The propeptide of preprosomatostatin mediates intracellular transport and secretion of α-globin from mammalian cells

Abstract

We have investigated the role of the somatostatin propeptide in mediating intracellular transport and sorting to the regulated secretory pathway. Using a retroviral expression vector, two fusion proteins were expressed in rat pituitary (GH3) cells: a control protein consisting of the β-lactamase signal peptide fused to chimpanzee α-globin (142 amino acids); and a chimera of the somatostatin signal peptide and proregion (82 amino acids) fused to α-globin. Control globin was translocated into the endoplasmic reticulum as determined by accurate cleavage of its signal peptide; however, α-globin was not secreted but was rapidly and quantitatively degraded intracellularly with a t 1/2 of 4-5 min. Globin degradation was insensitive to chloroquine, a drug which inhibits lysosomal proteases, but was inhibited at 16°C suggesting proteolysis occurred during transport to the cis-Golgi apparatus. In contrast to the control globin, ~30% of the somatostatin propeptide-globin fusion protein was transported to the distal elements of the Golgi apparatus where it was endoproteolytically processed. Processing of the chimera occurred in an acidic intracellular compartment since cleavage was inhibited by 25 μM chloroquine. 60% of the transported chimera was cleaved at the Arg-Lys processing site in native prosomatostatin yielding 'mature' α-globin. Most significantly, ~50% of processed α-globin was sorted to the regulated pathway and secreted in response to 8-Br-cAMP. We conclude that the somatostatin propeptide mediated transport of α-globin from the endoplasmic reticulum to the trans-Golgi network by protecting molecules from degradation and in addition, facilitated packaging of α-globin into vesicles whose secretion was stimulated by cAMP.