Previously, we found that bombesin receptor subtype 3 (BRS-3) significantly increased in an ozone-stressed airway hyperresponsiveness animal model and resulted in induced wound repair and protection from acute lung injury. In the present study, we determined molecular mechanisms of BRS-3 regulation in human BECs (bronchial epithelial cells) in response to ozone stress. Ten oligonucleotide probes corresponding to various regions of the BRS-3 promoter were used in EMSA (electrophoretic mobilityshift assays). Four were found to have an enhanced mobility shift with extracts from ozone-stressed cells. On the basis of the assay of mutated probes binding with extracts and antibody supershift, they were verified as MTF-1 (metal-regulatory-element-binding transcription factor-1), PPARα (peroxisome-proliferator-activated receptor α), AP-2α (activator protein 2α) and HSF-1 (heat-shock factor 1). Next, ChIP (chromatin immunoprecipitation) assay, site-directed mutagenesis technology and antisense oligonucleotide technology were used to observe these transcription factors associated with the BRS-3 promoter. Only AP-2α and PPARα increased ozone-inducible DNA binding on the BRS-3 promoter and BRS-3 expression. The time courses of AP-2α and PPARα activation, followed by BRS-3 expression, were also examined. It was shown that ozone-inducible BRS-3 expression and AP-2α- and PPARα-binding activity correlated over a 48 h period. The translocation of PPARα was observed by immunofluorescence assay, which showed that PPARα nuclear translocation increased after ozone exposure. Our data suggest that AP-2α and PPARα may be especially involved in this ozone-inducible up-regulation mechanism of BRS-3 expression.
PPARα and AP-2α regulate bombesin receptor subtype 3 expression in ozone-stressed bronchial epithelial cells
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Yu-rong Tan, Xiao-qun Qin, Yang Xiang, Tao Yang, Fei Qu, Yue Wang, Hui-jun Liu, H. Christian Weber; PPARα and AP-2α regulate bombesin receptor subtype 3 expression in ozone-stressed bronchial epithelial cells. Biochem J 1 July 2007; 405 (1): 131–137. doi: https://doi.org/10.1042/BJ20061754
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