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Transcriptional control by adenovirus E1A conserved region 3 via p300/CBP.

Pelka P, Ablack JN, Torchia J, Turnell AS, Grand RJ, Mymryk JS - Nucleic Acids Res. (2009)

Bottom Line: This is supported by the observation that the repressive effect of E1A 12S is reversed by expression of exogenous p300 or CBP, but not by a CBP mutant lacking actyltransferase activity.Furthermore, we show that transcriptional activation by 13S E1A is greatly reduced by siRNA knockdown of p300 and that CR3 binds p300 independently of the well-characterized N-terminal/CR1-binding site.These results identify a new functionally significant interaction between E1A CR3 and the p300/CBP acetyltransferases, expanding our understanding of the mechanism by which this potent transcriptional activator functions.

View Article: PubMed Central - PubMed

Affiliation: Department of Oncology, The University of Western Ontario, London Regional Cancer Centre, London, Ontario, Canada. peter.pelka@gmail.com

ABSTRACT
The human adenovirus type 5 (HAdV-5) E1A 13S oncoprotein is a potent regulator of gene expression and is used extensively as a model for transcriptional activation. It possesses two independent transcriptional activation domains located in the N-terminus/conserved region (CR) 1 and CR3. The protein acetyltransferase p300 was previously identified by its association with the N-terminus/CR1 portion of E1A and this association is required for oncogenic transformation by E1A. We report here that transcriptional activation by 13S E1A is inhibited by co-expression of sub-stoichiometric amounts of the smaller 12S E1A isoform, which lacks CR3. Transcriptional inhibition by E1A 12S maps to the N-terminus and correlates with the ability to bind p300/CBP, suggesting that E1A 12S is sequestering this limiting factor from 13S E1A. This is supported by the observation that the repressive effect of E1A 12S is reversed by expression of exogenous p300 or CBP, but not by a CBP mutant lacking actyltransferase activity. Furthermore, we show that transcriptional activation by 13S E1A is greatly reduced by siRNA knockdown of p300 and that CR3 binds p300 independently of the well-characterized N-terminal/CR1-binding site. Importantly, CR3 is also required to recruit p300 to the adenovirus E4 promoter during infection. These results identify a new functionally significant interaction between E1A CR3 and the p300/CBP acetyltransferases, expanding our understanding of the mechanism by which this potent transcriptional activator functions.

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Schematic of E1A isoforms and locations of binding sites for indicated proteins. (A) Schematic representation of E1A 12S and E1A 13S splice isoforms. (B) Binding sites for p300/CBP, pCAF, TBP, p400 and TRRAP on E1A are indicated.
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Figure 1: Schematic of E1A isoforms and locations of binding sites for indicated proteins. (A) Schematic representation of E1A 12S and E1A 13S splice isoforms. (B) Binding sites for p300/CBP, pCAF, TBP, p400 and TRRAP on E1A are indicated.

Mentions: Human adenovirus type 5 (HAdV-5) early region 1A (E1A) is the first viral gene to be transcribed upon infection and plays an essential role in activating transcription (1,2). The 13S and 12S E1A mRNAs encode two major products of 289 residues (R) and 243R, respectively (Figure 1A), and these share identical amino and carboxyl sequences. The only difference between them is the presence of an additional 46 amino acids in the 289R protein that arises as the result of differential splicing of the primary E1A transcript (2). The region unique to the 13S encoded E1A protein coincides with a region that is highly conserved amongst the E1A proteins of different adenovirus serotypes, referred to as conserved region 3 (CR3) (3–5). Of the two major E1A polypeptides, the larger is considered to be primarily responsible for transcriptional activation of gene expression. Indeed, alterations within CR3 generally abolish E1A transactivation (6–10). Interestingly, a synthetic CR3 peptide corresponding to residues 140–188 of E1A was sufficient to transactivate adenovirus early promoters when microinjected into HeLa cells (11). Later work identified an adjacent acidic region spanning residues 189–200, termed Auxiliary Region 1 (AR1) as essential for efficient transactivation of early viral promoters by E1A (12).Figure 1.


