Deletion or substitution within the alpha platelet-derived growth factor receptor kinase insert domain: effects on functional coupling with intracellular signaling pathways.

Abstract

The tyrosine kinase domains of the platelet-derived growth factor (PDGF) and colony-stimulating factor-i (CSF-1)/c-fins receptors are interrupted by kinase inserts (ki) which vary in length and amino acid sequence.To define the role of the ki in the human aPDGF receptor (aPDGFR), we generated deletion mutants, designated aRAki-1 and aRAki-2, which lacked 80 (710 to 789) and 95 (695 to 789) amino acids of the 104-amino-acid ki region, respectively. Their functional characteristics were compared with those of the wild-type aPDGFR following introduction into a naive hematopoietic cell line, 32D. Biochemical responses, including PDGF-stimulated PDGFR tyrosine phosphorylation, phosphatidylinositol (PI) turnover, and receptor-associated PI-3 kinase activity, were differentially impaired by the deletions. Despite a lack of any detectable receptor-associated PI-3 kinase activity, 32D cells expressing oxRAkl-1 showed only partially impaired chemotactic and mitogenic responses and were capable of sustained proliferation in vitro and in vivo under conditions of autocrine stimulation by the c-sis product. 32D transfectants expressing the larger ki deletion (aRAki-2) showed markedly decreased or abolished biochemical and biological responses. However, insertion of the highly unrelated smaller c-fms (685 to 750) ki domain into aRAki-2 restored each of these activities to wild-type aPDGFR levels. Since the CSF-1R does not normally induce PI turnover, the ability of the c-fms ki domain to reconstitute PI turnover in the aRAki-2 transfectant provides evidence that the ki domain of the aPDGFR does not directly couple with this pathway. Taken together, all of these findings imply that their ki domains have evolved to play very similar roles in the known signaling functions of PDGF and CSF-1 receptors.

SHP-2 is involved in heterodimer specific loss of phosphorylation of Tyr771 in the PDGF β-receptor

“…The PDGF-AB isoform has been reported to possess the strongest ability to induce a mitogenic response in cells expressing both a-and b-receptors (Heidaran et al, 1991;Rupp et al, 1994). We have therefore investigated the possible unique features of the heterodimeric PDGF receptor complex compared to the homodimeric complexes, and found that in the heterodimeric complex activation of Ras and Erk2 MAP kinase were stronger and more sustained compared to homodimeric complexes (Ekman et al, 1999).…”

Increased mitogenicity of an αβ heterodimeric PDGF receptor complex correlates with lack of RasGAP binding

“…Examples include heteromeric complexes of members of the epidermal growth factor receptor family Solto et al, 1994;Wada et al, 1989;Heidaran et al, 1991;Kanakaraj et al, 1991;Seifert et al, 1989). PDGF is a family of dimeric isoforms of related A-and Bpolypeptide chains, which exert their cellular eects by binding to two structurally similar tyrosine kinase receptors.…”

Oncogenic activation of the αPDGFR defines a domain that negatively regulates receptor dimerization

“…To determine the e ects of the deletion mutant on the ligand-dependent and independent tyrosine phosphorylation of a and bPDGFRs endogenously expressed in NIH3T3 cells (Heidaran et al, 1991;Seifert et al, 1989), total cell lysates prepared from untreated or PDGF-BB treated NIH3T3 or NIH3T3/ aR D235 ± 290 cells were subjected to immunoblot analysis using anti-P-Tyr. Consistent with the size of the deletion mutant, a protein species with molecular weight of about 180 kDa was speci®cally phosphorylated on tyrosine in NIH3T3/aR D235 ± 290 (Figure 4a, lane 3).…”

“…The PDGF system is complicated in that three PDGF isoforms comprised of homo or heterodimers of A and B chains di erentially interact with a and b PDGF receptors (PDGFRs) encoded by two distinct genes (Claesson-Welsh et al, 1988;Matsui et al, 1989a;Yarden et al, 1986). Moreover, there is accumulating evidence that PDGF activation involves recruitment of receptor dimers (Bishayee et al, 1989;Heidaran et al, 1991;Heldin et al, 1989;Seifert et al, 1989;Ueno et al, 1991). In ®broblasts where both PDGFRs are expressed, PDGF-BB activates ab, bb and aa receptor dimers.…”

“…In ®broblasts where both PDGFRs are expressed, PDGF-BB activates ab, bb and aa receptor dimers. PDGF-AA induces formation of only aa receptor homodimers, while PDGF-AB is capable of recruiting aa and ab dimers (Heidaran et al, 1991;Seifert et al, 1989).…”