We have been studying assembly of adenovirus with attention drawn to the polar encapsidation of the viral genome in vivo and the interactions of empty viral capsids with DNA molecules in vitro. Molecular cloning, sequence determination, and analysis of novel genome variants of adenovirus type 3 (Ad3) have delimited the left end DNA sequence recognized for polar encapsidation. Host range variants have enabled us to discern and exploit interference and cross-subgroup complementation in the expression of early region Ia, part of the adenovirus transformation/tumor antigen complex. We have further characterized the avid binding of empty adenovirus capsids with DNA in vitro, in particular with closed circular, superhelical DNA substrates. Most recently we have successfully used the capsid:DNA complexes as an efficient gene transfection system, demonstrating adsorption to a variety of cells in culture, expression of transfected genes, infectivity of transfected papovavirus DNA, and cellular transformation by a cloned thymidine kinase gene. One objective now proposed is to establish and transpose the DNA encapsidation sequence and evaluate the capacity for cross subgroup recognition of this site by the presumed viral proteins that play a role in DNA packaging. As part of this effort we want to evaluate the evolutionary constraints and divergence of the encapsidation site and the proximal (perhaps overlapping) sequences that regulate expression of early gene Ia. Methods of site directed mutagenesis and gene transfer that have been developed, some by this laboratory, will be used in this proposed investigation of the structure, functions and evolution of non-coding sequences near the left end of the adenovirus genome. This will be of immediate interest in terms of adenovirus biology, but also for the broader fields of gene expression and genetic modification of animal cells.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA034126-03
Application #
3171873
Study Section
Experimental Virology Study Section (EVR)
Project Start
1982-07-01
Project End
1986-12-31
Budget Start
1985-01-01
Budget End
1985-12-31
Support Year
3
Fiscal Year
1985
Total Cost
Indirect Cost
Name
Vanderbilt University Medical Center
Department
Type
Schools of Medicine
DUNS #
004413456
City
Nashville
State
TN
Country
United States
Zip Code
37203
Hamid, R; Cogan, J D; Jones, S N et al. (1995) Phenotypic determinants of adenovirus E1A gene autoregulation: variable region between conserved coding domains 2 and 3. Virology 213:666-70
Benamira, M; Johnson, K; Chaudhary, A et al. (1995) Induction of mutations by replication of malondialdehyde-modified M13 DNA in Escherichia coli: determination of the extent of DNA modification, genetic requirements for mutagenesis, and types of mutations induced. Carcinogenesis 16:93-9
Gruber, W C; Russell, D J; Tibbetts, C (1993) Fiber gene and genomic origin of human adenovirus type 4. Virology 196:603-11
Cogan, J D; Jones, S N; Hall, R K et al. (1992) Functional diversity of E1A gene autoregulation among human adenoviruses. J Virol 66:3833-45
Bowling, J M; Bruner, K L; Cmarik, J L et al. (1991) Neighboring nucleotide interactions during DNA sequencing gel electrophoresis. Nucleic Acids Res 19:3089-97
Jones, S N; Tibbetts, C (1989) Upstream DNA sequences determine different autoregulatory responses of the adenovirus types 5 and 3 E1A promoters. J Virol 63:1833-8
Larsen, P L; Tibbetts, C (1987) Adenovirus E1A gene autorepression: revertants of an E1A promoter mutation encode altered E1A proteins. Proc Natl Acad Sci U S A 84:8185-9
Tibbetts, C; Larsen, P L; Jones, S N (1986) Autoregulation of adenovirus E1A gene expression. J Virol 57:1055-64
Larsen, P L; McGrane, M M; Robinson, C C et al. (1986) An E1A mutant of adenovirus type 3: Ad3hr15 has reiterated DNA sequences 5' to its E1A gene. Virology 155:148-59
Larsen, P L; Tibbetts, C (1985) Spontaneous reiterations of DNA sequences near the ends of adenovirus type 3 genomes. Virology 147:187-200