Perturbation of Host Nuclear Membrane Component RanBP2 Impairs the Nuclear Import of Human Immunodeficiency Virus -1 Preintegration Complex (DNA)

Abstract

HIV-1 is a RNA virus that requires an intermediate DNA phase via reverse transcription (RT) step in order to establish productive infection in the host cell. The nascent viral DNA synthesized via RT step and the preformed viral proteins are assembled into pre-integration complex (PIC) in the cell cytoplasm. To integrate the viral DNA into the host genome, the PIC must cross cell nuclear membrane through the nuclear pore complex (NPC). RanBP2, also known as Nup358, is a major component of the cytoplasmic filaments that emanates from the nuclear pore complex and has been implicated in various nucleo-cytoplasmic transport pathways including those for HIV Rev-protein. We sought to investigate the role of RanBP2 in HIV-1 replication. In our investigations, we found that RanBP2 depletion via RNAi resulted in profound inhibition of HIV-1 infection and played a pivotal role in the nuclear entry of HIV DNA. More precisely, there was a profound decline in 2-LTR DNA copies (marker for nuclear entry of HIV DNA) and an unchanged level of viral reverse transcription in RanBP2-ablated HIV-infected cells compared to RanBP3-depleted or non-specific siRNA controls. We further demonstrated that the function of Rev was unaffected in RanBP2-depleted latently HIV infected cells (reactivated). We also serendipitously found that RanBP2 depletion inhibited the global ectopic gene expression. In conclusion, RanBP2 is a host factor that is involved in the nuclear import of HIV-1 PIC (DNA), but is not critical to the nuclear export of the viral mRNAs or nucleo-cytoplasmic shuttling of Rev. RanBP2 could be a potential target for efficient inhibition of HIV.

HIV-1 Resistance to the Capsid-Targeting Inhibitor PF74 Results in Altered Dependence on Host Factors Required for Virus Nuclear Entry

“…Infection by HIV-1 is dependent on the cellular factors Nup153, RanBP2, and TNPO3, which promote HIV-1 nuclear entry and/or integration (19,46). HIV-1 dependence on TNPO3 and Nup153 is altered by substitutions in the PF74 binding site (13,40,47), suggesting that the compound competes with host factor binding.…”

Cellular and viral determinants of retroviral nuclear entry

“…Several genome-wide RNA interference screening experiments identified numerous NUPs as potential nuclear transport factors for the Retroviridae family, including NUP358/RANBP2, NUP153, NUP98, and NUP214/ CAN (NUP 50,62,85,107,133,155,160,210, ELYS, and TRP) (Ebina et al 2004;Brass et al 2008;König et al 2008;Woodward et al 2009;Lee et al 2010;Zhang et al 2010;Engelman 2011, 2013a;Di Nunzio et al 2013;Matreyek et al 2013b). Among them, NUP358 and NUP214 are located exclusively at the cytoplasmic face.…”

“…Regulator of expression of the virion (Rev), one of the HIV-1 regulatory proteins, harbors both an NLS and NES and contributes to the export of Rev responsive elementcontaining spliced or unspliced mRNA (Cullen 2003;Strebel 2003;Zhang et al 2010). Interestingly, Rev can bind with IN and, as proposed in a recent model, might balance the movement of IN between the cytoplasm and nucleus (Levin et al 2010b).…”

“…Overexpression of Rev or Rev-IN complexes inhibits nuclear translocation of IN, while deletion mutants of Rev hamper nuclear import of IN (Levin et al 2010b), thereby suggesting the direct involvement of Rev in IN movement. Additionally, Rev relies on NUP BP2 for its own nuclear entry (Zhang et al 2010).…”

RANBP2 (RAN binding protein 2)

“…In summary, RANBP2 aids nuclear import by at least two mechanisms: i) by "capturing" transport receptors through the FG-repeats, it conveys them towards the NPC and reduces the effective concentration of import receptors required for efficient transport, while ii) by interacting with selected cargos in a receptor-independent manner, through the RANBP2 N-ter domain, it increases the overall efficiency of nuclear import. Interestingly, RANBP2 is also implicated in the nuclear delivery and integration of certain human viruses, including Herpes simplex (Copeland et al, 2009) and immunodeficiency virus-1 (HIV-1) (Zhang et al, 2010;Ocwieja et al, 2011;). …”