RESUMO
The product of the oncosuppressor adenomatous polyposis coli (APC) gene is involved in cell cycle arrest and apoptosis and its loss of function is associated with the development of colorectal carcinogenesis. Its transcriptional regulation seems rather complex and has not been completely elucidated up to now. In an attempt to identify the transcription start sites for the mouse Apc gene we have detected a novel transcript in mouse embryonic stem (ES) cells and colon tissue. This transcript contains an untranslated exon, whose flanking sequences exhibited strong promoter activity in transient transfection experiments. These results suggest that we have identified a novel promoter for the mouse Apc gene, localized about 40 kb upstream of the initiating methionine, which drives expression of the unique Apc transcript type detected in undifferentiated totipotent ES cells. Transcripts bearing the novel exon combined either with exon 1 or with exon 2 were detected in all mouse tissues tested.
Assuntos
Proteína da Polipose Adenomatosa do Colo/genética , Metionina/genética , Regiões Promotoras Genéticas , Sítio de Iniciação de Transcrição , Região 3'-Flanqueadora , Região 5'-Flanqueadora , Proteína da Polipose Adenomatosa do Colo/metabolismo , Animais , Sequência de Bases , Diferenciação Celular , Células Cultivadas , Colo/fisiologia , Éxons , Ordem dos Genes , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Células-Tronco Totipotentes/fisiologiaRESUMO
Filensin and phakinin are two lens-specific members of the intermediate filament (IF) superfamily of proteins. They coassemble to form a beaded submembraneous filamentous network, the beaded filaments (BFs). The low sequence homology and differences in assembly compared to other IF proteins do not allow their classification in any of the five IF subgroups. The organization of the phakinin gene exon/intron boundaries provides evidence that this partner may be sharing a common origin with type I cytokeratin genes. Here we report the molecular cloning, sequence and characterization of the mouse filensin gene. The filensin gene consists of 8 exons and 7 introns, with 6 introns interrupting its rod domain in a highly conserved manner characteristic of type III IF genes, like vimentin, desmin, or peripherin. Of the two tail domain exons the one adjacent to the rod domain, compares to exon 7 of the non-neuronal cytoplasmic IF gene of helix aspersa and to the lamin region bridging the end of the rod domain to the nuclear localization signal. Altogether, these observations indicate that the lens beaded filaments form an independent class of IF.
Assuntos
Evolução Molecular , Proteínas do Olho/genética , Genes/genética , Proteínas de Filamentos Intermediários/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Mapeamento Cromossômico , Clonagem Molecular , Sequência Conservada , DNA Complementar/genética , Éxons/genética , Feminino , Dosagem de Genes , Íntrons/genética , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Análise de Sequência de DNA , Homologia de Sequência de AminoácidosRESUMO
Ligand activated Glucocorticoid Receptor (GR), specifically inhibited the serum induced c-fos promoter activation in NIH3T3 fibroblasts. The negative control was mediated by the c-fos SRE and correlated with the relative abundance of active GR. Serum activated SRE was repressed 3-4-fold by glucocorticoids irrespective of the promoter context (heterologous or authentic). The suppressing ability of GR was absolutely dependent on its DNA binding domain (DBD), since deletion of this region left the serum induction unimpaired. The methylation interference pattern of GR revealed two distinct binding sites within the SRE and identified the GR contact bases, important also for binding and function of SRE targeted transcription factors, such as the Serum Response Factor (SRF) and the p62 Ternary Complex Factor (TCF). We conclude that GR binds to c-fos SRE and inhibits c-fos promoter activation by antagonizing the function of positive transcription factors targeting to overlapping or identical sites. Since the c-fos SRE is activated by multiple mitogenic signalling pathways, inactivation by GR could explain, at least in part, the growth inhibitory response of fibroblasts to glucocorticoid hormones.