Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Promiscuous gene expression in human medullary thymic epithelial cell subsets

ABSTRACT: Different human mTEC subsets (MUC1, CEACAM5 and SGLT1) were purified by sequential enzymatic digestion (collagenase/dispase, trypsin) followed by enrichment using magnetic beads (CD45 beads, Miltenyi Biotech) and FACS sorting. Cells of the surface phenotype CD45-, CDR2-, EpCAM+ were further subdivided into MUC1+/MUC1-, CEACAM5+/CEACAM5- and SGLT1+/SGLT1- fractions. RNA was isolated using μMACS™ SuperAmp™ protocol (Miltenyi Biotec) and hybridized to Illumina Whole-Genome Expression Beadchips. Gene expression of Antigen-positive and Antigen-negative mTEC subsets was compared. Total RNA was isolated from ex-vivo isolated human mTEC subsets using μMACS™ SuperAmp™ protocol (Miltenyi Biotec)

ORGANISM(S): Homo sapiens

SUBMITTER: Benedikt Brors

PROVIDER: E-GEOD-49625 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

Overlapping gene coexpression patterns in human medullary thymic epithelial cells generate self-antigen diversity.

Pinto Sheena S Michel Chloé C Schmidt-Glenewinkel Hannah H Harder Nathalie N Rohr Karl K Wild Stefan S Brors Benedikt B Kyewski Bruno B

Proceedings of the National Academy of Sciences of the United States of America 20130826 37

Promiscuous expression of numerous tissue-restricted self-antigens (TRAs) in medullary thymic epithelial cells (mTECs) is essential to safeguard self-tolerance. A distinct feature of promiscuous gene expression is its mosaic pattern (i.e., at a given time, each self-antigen is expressed only in 1-3% of mTECs). How this mosaic pattern is generated at the single-cell level is currently not understood. Here, we show that subsets of human mTECs expressing a particular TRA coexpress distinct sets of ...[more]

PMID: 23980163

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Project description:BACKGROUND: Orofacial development is a multifaceted process involving precise, spatio-temporal expression of a panoply of genes. MicroRNAs (miRNAs) constitute the largest family of noncoding RNAs involved in gene silencing, and represent critical regulators of cell and tissue differentiation. MicroRNA gene expression profiling is an effective means of acquiring novel and valuable information regarding the expression and regulation of genes, under the control of miRNA, involved in mammalian orofacial development. RESULTS: To identify differentially expressed miRNAs during mammalian orofacial ontogenesis, miRNA expression profiles from gestation day (GD) -12, -13 and -14 murine orofacial tissue were compared utilizing miRXplore™ microarrays from Miltenyi Biotech GmbH. TaqManTM quantitative Real-Time PCR was utilized for validation of gene expression changes. Cluster analysis of the microarray data was conducted with the clValid R package and the UPGMA (hierarchical) clustering method. Functional relationships between selected miRNAs were investigated using Ingenuity Pathway Analysis. Expression of over 26% of the approximately 588 murine miRNA genes examined was detected in murine orofacial tissues from GD 12, 13 and 14. Among these expressed genes several clusters were seen to be developmentally regulated. Differential expression of genes encoding miRNAs within such clusters were shown to target genes encoding proteins involved in cell proliferation, cell adhesion, differentiation, apoptosis and epithelial-mesenchymal transformation, all processes critical for normal orofacial development. Functional relationships between miRNAs differentially expressed were investigated using Ingenuity Pathway Analysis (IPA; Ingenuity Systems). CONCLUSIONS: Using miRNA microarray technology, unique gene expression signatures of hundreds of miRNAs in embryonic orofacial tissue were defined. Gene targeting and functional analysis revealed that the expression of numerous protein-encoding genes, crucial to normal orofacial ontogeny, may be regulated by specific miRNAs. Time-course experiment (Developmental Stages), ICR mice embryos on gestational days (GD) 12, 13 and 14. Biological replicates: For each day of gestation, 3 independent pools of 15 to 20 staged embryos were used to procure embryonic orofacial tissues for preparation of 3 distinct pools of RNA that were independently processed and applied to individual miRXplore™ microRNA Microarray chips (Miltenyi Biotec GmbH). Technology: 2-color spotted cDNA, Hy5 (experimental sample) vs. Hy3 (control - miRXplore Universal Reference).

Dataset Information

Promiscuous gene expression in human medullary thymic epithelial cell subsets

Publications

Overlapping gene coexpression patterns in human medullary thymic epithelial cells generate self-antigen diversity.

Similar Datasets

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