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Abstract
ASSEMBLY AND MOVEMENT OF GAG/GAG-POL COMPLEXES IN HIV-INFECTED CELLS.
HALWANI R, KHORCHID A, WAINBERGE M, KLEIMAN L
We have detected the cytoplasmic interaction between Pr55gagand Pr160gag-polin HIV-1 using antibody to integrase (anti-IN) to coimmunoprecipitate both Pr55gagand Pr160gag-polfrom lysates of COS-7 and 293T cells transfected with wild-type and mutant HIV-1 proviral DNA. The interaction occurs independently of membrane attachment (non-myristylated precursors interact), but it is dependent upon the presence of the major homology region in the capsid sequence. The interaction is also dependent upon the presence of RNA; i.e., coimmiunoprecipitation is disrupted by RNase but not by DNase.
Using pulse-chase experiments coupled with cell fractionation and membrane floatation techniques, we followed the movement of this Gag/Gag-Pol complex from the cytoplasm to the plasma membrane, the site of viral assembly. In cells that were transfected with a plasmid expressing Gag-Pol alone, however, the Gag-Pol precursor remained in the cytoplasmic fractions and did not move to the membrane. This indicates the requirement of a Gag/Gag-Pol interaction in the cytoplasm for the translocation of Gag-Pol to the plasma membrane. Mutations in basic amino acids upsteam of the first Cys-His box in the HIV-1 nucleocapsid sequence (R7R10K11S) also inhibit this interaction but act only through Pr55gagand not through Pr160gag-pol.
Mutations in basic amino acids in the linker region between the two Cys-His boxes [S3(32—34)] do not inhibit the Pr55gag/ Pr160gag-polinteraction, even though both mutations inhibit genomic RNA packaging by 80%. These differences in the ability of the two types of NC mutations to inhibit Pr55gag/Pr160gag-polinteraction are reflected in similar differences in the ability of the mutant virus to produce new viral particles. These data suggest that the Pr55gag/Pr160gag-polinteraction is dependent on the non-specific interaction of Pr55gag and not Pr160gag-pol, with RNA in the cell, through the basic amino acid sequences upstream of the first Cys-His box in Pr55gagnucleocapsid.
The 1st. IAS Conference on HIV Pathogenesis and Treatment
Abstract no.
118
Suggested Citation
"HALWANIR, et al.
ASSEMBLY AND MOVEMENT OF GAG/GAG-POL COMPLEXES IN HIV-INFECTED CELLS..
Oral Presentation:
The 1st. IAS Conference on HIV Pathogenesis and Treatment
:
Abstract no.
118"
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