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The C Terminus of the Herpes Simplex Virus UL25 Protein Is Required for Release of Viral Genomes from Capsids Bound to Nuclear Pores.

TitleThe C Terminus of the Herpes Simplex Virus UL25 Protein Is Required for Release of Viral Genomes from Capsids Bound to Nuclear Pores.
Publication TypeJournal Article
Year of Publication2017
AuthorsHuffman JB, Daniel GR, Falck-Pedersen E, Huet A, Smith GA, Conway JF, Homa FL
JournalJ Virol
Volume91
Issue15
Date Published2017 08 01
ISSN1098-5514
KeywordsAnimals, Capsid Proteins, Chlorocebus aethiops, DNA, Viral, Herpesvirus 1, Human, Microscopy, Mutant Proteins, Nuclear Pore, Sequence Deletion, Vero Cells, Virus Uncoating
Abstract

The herpes simplex virus (HSV) capsid is released into the cytoplasm after fusion of viral and host membranes, whereupon dynein-dependent trafficking along microtubules targets it to the nuclear envelope. Binding of the capsid to the nuclear pore complex (NPC) is mediated by the capsid protein pUL25 and the capsid-tethered tegument protein pUL36. Temperature-sensitive mutants in both pUL25 and pUL36 dock at the NPC but fail to release DNA. The uncoating reaction has been difficult to study due to the rapid release of the genome once the capsid interacts with the nuclear pore. In this study, we describe the isolation and characterization of a truncation mutant of pUL25. Live-cell imaging and immunofluorescence studies demonstrated that the mutant was not impaired in penetration of the host cell or in trafficking of the capsid to the nuclear membrane. However, expression of viral proteins was absent or significantly delayed in cells infected with the pUL25 mutant virus. Transmission electron microscopy revealed capsids accumulated at nuclear pores that retained the viral genome for at least 4 h postinfection. In addition, cryoelectron microscopy (cryo-EM) reconstructions of virion capsids did not detect any obvious differences in the location or structural organization for the pUL25 or pUL36 proteins on the pUL25 mutant capsids. Further, in contrast to wild-type virus, the antiviral response mediated by the viral DNA-sensing cyclic guanine adenine synthase (cGAS) was severely compromised for the pUL25 mutant. These results demonstrate that the pUL25 capsid protein has a critical role in releasing viral DNA from NPC-bound capsids. Herpes simplex virus 1 (HSV-1) is the causative agent of several pathologies ranging in severity from the common cold sore to life-threatening encephalitic infection. Early steps in infection include release of the capsid into the cytoplasm, docking of the capsid at a nuclear pore, and release of the viral genome into the nucleus. A key knowledge gap is how the capsid engages the NPC and what triggers release of the viral genome into the nucleus. Here we show that the C-terminal region of the HSV-1 pUL25 protein is required for releasing the viral genome from capsids docked at nuclear pores. The significance of our research is in identifying pUL25 as a key viral factor for genome uncoating. pUL25 is found at each of the capsid vertices as part of the capsid vertex-specific component and implicates the importance of this complex for NPC binding and genome release.

DOI10.1128/JVI.00641-17
Alternate JournalJ Virol
PubMed ID28490590
PubMed Central IDPMC5512264
Grant ListS10 OD019995 / OD / NIH HHS / United States
R21 AI132967 / AI / NIAID NIH HHS / United States
T32 GM008061 / GM / NIGMS NIH HHS / United States
R01 AI094050 / AI / NIAID NIH HHS / United States
R01 AI056346 / AI / NIAID NIH HHS / United States
R01 AI089803 / AI / NIAID NIH HHS / United States

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