Direct recruitment of CRK and GRB2 to VEGFR-3 induces proliferation, migration, and survival of endothelial cells through the activation of ERK, AKT, and JNK pathways.
Vascular endothelial growth factor receptor-3 (VEGFR-3) plays a key role for the remodeling of the primary capillary plexus in the embryo and contributes to angiogenesis and lymphangiogenesis in the adult. However, VEGFR-3 signal transduction pathways remain to be elucidated. Here we investigated VEGFR-3 signaling in primary human umbilical vein endothelial cells (HUVECs) by the systematic mutation of the tyrosine residues potentially involved in VEGFR-3 signaling and identified the tyrosines critical for its function. Y1068 was shown to be essential for the kinase activity of the receptor. Y1063 signals the receptor-mediated survival by recruiting CRKI/II to the activated receptor, inducing a signaling cascade that, via mitogen-activated protein kinase kinase-4 (MKK4), activates c-Jun N-terminal kinase-1/2 (JNK1/2). Inhibition of JNK1/2 function either by specific peptide inhibitor JNKI1 or by RNA interference (RNAi) demonstrated that activation of JNK1/2 is required for a VEGFR-3-dependent prosurvival signaling. Y1230/Y1231 contributes, together with Y1337, to proliferation, migration, and survival of endothelial cells. Phospho-Y1230/Y1231 directly recruits growth factor receptor-bonus protein (GRB2) to the receptor, inducing the activation of both AKT and extracellular signal-related kinase 1/2 (ERK1/2) signaling. Finally, we observed that Y1063 and Y1230/Y1231 signaling converge to induce c-JUN expression, and RNAi experiments demonstrated that c-JUN is required for growth factor-induced prosurvival signaling in primary endothelial cells.
Salameh
Ahmad
A
Dipartimento di Biologia Molecolare, Università degli Studi di Siena, Italy.
Galvagni
Federico
F
Bardelli
Monia
M
Bussolino
Federico
F
Oliviero
Salvatore
S
D002465:Cell Movement / Q000187:drug effects / Q000502:physiology*; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D042783:Endothelial Cells / Q000166:cytology / Q000502:physiology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D051380:GRB2 Adaptor Protein / Q000378:metabolism; D006801:Humans; D020928:Mitogen-Activated Protein Kinases; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D050737:Proto-Oncogene Proteins c-crk / Q000378:metabolism; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D014161:Transduction, Genetic / Q000379:methods; D014471:Umbilical Veins / Q000166:cytology / Q000502:physiology*; D040321:Vascular Endothelial Growth Factor Receptor-3 / Q000235:genetics / Q000378:metabolism*
D004791:Enzyme Inhibitors; D051380:GRB2 Adaptor Protein; D050737:Proto-Oncogene Proteins c-crk; D034741:RNA, Small Interfering; D040321:Vascular Endothelial Growth Factor Receptor-3; D017346:Protein Serine-Threonine Kinases; D020928:Mitogen-Activated Protein Kinases