Molecular cloning and functional expression of a phorbol ester-inducible 8S-lipoxygenase from mouse skin.
One of the effects of topical application of phorbol ester to mouse skin is the induction of an 8S-lipoxygenase in association with the inflammatory response. Here we report the molecular cloning and characterization of this enzyme. The cDNA was isolated by polymerase chain reaction from mouse epidermis and subsequently from a mouse epidermal cDNA library. The cDNA encodes a protein of 677 amino acids with a calculated molecular mass of 76 kDa. The amino acid sequence has 78% identity to a 15S-lipoxygenase cloned recently from human skin and approximately 40% identity to other mammalian lipoxygenases. When expressed in vaccinia virus-infected Hela cells, the mouse enzyme converts arachidonic acid exclusively to 8S-hydroperoxyeicosatetraenoic acid while linoleic acid is converted to 9S-hydroperoxy-linoleic acid in lower efficiency. Phorbol ester treatment of mouse skin is associated with strong induction of 8S-lipoxygenase mRNA and protein. By Northern analysis, expression of 8S-lipoxygenase mRNA was also detected in brain. Immunohistochemical analysis of phorbol ester-treated mouse skin showed the strongest reaction to 8S-lipoxygenase in the differentiated epidermal layer, the stratum granulosum. The inducibility may be a characteristic feature of the mouse 8S-lipoxygenase and its human 15S-lipoxygenase homologue.
Jisaka
M
M
Division of Clinical Pharmacology, Department of Pharmacology, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-6602, USA.
Kim
R B
RB
Boeglin
W E
WE
Nanney
L B
LB
Brash
A R
AR
D000595:Amino Acid Sequence; D000818:Animals; D001091:Arachidonate Lipoxygenases / Q000235:genetics* / Q000378:metabolism; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D012867:Skin / Q000201:enzymology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D018076:DNA, Complementary; D001091:Arachidonate Lipoxygenases; C054192:arachidonate 8-lipoxygenase; D013755:Tetradecanoylphorbol Acetate