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Distribution of elongating and total RNA polymerase II in xrn1 mutants using GRO and RPCC

ABSTRACT: This SuperSeries is composed of the SubSeries listed below. Refer to individual Series

ORGANISM(S): Saccharomyces cerevisiae

SUBMITTER: Jose Perez-Ortin

PROVIDER: E-GEOD-43605 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Gene expression is circular: factors for mRNA degradation also foster mRNA synthesis.

Haimovich Gal G Medina Daniel A DA Causse Sebastien Z SZ Garber Manuel M Millán-Zambrano Gonzalo G Barkai Oren O Chávez Sebastián S Pérez-Ortín José E JE Darzacq Xavier X Choder Mordechai M

Cell 20130501 5

Maintaining proper mRNA levels is a key aspect in the regulation of gene expression. The balance between mRNA synthesis and decay determines these levels. We demonstrate that most yeast mRNAs are degraded by the cytoplasmic 5'-to-3' pathway (the "decaysome"), as proposed previously. Unexpectedly, the level of these mRNAs is highly robust to perturbations in this major pathway because defects in various decaysome components lead to transcription downregulation. Moreover, these components shuttle ...[more]

PMID: 23706738

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Distribution of total RNA polymerase II along the 5'/3' regions in xrn1 mutants

Project description:Determination of 3' or 5' intragenic RNA pol II occupancy. Maintaining the proper mRNA levels is a key aspect in the regulation of gene expression. The balance between mRNA synthesis and decay determines these levels. Using a whole-genome analysis, we demonstrate that most yeast mRNAs are degraded by the 5'-to-3' pathway (the "decaysome"), as proposed previously. Unexpectedly, the level of these mRNAs is highly robust to perturbations in this major pathway, as defects in various decaysome components lead to transcription down-regulation. Moreover, these components shuttle between the cytoplasm and the nucleus, in a manner dependent on proper mRNA degradation. In the nucleus, they associate with chromatin - preferentially ~30bp upstream of transcription start-sites - and directly stimulate transcription initiation and elongation. Hence, the decaysome has a dual role in maintaining mRNA levels. Significantly, proper import of some decaysome components seems to play a key role in coupling the two functions. The gene expression process is therefore circular, whereby the hitherto first and last stages are interconnected. This study focuses on the distribution of RNA pol II in the 3' or 5' region of 377 S. cerevisiae ORFs. S. cerevisiae cells grown in YPD to exponential phase were subjected to chromatin immunoprecipitation (ChIP) with anti-Rpb3 antibody. Data were normalized using whole cell extract hybridized on the same array. Home-made macroarrays containing 300 bp from both the 5' or 3' ends of each ORF were used.

2013-05-22 | E-GEOD-43604 | biostudies-arrayexpress

ChIP-chip of Spo14 and Mvp1

Project description:Genome-wide localization of two new RNA pol II interacting proteins. We use C-LYTAG to map binding sites of Spo14 and Mvp1 along the S. cerevisiae genome. Two biological replicates of each experiment were done. Keywords: ChIp-chip Two independent biological replicates of each IP. We used three macroarrays: V5R2-L1-20, V5R2-L1-21 and V5R2-L1-22 that were hybridized with Whole Cell Extract (WCE) and then with two biological samples of immunoprecipitated (IP) Mvp1 (V5R2-L1-20 and V5R2-L1-21) and Spo14 (V5R2-L1-20 and V5R2-L1-22)

2011-10-31 | E-GEOD-16905 | biostudies-arrayexpress

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Project description:A time-course transcriptomic experiment was performed in three geographically different wine yeast strains in order to test differences in gene expression as response to different nitrogen availability mRNA amounts for different wine yeast strains at different nitrogen availability was determined along the fermentation of a syntethic must. mRNA determinations were made at different fermentation stages (12, 24, 36 and 96 h after innoculation) Please note that there are two replicates per each stage (i.e. two raw data files per each sample; R1.txt and R2.txt) and the data were combined to generate the normalized data for each stage (i.e. sample).

2015-03-01 | E-GEOD-63187 | biostudies-arrayexpress

Genomic Run-On (GRO): determination of the nascent transcriptional rate and mRNA amount in Xrn1 mutants

Project description:Maintaining the proper mRNA levels is a key aspect in the regulation of gene expression. The balance between mRNA synthesis and decay determines these levels. Using a whole-genome analysis, we demonstrate that most yeast mRNAs are degraded by the 5'-to-3' pathway (the "decaysome"), as proposed previously. Unexpectedly, the level of these mRNAs is highly robust to perturbations in this major pathway, as defects in various decaysome components lead to transcription down-regulation. Moreover, these components shuttle between the cytoplasm and the nucleus, in a manner dependent on proper mRNA degradation. In the nucleus, they associate with chromatin - preferentially ~30bp upstream of transcription start-sites - and directly stimulate transcription initiation and elongation. Hence, the decaysome has a dual role in maintaining mRNA levels. Significantly, proper import of some decaysome components seems to play a key role in coupling the two functions. The gene expression process is therefore circular, whereby the hitherto first and last stages are interconnected. This study focuses on the transcriptional activity of RNA pol II in 6035 S. cerevisiae ORFs. S. cerevisiae cells grown in YPD to exponential phase were subjected to Genomic Run-On. Data were normalized using gDNA hybridized on the same array and by the software ArrayStat. Home-made macroarrays containing 6035 ORFs were used.

