Endocrinology, Vol 119, 2097-2105, Copyright © 1986 by Endocrine Society

ARTICLES

Human growth hormone (GH)-releasing factor stimulates and somatostatin inhibits the release of rat GH variants

S Yokoya and HG Friesen

Two-dimensional polyacrylamide gel electrophoresis (2D PAGE) was used for the analysis of proteins secreted by male rat pituitary cells in monolayer culture in the presence of 10 nM human GH-releasing factor (hGRF) or 30 nM somatostatin (SRIF) or in the absence of these factors. More than 300 medium proteins were reproducibly detected either by fluorographic autoradiography or by silver staining. Immunoreactivity of each protein was examined after 2D PAGE followed by Western blotting and immunostaining with affinity-purified antirat GH (rGH) antibody. While there was a cluster of immunoreactive spots in the GH dimer range (40,000-50,000 mol wt), at least 16 medium proteins of mol wt 22,000 or less were also stained. Among these 16 proteins the release of 15 was stimulated and the release of 14 was inhibited by hGRF and SRIF, respectively. On the other hand, there were 3 proteins of approximate mol wt 16,000 whose secretion was regulated in a coordinate manner as rGH by the hypothalamic factors but which did not cross-react with anti- rGH antibodies. The increase or decrease in the radioactivity of each protein spot obtained from media after pituitary cells were incubated with [35S]methionine and hypothalamic factors was analyzed statistically. A pulse-chase study suggested that at least 7 of the hormonally regulated proteins, including rGH, were synthesized very rapidly. Finally, the 2D PAGE analysis of cell-free translation products of messenger RNA derived from male rat anterior pituitaries revealed the presence of about 40 rGH-immunoreactive proteins which included pre-GH. These data suggest that there are multiple forms of rGH-variants or rGH-related proteins. The biological significance(s) of all the rGH immunoreactive proteins and of the GRF- and SRIF-regulated pituitary proteins remains unclear. It is evident that a number of these proteins are synthesized and released rapidly by pituitary cells in culture. Furthermore, the presence of multiple genes for these rGH- related proteins is suggested by the large family of immunoreactive gene products identified after cell-free translation of messenger RNA derived from the pituitary.

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