Mechanisms Controlling the Unique Phenotype of Brain Endothelium

Sade, Hadassah S. (2009). Mechanisms Controlling the Unique Phenotype of Brain Endothelium. PhD thesis The Open University.



Endothelial cells from different tissues differ widely in the expression of junctional proteins like occludin and transporters like the transferrin receptor. The mechanism(s) responsible for the differential expression of these proteins is not known. In this project we have studied how the occludin promoter interacts with nuclear transcription factors (TFs) from brain and non-brain endothelium. EMSA data indicates the TFs Sp1, Sp3 and YY1 are responsible for the specific binding to the occludin promoter in hCMEC/D3 cells, a transformed brain endothelial cell line. Using ChIP assays, we confirmed the interaction between these three transcription factors and DNA as these complexes were active in live cultured cells from transformed and primary brain endothelium. We investigated the expression and localisation of Sp1, Sp3 and YY1 in these cells and compared with lung endothelial cells and report the specific association of the TFs Sp3 and YY1 in brain endothelium which is absent in non brain endothelium. In addition, we have compared the activity of the occludin promoter in hCMEC/D3 cell to that in primary human dermal and lung endothelial cells by transfection with reporter vectors under the control of the full length and fragments of the occludin promoter. Our work identifies a group of transcription factors present in brain endothelium which may regulate the expression of the tight junction protein occludin. We propose a model whereby the TF Sp3 is necessary for the transcription of the occludin promoter in brain endothelium and YY1 negatively regulates promoter activity in non-brain endothelium by controlling access of Sp3 to the initiation sites on the occludin promoter.

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