Herpes Simplex Virus Replication: Roles of Viral Proteins and Nucleoporins in Capsid-Nucleus Attachment

Abstract

Replication of herpes simplex virus type 1 (HSV-1) involves a step in which a parental capsid docks onto a host nuclear pore complex (NPC). The viral genome then translocates through the nuclear pore into the nucleoplasm, where it is transcribed and replicated to propagate infection. We investigated the roles of viral and cellular proteins in the process of capsid-nucleus attachment. Vero cells were preloaded with antibodies specific for proteins of interest and infected with HSV-1 containing a green fluorescent protein-labeled capsid, and capsids bound to the nuclear surface were quantified by fluorescence microscopy. Results showed that nuclear capsid attachment was attenuated by antibodies specific for the viral tegument protein VP1/2 (UL36 gene) but not by similar antibodies specific for UL37 (a tegument protein), the major capsid protein (VP5), or VP23 (a minor capsid protein). Similar studies with antibodies specific for nucleoporins demonstrated attenuation by antibodies specific for Nup358 but not Nup214. The role of nucleoporins was further investigated with the use of small interfering RNA (siRNA). Capsid attachment to the nucleus was attenuated in cells treated with siRNA specific for either Nup214 or Nup358 but not TPR. The results are interpreted to suggest that VP1/2 is involved in specific attachment to the NPC and/or in migration of capsids to the nuclear surface. Capsids are suggested to attach to the NPC by way of the complex of Nup358 and Nup214, with high-resolution immunofluorescence studies favoring binding to Nup358.

Recombinant Adeno-Associated Virus Utilizes Host Cell Nuclear Import Machinery To Enter the Nucleus

“…HSV-1 and adenovirus both directly bind to the cytoplasmic side of the NPC via nucleoporins (40,41,(68)(69)(70)(71). The adenoviral core protein protein VII has been shown to bind to several import proteins, including importin-␣, importin-␤, importin-7, and transportin, which are thought to mediate entry of the nucleoprotein through the NPC (39).…”

Cellular and viral determinants of retroviral nuclear entry

“…The transport receptor-cargo complex docks on NUPs, allowing passage across the NPC. Hepatitis B virus, herpes simplex virus, influenza virus, and adenovirus also use NUPs for nuclear entry (Trotman et al 2001;König et al 2008König et al , 2010Copeland et al 2009;Schmitz et al 2010), whereas poliovirus and cardiovirus induce modifications of NUPs (Gustin and Sarnow 2001;Porter and Palmenberg 2009). Previous LC-MS/MS studies revealed that KTN1, NUP43, NUP358, NUP45, and NUP54 expression is downregulated on HIV-1 infection while NUP358, NUP98, and TPR expression is upregulated (Chan et al 2007; Monette et al 2011).…”

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;). …”