Membrane expression of annexin I is enhanced by calcium and TPA in cultured human keratinocytes.
Annexin I plays an important role in the process of keratinization as a component of the cornified envelope. To elucidate the function of annexin I in keratinization, we investigated the effects of calcium, epinephrine, hydrocortisone, and 12-O-tetradecanoyl phorbol 13-acetate (TPA) on the expression and localization of annexin I in cultured human keratinocytes. Normal human keratinocytes were cultured in serum-free culture medium (0.15 mM calcium) until 70% confluence. After incubation with a higher concentration of calcium (1.8 mM), TPA (100 nM), epinephrine (50 microM), or hydrocortisone (10 microM) for 24 h, the expression of annexin I protein and mRNA was examined using immunofluorescence, Western blot, and Northern blot techniques. Immunofluorescence microscopy showed increased membrane staining of annexin I by calcium, which was inhibited by the addition of epinephrine or hydrocortisone. Western blotting confirmed elevated annexin I on the cell membrane. It was increased in the cell membrane fraction, but not in the cytosol fraction. Interestingly, the mRNA level of annexin I was slightly reduced after incubation with calcium, whereas TPA upregulated both membrane expression and the mRNA level. Secretion of annexin I was increased by TPA but inhibited by calcium. Because calcium and TPA are known to promote keratinization, our data suggest that annexin I expression on the cell membrane is involved in the process of keratinization.
Sato-Matsumura
K C
KC
Department of Dermatology, Hokkaido University School of Medicine, Kita 15, Nishi 7, Kita-ku, Sapporo, 060-8638, Japan.
Matsumura
T
T
Nakamura
H
H
Sawa
H
H
Nagashima
K
K
Koizumi
H
H
D017305:Annexin A1 / Q000096:biosynthesis* / Q000235:genetics; D015152:Blotting, Northern; D002118:Calcium / Q000494:pharmacology*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D004837:Epinephrine / Q000494:pharmacology; D006801:Humans; D006854:Hydrocortisone / Q000494:pharmacology; D015151:Immunoblotting; D007150:Immunohistochemistry; D015603:Keratinocytes / Q000187:drug effects / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D017305:Annexin A1; D012333:RNA, Messenger; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; D006854:Hydrocortisone; D004837:Epinephrine