Regulation of 13(S)-hydroxyoctadecadienoic acid biosynthesis in Syrian hamster embryo fibroblasts by the epidermal growth factor receptor tyrosine kinase.
Metabolism of arachidonic and linoleic acid can be regulated by polypeptide growth factors in a variety of cell types. In Syrian hamster embryo (SHE) fibroblasts, epidermal growth factor (EGF) stimulates the conversion of exogenous linoleic acid to 13(S)-hydroxyoctadecadienoic acid (HODE). Inhibition of 13-HODE biosynthesis blocks the EGF-mitogenic response in SHE cells, and 13-HODE and its hydroperoxy precursor are potent and highly specific enhancers of EGF-dependent DNA synthesis. We demonstrated that EGF stimulates a biphasic production and release of endogenous 13-HODE. Through development of a stable isotope-dilution GC/MS assay for 13-HODE, we observed 13-HODE production as early as 5 min after EGF stimulation, and this initial phase peaked at 1 hr. A second rise in 13-HODE formation was seen at 2-4 hr, and this phase plateaued at 4-6 hr at a level of 30-40 ng/10(6) cells. EGF stimulation of 13-HODE biosynthesis is not mediated by transcriptional or translational regulation of the inducible form of prostaglandin H synthase. Based on enzyme inhibitor studies and structural characterization of products, the linoleate metabolite is apparently formed by an n-6 lipoxygenase that remains to be characterized. EGF stimulation of 13-HODE formation is linked with activation of the EGF receptor tyrosine kinase. Inhibition of EGF receptor tyrosine kinase activity with methyl-2,5-dihydroxycinnamate blocked EGF-dependent linoleic acid metabolism and EGF-regulated DNA synthesis. Potentiation of the EGF receptor tyrosine phosphorylation cascade through treatment of SHE cells with the tyrosine phosphatase inhibitor vanadate resulted in a 3-fold increase in EGF-stimulated 13-HODE production and a corresponding enhancement of the EGF mitogenic response. The coupling of EGF-regulated linoleic acid metabolism with the EGF receptor tyrosine kinase activity suggests the importance of specific linoleate compounds in mediating mitogenic signal transduction.
Glasgow
W C
WC
Laboratory of Molecular Biophysics, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina 27709, USA. glasgow@niehs.nih.gov
Hill
E M
EM
McGown
S R
SR
Tomer
K B
KB
Eling
T E
TE
D000818:Animals; D001094:Arachidonate 5-Lipoxygenase / Q000502:physiology; D002478:Cells, Cultured; D006224:Cricetinae; D004815:Epidermal Growth Factor / Q000494:pharmacology; D066246:ErbB Receptors / Q000502:physiology*; D005347:Fibroblasts / Q000378:metabolism; D008041:Linoleic Acids / Q000096:biosynthesis*; D008647:Mesocricetus
D008041:Linoleic Acids; C024348:13-hydroxy-9,11-octadecadienoic acid; D004815:Epidermal Growth Factor; D001094:Arachidonate 5-Lipoxygenase; D066246:ErbB Receptors