Both the UL31 and UL34 proteins are essential for envelopment of herpes simplex virus 1 nucleocapsids at the inner nuclear membrane (INM). Data imply that these proteins play multiple roles in the envelopment reaction. The proposed studies in Aim 1 will investigate how pUL31 and pUL34 are targeted to the INM. This information will be relevant to understanding two separate aspects of the envelopment reaction that comprise the rest of the application: 1) Disruption of the nuclear lamina to provide nucleocapsids access to budding sites in the INM and 2) Construction of budding sites at the INM.
For Aim 2, preliminary data support the hypothesis that pUL31 and/or pUL34 proteins are able to disrupt the nuclear lamina when expressed alone or together. The hypothesis is that the pUL31/pUL34 complex interferes with interactions required for nuclear lamina integrity, thereby locally depolymerizing the lamina to allow virions access to budding sites in the inner nuclear membrane. The role of lamina association and putative depolymerizing activities identified in transient expression assays will be tested for their effects on nuclear egress of virions.
For Aim 3, preliminary data support a pUL34-dependent recruitment of viral glycoprotein D (gD) to the INM and indicate that pUL34 binds immature gD, an INM membrane protein known to become incorporated into perinuclear virions. Because this protein interacts with gB and gH which are also perinuclear virion components, we hypothesize that the pUL34/gD interaction helps orchestrate the proteome of INM budding sites and perinuclear virions. To test this possibility we propose to identify the pUL34 binding site in gD, and generate recombinant viruses bearing mutations that preclude the pUL34/gD interaction. The hypothesis predicts that gD should not localize efficiently in the INM of cells infected with the viral mutant, and should not be present in perinuclear virions as assessed by immunogold electron microscopy. Finally, preliminary data indicate that pUL31 is required for INM structures that stain densely with OsO4 and represent preferential nucleocapsid budding sites. Thus, pUL31 may act similarly to matrix proteins of other viruses to help orchestrate budding sites at the INM by alteration of localized lipid composition. To test this possibility, thin layer chromatography will be used to compare the lipid composition of perinuclear virions to those of nuclear and Golgi membranes. The hypothesis predicts preferential recruitment of unsaturated fatty acids to budding sites and perinuclear virions. How this putative alteration of lipid composition contributes to protein recruitment to INM budding sites will also be tested.

Public Health Relevance

These studies are applicable to public health because the mechanisms of envelopment, and the proteins involved are conserved in all herpesviruses and essential for viral replication. Thus any drugs that preclude the function of these proteins could have broad activities against all herpesviruses.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI052341-10
Application #
8489098
Study Section
Virology - A Study Section (VIRA)
Program Officer
Challberg, Mark D
Project Start
2002-07-01
Project End
2014-06-30
Budget Start
2013-07-01
Budget End
2014-06-30
Support Year
10
Fiscal Year
2013
Total Cost
$348,914
Indirect Cost
$118,590
Name
Cornell University
Department
Microbiology/Immun/Virology
Type
Schools of Veterinary Medicine
DUNS #
872612445
City
Ithaca
State
NY
Country
United States
Zip Code
14850
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Roller, Richard J; Haugo, Alison C; Yang, Kui et al. (2014) The herpes simplex virus 1 UL51 gene product has cell type-specific functions in cell-to-cell spread. J Virol 88:4058-68
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Selvarajan Sigamani, Sundaresan; Zhao, Haiyan; Kamau, Yvonne N et al. (2013) The structure of the herpes simplex virus DNA-packaging terminase pUL15 nuclease domain suggests an evolutionary lineage among eukaryotic and prokaryotic viruses. J Virol 87:7140-8
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Yang, Kui; Wills, Elizabeth G; Baines, Joel D (2012) Release of the herpes simplex virus 1 protease by self cleavage is required for proper conformation of the portal vertex. Virology 429:63-73
Mbong, Ekaette F; Woodley, Lucille; Frost, Elizabeth et al. (2012) Deletion of UL21 causes a delay in the early stages of the herpes simplex virus 1 replication cycle. J Virol 86:7003-7

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