Browse
Submit Data
Databases
API
Help

Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

15 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Sequence analysis of scnRNAs

ABSTRACT: The developmentally regulated 26- to 32-nt siRNAs (scnRNAs) are loaded to the Argonaute protein Twi1p and display a strong bias for uracil at the 5' end. In this study, we used deep sequencing to analyze loaded and unloaded populations of scnRNAs. We show that the size of the scnRNA is determined during a pre-loading process, whereas their 5' uracil bias is attributed to both pre-loading and loading processes. We also demonstrate that scnRNAs have a strong bias for adenine at the third base from the 3' terminus, suggesting that most scnRNAs are direct Dicer products. Furthermore, we show that the thermodynamic asymmetry of the scnRNA duplex does not affect the guide and passenger strand decision. Finally, we show that scnRNAs frequently have templated uracil at the last base without a strong bias for adenine at the second base indicating non-sequential production of scnRNAs from substrates. These findings provide a biochemical basis for the varying attributes of scnRNAs, which should help improve our understanding of the production and turnover of scnRNAs in vivo. We compared Twi1p-loaded scnRNAs to scnRNAs before they have been loaded into Twi1p by deep sequencing to understand how the two processes, the production of siRNAs by Dicer and the loading of siRNAs into Argonaute, shape the population of siRNAs in vivo.

ORGANISM(S): Tetrahymena thermophila

SUBMITTER: Kazufumi Mochizuki

PROVIDER: E-GEOD-47650 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

ACCESS DATA

Json Xml

Similar Datasets

Analysis of conjugation-specific 26-32nt siRNAs of Tetrahymena thermophila

Project description:Distinct classes of small RNAs are often selectively sorted to different Argonaute proteins. Various properties of small RNAs, such as length, terminal nucleotide, thermodynamic asymmetry and duplex mismatches, can impact sorting in different RNA silencing pathways in diverse eukaryotes. The developmentally regulated ~26-32 nt siRNAs, which are involved in programmed DNA elimination in Tetrahymena, show a strong bias for uracil at the 5' end. In this study, we analyzed loaded and unloaded populations of ~26-32 nt siRNAs by deep RNA sequencing. We show that the production process is the main determinant of size, whereas the 5' uracil bias is attributed not only to the process of loading siRNAs into the Argonaute protein Twi1p but also significantly to the initial processing of the siRNAs. We also show that both the loaded and the unloaded ~26-32 nt siRNAs have a strong bias for adenine as the 3rd base from the 3' terminus, suggesting that most of these siRNAs are direct Dicer products and little post-processing amplification of this class of siRNAs occurs. Further, we demonstrate that the siRNA-loading process in vivo can be deduced from the fraction of siRNAs with uracil as the first base. These findings provide biochemical bases for the attributes of ~26-32 nt siRNAs, which should help improve our understanding of their production and turnover in vivo. Examination of siRNA populations in wild-type and TWI1 KO Tetrahymena cells

2012-10-01 | E-GEOD-37696 | biostudies-arrayexpress

Sequence analysis of scnRNAs

Project description:The developmentally regulated 26- to 32-nt siRNAs (scnRNAs) are loaded to the Argonaute protein Twi1p and display a strong bias for uracil at the 5' end. In this study, we used deep sequencing to analyze loaded and unloaded populations of scnRNAs. We show that the size of the scnRNA is determined during a pre-loading process, whereas their 5' uracil bias is attributed to both pre-loading and loading processes. We also demonstrate that scnRNAs have a strong bias for adenine at the third base from the 3' terminus, suggesting that most scnRNAs are direct Dicer products. Furthermore, we show that the thermodynamic asymmetry of the scnRNA duplex does not affect the guide and passenger strand decision. Finally, we show that scnRNAs frequently have templated uracil at the last base without a strong bias for adenine at the second base indicating non-sequential production of scnRNAs from substrates. These findings provide a biochemical basis for the varying attributes of scnRNAs, which should help improve our understanding of the production and turnover of scnRNAs in vivo.

2013-06-05 | GSE47650 | GEO

Efficient and quantitative high-throughput transfer RNA sequencing

Project description:Despite its biological importance, transfer RNA (tRNA) could not be adequately sequenced due to the abundant presence of post-transcriptional modifications and extensive structure that interfere with cDNA synthesis and adapter ligation. We achieve efficient and quantitative tRNA sequencing by removing base methylations using engineered demethylases and using a highly processive thermo-stable reverse transcriptase without the need for adapter ligation (DMTRT-tRNA-seq). Our method should be applicable for biological investigations of tRNA in all organisms. Development of tRNA-Seq method

2015-07-01 | E-GEOD-66550 | biostudies-arrayexpress

Ultra-high throughput sequencing-based small RNA discovery and discrete statistical biomarker analysis in a collection of cervical tumors and matched controls

Project description:We prepared small RNA libraries from 29 tumor/normal pairs of human cervical tissue samples. Analysis of the resulting sequences (42 million in total) defined 64 new human microRNA (miRNA) genes. Both arms of the hairpin precursor were observed in twenty-three of the newly identified miRNA candidates. We tested several computational approaches for analysis of class differences between high throughput sequencing datasets, and describe a novel application of log linear model that has provided the most datasets, and describe a novel application of log linear model that has provided the most effective analysis for this data. This method resulted in the identification of 67 miRNAs that were differentially-expressed between the tumor and normal samples at a false discovery rate less than 0.001. A total of 29 tumor/normal pairs of human cervical tissue samples were analyzed. Two samples (G699N_2 and G761T_2) were performed in duplicates. No Fastq files for GSM532871 to GSM532889, GSM532929, and GSM532930. Sequence files are provided as text files for these 22 Sample records in GSE20592_RAW.tar. 38 samples with quality scores are available from SRA as SRP002/SRP002326 (see Supplementary file below).

