This application describes a collaborative project between the laboratories of Linda Gooding and William Wold on the relationship between tumor necrosis factor (TNF) and the E3 transcription unit of adenovirus. TNF is secreted by macrophages in response to inflammatory stimuli. It has diverse biological functions which include regulation of cells involved in immunity and inflammation, cytolysis of certain tumor cells, and inhibition of viral replication. We have found that TNF lyses cells infected by human group C adenoviruses with deletions in early region E3. Uninfected cells and cells infected by wild type adenovirus are not lysed. These results indicate that adenovirus renders cells susceptible to TNF, and that a product of region E3 protects against lysis. We have mapped TNF resistance to a gene encoding a 14.7K protein in region E3 and have definitively identified this protein species. Significantly, 14.7K shares sequence homology with interferon B2 (IFN B2). IFN B2 is induced in cells treated with TNF and its presence correlates with cellular resistance to lysis by TNF. We postulate that the 14.7K protein evolved to protect virus infected cells from TNF by mimicking IFN B2. Our major goals are to understand in molecular terms how the 14.7K protein protects against TNF, and how adenovirus induces susceptibility to lysis by TNF. This basic understanding eventually should help us to develop strategies for employing TNF as an anti-viral and an anti-cancer agent.
Our specific aims are to purify and characterize 14.7K; to map functional domains within the protein, to study the mechanism of 14.7K function, to determine the relevance of 14.7K expression to pathogenesis among adenovirus serotypes; and to map the adenovirus gene that induces cellular susceptibility to TNF.
These specific aims represent, in our view, an ideal merging of the expertise of two laboratories: that of L. Gooding in TNF and other areas of molecular and cellular immunology, and that of W. Wold in the molecular biology of adenovirus region E3.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA048219-02
Application #
3192295
Study Section
Virology Study Section (VR)
Project Start
1988-08-01
Project End
1992-06-30
Budget Start
1989-07-01
Budget End
1990-06-30
Support Year
2
Fiscal Year
1989
Total Cost
Indirect Cost
Name
Emory University
Department
Type
Schools of Medicine
DUNS #
042250712
City
Atlanta
State
GA
Country
United States
Zip Code
30322
Ranheim, T S; Shisler, J; Horton, T M et al. (1993) Characterization of mutants within the gene for the adenovirus E3 14.7-kilodalton protein which prevents cytolysis by tumor necrosis factor. J Virol 67:2159-67
White, E; Sabbatini, P; Debbas, M et al. (1992) The 19-kilodalton adenovirus E1B transforming protein inhibits programmed cell death and prevents cytolysis by tumor necrosis factor alpha. Mol Cell Biol 12:2570-80
Krajcsi, P; Tollefson, A E; Anderson, C W et al. (1992) The adenovirus E3 14.5-kilodalton protein, which is required for down-regulation of the epidermal growth factor receptor and prevention of tumor necrosis factor cytolysis, is an integral membrane protein oriented with its C terminus in the cytoplasm. J Virol 66:1665-73
Krajcsi, P; Wold, W S (1992) The adenovirus E3-14.5K protein which is required for prevention of TNF cytolysis and for down-regulation of the EGF receptor contains phosphoserine. Virology 187:492-8
Gooding, L R; Aquino, L; Duerksen-Hughes, P J et al. (1991) The E1B 19,000-molecular-weight protein of group C adenoviruses prevents tumor necrosis factor cytolysis of human cells but not of mouse cells. J Virol 65:3083-94
Gooding, L R; Ranheim, T S; Tollefson, A E et al. (1991) The 10,400- and 14,500-dalton proteins encoded by region E3 of adenovirus function together to protect many but not all mouse cell lines against lysis by tumor necrosis factor. J Virol 65:4114-23
Duerksen-Hughes, P J; Hermiston, T W; Wold, W S et al. (1991) The amino-terminal portion of CD1 of the adenovirus E1A proteins is required to induce susceptibility to tumor necrosis factor cytolysis in adenovirus-infected mouse cells. J Virol 65:1236-44
Horton, T M; Ranheim, T S; Aquino, L et al. (1991) Adenovirus E3 14.7K protein functions in the absence of other adenovirus proteins to protect transfected cells from tumor necrosis factor cytolysis. J Virol 65:2629-39
Boss, J M; Laster, S M; Gooding, L R (1991) Sensitivity to tumour necrosis factor-mediated cytolysis is unrelated to manganous superoxide dismutase messenger RNA levels among transformed mouse fibroblasts. Immunology 73:309-15
Wilson-Rawls, J; Saha, S K; Krajcsi, P et al. (1990) A 6700 MW membrane protein is encoded by region E3 of adenovirus type 2. Virology 178:204-12

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