THE LINOLEIC-ACID METABOLITE, (13S)-HYDROXYPEROXYOCTADECADIENOIC ACID, AUGMENTS THE EPIDERMAL GROWTH-FACTOR RECEPTOR SIGNALING PATHWAY BY ATTENUATION OF RECEPTOR DEPHOSPHORYLATION - DIFFERENTIAL RESPONSE IN SYRIAN-HAMSTER EMBRYO TUMOR-SUPPRESSOR PHENOTYPES

In Syrian hamster embryo (SHE) fibroblasts, epidermal growth factor re ceptor (EGFR) tyrosine kinase activity regulates the metabolism of end ogenous linoleic acid to (13S)-hydroperoxyoctadecadienoic acid (13S)-H PODE), (13S)-HPODE stimulates EGF-dependent mitogenesis in a SHE cell phenotype, which expresses tumor suppressor genes (supB(+)), but was n ot effective in a variant that does not express these suppressor genes (supB(-)). In the present study, we have investigated the potential e ffects of this lipid metabolite on the EGFR signaling pathways in thes e two SHE cell lines, Treatment of quiescent SHE cells with EGF produc ed a rapid, transient increase in the tyrosine phosphorylation of EGFR , Dependence on EGF concentration for EGFR tyrosine phosphorylation wa s similar in both SHE cell lines, but a more prolonged phosphorylation was detected in the supB(-) variant. Incubation of supB(+) cells with (13S)-HPODE and EGF increased EGFR autophosphorylation and tyrosine p hosphorylation on several signaling proteins with Src homology-a domai ns including GTPase-activating protein, The lipid metabolite did not s ignificantly alter EGF-dependent tyrosine phosphorylation in the supB( -) variant, Tyrosine phosphorylation of mitogen-activated protein (MAP ) kinase was also measured, The addition of (13S)-HPODE increased the extent and duration of MAP kinase tyrosine phosphorylation in supB(+) cells but not in the supB(-) variant, MAP kinase activity in supB(+) c ells, as measured in immunoprecipitates from cells after the addition of EGF, was increased by the presence of (13S)-HPODE. The addition of (1SS)-HPODE did not directly alter EGFR kinase activity or the interna lization of the EGFR, However, the addition of (13S)-HPODE to supB(+) cells extended the tyrosine phosphorylation of the EGFR in response to EGF, The dephosphorylation of the EGFR was measured directly, and a s lower rate was observed in the supB(-) compared with the supB(+) cells , Incubation of the supB(+) cells with (1SS)-HPODE attenuated the deph osphorylation of the EGFR. Thus, (13S)-HPODE stimulates EGF-dependent mitogenesis and up-regulation of EGF-dependent tyrosine phosphorylatio n by inhibiting the dephosphorylation of the EGFR. This study shows th at a metabolite of an essential dietary fatty acid, linoleic acid, can modulate tyrosine phosphorylation and activity of key signal transduc tion proteins in a growth factor mitogenic pathway.