RESUMO
Beta cell death may occur both after islet isolation and during infusion back into recipients undergoing total pancreatectomy with islet autotransplantation (TPIAT) for chronic pancreatitis. We measured the novel beta cell death marker unmethylated insulin (INS) DNA in TPIAT recipients before and immediately after islet infusion (n = 21) and again 90 days after TPIAT, concurrent with metabolic functional assessments (n = 25). As expected, INS DNA decreased after pancreatectomy (p = 0.0002). All TPIAT recipients had an elevated unmethylated INS DNA ratio in the first hours following islet infusion. In four samples (three patients), INS DNA was also assessed immediately after islet isolation and again before islet infusion to assess the impact of the isolation process: Unmethylated and methylated INS DNA fractions both increased over this interval, suggesting death of beta cells and exocrine tissue before islet infusion. Higher glucose excursion with mixed-meal tolerance testing was associated with persistently elevated INS DNA at day 90. In conclusion, we observed universal early elevations in the beta cell death marker INS DNA after TPIAT, with pronounced elevations in the islet supernatant before infusion, likely reflecting beta cell death induced by islet isolation. Persistent posttransplant elevation of INS DNA predicted greater hyperglycemia at 90 days.
Assuntos
Metilação de DNA , DNA/química , Diabetes Mellitus Tipo 1/cirurgia , Células Secretoras de Insulina/patologia , Insulina/genética , Transplante das Ilhotas Pancreáticas , Pancreatectomia/efeitos adversos , Pancreatite Crônica/cirurgia , Adolescente , Adulto , Biomarcadores/metabolismo , Criança , DNA/genética , Feminino , Rejeição de Enxerto , Sobrevivência de Enxerto , Humanos , Células Secretoras de Insulina/metabolismo , Masculino , Complicações Pós-Operatórias , Prognóstico , Estudos Prospectivos , Fatores de Risco , Transplante Autólogo , Adulto JovemRESUMO
Transcription factor (TF)-cofactor (COF) interactions define dynamic, cell-specific networks that govern gene expression; however, these networks are understudied due to a lack of methods for high-throughput profiling of DNA-bound TF-COF complexes. Here we describe the Cofactor Recruitment (CoRec) method for rapid profiling of cell-specific TF-COF complexes. We define a lysine acetyltransferase (KAT)-TF network in resting and stimulated T cells. We find promiscuous recruitment of KATs for many TFs and that 35% of KAT-TF interactions are condition specific. KAT-TF interactions identify NF-κB as a primary regulator of acutely induced H3K27ac. Finally, we find that heterotypic clustering of CBP/P300-recruiting TFs is a strong predictor of total promoter H3K27ac. Our data supports clustering of TF sites that broadly recruit KATs as a mechanism for widespread co-occurring histone acetylation marks. CoRec can be readily applied to different cell systems and provides a powerful approach to define TF-COF networks impacting chromatin state and gene regulation.
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Fungal communities form close beneficial (mutualists) or detrimental (pathogens) associations with their plant hosts. Their diversity and abundance can be affected by agricultural practices which include cropping systems such as rotations and intercropping. Despite the importance of cropping systems in increasing productivity, knowledge of the fungal mycobiome and the core inhabitants for under-utilised cereal and legume crops, particularly over a period, is still limited. The core mycobiomes in plant tissues and bulk soils of a cereal-legume intercrop were characterized over two years using high-throughput sequencing. The intercropping trial consisted of sorghum, Bambara groundnut, cowpea, dry bean, and soybean. A greater number of molecular operational taxonomic units (MOTUs) were found in plant tissues compared to those from the soils and between year one and year two. Principal coordinate analyses revealed that fungal communities for each year were relatively distinct, particularly for the soils. The core mycobiome was dominated by a Davidiellaceae sp. (Cladosporium), Didymellaceae sp. 1 (Phoma), Didymellaceae sp. 2 (Epicoccum), Fusarium sp. 2, Unidentified (Ascomycota), and Cryptococcus MOTUs that were present in all plant tissues and soils of year one and two. Other key MOTUs were only specific to a year, substrate, or crop. Although the mycobiome of sorghum were more distinct than the cores of the legumes, there were still MOTUs dominant across all of the crops. Characterization of this baseline core across two years provides insight into those fungi that are always present in these crops, and that could be utilized in improving crop performance and productivity.
