RESUMO
Many biological phenomena are dependent on mechanisms that fine-tune the expression levels of particular genes. This can be achieved by altering the relative activity of a single transcription factor, by post-translational modifications or by interaction with regulatory molecules. An alternative mechanism is based on competition between two or more differently active isoforms of the same transcription factor. We found that FHX, a recently characterized human fork-head transcriptional activator, may show such a mechanism for balancing its activity by expressing two differently sized isoforms, FHX.S and FHX.L, encoded by a single gene located on human chromosome 12. FHX. L and FHX.S showed different transcriptional capacities, the larger form, FHX.L, behaving as the more potent transactivator. A transactivation domain of the acidic type present only in FHX.L would account for this functional difference. The relative concentrations of these two FHX isoforms appear to vary in a number of cell types, a circumstance that may regulate the final activity of this transcription factor.
Assuntos
Processamento Alternativo , Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/metabolismo , Transativadores/química , Transativadores/metabolismo , Ativação Transcricional , Processamento Alternativo/genética , Motivos de Aminoácidos , Sequência de Aminoácidos , Sequência de Bases , Mapeamento Cromossômico , Cromossomos Humanos Par 12/genética , Clonagem Molecular , Proteínas de Ligação a DNA/genética , Fatores de Transcrição Forkhead , Perfilação da Expressão Gênica , Humanos , Dados de Sequência Molecular , Peso Molecular , Especificidade de Órgãos , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Estrutura Terciária de Proteína , RNA Mensageiro/análise , RNA Mensageiro/genética , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Transativadores/genéticaRESUMO
The HNF3/fork head family includes a large number of transcription factors that share a structurally related DNA binding domain. Fork head factors have been shown to play important roles both during development and in the adult. We now describe the cloning of a novel mammalian fork head factor that we have named FHX (fork head homologous X (FHX), which is expressed in many adult tissues. In the embryo, FHX expression showed a very early onset during the cleavage stages of preimplantation development. Polymerase chain reaction-assisted site selection experiments showed that FHX bound DNA with a dual sequence specificity. Sites recognized by FHX could be classified into two different types according to their sequences. Binding of FHX to sequences of each type appeared to occur independently. Our data suggest that either different regions of the fork head domain or different molecular forms of this domain could be involved in binding of FHX to each type of site. In transfection assays, FHX was capable of activating transcription from promoters containing FHX sites of either type.
Assuntos
Proteínas de Ligação a DNA/química , Proteínas de Ligação a DNA/genética , Adulto , Sequência de Aminoácidos , Sequência de Bases , Encéfalo/metabolismo , Clonagem Molecular , DNA Complementar/metabolismo , Proteínas de Ligação a DNA/metabolismo , Fatores de Transcrição Forkhead , Biblioteca Gênica , Humanos , Fígado/embriologia , Fígado/metabolismo , Dados de Sequência Molecular , Regiões Promotoras Genéticas , Ligação Proteica , Estrutura Terciária de Proteína , Proteínas Recombinantes de Fusão/metabolismo , Transcrição Gênica , TransfecçãoRESUMO
To elucidate whether pancreatic acinar cell submitted to stress is able to express TNF-alpha, we studied TNF-alpha mRNA expression by Northern blot and in situ hybridization in healthy pancreas, in tissue from caerulein-induced pancreatitis and after lipopolysaccharide (LPS) treatment. In specimens from normal pancreas, TNF-alpha mRNA expression, as judged by both Northern blot and in situ hybridization, was negative, whereas a strong but transient expression was observed in acinar cells from caerulein pancreatitis and LPS treatment. TNF-alpha mRNA appeared as rapidly as 30 min after treatment, and was maximal 6 h after. At this time, there was mild infiltration consisting mostly of polymorphonuclear leukocytes (PMNL) and no signal of TNF-alpha transcript was found in their cytoplasm. Our results strongly indicate that pancreatic acinar cell is the source of TNF-alpha early in the course of acute pancreatitis and LPS treatment, and suggest that the expression of this cytokine is a part of a general response of the acinar cell to aggression.
