Enhancer ID: | E_02_148 | |
Enhancer symbol: | CNS1 Enhancer | |
Species: | Mouse | |
Position : | chr10:78046488-78046636 |
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Biosample name: | Thymic Medullary Epithelial Cells (mTECs) | |
Experiment class : | Low throughput |
Enhancer type: | Enhancer | |
Disease: | -- | |
DO: | -- | |
Mesh: | -- | |
Distance from TSS: | >2KB | |
Pubmed ID: | 26364592 |
Enhancer experiment: | qPCR | |
Enhancer experiment description: | To determine the in vivo function of the Aire enhancer region,we generated knockout mice (CNS1-KO) with a deletion of 45 bp within CNS1 including both NF-κB sites.We first studied thymic expression of Aire in purified thymic epithelial cell subsets-cTEC, mTEClo, and mTEChi-from homozygous CNS1-KO mice by qPCR targeting exons 7–9. Strik_x0002_ingly, Aire was not expressed in mTEChi from CNS1-KO mice,whereas the gene was highly expressed in mTEChi from WT mice(Fig. 2A). |
Target gene : | Aire(Aire) | |
Strong evidence: | -- | |
Less strong evidence: | qPCR,Immunofluorescence,RT-PCR | |
Target gene experiment description: | To determine whether the upstream CNS1 region regulates these variants, we analyzed the expression levels of the majority of exons in the Aire coding region by qPCR.Aire-KO mouse results are shown in Supporting Information Fig.2B), indicating that CNS1 is required for the thymic expression of the majority of, if not all, Aire splice variants.Deficiency of Aire protein expression was confirmed in situ by immunofluorescence staining, which showed no Aire protein in the thymi of CNS1-KO(Fig. 2D). These results established that the CNS1 region is critical for Aire mRNA and protein expression in the mouse thymus. |
TF name : | Nfkb1(CVID12,EBP-1,KBF1,NF-kB,NF-kB1,NF-kappa-B1,NF-kappaB,NFKB-p105,NFKB-p50,NFkappaB,p105,p50) | |
TF experiment: | Luciferase Reporter Assay,EMSA | |
TF experiment description: | We next analyzed the CNS1 region for its responsiveness to NF-κB stimulation in luciferase reporter assays in vitro.We inserted a CNS1 fragment containing the two NF-κB binding sites upstream of the IFN-β minimal promoter and luciferase coding region and then introduced specific mutations into the NF-κB binding sites A and B,either individually or in combination. |
Enhancer function : | CNS1 is indispensable for RANK-induced Aire expression and that CNS1 is activated by NF-κB pathway complexes containing RelA. |
Enhancer function experiment: | 3C,Luciferase Reporter Assay,EMSA |
Enhancer function experiment description: |
To further examine the role of CNS1 in Aire gene regulation, we evaluated the spa_x0002_tial proximity between the CNS1 Enhancer and Aire promoter by chromosome conformation capture (3C) method.This result suggests that these distant regulatory regions are brought close to each other in intact chromatin. |
SNP ID: | -- | |
SNP position: | -- |
SNP experiment: | -- |
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