Positive and negative modulatory roles of Platelet Endothelial Cell Adhesion Molecule-1 signaling in endothelial cell migration: coordination of Rho signaling and targeting of SHP-2 tyrosine phosphatase activity
Date of Award
Open Access Dissertation
Doctor of Philosophy (PhD)
Madri, Joseph A.
Platelet Endothelial Cell Adhesion Molecule-1 (PECAM-1), an immunoglobulin family adhesion molecule, is involved in endothelial migration and angiogenesis. We found that PECAM-1 deficient endothelial cells exhibit enhanced nondirected single cell motility and extension formation at the expense of wound-healing migration. These in vitro endothelial behaviors model aspects of highly developmentally and pathophysiologically relevant processes. Wound healing migration may model the reendothelialization of denuded vasculature following balloon angioplasty or the remodeling of vascular endothelium under variant flow conditions, while single cell motility is prominently involved in developmental processes such as endocardial-mesenchymal transition within the cardiac cushion. A specific deficiency of RhoGTP in PECAM-1 deficient endothelial cells accounted for their migratory phenotype. The serum sphingolipid sphingosine-1-phosphate (S1P) drives Rho-dependent migration and angiogenesis. Indeed PECAM-1 was necessary for an appropriate wound healing response to S1P. We have found that PECAM-1 localizes to rafts, and that in its absence heterotrimeric G protein components are differentially recruited to rafts, providing a potential mechanism for PECAM-1-mediated coordination of S1P signaling.PECAM-1 activates the tyrosine phosphatase SHP-2 via its phosphorylated immunoreceptor tyrosine-based inhibitory motif (ITIM). ITIM signaling is not required for the positive Rho-activating role of PECAM-1. Instead we found that ITIM signaling retards wound healing migration, and that during migration the second ITIM tyrosine becomes selectively dephosphorylated. The second ITIM tyrosine proved important for recruitment but not activation of SHP-2. Any SHP-2 that remains transiently associated with partially dephosphorylated PECAM-1 could amplify the migration signal by dephosphorylating adjacent PECAM-1 ITIMs. We found that PECAM-1 helps localize SHP-2 to the actin cytoskeleton, and to a subset of phosphoproteins. In fact, interfering with PECAM-1 ITIM signaling not only increased wound healing migration but decreased dephosphorylation of beta-catenin and focal adhesion kinase. SHP-2 targetting is instrumental in controlling endothelial wound healing migration, potentially via effects on adherens junction stability and focal contact dynamics.
Gratzinger, Dita, "Positive and negative modulatory roles of Platelet Endothelial Cell Adhesion Molecule-1 signaling in endothelial cell migration: coordination of Rho signaling and targeting of SHP-2 tyrosine phosphatase activity" (2003). Yale Medicine Thesis Digital Library. 2223.