Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Identification of gene targets of Meis2

ABSTRACT: The homeodomain protein Meis1 is essential for definitive hematopoiesis and vascular patterning in the mouse embryo. Meis2, another member of the same family, shares 82% protein identities with Meis1. Our present study suggested Meis2 exerts two distinguishable effects in differentiating ES cells. First, it increases the numbers of hematopoietic progenitors and extends their persistence in culture. Second, Meis2 skews hematopoietic differentiation by suppressing erythroid while enhancing megakaryocytic progenitor differentiation. To identify the underlying transcriptional bases of these actions, we carried out microarray analysis to compare the various populations of cells developing in ES differentiation cultures in the presence and absence of Meis2 induction. ES cells with dox-inducible Meis2 (A2lox.Meis2) were differentiated as embryoid bodies (EBs) for 6 days before plating on OP9-GFP cell monolayers and cytokines, and treated with (+) or without (-) doxycycline (dox). Cells were purified by cell sorting on day 7 or 8 into various populations based on levels of CD41 expression: GFP-CD41-, GFP-CD41+ (day 7) and GFP-CD41-,GFP-CD41int, and GFP-CD41hi (day 8). Gene expression of these purified populations was determined by microarray analysis.

ORGANISM(S): Mus musculus

SUBMITTER: Mi Cai

PROVIDER: E-GEOD-34541 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Dual actions of Meis1 inhibit erythroid progenitor development and sustain general hematopoietic cell proliferation.

Cai Mi M Langer Ellen M EM Gill Jennifer G JG Satpathy Ansuman T AT Albring Jörn C JC KC Wumesh W Murphy Theresa L TL Murphy Kenneth M KM

Blood 20120604 2

Myeloid ecotropic viral integration site 1 (Meis1) forms a heterodimer with Pbx1 that augments Hox-dependent gene expression and is associated with leukemogenesis and HSC self-renewal. Here we identified 2 independent actions of Meis1 in hematopoietic development: one regulating cellular proliferation and the other involved in megakaryocyte lineage development. First, we found that endogenous Mesp1 indirectly induces Meis1 and Meis2 in endothelial cells derived from embryonic stem cells. Overexp ...[more]

PMID: 22665933

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Project description:In order to detect transcriptional differences between primitive and definitive hematopoietic stem and progenitor cells during regular development in the zebrafish embryo, gata1-GFP+/+(18 somites), lmo2-GFP+/+ (18 somites and 35 hpf)1 and cd41-GFP+/+ (35 hpf) cells from transgenic embryos were individually separated from GFP-/- cells by flow cytometry at the indicated stages. For each individual population, pools of 600 - 1500 transgenic embryos were collected. After RNA extraction, labelled cRNA was hybridized onto Affymetrix microarrays. Individual experiments were performed with 2 or 3 biological replicates. Experiment Overall Design: gata1-GFP+/+(18 somites), lmo2-GFP+/+ (18 somites and 35 hpf)1 and cd41-GFP+/+ (35 hpf)2 cells were separated from GFP-/- cells Experiment Overall Design: by flow cytometry at the indicated stages. Sorting was based on propidium iodide exclusion, forward scatter, and GFP Experiment Overall Design: fluorescence, using a FACSVantage flow cytometer (Beckton Dickinson). Sorted cell populations were run twice to optimize cell Experiment Overall Design: purity. Total RNA from cell-sorted populations was extracted with TRIzol reagent (Invitrogen) and purified on RNeasy resins Experiment Overall Design: (Qiagen) according to the manufacturerâs recommendations. Total RNA was subjected to two rounds of linear amplification Experiment Overall Design: (Ambion) and hybridized to Affymetrix zebrafish Gene Chips, according to Affymetrix guidelines. After staining with a Experiment Overall Design: streptavidin-phycoerythrin conjugate (Molecular Probes), the fluorescence of bound RNA was quantitated by using a Gene Chip Experiment Overall Design: scanner (Affymetrix). The raw expression data were calculated and, after pairing of GFP+/+ and GFP-/- samples, statistically Experiment Overall Design: analyzed using methods implemented in Bioconductorâs âaffyâ package and available custom scripts 3,4. Experiment Overall Design: references: Experiment Overall Design: 1. Zhu, H. et al. Regulation of the lmo2 promoter during hematopoietic and vascular development in zebrafish. Dev Biol 281, Experiment Overall Design: 256-69 (2005). Experiment Overall Design: 2. Lin, H. F. et al. Analysis of thrombocyte development in CD41-GFP transgenic zebrafish. Blood 106, 3803-10 (2005). Experiment Overall Design: 3. Choe, S. E., Boutros, M., Michelson, A. M., Church, G. M. & Halfon, M. S. Preferred analysis methods for Affymetrix Experiment Overall Design: GeneChips revealed by a wholly defined control dataset. Genome Biol 6, R16 (2005). Experiment Overall Design: 4. Weber, G. J. et al. Mutant-specific gene programs in the zebrafish. Blood 106, 521-30 (2005).

Dataset Information

Identification of gene targets of Meis2

Publications

Dual actions of Meis1 inhibit erythroid progenitor development and sustain general hematopoietic cell proliferation.

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