Control of adenovirus early gene expression: accumulation of viral mRNA after infection of transformed cells

Abstract

We studied the accumulation of viral mRNA in the presence of inhibitors of protein synthesis in an adenovirus type 5-transformed cell line (line 293 cells). An analysis of the endogenous viral mRNA's and proteins revealed that only early (gions 1A and 1B were expressed in uninfected 293 cells. However, viral mRNA's from early regions 2, 3, and 4, as well as mRNA's from early regions 1A and 1B, accumulated in 293 cells after infection with adenovirus type 2. Cells treated with anisomycin befo- .-e infection showed a drastic enhancement of mRNA from early region 4 compared with drug-free controls. This increase in viral mRNA was detected by using filter hybridization, Si endonuclease mapping, and in vitro translation. The rate of transcription of early region 4 nuclear RNA also increased significantly in the presence of anisomycin. In contrast to the results with early region 4, the levels of cytoplasmic mRNA's from early regions 2 and 3 did not increase in cells treated with inhibitors. These results suggested that multiple virus encoded controls operate on the early regions of the adenovirus genome.

Cycloheximide stimulates early adenovirus transcription if early gene expression is allowed before treatment

“…No strong effect on transcription was seen when CH was added before the virus, as was reported earlier (16). The lack of stimulation of transcription by inhibiting protein synthesis before infection is in accord with the results from other laboratories with other protein synthesis inhibitors (11,18,19). Thus, protein synthesis inhibition can cause a strong stimulation of viral transcription and RNA accumulation, but only if a period of uninhibited early infection is allowed before treatment.…”

“…Persson et al (19) showed that in 293 cells, which express 1A products constitutively, blocking protein synthesis before infection causes a large stimulation of E4 transcription only; other early regions are not stimulated. This suggests that whereas E4 may be directly stimulated by inhibiting protein synthesis in the presence of ElA products, the CH stimulation of other early regions (1A, 1B, E2, E3, and L1) which we demonstrated may depend on the previous expression of other early genes besides 1A.…”

“…Additional studies using variations in virus input and addition of various inhibitors of protein synthesis have been described (12, 18, 19), but no completely consistent picture of early virus mRNA control has emerged. When CH is added before infection at a high MOI, transcription of all early virus transcription units (except region 4) still apparently achieves a normal rate (16), but excess amounts of mRNA as detected by translation do not accumulate (19). Anisomycin has a similar effect when a high MOI is used (11,12,18), but the accumulation of individual mRNA species as judged by translation is not uniform.…”

Control of Adenovirus Gene Expression: Cellular Gene Products Restrict Expression of Adenovirus Host Range Mutants in Nonpermissive Cells

“…These results led to a model to explain how the viral proteins encoded in region ElA can enhance accumulation of viral mRNA from other early regions. The ElA proteins may inactivate a cellular protein which, when present, decreases the amount of early viral mRNA (13,22,23). This model is consistent with all of the results obtained from experiments with protein synthesis inhibitors and furthermore suggests that the host range mutants from region ElA fail to inactivate the cellular protein, which leads to a drastic reduction in the accumulation of early viral mRNA.…”

“…If the regulatory element encoded in ElA is a protein, the addition of inhibitors of protein synthesis shortly before infection should effect the accumulation of early viral mRNA. Studies from our laboratory and others (13,16,22,23) have tested this hypothesis, and the accumulation of viral mRNA was reduced under these conditions. However, the addition of the drug at 1 h postinfection or later has the opposite effect, i.e., early viral mRNA accumulation is enhanced (5, 23).…”