Distinctive regulation of v-Src-associated phosphatidylinositol 3-kinase during PC12 cell differentiation.
In chicken embryo fibroblasts, the binding of v-Src to PtdIns 3-kinase requires Src homology domains, SH3, SH2 and the SH1 or kinase domain, which induces the cytoskeletal disruption associated with fibroblast transformation. In the rat phaeochromocytoma PC12 cell line, v-Src has a different effect on the cytoskeleton, inducing neurite extension rather than cytoskeletal disruption. Here we show that v-Src-induced neurite outgrowth is suppressed by the selective PtdIns 3-kinase inhibitor LY294002, suggesting that this effect of v-Src in PC12 cells also requires the activity of the lipid kinase. However, in contrast with chicken embryo fibroblasts, the association of PtdIns 3-kinase with v-Src in PC12 cells is delayed until several hours after activating the v-Src tyrosine kinase. Furthermore the v-Src-associated p85 regulatory subunit of PtdIns 3-kinase is not phosphorylated on tyrosine in PC12 cells and associates only weakly with isolated v-Src homology domains (SH3/SH2) in a Src kinase-independent manner. However, p85 and v-Src both associate with an unidentified protein (of molecular mass approx. 68 kDa; termed p68), which becomes tyrosine phosphorylated concomitantly with the association of both p85 and PtdIns 3-kinase with v-Src in PC12 cells. Thus we conclude that the mode of regulation of v-Src-associated PtdIns 3-kinase is cell-context-dependent and that p68 might act as an adaptor protein to mediate the association of p85 and v-Src in PC12 cells. The different regulation of PtdIns 3-kinase in PC12 and in chicken embryo fibroblasts in response to v-Src activity might reflect the different cytoskeletal rearrangements induced by this oncoprotein in the two cell types.
Haefner
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The Beatson Institute for Cancer Research, Cancer Research Campaign Beatson Laboratories, Garscube Estate, Switchback Road, Bearsden, Glasgow G61 1BD, Scotland, U.K.
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D000818:Animals; D002867:Chromones / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000378:metabolism; D005786:Gene Expression Regulation; D018507:Gene Expression Regulation, Developmental; D015971:Gene Expression Regulation, Enzymologic; D008954:Models, Biological; D009025:Morpholines / Q000494:pharmacology; D016501:Neurites / Q000502:physiology*; D015688:Oncogene Protein pp60(v-src) / Q000378:metabolism*; D016716:PC12 Cells; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D019000:Phosphotyrosine / Q000032:analysis; D011485:Protein Binding; D051381:Rats; D018909:src Homology Domains
D002867:Chromones; D004791:Enzyme Inhibitors; D009025:Morpholines; D010750:Phosphoproteins; D019000:Phosphotyrosine; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D019869:Phosphatidylinositol 3-Kinases; D015688:Oncogene Protein pp60(v-src)