Project description:Plant organs are comprised of distinct cell types with unique assemblages of mRNAs. This is a collection of CEL files of mRNA profiles of the total steady-state mRNAs and polysomal mRNAs of distinct cell types of the root tip of 7-d-old Arabidopsis thaliana seedlings. The cell type specific mRNA populations are those present in ribosome-mRNA complexes. This sub-population of mRNAs was obtained by first establishing a collection of Arabidopsis lines that express a FLAG-epitope tagged ribosomal protein L18 (RPL18) directed by promoters expressed in specific cell types and regions. Four different promoter:FLAG-RPL18 lines were used. The targeted cell types and promoters included root endodermis (pSCR) and root stelar xylem and pericycle (pWOL, pSHR). A CaMV 35S promoter:FLAG-RPL18 line was used to obtain the polysomal mRNA of multiple cell types. The immunopurification of ribosome-mRNA complexes of specific cell types was accomplished by the method described in Zanetti et al. (Plant Physiology, 138, 624-635; 2005). Hybridization of the immunopurified mRNAs to the Affymetrix ATH1 DNA microarray platform and subsequent data analysis permitted the identification of transcripts that are enriched or depleted in specific cell types of root tips. The dataset includes samples from cell types from seedlings grown under control conditions and cell types of seedlings exposed to low oxygen stress (hypoxia) for 2 h. Keywords: cell-type specific expression, hypoxic stress, polysomal mRNA, abiotic stress, endodermis, stele Overall design: 20 samples, 2 conditions (2 h hypoxia stress, 2 h non-stress), 2 RNA pools (Total mRNA and polysomal mRNA), 4 promoter lines, 2 replicates
Project description:Plant organs are comprised of distinct cell types with unique assemblages of mRNAs. This is a collection of CEL files of mRNA profiles of the total steady-state mRNAs and polysomal mRNAs of distinct cell types of the whole root and shoot of 7-d-old Arabidopsis thaliana seedlings. The cell type specific mRNA populations are those present in ribosome-mRNA complexes. This sub-population of mRNAs was obtained by first establishing a collection of Arabidopsis lines that express a FLAG-epitope tagged ribosomal protein L18 (RPL18) directed by promoters expressed in specific cell types and regions. Thirteen different promoter:FLAG-RPL18 lines were used. The targeted cell types and promoters included root atrichoblast (non-hair) epidermal cells (pGL2), root endodermis (pSCR), root stelar xylem and pericycle (pWOL, pSHR), root phloem companion cells (phloem CC) (pSUC2, pSultr2;2), root proliferating cells (pRPL11C), root cortex meristematic cells (pCO2), root cortex elongation/maturation cells (pPEP), shoot mesophyll (pRBCS), shoot epidermis (pCER5), shoot guard cells (pKAT1), shoot bundle sheath (pSultr2;2), shoot phloem CC (pSUC2) and shoot trichomes (pGL2). A CaMV 35S promoter:FLAG-RPL18 line was used to obtain the polysomal mRNA of multiple cell types. The immunopurification of ribosome-mRNA complexes of specific cell types/regions was accomplished by the method described in Zanetti et al. (Plant Physiology, 138, 624-635; 2005). Hybridization of the immunopurified mRNAs to the Affymetrix ATH1 DNA microarray platform and subsequent data analysis permitted the identification of transcripts that are enriched or depleted in specific cell types/regions of roots and shoots. The dataset includes samples from cell types/regions from seedlings grown under control conditions and cell types/regions of seedlings exposed to low oxygen stress (hypoxia) for 2 h. Keywords: cell-type specific expression, hypoxic stress, polysomal mRNA, abiotic stress, atrichoblasts, epidermis, cortex, endodermis, stele, phloem companion cells, guard cells, mesophyll Overall design: 79 samples, 2 conditions (2 h hypoxia stress, 2 h non-stress), 2 RNA pools (Total mRNA and polysomal mRNA), 2 organs, 13 promoter lines, 2-4 replicates
Project description:Nearly all mitochondrial proteins are nuclear-encoded and are targeted to their mitochondrial destination from the cytosol. Here, we used proximity-specific ribosome profiling to comprehensively measure translation at the mitochondrial surface in yeast. The majority of inner membrane proteins were co-translationally targeted to mitochondria, reminiscent of proteins entering the endoplasmic reticulum (ER). Comparison between mitochondrial and ER localization demonstrated that the vast majority of proteins were targeted to a specific organelle. A prominent exception was the fumarate reductase Osm1, known to reside in mitochondria. We identified a conserved ER isoform of Osm1, which contributes to the oxidative protein folding capacity of the organelle. This dual localization was enabled by alternative translation initiation sites encoding distinct targeting signals. These findings highlight the exquisite in vivo specificity of organellar targeting mechanisms. Overall design: Ribosome profiling of whole cell or streptavidin-purified ribosomes biotinylated by cytosolic, ER or mitochondrially localized biotin ligase in yeast and mammalian cells
Project description:We report cell-type specific ribosome profiling in a mouse glioma model. We report a strategy for cell-type specific ribosome profiling in vivo. Our strategy was applied to the characterization of a mouse glioma model.
