| Summary: | 碩士 === 國立成功大學 === 藥理學研究所 === 85 === Arachidonate 12-lipoxygenase in mammalian cells catalyzed
the enzymatic reaction of arachidonic acid converting to
12(S)-hydroperoxyeicosatetraenoic acid, which is then converted
to 12(S)-hydroxyeicosatetraenoic acid (12(S)-HETE) by a
glutathione-dependent peroxi dase. Transcription of the
12-lipoxygenase gene gives rise to a 3.1 kb mRNA, which is
subsequently translated into a 72 kd enzyme. We previously
reported that epidermal growth factor (EGF) increased
12-lipoxygenase mRNA level by about 2-fold with a lag period of
10 h in human epidermoid carcinoma A431 cells. In the present
study, inhibition of EGF-induced expression of 12-lipoxygenase
mRNA by protein synthesis inhibitors was observed. Induction
of 12-lipoxygenase enzyme activity and mRNA expressionby EGF was
completely blocked by 10 mg / ml puromycin and by 2.5 mg / ml
anisomycin, if present in culture medium during EGF treatment,
indicating that a de novo protein biosynthesis was essential for
EGF-induced 12-lipoxygenase expression. Ras activation is at the
downstream of the EGF-signaling pathway, andmediates the
activation of mitogen-activated protein kinase. Overexpression
of Ha-ras in A431 cells increased the 12-lipoxygenase promoter
activity and mRNA expression when low dose (0.5 or 1 micro
gram) pSV2ras plasmid was transiently transfected, which
correlated closely with the cellular expression of Ras protein
in a time-dependent manner. However, under higher dose (4.0
micro gram) pSV2ras transfection, the 12-lipoxygenase protein
expression and enzyme activity was not increased in a dose-
dependent manner. So far these results suggest that Ras-
signaling pathway is at least involved in the
transcriptionalregulation of EGF-induced 12-lipoxygenase
expression, though we need more evidence to elucidate the role
of Ras.
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