Characterization of rat somatotroph and mammotroph secretory granules. Presence of sulfated molecules

Abstract

The contents of a purified somatotroph and mammotroph secretory granule fraction isolated from rat anterior pituitaries were solubilized in 4 M urea and analyzed by PAGE. In gels electrophoresed under a variety of conditions and stained with Coomassie Blue only two major bands, identified as GH and PRL, were present. In gels stained with Stains-All (which stains anionic substances), several additional bands were detected. When quarter pituitaries were labeled with a [3H]amino acid mixture, GH and PRL accounted for greater than 95% of the radioactivity incorporated into the granules. After labeling with [35S]sulfate, two classes of radiolabeled sulfated components were detected in the granules: a class of trypsin-sensitive macromolecular components which were coincident with two of the bands seen after Stains-All, and a class of low molecular weight components. In order to examine the distribution of the two classes of sulfated components within somatotroph and mammotroph granules, granules were suspended in 0.4% (w/v) Lubrol PX at pH 4.0, a treatment which has been shown to selectively solubilize somatotroph granule contents leaving mammotroph granule cores intact. This treatment was found to solubilize greater than 95% of the GH and greater than 99% of the radiolabeled, low molecular weight sulfated components; in contrast, there was virtually no solubilization of either PRL or macromolecular sulfated components. The findings indicate (a) that [35S]sulfate is incorporated into both somatotroph and mammotroph granules, and (b) that the low molecular weight sulfated components are associated with somatotroph granules whereas the macromolecular sulfated components are associated with mammotroph granules.