The effect of estrogen on phosphorylation of prolactin in the mouse pituitary gland

Several studies have indicated that prolactin (PRL) assumes oligomeric, proteolytically cleaved, phosphorylated and glycosylated forms. Phosphorylated PRL (PPRL) is considered to be the most important posttranslationally modified form in the rat. In the present study, we examined whether or not PRL is present in the mouse pituitary gland in the phosphorylated form. Mouse pituitary PRL was digested with acid phosphatase, resolved by two-dimensional gel electrophoresis, stained with Coomassie brilliant blue, and then immunoblotted against the anti-PRL, anti-phosphoserine and anti-phosphothreonine antibodies. We also examined whether PRL is phosphorylated by protein kinases and semi-quantified the ratios of PPRL to PRL in the pituitary gland. The results indicated that three types of PRL are present in the pituitary glands of both male and female mice. One was non-phosphorylated (isoform 1), and the other two were immunoreactive to anti-phosphoserine (isoform 2) and/or anti-phosphothreonine (isoform 3) antibodies. The ratio between isoforms 2 and 1 of the 30-day-old female mice was higher than that of the 20-day-old female mice. However, the ratios among the three isoforms in the male pituitary glands did not differ with age. The ratio of PPRL to isoform 1 was obviously reduced after ovariectomy (OVX), and it recovered with estrogen replacement. These results suggest that estrogen influences PRL phosphorylation in female mice.