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'Gain of function' phenotype of tumor-derived mutant p53 requires the oligomerization/nonsequence-specific nucleic
1Department of Microbiology, University of Texas Health Science Center at San Antonio, 78284, USA.
Abstract:
Tumor-derived p53 mutants can transcriptionally activate a number of promoters of genes involved in cellular proliferation. For this transactivation, mutant p53 does not use the wild-type p53 DNA-binding site, suggesting a mechanism of transactivation that is independent of direct DNA binding. Here we describe our analysis of the domain requirements for mutant p53 to transactivate promoters of the human epidermal growth factor receptor (EGFR), human multiple drug resistance 1 (MDR-1) and human proliferating cell nuclear antigen (PCNA) genes. We also report the identification of a structural domain required for the 'gain of function' property of mutant p53-281G. 'Gain of function' is measured as the tumorigenicity (in nude mice) of 10(3) murine cells expressing mutant p53 constitutively. We have generated internal deletion mutants of p53-281G deleting conserved domains I, II, III, IV and V, individually. We have also generated one deletion mutant eliminating amino acids 100 through 300 that removes four of the five conserved domains (II - V); another mutant, p53-281G del 393-327, deletes the oligomerization and nonsequence-specific nucleic acid-binding domains of p53. For the EGFR and MDR-1 promoters, all these mutants have significantly lower transactivation ability than intact p53-281G. These deletion mutants, however, significantly activated the pCNA promoter, suggesting that the mechanism of transactivation of the PCNA promoter is different from that of the EGFR and MDR-1 promoters. When expressed constitutively in 10(3) cells, p53-281G del 393-327 was found to be defective in inducing tumor formation in nude mice although intact p53-281G was very efficient. Thus, our results suggest that structural domains near the C-terminus are needed for 'gain of function'.
Insights
Tumor-derived mutant p53 activates proliferation genes independently of DNA binding. Structural domains near the C-terminus are crucial for this
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Tumorigenic p53 mutants exhibit gain-of-function (GOF) properties, including transcriptional activation of proliferation-associated genes.
- This transactivation can occur independently of the canonical p53 DNA-binding site, suggesting alternative mechanisms.
Purpose of the Study:
- To investigate the domain requirements for mutant p53-mediated transactivation of specific gene promoters (EGFR, MDR-1, PCNA).
- To identify the structural domain responsible for the GOF tumorigenicity of mutant p53-281G.
Main Methods:
- Generation and analysis of p53-281G internal deletion mutants targeting conserved domains.
- Assessment of transactivation capacity on EGFR, MDR-1, and PCNA promoters.
- Evaluation of tumorigenicity in nude mice for specific deletion mutants.
Main Results:
- All deletion mutants showed reduced transactivation of EGFR and MDR-1 promoters compared to intact p53-281G.
- PCNA promoter transactivation was maintained by deletion mutants, indicating a distinct mechanism.
- A specific mutant (p53-281G del 393-327), lacking C-terminal domains, was defective in inducing tumor formation.
Conclusions:
- Structural domains near the C-terminus of p53 are essential for its gain-of-function (GOF) tumorigenicity.
- The mechanisms of transactivation for PCNA differ from those for EGFR and MDR-1 promoters.