The UL14 Tegument Protein of Herpes Simplex Virus Type 1 Is Required for Efficient Nuclear Transport of the Alpha Transinducing Factor VP16 and Viral Capsids

Abstract

The protein encoded by the UL14 gene of herpes simplex virus type 1 (HSV-1) and HSV-2 is expressed late in infection and is a minor component of the virion tegument. An UL14-deficient HSV-1 mutant (UL14D) forms small plaques and exhibits an extended growth cycle at low multiplicities of infection (MOI) compared to wild-type virus. Although UL14 is likely to be involved in the process of viral maturation and egress, its precise role in viral replication is still enigmatic. In this study, we found that immediate-early viral mRNA expression was decreased in UL14D-infected cells. Transient coexpression of UL14 and VP16 in the absence of infection stimulated the nuclear accumulation of both proteins. We intended to visualize the fate of VP16 released from the infected virion and constructed UL14-null (14D-VP16G) and rescued (14R-VP16G) viruses that expressed a VP16-green fluorescent protein (GFP) fusion protein. Synchronous high-multiplicity infection of the viruses was performed at 4°C in the absence of de novo protein synthesis. We found that the presence of UL14 in the virion had an enhancing effect on the nuclear accumulation of VP16-GFP. The lack of UL14 did not significantly alter virus internalization but affected incoming capsid transport to the nuclear pore. These observations suggested that UL14 (i) enhanced VP16 nuclear localization at the immediately early phase, thus indirectly regulating the expression of immediate-early genes, and (ii) was associated with efficient nuclear targeting of capsids. The tegument protein UL14 could be part of the machinery that regulates HSV-1 replication.

Herpes simplex virus induces extensive modification and dynamic relocalisation of the nuclear mitotic apparatus (NuMA) protein in interphase cells

“…14R-VP16G is a wild-type HSV-1 virus that encodes VP16, the major tegument protein, fused to GFP and replicates normally in cultured HEp-2 cells (Yamauchi et al, 2008). HEp-2 cells were infected with 14R-VP16G at a MOI of 1 PFU/cell and fixed 8 hours post infection.…”

“…The HSV-1 wild-type virus F and F-VP26/GFP were kind gifts from Bernard Roizman (University of Chicago, IL) and David Johnson (Oregon Health & Sciences University, Portland, OR), respectively. The VP16-GFP expressing HSV-1 wild-type virus, 14R-VP16G, (Yamauchi et al, 2008) and HSV-2 wild-type virus 186 were also used. Viral stocks were propagated and titrated on Vero cell monolayers.…”

“…All procedures were performed as described previously (Yamauchi et al, 2008). Briefly, cells grown on coverslips were washed in PBS and then fixed in -20°C methanol for 10 minutes.…”

DNA-tumor virus entry—From plasma membrane to the nucleus

“…Interestingly, the 32 kDa tegument protein UL14 has been implicated in enhancing nuclear import of UL48 (VP16), and it enhanced nuclear targeting of incoming HSV1 (154). In the nucleus, it forms a regulatory complex with HCF1 and the POU-domain transcription factor Oct1, and thereby activates transcription of immediate early viral genes (155).…”

Virus entry at a glance

“…Many associate with microtubule-based motors, such as dynein and dynactin, and move along microtubules towards the nucleus (Bremner et al, 2009;Dodding and Way, 2011;Döhner et al, 2002;Engelke et al, 2011;Leopold et al, 2000;Radtke et al, 2010;Sodeik et al, 1997;Suomalainen et al, 1999;Yamauchi et al, 2008). For transport into the nucleus, they bind nuclear import receptors, such as importins, by using them to target their capsids or genome to nuclear pore complexes (NPCs) (Darshan et al, 2004;Klucevsek et al, 2006;Nakanishi et al, 2002;Schmitz et al, 2010;Wodrich et al, 2006) (reviewed by Puntener and Greber, 2009).…”