Increased peptidylarginine deiminase expression during induction of prolactin biosynthesis in a growth-hormone-producing rat pituitary cell line, MtT/S.
Insulin and type I insulin-like growth factor (IGF-I) suppressed growth hormone (GH) expression followed by the induction of prolactin (PRL) biosynthesis in MtT/S cells cultured with normal sera. Insulin also increased the peptidylarginine deiminase activity in a dose-dependent manner. The increase was detectable at 1 ng/ml and reached a maximum (about 16-fold higher than the control) at 1 micrograms/ml. IGF-I showed similar but less prominent effects. The enzyme activity started to increase by 15 hr after the addition of insulin (500 ng/ml), and reached a plateau level at 48 hr. There were concurrent increases in the enzyme mRNA level, enzyme biosynthesis, and enzyme protein contents detected by Northern blot hybridization, [35S]-amino-acid incorporation, and Western immunoblot analysis, respectively. Two-color immunofluorescence staining at 1 day after the insulin addition detected a small number of peptidylarginine-deiminase-positive cells (about 1% of the total cells) which were also GH-positive. The enzyme-positive cells increased to 12% on day 2 and to 24-26% on days 4-6. PRL-positive cells first appeared in the enzyme-positive cell population on day 2, and PRL-positive, enzyme-negative cells appeared later. These results suggest that peptidylarginine deiminase expression increases in association with the hormone switching in MtT/S cells. When the cells were cultured in a steroid-depleted medium, insulin failed to increase the enzyme activity. The insulin action could be specifically restored by estrogen, indicating estrogen-insulin synergism in regulation of the enzyme expression.
Nagata
S
S
Department of Cell Chemistry, Tokyo Metropolitan Institute of Gerontology, Japan.
Uehara
T
T
Inoue
K
K
Senshu
T
T
D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D004967:Estrogens / Q000494:pharmacology; D005455:Fluorescent Antibody Technique; D013006:Growth Hormone / Q000096:biosynthesis; D006867:Hydrolases / Q000235:genetics / Q000378:metabolism*; D066298:In Vitro Techniques; D007328:Insulin / Q000494:pharmacology; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D010902:Pituitary Gland / Q000166:cytology / Q000378:metabolism*; D011388:Prolactin / Q000096:biosynthesis*; D000080002:Protein-Arginine Deiminase Type 4; D000076342:Protein-Arginine Deiminases; D012333:RNA, Messenger / Q000235:genetics; D051381:Rats
D004967:Estrogens; D007328:Insulin; D012333:RNA, Messenger; D007334:Insulin-Like Growth Factor I; D011388:Prolactin; D013006:Growth Hormone; D006867:Hydrolases; D000080002:Protein-Arginine Deiminase Type 4; D000076342:Protein-Arginine Deiminases