Molecular regulation of VEGFR-2 expression and activation in endothelial cells
MetadataShow full item record
Angiogenesis, a hallmark step in tumor growth and metastasis, is primarily driven by the action of vascular endothelial growth factor (VEGF) on its receptor, VEGFR-2. Central to its regulation, the abundance of VEGFR-2 on the surface of endothelial cells determines VEGF's ability to stimulate vasculogenesis and pathological angiogenesis. By using a yeast two-hybrid system we have identified PDCL3 (Phosducin like 3) as a novel VEGFR-2 interacting protein that serves as a chaperone protein. PDCL3 expression is highly up-regulated in pathological neovascularization in a mouse model of oxygen-induced proliferative retinopathy and its expression correlates with VEGFR-2 expression. Knockdown of PDCL3 in endothelial cells significantly reduced hypoxia-induced expression of VEGFR-2. Further studies showed that PDCL3 binds to both nascent and mature VEGFR-2 through recognition of its juxtamembrane (JM) domain. Preventing binding of PDCL3 to the JM domain, and manipulating expression of PDCL3 by over-expression and siRNA in endothelial cells, showed that PDCL3 controls the abundance of VEGFR-2 through inhibition of its ubiquitination and degradation. Various in vivo and in vitro angiogenesis assays showed that PDCL3 activity is required for VEGF-mediated endothelial cell proliferation, capillary tube formation, and angiogenesis in developing zebrafish embryos. Further investigation of the molecular mechanisms of VEGFR-2 protein homeostasis revealed that its PEST domain controls degradation and ubiquitination of mature VEGFR-2. The data demonstrate that β-Trcp1 ubiquitin E3 ligase is recruited to the PEST domain and mediates ubiquitination and degradation of VEGFR-2. The PEST domain via phospho-Y1173 also contributes to activation of p38 mitogen-activated protein kinase (p38 MAPK), which results in the stabilization of VEGFR-2. The work demonstrates that VEGFR-2 protein homeostasis in endothelial cells is controlled at multiple levels involving co-chaperone functions of PDCL3, βTrcp1 ubiquitin E3 ligase and p38 MAPK pathway.
Thesis (Ph.D.)--Boston University PLEASE NOTE: Boston University Libraries did not receive an Authorization To Manage form for this thesis or dissertation. It is therefore not openly accessible, though it may be available by request. If you are the author or principal advisor of this work and would like to request open access for it, please contact us at email@example.com. Thank you.