Browse
Submit Data
Databases
API
Help

Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

A Tetrahymena Piwi protein and the exonuclease Xrn2 form a complex that is essential for ribosome biogenesis

ABSTRACT: Emerging evidence suggests that Ago/Piwi proteins function in the nucleus as well as the cytoplasm to control gene expression, but the mechanisms they employ in the nucleus remain poorly defined. The Tetrahymena thermophila Ago/Piwi protein Twi12 is essential for growth and functions in the nucleus. We show that Twi12 interacts with the exonuclease Xrn2. Twi12 functions to stabilize and localize Xrn2 in vivo, as well as activate its exonuclease activity in vitro. When Twi12 or Xrn2 are depleted, pre-rRNA processing intermediates accumulate and mature rRNA levels decline. RNA polymerase I and II (RNAP I and II) transcripts also accumulate. Twi12 function depends on small RNA (sRNA) binding, which is required for its nuclear import. Twi12-bound sRNAs are Dicer-independent 3' tRNA fragments that we propose may not be involved in sequence-specific base pairing to targets. Our findings suggest a role for tRNA fragments and an Ago/Piwi protein in global control of gene expression through interaction with a conserved exonuclease. One library is analyzed here: small RNAs associated with ZZF-tagged Twi12. Twi12 is the full-length version, which is extended by 17 amino acids at the N-terminus compared to the previously-reported Twi12. Specifically, the 16-22 nt reads were analyzed here.

ORGANISM(S): Tetrahymena thermophila

SUBMITTER: Mary Couvillion

PROVIDER: E-GEOD-38507 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

ACCESS DATA

Json Xml

Similar Datasets

A Tetrahymena Piwi Bound to Mature tRNA 3' Fragments Activates the Exonuclease Xrn2 for RNA Processing in the Nucleus

Project description:Emerging evidence suggests that Ago/Piwi proteins function in the nucleus as well as the cytoplasm to control gene expression, but the mechanisms they employ in the nucleus remain poorly defined. The Tetrahymena thermophila Ago/Piwi protein Twi12 is essential for growth and functions in the nucleus. We show that Twi12 interacts with the exonuclease Xrn2. Twi12 functions to stabilize and localize Xrn2 in vivo, as well as activate its exonuclease activity in vitro. When Twi12 or Xrn2 are depleted, pre-rRNA processing intermediates accumulate and mature rRNA levels decline. RNA polymerase I and II (RNAP I and II) transcripts also accumulate. Twi12 function depends on small RNA (sRNA) binding, which is required for its nuclear import. Twi12-bound sRNAs are Dicer-independent 3' tRNA fragments that we propose may not be involved in sequence-specific base pairing to targets. Our findings suggest a role for tRNA fragments and an Ago/Piwi protein in global control of gene expression through interaction with a conserved exonuclease.

2012-08-31 | GSE38507 | GEO

Effects of transcription elongation rate and Xrn2 exonuclease activity on RNA polymerase II termination suggest widespread kinetic competition

Project description:Transcription termination was analyzed by anti RNA pol II ChIP-seq in isogenic human HEK293 cell lines that inducibly express a-amanitin resistant mutants of the RNA polymerase II large subunit with slow and fast elongation rates and in lines that inducbily over-express WT or an active site mutant of the RNA exonuclease "torpedo" Xrn2. Transcription termination zones were mapped by anti-pol II ChIP-seq under conditions where transcription elongation rate was increased or decreased by point mutations in the large subunit of the enzyme. Termination was also assayed under conditions where Xrn2 exonuclease activity was inhibited by over-expression of an active site mutant (D235A).

2015-11-19 | E-GEOD-72800 | biostudies-arrayexpress

mRNA decapping factors and the exonuclease Xrn2 function in widespread premature termination of RNA polymerase II transcription

Project description:We report a function of human mRNA decapping factors in control of transcription by RNA polymerase II. Decapping proteins Edc3, Dcp1a and Dcp2 and the termination factor TTF2 co-immunoprecipitate with Xrn2, the nuclear 5'-3' exonuclease torpedo that facilitates transcription termination at the 3' ends of genes. Dcp1a, Xrn2 and TTF2 localize near transcription start sites (TSSs) by ChIP-Seq. At genes with 5' peaks of paused pol II, knockdown of decapping or termination factors, Xrn2 and TTF2, shifted polymerase away from the TSS toward upstream and downstream distal positions. This re-distribution of pol II is similar in magnitude to that caused by depletion of the elongation factor Spt5. We propose that coupled decapping of nascent transcripts and premature termination by the torpedo mechanism is a widespread mechanism that limits bidirectional pol II elongation. Regulated co-transcriptional decapping near promoter-proximal pause sites followed by premature termination could control productive pol II elongation. RNA pol II (GSE30895: GSM766171), Xrn2, TTF2 and Dcp1a were localized by ChIP-Seq in HeLa cells. RNA pol II was localized in control HEK293 cells and cells infected with lentiviruses expressing a scrambled control shRNA (scr), and shRNAs targeting the following proteins: Xrn2, TTF2, Xrn2+TTF2, Edc3, Dcp1a, and Dcp2.

