HIV-1 Vpr Abrogates the Effect of TSG101 Overexpression to Support Virus Release

Abstract

HIV-1 budding requires interaction between Gag and cellular TSG101 to initiate viral particle assembly and release via the endosomal sorting complexes required for transport (ESCRT) pathway. However, some reports show that overexpression of TSG101 inhibits virus release by disruption of Gag targeting process. Since a HIV-1 accessory protein, Vpr binds to Gag p6 domain at the position close to the binding site for TSG101, whether Vpr implicates TSG101 overexpression effect has not been investigated. Here, we found that Vpr abrogates TSG101 overexpression effect to rescue viral production. Co-transfection of TSG101 and Gag with Vpr prevented TSG101-induced Gag accumulation in endosomes and lysosomes. In addition, Vpr rescued virus-like particle (VLP) production in a similar manner as a lysosomal inhibitor, Bafilomycin A1 indicating that Vpr inhibits TSG101-induced Gag downregulation via lysosomal pathway. Vpr and Gag interaction is required to counteract TSG101 overexpression effect since Vpr A30F mutant which is unable to interact with Gag and incorporate into virions, reduced ability to prevent Gag accumulation and to rescue VLP production. In addition, GST pull-down assays and Biacore analysis revealed that Vpr competed with TSG101 for Gag binding. These results indicate that Vpr overcomes the effects of TSG101 overexpression to support viral production by competing with TSG101 to bind Gag.

Vpr Co-assembles with Gag during HIV-1 Assembly

“…The delay of assembly of Vpr compared to Gag suggests that Vpr might be recruited to Gag only after Gag is at the membrane rather than Vpr being attached to Gag before arriving at to the membrane. It has been shown previously that Vpr and Tsg101 can competitively bind to Gag since their recruitment sites are close together (41). The competition with Tsg101 may also contribute to the delay of Vpr assembly compared to Gag.…”

Huntingtin-Interacting Protein 1 Promotes Vpr-Induced G2 Arrest and HIV-1 Infection in Macrophages

“…Vpr is an accessory gene product of the human immunodeficiency virus type 1 (HIV-1) and a small 15-kDa protein with multiple biological functions, including splicing regulation [1][2][3], support of virus release [4], nuclear import of the viral preintegration complex in macrophages [5][6][7], enhanced expression and processing of the envelope glycoprotein in macrophages [8][9][10], sustaining interleukin 6 expression to enhance HIV-1 replication [11], antagonism of exonuclease 1-and helicase-like transcription factor-mediated restriction in T cells through degradation of these proteins [12][13][14][15], regulation of apoptosis in both a positive and negative manner, and the induction of cell cycle arrest at the G2 phase in dividing cells [16][17][18][19][20][21]. Multiple functions of Vpr are exerted through interactions with various host factors, such as DNA damage-binding protein 1 (DDB1)-and cullin 4 (CUL4)-associated factor 1 (DCAF1), spliceosome-associated protein 145, p300, synthetic lethal of unknown (X) function 4 (SLX4), protein arginine N-methyltransferase 5, importin α, mini-chromosome maintenance protein10, and coiled-coil domain-containing-137 (CCDC137) [2,6,7,13,20,[22][23][24][25][26][27].…”