About Enhancer
| Enhancer ID: | E_02_0798 |
| Species: | human |
| Position : | chr18:61533339-61545796   |
| Biosample name: | |
| Experiment class : | Low+High throughput |
| Enhancer type: | Enhancer |
| Disease: | -- |
| Pubmed ID: | 28578223 |
| Enhancer experiment: | ChIP-seq,Luciferase Reporter Assay,qRT-PCR |
| Enhancer experiment description: | A cluster of RNAs, not previously annotated, was found approximately 21 kb upstream of the SERPINB2 gene in SLE monocytes (Shi et al.,2014). The RNAs (772, 774, 775) formed a cluster near an enhancer, de?ned by H3K27ac in UCSC Genome Browser. 774 is centered on the H3K27ac peak, 775 is at the proximal edge of the H3K27ac peak and 772 is at the distal shores of the H3K27ac peak. We will refer to this region as Enhancer 1. We tested the DNA corresponding to the 775 ncRNA in an enhancer assay by sub-cloning it downstream of a minimal thymidine kinase (TK) promoter. The DNA corresponding to the 775 ncRNA location was capable of augmenting luciferase production in this transient transfection assay in two di?erent cell types (K562 and HEK293), thereby assigning it as a probable enhancer (Fig. 2A and B). For this reason, we will refer to the ncRNAs subsequently as eRNAs. Enhancer RNAs are predicted to have chromatin localization. To validate the remaining eRNAs, we used qRT-PCR from three di?erent cell types to demonstrate tissue speci?city and analyzed eRNA sub-cellular localization by fractionating cells. The eRNAs were found predominantly in monocytes and in the chromatin and nucleoplasm fractions predominantly (Fig. 2C and D). This demonstration of enhancer activity of the DNA and chromatin localization support the assignment of the ncRNAs as eRNAs. |
About Target gene
| Target gene : | SERPINB2(HsT1201,PAI,PAI-2,PAI2,PLANH2) |
| Strong evidence: | -- |
| Less strong evidence: | ChIP-seq,Luciferase Reporter Assay,qRT-PCR |
| Target gene experiment description: | A cluster of RNAs, not previously annotated, was found approximately 21 kb upstream of the SERPINB2 gene in SLE monocytes (Shi et al.,2014). The RNAs (772, 774, 775) formed a cluster near an enhancer, de?ned by H3K27ac in UCSC Genome Browser. 774 is centered on the H3K27ac peak, 775 is at the proximal edge of the H3K27ac peak and 772 is at the distal shores of the H3K27ac peak. We will refer to this region as Enhancer 1. We tested the DNA corresponding to the 775 ncRNA in an enhancer assay by sub-cloning it downstream of a minimal thymidine kinase (TK) promoter. The DNA corresponding to the 775 ncRNA location was capable of augmenting luciferase production in this transient transfection assay in two di?erent cell types (K562 and HEK293), thereby assigning it as a probable enhancer (Fig. 2A and B). For this reason, we will refer to the ncRNAs subsequently as eRNAs. Enhancer RNAs are predicted to have chromatin localization. To validate the remaining eRNAs, we used qRT-PCR from three di?erent cell types to demonstrate tissue speci?city and analyzed eRNA sub-cellular localization by fractionating cells. The eRNAs were found predominantly in monocytes and in the chromatin and nucleoplasm fractions predominantly (Fig. 2C and D). This demonstration of enhancer activity of the DNA and chromatin localization support the assignment of the ncRNAs as eRNAs. |
About TF
| TF name : | NSMF(HH9,NELF)CDK9(C-2k,CDC2L4,CTK1,PITALRE,TAK) |
| TF experiment: | ChIP |
| TF experiment description: | To examine whether NELF and its major kinase are localized to the promoter and Enhancer of SERPINB2, we performed ChIP studies for NELF and CDK9. |
About Function
| Enhancer function : | Knock-down of the enhancer RNAs compromised stimulus induction of promoter and enhancer chromatin changes. Conversely,over-expression was associated with enhanced recruitment of c-JUN and increased expression of SERPINB2 mRNA expression. |
| Enhancer function experiment: | qRT-PCR |
| Enhancer function experiment description: |
We knocked down 774 and 775 as SERPINB2-speci?c eRNAs. Knock-down was validated by qRT-PCR of the target eRNAs. Knock-down of 774 and 775 led to inhibition of SERPINB2 mRNAs but not SERPINB10, which is not co-regulated with SERPINB2 (Fig. 3B). We then over-expressed 774 and 775 to determine whether they could regulate SERPIN gene expression when provided in trans. Each over-expression construct was validated as expressing the correct RNA species (Fig. 3C). Over-expression of both 774 and 775 led to higher SERPINB2 RNA but not the distinctly regulated SERPINB10.Ano?-target, control RNA, 477, had no e?ect. Enhancer RNAs have been hypothesized to regulate gene expression via diverse potential mechanisms. We tested whether they regulated histone modi?cations. To de?ne e?ects related to eRNAs, we used anti-sense oligonucleotides to knock-down 774 and 775.We examined chromatin marks of gene activation in MonoMac6 cells at baseline and after treatment with LPS (Fig. 4CF). We compared the e?ects of 774 and 775 knock-down with GFP knock-down. Non-transfected cells were included as assay controls for each experiment but are not shown to streamline the ?gures.The GAPDH promoter was una?ected by LPS stimulation and serves as a control. The anti-sense oligonucleotides to 774 and 775 were associated with diminished acquisition of the chromatin marks seen with LPS stimulation at Enhancer 1. The e?ect was limited to LPS-stimulated cells and was not observed in resting cells. Knock-down of 774 and 775 primarily a?ected H3K4me3 and H3K27ac at the promoter. These data demonstrate that eRNAs regulate histone modi?cations not just at the enhancer but also at their target gene promoter. |
About SNP
| SNP ID: | -- |
Upstream Pathway Annotation of TF
Enhancer associated network
The number on yellow line represents the distance between enhancer and
target gene