Morphological analysis of the transfer of VSV ts-045 G glycoprotein from the endoplasmic reticulum to the intermediate compartment in vero cells.
Vero cells were infected with the ts-045 strain of vesicular stomatitis virus, and the cells were incubated at 39 degrees C to accumulate the mutant G glycoprotein in the ER as a misfolded aggregate. Cycloheximide was added to the culture medium 3.5 h after infection to prevent further protein synthesis, and the temperature was lowered to 10, 15, or 31 degrees C. At these temperatures, the mutant G glycoprotein correctly folds and oligomerizes. Immunofluorescence light microscopy showed that the G glycoprotein was exported to the Golgi complex at 31 degrees C and to the intermediate compartment (IC) at 15 degrees C, but no export was observed at 10 degrees C. However, incubations at 10 degrees C followed by shift to 15 or 31 degrees C resulted in the normal transfer of the glycoprotein to the IC and the Golgi, respectively. Immunoelectron microscopical analysis confirmed all these results, but showed also that the glycoprotein was frequently clustered in the ER at 10 degrees C. Conventional electron microscopy showed that the morphology of the ER, IC, and Golgi complex remained essentially unchanged at all temperatures. The only significant difference detectable in cells incubated at 10 degrees C was the increased number of partially coated ER protrusions, longer than those detected at higher temperatures. These results demonstrate that the transport toward the Golgi complex of G glycoprotein can be arrested at a step preceding the entry into the IC, thus suggesting that ER and IC are separate stations in the exocytic pathway.
Lotti
L V
LV
Dipartimento di Medicina Sperimentale, Università di Roma La Sapienza, Italy.
Torrisi
M R
MR
Erra
M C
MC
Bonatti
S
S
D000818:Animals; D001692:Biological Transport / Q000502:physiology; D002451:Cell Compartmentation / Q000502:physiology*; D002522:Chlorocebus aethiops; D004721:Endoplasmic Reticulum / Q000378:metabolism* / Q000648:ultrastructure; D005455:Fluorescent Antibody Technique; D006023:Glycoproteins / Q000378:metabolism; D006056:Golgi Apparatus / Q000378:metabolism / Q000648:ultrastructure; D008562:Membrane Glycoproteins; D016253:Microscopy, Immunoelectron; D013696:Temperature; D014709:Vero Cells; D014721:Vesicular stomatitis Indiana virus / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D014759:Viral Envelope Proteins / Q000378:metabolism*
C028391:G protein, vesicular stomatitis virus; D006023:Glycoproteins; D008562:Membrane Glycoproteins; D014759:Viral Envelope Proteins