Identification of TNF-α-responsive promoters and enhancers in the intestinal epithelial cell model Caco-2.
Bottom Line: We found 520 promoters that significantly changed their usage level upon TNF-α stimulation; of these, 52% are not annotated.These enhancers share motif enrichments with similarly responding gene promoters.As a case example, we characterize an enhancer regulating the laminin-5 γ2-chain (LAMC2) gene by nuclear factor (NF)-κB binding.
Affiliation: The Bioinformatics Centre, Department of Biology & Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Ole Maaloes Vej 5, Copenhagen DK-2200, Denmark.Show MeSH
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Mentions: To verify the findings, we randomly selected 18 TNF-α(+) and 2 TNF-α(−) TCs within different classes: known promoters, novel alternative promoters and novel intergenic promoters. We validated their response to TNF-α by qPCR in quadruplicates and four different TNF-α concentrations. For validating alternative promoters, we used two primer pairs: one pair located immediately downstream of the TC of interest and the other pair just downstream of the most upstream annotated promoter. In the large majority (16 of 20) of cases, the qPCR results validated the CAGE data as well as the response to TNF-α (Fig. 2 and Supplementary Figure S1). We highlight a few examples below; fold changes and P-values refer to qPCR validations at 10 nM TNF-α, two-sided t-tests.Figure 2.
Affiliation: The Bioinformatics Centre, Department of Biology & Biotech Research and Innovation Centre (BRIC), University of Copenhagen, Ole Maaloes Vej 5, Copenhagen DK-2200, Denmark.