Identification of two C-terminal autophosphorylation sites in the PDGF beta-receptor: involvement in the interaction with phospholipase C-gamma.

Abstract

Two novel sites of autophosphorylation were localized to the C‐terminal tail of the PDGF beta‐receptor. To evaluate the importance of these phosphorylation sites, receptor mutants in which Tyr1009, Tyr1021 or both were replaced with phenylalanine residues, were expressed in porcine aortic endothelial (PAE) cells. These mutants were similar to the wild type receptor with regard to protein tyrosine kinase activity and ability to induce mitogenicity in response to PDGF‐BB. However, both the Y1009F and Y1021F mutants showed a decreased ability to mediate association with and the tyrosine phosphorylation of phospholipase C‐gamma (PLC‐gamma) compared to the wild type PDGF beta‐receptor; in the case of the Y1009F/Y1021F double mutant, no association or phosphorylation of PLC‐gamma could be detected. These data show that tyrosine phosphorylation of PLC‐gamma is dependent on autophosphorylation of the PDGF beta‐receptor at Tyr1009 and Tyr1021.

Fibroblast growth factor receptor-1 mediates chemotaxis independently of direct SH2-domain protein binding

“…It is known that many di erent growth factors stimulate cell migration. Signal transduction through the PDGF b-receptor leading to chemotaxis has been carefully dissected and shown to depend on activation on PI3-kinase as well as PLC-g (Kundra et al, 1994;RoÈ nnstrand et al, 1992;WennstroÈ m et al, 1994). In this paper, we have studied chemotactic signaling via the FGFR-1, and identi®ed a stretch between amino acid residues 759 and 773, located immediately following the second kinase domain, as critical for FGFR-1-dependent migration.…”

Activation of Stat5 by platelet-derived growth factor (PDGF) is dependent on phosphorylation sites in PDGF β-receptor juxtamembrane and kinase insert domains

“…The number of binding sites per cell, as determined by Scatchard analysis, were as follows: wildtype PDGF a-receptor, 40 000 ; wildtype PDGF b-receptor, 42 000 (Mori et al, 1993); PDGF b-receptor mutants, K634A, 40 000 (Sorkin et al, 1991), Y579F, 35 000; Y581F, 34 000 (Mori et al, 1993); Y716F, 32 000 ; Y740/751F, 37 000 ; and Y1009/1021F, 34 000 (RoÈ nnstrand et al, 1992). For Y771F, Y775F, Y778F mutant b-receptors, the levels of receptor expression was very similar to that of cells expressing wildtype receptors as determined by immunoblotting (Ruusala et al, manuscript in preparation).…”

“…Autophosphorylation sites in the kinase insert bind Grb2 (Tyr716; Arvidsson et al, 1994), the regulatory subunit (p85) of phosphatidylinositol 3-kinase (Tyr740 and Tyr751; Fantl et al, 1992;Kashishian et al, 1992), Nck (Tyr751; Nishimura et al, 1993), and the GTPase-activating protein of Ras (Tyr771; Fantl et al, 1992;Kashishian et al, 1992). Two autophosphorylation sites in the C-terminal tail mediate binding of phospholipase C-g (Tyr1009 and Tyr1021; Kashishian and Cooper, 1993;RoÈ nnstrand et al, 1992;Valius et al, 1993), Tyr1009 in addition binds the tyrosine phosphatase SHP2 .…”

New insights into protein‐tyrosine kinase receptor signaling complexes

“…These include tyrosine 771 in the kinase insert region, the site of interaction with RasGAP (Kazlauskas et al, 1992); tyrosines 1,009 and 1,021 in the C-terminal tail for interaction with PLCy (Ronnstrand et al, 1992;Kashishian & Cooper, 1993;; tyrosine 1,009, the binding site for the phosphotyrosine phosphatase, Syp (Feng et al, 1993;; and tyrosines 579 and 581 in the juxtamembrane segment N-terminal to the catalytic domain, as the binding site for Src family PTKs (Mori et al, 1993). The observation that a single receptor can use the same mechanism to interact with such a diverse range of signaling molecules (and this list is probably not complete) demonstrates the flexibility and functional diversity of SH2 domain-phosphotyrosine interactions.…”