either untreated or depleted of NOP58 or RBFOX2 using specific siRNAs. The resulting libraries were multiplexed and paired-end sequenced using Illumina HiSeq.

2014-07-01 | E-GEOD-55946 | biostudies-arrayexpress

Orphan box C/D snoRNAs guide rRNA base acetylations in yeast

Project description:Box C/D snoRNAs snR4 and snR45, whose role was unknown, guide 18S rRNA acetylations by Kre33 in yeast. UV-crosslinking (CRAC) was used to determine binding sites of Kre33 on RNA.

2017-07-25 | GSE87480 | GEO

Next generation sequencing identifies discrete classes of box C/D snoRNAs featuring different ends and RNA binding protein dependency

2014-07-01 | GSE55946 | GEO

High-grade Ovarian Cancer Associated H/ACA snoRNAs Promote Cancer Cell Proliferation and Survival

Project description:Small nucleolar RNAs (snoRNAs) are an omnipresent class of non-coding RNAs involved in the modification and processing of ribosomal RNA (rRNA). There are two types of snoRNAs, box C/D, which typically guide 2’-O-methylation, and box H/ACA, which guide pseudouridylation. As snoRNAs are required for ribosome production, the increase of which is a hallmark of cancer development, their expression would be expected to increase in proliferating cancer cells. However, the contributions of H/ACA box and C/D box snoRNAs to the biology of cancer cells remain unclear. To understand the contribution of snoRNAs to cancer progression and aggressiveness we examined their abundance patterns in high-grade serous ovarian carcinomas (HGSC) and serous borderline tumors (SBT), using a newly developed snoRNA sensitive sequencing technique. Surprisingly, the results indicate that while abundance of a selected group of H/ACA snoRNAs increases, most C/D snoRNAs remain stable or decrease in HGSC compared to SBT. This suggests that induction of ribosome biogenesis and translation in HGSC does not require a general increase in snoRNAs expression but depends upon the upregulation of a handful of key H/ACA snoRNAs. This subgroup accurately discriminates between SBT and HGSC underlining their potential as biomarkers of tumor aggressiveness. Remarkably, knockdown of HGSC associated H/ACA snoRNAs, but not their host genes, inhibits cell proliferation and induces apoptosis of model ovarian cancer cell lines. Together our data indicate that specific H/ACA snoRNAs promote tumor aggressiveness through the induction of cell proliferation and resistance to apoptosis.

2021-12-08 | GSE181496 | GEO

Cytosolic Accumulation of Small Nucleolar RNAs (snoRNAs) is Dynamically Regulated by NADPH Oxidase

Project description:Small nucleolar RNAs (snoRNAs) guide nucleotide modifications of cellular RNAs in the nucleus. We have previously shown that box C/D snoRNAs from the Rpl13a locus are unexpected mediators of physiologic oxidative stress, independent of their predicted ribosomal RNA modifications. Here, we demonstrate that oxidative stress induced by doxorubicin causes rapid cytoplasmic accumulation of the Rpl13a snoRNAs through a mechanism that requires superoxide and a nuclear splice variant of NADPH oxidase. RNA-sequencing analysis reveals that box C/D snoRNAs as a class are present in the cytoplasm, where their levels are dynamically regulated by NADPH oxidase. These findings suggest that snoRNAs may orchestrate the response to environmental stress through molecular interactions outside of the nucleus.

2015-03-20 | GSE67050 | GEO

Transcriptomic analysis of the abundance of snoRNAs and their host genes in 4 different cell lines, using low structure bias TGIRT-seq

Project description:Small nucleolar RNAs (snoRNAs) are highly abundant non-coding RNAs whose canonical function is to guide the 2’-O-ribose methylation or pseudouridylation of ribosomal RNAs. They are often encoded in longer genes referred to as their host genes, the expression of which is required for their own biogenesis. Due to their very stable structure, reverse transcription of snoRNAs using standard viral reverse transcriptases is very inefficient, and does not provide an adequate view of the abundance of these small RNAs. We thus employed high-throughput, low structure bias TGIRT-seq sequencing to quantified the abundance of the entire ribodepleted transcriptome, including snoRNAs and their host genes, in 4 different cell lines.

2022-08-10 | GSE209924 | GEO

Insights into snoRNA biogenesis and processing from PAR-CLIP of snoRNA core proteins and small RNA sequencing

Project description:Background: In recent years, a variety of small RNAs derived from other RNAs with well-known functions such as tRNAs and snoRNAs, have been identified. The functional relevance of these RNAs is largely unknown. To gain insight into the complexity of snoRNA processing and the functional relevance of snoRNA-derived small RNAs, we sequenced long and short RNAs, small RNAs that co-precipitate with the Argonaute 2 protein and RNA fragments obtained in photoreactive nucleotide-enhanced crosslinking and immunoprecipitation (PAR-CLIP) of core snoRNA-associated proteins. Results: Analysis of these data sets revealed that many loci in the human genome reproducibly give rise to C/D box-like snoRNAs, whose expression and evolutionary conservation are typically less pronounced relative to the snoRNAs that are currently catalogued. We further found that virtually all C/D box snoRNAs are specifically processed inside the regions of terminal complementarity, retaining in the mature form only 4-5 nucleotides upstream of the C box and 2-5 nucleotides downstream of the D box. Sequencing of the total and Argonaute 2-associated populations of small RNAs revealed that despite their cellular abundance, C/D box-derived small RNAs are not efficiently incorporated into the Ago2 protein. Conclusions: We conclude that the human genome encodes a large number of snoRNAs that are processed along the canonical pathway and expressed at relatively low levels. Generation of snoRNA-derived processing products with alternative, particularly miRNA-like, functions appears to be uncommon. PAR-CLIP profiling for snoRNP core proteins NOP56, NOP58, Fibrillarin, and Dyskerin in HEK293 cells. Small RNA profiling using RNA-seq in HEK293 and HeLa cells, small RNA profiling using IP-seq of Ago2 associated small RNAs.

2013-05-08 | E-GEOD-43666 | biostudies-arrayexpress

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Project description:We generated a SNORD71 KO chondrocyte cell pool using CRISPR/Cas9 gene editing. A CRISPR control cell line was generated and used as a control. Levels of 2’-O-methylation of human rRNAs in SNORD71 KO cell pool and CRISPR control cells were evaluated by RiboMethSeq.

2022-03-05 | E-MTAB-11470 | biostudies-arrayexpress

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