RESUMO
Plant-associated fungi, or the mycobiome, inhabit plant surfaces above ground, reside in plant tissues as endophytes, or are rhizosphere in the narrow zone of soil surrounding plant roots. Studies have characterized mycobiomes of various plant species, but little is known about the sorghum mycobiome, especially in Africa, despite sorghum being one of the most important indigenous and commercial cereals in Africa. In this study, the mycobiome associated with above- and below-ground tissues of three commercial sorghum cultivars, as well as from rhizosphere and surrounding bulk soil samples, were sequenced using targeted sequencing with the Illumina MiSeq platform. Relative abundance differences between fungal communities were found between above-ground and below-ground niches, with most differences mostly in the dominant MOTUs, such as Davidiellaceae sp. (Cladosporium), Didymellaceae sp. 1 (Phoma), Fusarium, Cryptococcus and Mucor. Above-ground communities also appeared to be more diverse than below-ground communities, and plants harboured the most diversity. A considerable number of MOTUs were shared between the cultivars although, especially for NS5511, their abundances often differed. Several of the detected fungal groups include species that are plant pathogens of sorghum, such as Fusarium, and, at low levels, Alternaria and the Ustilaginomycetes. Findings from this study illustrate the usefulness of targeted sequencing of the ITS rDNA gene region (ITS2) to survey and monitor sorghum fungal communities and those from associated soils. This knowledge may provide tools for disease management and crop production and improvement.
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The human c-rel proto-oncogene (REL) encodes a subunit of the nuclear factor-kappaB (NF-kappaB) transcription factor. In this report, we have identified an identical point mutation in two human B-cell lymphomas (follicular (FL) and mediastinal) that changes serine (Ser)525 (TCA) to proline (Pro) (CCA) within the REL transactivation domain. This mutation was not identified in a similarly sized cohort of healthy individuals. In the mediastinal B-cell lymphoma, the mutation in REL is of germ-line origin. In both tumors, the S525P mutant allele is over-represented. REL-S525P shows enhanced in vitro transforming activity in chicken spleen cells. REL-S525P has a reduced ability to activate the human manganese superoxide dismutase (MnSOD) promoter in A293 cells; however, the MnSOD protein shows increased expression in REL-S525P-transformed chicken spleen cells as compared to wild-type REL-transformed cells. Ser525 is a site for phosphorylation by IkappaB kinase (IKK) in vitro. The S525P mutation reduces IKKalpha- and tumor necrosis factor (TNF)alpha-stimulated transactivation by a GAL4-REL protein. Furthermore, REL-S525P-transformed chicken spleen cells are more resistant to TNFalpha-induced cell death than cells transformed by wild-type REL. These results suggest that the S525P mutation contributes to the development of human B-cell lymphomas by affecting an IKKalpha-regulated transactivation activity of REL.
Assuntos
Transformação Celular Viral , Quinase I-kappa B/fisiologia , Linfoma de Células B/genética , Mutação Puntual/genética , Proteínas Proto-Oncogênicas c-rel/genética , Sequência de Aminoácidos , Animais , Western Blotting , Galinhas , Ensaio de Desvio de Mobilidade Eletroforética , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Hibridização in Situ Fluorescente , Rim/metabolismo , Luciferases/metabolismo , Linfoma de Células B/metabolismo , Linfoma Folicular/genética , Linfoma Folicular/metabolismo , Neoplasias do Mediastino/genética , Neoplasias do Mediastino/metabolismo , Dados de Sequência Molecular , NF-kappa B/genética , NF-kappa B/metabolismo , Fosforilação , Regiões Promotoras Genéticas/genética , Proto-Oncogene Mas , Proteínas Proto-Oncogênicas c-rel/metabolismo , Homologia de Sequência de Aminoácidos , Baço/metabolismo , Baço/virologia , Ativação Transcricional , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
This article serves as an introduction to the collection of reviews on nuclear factor-kappaB (NF-kappaB). It provides an overview of the discovery and current status of NF-kappaB as a research topic. Described are the structures, activities and regulation of the proteins in the NF-kappaB family of transcription factors. NF-kappaB signaling is primarily regulated by inhibitor kappaB (IkappaB) proteins and the IkappaB kinase complex through two major pathways: the canonical and non-canonical NF-kappaB pathways. The organization and focus of articles included in the following reviews are described, as well as likely future areas of research interest on NF-kappaB.
