Abstract
The rat pituitary cell line GH4C1 secretes granins (chromogranin B and secretogranin II) and prolactin by the regulated secretory pathway. The intracellular storage of prolactin is preferentially induced by hormone treatment with estradiol, insulin, and epidermal growth factor. The goal of this study was to determine the effect of hormone treatment on storage of granins and constitutive secretory markers. The granins were efficiently stored in both hormone-treated and -untreated cells (17% of total secreted in 4 h). Secreted alkaline phosphatase (SEAP), a truncated membrane protein that would not be expected to enter secretory granules, and glycosaminoglycan, a marker for the constitutive secretory pathway, exhibited 70 80% secretion under both conditions. In comparison, the relative prolactin secretion was 31 and 68% from hormone-treated and -untreated cells, respectively. Phorbol ester and KCl stimulated prolactin secretion 2.3-fold from untreated cells and 5. 5-fold from hormone-treated cells. In contrast, SEAP secretion was stimulated 1.5-fold from both treated and untreated cells, consistent with secretion by the constitutive secretory pathway. Stimulated secretion of granins was detected from both hormone-treated and -untreated cells. These results suggest that granin and prolactin storage are differentially regulated and that the constitutive secretory pathway is not affected by hormone treatment.
Highlights
Endocrine cells exhibit a constitutive secretory pathway, common to all eukaryotic cells, as well as a regulated secretory pathway (Kelly, 1985)
When insulin is expressed in transfected GH4C1 cells, the peptide is sorted to the regulated secretory pathway, but insulin does not exhibit a preferential increase in storage in hormone-treated cells (Reaves et al, 1990)
Rat pituitary GH4C1 cells were cultured for 4 days with or without hormone treatment (EIE: 1 nM estradiol, 300 nM insulin, 10 nM epidermal growth factor (EGF))
Summary
Endocrine cells exhibit a constitutive secretory pathway, common to all eukaryotic cells, as well as a regulated secretory pathway (Kelly, 1985). It has been suggested that granins play a direct role in the sorting and packaging of peptide hormones in secretory granules (Rosa et al, 1985; Gorr et al, 1987a; Huttner et al, 1991; Scammell, 1993) In support of this hypothesis, granins exhibit calcium-induced aggregation at low pH, i.e. the conditions found in the trans-Golgi network and secretory granules of endocrine cells (Gorr et al, 1987b, 1988, 1989; Gerdes et al, 1989; Chanat and Huttner, 1991; Thompson et al, 1992). The expression of prolactin and granins is induced by treating the cells with a combination of estradiol, insulin, and epidermal growth factor (EGF) (Scammell et al, 1986, 1990a; Thompson et al, 1992) This hormone treatment preferentially induces prolactin granulogenesis and the intracellular storage of prolactin (Scammell et al, 1986; Reaves et al, 1990). The secretion and storage of secreted placental alkaline phosphatase (SEAP) and glycosaminoglycans was not affected by hormone-treatment
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