RESUMEN
The Toll/IL-1R-domain-containing adaptor protein SARM1 is expressed primarily in the brain, where it mediates axonal degeneration. Roles for SARM1 in TLR signaling, viral infection, inflammasome activation, and chemokine and Xaf1 expression have also been described. Much of the evidence for SARM1 function relies on SARM1-deficient mice generated in 129 ESCs and backcrossed to B6. The Sarm1 gene lies in a gene-rich region encompassing Xaf1 and chemokine loci, which remain 129 in sequence. We therefore generated additional knockout strains on the B6 background, confirming the role of SARM1 in axonal degeneration and WNV infection, but not in VSV or LACV infection, or in chemokine or Xaf1 expression. Sequence variation in proapoptotic Xaf1 between B6 and 129 results in coding changes and distinct splice variants, which may account for phenotypes previously attributed to SARM1. Reevaluation of phenotypes in these strains will be critical for understanding the function of SARM1.
Asunto(s)
Proteínas del Dominio Armadillo/genética , Proteínas del Dominio Armadillo/metabolismo , Proteínas del Citoesqueleto/genética , Proteínas del Citoesqueleto/metabolismo , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Animales , Apoptosis/genética , Proteínas Reguladoras de la Apoptosis/metabolismo , Axones/metabolismo , Encéfalo/metabolismo , Encefalitis de California/genética , Enfermedades Hereditarias del Ojo , Femenino , Virus La Crosse , Masculino , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Mutación/genética , Fenotipo , Receptores de Interleucina-1/metabolismo , Degeneración Retiniana , Transducción de Señal/genética , Receptores Toll-Like/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Trastornos de la Visión , Fiebre del Nilo Occidental/genéticaRESUMEN
Four of the five members of the Toll/IL-1R domain-containing adaptor family are required for signaling downstream of TLRs, promoting innate immune responses against different pathogens. However, the role of the fifth member of this family, sterile α and Toll/IL-1R domain-containing 1 (SARM), is unclear. SARM is expressed primarily in the CNS where it is required for axonal death. Studies in Caenorhabditis elegans have also shown a role for SARM in innate immunity. To clarify the role of mammalian SARM in innate immunity, we infected SARM(-/-) mice with a number of bacterial and viral pathogens. SARM(-/-) mice show normal responses to Listeria monocytogenes, Mycobacterium tuberculosis, and influenza virus, but show dramatic protection from death after CNS infection with vesicular stomatitis virus. Protection correlates with reduced CNS injury and cytokine production by nonhematopoietic cells, suggesting that SARM is a positive regulator of cytokine production. Neurons and microglia are the predominant source of cytokines in vivo, supporting a role for SARM as a link between neuronal injury and innate immunity.
Asunto(s)
Proteínas del Dominio Armadillo/metabolismo , Enfermedades Virales del Sistema Nervioso Central/inmunología , Proteínas del Citoesqueleto/metabolismo , Infecciones por Rhabdoviridae/inmunología , Virus de la Estomatitis Vesicular Indiana/inmunología , Animales , Proteínas del Dominio Armadillo/genética , Proteínas del Dominio Armadillo/inmunología , Células de la Médula Ósea , Células Cultivadas , Sistema Nervioso Central/metabolismo , Sistema Nervioso Central/microbiología , Sistema Nervioso Central/virología , Enfermedades Virales del Sistema Nervioso Central/metabolismo , Citocinas/biosíntesis , Proteínas del Citoesqueleto/genética , Proteínas del Citoesqueleto/inmunología , Inmunidad Innata , Virus de la Influenza A/inmunología , Listeria monocytogenes/inmunología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Microglía/metabolismo , Mycobacterium tuberculosis/inmunología , Neuronas/metabolismo , Infecciones por Rhabdoviridae/metabolismoRESUMEN
Axonal and synaptic degeneration is a hallmark of peripheral neuropathy, brain injury, and neurodegenerative disease. Axonal degeneration has been proposed to be mediated by an active autodestruction program, akin to apoptotic cell death; however, loss-of-function mutations capable of potently blocking axon self-destruction have not been described. Here, we show that loss of the Drosophila Toll receptor adaptor dSarm (sterile α/Armadillo/Toll-Interleukin receptor homology domain protein) cell-autonomously suppresses Wallerian degeneration for weeks after axotomy. Severed mouse Sarm1 null axons exhibit remarkable long-term survival both in vivo and in vitro, indicating that Sarm1 prodegenerative signaling is conserved in mammals. Our results provide direct evidence that axons actively promote their own destruction after injury and identify dSarm/Sarm1 as a member of an ancient axon death signaling pathway.
