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Adenovirus E1A proteins inhibit activation of transcription by p53
W T Steegenga1, T van Laar, N Riteco
1Laboratory of Molecular Carcinogenesis, Sylvius Laboratories, Leiden University, The Netherlands.
Abstract:
p53 stimulates the transcription of a number of genes, such as MDM2, Waf1, and GADD45. We and others have shown previously that this activity of p53 can be inhibited by adenovirus type 2 or 12 large E1B proteins. Here we show that the adenovirus E1A proteins also can repress the stimulation of transcription by p53, both in transient transfections and in stably transfected cell lines. The inhibition by E1A occurs without a significant effect on the DNA-binding capacity of p53. Furthermore, the activity of a fusion protein containing the N-terminal part of p53 linked to the GAL4 DNA-binding domain can be suppressed by E1A. This indicates that E1A affects the transcription activation domain of p53, although tryptic phosphopeptide mapping revealed that the level of phosphorylation of this domain does not change significantly in E1A-expressing cell lines. Gel filtration studies, however, showed p53 to be present in complexes of increased molecular weight as a result of E1A expression. Apparently, E1A can cause increased homo- or hetero-oligomerization of p53, which might result in the inactivation of the transcription activation domain of p53. Additionally, we found that transfectants stably expressing E1A have lost the ability to arrest in G1 after DNA damage, indicating that E1A can abolish the normal biological function of p53.
Insights
Adenovirus E1A proteins inhibit p53
Area of Science:
- Molecular Biology
- Virology
- Cancer Research
Background:
- p53 is a tumor suppressor protein that regulates gene transcription.
- Adenovirus E1B proteins are known inhibitors of p53 transcriptional activity.
- The role of adenovirus E1A proteins in modulating p53 function was not fully understood.
Purpose of the Study:
- To investigate the effect of adenovirus E1A proteins on p53-mediated gene transcription.
- To elucidate the mechanism by which E1A affects p53 activity.
- To determine if E1A can abolish the biological functions of p53.
Main Methods:
- Transient and stable transfections of cell lines.
- Assays to measure p53 DNA-binding capacity.
- GAL4-p53 fusion protein activity assays.
- Tryptic phosphopeptide mapping.
- Gel filtration chromatography.
- Analysis of G1 cell cycle arrest after DNA damage.
Main Results:
- Adenovirus E1A proteins repress p53-stimulated transcription without affecting p53 DNA binding.
- E1A suppresses the activity of the p53 transcription activation domain.
- E1A expression leads to increased p53 oligomerization.
- E1A-expressing cells lose the ability to arrest in G1 after DNA damage.
Conclusions:
- Adenovirus E1A proteins inactivate p53 transcriptional activity, likely through promoting p53 oligomerization.
- E1A disrupts the normal tumor-suppressive functions of p53, including cell cycle arrest.
- These findings reveal a novel mechanism of viral oncoprotein-mediated subversion of host cell pathways.