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.