RESUMEN
Signal transduction pathways are modular composites of functionally interdependent sets of proteins that act in a coordinated fashion to transform environmental information into a phenotypic response. The pro-inflammatory cytokine tumour necrosis factor (TNF)-alpha triggers a signalling cascade, converging on the activation of the transcription factor NF-kappa B, which forms the basis for numerous physiological and pathological processes. Here we report the mapping of a protein interaction network around 32 known and candidate TNF-alpha/NF-kappa B pathway components by using an integrated approach comprising tandem affinity purification, liquid-chromatography tandem mass spectrometry, network analysis and directed functional perturbation studies using RNA interference. We identified 221 molecular associations and 80 previously unknown interactors, including 10 new functional modulators of the pathway. This systems approach provides significant insight into the logic of the TNF-alpha/NF-kappa B pathway and is generally applicable to other pathways relevant to human disease.
Asunto(s)
Proteínas de Drosophila , FN-kappa B/metabolismo , Transducción de Señal/fisiología , Factor de Necrosis Tumoral alfa/metabolismo , Animales , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Línea Celular , Chaperoninas , Cromatografía de Afinidad/métodos , Activación Enzimática , Proteínas HSP90 de Choque Térmico/genética , Proteínas HSP90 de Choque Térmico/metabolismo , Humanos , Proteínas I-kappa B/aislamiento & purificación , Proteínas I-kappa B/metabolismo , MAP Quinasa Quinasa Quinasa 3 , Quinasas Quinasa Quinasa PAM/genética , Quinasas Quinasa Quinasa PAM/metabolismo , Sustancias Macromoleculares , Espectrometría de Masas/métodos , Modelos Biológicos , Chaperonas Moleculares/genética , Chaperonas Moleculares/metabolismo , FN-kappa B/genética , FN-kappa B/aislamiento & purificación , Proteoma/análisis , Interferencia de ARN , Receptores del Factor de Necrosis Tumoral/metabolismo , Proteínas de Unión a Tacrolimus/genética , Proteínas de Unión a Tacrolimus/metabolismo , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/aislamiento & purificación , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismoRESUMEN
Benzimidazole-N-oxide modifications of potent lipophilic dihydrofolate reductase (DHFR) inhibitors (e.g., methylbenzoprim 1 and dichlorobenzoprim 2) have been prepared by base-promoted cyclization of the nitrophenylbenzylamino groups to explore the possibility that abrogation of DHFR-inhibitory activity might reveal clues to an alternative anti-ras mechanism. Examples of the new series had only low growth inhibitory activities (GI(50) generally >50 microM) against colon HCT116 and lung HT29 cell lines but, unlike methylbenzoprim, this activity was unaffected by hypoxanthine/thymidine rescue.