Formation of virus assembly intermediate complexes in the cytoplasm by wild-type and assembly-defective mutant human immunodeficiency virus type 1 and their association with membranes

Young Min Lee, Bindong Liu, Xiao Fang Yu

Research output: Contribution to journalArticle

Abstract

We have previously identified two distinct forms of putative viral assembly intermediate complexes, a detergent-resistant complex (DRC) and a detergent-sensitive complex (DSC), in human immunodeficiency virus type 1 (HIV-1)-infected CD4+ T cells (Y. M. Lee and X. F. Yu, Virology 243:78-93, 1998). In the present study, the intracellular localization of these two viral assembly intermediate complexes was investigated by use of a newly developed method of subcellular fractionation. In wild-type HIV-1-infected H9 cells, the DRC fractionated with the soluble cytoplasmic fraction, whereas the DSC was associated with the membrane fraction. The DRC was also detected in the cytoplasmic fraction in H9 cells expressing HIV-1 Myr-mutant Gag. However, little of the unmyristylated Gag and Gag-Pol proteins was found in the membrane fraction. Furthermore, HIV-1 Gag proteins synthesized in vitro in a rabbit reticulocyte lysate system in the absence of exogenous lipid membrane were able to assemble into a viral Gag complex similar to that of the DRC identified in infected H9 cells. The density of the viral Gag complex was not altered by treatment with the nonionic detergent Triton X-100, suggesting a lack of association of this complex with endogenous lipid. Formation of the DRC was not significantly affected by mutations in assembly domains M and L of the Gag protein but was drastically inhibited by a mutation in the assembly I domain. Purified DRC could be disrupted by high- salt treatment, suggesting electrostatic interactions are important for stabilizing the DRC. The Gag precursor proteins in the DRC were more sensitive to trypsin digestion than those in the DSC. These findings suggest that HIV-1 Gag and Gag-Pol precursors assemble into DRC in the cytoplasm, a process which requires the protein-protein interaction domain (I) in NCp7; subsequently, the DRC is transported to the plasma membrane through a process mediated by the M domain of the matrix protein. It appears that during this process, a conformational change might occur in the DRC either before or after its association with the plasma membrane, and this change is followed by the detection of virus budding structure at the plasma membrane.

Original languageEnglish (US)
Pages (from-to)5654-5662
Number of pages9
JournalJournal of Virology
Volume73
Issue number7
StatePublished - 1999

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Virus Assembly
Human immunodeficiency virus 1
detergents
Detergents
HIV-1
Cytoplasm
cytoplasm
mutants
Membranes
gag Gene Products
plasma membrane
Cell Membrane
virus assembly
proteins
Human Immunodeficiency Virus gag Gene Products
gag-pol Fusion Proteins
viral morphology
Protein Interaction Domains and Motifs
Virus Release
mutation

ASJC Scopus subject areas

  • Immunology

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Formation of virus assembly intermediate complexes in the cytoplasm by wild-type and assembly-defective mutant human immunodeficiency virus type 1 and their association with membranes. / Lee, Young Min; Liu, Bindong; Yu, Xiao Fang.

In: Journal of Virology, Vol. 73, No. 7, 1999, p. 5654-5662.

Research output: Contribution to journalArticle

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