Assuntos
NF-kappa B/fisiologia , NF-kappa B/metabolismo , Transdução de Sinais , Terminologia como AssuntoRESUMO
The nuclear factor-kappa B (NF-kappaB) signaling pathway is a multi-component pathway that regulates the expression of hundreds of genes that are involved in diverse and key cellular and organismal processes, including cell proliferation, cell survival, the cellular stress response, innate immunity and inflammation. Not surprisingly, mis-regulation of the NF-kappaB pathway, either by mutation or epigenetic mechanisms, is involved in many human and animal diseases, especially ones associated with chronic inflammation, immunodeficiency or cancer. This review describes human diseases in which mutations in the components of the core NF-kappaB signaling pathway have been implicated and discusses the molecular mechanisms by which these alterations in NF-kappaB signaling are likely to contribute to the disease pathology. These mutations can be germline or somatic and include gene amplification (e.g., REL), point mutations and deletions (REL, NFKB2, IKBA, CYLD, NEMO) and chromosomal translocations (BCL-3). In addition, human genetic diseases are briefly described wherein mutations affect protein modifiers or transducers of NF-kappaB signaling or disrupt NF-kappaB-binding sites in promoters/enhancers.
Assuntos
Doenças Genéticas Inatas/metabolismo , Mutação , NF-kappa B/metabolismo , Transdução de Sinais , HumanosRESUMO
Nuclear factor kappa B (NF-kappaB) transcription factors regulate several important physiological processes, including inflammation and immune responses, cell growth, apoptosis, and the expression of certain viral genes. Therefore, the NF-kappaB signaling pathway has also provided a focus for pharmacological intervention, primarily in situations of chronic inflammation or in cancer, where the pathway is often constitutively active and plays a key role in the disease. Now that many of the molecular details of the NF-kappaB pathway are known, it is clear that modulators of this pathway can act at several levels. As described herein, over 750 inhibitors of the NF-kappaB pathway have been identified, including a variety of natural and synthetic molecules. These compounds include antioxidants, peptides, small RNA/DNA, microbial and viral proteins, small molecules, and engineered dominant-negative or constitutively active polypeptides. Several of these molecules act as general inhibitors of NF-kappaB induction, whereas others inhibit specific pathways of induction. In addition, some compounds appear to target multiple steps in the NF-kappaB pathway. Compounds designed as specific NF-kappaB inhibitors are not yet in clinical use, but they are likely to be developed as treatments for certain cancers and neurodegenerative and inflammatory diseases. Moreover, the therapeutic and preventative effects of many natural products may, at least in part, be due to their ability to inhibit NF-kappaB.
Assuntos
NF-kappa B/metabolismo , Transdução de Sinais/efeitos dos fármacos , Animais , Bactérias/metabolismo , Fungos/metabolismo , Humanos , Hidrólise , Quinase I-kappa B/metabolismo , NF-kappa B/antagonistas & inibidores , Vírus/metabolismoRESUMO
Complexes formed from the nuclear factor kappaB (NF-kappaB) family of transcription factors are ubiquitously expressed and are induced by a diverse array of stimuli. This results in their becoming activated in a wide variety of different settings. While the functions of NF-kappaB in many of these contexts have been the subject of intense research and are now well established, it is also clear that there is great diversity in the effects and consequences of NF-kappaB activation. NF-kappaB subunits do not necessarily regulate the same genes, in an identical manner, in all of the different circumstances in which they are induced. This review will discuss the different functions of NF-kappaB, the pathways that modulate NF-kappaB subunit activity and, in contrast to its more commonly thought of role as a promoter of cancer cell growth and survival, the ability of NF-kappaB, under some circumstances, to behave as a tumor suppressor.
Assuntos
Modelos Biológicos , NF-kappa B/metabolismo , Neoplasias/terapia , Transdução de Sinais , Proteínas Supressoras de Tumor/metabolismo , Carcinógenos/metabolismo , Humanos , Quinase I-kappa B/metabolismo , NF-kappa B/fisiologiaRESUMO
A newly described family of transcriptional regulatory proteins, the Rel family, has recently been the subject of much interest. The Rel family includes proteins known to be important in Drosophila development, replication of HIV-1, oncogenesis and general transcriptional control. Nevertheless, there is still much to be learned about their precise mechanism of action, including the process by which the original member of this family, v-Rel, malignantly transforms cells.
