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

angles) . Because of their molecular mass, we suspected that the 78-and 54-kD protein were the ER-resident proteins BiP and PDI, respectively, present in ERderived vesicles contaminating the preparation of [35 S]sulfate-labeled TGN vesicles. We investigated this possibility by immunoblotting using antibodies (Vaux et al ., 1990) raised against synthetic peptides corresponding to the carboxy-terminal sequences of BiP and PDI . Fig . 5 (left panels) shows that the bands of 78 kD (see lowerdot in Fig . 1, right) and 54 kD (see lowertriangle in Fig . 1, right) released from vesicles upon incubation with saponin were indeed BiP and PDI, respectively. In addition, the anti-BiP antibody also detected a 99-kD protein (upper dot in Fig . 1, right and Fig . 5, top left) , and the anti-PDI antibody also detected a 60-kD protein (upper triangle in Fig . 1, right and Fig . 5, bottom left) ; both proteins are acidic ER-resident proteins containing a carboxy-terminal 151 3 -KDEL sequence (S. D. Fuller, manuscript in preparation) . Although the release of these four ERresident proteins was incomplete and varied between the individual proteins, it nevertheless showed that ER-derived vesicles, despite their lower concentration of cholesterol, were at least in part permeabilized with saponin .