Human cytomegalovirus (HCMV) is a ubiquitous, normally benign virus that establishes lifelong persistence and/or latency. However, infections in immunodeficient or immunosuppressed patients often cause severe disease including pneumonia, gastrointestinal syndromes, hepatitis, retinitis, and encephalitis. Importantly, HCMV frequently contributes to the rejection of transplanted organs. In immunologically naive children, HCMV can cause developmental defects of the CNS, especially deafness. Moreover, clinically recognized cases of HCMV in young children may represent only the "tip of the iceberg", many other infants with less well pronounced HCMV in utero may suffer more subtle developmental defects. HCMV causes a diverse spectrum of diseases, in part, related to a capacity to infect a broad array of cell types including epithelial, endothelial and glial cells, fibroblasts, monocyte-macrophages (M/M) and neurons. In order to infect all these different cell types, HCMV must bind onto cell surfaces and enter the cells. This fundamentally important first step in HCMV replication play an important role in virus tropism and pathogenesis, yet is poorly understood. In part, gaps in understanding HCMV entry, relate to the fact that extensively studied laboratory strains of HCMV fail to enter M/M, epithelial and endothelial cells. Clinical strains of HCMV enter all these diverse cell types, apparently because these viruses express two entry glycoproteins: gH/gL/UL128-131, required for entry into epithelial and endothelial cells and gH/gL/gO, required for entry into fibroblasts. Lab strains of HCMV (propagated long term on fibroblasts) express only gH/gL/gO. When we transduced epithelial cells to cause expression of gH/gL/UL128-131, HCMV entry into the cells was blocked. This "interference" with virus entry provides principal support for our hypothesis that gH/gL/UL128-131 functions to bind epithelial cell receptors. We are testing the related hypothesis that gH/gL/gO acts to bind fibroblast receptors. We also found that expression of gH/gL (without UL128-131) and a second HCMV glycoprotein, gB, caused fusion of epithelial cells, surprisingly, even when expressed in trans, i.e. gB in one set of cells and gH/gL in other cells. These observations have important conceptual implications for models of how viral proteins fuse membranes. Our ongoing research is focused on understanding how the two HCMV glycoproteins gH/gL/UL128-131 and gH/gL/gO are assembled and incorporated into virus particles, and how these proteins function in virus entry. Moreover, we will study the molecular mechanisms of gB and gH/gL-mediated membrane fusion. Other efforts will focus on the identification of novel cellular receptors that will explain clinical HCMV entry into epithelial and endothelial cells. Information about viral and cellular entry mediators is key to a better understanding of HCMV pathogenesis and for the design of vaccines and anti-virals.

Public Health Relevance

Human cytomegalovirus (HCMV) causes substantial disease in several vulnerable populations: developing children, transplant patients, and AIDS patients. HCMV can infect a very broad spectrum of human cell types including retinal and gut epithelial cells, endothelial cells, hepatocytes, monocyte-macrophages and glial cells. Our studies will provide a better picture of how HCMV gains entry into biologically relevant epithelial and endothelial cells to facilitate the design of better anti-HCMV drugs and vaccines.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI081517-17
Application #
8313972
Study Section
Special Emphasis Panel (ZRG1-IDM-P (02))
Program Officer
Beisel, Christopher E
Project Start
2009-09-07
Project End
2014-08-31
Budget Start
2012-09-01
Budget End
2013-08-31
Support Year
17
Fiscal Year
2012
Total Cost
$436,883
Indirect Cost
$153,193
Name
Oregon Health and Science University
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
096997515
City
Portland
State
OR
Country
United States
Zip Code
97239
Wille, Paul T; Wisner, Todd W; Ryckman, Brent et al. (2013) Human cytomegalovirus (HCMV) glycoprotein gB promotes virus entry in trans acting as the viral fusion protein rather than as a receptor-binding protein. MBio 4:e00332-13
Sharma, Sapna; Wisner, Todd W; Johnson, David C et al. (2013) HCMV gB shares structural and functional properties with gB proteins from other herpesviruses. Virology 435:239-49
Vanarsdall, Adam L; Johnson, David C (2012) Human cytomegalovirus entry into cells. Curr Opin Virol 2:37-42
Johnson, David C; Baines, Joel D (2011) Herpesviruses remodel host membranes for virus egress. Nat Rev Microbiol 9:382-94
Vanarsdall, Adam L; Chase, Marie C; Johnson, David C (2011) Human cytomegalovirus glycoprotein gO complexes with gH/gL, promoting interference with viral entry into human fibroblasts but not entry into epithelial cells. J Virol 85:11638-45
Wille, Paul T; Knoche, Amber J; Nelson, Jay A et al. (2010) A human cytomegalovirus gO-null mutant fails to incorporate gH/gL into the virion envelope and is unable to enter fibroblasts and epithelial and endothelial cells. J Virol 84:2585-96
Ryckman, Brent J; Chase, Marie C; Johnson, David C (2010) Human cytomegalovirus TR strain glycoprotein O acts as a chaperone promoting gH/gL incorporation into virions but is not present in virions. J Virol 84:2597-609