Localization of the Lys, Asp, Glu, Leu tetrapeptide receptor to the Golgi complex and the intermediate compartment in mammalian cells

We have recently demonstrated that the COOH-terminal 21-residue sequence of the KDEL-R (the peptide epitope) is on the cytoplasmic side of the membrane (Singh et al., 1993) and that some cytosolic factor(s) may specifically mask the epitope (Tang et al., 1994) . These observations suggest that some KDEL-R epitopes might not be accessible to the antibody because of masking by either cytosolic factor(s) or, in some structures, because of the high packing of membrane (e.g., the rough ER system in the PAC). To overcome these potential problems, we used an alternative approach, described recently (Krijnse-Locker et al., 1994) , to investigate the subcellular localization of the KDEL-R. Mouse L cells, both uninfected as well as after infection with MHV for 6 h, were permeabilized with SLO to remove the bulk of the cytosol and the putative soluble masking factor(s). The ceils were then fixed and thawed cryosections were processed for immunogold labeling. This approach not only facilitates the visualization of detailed membrane structures (Krijnse-Locker et al., 1994) , but it also allows us to identify the intermediate compartment, marked by the budding profiles of MHV. There was variable labeling throughout the Golgi stack (Fig. 3, D and E) . In most cases, essentially the entire Golgi stack was heavily labeled with gold particles (Fig. 3 E) . There was also a strong labeling of the IC/MHV budding compartment (Fig. 3, A-C) . A significant amount of gold particles were also detected in the rough ER (Fig. 3 , A and D) and the nuclear envelope ( Fig. 3 E) , although the extent of the labeling was clearly less than that seen over the IC and Golgi stack.