Here, we report on experiments in double-transgenic mice, in which RFP is expressed in all Foxp3+ Treg cells, whereas Foxp3-dependent GFP expression is exclusively confined to intrathymically induced Foxp3+ Treg cells. This novel molecular genetic tool enabled us to faithfully track and characterize naturally induced Treg cells of intrathymic (RFP+GFP+) and extrathymic (RFP+GFP) origin in otherwise unmanipulated mice.
Fluorochrome-based definition of naturally occurring Foxp3(+) regulatory T cells of intra- and extrathymic origin.
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View SamplesMicroarray experiments were performed using FAC-sorted young photoreceptors to analyze their transcriptome in comparison to remaining retinal cells at same developmental stage and retinal progenitors.
Increased integration of transplanted CD73-positive photoreceptor precursors into adult mouse retina.
Specimen part
View SamplesEmbryonic stem cells (ESCs) cells run a self-renewal gene expression program, requiring the expression of certain transcription factors accompanied by a particular chromosome organization to maintain a balance between pluripotency and the capacity for rapid differentiation. However, how transcriptional regulation is linked to chromosome organization in ESCs remains enigmatic. Here we show that Cohesin exhibits a functional role in maintaining ESC identity through association with the pluripotency transcriptional network. ChIP-seq analyses of the cohesin subunit Rad21 reveal an ESC specific cohesin binding pattern that is characterized by a CTCF independent colocalization of cohesin with pluripotency related transcription factors. Upon ESC differentiation, these binding sites disappear and instead new CTCF independent Rad21 binding sites emerge, which are enriched for binding sites of transcription factors implicated in early differentiation. Furthermore, knock-down of cohesin subunits causes expression changes that are reminiscent of the depletion of key pluripotency transcription factors, demonstrating the functional relevance of the cohesin - pluripotency transcriptional network association. Finally, we show that Nanog physically interacts with the cohesin interacting proteins Stag1 and Wapl, further substantiating this association. Based on these findings we propose that a dynamic placement of cohesin by pluripotency transcription factors contributes to a chromosome organization supporting the ESC expression program.
RAD21 cooperates with pluripotency transcription factors in the maintenance of embryonic stem cell identity.
Specimen part
View SamplesThe Cohesin complex has recently been described to regulate gene expression. We wanted to determine the gene expression profile specific in mouse ES cells after depletion of the Cohesin subunit Rad21.
No associated publication
Specimen part
View SamplesLong non-coding RNAs (lncRNAs) regulate diverse biological pathways. Unlike protein coding genes, where methods to comprehensibly study their functional roles in cellular systems are available, techniques to systematically investigate lncRNAs have largely remained unexplored. Here, we report a technology for combined Knockdown and Localization Analysis of Non-coding RNAs (c-KLAN) that merges phenotypic characterization and localization approaches to study lncRNAs. Using a library of endoribonuclease prepared short interfering RNAs (esiRNAs) coupled with a pipeline for synthesizing labeled riboprobes for RNA fluorescence in situ hybridization (FISH), we demonstrate the utility of c-KLAN by identifying a novel transcript Panct1 (Pluripotency associated non-coding transcript 1) that regulates embryonic stem cell identity. We postulate that c-KLAN should be generally useful in the discovery of lncRNAs implicated in various biological processes.
Combined RNAi and localization for functionally dissecting long noncoding RNAs.
Specimen part
View SamplesTranscriptome analysis of murine foetal NSCs (E14) after short-term (48 hours) and long-term (13 days) hypoxic (3% oxygen) culture compared to normoxic culture (21% oxygen)
No associated publication
Specimen part
View SamplesFetal liver of E14.5 RNaseh2b KOF and Rnaseh2b wild type embryos was isolated, RNA was extracted and microarray analysis using Affymetrix Mouse 430 2.0 gene chip was performed
Mammalian RNase H2 removes ribonucleotides from DNA to maintain genome integrity.
Specimen part
View SamplesPHD4 regulates the expression of Hypxia-inducible Factor 2 (HIF-2) alpha in LM8 osteosarcoma cells. PHD4 overexpression inhibits the growth of experimental tumor in syngenic mice but stimulates angiogenesis via Transforming Growth-Factor (TGF)-alpha.
PHD4 stimulates tumor angiogenesis in osteosarcoma cells via TGF-α.
Cell line, Treatment
View SamplesThe basic helix-loop-helix (bHLH) transcription factor hairy and enhancer of split (Hes3) is a member of the Hes/Hey gene family that regulates developmental processes in progenitor cells from various tissues. We demonstrated the Hes3 expression in mouse pancreatic tissue, suggesting it may have a role in modulating beta-cell function. We employed the mouse insulinoma cell line MIN6 to perform gene expression profiles in conditions known to modulate Hes3 based upon our previous work using neural stem cell cultures. In these conditions, cells showed elevated Hes3 expression and nuclear localization, grew efficiently and showed higher evoked insulin release responses, compared to serum-containing conditions. They also exhibited higher expression of the transcription factor Pdx1 and insulin.
No associated publication
Specimen part, Treatment
View SamplesThe basic helix-loop-helix (bHLH) transcription factor hairy and enhancer of split (Hes3) is a member of the Hes/Hey gene family that regulates developmental processes in progenitor cells from various tissues. We demonstrated the Hes3 expression in mouse pancreatic tissue, suggesting it may have a role in modulating beta-cell function. We employed a transfection approach to address specific functions of Hes3. Hes3 RNA interference opposed the growth of the mouse insulinoma cell line Min6. Western blotting and PCR approaches specifically showed that Hes3 RNA interference opposes the expression of Pdx1 and insulin. Likewise, Hes3 knock down reduced evoked insulin release from Min6 cells.
Hes3 is expressed in the adult pancreatic islet and regulates gene expression, cell growth, and insulin release.
Specimen part
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