c-Jun positively regulates transcription of thioredoxin reductase 1 gene through direct binding to its promoter

AIM: To investigate whether the transcription factor activator protein-1 (AP-1) family member c-Jun regulates the transcription of thioredoxin reductase 1 ( TrxR1 ). METHODS: The interaction of transcription factors with promoter region of TrxR1 was analyzed by bioinformatics methods. A series of plasmids containing truncated TrxR1 promoter regions with luciferase reporter gene were constructed. A c-Jun eukaryotic expression vector pcDNA3.1 (+)-c-Jun was also prepared. The plasmids containing truncated TrxR1 promoter regions were transfected into human embryonic kidney HEK293 cells and rat myocardium H9c2 cells for determining the luciferase activity. A putative AP-1 binding site located at TrxR1 promoter region -1 612~-1 600 was mutated by the method of site-directed mutagenesis. A c-Jun expression vector was transiently transfected into the 2 cell lines, and luciferase activity in each group was detected. Chromatin immunoprecipitation (ChIP) assay was used to demonstrate the binding ability of c-Jun to the promoter of TrxR1 . RESULTS: Restriction digestion analysis and sequencing results showed that 3 fragments of TrxR1 promoter with different lengths were obtained and the clones were consistent with the sequence in GenBank DNA database. The truncated TrxR1 promoter regions showed significant promoter activity. Transient transfection of c-Jun expression vector into HEK293 and H9c2 cells elevated TrxR1 promoter activity. However, the promoter activity disappeared when the AP-1 binding site was mutated or deleted. Similarly, ChIP analysis confirmed that c-Jun bound to TrxR1 promoter region. CONCLUSION: These results provide evidence that AP-1 family member c-Jun positively regulates TrxR1 gene expression through direct binding to its promoter.