Regulation of endothelial cell activation by TGF-β family signaling

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We have already shown that Id1 and Herp2 promote and block angiogenic responses in endothelial cells, respectively. In present study, we tried to isolate Id1-and Herp2-interacting molecules which modulate function of Id1 and Herp2 by yeast two hybrid method. Among molecules isolated, E2-2, Id2, CGI-128, FHL2 and FLJ13861 could interact with either Id1 or Herp2 in mammalian cells. Out of them, we focused on E2-2 in further experiments because the heterodimer complex formation between E2-2 and Id1 was the strongest. Indeed, Id1 efficiently inhibited E2-2-induced luciferase activity. When E2-2 was over-expressed in endothelial cells, serum-induced proliferation and network formation in endothelial cells was suppressed in contrast with expression of Id1 in endothelial cells. To elucidate the mechanism by which E2-2 blocks angiogenic responses in endothelial cells, we tested the expression of VEGFR2, of which expression is known to be induced during endothelial activation, in endothelial ce … More lls when E2-2 was expressed in cells. As expected, expression of VEGFR2 mRNA was inhibited by E2-2, whereas E2-2-mediated decrease of VEGFR2 expression was improved by introduction of Id1 in the cells. Consistent with reduction of VEGFR2 mRNA, VEGFR2-lucifease activity was blocked by E2-2. Thus, it is possible that Id1 potentiates angiogenic responses in endothelial cells due to making heterodimer with E2-2 which substantially suppresses endothelial cell activation by blocking of VEGFR2 transcript.We also made ALK5 knock-in mice which can not transduce TGF-β/ALK5 signaling, but still possess TGF-β/ALK1 signaling. ALK5 knock-in mice die at E10.5 like ALK5 knock-out mice. We could not observe any mature vessel formation in yolk sac in ALK5 knock-in mice. The phenotype of yolk sac from ALK5 knock-in mice was quite similar to that from ALK5 knock-out mice. However, labyrinth formation in placenta from ALK5 knock-in mice could be detected in contrast with ALK5 knock-out mice. Thus, TGF-β/ALK1 signaling might improve defect of labyrinth formation seen in ALK5 knock-out mice. Less