2013-05-22 | E-GEOD-29519 | biostudies-arrayexpress

Genomic Run On (GRO): determination of the nascent transcriptional rates and mRNA levels in several yeast mutants.

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2015-12-31 | E-GEOD-57467 | biostudies-arrayexpress

Genomic Run On (GRO): determination of the nascent transcriptional rates and mRNA levels in several yeast mutants (SAC3, SUS1, SRC1)

Project description:We analyzed the effect of SAC3, SUS1 and SRC1 on the transcription rates, mRNA stabilities and mRNA levels by doing GRO experiments in a deletion mutants and the partial C-truncated version of Sac3 comparing with a wild type. Some data for mRNA amounts (sus1 ans src1 mutants) are not included because were already in GEO database: GSE920 and GSE6370 accession numbers. This study focus on the transcriptional activity of RNA pol II in yeast genes. S. cerevisiae cells grown in YPD to exponential phase were subjected to Genomic Run On. Data were normalized by the ArrayStat software. Home-made macroarrays containing entire ORFs were used.

2015-12-31 | E-GEOD-63769 | biostudies-arrayexpress

Genomic Run On (GRO): determination of the nascent transcriptional rates in a spt16 mutant

Project description:We analyzed the effect of SPT16 on the transcription rates, mRNA stabilities and mRNA levels by doing GRO experiments in a thermosensitive spt16 point mutant (G132D) mutant at 35 ÂºC comparing with permisive temperature (30 ÂºC) and a wt treated in the same way. Cells (wt or spt16 mutant) were grown in SC-ura at 30 ÂºC to 0.25 OD600. At that time Î± factor was added and let to stand for 160 min in the same conditions. Then each sample (wt or mutant) was separated in two aliquots. One of them remain at 30ÂºC and the other was incubated at 35ÂºC. Both aliquots in the presence of Î± factor for 1h more. Finally, samples were processed for GRO experiment as described in Garcia-Marinez et al (2004). Three replicates of the whole experiment were done.

2016-01-05 | E-GEOD-64006 | biostudies-arrayexpress

Genomic Run On (GRO): determination of the nascent transcriptional rates in a series of a mex67 mutant.

Project description:The goal of the project was to check the effect on transcription rate of a sudden depletion of Mex67 protein by using a thermosensitive mutant: mex67-5 Cells were grown in YPD at 26ÂºC to 0.5 OD600. At that time cells were shifted to 37ÂºC to induce Mex67 protein inactivation and recovered after, 0, 5, 20, 40, 60 and 150 min.

2015-12-31 | E-GEOD-65228 | biostudies-arrayexpress

Distribution of elongating RNA polymerase II along the 5'/3' regions in xrn1 mutants

Project description:Determination of the 3' or 5' intragenic nascent transcriptional rate. Maintaining the proper mRNA levels is a key aspect in the regulation of gene expression. The balance between mRNA synthesis and decay determines these levels. Using a whole-genome analysis, we demonstrate that most yeast mRNAs are degraded by the 5'-to-3' pathway (the "decaysome"), as proposed previously. Unexpectedly, the level of these mRNAs is highly robust to perturbations in this major pathway, as defects in various decaysome components lead to transcription down-regulation. Moreover, these components shuttle between the cytoplasm and the nucleus, in a manner dependent on proper mRNA degradation. In the nucleus, they associate with chromatin - preferentially ~30bp upstream of transcription start-sites - and directly stimulate transcription initiation and elongation. Hence, the decaysome has a dual role in maintaining mRNA levels. Significantly, proper import of some decaysome components seems to play a key role in coupling the two functions. The gene expression process is therefore circular, whereby the hitherto first and last stages are interconnected. This study focuses on the transcriptional activity of RNA pol II in the 3' or 5' region of 377 S. cerevisiae ORFs. S. cerevisiae cells grown in YPD to exponential phase were subjected to Genomic Run-On. Data were normalized using gDNA hybridized on the same array. Home-made macroarrays containing 300 bp from both the 5' or 3' ends of each ORF were used.

2013-05-22 | E-GEOD-43602 | biostudies-arrayexpress

Genomic Run On (GRO): determination of the nascent transcriptional rates in a series of Hrp1 mutants.

Project description:The goal of the project was to check the differential effect on transcription rate of a sudden degron-induced depletion of Hpr1 protein and its comparison towards the effect of a constitutive absence of Hpr1 protein in a deletion strain. Cells were grown in YP-Raffinose + 0.1 mM CuSO4 at 26ÂºC to 0.5 OD600. At that cells were spun-down and changed to YP-Galactose without CuSO4 for 30 min at 26ÂºC. Then cells were shifted to 37ÂºC to induce degron-mediated proteolysis and recovered after 30 min. Samples include wt, hpr1delta and hpr1-degron after 0 or 30 min growth at 30ÂºC.

2015-12-31 | E-GEOD-65225 | biostudies-arrayexpress

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