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

For quantifying the amount the KDEL-R in the cis versus the trans side of the Golgi stack, we used the HeLa-SA:48 cells, either uninfected or after infection for 9 h with the WR strain of vaccinia virus. In both cases, the cells were treated with SLO before fixation (as above), cryosectioned, and double labeled with anti-KDEL-R and a rabbit antibody against the P4D5 cytoplasmic domain epitope of the VSV-G protein. Golgi stacks cut perpendicularly to the cisternal membranes were systematically sampled and photographed at a primary magnification of 28,000. The labeling for the G protein (indicative of the sialyltransferase, a TGN marker) was used to distinguish the trans side of the stack from the cis. An arbitrary line was then drawn through the stack to split the stack into cis and trans sides. The amount of KDEL-R labeling associated with the two sides was then evaluated.