Transcription of the occludin and claudin-5 genes has been investigated in the interest of understanding the mechanism of barrier induction and to improve therapeutic options for diseases that involve loss of the BBB and BRB. Previous work has identified a number of
cis-sequences of the occludin and claudin-5 promoters, as well as transcription factors that regulate their expression (
Figs. 5A,
5B). Cytokines, such as TNF-α and IFN-γ reduce expression from the occludin promoter in a luciferase reporter assay.
46 Transcription of occludin is inhibited by YY1 binding sites clustered at approximately −1200 to −1700,
47 whereas SP3 sites present directly upstream of the transcription start site have been postulated to induce transcription of the gene,
47 thus providing a potential model for differential expression of occludin in brain versus lung vascular endothelium. Additionally, TTF-1, an oncogene amplified in lung cancers, interacts with a sequence at −27 to −37 of the occludin promoter to increase expression and paradoxically decrease metastatic potential.
48 Our group and others have previously demonstrated GC induction of occludin expression. A distal imperfect glucocorticoid response element (GRE) sequence present in the occludin promoter has been postulated to direct this GC response.
49 In the current studies, a smaller promoter was used that did not include the previously mentioned GRE-like sequence. This shorter promoter maintained GC responsiveness and mutations of potential GRE half-sites did not abolish the GC effect.
28 Instead, ChIP analysis demonstrated that p54 and PSF transcription factors bind the OEE of occludin and homologous OEEs of claudin-5 and cadherin-9, whereas GR fails to bind. Further, siRNA knockdown experiments revealed that p54 is required for the steroid induction of barrier properties, suggesting p54/PSF binding to the OEE contributes to gene regulation of occludin and claudin-5. However, we did not observe an increase in p54 or PSF protein content in nuclear extracts with steroid treatment, suggesting that GCs do not act by simply increasing p54 expression. Alternatively, GC treatment may increase the specific DNA binding activity of the p54/PSF complex or induce another
trans-acting protein that facilitates p54/PSF binding to the OEE.