Project description:Low oxygen stress dynamically regulates the translation of cellular mRNAs as a means of energy conservation in seedlings of Arabidopsis thaliana. Most of the highly hypoxia-induced mRNAs are recruited to polysomes and actively translated, whereas other cellular mRNAs become translationally inactive and are either targeted for stabilization or degradation. Here we identify the involvement of OLIGOURIDYLATE BINDING PROTEIN 1 (UBP1), a triple RNA Recognition Motif protein, in dynamic and reversible aggregation of translationally repressed mRNAs during hypoxia. Mutation or downregulation of UBP1C interferes with seedling establishment and reduces survival of low oxygen stress. By use of messenger ribonucleoprotein immunopurification, we show that UBP1C constitutively binds a subpopulation of mRNAs characterized by U-rich 3’-untranslated regions under normoxic conditions. During hypoxia, UBP1C association with non-U-rich mRNAs is enhanced concomitant with its aggregation into microscopically visible cytoplasmic foci, referred to as UBP1 stress granules (SGs). This UBP1C-mRNA association occurs as global levels of protein synthesis decline. Upon reoxygenation, rapid UBP1 SG disaggregation coincides with the return of the stabilized mRNAs to polysomes. The mRNAs that are highly induced and translated during hypoxia largely circumvent UBP1C sequestration. Thus, UBP1 is established as a component of dynamically assembled cytoplasmic mRNPs that sequester mRNAs that are poorly translated during a transient low energy stress. Immunoprecipated RNA associated with Arabidopsis UBP1C (IP) was compared with total cellular RNA from light (L), mock dark (D), 2 h hypoxia, and 2 h hypoxia + 20 min reoxygenation treated samples with duplicate hybridizations to the Affymetrix ATH1 Genechip array.
Project description:Post-transcriptional regulation including mRNA binding to ribosomes plays an important role in determining cell-type-specific gene expression patterns. Here, we applied an approach that profiles cell-type-specific mRNAs. The Translating Ribosome Affinity Purification method (TRAP; Heiman et al., Cell, 2008 and Doyle et al., Cell, 2008) was developed in mice and has been combined with the UAS/Gal4 system in Drosophila (Thomas et al., PLoS ONE, 2012). TRAP is a powerful method to find cell-type-specific differences at the level of the 'translatome' (Dougherty, Schmidt, Nakajima, & Heintz, Nucleic Acids Research, 2010). In parallel to published efforts, we developed and implemented the method for the fly and compared distinct head cell types and identified cell-type-specific transcript classes with neuronal (e.g. receptor-, neuropeptide- or hormone activity) or glial function (e.g. transporter activity). Neuronal TRAP genes are over-represented in the brain, larval CNS and thoracico-abdominal ganglion (Chintapalli, Wang, & Dow, Nature Genetics, 2007). Using cell-type-to-cell-type comparisons (e.g. neurons vs. glia), instead of a given cell population to the total (e.g. neurons vs. head), the differences could be identified with greater resolution. TRAP uncovered more neuronal genes compared to neuronal RNA polymerase II ChIP-seq data (Schauer et al., Cell Reports, 2013). Thus, TRAP data confirm the importance of post-transcriptional regulation in defining cell identity. TRAP is one of the best methods to reveal differential "omics" data among distinct cell types by profiling ribosome-bound mRNAs. TRAP is a promising tool to reveal cell-type-specific transcriptional and translational changes in a perturbed environment. Overall design: This dataset contains cell-type-specific ribosome-bound mRNA-seq profiles. Input and IP samples from Drosophila adult head neurons, glia and fat body.
Project description:Post-transcriptional gene regulation plays a significant role in the response to Pi starvation. Here, we utilized advances in next-generation sequencing technology to examine changes in transcriptional control, RNA association with translating ribosomes in 14-day-old Arabidopsis seedlings subjected to 7 days of Pi starvation. Overall design: 14 samples, 2 conditions 500uM NaH2PO4, 0uM NaH2PO4. 2 bioreplicates of 4 RNA pools (total mRNA, ribosome footprints, small RNA, double stranded RNA)
Project description:The aim of this study is to assess translational changes in macrophages over a time course of Salmonella infection.
This data is part of a pre-publication release. For information on the proper use of pre-publication data shared by the Wellcome Trust Sanger Institute (including details of any publication moratoria), please see http://www.sanger.ac.uk/datasharing/
Project description:Nucleosome organization and dynamics play a central role in controlling the DNA accessibility to regulatory factors of many critical cellular functions, especially gene regulation. However, despite extensive studies, the main factors determining nucleosome positioning and its fluctuation during cell cycle still remain elusive. Here, we present a large-scale study of nucleosome plasticity throughout the cell cycle and its interplay with gene expression based on genome-wide nucleosome positioning and mRNA abundance. We have clusterized distinct nucleosome architectures around transcription start sites and replication origins and studied their dynamics during the cell cycle progression. The most significant cell cycle-dependent changes occur at G1-S and G2-M transitions due to a large changes in gene expression in cell cycle regulatory genes. Taken together, our accurate study provides a dynamic picture of chromatin organization along cell cycle and its interplay with gene expression.