Human cytomegalovirus (HCMV) infects up to 90% of the U.S. population, resulting in lifelong chronic infection. In immunosuppressed individuals, the outcome is more severe, with lifelong persistent infection associated with various pathologies. HCMV is also a leading viral cause of congenital birth defects. Several antiviral treatments exist, but their use is limited due to significant toxicity, teratogenic effects, poor oral availability and emergence of antiviral resistance. An HCMV vaccine has yet to be licensed. Thus there is a need to understand the mechanism of HCMV infection to establish new targets and approaches for vaccines and therapies. Persistent HCMV infection is complex, involving a balance between productive lytic replication and a silent latent infection involving genome maintenance without replication. The long-term goal is to elucidate mechanisms used by HCMV proteins to control replication and cellular stress responses that influence the fate of infection. Defining these molecular events will augment understanding of the HCMV infected cell and facilitate development of antiviral strategies to manage infection. The experiments presented herein will define the dynamic and antagonistic regulatory relationships between the HCMV kinase pUL97 and pUL27, and determine how this network controls HCMV viral DNA synthesis. HCMV pUL97 regulates pUL27, Tip60 acetyl-transferase and the downstream effector, p21Cip1. The hypothesis is that pUL97 kinase activates Tip60-dependent chromatin remodeling while also inhibiting the pUL27-mediated degradation of Tip60 and the corresponding p21Cip1 expression to promote viral DNA synthesis. This will be tested in three specific aims:
Aim 1 Test the hypothesis that pUL27 induces p21Cip1 and prolonged p21Cip1 expression inhibits factors involved in viral DNA synthesis;
Aim 2 Test the hypothesis that pUL97 activation of Tip60 promotes viral DNA synthesis in a manner that involves chromatin modifications;
and AIM 3 Test the hypothesis that the pUL27-Tip60-pUL97 axis regulates entry into and exit from HCMV latency. A comprehensive repertoire of cellular and molecular biological techniques, viral genetic analysis, transcriptomics, and quantitative ?bottom up? and ?top down? proteomics will be used to study both lytic and latent models of HCMV infection. These studies will define how pUL97 kinase promotes viral DNA synthesis and how the antagonistic activities of two HCMV proteins, pUL97 and pUL27, regulate Tip60 and p21Cip1 expression and control HCMV replication. Understanding this regulatory switch will elucidate the mechanism by which therapeutics may silence HCMV replication.

Public Health Relevance

The beta-herpesvirus human cytomegalovirus (HCMV) infects the majority of the world population causing life-threatening disease in immunosuppressed patients and congenital birth defects. Our goal is to uncover the mechanisms whereby HCMV proteins manipulate cellular processes to promote infection. Defining the landscape of molecular events (e.g. protein-protein interaction networks) that occur during infection will augment our understanding of the infected cell and facilitate the development of new antiviral strategies to manage HCMV infection.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI083281-07
Application #
9517647
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Beisel, Christopher E
Project Start
2011-07-01
Project End
2021-06-30
Budget Start
2018-07-01
Budget End
2019-06-30
Support Year
7
Fiscal Year
2018
Total Cost
Indirect Cost
Name
Medical College of Wisconsin
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
937639060
City
Milwaukee
State
WI
Country
United States
Zip Code
53226
Westdorp, Kristen N; Terhune, Scott S (2018) Impact of RNA polymerase I inhibitor CX-5461 on viral kinase-dependent and -independent cytomegalovirus replication. Antiviral Res 153:33-38
Rak, Michael A; Buehler, Jason; Zeltzer, Sebastian et al. (2018) Human Cytomegalovirus UL135 Interacts with Host Adaptor Proteins To Regulate Epidermal Growth Factor Receptor and Reactivation from Latency. J Virol 92:
Forte, Eleonora; Swaminathan, Suchitra; Schroeder, Mark W et al. (2018) Tumor Necrosis Factor Alpha Induces Reactivation of Human Cytomegalovirus Independently of Myeloid Cell Differentiation following Posttranscriptional Establishment of Latency. MBio 9:
Greseth, Matthew D; Carter, Dominique C; Terhune, Scott S et al. (2017) Proteomic Screen for Cellular Targets of the Vaccinia Virus F10 Protein Kinase Reveals that Phosphorylation of mDia Regulates Stress Fiber Formation. Mol Cell Proteomics 16:S124-S143
Gonyo, P; Bergom, C; Brandt, A C et al. (2017) SmgGDS is a transient nucleolar protein that protects cells from nucleolar stress and promotes the cell cycle by regulating DREAM complex gene expression. Oncogene 36:6873-6883
Westdorp, Kristen N; Sand, Andrea; Moorman, Nathaniel J et al. (2017) Cytomegalovirus Late Protein pUL31 Alters Pre-rRNA Expression and Nuclear Organization during Infection. J Virol 91:
Buehler, Jason; Zeltzer, Sebastian; Reitsma, Justin et al. (2016) Opposing Regulation of the EGF Receptor: A Molecular Switch Controlling Cytomegalovirus Latency and Replication. PLoS Pathog 12:e1005655
Bigley, Tarin M; McGivern, Jered V; Ebert, Allison D et al. (2016) Impact of a cytomegalovirus kinase inhibitor on infection and neuronal progenitor cell differentiation. Antiviral Res 129:67-73
Bigley, Tarin M; Reitsma, Justin M; Terhune, Scott S (2015) Antagonistic Relationship between Human Cytomegalovirus pUL27 and pUL97 Activities during Infection. J Virol 89:10230-46
Carter, Dominique M; Westdorp, Kristen; Noon, Kathleen R et al. (2015) Proteomic identification of nuclear processes manipulated by cytomegalovirus early during infection. Proteomics 15:1995-2005

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