The retinoic acid receptor-related orphan receptor α positively regulates tight junction protein claudin domain-containing 1 mRNA expression in human brain endothelial cells

Members of the claudin family play important roles in the formation of tight junctions (TJs) in several tissues. Claudin domain containing 1 (CLDND1) is homologous to this family and localizes to TJs and the cytoplasm when exogenously expressed in cultured epithelial cell lines. Furthermore, serum antibody levels of CLDND1-derived peptides are elevated in patients with cerebral infection, cardiovascular disease or diabetes mellitus as compared to healthy controls. However, CLDND1 transcriptional regulation remains poorly analyzed and most regional transcription factor binding sites remain to be defined. Notably, the CLDND1 promoter contains a putative response element for retinoic acid receptor-related orphan receptor alpha (ROR alpha), which is involved in the above-mentioned disorders. In this study, we found that Cldnd1 and Rora mRNA levels are correlated in rat tissues and that ROR alpha overexpression in human brain endothelial cells enhanced CLDND1 transcript expression. In addition, siRNA-mediated knockdown of ROR alpha significantly decreased CLDND1 transcription. An electrophoresis mobility shift assay indicated that ROR alpha binds to the identified response element in a sequence-specific manner. Furthermore, luciferase reporter assays confirmed that ROR alpha interacts with the CLDND1 promoter to enhance transcription. Taken together, our findings strongly suggest that CLDND1 is a direct ROR alpha target.