Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Transcription profiling of Arabidopsis root tips in response to hypoxia using immunopurified mRNA-ribosome complexes

ABSTRACT: 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. Experiment 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

ORGANISM(S): Arabidopsis thaliana

SUBMITTER: Angelika Mustroph

PROVIDER: E-GEOD-14493 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Profiling translatomes of discrete cell populations resolves altered cellular priorities during hypoxia in Arabidopsis.

Mustroph Angelika A Zanetti M Eugenia ME Jang Charles J H CJ Holtan Hans E HE Repetti Peter P PP Galbraith David W DW Girke Thomas T Bailey-Serres Julia J

Proceedings of the National Academy of Sciences of the United States of America 20091020 44

Multicellular organs are composed of distinct cell types with unique assemblages of translated mRNAs. Here, ribosome-associated mRNAs were immunopurified from specific cell populations of intact seedlings using Arabidopsis thaliana lines expressing a FLAG-epitope tagged ribosomal protein L18 (FLAG-RPL18) via developmentally regulated promoters. The profiling of mRNAs in ribosome complexes, referred to as the translatome, identified differentially expressed mRNAs in 21 cell populations defined by ...[more]

PMID: 19843695

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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. Experiment 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: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

Project description:Translatome analysis by sucrose gradient centrifugation of cell lysates followed by microarray profiling of the polysomal and subpolysomal RNA fractions represents a way of both studying translational control networks and better approximating the proteomic representation of cells. It is an established notion that translational control takes place essentially at the translation initiation level, therefore the variation in abundance of a given mRNA species on polysomes can be directly related to the variation in abundance of the corresponding protein. Comparison of translatome profile changes with corresponding transcriptome profile changes can provide a measure of the degree of concordance between cellular controls affecting mRNA abundance and cellular controls affecting mRNA availability to translation. To provide a direct experimental evaluation of the phenomenon, we decided to study a classical example of transcriptional reprogramming of gene expression: Epidermal Growth Factor (EGF) treatment. This stimulus triggers a well known chain of intracellular transduction events, ultimately resulting in a multifaceted phenotypic spectrum of changes with prevalent induction of cell growth and proliferation. We subjected HeLa cells to serum starvation for 12h and then we added EGF at final concentration of 1 Î¼g/ml, profiling before and after 40 minutes of treatment the transcriptome, the translatome, coming from the polysomal pool of mRNAs after sucrose gradient separation, and also the mRNA content of the subpolysomal pool, expected not to be actively translated. Keywords: translatome profiling, polysomal profiling, polysomal RNA, translational control, translational profiling, polysome profiling, post-transcriptional regulation, EGF starvation release. The comparison between transcriptional and polysomal profiling was used for the discovery of general and mRNA-specific changes in the translation state of the serum starved HeLa cells transcriptome in response to EGF stimulus. To identify translationally regulated mRNA molecules, gene expression signals derived from the polysomal and subpolysomal RNA populations were compared by microarrays analysis to those obtained from unfractionated total RNAs. Polysomal RNA, subpolysomal RNA and total RNA were isolated from HeLa cells serum starved and treated with EGF. Cells lysates were collected before (t = 0 min) and after (t = 40 min) EGF treatment. All experiments were run in triplicates.

Dataset Information

Transcription profiling of Arabidopsis root tips in response to hypoxia using immunopurified mRNA-ribosome complexes

Publications

Profiling translatomes of discrete cell populations resolves altered cellular priorities during hypoxia in Arabidopsis.

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