RESUMEN
Endoplasmic reticulum (ER) stress activates the Unfolded Protein Response, a compensatory signaling response that is mediated by the IRE-1, PERK/PEK-1, and ATF-6 pathways in metazoans. Genetic studies have implicated roles for UPR signaling in animal development and disease, but the function of the UPR under physiological conditions, in the absence of chemical agents administered to induce ER stress, is not well understood. Here, we show that in Caenorhabditis elegans XBP-1 deficiency results in constitutive ER stress, reflected by increased basal levels of IRE-1 and PEK-1 activity under physiological conditions. We define a dynamic, temperature-dependent requirement for XBP-1 and PEK-1 activities that increases with immune activation and at elevated physiological temperatures in C. elegans. Our data suggest that the negative feedback loops involving the activation of IRE-1-XBP-1 and PEK-1 pathways serve essential roles, not only at the extremes of ER stress, but also in the maintenance of ER homeostasis under physiological conditions.
Asunto(s)
Proteínas de Caenorhabditis elegans/genética , Caenorhabditis elegans/crecimiento & desarrollo , Caenorhabditis elegans/inmunología , Proteínas Portadoras/genética , Proteínas de Ciclo Celular/genética , Inmunidad/genética , MAP Quinasa Quinasa 1/genética , Proteínas Serina-Treonina Quinasas/genética , Animales , Caenorhabditis elegans/genética , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas Portadoras/metabolismo , Proteínas de Ciclo Celular/metabolismo , Estrés del Retículo Endoplásmico/efectos de los fármacos , Estrés del Retículo Endoplásmico/genética , Retroalimentación Fisiológica , Regulación del Desarrollo de la Expresión Génica , Homeostasis/genética , Homeostasis/fisiología , Larva/genética , Larva/crecimiento & desarrollo , MAP Quinasa Quinasa 1/metabolismo , Proteínas Mutantes/genética , Proteínas Mutantes/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Transducción de Señal , Temperatura , Tunicamicina/farmacología , Respuesta de Proteína Desplegada/genética , Respuesta de Proteína Desplegada/fisiologíaRESUMEN
mTOR complex 2 (mTORC2) contains the mammalian target of rapamycin (mTOR) kinase and the Rictor regulatory protein and phosphorylates Akt. Whether this function of mTORC2 is critical for cancer progression is unknown. Here, we show that transformed human prostate epithelial cells lacking PTEN require mTORC2 to form tumors when injected into nude mice. Furthermore, we find that Rictor is a haploinsufficient gene and that deleting one copy protects Pten heterozygous mice from prostate cancer. Finally, we show that the development of prostate cancer caused by Pten deletion specifically in prostate epithelium requires mTORC2, but that for normal prostate epithelial cells, mTORC2 activity is nonessential. The selective requirement for mTORC2 in tumor development suggests that mTORC2 inhibitors may be of substantial clinical utility.
Asunto(s)
Proteínas Portadoras/metabolismo , Fosfohidrolasa PTEN/metabolismo , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Neoplasias de la Próstata/fisiopatología , Proteínas Quinasas/metabolismo , Factores de Transcripción/metabolismo , Animales , Proteínas Portadoras/genética , Transformación Celular Neoplásica , Células Cultivadas , Células Epiteliales/patología , Células Epiteliales/fisiología , Fibroblastos/citología , Fibroblastos/fisiología , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Trasplante de Neoplasias , Fosfohidrolasa PTEN/genética , Fenotipo , Fosfotransferasas (Aceptor de Grupo Alcohol)/genética , Próstata/citología , Próstata/metabolismo , Próstata/patología , Neoplasias de la Próstata/patología , Proteínas Quinasas/genética , Proteínas Serina-Treonina Quinasas/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Piruvato Deshidrogenasa Quinasa Acetil-Transferidora , Interferencia de ARN , Proteína Asociada al mTOR Insensible a la Rapamicina , Transducción de Señal/fisiología , Serina-Treonina Quinasas TOR , Factores de Transcripción/genética , Trasplante HeterólogoRESUMEN
Inward rectifier potassium channels (Kir) play critical roles in cell physiology. Despite representing the simplest tetrameric potassium channel structures, the pharmacology of this channel family remains largely undeveloped. In this respect, tertiapin (TPN), a 21 amino acid peptide isolated from bee venom, has been reported to inhibit Kir1.1 and Kir3.1/3.4 channels with high affinity by binding to the M1-M2 linker region of these channels. The features of the peptide-channel interaction have been explored electrophysiologically, and these studies have identified ways by which to alter the composition of the peptide without affecting its biological activity. In the present study, the TPN derivative, TPN-Y1/K12/Q13, has been synthesized and radiolabeled to high specific activity with (125)I. TPN-Y1/K12/Q13 and mono-iodo-TPN-Y1/K12/Q13 ([(127)I]TPN-Y1/K12/Q13) inhibit with high affinity rat but not human Kir1.1 channels stably expressed in HEK293 cells. [(125)I]TPN-Y1/K12/Q13 binds in a saturable, time-dependent, and reversible manner to HEK293 cells expressing rat Kir1.1, as well as to membranes derived from these cells, and the pharmacology of the binding reaction is consistent with peptide binding to Kir1.1 channels. Studies using chimeric channels indicate that the differences in TPN sensitivity between rat and human Kir1.1 channels are due to the presence of two nonconserved residues within the M1-M2 linker region. When these results are taken together, they demonstrate that [(125)I]TPN-Y1/K12/Q13 represents the first high specific activity radioligand for studying rat Kir1.1 channels and suggest its utility for identifying other Kir channel modulators.