RESUMO
The expression of the APP (amyloid precursor protein), which plays a key role in the development of AD (Alzheimer's disease), is regulated by a variety of cellular mediators in a cell-dependent manner. In this study, we present evidence that p53 regulates the expression of the APP gene in neuroblastoma cells. Transient expression of ectopic p53, activation of endogenous p53 by the DNA-damaging drug camptothecin or Mdm2 (murine double minute 2) depletion decreases the intracellular levels of APP in murine N2abeta neuroblastoma cells. This effect was also observed in primary cultures of rat neurons as well as in SH-SY5Y cells, a human neuroblastoma cell line. Transient transfection studies using plasmids that contain progressive deletions of the 5' region of the gene demonstrate that p53 represses APP promoter activity through a mechanism that is mediated by DNA sequences located downstream of the transcription start site (+55/+101). Accordingly, expression of a dominant-negative p53 mutant significantly increases the transcriptional activity of the APP promoter. In addition, results obtained in gel mobility-shift assays show that p53 does not bind to the +55/+101 APP region, although it reduces binding of the transcription factor Sp1 (stimulating protein 1). Reduction of Sp1 binding after activation of p53 with camptothecin was also observed in chromatin immunoprecipitation assays. Altogether, our results strongly suggest a mechanism by which p53 precludes binding of Sp1 to DNA, and therefore the stimulation of the APP promoter by this transcription factor.
Assuntos
Precursor de Proteína beta-Amiloide/biossíntese , Proteína Supressora de Tumor p53/fisiologia , Animais , Camptotecina/farmacologia , Linhagem Celular Tumoral , Imunoprecipitação da Cromatina , DNA/efeitos dos fármacos , Ensaio de Desvio de Mobilidade Eletroforética , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos , Neuroblastoma/metabolismo , Regiões Promotoras Genéticas/fisiologia , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Ratos , Fator de Transcrição Sp1/metabolismoRESUMO
As a precursor of the neurotoxic amyloid-beta peptide, APP plays a central role in Alzheimer's disease. We have recently reported that the tumor suppressor p53 inhibits APP gene transcription through the same DNA sequences that mediate an inhibitory effect of thyroid hormones. Now, we have analyzed whether the thyroid hormone T3 can modulate the effects of p53 on APP expression. Exposition to UVC radiation leads to a marked decrease of intracellular APP levels that is paradoxically reversed by T3. Repression by UVC and reversion by the hormone are not observed in cells depleted of p53, demonstrating a p53-dependent mechanism. These results suggest the existence of a cross-talk between p53 and T3 that could play an important role in Alzheimer s disease.