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p53 oligomerization and DNA looping are linked with transcriptional activation
J E Stenger1, P Tegtmeyer, G A Mayr
1Department of Molecular Genetics and Microbiology, State University of New York, Stony Brook 11794.
Abstract:
We examined the role of p53 oligomerization in DNA binding and in transactivation. By conventional electron microscopy (EM) and scanning transmission EM, we find that wild-type tetramers contact 18-20 bp at single or tandem 19 bp consensus sequences and also stack in apparent register, tetramer on top of tetramer. Stacked tetramers link separated DNA binding sites with DNA loops. Interestingly, the p53(1-320) segment, which lacks the C-terminal tetramerization domain, binds DNA consensus sites as stacked oligomers. Although the truncated protein binds DNA with reduced efficiency, it nevertheless induces DNA looping by self-association. p53, therefore, has a C-terminal tetramerization domain that enhances DNA binding and a non-tetrameric oligomerization domain that stacks p53 at consensus sites and loops separated consensus sites via protein-protein interactions. Using model promoters, we demonstrate that wild-type and tetramerization-deficient p53s activate transcription well when tandem consensus sites are proximal to TATA sequences and poorly when tandem sites are distal. In the presence of proximal sites, however, stimulation by distal sites increases 25-fold. Tetramerization and stacking of tetramers, therefore, provide dual mechanisms to augment the number of p53 molecules available for activation through p53 response elements. DNA looping between separated response elements further increases the concentration of local p53 by translocating distally bound protein to the promoter.
Insights
The tumor suppressor p53 protein uses tetramerization and stacking to bind DNA and activate transcription. These mechanisms enhance DNA binding and facilitate DNA looping, increasing p53
Area of Science:
- Molecular Biology
- Protein-DNA Interactions
- Gene Regulation
Background:
- The tumor suppressor protein p53 plays a critical role in cellular responses to stress.
- p53's function is regulated by its ability to bind DNA and activate transcription.
- Oligomerization of p53 is known to be important for its DNA-binding activity.
Purpose of the Study:
- To investigate the role of p53 oligomerization in DNA binding and transcriptional activation.
- To elucidate the mechanisms by which p53 interacts with DNA and regulates gene expression.
Main Methods:
- Conventional electron microscopy (EM) and scanning transmission EM were used to visualize p53-DNA complexes.
- Studies involved wild-type p53 and a truncated p53(1-320) mutant lacking the tetramerization domain.
- Functional assays using model promoters assessed transcriptional activation.
Main Results:
- Wild-type p53 tetramers bind to DNA consensus sequences and can stack, forming tetramer-on-tetramer structures.
- Stacked p53 tetramers facilitate DNA looping, linking separated DNA binding sites.
- A p53 segment lacking the tetramerization domain also forms stacked oligomers on DNA, inducing looping and enhancing transcription when sites are proximal.
Conclusions:
- p53 utilizes both its C-terminal tetramerization domain and a separate oligomerization domain for DNA binding and transcriptional regulation.
- Tetramerization enhances DNA binding, while stacking and DNA looping provide additional mechanisms to increase p53 concentration at response elements.
- These dual mechanisms allow p53 to effectively regulate gene expression through cooperative binding and DNA looping.