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

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Abstract
The tyrosine kinase domains of the platelet-derived growth factor (PDGF) and colony-stimulating factor-1 (CSF-1)/c-fms 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 alpha PDGF receptor (alpha PDGFR), we generated deletion mutants, designated alpha R delta ki-1 and alpha R delta ki-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 alpha PDGFR 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 alpha R delta ki-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 (alpha R delta ki-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 alpha R delta ki-2 restored each of these activities to wild-type alpha PDGFR 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 alpha R delta ki-2 transfectant provides evidence that the ki domain of the alpha PDGFR does not directly couple with this pathway. Taken together, all od these bindings imply that their ki domains have evolved to play very similar roles in the known signaling functions PDGF and CSF-1 receptors.