Endothelial epsin deficiency decreases tumor growth by enhancing VEGF signaling
Satish Pasula, … , Wang Min, Hong Chen
Satish Pasula, … , Wang Min, Hong Chen
Published November 26, 2012
Citation Information: J Clin Invest. 2012;122(12):4424-4438. https://doi.org/10.1172/JCI64537.
View: Text | PDF
Research Article

Endothelial epsin deficiency decreases tumor growth by enhancing VEGF signaling

Abstract

Epsins are a family of ubiquitin-binding, endocytic clathrin adaptors. Mice lacking both epsins 1 and 2 (Epn1/2) die at embryonic day 10 and exhibit an abnormal vascular phenotype. To examine the angiogenic role of endothelial epsins, we generated mice with constitutive or inducible deletion of Epn1/2 in vascular endothelium. These mice exhibited no abnormal phenotypes under normal conditions, suggesting that lack of endothelial epsins 1 and 2 did not affect normal blood vessels. In tumors, however, loss of epsins 1 and 2 resulted in disorganized vasculature, significantly increased vascular permeability, and markedly retarded tumor growth. Mechanistically, we show that VEGF promoted binding of epsin to ubiquitinated VEGFR2. Loss of epsins 1 and 2 specifically impaired endocytosis and degradation of VEGFR2, which resulted in excessive VEGF signaling that compromised tumor vascular function by exacerbating nonproductive leaky angiogenesis. This suggests that tumor vasculature requires a balance in VEGF signaling to provide sufficient productive angiogenesis for tumor development and that endothelial epsins 1 and 2 negatively regulate the output of VEGF signaling. Promotion of excessive VEGF signaling within tumors via a block of epsin 1 and 2 function may represent a strategy to prevent normal angiogenesis in cancer patients who are resistant to anti-VEGF therapies.

Authors

Satish Pasula, Xiaofeng Cai, Yunzhou Dong, Mirko Messa, John McManus, Baojun Chang, Xiaolei Liu, Hua Zhu, Robert Silasi Mansat, Seon-Joo Yoon, Scott Hahn, Jacob Keeling, Debra Saunders, Genevieve Ko, John Knight, Gail Newton, Francis Luscinskas, Xiaohong Sun, Rheal Towner, Florea Lupu, Lijun Xia, Ottavio Cremona, Pietro De Camilli, Wang Min, Hong Chen

×

Figure 5

VEGF induces VEGFR2 ubiquitination and binding to epsins 1 and 2, and interaction of epsin and VEGFR2 requires epsin UIM.

Options: View larger image (or click on image) Download as PowerPoint
VEGF induces VEGFR2 ubiquitination and binding to epsins 1 and 2, and in...
(A) Schematic drawing of epsin depicts interactions between ENTH domain and PI(4,5)P2, UIM, and ubiquitin, COOH-terminal domain and clathrin, AP-2, and EH domain-containing proteins. (B) Lysates from BAECs stimulated with VEGF-A were immunoprecipitated with anti–epsin 1 or control IgG and Western blotted with anti-VEGFR2. Quantifications showing epsin 1 bound to VEGFR2 and total VEGFR2 are on right. (C) Lysates from BAECs stimulated with VEGF-A for 2 minutes were immunoprecipitated with anti-VEGFR2 or control IgG and Western blotted with anti–epsin 2. (D) Lysates from HEK 293T cells expressing indicated plasmids stimulated with VEGF-A for 2 minutes were immunoprecipitated with anti-FLAG and Western blotted with indicated antibodies. (E) Lysates from HEK 293T cells expressing indicated plasmids stimulated with VEGF-A for 2 minutes were immunoprecipitated with anti-VEGFR2 and Western blotted with indicated antibodies. (F) Lysates from BAECs stimulated with VEGF-A were immunoprecipitated with anti-VEGFR2 or control IgG and Western blotted with the indicated antibodies. (G) Lysates from HEK 293T cells expressing indicated plasmids stimulated with VEGF-A for 5 minutes were first immunoprecipitated with anti-FLAG and Western blotted with indicated antibodies. Immunoprecipitates were eluted and subjected to a second immunoprecipitation with anti-VEGFR2 and Western blotted with indicated antibodies. (H) Lysates from HEK 293T cells expressing indicated plasmids were stimulated with VEGF-A for 5 minutes and were immunoprecipitated with anti-HA and Western blotted with antibodies indicated. 50 ng/ml VEGF-A was used in B–H.