Milieu-induced, selective aggregation of regulated secretory proteins in the trans-Golgi network

Aggregative milieu not only allowed the recovery ofgranin aggregates from the TGN and immature secretory granules (and also from mature secretory granules; data not shown), compartments in which such aggregates are known to exist, but was sufficient to induce their formation in the more proximal compartments of the secretory pathway which normally do not contain dense cores of regulated secretory proteins . When the membrane of the RER was solubilized in aggregative rather than nonaggregative milieu, the granins were found in the pellet . This induction of granin aggregation is what would be expected if a change in the milieu is the cause of aggregation in vivo as secretory proteins travel along the secretory pathway, and if the aggregative milieu used in this study mimicked the lumenal milieu in the TGN . The aggregation ofthe granins probably occurred simultaneously with the solubilization of the RER membrane, i.e., when they were present at a relatively high concentration (presumably Chanat and Huttner Aggregation-mediating Sorting of Secretory Proteins several mg/ml) since no aggregation was observed when these proteins were exposed to aggregative milieu at much lower concentration, after their diffusion from the TGN lumen or after their partial purification . It therefore appears that these proteins must be present at a certain concentration for the aggregative milieu to be effective . This conclusion is consistent with previous morphological observations in vivo andwith results obtained on the aggregation ofthe granins in vitro (Gory et al., 1989) . A role ofprotein concentration in the aggregation process is further supported by our finding that after hormone treatment ofGH4C1 cells, which was found to increase granin synthesis and their concentration in the TGN, a greater proportion of SgII in the TGN was recovered in an aggregated state than for control cells.