Abstract
Binding of the platelet-derived growth factor (PDGF)-B to its receptor PDGFRbeta promotes proliferation, migration, and recruitment of pericytes and smooth muscle cells to endothelial cells, serving to stabilize developing blood vessels. The main goals of this study were to determine whether the extracellular domain of the PDGFRbeta can be proteolytically released from cell membranes and, if so, to identify the responsible sheddase and determine whether activation of the PDGFRbeta stimulates its shedding and potentially that of other membrane proteins. We found that the PDGFRbeta is shed from cells by a metalloproteinase and used loss-of-function experiments to identify ADAM10 as the sheddase responsible for constitutive and ionomycin-stimulated processing of the PDGFRbeta. Moreover, we showed that ligand-dependent activation of the PDGFRbeta does not trigger its own shedding by ADAM10, but instead it stimulates ADAM17 and shedding of substrates of ADAM17, including tumor necrosis factor alpha and transforming growth factor alpha. Finally, we demonstrated that treatment of mouse embryonic fibroblasts with PDGF-B triggers a metalloproteinase-dependent cross-talk between the PDGFRbeta and the epidermal growth factor receptor (EGFR)/ERK1/2 signaling axis that is also critical for PDGF-B-stimulated cell migration, most likely via ADAM17-dependent release and activation of ligands of the EGFR. This study identifies the principal sheddase for the PDGFRbeta and provides new insights into the mechanism of PDGFRbeta-dependent signal transduction and cross-talk with the EGFR.
Highlights
Signaling activated by the platelet-derived growth factor (PDGF)3-B and its receptor on pericytes, the PDGF receptor 
Based on our previous studies, the stimulation of PDGFR-alkaline phosphatase (AP) shedding by IM but not PMA was indicative of a role for ADAM10 in this process, because ADAM10 responds to short term stimulation (ϳ30 – 60 min) with 2.5 M of IM but not to short term stimulation with 25 ng/ml PMA [26, 27], which activates the metalloproteinase ADAM17
Shedding of PDGFR-AP transfected into wild type mouse embryonic fibroblasts (mEFs) was stimulated by 2.5 M IM and inhibited by 4 M MM, whereas shedding of PDGFR-AP transfected into Adam10Ϫ/Ϫ mEFs was only weakly induced by IM (Fig. 1B)
Summary
Signaling activated by the platelet-derived growth factor (PDGF)3-B and its receptor on pericytes, the PDGF receptor . Protein ectodomain shedding is a regulated process, and members of the ADAM (a disintegrin and metalloproteinase) family of membrane-anchored metalloproteinases [6, 7] have often been identified as the enzymes involved in the constitutive and stimulated release of integral membrane substrate proteins such as the pro-inflammatory cytokine TNF␣ (8 –10), ligands of the EGFR including EGF, HB-EGF, and TGF␣ [11,12,13,14,15] and growth factor receptors such as VEGFR2 [16] or the p75NTR [17]. The second component likely depends on stimulation of ADAM17 and the release of EGFR ligands that can activate ERK1/2 by binding to the EGFR [16] These findings prompted us to test whether a similar two-stage mechanism of ERK1/2 activation might be triggered by stimulation of the PDGFR with PDGF-B
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