Induction of arachidonate 12-lipoxygenase mRNA by epidermal growth factor in A431 cells.
12(S)-Hydroxyeicosatetraenoic acid is biosynthesized from arachidonic acid by the microsomal fraction of human epidermoid carcinoma A431 cells, and the microsomal 12-lipoxygenase activity is enhanced by about 2-fold by epidermal growth factor (EGF) with a 10-h lag period (Chang, W.C., Ning, C.C., Lin, M.T., and Huang, J.D. (1992) J. Biol. Chem. 267, 3657-3666). The microsomal 12-lipoxygenase in A431 cells was only 3% active with linoleic acid as compared with arachidonic acid. The enzyme was immunoprecipitated by a monoclonal antibody against human platelet 12-lipoxygenase but not by that against porcine leukocyte enzyme. A 3.1-kilobase mRNA was detected in A431 cells by Northern blot analyses using cDNA probe of human platelet 12-lipoxygenase. EGF could increase the 12-lipoxygenase mRNA level by about 2-fold with a lag period of 10 h, which was well parallel with the increase in the enzyme activity. The induction of the 12-lipoxygenase mRNA by EGF was completely blocked by 35 microM cycloheximide, if present in culture medium during EGF treatment, indicating that a de novo protein biosynthesis was essential for EGF-induced 12-lipoxygenase mRNA expression. Our data provide the first evidence for the inducibility of human 12-lipoxygenase gene expression by a growth factor.
Chang
W C
WC
Department of Pharmacology, College of Medicine, National Cheng Kung University, Tainan, Taiwan, Republic of China.
Liu
Y W
YW
Ning
C C
CC
Suzuki
H
H
Yoshimoto
T
T
Yamamoto
S
S
D000911:Antibodies, Monoclonal / Q000276:immunology; D000918:Antibody Specificity; D001092:Arachidonate 12-Lipoxygenase / Q000235:genetics*; D016718:Arachidonic Acid / Q000378:metabolism; D001792:Blood Platelets / Q000201:enzymology; D015152:Blotting, Northern; D003513:Cycloheximide / Q000494:pharmacology; D015342:DNA Probes; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D006801:Humans; D007163:Immunosorbent Techniques; D019787:Linoleic Acid; D008041:Linoleic Acids / Q000378:metabolism; D008861:Microsomes / Q000201:enzymology; D012333:RNA, Messenger / Q000096:biosynthesis*; D013379:Substrate Specificity; D014407:Tumor Cells, Cultured
D000911:Antibodies, Monoclonal; D015342:DNA Probes; D008041:Linoleic Acids; D012333:RNA, Messenger; D016718:Arachidonic Acid; D004815:Epidermal Growth Factor; D003513:Cycloheximide; D019787:Linoleic Acid; D001092:Arachidonate 12-Lipoxygenase