Transcriptional control by adenovirus E1A conserved region 3 via p300/CBP.

Pelka P, Ablack JN, Torchia J, Turnell AS, Grand RJ, Mymryk JS - Nucleic Acids Res. (2009)

Schematic of E1A isoforms and locations of binding sites for indicated proteins. (A) Schematic representation of E1A 12S and E1A 13S splice isoforms. (B) Binding sites for p300/CBP, pCAF, TBP, p400 and TRRAP on E1A are indicated.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2651774&req=5

Figure 1: Schematic of E1A isoforms and locations of binding sites for indicated proteins. (A) Schematic representation of E1A 12S and E1A 13S splice isoforms. (B) Binding sites for p300/CBP, pCAF, TBP, p400 and TRRAP on E1A are indicated.
Mentions: Human adenovirus type 5 (HAdV-5) early region 1A (E1A) is the first viral gene to be transcribed upon infection and plays an essential role in activating transcription (1,2). The 13S and 12S E1A mRNAs encode two major products of 289 residues (R) and 243R, respectively (Figure 1A), and these share identical amino and carboxyl sequences. The only difference between them is the presence of an additional 46 amino acids in the 289R protein that arises as the result of differential splicing of the primary E1A transcript (2). The region unique to the 13S encoded E1A protein coincides with a region that is highly conserved amongst the E1A proteins of different adenovirus serotypes, referred to as conserved region 3 (CR3) (3–5). Of the two major E1A polypeptides, the larger is considered to be primarily responsible for transcriptional activation of gene expression. Indeed, alterations within CR3 generally abolish E1A transactivation (6–10). Interestingly, a synthetic CR3 peptide corresponding to residues 140–188 of E1A was sufficient to transactivate adenovirus early promoters when microinjected into HeLa cells (11). Later work identified an adjacent acidic region spanning residues 189–200, termed Auxiliary Region 1 (AR1) as essential for efficient transactivation of early viral promoters by E1A (12).Figure 1.

Bottom Line: This is supported by the observation that the repressive effect of E1A 12S is reversed by expression of exogenous p300 or CBP, but not by a CBP mutant lacking actyltransferase activity.Furthermore, we show that transcriptional activation by 13S E1A is greatly reduced by siRNA knockdown of p300 and that CR3 binds p300 independently of the well-characterized N-terminal/CR1-binding site.These results identify a new functionally significant interaction between E1A CR3 and the p300/CBP acetyltransferases, expanding our understanding of the mechanism by which this potent transcriptional activator functions.

View Article: PubMed Central - PubMed

Affiliation: Department of Oncology, The University of Western Ontario, London Regional Cancer Centre, London, Ontario, Canada. peter.pelka@gmail.com

ABSTRACT
The human adenovirus type 5 (HAdV-5) E1A 13S oncoprotein is a potent regulator of gene expression and is used extensively as a model for transcriptional activation. It possesses two independent transcriptional activation domains located in the N-terminus/conserved region (CR) 1 and CR3. The protein acetyltransferase p300 was previously identified by its association with the N-terminus/CR1 portion of E1A and this association is required for oncogenic transformation by E1A. We report here that transcriptional activation by 13S E1A is inhibited by co-expression of sub-stoichiometric amounts of the smaller 12S E1A isoform, which lacks CR3. Transcriptional inhibition by E1A 12S maps to the N-terminus and correlates with the ability to bind p300/CBP, suggesting that E1A 12S is sequestering this limiting factor from 13S E1A. This is supported by the observation that the repressive effect of E1A 12S is reversed by expression of exogenous p300 or CBP, but not by a CBP mutant lacking actyltransferase activity. Furthermore, we show that transcriptional activation by 13S E1A is greatly reduced by siRNA knockdown of p300 and that CR3 binds p300 independently of the well-characterized N-terminal/CR1-binding site. Importantly, CR3 is also required to recruit p300 to the adenovirus E4 promoter during infection. These results identify a new functionally significant interaction between E1A CR3 and the p300/CBP acetyltransferases, expanding our understanding of the mechanism by which this potent transcriptional activator functions.

Show MeSH
Related in: MedlinePlus