2012-04-04 | E-GEOD-36185 | biostudies-arrayexpress

An Ancient Transcription Factor An Ancient Transcription Factor During Early Meiosis in Mouse Testes (PAS-Seq and CAGE)

Project description:Animal germ cells produce PIWI-interacting RNAs (piRNAs), small silencing RNAs that suppress transposons and enable gamete maturation. Mammalian transposon-silencing piRNAs accumulate early in spermatogenesis, whereas pachytene piRNAs are produced later during post-natal spermatogenesis and account for >95% of all piRNAs in the adult mouse testis. Mutants defective for pachytene piRNA pathway proteins fail to produce mature sperm, but neither the piRNA precursor transcripts nor the trigger for pachytene piRNA production is known. Here, we show that the transcription factor A-MYB initiates pachytene piRNA production. A-MYB drives transcription of both pachytene piRNA precursor RNAs and the mRNAs for core piRNA biogenesis factors, including MIWI, the protein through which pachytene piRNAs function. A-MYB regulation of piRNA pathway proteins and piRNA genes creates a coherent feed-forward loop that ensures the robust accumulation of pachytene piRNAs. This regulatory circuit, which can be detected in rooster testes, likely predates the divergence of birds and mammals. PAS-Seq and CAGE in mouse testes

2013-03-27 | E-GEOD-45048 | biostudies-arrayexpress

Mouse MOV10L1 associates with Piwi proteins and is an essential component of the piRNA pathway

Project description:Piwi-interacting RNAs (piRNAs) are essential for silencing of transposable elements in the germline but their biogenesis is poorly understood. Here we demonstrate that MOV10L1, a germ cell-specific putative RNA helicase, is associated with Piwi proteins. Genetic disruption of the MOV10L1 RNA helicase domain in mice renders both MILI and MIWI2 devoid of piRNAs. Absence of a functional piRNA pathway in Mov10l1 mutant testes causes loss of DNA methylation and subsequent de-repression of retrotransposons in germ cells. The Mov10l1 mutant males are sterile due to complete meiotic arrest. This is the first mouse mutant that expresses Piwi proteins but lacks piRNAs, suggesting that MOV10L1 is required for piRNA biogenesis and/or loading to Piwi proteins. Examination of piRNAs in Mov10l IP and differences in total small RNA profiles in RNA helicase Mov10l+/- and Mov10l-/- mutants.

2010-06-12 | E-GEOD-21763 | biostudies-arrayexpress

Xrn2 substrate mapping identifies torpedo loading sites and extensive premature termination of RNA pol II transcription

Project description:The exonuclease torpedo Xrn2 loads onto nascent RNA 5’-PO4 ends and chases down pol II to promote termination downstream of polyA sites. We report that Xrn2 is recruited to pre-initiation complexes and “travels” to 3’ ends of genes. Mapping of 5’-PO4 ends in nascent RNA identified Xrn2 loading sites stabilized by an active site mutant, Xrn2(D235A). Xrn2 loading sites are ~2-20 bases downstream of where CPSF73 cleaves at polyA sites and histone 3’ ends. We propose that processing of all mRNA 3’ ends comprises cleavage and limited 5’-3’ trimming by CPSF73 followed by hand-off to Xrn2. A similar hand-off occurs at tRNA 3’ ends where co-transcriptional RNAseZ cleavage generates novel Xrn2 substrates. Exonuclease-dead Xrn2 increased transcription in 3’ flanking regions by inhibiting polyA site-dependent termination. Surprisingly, the mutant Xrn2 also rescued transcription in promoter-proximal regions to the same extent as in 3’ flanking regions. eNET-seq revealed Xrn2-mediated degradation of sense and anti-sense nascent RNA within a few bases of the TSS where 5’-PO4 ends may be generated by decapping or endonucleolytic cleavage. These results suggest that a major fraction of pol II complexes terminate prematurely close to the start site under normal conditions by an Xrn2-mediated torpedo mechanism.