Assuntos
Transformação Celular Neoplásica , Mutação , Oncogenes , Proteínas Oncogênicas de Retroviridae/genética , Fatores de Transcrição/genética , Sequência de Aminoácidos , Animais , Humanos , Dados de Sequência Molecular , Proteínas Oncogênicas v-rel , Proteínas Oncogênicas de Retroviridae/metabolismo , Fatores de Transcrição/metabolismoRESUMO
The I kappa B proteins bind to Rel/NF-kappa B transcription factors and modulate their activities. Although originally described only as cytoplasmic inhibitors of Rel/NF-kappa B transcription complexes, it is now clear that I kappa B proteins also have other functions.
Assuntos
Proteínas de Ligação a DNA/fisiologia , Proteínas Proto-Oncogênicas/fisiologia , Fatores de Transcrição/fisiologia , Sequência de Aminoácidos , Proteínas I-kappa B , Dados de Sequência MolecularRESUMO
The v-Rel oncoprotein of the avian Rev-T retrovirus is a member of the Rel/NF-kappa B family of transcription factors. The mechanism by which v-Rel malignantly transforms chicken spleen cells is not precisely known. To gain a better understanding of functions needed for transformation by v-Rel, we have now characterized the activities of mutant v-Rel proteins that are defective for specific protein-protein interactions. Mutant v-delta NLS, which has a deletion of the primary v-Rel nuclear localizing sequence, does not interact efficiently with I kappa B-alpha but still transforms chicken spleen cells approximately as well as wild-type v-Rel, indicating that interaction with I kappa B-alpha is not essential for the v-Rel transforming function. A second v-Rel mutant, v-SPW, has been shown to be defective for the formation of homodimers, DNA binding, and transformation. However, we now find that v-SPW can form functional DNA-binding heterodimers in vitro and in vivo with the cellular protein NF-kappa B p-52. Most strikingly, coexpression of v-SPW and p52 from a retroviral vector can induce the malignant transformation of chicken spleen cells, whereas expression of either protein alone cannot. Our results are most consistent with a model wherein Rel homodimers or heterodimers must bind DNA and alter gene expression in order to transform lymphoid cells.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Proteínas I-kappa B , NF-kappa B/metabolismo , Proteínas Oncogênicas de Retroviridae/metabolismo , Animais , Sequência de Bases , Transformação Celular Neoplásica , Células Cultivadas , Galinhas , Dados de Sequência Molecular , Mutação , Inibidor de NF-kappaB alfa , Proteínas Oncogênicas v-rel , Ligação Proteica , Proteínas Oncogênicas de Retroviridae/genéticaRESUMO
We have examined the phosphorylation of a 50,000-dalton cellular polypeptide associated with the Rous sarcoma virus (FSV) transforming protein pp60-src. It has been shown that pp60src forms a complex with two cellular polypeptides, an 89,000-dalton heat-shock protein (89K) and a 50,000-dalton phosphoprotein (50K). The pp60src-associated protein kinase activity phosphorylates at tyrosine residues, and the 50K polypeptide present in the complex contains phosphotyrosine and phosphoserine. These observations suggest that the 50K polypeptide may be a substrate for the protein kinase activity of pp60src. To examine this possibility, we isolated the 50K polypeptide by two-dimensional polyacrylamide gel electrophoresis from lysates of uninfected or virally infected cells. Tryptic phosphopeptide analysis indicated that the 50K polypeptide isolated by this method was the same polypeptide as that complexed to pp60src. In uninfected cells or cells infected by a transformation-defective mutant, the 50K polypeptide contained phosphoserine but little or no phosphotyrosine. In cells infected by Schmidt-Ruppin or Prague RSV, there was a 40- to 50-fold increase in the quantity of phosphotyrosine in the 50K protein. Thus, the phosphorylation of the 50K polypeptide at tyrosine is dependent on the presence of pp60src. However, the 50K polypeptide isolated from cells infected by temperature-sensitive mutants of RSV was found to be phosphorylated at tyrosine at both permissive and nonpermissive temperatures; this behavior is different from that of other substrates or putative substrates of the pp60src kinase activity. It is possible that the 50K polypeptide is a high-affinity substrate of pp60src.