2010-04-28 | E-GEOD-20592 | biostudies-arrayexpress

Biased transcription and selective degradation of small RNAs shape the pattern of DNA elimination in Tetrahymena

Project description:The ciliated protozoan Tetrahymena undergoes extensive programmed DNA elimination when the germline micronucleus produces the new macronucleus during sexual reproduction. DNA elimination is epigenetically controlled by DNA sequences of the parental macronuclear genome, and this epigenetic regulation is mediated by small RNAs (scnRNAs) of approximately 28-30 nucleotides that are produced and function by an RNAi-related mechanism. Here, we examine scnRNA production and turnover by deep sequencing. scnRNAs are produced exclusively from the micronucleus and non-homogeneously from a variety of chromosomal locations. scnRNAs are preferentially derived from the eliminated sequences, and this preference is mainly determined at the level of transcription. Despite this bias, a significant fraction of scnRNAs is also derived from the macronuclear-destined sequences, and these scnRNAs are degraded during the course of sexual reproduction. These results indicate that the pattern of DNA elimination in the new macronucleus is shaped by the biased transcription in the micronucleus and by the selective degradation of scnRNAs in the parental macronucleus. GRO-Seq and Examination of siRNAs in wild-type,nullisomic 4, EMA1 KO, and TWI1 KO Tetrahymena cells

2012-06-21 | E-GEOD-38768 | biostudies-arrayexpress

Argonaute and Trimmer generate Dicer-independent priRNAs and nucleate RNAi

Project description:RNA interference is involved in silencing of transposable and repetitive elements. How these elements are initially recognized by RNAi is a fundamental and unanswered question. We previously identified a class of Dicer-independent small RNAs, called primal small RNAs (priRNAs), in fission yeast. The mechanism by which Dicer-independent small RNAs are generated is not clear for any species. Here we reconstitute priRNA biogenesis in vitro and demonstrate that priRNAs can nucleate RNAi in vivo. We identify 3'-5' exonuclease Trimmer and show that Argonaute, loaded with longer RNA precursors, recruits Trimmer to generate the new 3' end. Next, we show that antisense priRNAs accumulate in rrp6M-NM-^T cells and nucleate RNAi at subset of protein coding genes in a Trimmer- and priRNA-dependent manner. Thus, Rrp6-mediated degradation of antisense transcripts and priRNA precursors protects the genome from spurious RNAi. Our results suggest that Argonaute association with random RNA degradation products triggers RNAi in a process of transcriptome surveillance. small RNA profiling in wild type S. pombe cells and in mutant cells

2014-04-21 | E-GEOD-38636 | biostudies-arrayexpress

Analyses of Early- and Late-scnRNAs of Tetrahymena

Project description:In this study, we demonstrated that there is a novel, unanticipated mechanism regulating programmed DNA elimination: a genome-wide trans-recognition network for IES identification. In this mechanism, Early-scnRNAs produced from Type-A IESs in the MIC identify not only the IESs from which they are derived but also other IESs in trans to trigger the cis-spreading of Late-scnRNA production in the IESs. This cis-spreading of Late-scnRNA production requires heterochromatin formation . Furthermore, these Late-scnRNAs can recognize other IESs in trans. This “chain reaction” of Late-scnRNA production by the trans-recognition network most likely provides strong robustness in DNA elimination by buffering cell-to-cell variability in the initial Early-scnRNA populations. 26 to 32-nt small RNAs from various mutants or from immuno precipitated with Argonaute proteins were analyzed by high-throughput sequencing

2015-06-15 | E-GEOD-68455 | biostudies-arrayexpress

In vitro evolution of Pseudomonas aeruginosa AA2 biofilms in the presence of cystic fibrosis lung microbiome members

Project description:P. aeruginosa was cultured in a MultiScreen-Mesh plate, which has a filter at the bottom of the wells. The plate was immersed in either in medium alone (control) or in medium inoculated with a mixture of five bacterial strains commonly found in cystic fibrosis sputum (\"microbiome\"). The filter prevented physical contact between P. aeruginosa and the other bacteria, yet soluble products could migrate through the filter into the P. aeruginosa biofilm. P. aeruginosa was then allowed to form biofilms in the wells for 72h, then the biofilm was harvested and a fraction of the harvested cells were used for re-inoculations. This was repeated for 18 cycles for a total of 54 days.

2019-09-13 | E-MTAB-7331 | biostudies-arrayexpress

PatchSeq experiment on somatostatin-expressing (Sst) neurons from mouse ventral tegmental area (VTA). Full-length single cell sequencing.

Project description:Experiment was done as a part of multi-methodological description of Sst-neurons subtypes in mouse VTA, aiming to reveal their transcriptional profiles and neurotransmitter nature.

2020-06-02 | E-MTAB-8780 | biostudies-arrayexpress

Tetrahymena thermophila

Project description:Twi1p-loaded scnRNAs in GIW1 KO Tetrahymena cells

| PRJNA261680 | ENA