RESUMO
The Tumour Necrosis Factor (TNF) Receptor-associated factor-1 (TRAF1) adaptor protein is a key component in initiating intracellular signalling pathways downstream of TNF receptors (TNFR). More importantly, TRAF1 has a pattern of expression restricted primarily to lymphoid cells and plays an important role in lymphocyte survival. TRAF1 has been shown to be phosphorylated on Serine 139, consequently inhibiting NF-κB activation downstream of TNFR2 when expressed in HeLa cells. We have previously demonstrated that TRAF1 cooperates with the TNFR family member 4-1BB to mediate signalling in T cells. However, the impact of TRAF1 phosphorylation on events downstream of 4-1BB in T cells remained to be defined. Using a proteomics approach we demonstrate that TANK-binding kinase 1 (TBK1) preferentially associates with the TRAF1 Serine 139 to Alanine (S139A) mutant. TBK1 is a kinase that functions upstream of NIK and IKK in the activation of the NF-κB pathway. When TRAF1-deficient CD8 T cells were reconstituted with the TRAF1 S139A mutant, we observed more sustained levels of IκBα degradation in response to 4-1BB stimulation in contrast to cells expressing either TRAF1 wild-type or TRAF1 S139D phospho-mimetic mutant. Together, these findings define the importance of the basal phosphorylation state of the TRAF1 Serine 139 residue in coordinating signalling events downstream of 4-1BB in primary T cells.
Assuntos
NF-kappa B/metabolismo , Linfócitos T/metabolismo , Fator 1 Associado a Receptor de TNF/metabolismo , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismo , Animais , Proteínas I-kappa B , Camundongos , Camundongos Mutantes , Inibidor de NF-kappaB alfa , Fosforilação , Proteínas Serina-Treonina Quinases/metabolismo , Serina/metabolismo , Transdução de Sinais , Fator 1 Associado a Receptor de TNF/genéticaRESUMO
Coordinated development of excitatory and inhibitory synapses is essential for higher brain function, and impairment in this development is associated with neuropsychiatric disorders. In contrast to the large body of accumulated evidence regarding excitatory synapse development, little is known about synaptic adhesion and organization mechanisms underlying inhibitory synapse development. Through unbiased expression screens and proteomics, we identified immunoglobulin superfamily member 21 (IgSF21) as a neurexin2α-interacting membrane protein that selectively induces inhibitory presynaptic differentiation. IgSF21 localizes postsynaptically and recruits axonal neurexin2α in a trans-interaction manner. Deleting IgSF21 in mice impairs inhibitory presynaptic organization, especially in the hippocampal CA1 stratum radiatum, and also diminishes GABA-mediated synaptic transmission in hippocampal CA1 neurons without affecting their excitatory synapses. Finally, mice lacking IgSF21 show a sensorimotor gating deficit. These findings suggest that IgSF21 selectively regulates inhibitory presynaptic differentiation through interacting with presynaptic neurexin2α and plays a crucial role in synaptic inhibition in the brain.Molecular mechanisms regulating the development of inhibitory synapses are poorly understood. Here the authors show that IgSF21 interacts with neurexin2α to induce presynaptic differentiation of inhibitory synapses, and that mice lacking IgSF21 exhibit deficits in inhibitory synaptic transmission.
Assuntos
Proteínas de Membrana/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Sinapses/metabolismo , Transmissão Sináptica , Animais , Encéfalo/metabolismo , Células COS , Adesão Celular , Diferenciação Celular , Chlorocebus aethiops , Deleção de Genes , Células HEK293 , Hipocampo/metabolismo , Homozigoto , Humanos , Masculino , Camundongos , Neurônios/metabolismo , Plasmídeos/metabolismo , Ligação Proteica , Isoformas de Proteínas , Ratos , Receptores Pré-Sinápticos/metabolismoRESUMO
4-1BB is a member of the TNFR superfamily, which contributes to the activation of signaling pathways required for the survival of activated and memory T cells. We have shown previously that TRAF1, an adaptor protein recruited to 4-1BB, is required for 4-1BB-mediated CD8 T cell survival in vivo. With the use of a proteomics approach in primary T cells, we have identified LSP1 as a novel protein recruited to the 4-1BB signaling complex in a TRAF1-dependent manner. Further characterization of the interaction between TRAF1 and LSP1 revealed that LSP1 requires the TRAF-N domain of TRAF1 for direct association. Similarly to TRAF1(-/-) T cells, LSP1(-/-) T cells exhibit impaired ERK activation following stimulation through 4-1BB and consequently, are unable to down-modulate expression of the proapoptotic Bcl-2 family member Bim. Moreover, we demonstrate that the absence of LSP1 expression leads to defective expansion and survival of T cells in response to 4-1BB stimulation. Thus, we have identified LSP1 as a new mediator involved in 4-1BB signaling and T cell survival. Collectively, our work shows that TRAF1 and LSP1 cooperate downstream of 4-1BB to activate ERK signaling and down-modulate the levels of Bim leading to enhanced T cell survival.