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Endostatin blocks vascular endothelial growth factor-mediated signaling via direct interaction with KDR/Flk-1
Young-Mi Kim1, Sewook Hwang, Young-Myoeng Kim
1Department of Biochemistry, College of Natural Sciences, Kangwon National University, Chunchon, Kangwon-Do 200-701, Korea.
Abstract:
Endostatin, a fragment of collagen XVIII, is a potent anti-angiogenic protein, but the molecular mechanism of its action is not yet clear. We examined the effects of endostatin on the biological and biochemical activities of vascular endothelial growth factor (VEGF). Endostatin blocked VEGF-induced tyrosine phosphorylation of KDR/Flk-1 and activation of ERK, p38 MAPK, and p125(FAK) in human umbilical vein endothelial cells. Endostatin also inhibited the binding of VEGF(165) to both endothelial cells and purified extracellular domain of KDR/Flk-1. Moreover, the binding of VEGF(121) to KDR/Flk-1 and VEGF(121)-stimulated ERK activation were blocked by endostatin. The direct interaction between endostatin and KDR/Flk-1 was confirmed by affinity chromatography. However, endostatin did not bind to VEGF. Our findings suggest that a direct interaction of endostatin with KDR/Flk-1 may be involved in the inhibitory function of endostatin toward VEGF actions and responsible for its potent anti-angiogenic and anti-tumor activities in vivo.
Insights
Endostatin directly binds to KDR/Flk-1, inhibiting vascular endothelial growth factor (VEGF) signaling. This interaction explains endostatin's potent anti-angiogenic and anti-tumor effects.
Area of Science:
- Molecular Biology
- Cell Biology
- Biochemistry
Background:
- Endostatin, a collagen XVIII fragment, is a known anti-angiogenic protein.
- The precise molecular mechanism of endostatin's action remains unclear.
- Vascular endothelial growth factor (VEGF) plays a crucial role in angiogenesis.
Purpose of the Study:
- To investigate the molecular mechanism of endostatin's anti-angiogenic activity.
- To examine the effects of endostatin on VEGF-mediated biological and biochemical processes.
- To determine if endostatin interacts directly with VEGF or its receptor.
Main Methods:
- Utilized human umbilical vein endothelial cells (HUVECs).
- Assessed VEGF-induced tyrosine phosphorylation of KDR/Flk-1 and activation of downstream signaling pathways (ERK, p38 MAPK, p125FAK).
- Performed binding assays with VEGF (isoforms 165 and 121) and purified KDR/Flk-1 extracellular domain; confirmed direct interaction via affinity chromatography.
Main Results:
- Endostatin inhibited VEGF-induced phosphorylation of KDR/Flk-1 and activation of ERK, p38 MAPK, and p125FAK in HUVECs.
- Endostatin blocked the binding of VEGF165 to endothelial cells and purified KDR/Flk-1.
- Endostatin also inhibited VEGF121 binding to KDR/Flk-1 and subsequent ERK activation.
- Direct binding of endostatin to KDR/Flk-1 was confirmed, but no direct interaction with VEGF was observed.
Conclusions:
- Endostatin directly interacts with the KDR/Flk-1 receptor.
- This direct interaction inhibits VEGF binding and downstream signaling.
- The findings suggest endostatin's anti-angiogenic and anti-tumor activities are mediated by its interaction with KDR/Flk-1, blocking VEGF actions.