RESUMO
The regulation of cytokine signaling is critical for controlling cellular proliferation and activation during an immune response. SOCS-1 is a potent inhibitor of Jak kinase activity and of signaling initiated by several cytokines. SOCS-1 protein levels are tightly regulated, and recent data suggest that SOCS-1 may regulate the protein levels of some signaling proteins by the ubiquitin proteasome pathway; however, the cellular mechanism by which SOCS-1 directs proteins for degradation is unknown. In this report, SOCS-1 is found to colocalize and biochemically copurify with the microtubule organizing complex (MTOC) and its associated 20S proteasome. The SOCS-1 SH2 domain is required for the localization of SOCS-1 to the MTOC. Overexpression of SOCS-1 targets Jak1 in an SH2-dependent manner to a perinuclear distribution resembling the MTOC-associated 20S proteasome. Analysis of MTOCs fractionated from SOCS-1-deficient cells demonstrates that SOCS-1 may function redundantly to regulate the localization of Jak1 to the MTOC. Nocodazole inhibits the protein turnover of SOCS-1, demonstrating that the minus-end transport of SOCS-1 to the MTOC-associated 20S proteasome is required to regulate SOCS-1 protein levels. These data link SOCS-1 directly with the proteasome pathway and suggest another function for the SH2 domain of SOCS-1 in the regulation of Jak/STAT signaling.
Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Centro Organizador dos Microtúbulos/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteínas Repressoras/metabolismo , Animais , Fracionamento Celular , Linhagem Celular , Centrossomo/química , Centrossomo/metabolismo , Chlorocebus aethiops , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Janus Quinase 1 , Microtúbulos/metabolismo , Estrutura Terciária de Proteína , Proteínas Tirosina Quinases/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Proteínas Repressoras/genética , Frações Subcelulares/metabolismo , Proteína 1 Supressora da Sinalização de Citocina , Proteínas Supressoras da Sinalização de CitocinaRESUMO
Recent studies have suggested that signaling initiated by the activation of Ag receptors and signaling activated through cytokine receptors may be regulated by a common set of inhibitory proteins. Suppressor of cytokine signaling 3 (SOCS-3), which has previously been demonstrated to inhibit cytokine signaling, is induced on TCR ligation. Overexpression of SOCS-3 can inhibit transcription driven by the IL-2 promoter in response to T cell activation. This inhibitory activity correlates with the suppression of calcineurin-dependent dephosphorylation and activation of the IL-2 promoter binding transcription factor, NFATp. Infection of primary murine T cells with a retrovirus encoding SOCS-3 blocks their IL-2 production in response to activation. Interestingly, SOCS-3 was found to coimmunoprecipitate with the catalytic subunit of calcineurin. These studies suggest that SOCS-3 may regulate T cell function as part of a negative feedback loop.
Assuntos
Proteínas de Ligação a DNA/metabolismo , Proteínas Nucleares , Proteínas/fisiologia , Proteínas Repressoras , Linfócitos T/imunologia , Fatores de Transcrição/metabolismo , Animais , Calcineurina/metabolismo , Células Cultivadas , Proteínas de Ligação a DNA/antagonistas & inibidores , Humanos , Interleucina-2/genética , Células Jurkat , Camundongos , Camundongos Endogâmicos C57BL , Fatores de Transcrição NFATC , Fosforilação , Regiões Promotoras Genéticas , Proteínas/genética , RNA Mensageiro/biossíntese , Receptores de Antígenos de Linfócitos T/metabolismo , Proteína 3 Supressora da Sinalização de Citocinas , Proteínas Supressoras da Sinalização de Citocina , Fatores de Transcrição/antagonistas & inibidores , Transcrição Gênica , Regulação para CimaRESUMO
Members of the suppressor of cytokine signaling (SOCS) family of signaling molecules regulate the activation of cytokine signaling. Experimental evidence indicates that SOCS expression is induced by cytokines and pro-inflammatory stimuli and is controlled at both the transcriptional and post-transcriptional levels. SOCS proteins are unstable and seem to be rapidly degraded by proteasomal pathways. However, the mechanisms by which SOCS protein levels are regulated remain unclear. Here, we show that TRIM8/GERP, a RING finger protein, interacts with SOCS-1 in vitro and in vivo. TRIM8/GERP, previously identified as a new member of the family of proteins containing a tripartite motif (TRIM), is a 551-amino acid RING finger protein conserved across species. TRIM8/GERP expression can be induced by interferon-gamma in epithelial and lymphoid cells. Coexpression of TRIM8/GERP with SOCS-1 decreases SOCS-1 protein stability and levels. Functionally, expression of TRIM8/GERP decreases the repression of interferon-gamma signaling mediated by SOCS-1. These data suggest that TRIM8/GERP may be a regulator of SOCS-1 function.
Assuntos
Proteínas de Transporte/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular , Proteínas do Tecido Nervoso/metabolismo , Proteínas Repressoras , Sequência de Aminoácidos , Anticorpos , Proteínas de Transporte/química , Proteínas de Transporte/genética , Clonagem Molecular , Sequência Conservada , Humanos , Dados de Sequência Molecular , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Transdução de Sinais , Proteína 1 Supressora da Sinalização de Citocina , Proteínas Supressoras da Sinalização de Citocina , Transfecção , Dedos de ZincoRESUMO
The pim family of proto-oncogenes encodes three serine-threonine kinases that have been implicated in the development of malignancies in mice and in humans. Expression of the Pim protein kinases is tightly regulated at the transcriptional, post-transcriptional, and translational levels. Dysregulation of pim transcription and pim mRNA stability have been implicated in Pim-mediated transformation. The data presented herein demonstrate that expression of the Pim kinases is additionally regulated at the post-translational level, by the serine-threonine phosphatase protein phosphatase 2A (PP2A). The catalytic subunit of PP2A associates with the Pim kinases in vivo, and the Pim kinases are substrates of PP2A phosphatase activity in vitro. Furthermore, overexpression of PP2A reduces the levels of the Pim proteins, whereas inhibition of PP2A activity by the protein phosphatase inhibitor okadaic acid stabilizes the Pim proteins. Finally, the effects of PP2A on the expression of the Pim proteins can affect Pim function. Taken together, these data suggest that PP2A activity is important for the regulation of the stability and function of the Pim kinases.
Assuntos
Fosfoproteínas Fosfatases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Domínio Catalítico/fisiologia , Linhagem Celular , Estabilidade Enzimática , Humanos , Proteína Fosfatase 2 , Processamento de Proteína Pós-Traducional , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas c-pim-1RESUMO
Studies of SOCS-1-deficient mice have implicated Socs-1 in the suppression of JAK-STAT (Janus tyrosine kinase-signal transducers and activators of transcription) signaling and T cell development. It has been suggested that the levels of Socs-1 protein may be regulated through the proteasome pathway. Here we show that Socs-1 interacts with members of the Pim family of serine/threonine kinases in thymocytes. Coexpression of the Pim kinases with Socs-1 results in phosphorylation and stabilization of the Socs-1 protein. The protein levels of Socs-1 are significantly reduced in the Pim-1(-/-), Pim-2(-/-) mice as compared with wild-type mice. Similar to Socs-1(-/-) mice, thymocytes from Pim-1(-/-), Pim-2(-/-) mice showed prolonged Stat6 phosphorylation upon IL-4 stimulation. These data suggest that the Pim kinases may regulate cytokine-induced JAK-STAT signaling through modulation of Socs-1 protein levels.