Assuntos
Vírus do Sarcoma Aviário/metabolismo , Transformação Celular Viral , Fosfoproteínas/metabolismo , Tirosina/metabolismo , Proteínas Virais/metabolismo , Animais , Células Cultivadas , Embrião de Galinha , Vírus Defeituosos/metabolismo , Fibroblastos , Mutação , Proteína Oncogênica pp60(v-src) , Fosforilação , Proteínas Quinases/metabolismo , Serina/metabolismo , TemperaturaRESUMO
The Rel family of proteins includes a number of proteins involved in transcriptional control, such as the retroviral oncoprotein v-Rel, c-Rel, the Drosophila melanogaster developmental protein Dorsal, and subunits of the transcription factor NF-kappa B. These proteins are related through a highly conserved domain of approximately 300 amino acids, called the Rel homology domain, that contains dimerization, DNA binding, and nuclear targeting functions. Also within the Rel homology domain, there is a conserved consensus sequence (Arg-Arg-Pro-Ser) for phosphorylation by cyclic AMP-dependent protein kinase (PKA). We used linker insertion mutagenesis and site-directed mutagenesis to determine the importance of this sequence for the transformation of avian spleen cells by v-Rel and the subcellular localization of c-Rel in chicken embryo fibroblasts (CEF). The insertion of 2 amino acids (Pro-Trp) within this sequence completely abolished transformation and transcriptional repression by v-Rel and resulted in a shift in the localization of c-Rel from cytoplasmic to nuclear in CEF. When the conserved Ser within the PKA recognition sequence was replaced by Ala, there was no significant effect on transformation and transcriptional repression by v-Rel or on cytoplasmic retention of c-Rel. However, when this Ser was changed to Asp or Glu, transformation and transcriptional repression by v-Rel were significantly inhibited and c-Rel showed a diffuse nuclear and cytoplasmic localization in CEF. Although a peptide containing the recognition sequence from v-Rel can be phosphorylated by PKA in vitro, this site is not constitutively phosphorylated to a high degree in vivo in transformed spleen cells incubated with okadaic acid. Our results indicate that the transforming and transcriptional repressing activities of v-Rel and the cytoplasmic retention of c-Rel are dependent on the structure of the conserved PKA recognition motif. In addition, they suggest that phosphorylation at the conserved PKA site could have a negative effect on transformation and transcriptional repression by v-Rel and induce the nuclear localization of c-Rel.
Assuntos
Transformação Celular Neoplásica , Proteínas Quinases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Oncogênicas de Retroviridae/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Western Blotting , Galinhas , Citoplasma/metabolismo , DNA , Dados de Sequência Molecular , Mutação , Proteínas Oncogênicas v-rel , Proteínas Proto-Oncogênicas/genética , Proteínas Proto-Oncogênicas c-rel , Especificidade por Substrato , Transcrição Gênica , TripsinaRESUMO
The mechanism by which the products of the v-rel oncogene, the corresponding c-rel proto-oncogene, and the related dorsal gene of Drosophila melanogaster exert their effects is not clear. Here we show that the v-rel, chicken c-rel, and dorsal proteins activated gene expression when fused to LexA sequences and bound to DNA upstream of target genes in Saccharomyces cerevisiae. We have defined two distinct activation regions in the c-rel protein. Region I, located in the amino-terminal half of rel and dorsal proteins, contains no stretches of glutamines, prolines, or acidic amino acids and therefore may be a novel activation domain. Lesions in the v-rel protein that diminished or abolished oncogenic transformation of avian spleen cells correspondingly affected transcription activation by region I. Region II, located in the carboxy terminus of the c-rel protein, is highly acidic. Region II is not present in the v-rel protein or in a transforming mutant derivative of the c-rel protein. Our results show that the oncogenicity of Rel proteins requires activation region I and suggest that the biological function of rel and dorsal proteins depends on transcription activation by this region.
