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dc.contributor.authorWilliamson, Andrew J K
dc.contributor.authorSmith, Duncan L
dc.contributor.authorBlinco, David
dc.contributor.authorUnwin, Richard D
dc.contributor.authorPearson, Stella
dc.contributor.authorWilson, Claire L
dc.contributor.authorMiller, Crispin J
dc.contributor.authorLancashire, Lee J
dc.contributor.authorLacaud, Georges
dc.contributor.authorKouskoff, Valerie
dc.contributor.authorWhetton, Anthony D
dc.date.accessioned2009-05-22T13:14:06Z
dc.date.available2009-05-22T13:14:06Z
dc.date.issued2008-03
dc.identifier.citationQuantitative proteomics analysis demonstrates post-transcriptional regulation of embryonic stem cell differentiation to hematopoiesis. 2008, 7 (3):459-72 Mol. Cell Proteomicsen
dc.identifier.issn1535-9484
dc.identifier.pmid18045800
dc.identifier.doi10.1074/mcp.M700370-MCP200
dc.identifier.urihttp://hdl.handle.net/10541/68796
dc.description.abstractEmbryonic stem (ES) cells can differentiate in vitro to produce the endothelial and hematopoietic precursor, the hemangioblasts, which are derived from the mesoderm germ layer. Differentiation of Bry(GFP/+) ES cell to hemangioblasts can be followed by the expression of the Bry(GFP/+) and Flk1 genes. Proteomic and transcriptomic changes during this differentiation process were analyzed to identify mechanisms for phenotypic change during early differentiation. Three populations of differentiating Bry(GFP) ES cells were obtained by flow cytometric sorting, GFP-Flk1- (epiblast), GFP+Flk1- (mesoderm), and GFP+Flk1+ (hemangioblast). Microarray analyses and relative quantification two-dimensional LCLC-MS/MS on nuclear extracts were performed. We identified and quantified 2389 proteins, 1057 of which were associated to their microarray probe set. These included a variety of low abundance transcription factors, e.g. UTF1, Sox2, Oct4, and E2F4, demonstrating a high level of proteomic penetrance. When paired comparisons of changes in the mRNA and protein expression levels were performed low levels of correlation were found. A strong correlation between isobaric tag-derived relative quantification and Western blot analysis was found for a number of nuclear proteins. Pathway and ontology analysis identified proteins known to be involved in the regulation of stem cell differentiation, and proteins with no described function in early ES cell development were also shown to change markedly at the proteome level only. ES cell development is regulated at the mRNA and protein level.
dc.language.isoenen
dc.subject.meshAnimals
dc.subject.meshCell Differentiation
dc.subject.meshEmbryonic Stem Cells
dc.subject.meshFetal Proteins
dc.subject.meshGene Expression Profiling
dc.subject.meshGene Expression Regulation, Developmental
dc.subject.meshGreen Fluorescent Proteins
dc.subject.meshHematopoiesis
dc.subject.meshMetabolic Networks and Pathways
dc.subject.meshMice
dc.subject.meshProtein Array Analysis
dc.subject.meshProteomics
dc.subject.meshRNA, Messenger
dc.subject.meshReproducibility of Results
dc.subject.meshT-Box Domain Proteins
dc.subject.meshTandem Mass Spectrometry
dc.subject.meshTranscription, Genetic
dc.subject.meshVascular Endothelial Growth Factor Receptor-2
dc.titleQuantitative proteomics analysis demonstrates post-transcriptional regulation of embryonic stem cell differentiation to hematopoiesis.en
dc.typeArticleen
dc.contributor.departmentStem Cell and Leukemia Proteomics Laboratory, Faculty of Medical and Human Sciences, University of Manchester, Kinnaird House, Kinnaird Road, Manchester M20 4QL, United Kingdom.en
dc.identifier.journalMolecular & Cellular Proteomicsen
html.description.abstractEmbryonic stem (ES) cells can differentiate in vitro to produce the endothelial and hematopoietic precursor, the hemangioblasts, which are derived from the mesoderm germ layer. Differentiation of Bry(GFP/+) ES cell to hemangioblasts can be followed by the expression of the Bry(GFP/+) and Flk1 genes. Proteomic and transcriptomic changes during this differentiation process were analyzed to identify mechanisms for phenotypic change during early differentiation. Three populations of differentiating Bry(GFP) ES cells were obtained by flow cytometric sorting, GFP-Flk1- (epiblast), GFP+Flk1- (mesoderm), and GFP+Flk1+ (hemangioblast). Microarray analyses and relative quantification two-dimensional LCLC-MS/MS on nuclear extracts were performed. We identified and quantified 2389 proteins, 1057 of which were associated to their microarray probe set. These included a variety of low abundance transcription factors, e.g. UTF1, Sox2, Oct4, and E2F4, demonstrating a high level of proteomic penetrance. When paired comparisons of changes in the mRNA and protein expression levels were performed low levels of correlation were found. A strong correlation between isobaric tag-derived relative quantification and Western blot analysis was found for a number of nuclear proteins. Pathway and ontology analysis identified proteins known to be involved in the regulation of stem cell differentiation, and proteins with no described function in early ES cell development were also shown to change markedly at the proteome level only. ES cell development is regulated at the mRNA and protein level.


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