title: Redox-mediated regulation of p21(waf1/cip1) expression involves a post-transcriptional mechanism and activation of the mitogen-activated protein kinase pathway.
abstract: p21(waf1/cip1) gene expression is induced by DNA damage in cells with wild-type p53 and contributes to the arrest of cell growth. It was demonstrated that under many experimental conditions, including oxidative stress, p21(waf1/cip1) expression can be induced through p53-independent pathways. Since most of these experimental conditions induce the phosphorylation of mitogen-activated protein kinase (MAPK) and thus its activation, we evaluated p21(waf1/cip1) mRNA levels in cells exposed to an oxidative stress, induced by diethylmaleate (Et2Mal), and in which the MAPK pathway was blocked. The expression of a dominant-negative mutant of MEK, the MAPK kinase that phosphorylates and activates MAPK, and of a dominant-negative [Asn17]Ras mutant prevented the Et2Mal-induced accumulation of p21(waf1/cip1) mRNA. Similarly, the expression of MEK- and of [Asn17]Ras mutants decreased the 12-O-tetradecanoyl-phorbol 13-acetate (TPA)-mediated p21(waf1/cip1) induction. Furthermore, TPA-induced and serum-induced p21(waf1/cip1) mRNA accumulation was blocked by pretreating the cells with the antioxidant compound N-acetylcysteine, suggesting that oxidative stress is involved in these responses. p21(waf1/cip1) mRNA levels reached a maximum within 2 h of adding Et2Mal or TPA; however, the rate of transcription from a p21(waf1/cip1)-promoter construct did not increase during this period. In contrast, cells treated with actinomycin D show an increase of p21(waf1/cip1) mRNA stability after Et2Mal treatment. This result suggests that the increase in p21(waf1/cip1) mRNA at early times results from post-transcriptional regulatory events. Longer exposure to TPA may activate p21(waf1/cip1) gene transcription through an Sp1-dependent mechanism, while Et2Mal treatment gradually inhibits p21(waf1/cip1) gene transcription through oxidative changes that affect Sp1 binding to DNA.
authors.affiliation: Dipartimento di Biochimica e Biotecnologie Mediche, Università degli Studi di Napoli Federico II, Naples, Italy.
authors.forename: F
authors.initials: F
authors.lastname: Esposito
authors.forename: F
authors.initials: F
authors.lastname: Cuccovillo
authors.forename: M
authors.initials: M
authors.lastname: Vanoni
authors.forename: F
authors.initials: F
authors.lastname: Cimino
authors.forename: C W
authors.initials: CW
authors.lastname: Anderson
authors.forename: E
authors.initials: E
authors.lastname: Appella
authors.forename: T
authors.initials: T
authors.lastname: Russo
mesh_terms: D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000235:genetics* / Q000378:metabolism; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D006367:HeLa Cells; D006801:Humans; D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000235:genetics*; D012323:RNA Processing, Post-Transcriptional; D012333:RNA, Messenger / Q000032:analysis; D017382:Reactive Oxygen Species; D015398:Signal Transduction / Q000235:genetics*; D016159:Tumor Suppressor Protein p53 / Q000235:genetics
chemical_list: C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D016159:Tumor Suppressor Protein p53; D017871:Calcium-Calmodulin-Dependent Protein Kinases