Neurite extension and protein tyrosine phosphorylation elicited by inducible expression of the v-src oncogene in a PC12 cell line.
Tyrosine-specific protein kinase activity in neuronal differentiation was studied in a PC12 pheochromocytoma cell line (PC12-B9) produced by stable transfection with an inducible v-src gene encoding an activated tyrosine kinase (pp60v-src) under the transcriptional control of the mouse metallothionine I gene promoter. Induction of pp60v-src expression with Cd2+ and Zn2+ resulted in the reversible differentiation of PC12-B9 cells into neuron-like cells. pp60v-src elicited morphological differentiation with apparent first order kinetics at the same rate as NGF-directed neurite outgrowth in PC12-B9 cells. v-src gene expression enhanced the rate of NGF-directed neurite extension in an additive manner. Induction of pp60v-src alone constitutively increased the levels of phosphotyrosine-modified proteins (130-120, 90, 83, 65, 60/59, 36 kDa) detected by immunoblotting with phosphotyrosine antibodies. NGF treatment of PC12-B9 cells transiently increased the levels of distinct phosphotyrosine-modified proteins (108, 46, 42 kDa), as well as common substrates, including a 59-kDa protein that comigrated with alpha-tubulin. Phosphotyrosine-modified proteins were not synergistically increased in PC12-B9 cells induced for both v-src and NGF. The nonsynergistic effects of v-src gene expression on neurite outgrowth and phosphorylation suggest that pp60v-src induces PC12 cell differentiation by an intracellular signaling pathway that is largely distinct from that induced by NGF.
Cox
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Department of Biochemistry, University of North Carolina, Chapel Hill.
Maness
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PF
D000310:Adrenal Gland Neoplasms; D000818:Animals; D002104:Cadmium / Q000494:pharmacology; D002454:Cell Differentiation; D005786:Gene Expression Regulation / Q000187:drug effects; D016391:Genes, src; D007700:Kinetics; D009414:Nerve Growth Factors / Q000187:drug effects; D009474:Neurons / Q000166:cytology* / Q000187:drug effects; D015688:Oncogene Protein pp60(v-src) / Q000235:genetics; D010673:Pheochromocytoma; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D051381:Rats; D013379:Substrate Specificity; D014162:Transfection; D014407:Tumor Cells, Cultured; D014443:Tyrosine / Q000378:metabolism; D015032:Zinc / Q000494:pharmacology
D009414:Nerve Growth Factors; D002104:Cadmium; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D015688:Oncogene Protein pp60(v-src); D015032:Zinc