Control of adenovirus early gene expression: posttranscriptional control mediated by both viral and cellular gene products.

102

The function of the adenovirus E1Aa protein (the product of the 13S ElA mRNA) during a productive viral infection is to activate transcription of the six early viral transcription units. To study the mechanism of action of this protein, a peptide which was 13 amino acids long and had a sequence unique to the protein product of the adenovirus 13S ElA mRNA (pElAa) was coupled to keyhole limpet hemocyanin and used to raise an antibody in rabbits. The resulting antiserum was specific to this protein and did not react with the protein product of the 12S ElA mRNA, which shares considerable sequence with the E1Aa protein.This antiserum was used to probe for the E1Aa protein in situ by indirect immunofluorescence and in extracts of infected HeLa cells. We found that the protein was associated with large cellular structures both in the nucleus and in the cytoplasm. The nuclear form of the protein was analyzed further and was found to purify with the nuclear matrix.

mentioning

confidence: 99%

“…There is also evidence that protein synthesis inhibition may have a posttranscriptional effect and may indicate that the ElA gene has more than one function (19). (ii) Early viral genes can be transcribed in HeLa cells in the absence of ElA function by using a high multiplicity of input virus for infection (32,40,41).…”

mentioning

confidence: 99%

Localization of the adenovirus E1Aa protein, a positive-acting transcriptional factor, in infected cells infected cells.

Feldman¹,

Nevins²

1983

102

“…It has been suggested that ElA acts by inactivating a cellular repressor of transcription (13,14,16,20,21). This has been used to explain the ability of ElA mutants to synthesize early mRNA and protein at high multiplicity in HeLa cells (16,20,27 …”

Section: Discussionmentioning

confidence: 99%

“…6). Because ElA has been reported to affect mRNA accumulation and translation as well as transcription (14,22), we also looked for E2A mRNA in rat cells infected by 1,000 IU of d1312 per cell but found none, although it was readily detectable in rat cells infected by WT Ad5 at 10 IU per cell (Fig. 6).…”

mentioning

confidence: 99%

Control functions of adenovirus transformation region E1A gene products in rat and human cells.

Bellett¹,

Li²,

David³

et al. 1985

Altered control of the rat cell cycle induced by adenovirus requires expression of transformation region ElA, but not of E1B, E2A, E2B, or late genes. We show here that neither E3 nor E4 is required, so the effect results directly from an ElA product. Mutants with defects in the 289-amino-acid (aa) ElA product had little or no effect on the rat cell cycle even at 1,000 IU The adenovirus transformation gene ElA has distant sequence homology with the retrovirus oncogenes v-myc and v-myb and can replace myc in transformation of primary cells (24,26). ElA induces expression of other viral early genes (2,12,27), and at least two cell genes, thymidine kinase and the 70,000-molecular-weight (70K) heat shock protein (1,3,8,11,21). The function of ElA in transformation appears to be "immortalization" of the cell and reduction of requirements for serum growth factors. The ability of adenovirus to induce a growth cycle in rodent cells arrested in the Gi phase by deprivation of serum or reduction of Ca2+ concentration, and to alter cycle progression in growing cells (1,5,8,18), may be related to the ElA transformation functions. Experiments with mutant viruses showed that alteration of control of the rat cell growth cycle requires expression of ElA, but not of E1B, E2A, E2B, or late genes (1,5,8,19). However, because ElA induces all of the other early regions, it remained possible that the cell cycle effects were actually due to E3 or E4, under the control of ElA. In this paper, we show that expression of human adenovirus type 5 (AdS) regions E3 and E4 is not necessary for alteration of rat cell cycle progression, which must therefore be a direct effect of ElA. Experiments with six mutants having different defects in the ElA 289-amino-acid (aa) product showed complete or almost complete defects in alteration of cell cycle progression even at 1,000 IU per cell. In this respect, these mutants lack the "multiplicity-dependent leakiness" reported for ElA mutants in human cells (20,27 ElA function in rat cells. The leakiness of some ElA mutants at high multiplicity in rat cells appears to be due to residual function of the mutant proteins. Additional leak in HeLa cells may be due to a cellular ElA-like function. MATERIALS AND METHODSViruses and cells. Ad2/5 pm975 (17) was a generous gift from A. J. Berk, University of California, Los Angeles; AdS hrA (28) was from D. Solnick, Yale University, New Haven, Conn.; and inSOO (6) was from N. C. Jones, Purdue University, West Lafayette, Ind. The origins of other AdS ElA mutants have been described previously (3). Locations of the ElA mutations are shown in relation to the early El mRNA species in Fig. 1. ElA mutants were grown in 293 cells and titrated by methods described previously (4,5,18,19), and with the exception of pm975 had a ratio of titer on 293 cells to that on HeLa cells of 103 to 104.Mutant d1808, deleted from between 91.4 and 92.0 to between 97.2 and 98.4 map units (m.u.) (7), and thus defective in subregions 1 or 2 through 7 of early region E4, was kindly supplied by G...

“…This represents a significant acceleration, since the normal latent period is only 4 to 5 h at 33°C. In this respect, TPA action resembles that of the AdS Ela regulatory product (5,26), the absence of which (in Ela mutants such as hrl) considerably delays early gene expression (12,19,20,26).…”

mentioning

confidence: 99%

Accelerated onset of viral transcription in adenovirus-infected HeLa cells treated with the tumor promoter 12-O-tetradecanoyl-phorbol-13-acetate.

Carter¹,

Milovanovic²,

Babiss³

et al. 1984

When adenovirus type 5-infected HeLa cells were exposed to the tumor promoter 12-0-tetradecanoylphorbol-13-acetate, short pulse-labeling with [3H]uridine in vivo and [3H]UTP incorporation by isolated nuclei in vitro were both consistent with a decreased latent period before initiation by RNA polymerase at early viral promoters. Acceleration was not dependent upon concurrent protein synthesis and could not be attributed to rapid entry of virus into the cell nucleus. 12-O-tetradecanoyl-phorbol-13-acetate suppressed the transcription-delay phenotype of the Ela mutant, hrl, without restoring its ability to replicate.