Effects of a cytosolic protein on the interaction of rat pancreatic zymogen granules in vitro.

Photon correlation spectroscopy has been used to study the kinetics of aggregation of isolated rat pancreatic zymogen granules in vitro by monitoring time-dependent changes in mean particle size derived from the photon count autocorrelation function, g2(tau). Isolated granules were stable in isotonic sucrose (pH 5.4-7.0). At pH 6.0 they maintained a mean diameter of 1225 +/- 18 nm with a polydispersity index of 0.199 +/- 0.007. The mean granule diameter showed a limited decrease (approx. 20%) with increasing pH within the range 5.4-7.0, but the polydispersity index was unaltered. At pH greater than 7.0 granule instability was indicated by a rapid reduction in total photon counts. In solutions of monovalent cations ([M+] greater than 10 mM) and divalent cations ([M2+] greater than 0.5 mM) zymogen granules aggregated at a rate dependent upon both ion and granule concentration. These effects were consistent with the bimolecular nature of the interaction mechanism and were clearly distinguishable from the limited size changes associated with osmolarity. At concentrations of Na+ or K+ salts greater than 50 mM granule aggregation was accompanied by anion-dependent solubilisation. A soluble protein fraction separated from the pancreatic acinar cell cytosol by gel filtration reduced the mean diameter and polydispersity index of zymogen granules suspended in isotonic sucrose, inhibited cation-induced aggregation and stabilised granules to solubilisation induced by raising pH greater than 7.0 or exposure to high ionic strength media. The inhibitory effects of this protein were apparent at concentrations less than or equal to 10 micrograms X ml-1 (i.e. at inhibitor: granule protein ratios less than 1:20) and could not be mimicked by bovine serum albumin, the Ca2+-binding proteins calmodulin and troponin C (less than or equal to 100 micrograms X ml-1), nor the highly negatively charged polymer polyglutamate (less than or equal to 10 micrograms X ml-1). Inhibitory activity was also absent from fractions of rat liver cytosol prepared identically to pancreatic acinar cytosol. These observations are consistent with the presence in pancreatic acinar cells of a specific cytosolic granule stabilisation factor (or factors) that normally restricts zymogen granule interaction and may therefore play an important role in the regulation of granule mobility and exocytosis.