A conserved α-helix at the amino terminus of prosomatostatin serves as a sorting signal for the regulated secretory pathway

Mammalian prosomatostatin (PSST) contains the bioactive peptides SST-14 and SST-28 at the COOH-terminal end of the molecule and a putative sorting signal in the propeptide segment for targeting the precursor to the regulated secretory pathway. The NH2-terminal segment of PSST consists of an amphipathic cu-helix, which has been totally conserved throughout vertebrate evolution. We have analyzed the PSST-(3-15) region for sorting function by alanine scanning and deletional mutagenesis, Mutants created were stably expressed in AtT-SO cells. Regulated secretion was studied by analyzing basal and stimulated release of SST-14 LI and by immunocytochemistry for staining of SST-14 LI in punctate granules. Deletion of the PSST-(3-15) segment blocked regulated secretion and rerouted PSST for constitutive secretion as unprocessed precursor. Alanine scanning mutagenesis identified the region Pro(5)-Gln(12) as being important in precursor targeting, with Leu(7) and Leu(11) being critical. Molecular modeling demonstrated that these two residues are located in close proximity on a hydrophobic surface of the alpha -helix. Disruption of the alpha -helix did not impair the ability of PSST to be processed at the COOH terminus to SST-14 and SST-28. Processing, however, was shifted to the early compartments of the secretory pathway rather than storage granules and was relatively inefficient.