Assuntos
Transformação Celular Neoplásica , Regulação da Expressão Gênica , Oncogenes , Proteínas Oncogênicas de Retroviridae/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Galinhas , Drosophila melanogaster/genética , Escherichia coli/genética , Dados de Sequência Molecular , Proteínas Oncogênicas v-rel , Fosfoproteínas/genética , Plasmídeos , Proto-Oncogenes , Mapeamento por Restrição , Saccharomyces cerevisiae/genética , Transcrição GênicaRESUMO
Extracts of bakers' yeast (Saccharomyces cerevisiae) contain protein-tyrosine kinase activity that can be detected with a synthetic Glu-Tyr copolymer as substrate (G. Schieven, J. Thorner, and G.S. Martin, Science 231:390-393, 1986). By using this assay in conjunction with ion-exchange and affinity chromatography, a soluble tyrosine kinase activity was purified over 8,000-fold from yeast extracts. The purified activity did not utilize typical substrates for mammalian protein-tyrosine kinases (enolase, casein, and histones). The level of tyrosine kinase activity at all steps of each preparation correlated with the content of a 40-kDa protein (p40). Upon incubation of the most highly purified fractions with Mn-ATP or Mg-ATP, p40 was the only protein phosphorylated on tyrosine. Immunoblotting of purified p40 or total yeast extracts with antiphosphotyrosine antibodies and phosphoamino acid analysis of 32P-labeled yeast proteins fractionated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that the 40-kDa protein is normally phosphorylated at tyrosine in vivo. 32P-labeled p40 immunoprecipitated from extracts of metabolically labeled cells by affinity-purified anti-p40 antibodies contained both phosphoserine and phosphotyrosine. The gene encoding p40 (YPK1) was cloned from a yeast genomic library by using oligonucleotide probes designed on the basis of the sequence of purified peptides. As deduced from the nucleotide sequence of YPK1, p40 is homologous to known protein kinases, with features that resemble known protein-serine kinases more than known protein-tyrosine kinases. Thus, p40 is a protein kinase which is phosphorylated in vivo and in vitro at both tyrosine and serine residues; it may be a novel type of autophosphorylating tyrosine kinase, a bifunctional (serine/tyrosine-specific) protein kinase, or a serine kinase that is a substrate for an associated tyrosine kinase.
Assuntos
Genes Fúngicos , Fosfoproteínas/genética , Proteínas Tirosina Quinases/genética , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Quinase 3 da Glicogênio Sintase , Humanos , Cinética , Dados de Sequência Molecular , Fragmentos de Peptídeos/isolamento & purificação , Fosfoproteínas/isolamento & purificação , Fosfoproteínas/metabolismo , Fosfotirosina , Proteínas Quinases/genética , Proteínas Tirosina Quinases/isolamento & purificação , Proteínas Tirosina Quinases/metabolismo , Saccharomyces cerevisiae/enzimologia , Homologia de Sequência do Ácido Nucleico , Serina Endopeptidases , Tirosina/análogos & derivados , Tirosina/análiseRESUMO
The avian Rev-T retrovirus encodes the v-Rel oncoprotein, which is a member of the Rel/NF-kappaB transcription factor family. v-Rel induces a rapidly fatal lymphoma/leukemia in young birds, and v-Rel can transform and immortalize a variety of avian cell types in vitro. Although Rel/NF-kappaB transcription factors have been associated with oncogenesis in mammals, v-Rel is the only member of this family that is frankly oncogenic in animal model systems. The potent oncogenicity of v-Rel is the consequence of a number of mutations that have altered its activity and regulation: for example, certain mutations decrease its ability to be regulated by IkappaBalpha, change its DNA-binding site specificity, and endow it with new transactivation properties. The study of v-Rel will continue to increase our knowledge of how cellular Rel proteins contribute to oncogenesis by affecting cell growth, altering cell-cycle regulation, and blocking apoptosis. This review will discuss biological and molecular activities of v-Rel, with particular attention to how these activities relate to structure - function aspects of the Rel/NF-kappaB transcription factors.
Assuntos
Transformação Celular Neoplásica , Proteínas Oncogênicas v-rel/fisiologia , Sequência de Aminoácidos , Animais , Apoptose , Divisão Celular , DNA/metabolismo , Humanos , Dados de Sequência Molecular , Mutação , Proteínas Oncogênicas v-rel/análise , Proteínas Oncogênicas v-rel/química , Processamento de Proteína Pós-Traducional , Proteínas Proto-Oncogênicas c-rel/análise , Relação Estrutura-Atividade , Ativação TranscricionalRESUMO
Rel/NF-kappaB transcription factors regulate several important physiological processes, including developmental processes, inflammation and immune responses, cell growth, cancer, apoptosis, and the expression of certain viral genes. Therefore, they have also been sought-after molecular targets for pharmacological intervention. As details of the Rel/NF-kappaB signal transduction pathway are revealed, it is clear that modulators of this pathway can act at several levels. Inhibitors of the Rel/NF-kappaB pathway include a variety of natural and designed molecules, including anti-oxidants, proteasome inhibitors, peptides, small molecules, and dominant-negative or constitutively active polypeptides in the pathway. Several of these molecules act as general inhibitors of Rel/NF-kappaB induction, whereas others inhibit specific pathways of induction. Inhibitors of Rel/NF-kappaB are likely to gain stature as treatments for certain cancers and neurodegenerative and inflammatory diseases.
