Wild-Type Levels of Nuclear Localization and Human Immunodeficiency Virus Type 1 Replication in the Absence of the Central DNA Flap

Abstract

Numerous factors have been implicated in the nuclear localization of retroviral preintegration complexes. Whereas sequences in human immunodeficiency virus type 1 (HIV-1) matrix, Vpr, and integrase proteins were initially reported to function specifically in nondividing cells, other recently identified sequences apparently function in dividing cells as well. One of these, the central DNA flap formed during reverse transcription, is specific to lentiviruses. It was previously reported that flap-negative (F−) HIV-1LAI was completely defective for viral spread in the MT-4 T-cell line, yet F− HIV-1 vectors were only 2- to 10-fold defective in various single-round transduction assays. To address these different findings, we analyzed the infectivity and nuclear localization phenotypes of two highly related T-cell-tropic strains, HIV-1NL4-3 and a derivative of HIV-1HXBc2 deficient for both Vpr and Nef. In stark contrast to the previous report, F− derivatives of both strains replicated efficiently in MT-4 cells. F− HIV-1NL4-3 also spread like wild-type HIV-1NL4-3 in infected Jurkat and primary T-cell cultures. In contrast, F− HIV-1HXBc2 was replication defective in primary T cells. Results of real-time quantitative PCR assays, however, indicated that F− HIV-1HXBc2 entered primary T-cell nuclei as efficiently as its wild-type counterpart. Thus, the F− HIV-1HXBc2 growth defect did not appear to correlate with defective nuclear import. Consistent with this observation, wild-type nef restored replication to F− HIV-1HXBc2 in primary T cells. Our results indicate that the central DNA flap does not play a major role in either preintegration complex nuclear import or HIV-1 replication in a variety of cell types

Macrophages and HIV-1: dangerous liaisons

“…However, some studies have shown that cPPT mutant HIV-1 can replicate in non-dividing cells including primary macrophages, albeit less efficiently than the parental virus (Dvorin et al, 2002;Limon et al, 2002b). Viral proteins are also involved in the nuclear transport of HIV-1 PIC (Fouchier and Malim, 1999).…”

FIV: from lentivirus to lentivector

“…Studies with replicating virus and replicationdefective HIV-1 vectors have lent support to the hypothesis [104,123,124]. However, recent reassessments have challenged this functional assignment [125,126].…”

“…Two similar motifs have also been found to act as strong pause determinants for purified EIAV reverse transcriptase on in vitro templates [119,120]. Given the conflicting data about the function of the flap in HIV-1 [104,[123][124][125][126], it will be of interest to determine the functional role in FIV.…”

LEDGF/p75 Is Essential for Nuclear and Chromosomal Targeting of HIV-1 Integrase in Human Cells

“…The latter has been reported to be essential for nuclear import of HIV PICs and viral replication (9). Although the effect of the central DNA flap appears to be viral strain-and host cell-dependent (10,11), its insertion in HIV-derived lentiviral vectors clearly augments transduction efficiency (12) and nuclear import (13). The karyophilic properties of MA and its role in HIV nuclear import (14) are unclear (15)(16)(17).…”