Conserved amino acids of the human immunodeficiency virus type 2 Vpx nuclear localization signal are critical for nuclear targeting of the viral preintegration complex in non-dividing cells

Abstract

The HIV-2 viral accessory protein Vpx is related to, but distinct from the Vpr protein of HIV-1. Vpx is packaged into virions and as a component of the viral preintegration complex (PIC) is required for efficient virus replication in non-dividing cells. We have previously reported that the minimal transferable region of Vpx that contained karyophilic properties was aa 65 to 72. Analysis of Vpx sequences from various HIV-2/SIV strains reveals that this region contains highly conserved amino acids, including two basic residues (K68, R70) and three tyrosines (Y66, Y69, Y71). Here, we demonstrate that mutation of the basic or tyrosine residues abolishes PIC nuclear import in arrested cells as assessed by PCR detection of viral integration. Examination of cell-free virus by Western blot indicated that all mutant proteins were incorporated into virions, suggesting that the lack of replication in arrested cells was not due to a loss of Vpx in target cells. Together, these studies map critical residues of the Vpx nuclear localization signal that are required for efficient infection of non-dividing cells.

Hsp40 Facilitates Nuclear Import of the Human Immunodeficiency Virus Type 2 Vpx-Mediated Preintegration Complex

“…DNA was then isolated with the DNeasy tissue kit (Qiagen). Integrated proviral DNA was detected by a nested PCR as described previously (2,4,21). The first-round PCR was carried out for 12 cycles with the primer 5Ј-ATGCC ACGTAAGCGAAACTGCGAGGCTGGCAGATTGAGCCCTG-3Ј(L-Rod) (the 5Ј underlined sequence is a linker and is followed by a sequence complementary to the long terminal repeat [LTR] of HIV-2) and two Alu primers (5Ј-TCCCAGCTACTGGGGAGGCTGAGG and 5Ј-GCCTCCCAAAGTGCT GGGATTACAG).…”

Characterization of Simian Immunodeficiency Virus SIV SM /Human Immunodeficiency Virus Type 2 Vpx Function in Human Myeloid Cells

“…To identify residues that are involved in the function of Vpx, alanine-scanning mutagenesis was carried out in the context of a FLAG-tagged SIV MAC Vpx fusion protein. Several mutations that were previously described to affect nuclear localization or to alter potential phosphorylation sites of Vpx were examined in this context, together with a deletion removing the C-terminal proline-rich tail of Vpx (⌬Pro) (amino acids 1 to 101) (4,26,32,35).…”

Human Immunodeficiency Virus Type 1 Vpr Links Proteasomal Degradation and Checkpoint Activation

“…This raises the question of whether or not the ability of Vpx to interact with DCAF1 is necessary for Vpx to promote infection of nondividing cells (7,13,17). And if so, is this function exerted through manipulation of the corresponding E3 ubiquitin ligase?…”