Redundant control of adenovirus late gene expression by early region 4

Abstract

A series of human adenovirus type 5 derivatives carrying deletion mutations in early region 4 (E4) were constructed and characterized with respect to viral late protein synthesis, viral cytoplasmic late message accumulation, viral DNA accumulation, and plaquing ability. Viral late protein synthesis was essentially normal in cells infected by mutants expected to produce either the E4 open reading frame (ORF) 3 product or the E4 ORF 6 product. In cells infected by mutants lacking both ORF 3 and ORF 6, late protein synthesis was dramatically reduced. The basis for this reduction appears to be a concomitant reduction in cytoplasmic late message levels. Our results suggest that the products of ORFs 3 and 6 are redundant, since they are individually able to satisfy the requirement for E4 in late gene expression. Two of the mutants examined were defective for viral late protein synthesis but showed no measurable defect in viral DNA accumulation. The defect in late gene expression is not, therefore, a reflection of a primary defect in viral DNA synthesis. Finally, mutants expected to express ORF 3 or ORF 6 formed plaques with normal or only modestly reduced efficiency, whereas mutants expected to express neither ORF formed plaques with an efficiency less than 10-6that of wild-type virus. Thus, plaque-forming ability reflected late protein synthetic ability, suggesting that among these mutants late protein synthetic proficiency is the principle determinant of plaquing efficiency.

Stabilization of transgenes delivered by recombinant adenovirus vectors through extrachromosomal replication

“…Indeed, as has been demonstrated previously [19], at least some adenovirus mutants with deletion of the entire E4 region are not completely defective in DNA replication in 293 cells. The presence of the unexcised AdP2-8c genome might re¯ect, at least in part, the fact that the level of Cre expression was insuf®cient to mediate episome excision from all vector genomes.…”

Targeting of adenovirus E1A and E4-ORF3 proteins to nuclear matrix-associated PML bodies.

“…4 A) (Freyer et al, 1984;Virtanen et al, 1984), we made use of distinct deletion mutations of E4 in order to identify the protein(s) required for PML redistribution. The following E4 mutations were tested (Bridge and Ketner, 1989): (a) H5dl 1005, which deletes ORFs 1 and 2; (b) H5dl 1006, which deletes ORFs 1, 2, and 3; and (c) H5dl 1007, which deletes ORF3 but also affects the ORF6 product. The results show that, whereas mutant dl 1005 induces the typical filamentous structures with an efficiency similar to that of the wild-type virus (Fig.…”

“…Two of them, ORF3 and ORF6, have redundant effects in viral lytic infection (Huang and Hearing, 1989;Bridge and Ketner, 1989). While mutation in either of the two proteins essentially results in a normal productive infection, a doublemutant, defective in both ORF3 and ORF6 expression, grows extremely poorly and shows multiple lesions including defects in viral DNA replication, late viral mRNA accumulation and protein synthesis as well as failure in the shut-off of host cell protein synthesis (Halbert et al, 1985;Weinberg and Ketner, 1986;Huang and Hearing, 1989;Bridge and Ketner, 1989). Little is known concerning the biological activities of the ORF3 and ORF6 proteins.…”

RNA‐containing adenovirus/polyethylenimine transfer complexes effectively transduce dendritic cells and induce antigen‐specific T cell responses

“…Adenoviruses E4-defective human adenovirus (dl1014) [38] of serotype 5 was used for Ad/PEI transfection. dl1014 was grown in W162 cells [39] and purified by double banding in CsCl.…”