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Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Transcription and translation profiling by array of yeast caf20 mutants

ABSTRACT: One common form of translational control is mediated by proteins that bind to the mRNA 5' cap-binding protein eIF4E. These proteins are collectively called 4E binding proteins (4EBPs). Saccharomyces cerevisiae possesses two 4EBPs that are encoded by non-essential genes called CAF20 and EAP1. To determine the impact of gene deletion on gene expression, we monitored the transcript level and also the translation status for each RNA using cycloheximide to freeze elongating ribosomes in wild-type, caf20 and eap1 cells. Polyribosome fractionation of cell extracts was used to separate highly translated and poorly translated mRNAs that were then separately analyzed.

ORGANISM(S): Saccharomyces cerevisiae

SUBMITTER: Ioulia Smirnova

PROVIDER: E-MEXP-1308 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Identifying eIF4E-binding protein translationally-controlled transcripts reveals links to mRNAs bound by specific PUF proteins.

Cridge Andrew G AG Castelli Lydia M LM Smirnova Julia B JB Selley Julian N JN Rowe William W Hubbard Simon J SJ McCarthy John E G JE Ashe Mark P MP Grant Christopher M CM Pavitt Graham D GD

Nucleic acids research 20100812 22

eIF4E-binding proteins (4E-BPs) regulate translation of mRNAs in eukaryotes. However the extent to which specific mRNA targets are regulated by 4E-BPs remains unknown. We performed translational profiling by microarray analysis of polysome and monosome associated mRNAs in wild-type and mutant cells to identify mRNAs in yeast regulated by the 4E-BPs Caf20p and Eap1p; the first-global comparison of 4E-BP target mRNAs. We find that yeast 4E-BPs modulate the translation of >1000 genes. Most target m ...[more]

PMID: 20705650

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Transcription and translation profiling by array of yeast eap1 mutants

Project description:One common form of translational control is mediated by proteins that bind to the mRNA 5' cap-binding protein eIF4E. These proteins are collectively called 4E binding proteins (4EBPs). Saccharomyces cerevisiae possesses two 4EBPs that are encoded by non-essential genes called CAF20 and EAP1. To determine the impact of gene deletion on gene expression, we monitored the transcript level and also the translation status for each RNA using cycloheximide to freeze elongating ribosomes in wild-type, caf20 and eap1 cells. Polyribosome fractionation of cell extracts was used to separate highly translated and poorly translated mRNAs that were then separately analyzed.

2008-03-01 | E-MEXP-1309 | biostudies-arrayexpress

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Project description:Post-transcriptional regulation by RNA binding proteins (RBPs) plays prominent roles in a variety of biological processes. In this study, by analyzing the global regulatory relationship between RBPs and their target mRNAs in yeast, we discovered that most RBP genes are co-regulated with their target genes, but the RBPs tend to dampen expression variation among their target mRNAs. We further examined a well-studied RBP gene, PUF3, and found that the protein decreases the variation of its target mRNAs by differentially affecting their decay. Our results provided new insights into the functional importance of RBPs in coordinating their target genes expression. Both the PUF3 deletion strain and wild type BY4742 (MATa, his3Δ1, leu2Δ0, met15Δ0, ura3Δ0) strain were cultured overnight in YPD media. 0.01 OD cells were transferred into fresh YPEG media until 1.0 OD. Total RNA was extracted by the standard Trizol protocol. mRNA was then purified using oligo-dT DynaBeads. cDNA sequencing library was constructed according to the protocol described by Wang et al. After sequencing, the reads were also mapped into S. cerevisiae genome by SOAP2 with no more than two mismatches. RPKM was used to represent the expression level of each gene.

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Efficiency of isotopic labbeling in whole Drosophila body

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Project description:The regulation of translation elongation plays a vital role in protein folding; an adequate translational pause provides time and cellular environments for the co-translational folding of nascent peptides. However, the genomic landscape, sequence determinants, and molecular consequences of translational pausing remain mostly unknown. In this study, we performed disome-seq that sequenced mRNA fragments protected by two consecutive ribosomes – a product of severe translational pauses during which the upstream ribosome collides into the paused one. We detected severe translational pauses on ~75% of yeast genes. These pauses were often explained by one of the three mechanisms: 1) slow ribosome releasing at stop codons, 2) slow peptide formation from proline, glycine, asparagine, and cysteine, and 3) slow leaving of polylysine from the exit tunnel of ribosomes. Notably, these amino acids also terminate the α-helical conformation. Such dual roles of amino acids establish an inborn coupling between the synthetic completion of a structural motif and a translational pause. Furthermore, paused ribosomes often recruit chaperones to assist protein folding. As a consequence, emergent protein structures during evolution should be ready to be correctly folded. Collectively, our study shows widespread translational pauses and sheds lights on a better understanding of the regulation of co-translational protein folding.

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Selective inhibition of yeast regulons by daunorubicin

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