The long term objectives of the proposed project are three-fold. First, new substitution, deletion, insertion, and point mutations in the human adenovirus specified DNA binding protein (DBP) will be used to provide additional supporting evidence or new refuting evidence for the myriad of purported functions of DBP (e.g., in early gene turn on and off, DNA replication, late gene turn on, viral assembly, and oncogenic transformation). Second, an extensive collection of DBP mutants, which is in the process of being made, will be utilized to obtain a better understanding of the interrelationships of the many DBP functions. Third, chemical, biochemical and cell biological studies will be continued in order to better understand the mechanism(s) DBP uses to carry out its diverse set of roles, particularly as they apply to gene expression. In the latter, special emphasis will be directed to (i) understanding DBP interaction with RNA, (ii) deciphering the mechanism(s) by which this protein kills cells and (iii) determining its role in early gene expression. To achieve the first objective, our complementing, DBP expressing human lines will be utilized to construct many new DBP mutants. Two new schemes for mutagenesis will be attempted. They should facilitate the rapid construction of large numbers of diverse mutants in other adenovirus genes as well as DBP.

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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Rutgers University
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New Brunswick
United States
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Ricigliano, J W; Brough, D E; Klessig, D F (1994) Identification of a high-molecular-weight cellular protein complex containing the adenovirus DNA binding protein. Virology 202:715-23
Cleghon, V; Piderit, A; Brough, D E et al. (1993) Phosphorylation of the adenovirus DNA-binding protein and epitope mapping of monoclonal antibodies against it. Virology 197:564-75
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Eagle, P A; Klessig, D F (1992) A zinc-binding motif located between amino acids 273 and 286 in the adenovirus DNA-binding protein is necessary for ssDNA binding. Virology 187:777-87
Brough, D E; Cleghon, V; Klessig, D F (1992) Construction, characterization, and utilization of cell lines which inducibly express the adenovirus DNA-binding protein. Virology 190:624-34
Cleghon, V; Klessig, D F (1992) Characterization of the nucleic acid binding region of adenovirus DNA binding protein by partial proteolysis and photochemical cross-linking. J Biol Chem 267:17872-81
Vos, H L; Brough, D E; Van der Lee, F M et al. (1989) Characterization of adenovirus type 5 insertion and deletion mutants encoding altered DNA binding proteins. Virology 172:634-42
Silverman, L; Cleghon, V; Klessig, D F (1989) Increased permissivity of monkey cells to human adenovirus multiplication is affected by culturing conditions and correlates with both synthesis of virion fiber protein and altered splicing of its mRNA. Virology 173:109-19
Silverman, L; Klessig, D F (1989) Characterization of the translational defect to fiber synthesis in monkey cells abortively infected with human adenovirus: role of ancillary leaders. J Virol 63:4376-85

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