Abstract 15705: Novel Epsin-VEGFR2 Interactions Facilitated by c-Cbl Ubiquitination of Epsin and VEGFR2 Regulate VEGFR2 Signaling and Physiological and Pathological Angiogenesis
Angiogenesis is the process of developing new vasculature from pre-existing vasculature to ensure adequate oxygen and nutrient delivery to growing or damaged tissues. VEGF is a key pro-angiogenic growth factor that promotes new vessel sprouting via enhanced endothelial cell (EC) survival, proliferation and migration. VEGF signaling is regulated, in part, by the activation and degradation of VEGFR2. We recently reported that ubiquitin-interacting endocytic adaptor protein, epsin, binds ubiquitinated VEGFR2 via its ubiquitin interacting motif (UIM) and is required for degradation and signaling attenuation of activated VEGFR2. However, the molecular mechanism for epsin-dependent regulation of VEGFR2 activity is not understood. In this study, we report that activated VEGFR2 is ubiquitinated by E3 ligase, c-Cbl. Epsin UIM binds to the ubiquitin moieties on VEGFR2 which triggers ubiquitination of epsin by c-Cbl and facilitates the interaction of epsin with a novel ubiquitin-binding surface on VEGFR2. Additionally, we identified critical residues in VEGFR2 which are predicted to form hydrogen bonds with residues within the epsin UIM. Using site-directed mutagenesis, we determined the essential residues, His 816 and 891 and Arg 1080, 1022 and 1027 in VEGFR2, as well as Glu 183, 184 and 185 in epsin UIM are responsible for the specific epsin and VEGFR2 interaction. UIM peptides harboring mutations to said residues were used to investigate the requirement of above interactions in VEGF-VEGFR2 signaling and in vitro angiogenesis assays including EC proliferation, migration, tube formation and sprouting. We also determined their importance in regulating physiological and pathological angiogenesis using retina, matrigel plug, wound healing and xenograft tumor mouse models. Disrupting these novel interactions by inhibitory peptides could serve as potential therapeutics in vascular diseases.
Author Disclosures: H. Song: None. S. Pasula: None. M. Brophy: None. K.L. Tessneer: None. H. Wu: None. Y. Dong: None.
This research has received full or partial funding support from the American Heart Association
- © 2014 by American Heart Association, Inc.