title: Cytoplasmic protein phosphorylation related to multihormonal regulation of prolactin in pituitary cells.
abstract: The specialized functions of mammotropic cells from the anterior pituitary and of the pituitary tumor derived GH cell lines can be regulated by a large number of neurohormones and other substances. Although the biological responses to many of these regulatory signals are well documented, much less is known about the intracellular mechanisms involved in their action. Various "second messengers" are generated, which in turn regulate other intracellular activities like protein phosphorylation reactions. Thyrotropin-releasing hormone stimulates both prolactin synthesis and release by mammotrophs of the anterior pituitary, or by GH(4)C(1) cells in culture. Thyrotropin-releasing hormone simultaneously enhances the phosphorylation of a small number of cytoplasmic proteins in GH(4)C(1) cells, as revealed by two-dimensional polyacrylamide gel electrophoresis after (32)PO(4)(3?) labelling. On a pharmacological basis, two distinct subsets were identified among the seven phosphoproteins affected by thyrotropin-releasing hormone: set I, proteins 1-6, whose phosphorylation could be related to the regulation of prolactin synthesis; set II, proteins 7 and 8, whose phosphorylation was related to the modulation of prolactin release (Sobel A. and Tashjian A. H. Jr. (1983) J. Biol. Chem.258, 10312-10324). Phosphorylation of all these proteins but protein 1 was alkali-resistant, indicating that it might take place on tyrosine residues. Protein 1, which could be visualized by silver-staining and whose phosphorylation resulted in a shift of its position on two-dimensional gels, was present also in normal rat pituitary cells in primary culture. Moreover, thyrotropin-releasing hormone induced its phosphorylation as in GH(4)C(1) cells, thus indicating that this reaction is a normal regulatory process, not related to the tumoral origin of GH(4)C(1) cells. This observation allowed us to study some physiologically important regulatory mechanisms which were lost in GH cells as, for example, the inhibition of prolactin synthesis and release by dopamine: in normal anterior pituitary cells in culture, dopamine also prevented the thyrotropin-releasing hormone induced phosphorylation of protein 1. This result is in good agreement with those obtained with GH(4)C(1) cells, and extends them. The same technique allowed us to detect a protein with two-dimensional gel migration properties identical to those of protein 1 in certain tissues, like the adrenal medulla, but not in others, like the adrenal cortex. This tissue specificity might be related to the specific regulatory function of protein 1 in the cell.
authors.affiliation: Groupe de Biologie et Pathologie Neuromusculaires, INSERM U 153, 17 rue du Fe à Moulin, 75005 Paris, France.
authors.forename: A
authors.initials: A
authors.lastname: Sobel
authors.forename: M C
authors.initials: MC
authors.lastname: Boutterin