Proteins brefeldin blocking

Fig. 5. A scheme for transformation of mutant PrPs to a PrP state. Mutant PrPs are initially synthesized in the PrP state and acquire PrP properties in a stepwise fashion as they traverse different cellular compartments. PIPLC resistance, which develops in the ER, reflects folding of the polypeptide chain into the PrP conformation. Detergent insolubility and protease resistance, which develop on arrival at the plasma membrane or along an endocytic pathway, result from intermolecular aggregation ( maturation ). The times given underneath the boxes indicate when after pulse-labeling the corresponding property is detected. Addition of brefeldin A (BFA) to cells or incubation at 18°C, treatments that block movement of proteins beyond the Golgi apparatus, inhibit acquisition of detergent insolubility and protease resistance, but not PIPLC resistance. (Adapted with permission from Daude et al., 1997).

Our kinetic studies of mutant PrPs synthesized in CHO cells suggest that individual steps in formation of PrP may take place in at least two different cellular locations (Fig. 5). Because mutant PrPs become PIPLC-resistant within minutes of synthesis in pulse-labeling experiments, this early step must take place in the ER. Consistent with this conclusion, acquisition of PIPLC resistance is not affected by treatment of cells with brefeldin A or by incubation at 18°C, manipulations that block exit of proteins beyond the Golgi (Daude et al, 1997). In contrast, detergent insolubility and protease resistance, which do not develop until later times of chase, and are reduced by brefeldin A and 18°C incubation, are likely to be acquired after arrival of the protein at the cell surface, either on the plasma membrane itself or in endocytic compartments. Raft domains may be involved in these changes (unpublished data). 

---