Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Affinity purification of ribosomes and associated RNAs from stress-treated cells

ABSTRACT: In this study, we systematically identified ribosome associated RNAs. To identify ribosome associated RNAs, C-terminal ZZ-tagged Rpl16a, expressed under control of its native promoter, was affinity purified from whole cell extracts of cultures grown to mid-log phase. Extracts were incubated with immunoglobulin G (IgG) coupled microbeads, washed, and ribosomes were eluted by tobacco etch virus (TEV) protease treatment. To analyze how changes in steady-state mRNA levels (= transcriptome) are related to changes of respective messages in the translatome, total RNA and ribosome associated RNA from stress-treated and untreated cells were analyzed with yeast cDNA microarrays. To achieve this, fluorescently labeled (Cy5) cDNA was prepared from total RNA isolated from yeast cell extracts and from affinity-purified ribosomes, and each sample was mixed with differentially labeled (Cy3) cDNA prepared from a common reference RNA pool, and competitively hybridized on yeast cDNA microarrays. We performed five independent experiments with untreated cells grown in minimal medium (t=0 reference) and three or more independent biological replicates of treated cells. Altogether, we sampled the cell's response to five different "stress treatments" that were applied for relatively short periods of time (10 or 20 minutes) to avoid secondary effects triggered by transcriptional adaption. A stimulus or stress experiment design type is where that tests response of an organism(s) to stress/stimulus. e.g. osmotic stress, behavioral treatment Time: Duration of the treatment Growth Condition: Type of treatment stimulus_or_stress_design

ORGANISM(S): Saccharomyces cerevisiae

SUBMITTER: Andre Gerber

PROVIDER: E-GEOD-13653 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Stress-dependent coordination of transcriptome and translatome in yeast.

Halbeisen Regula E RE Gerber André P AP

PLoS biology 20090505 5

Cells rapidly alter gene expression in response to environmental stimuli such as nutrients, hormones, and drugs. During the imposed "remodeling" of gene expression, changes in the levels of particular mRNAs do not necessarily correlate with those of the encoded proteins, which could in part rely on the differential recruitment of mRNAs to translating ribosomes. To systematically address this issue, we have established an approach to rapidly access the translational status of each mRNA in the yea ...[more]

PMID: 19419242

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Dataset Information

Affinity purification of ribosomes and associated RNAs from stress-treated cells

Publications

Stress-dependent coordination of transcriptome and translatome in yeast.

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