2022-11-17 | GSE217033 | GEO

Deep Biosphere - Beatrix 326 BH2 (270712)

Project description:This project aims to investigate the metabolic pathways expressed by the active microbial community occurring at the deep continental subsurface. Subsurface chemoLithoautotrophic Microbial Ecosystems (SLiMEs) under oligotrophic conditions are supported by H2; however, the overall ecological trophic structures of these communities are poorly understood. Some deep, fluid-filled fractures in the Witwatersrand Basin, South Africa appear to support inverted trophic pyramids wherein methanogens contributing <5% of the total DNA apparently produce CH4 that supports the rest of the community. Here we show the active metabolic relationships of one such trophic structure by combining metatranscriptomic assemblies, metaproteomic and stable isotopic data, and thermodynamic modeling. Four autotrophic β-proteobacteria genera that are capable of oxidizing sulfur by denitrification dominate. They co-occur with sulfate reducers, anaerobic methane oxidizers and methanogens, which each comprises <5% of the total community. Defining trophic levels of microbial chemolithoautotrophs by the number of transfers from the initial abiotic H2-driven CO2 fixation, we propose a top-down cascade influence of the metabolic consumers that enhances the fitness of the metabolic producers to explain the inverted biomass pyramid of a multitrophic SLiME. Symbiotic partnerships are pivotal in the deep biosphere on and potentially beyond the Earth.

2018-10-27 | PXD004634 | Pride

High-throughput sequencing of AGO-immunoprecipitating miRs in human senescent fibroblast WI-38

Project description:In humans, there are four Ago proteins (Ago1M-bM-^@M-^S4) and AGO1- and 2 were previously implicated in TGS induced by exogenous siRNAs and microRNAs (miRs) directed against gene promoter transcripts via promotion of changes in histone covalent modifications and DNA methylation. Not-with-standing, many mechanistic details of this process remain poorly defined in human cells, and very little is known about the identity of possible endogenous signals, which may drive this process in human cells. Given the evolutionary conserved role of siRNAs and AGO proteins in TGS and heterochromatin formation, we set out to analyse their possible involvement in senesence-associated repression of E2F target genes. To obtain a detailed picture of AGO-immunoprecipitating miRs (RIP) in senescent cells, we used next-generation sequencing (NGS)(RIP-Seq). We also included histone H3 dimethylated on lysine 9 (H3K9me2) in this analysis to assign potential AGO2-interacting miRs to a repressive chromatin state and unfractionated, cellular RNA from senescent cells for normalisation. Determination of AGO AGO-immunoprecipitating miRs in WI-38 senescent human fibroblast

2011-12-17 | E-GEOD-34494 | biostudies-arrayexpress

Next generation sequencing of small RNAs isolated from exosomes in human semen

Project description:Preparation of exosomes isolated from semen contain a substantial amount of RNA, mostly from 20 to 100 nucleotides in length. We sequenced separately 20-40 and 40-100 nucleotide fractions of RNA from exosomes isolated from semenal fluid from six healthy donors. We found various classes of small non-coding RNA, including mature microRNA and piwi-RNA, as well as abundant Y RNAs and tRNAs present in both full length and fragmented forms. Specific RNAs were consistently present in all donors. For example, fifteen (of ~2,600 known) microRNAs constituted over 80% of mature microRNA in SE. Additionally, tRNA fragments were strongly enriched for 5M-bM-^@M-^Y-ends of 18-19 or 30-34 nucleotides in length. Size-fractionated small RNA profiles from exosomes isolated from the seminal fluid of six healthy donors

2014-06-04 | E-GEOD-56076 | biostudies-arrayexpress

Genome?wide analysis of pre?mRNA 3' end processing reveals a decisive role of human cleavage factor I in the regulation of 3' UTR length: A-seq

Project description:Through alternative polyadenylation, human mRNAs acquire longer or shorter 3' untranslated regions, the latter typically associated with higher transcript stability and increased protein production. To understand the dynamics of polyadenylation site usage, we mapped transcriptome?wide both binding sites of 3' end processing factors CPSF?160, CPSF?100, CPSF?73, CPSF?30, Fip1, CstF?64, CstF-64tau, CF Im25, CF Im59, and CF Im68 and 3' end processing sites in HEK293 cells. We found that although binding sites of these factors generally cluster around the poly(A) sites most frequently used in cleavage, CstF?64/CstF-64tau and CF Im proteins have much higher positional specificity compared to CPSF components. Knockdown of CF Im68 induced a systematic use of proximal polyadenylation sites, indicating that changes in relative abundance of a single 3' end processing factor can modulate the length of 3' untranslated regions transcriptome-wide, and suggesting a mechanism behind the previously observed increase in tumor cell invasiveness upon CF Im68 knockdown. 3' ends of transcripts were profiled by high-throughput sequencing in HEK 293 cells under normal conditions, and in HEK 293 cells depleted of 3' end processing factors CF Im 68 and CstF-64.

2012-05-15 | E-GEOD-37037 | biostudies-arrayexpress

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data