Assuntos
NF-kappa B/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Animais , Antioxidantes/farmacologia , Cisteína Endopeptidases/fisiologia , DNA/metabolismo , Humanos , Complexos Multienzimáticos/fisiologia , Fosforilação , Complexo de Endopeptidases do Proteassoma , Proteínas Proto-Oncogênicas/metabolismo , Fator de Transcrição RelA , Fator de Transcrição RelB , Fatores de Transcrição/metabolismo , Ativação TranscricionalRESUMO
Apoptosis is a physiological process critical for organ development, tissue homeostasis, and elimination of defective or potentially dangerous cells in complex organisms. Apoptosis can be initiated by a wide variety of stimuli, which activate a cell suicide program that is constitutively present in most vertebrate cells. In diverse cell types, Rel/NF-kappaB transcription factors have been shown to have a role in regulating the apoptotic program, either as essential for the induction of apoptosis or, perhaps more commonly, as blockers of apoptosis. Whether Rel/NF-kappaB promotes or inhibits apoptosis appears to depend on the specific cell type and the type of inducer. An understanding of the role of Rel/NF-kappaB transcription factors in controlling apoptosis may lead to the development of therapeutics for a wide variety of human diseases, including neurodegenerative and immune diseases, and cancer.
Assuntos
Apoptose , NF-kappa B/fisiologia , Animais , Ciclo Celular , Humanos , Sistema Imunitário/fisiologia , Neoplasias/terapia , Neurônios/fisiologia , Fator de Transcrição RelA , Fator de Necrose Tumoral alfa/farmacologia , Replicação ViralRESUMO
The avian retroviral oncoprotein v-Rel and its cellular homolog c-Rel are members of a family of related site-specific DNA-binding proteins. Towards the carboxy-terminal end of the highly conserved Rel homology (RH) domain in the majority of Rel proteins, there is a consensus recognition sequence for protein kinase A (PK-A). We have investigated the importance of this sequence (Arg-Arg-Pro-Ser) for several functional properties of v-Rel and c-Rel. Disruption of the PK-A sequence by a two amino acid insertion between the arginine and the proline residues completely abolished the ability of v-Rel and c-Rel to bind a kappa B site in vitro. When the phosphorylatable serine in this sequence (Ser-275 in v-Rel, Ser-266 in c-Rel) was replaced by an alanine, DNA binding by v-Rel was not affected, whereas the ability of c-Rel to bind DNA was reduced approximately fourfold by this mutation. Similarly, a serine to tryptophan change greatly reduced the DNA-binding ability of c-Rel, whereas v-Rel was not appreciably affected by this change. When this serine was replaced by an acidic amino acid, DNA binding by v-Rel was reduced approximately twofold and the DNA-binding activity of c-Rel was nearly abolished. Glutaraldehyde cross-linking experiments indicated that mutations at the PK-A recognition site that reduced DNA binding also negatively affected protein oligomerization, which is likely to be responsible for the reduced ability of mutant v-Rel and c-Rel proteins to bind DNA. Domain-swapping experiments showed that structural differences between v-Rel and c-Rel in the central region of the proteins are primarily responsible for the higher sensitivity of c-Rel to a serine to alanine mutation in the PK-A site. One difference between v-Rel and c-Rel, a glutamine to alanine change in v-Rel located three amino acids carboxy-terminal to the PK-A phosphorylatable serine (Ala-278 in v-Rel; Glu-269 in c-Rel), is mainly responsible for the lack of an effect on DNA binding by v-Rel when Ser-275 is replaced by alanine. That is, a v-Rel double mutant (v-275A/278E) showed reduced DNA-binding and transforming abilities as compared with v-Rel and v-275A. Similarly, the mutations in c-Rel that affected DNA binding showed a corresponding effect on the ability of c-Rel proteins to activate transcription in yeast from a reporter gene containing upstream Rel binding sites.(ABSTRACT TRUNCATED AT 400 WORDS)