Summary auto-generated
This study investigated how intracellular retention affects HIV-1 glycoprotein processing and virus particle assembly. Researchers engineered HIV-1 glycoproteins with endoplasmic reticulum (ER) or Golgi retention signals, creating mutants called Env-TM/Golgi and Env856ER. These modified glycoproteins remained trapped in intracellular compartments and were not proteolytically processed to their mature gp120 and gp41 forms, indicating that cleavage occurs downstream during intracellular transport, likely in or near the trans-Golgi network. Despite the glycoproteins being sequestered intracellularly, virus particles were still released from cells in equivalent amounts compared to wild-type, and electron microscopy confirmed that assembly and budding occurred exclusively at the plasma membrane. However, the retained glycoproteins were not incorporated into virions, and released particles were completely non-infectious. The findings demonstrate that while viral glycoprotein localization influences its processing and incorporation into particles, it does not determine the subcellular site of HIV-1 particle assembly in COS-7 cells.
Key findings
- Intracellular retention of HIV-1 glycoprotein prevents its proteolytic processing to gp120 and gp41, indicating cleavage occurs late in the transport pathway at or near the trans-Golgi network
- Retention of glycoproteins in the ER or Golgi compartments blocks their transport to the cell surface and prevents incorporation into virions
- HIV-1 particle assembly and budding remain exclusively at the plasma membrane regardless of glycoprotein subcellular localization, indicating glycoprotein location does not determine assembly site in COS-7 cells
- Particles released when glycoproteins are intracellularly retained are non-infectious despite normal amounts of structural proteins being released
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Abstract
In this study, specific signals known to mediate endoplasmic reticulum or Golgi localization of transmembrane proteins have been transferred to the human immunodeficiency virus type 1 (HIV-1) env gene product. The intracellularly retained recombinant glycoproteins were not proteolytically processed to gp120 and gp41, which is further evidence that this process occurs at a later stage in the transport pathway, presumably within or near the trans-Golgi network. Since the subcellular localization of the viral glycoproteins of enveloped viruses can be one of the factors determining the cellular site of particle assembly and release, experiments were performed to determine if this property was altered by coexpression of the recombinant HIV-1 glycoproteins. When wild-type virus was compared to mutant virus encoding the intracellularly retained glycoproteins, the extent of HIV- 1 particle release into the extracellular medium remained unaffected, and electron-microscopic analysis did not reveal any significant alteration in the cellular sites of particle assembly and budding. Thus, in COS-7 cells, altered subcellular localization of the viral glycoprotein does not exert a dominant influence on the assembly site of the HIV-1 particle.