The long term objective is to understand the molecular basis of cytomegalovirus (CMV) replication and use that information to develop new ways to diagnose and treat CMV-related diseases of man. This will be accomplished by identifying and studying the synthesis, structure, and function of specific viral proteins that are essential for virus replication. In this application for renewed support (20th year) there are five specific aims intended to provide a better understanding of the assembly and structure of the CMV virion and its precursors. The rationale for studying virus structure and assembly is that most aspects of virus replication are directly or indirectly coupled to the assembly process; therefore, it ultimately represents a major and largely untapped source of new targets for antivirals. CMV has been selected as a model system because of its increasing medical relevance in association with AIDS, organ transplantation, sexually transmitted disease and birth defects; and because of the need to determine molecular similarities and differences between herpes group viruses in order to understand their biological differences. In this proposal, a major research emphasis is placed on a """"""""scaffolding""""""""-like protein, called the assembly protein precursor, which together with its genetically related proteinase, is intimately involved in the process of capsid formation and possibly DNA packaging. Experiments are outlined to determine the function of five newly discovered domains on this protein: two are nuclear localization signals, two mediate protein-protein interactions in the capsid, and one is a CKII-like phosphorylation site. A second major research focus will be on the protein product of the largest open reading frame in the viral genome. This protein is a virion tegument constituent and forms a high molecular weight complex with a 115-kDa protein in both cells and virions. Genetic, biochemical, and immunologic techniques will be used to determine the structure and function of these proteins and the complex that they form.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Virology Study Section (VR)
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Beisel, Christopher E
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Johns Hopkins University
Schools of Medicine
United States
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Tullman, Jennifer A; Harmon, Mary-Elizabeth; Delannoy, Michael et al. (2014) Recovery of an HMWP/hmwBP (pUL48/pUL47) complex from virions of human cytomegalovirus: subunit interactions, oligomer composition, and deubiquitylase activity. J Virol 88:8256-67
Fernandes, Steve M; Brignole, Edward J; Taori, Kanchan et al. (2011) Cytomegalovirus capsid protease: biological substrates are cleaved more efficiently by full-length enzyme (pUL80a) than by the catalytic domain (assemblin). J Virol 85:3526-34
Margulies, Barry J; Gibson, Wade (2007) The chemokine receptor homologue encoded by US27 of human cytomegalovirus is heavily glycosylated and is present in infected human foreskin fibroblasts and enveloped virus particles. Virus Res 123:57-71
Loveland, Amy N; Nguyen, Nang L; Brignole, Edward J et al. (2007) The amino-conserved domain of human cytomegalovirus UL80a proteins is required for key interactions during early stages of capsid formation and virus production. J Virol 81:620-8
Wang, Jianlei; Loveland, Amy N; Kattenhorn, Lisa M et al. (2006) High-molecular-weight protein (pUL48) of human cytomegalovirus is a competent deubiquitinating protease: mutant viruses altered in its active-site cysteine or histidine are viable. J Virol 80:6003-12
McCartney, Stephen A; Brignole, Edward J; Kolegraff, Keli N et al. (2005) Chemical rescue of I-site cleavage in living cells and in vitro discriminates between the cytomegalovirus protease, assemblin, and its precursor, pUL80a. J Biol Chem 280:33206-12
Loveland, Amy N; Chan, Chee-Kai; Brignole, Edward J et al. (2005) Cleavage of human cytomegalovirus protease pUL80a at internal and cryptic sites is not essential but enhances infectivity. J Virol 79:12961-8
Chan, Chee-Kai; Brignole, Edward J; Gibson, Wade (2002) Cytomegalovirus assemblin (pUL80a): cleavage at internal site not essential for virus growth; proteinase absent from virions. J Virol 76:8667-74
Baxter, M K; Gibson, W (2001) Cytomegalovirus basic phosphoprotein (pUL32) binds to capsids in vitro through its amino one-third. J Virol 75:6865-73
Plafker, S M; Woods, A S; Gibson, W (1999) Phosphorylation of simian cytomegalovirus assembly protein precursor (pAPNG.5) and proteinase precursor (pAPNG1): multiple attachment sites identified, including two adjacent serines in a casein kinase II consensus sequence. J Virol 73:9053-62

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