RESUMO
Most of cells exhibit low nuclear level of NF-kappaB. However, in some cell lines and tissues aberrantly activated NF-kappaB is playing an important role in cell motility, growth control and survival. Here we describe the result of decrease of constitutive NF-kappaB level in different adenocarcinoma cell lines. Treatment of mouse adenocarcinoma cell line CSML-100 with both synthetic (TPCK or PDTC) or natural (I(kappaB)-alpha) NF-kappaB inhibitors caused apoptotic death. Low doses of TPCK were harmless for CSML100 cells but sensitized them to TNF-induced apoptosis. Death of CSML100 cells in the presence of high concentration TPCK was not accompanied with significant changes in c-myc activity but strongly correlated with rapid decrease in p53 level. Thus, mutual behavior p53 and NF-kappaB represented a unique feature of TPCK-induced apoptosis in CSML-100 adenocarcinoma cells.
Assuntos
Adenocarcinoma/metabolismo , Adenocarcinoma/patologia , Proteínas I-kappa B , NF-kappa B/metabolismo , Animais , Apoptose/efeitos dos fármacos , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Núcleo Celular/patologia , Sobrevivência Celular , Proteínas de Ligação a DNA/genética , Proteínas de Ligação a DNA/metabolismo , Relação Dose-Resposta a Droga , Regulação para Baixo/efeitos dos fármacos , Citometria de Fluxo , Células Gigantes/metabolismo , Células Gigantes/patologia , Camundongos , Inibidor de NF-kappaB alfa , NF-kappa B/química , Prolina/análogos & derivados , Prolina/farmacologia , Ligação Proteica/efeitos dos fármacos , Subunidades Proteicas , Proteínas Proto-Oncogênicas c-myc/metabolismo , Tiocarbamatos/farmacologia , Tosilfenilalanil Clorometil Cetona/farmacologia , Transfecção , Células Tumorais Cultivadas , Fator de Necrose Tumoral alfa/farmacologia , Proteína Supressora de Tumor p53/metabolismoRESUMO
We describe a novel mammalian DNA binding activity that requires at least two symmetrically methylated CpG dinucleotides in its recognition sequence, preferably within the sequence 5'CGCG. A key component of the activity is Kaiso, a protein with POZ and zinc-finger domains that is known to associate with p120 catenin. We find that Kaiso behaves as a methylation-dependent transcriptional repressor in transient transfection assays. Kaiso is a constituent of one of two methyl-CpG binding complexes originally designated as MeCP1. The data suggest that zinc-finger motifs are responsible for DNA binding, and may therefore target repression to specific methylated regions of the genome. As Kaiso associates with p120 catenin, Kaiso may link events at the cell surface with DNA methylation-dependent gene silencing.
Assuntos
Moléculas de Adesão Celular/metabolismo , Proteínas de Ligação a DNA/metabolismo , Histona Desacetilases , Fosfoproteínas/metabolismo , Proteínas Repressoras/metabolismo , Fatores de Transcrição/metabolismo , Animais , Cateninas , Extratos Celulares , Linhagem Celular , Núcleo Celular/metabolismo , Ilhas de CpG , Metilação de DNA , Proteínas de Ligação a DNA/genética , Humanos , Camundongos , Coelhos , Proteínas Repressoras/genética , Transdução de Sinais , Fatores de Transcrição/genética , Transcrição Gênica , Dedos de Zinco , delta CateninaRESUMO
Expression of the mts1 gene is often associated with malignant transformation of tumor cells. Transcription of the gene is controlled by a number of positive and negative regulatory elements, all of them being localized in the first intron (+38 to +1215) of the mts1 gene. Through analysis of the distribution of DNase I hypersensitive sites in the first intron of the gene we revealed a structurally conserved region that consisted of a non-canonical NFkB binding site and a minisatellite "core" element. Deletion of the minisatellite core DNA in the context of the first intron had no effect on its regulatory capacity when assayed in transient transfections, while a fivefold decrease was observed in a pool of stably transfected cells. The minisatellite core sequence CTGGGCAGGCAG is involved in DNA-protein interactions in vivo, and is similar to a binding site for the previously identified minisatellite DNA sequence binding protein (Msbp-1). The core DNA interacted in vitro with a protein that had an apparent molecular mass of 40 kDa. These data indicate that the minisatellite DNA represents the novel, chromatin-specific element in the mts1 complex enhancer.