Asunto(s)
Proteínas del Dominio Armadillo/genética , Proteínas del Dominio Armadillo/fisiología , Axones/fisiología , Proteínas del Citoesqueleto/genética , Proteínas del Citoesqueleto/fisiología , Proteínas de Drosophila/genética , Proteínas de Drosophila/fisiología , Neuronas/fisiología , Degeneración Walleriana , Animales , Animales Modificados Genéticamente , Apoptosis , Proteínas del Dominio Armadillo/análisis , Axones/ultraestructura , Axotomía , Supervivencia Celular , Células Cultivadas , Proteínas del Citoesqueleto/análisis , Desnervación , Drosophila/embriología , Drosophila/genética , Drosophila/fisiología , Proteínas de Drosophila/análisis , Ratones , Mutación , Nervio Ciático/lesiones , Nervio Ciático/fisiología , Transducción de Señal , Ganglio Cervical Superior/citología , Técnicas de Cultivo de TejidosRESUMEN
The mammalian Nit1 protein is homologous to plant and bacterial nitrilases. In flies and worms, Nit1 is fused to the 5' end of Fhit, suggesting that Nit1 may functionally interact with the Fhit pathway. Fhit has been shown to play a role of a tumor suppressor. Somatic loss of Fhit in human tissues is associated with a wide variety of cancers. Deletion of Fhit results in a predisposition to induced and spontaneous tumors in mice. It has been suggested that Nit1 collaborates with Fhit in tumor suppression. Similar to mice lacking Fhit, Nit1-deficient mice are more sensitive to carcinogen-induced tumors. It was previously shown that ectopic expression of Nit1 or Fhit led to caspase activation and apoptosis, and that both proteins may play a role in DNA damage-induced apoptosis. In this study, we analyzed the physiological function of Nit1 in T cells using Nit1-knockout mice. Nit1-deficient T cells can undergo apoptosis induced by DNA damage due to irradiation and chemical treatment. However, apoptosis induced by Fas or Ca(++) signals appeared to be compromised. Additionally, Nit1 deficiency resulted in T cell hyperproliferative responses induced by TCR stimulation. The expressions of T cell activation markers were elevated in Nit1(-/-) T cells. There was a spontaneous cell cycle entry and enhanced cell cycle progression in Nit1(-/-) T cells. These data indicate that Nit1 is a novel negative regulator in primary T cells.
Asunto(s)
Aminohidrolasas/inmunología , Proliferación Celular , Retroalimentación Fisiológica , Linfocitos T/metabolismo , Aminohidrolasas/genética , Aminohidrolasas/metabolismo , Animales , Apoptosis/genética , Apoptosis/inmunología , Calcio/inmunología , Calcio/metabolismo , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/inmunología , Proteínas de Ciclo Celular/metabolismo , Daño del ADN/inmunología , Activación de Linfocitos , Ratones , Ratones Noqueados , Homología de Secuencia de Aminoácido , Linfocitos T/inmunología , Linfocitos T/patología , Timo/patología , Receptor fas/inmunología , Receptor fas/metabolismoRESUMEN
The Fas-associated death domain protein (FADD)/Mort1 is a signaling adaptor protein which mediates the activation of caspase 8 during death receptor-induced apoptosis. Disruption of FADD in germ cells results in death receptor-independent embryonic lethality in mice. Previous studies indicated that in addition to its function in apoptosis, FADD is also required in peripheral T cell homeostasis and TCR-induced proliferative responses. In this report, we generated B cell-specific FADD-deficient mice and showed that deletion of FADD at the pro-B cell stage had minor effects on B cell development in the bone marrow, and resulted in increased splenic and lymph node B cell numbers and decreased peritoneal B1 cell numbers. As in T cells, a FADD deficiency inhibited Fas-induced apoptosis in B cells. However, B cell-proliferative responses induced by stimulation of the BCR and CD40 using anti-IgM or anti-CD40 Abs were unaffected by the absence of FADD. Further analyses revealed that FADD-deficient B cells were defective in proliferative responses induced by treatments with dsRNA and LPS which stimulate TLR3 and TLR4, respectively. Therefore, in addition to its apoptotic function, FADD also plays a role in TLR3- and TLR4-induced proliferative responses in B cells.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/fisiología , Apoptosis/inmunología , Linfocitos B/citología , Linfocitos B/inmunología , Proliferación Celular , Receptor Toll-Like 3/fisiología , Receptor Toll-Like 4/fisiología , Proteínas Adaptadoras Transductoras de Señales/deficiencia , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Antígenos de Diferenciación de Linfocitos B/biosíntesis , Linfocitos B/metabolismo , Diferenciación Celular/genética , Diferenciación Celular/inmunología , Células Cultivadas , Epítopos de Linfocito B/inmunología , Proteína de Dominio de Muerte Asociada a Fas , Inmunoglobulina D/sangre , Inmunoglobulina M/sangre , Lipopolisacáridos/farmacología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Transducción de Señal/genética , Transducción de Señal/inmunologíaRESUMEN
Fas-associated death domain protein (FADD)/mediator of receptor-induced toxicity-1 is required for signaling induced by death receptors such as Fas. In earlier studies, FADD-deficient mice died in utero, and a FADD deficiency in embryonic stem cells inhibited T cell production in viable FADD-/- -->RAG-1-/- chimeras. To analyze the temporal requirement of FADD in the development and function in the T lineage, it is necessary to establish viable mutant mice producing detectable FADD-deficient T cells. We generated mice that express a functional FADD:GFP fusion gene reconstituting normal embryogenesis and lymphopoiesis in the absence of the endogenous FADD. Efficient T cell-specific deletion of FADD:GFP was achieved, as indicated by the presence of a high percentage of GFP-negative thymocytes and peripheral T cells in mice expressing Lck-Cre or CD4-Cre. Sorted GFP-negative thymocytes and peripheral T cells contained undetectable levels of FADD and were resistant to apoptosis induced by Fas, TNF, and TCR restimulation. These T cell-specific FADD-deficient mice contain normal thymocyte numbers, but fewer peripheral T cells. Purified peripheral FADD-deficient T cells failed to undergo extensive homeostatic expansion after adoptive transfer into lymphocyte-deficient hosts, and responded poorly to proliferation induced by ex vivo TCR stimulation. Furthermore, deletion of FADD in preactivated mature T cells using retrovirus-Cre resulted in no proliferation. These results demonstrate that FADD plays a dispensable role during thymocyte development, but is essential in maintaining peripheral T cell homeostasis and regulating both apoptotic and proliferation signals.