miR-126 Is Regulated by the Transcription Factor, NF-κB3, via a Promoter Independent of Its Host Gene, EGFL7, in Ulcerative Colitis

Our previous studies showed that miR-126 was significantly upregulated in ulcerative colitis (UC) and promoted inflammatory responses by activating the nuclear factor-κB (NF-κB) signalling pathway. This study aimed to explore the transcriptional mechanisms involved in miR-126 upregulation in a cellular inflammation model. miRNA and mRNA expression were measured using qRT-PCR. Protein amounts were measured using western blotting. The core promoter sequences of miR-126 and its host gene EGF-like domain multiple 7 ( EGFL7 ) were determined through a series of plasmid DNA constructions via the luciferase reporter system. The binding of NF-κB3 to the core promoter region of miR-126 was detected using an electrophoretic mobility shift assay. Knockdown of NF-κB3 was performed using siRNA. miR-126 and EGFL7 mRNA were found to be upregulated in UC tissues. However, there was no significant correlation between miR-126 and EGFL7 . In the HT-29 cell line, stimulation of TNFα, IL-1β, lipopolysaccharide, MDP, HKM, and ODN2006 led to a discordant expression pattern of miR-126 and EGFL7 , while IFN-γ or FLA-ST treatment resulted in a concordant expression pattern of miR-126 and EGFL7 . Luciferase activity analysis revealed that miR-126 has its own independent promoter. The upregulated expression and increased phosphorylation of the transcription factor, NF-κB3, were observed in UC tissues. NF-κB3 could directly bind to the core promoter region of miR-126, and regulate the expression of miR-126 and EGFL7 . We demonstrated that miR-126 may have its own independent promoter and could be directly regulated by NF-κB3, partially by its host gene, EGFL7 , and by several molecular modulators. Our study provides further insights into the regulatory mechanisms of the upregulation of miRNA in inflammatory conditions like UC.