Abstract
In oxygenated cells, hypoxia‐inducible factor‐1 (HIF‐1) α subunits are rapidly degraded by a mechanism that involves ubiquitination by the von Hippel–Lindau tumor suppressor E3 ligase complex using 2‐oxoglutarate as a substrate. We examined the effect of 2‐oxoglutarate on the production of erythropoietin and vascular endothelial growth factor (VEGF). The expression of erythropoietin and VEGF protein were dose‐dependently downregulated in Hep3B cells by the addition of 2‐oxoglutarate. The promoter activity of VEGF‐luciferase was dose‐dependently downregulated by the addition of 2‐oxoglutarate. Gel mobility shift assays revealed that the addition of 2‐oxoglutarate dose‐dependently inhibited HIF‐1 binding activity, but did not affect GATA binding activity. Western blot analysis revealed that 2‐oxoglutarate dose‐dependently inhibited the HIF‐1α protein level in Hep3B cells in hypoxic conditions. However, MG132 (the proteasome inhibitor) rescued the inhibition of HIF‐1α protein expression by 2‐oxoglutarate. Furthermore, under hypoxic conditions, 2‐oxoglutarate dose‐dependently inhibited tube formation in in vitro angiogenesis assays. These results indicate that 2‐oxoglutarate treatment may be useful for the inhibition of angiogenesis. J. Cell. Physiol. 209: 333–340, 2006. © 2006 Wiley‐Liss, Inc.