RESUMO
Recently discovered type III IFNs (IFN-lambda) exert their antiviral and immunomodulatory activities through a unique receptor complex composed of IFN-lambdaR1 and interleukin-10 receptor 2. To further study type III IFNs, we cloned and characterized mouse IFN-lambda ligand-receptor system. We showed that, similar to their human orthologues, mIFN-lambda2 and mIFN-lambda3 signal through the IFN-lambda receptor complex, activate IFN stimulated gene factor 3, and are capable of inducing antiviral protection and MHC class I antigen expression in several cell types including B16 melanoma cells. We then used the murine B16 melanoma model to investigate the potential antitumor activities of IFN-lambdas. We developed B16 cells constitutively expressing murine IFN-lambda2 (B16.IFN-lambda2 cells) and evaluated their tumorigenicity in syngeneic C57BL/6 mice. Although constitutive expression of mIFN-lambda2 in melanoma cells did not affect their proliferation in vitro, the growth of B16.IFN-lambda2 cells, when injected s.c. into mice, was either retarded or completely prevented. We found that rejection of the modified tumor cells correlated with their level of IFN-lambda2 expression. We then developed IFN-lambda-resistant B16.IFN-lambda2 cells (B16.IFN-lambda2Res cells) and showed that their tumorigenicity was also highly impaired or completely abolished similar to B16.IFN-lambda2 cells, suggesting that IFN-lambdas engage host mechanisms to inhibit melanoma growth. These in vivo experiments show the antitumor activities of IFN-lambdas and suggest their strong therapeutic potential.
Assuntos
Interferons/imunologia , Melanoma Experimental/imunologia , Melanoma Experimental/terapia , Receptores de Interferon/imunologia , Sequência de Aminoácidos , Animais , Células COS , Chlorocebus aethiops , Clonagem Molecular , Cricetinae , Feminino , Humanos , Interferons/genética , Ligantes , Melanoma Experimental/genética , Camundongos , Camundongos Endogâmicos C57BL , Dados de Sequência Molecular , Receptores de Interferon/genética , Transdução de Sinais , TransfecçãoRESUMO
The receptor for IL-26 (AK155), a cytokine of the IL-10 family, has not previously been defined. We demonstrate that the active receptor complex for IL-26 is a heterodimer composed of two receptor proteins: IL-20R1 and IL-10R2. Signaling through the IL-26R results in activation of STAT1 and STAT3 which can be blocked by neutralizing Abs against IL-20R1 or IL-10R2. IL-10R2 is broadly expressed on a wide variety of tissues, whereas only a limited number of tissues express IL-20R1. Therefore, the ability to respond to IL-26 is restricted by the expression of IL-20R1. IL-10, IL-19, IL-20, IL-22, and IL-24 fail to signal through the combination of IL-10R2 and IL-20R1 proteins, demonstrating that this receptor combination is unique and specific for IL-26.
Assuntos
Interleucina-10/metabolismo , Interleucinas/metabolismo , Interleucinas/fisiologia , Receptores de Interleucina/fisiologia , Transdução de Sinais/imunologia , Linhagem Celular , Linhagem Celular Tumoral , Proteínas de Ligação a DNA/metabolismo , Dimerização , Células HT29 , Humanos , Especificidade de Órgãos/genética , Especificidade de Órgãos/imunologia , Isoformas de Proteínas/biossíntese , Isoformas de Proteínas/genética , Isoformas de Proteínas/fisiologia , Receptores de Interleucina/biossíntese , Receptores de Interleucina/genética , Receptores de Interleucina-10 , Fator de Transcrição STAT1 , Fator de Transcrição STAT3 , Transativadores/metabolismoRESUMO
Previous studies have demonstrated two different biosynthetic pathways to quinolinate, the universal de novo precursor to the pyridine ring of NAD. In prokaryotes, quinolinate is formed from aspartate and dihydroxyacetone phosphate; in eukaryotes, it is formed from tryptophan. It has been generally believed that the tryptophan to quinolinic acid biosynthetic pathway is unique to eukaryotes; however, this paper describes the use of comparative genome analysis to identify likely candidates for all five genes involved in the tryptophan to quinolinic acid pathway in several bacteria. Representative examples of each of these genes were overexpressed, and the predicted functions are confirmed in each case using unambiguous biochemical assays.
Assuntos
Bactérias/enzimologia , Bactérias/genética , NAD/biossíntese , Ácido Quinolínico/metabolismo , Triptofano/metabolismo , Arilformamidase/genética , Arilformamidase/metabolismo , Bactérias/metabolismo , Cromossomos Bacterianos/genética , Regulação Bacteriana da Expressão Gênica , Genes Bacterianos/genética , Teste de Complementação Genética , Hidrolases/genética , Hidrolases/metabolismo , Cinética , Dados de Sequência Molecular , NAD/química , Oxirredutases/genética , Oxirredutases/metabolismo , Ácido Quinolínico/química , Triptofano/químicaRESUMO
Novel drug targets are required in order to design new defenses against antibiotic-resistant pathogens. Comparative genomics provides new opportunities for finding optimal targets among previously unexplored cellular functions, based on an understanding of related biological processes in bacterial pathogens and their hosts. We describe an integrated approach to identification and prioritization of broad-spectrum drug targets. Our strategy is based on genetic footprinting in Escherichia coli followed by metabolic context analysis of essential gene orthologs in various species. Genes required for viability of E. coli in rich medium were identified on a whole-genome scale using the genetic footprinting technique. Potential target pathways were deduced from these data and compared with a panel of representative bacterial pathogens by using metabolic reconstructions from genomic data. Conserved and indispensable functions revealed by this analysis potentially represent broad-spectrum antibacterial targets. Further target prioritization involves comparison of the corresponding pathways and individual functions between pathogens and the human host. The most promising targets are validated by direct knockouts in model pathogens. The efficacy of this approach is illustrated using examples from metabolism of adenylate cofactors NAD(P), coenzyme A, and flavin adenine dinucleotide. Several drug targets within these pathways, including three distantly related adenylyltransferases (orthologs of the E. coli genes nadD, coaD, and ribF), are discussed in detail.
