A high molecular weight form of somatostatin-28 (1-12)-like immunoreactive substance without somatostatin-14 immunoreactivity in the rat pancreas. Evidence that somatostatin-14 synthesis can occur independently of somatostatin-28.
Synthesis of somatostatin-14 (S-14) could occur through direct enzymatic processing of precursor somatostatin (prosomatostatin) or via sequential breakdown of prosomatostatin leads to somatostatin-28 (S-28) leads to S-14. If direct processing is important, it should theoretically generate S-14 and a molecule equivalent to prosomatostatin without the S-14 sequence. In an attempt to identify such a molecule, I characterized the molecular forms of S-28(1-12)-like immunoreactivity (S-28(1-12) LI) in the rat pancreas and compared the relative amounts of these forms with those of S-14-like immunoreactivity (S-14 LI). Pancreatic extracts were chromatographed on Sephadex G-50 and Sephadex G-75 columns (Pharmacia Fine Chemicals Inc., Piscataway, NJ) under denaturing conditions and immunoreactivity in the eluting fractions was analyzed by region-specific radioimmunoassays (RIAs). For RIA of S-28(1-12) LI we used a newly developed rabbit antibody R 21 B, 125I-Tyr12 S-28(1-14), and S-28(1-12) standards. This system detects S-28, S-28(1-12), high molecular weight forms of S-28(1-12), but not S-14. S-14 LI was measured using antibody R149, which detects S-14, S-28, and higher molecular weight S-14-like substances, but not S-28(1-12). Three forms of S-28(1-12) LI were identified: Mr 9,000-11,000, Mr 1,200 (corresponding to S-28(1-12), and Mr less than 1,000, comprising, respectively, 35, 53, and 12% of total immunoreactivity. The relative abundance of the 9,000-11,000 mol wt S-28(1-12) LI material was unchanged following removal of S-14 LI from pancreatic extracts by affinity chromatography before gel filtration. Serial dilutions of fractions containing 9-11,000 and 1,200 mol wt materials exhibited parallelism with synthetic S-28(1-12). The total pancreatic concentration of S-28(1-12) LI was 1.56 pmol/mg protein, of which S-28(1-12) accounted for 0.83 pmol/mg protein and 9-11,000 S-28(1-12) LI comprised 0.55 pmol/mg protein. Pancreatic S-14 LI concentration was 2.07 pmol/mg protein, of which 98% corresponded to S-14. S-28-related peaks accounted for <1% of immunoreactivity in both RIAs. I concluded that (a) S-14 is the main form of pancreatic S-14 LI; (b) S-28 is present in very small quantities, in the pancreas; (c) S-28(1-12) LI consist mainly of S-28(1-12) and 9-11,000 mol wt S-28(1-12) LI; (d) 9-11,000 l wt S-28(1-12) LI could represent prosomatostatin without the S-14 sequence; (e) the finding of high concentrations of 9-11,000 mol wt S-28(1-12) LI suggests that S-14 synthesis can occur independently of S-28 and that direct processing of prosomatostatin is an important pathway for S-14 synthesis in the pancreas.
D000818:Animals; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D008297:Male; D008970:Molecular Weight; D010179:Pancreas / Q000378:metabolism*; D011863:Radioimmunoassay; D051381:Rats; D011919:Rats, Inbred Strains; D013004:Somatostatin / Q000096:biosynthesis*; D053561:Somatostatin-28
D013004:Somatostatin; D053561:Somatostatin-28