Repression of the blood endothelial marker CD146 by the homeobox gene PROX1

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CD146 is a cell adhesion molecule that has been shown to regulate cell adhesion, migration and proliferation of different cell types. It is highly expressed in blood endothelial cells (BECs), but is only lowly expressed in lymphatic endothelial cells (LECs). The PROX1 homeobox gene is a master regulator of lymphangiogenesis and its expression is necessary and sufficient to drive venous endothelial cells into a LEC phenotype. The highly permeable nature of the lymphatic vessels may partially derive from PROX1 mediated repression of CD146 transcription. We hypothesize that PROX1 promotes lymphatic differentiation by repressing CD146 transcription. In gain of function studies, Human Umbilical Vein Endothelial Cells (HUVECs) were infected with adenoviruses encoding EGFP, wild type PROX1 (AdProx1) or a Homeo-Prospero domain deleted version of PROX1 (AdHDPD), which cannot bind DNA. In order to knockdown PROX1, LECs were transfected with PROX1 specific siRNA. When compared to EGFP infected HUVECs, AdProx1 infected HUVECs had decreased CD146 expression both at protein and mRNA levels. In contrast, AdHDPD infected HUVECs had increased levels of CD146 expression. In support of a role for PROX1 in repressing CD146, PROX1 siRNA transfected LECs express higher levels of CD146 as compared to mock transfected LECs or LECs transfected with control siRNA. Based on these results, we predict that CD146 expression is kept at basal levels by an unknown repressor bound to the CD146 promoter. By interacting with this unknown repressor, PROX1 further represses CD146 expression. On the other hand, the DNA binding-deficient ΔHDPD version of PROX1 binds the unknown repressor and sequesters it from the CD146 promoter, thereby relieving the repression of CD146 expression in ECs. Different levels of CD146 expression between BECs and LECs might reflect the structural and functional differences between blood and lymphatic vessels. Since CD146 plays a critical role in EC adhesion, regulation of CD146 expression in ECs might be one of the key factors regulating vessel permeability.
lymphatic differentiation, endothelial cell fate