With regard to oncology, it is critical to know how the same viral gene performs a transforming function in one cell species and yet a regulatory function during infection in another. This has been shown to be true for the EIA gene of adenovirus. The long-term objective is to understand how expression of adenovirus genes is regulated in cells infected and transformed by the virus. An immediate goal is to learn how the large acidic protein of the EIA gene activates expression of early viral gene transcription. Two approaches will be taken. The first is to study the biochemical features of the EIA protein and the second is to construct and analyze specific EIA viral mutants. The objective of these combined approaches is to define the functional properties and domains of the EIA protein required for modulation of transcription and transformation.
The specific aims will be to purify EIA proteins using our recently produced monospecific antibodies. Both purified EIA proteins and EIA antibodies will be used to determine whether EIA proteins: bind to DNA; associate with other cellular proteins; enhance in vitro transcription; as well as to determine: the size of native EIA proteins; the steady state levels of EIA proteins throughout infection; and the identity of those amino acids of EIA which are phosphorylated.
We aim to produce more EIA monoclonal antibodies to be used in these assays. Secondly, we intend to further resolve the functions of EIA by genetic analysis. The mutation within the EIA DNA of the putatively missense Ad5 host range mutants have been functionally located by analysis of wild-type/host range EIA hybrid genes; the defective region of DNA will now be sequenced. We are furthermore constructing specific EIA point mutations in Ad5 and Ad12 by oligonucleotide-directed mutagenesis. In addition, we will pursue our most recent finding that expression of class I major histocompatibility (MHC) antigens is abolished on the surface of Ad12 transformed mouse cells (highly oncogenic) but not Ad5 transformed cells (weakly oncogenic) and that only Ad12 transformed cells resist being killed in a cell-mediated cytotoxicity assay. We will investigate why H-2 class I surface antigens fail to be expressed in Ad12 transformed cells and study whether the EIA or the EIB genes of Ad12 may independently achieve MHC restriction in mouse cells.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA029797-06
Application #
3168876
Study Section
Experimental Virology Study Section (EVR)
Project Start
1981-04-01
Project End
1987-11-30
Budget Start
1986-12-01
Budget End
1987-11-30
Support Year
6
Fiscal Year
1987
Total Cost
Indirect Cost
Name
Wistar Institute
Department
Type
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Guan, Hancheng; Ricciardi, Robert P (2012) Transformation by E1A oncoprotein involves ubiquitin-mediated proteolysis of the neuronal and tumor repressor REST in the nucleus. J Virol 86:5594-602
Heyward, Christa Y; Patel, Rajen; Mace, Emily M et al. (2012) Tumorigenic adenovirus 12 cells evade NK cell lysis by reducing the expression of NKG2D ligands. Immunol Lett 144:16-23
Jiao, Junfang; Guan, Hancheng; Lippa, Andrew M et al. (2010) The N terminus of adenovirus type 12 E1A inhibits major histocompatibility complex class I expression by preventing phosphorylation of NF-kappaB p65 Ser276 through direct binding. J Virol 84:7668-74
Guan, Hancheng; Williams, Jim F; Ricciardi, Robert P (2009) Induction of neuronal and tumor-related genes by adenovirus type 12 E1A. J Virol 83:651-61
Guan, Hancheng; Jiao, Junfang; Ricciardi, Robert P (2008) Tumorigenic adenovirus type 12 E1A inhibits phosphorylation of NF-kappaB by PKAc, causing loss of DNA binding and transactivation. J Virol 82:40-8
Guan, Hancheng; Hou, Shihe; Ricciardi, Robert P (2005) DNA binding of repressor nuclear factor-kappaB p50/p50 depends on phosphorylation of Ser337 by the protein kinase A catalytic subunit. J Biol Chem 280:9957-62
Williams, J F; Zhang, Y; Williams, M A et al. (2004) E1A-based determinants of oncogenicity in human adenovirus groups A and C. Curr Top Microbiol Immunol 273:245-88
Guan, Hancheng; Smirnov, Denis A; Ricciardi, Robert P (2003) Identification of genes associated with adenovirus 12 tumorigenesis by microarray. Virology 309:114-24
Zhao, Biwei; Hou, Shihe; Ricciardi, Robert P (2003) Chromatin repression by COUP-TFII and HDAC dominates activation by NF-kappaB in regulating major histocompatibility complex class I transcription in adenovirus tumorigenic cells. Virology 306:68-76
Hou, Shihe; Guan, Hancheng; Ricciardi, Robert P (2003) Phosphorylation of serine 337 of NF-kappaB p50 is critical for DNA binding. J Biol Chem 278:45994-8

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