Retention of the human immunodeficiency virus type 1 envelope glycoprotein in the endoplasmic reticulum does not redirect virus assembly from the plasma membrane.
The envelope glycoprotein (Env) of human immunodeficiency virus type 1 (HIV-1) has been shown to redirect the site of virus assembly in polarized epithelial cells. To test whether localization of the glycoprotein exclusively to the endoplasmic reticulum (ER) could redirect virus assembly to that organelle in nonpolarized cells, an ER -retrieval signal was engineered into an epitope-tagged variant of Env. The epitope tag, attached to the C terminus of Env, did not affect the normal maturation and transport of the glycoprotein or the incorporation of Env into virions. The epitope-tagged Env was also capable of mediating syncytium formation and virus entry with a similar efficiency to that of wild-type Env. When the epitope was modified to contain a consensus K(X)KXX ER retrieval signal, however, the glycoprotein was no longer proteolytically processed into its surface and transmembrane subunits and Env could not be detected at the cell surface by biotinylation. Endoglycosidase H analysis revealed that the modified Env was not transported to the Golgi apparatus. Immunofluorescent staining patterns were also consistent with the exclusion of Env from the Golgi. As expected, cells expressing the modified Env failed to form syncytia with CD4(+) permissive cells. Despite this tight localization of Env to the ER, when the modified Env was expressed in the context of virus, virions continued to be produced efficiently from the plasma membrane of transfected cells. However, these virions contained no detectable glycoprotein and were noninfectious. Electron microscopy revealed virus budding from the plasma membrane of these cells, but no virus was seen assembling at the ER membrane and no assembled virions were found within the cell. These results suggest that the accumulation of Env in an intracellular compartment is not sufficient to redirect the assembly of HIV Gag in nonpolarized cells.
Department of Microbiology, University of Alabama at Birmingham, Birmingham, Alabama 35294, USA.
K
K
Salzwedel
J T
JT
West
M J
MJ
Mulligan
E
E
Hunter
D000818:Animals; D001692:Biological Transport; D019556:COS Cells; D002460:Cell Line; D002461:Cell Line, Transformed; D002462:Cell Membrane / Q000378:metabolism; D002522:Chlorocebus aethiops; D010450:Endopeptidases / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D005110:Extracellular Space; D015752:Genes, env; D015726:Giant Cells; D015699:HIV Envelope Protein gp120 / Q000378:metabolism; D015497:HIV-1 / Q000235:genetics* / Q000502:physiology*; D006801:Humans; D011499:Protein Processing, Post-Translational; D021382:Protein Sorting Signals; D014771:Virion; D019065:Virus Assembly
D015699:HIV Envelope Protein gp120; D021382:Protein Sorting Signals; D010450:Endopeptidases