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1.
PLoS Pathog ; 17(2): e1009317, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33600488

RESUMO

The transmembrane protein 33 (TMEM33) was originally identified as an endoplasmic reticulum (ER) protein that influences the tubular structure of the ER and modulates intracellular calcium homeostasis. However, the role of TMEM33 in antiviral immunity in vertebrates has not been elucidated. In this article, we demonstrate that zebrafish TMEM33 is a negative regulator of virus-triggered interferon (IFN) induction via two mechanisms: mitochondrial antiviral signaling protein (MAVS) ubiquitination and a decrease in the kinase activity of TANK binding kinase 1 (TBK1). Upon stimulation with viral components, tmem33 was remarkably upregulated in the zebrafish liver cell line. The IFNφ1 promoter (IFNφ1pro) activity and mRNA level induced by retinoic acid-inducible gene (RIG)-I-like receptors (RLRs) were significantly inhibited by TMEM33. Knockdown of TMEM33 increased host ifn transcription. Subsequently, we found that TMEM33 was colocalized in the ER and interacted with the RLR cascades, whereas MAVS was degraded by TMEM33 during the K48-linked ubiquitination. On the other hand, TMEM33 reduced the phosphorylation of mediator of IFN regulatory factor 3 (IRF3) activation (MITA)/IRF3 by acting as a decoy substrate of TBK1, which was also phosphorylated. A functional domain assay revealed that the N-terminal transmembrane domain 1 (TM1) and TM2 regions of TMEM33 were necessary for IFN suppression. Finally, TMEM33 significantly attenuated the host cellular antiviral capacity by blocking the IFN response. Taken together, our findings provide insight into the different mechanisms employed by TMEM33 in cellular IFN-mediated antiviral process.

2.
Dev Comp Immunol ; 115: 103876, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32987012

RESUMO

In mammals, cyclic GMP-AMP synthase (cGAS) is a crucial cytosolic DNA sensor responsible for activating the interferon (IFN) response. A cGAS-like (cGASL) gene was previously identified from grass carp Ctenopharyngodon idellus, which is evolutionarily closest to cGAS but not a true ortholog of cGAS. Here, we found that grass carp cGASL targets mitochondrial antiviral signaling protein (MAVS) for autophagic degradation to negatively regulate fish IFN response. Firstly, the transcriptional level of cellular cgasl was upregulated by poly I:C stimulation, and overexpression of cGASL significantly decreased poly I:C- and MAVS-induced promoter activities and transcriptional levels of IFN and IFN-stimulated genes (ISGs). In addition, cGASL associated with MAVS and prompted autophagic degradation of MAVS in a dose-dependent manner. Finally, overexpression of cGASL attenuated MAVS-mediated cellular antiviral response. These results collectively indicate that cGASL negatively regulates fish IFN response by triggering autophagic degradation of MAVS.

3.
Dev Comp Immunol ; 114: 103805, 2021 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-32755617

RESUMO

TANK-binding kinase 1 (TBK1) plays a vital role in activating interferon (IFN) production and positively regulating antiviral response in mammals. Research on more species of fish is necessary to clarify whether the function of fish TBK1 is conserved compared to that in mammals. Here, a cyprinid fish (Ancherythroculter nigrocauda) TBK1 (AnTBK1) was functionally identified and characterized. The full-length open reading frame (ORF) of AnTBK1 consists of 2184 nucleotides encoding 727 amino acids and contains a conserved Serine/Threonine protein kinase catalytic domain (S_TKc) in the N-terminal, similar to TBK1 in other species. The transcripts of AnTBK1 were found in all the tissues evaluated and the cellular distribution indicated that AnTBK1 was localized in the cytoplasm. In terms of functional identification, AnTBK1 induced a variety of IFN promoter activities as well as the expression of downstream IFN-stimulated genes (ISGs). In addition, AnTBK1 interacted with and significantly phosphorylated IFN regulatory factor 3 (IRF3), exhibiting the canonical kinase activity of TBK1. Finally, AnTBK1 presented strong antiviral activity against spring viremia of carp virus (SVCV) infection. Taken together, our research on the features and functions of AnTBK1 demonstrated that AnTBK1 plays a central role in IFN induction against SVCV infection.

4.
Front Immunol ; 11: 545302, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33193312

RESUMO

Viruses typically target host RIG-I-like receptors (RLRs), a group of key factors involved in interferon (IFN) production, to enhance viral infection. To date, though immune evasion methods to contradict IFN production have been characterized for a series of terrestrial viruses, the strategies employed by fish viruses remain unclear. Here, we report that all grass carp reovirus (GCRV) proteins encoded by segments S1 to S11 suppress mitochondrial antiviral signaling protein (MAVS)-mediated IFN expression. First, the GCRV viral proteins blunted the MAVS-induced expression of IFN, and impair MAVS antiviral capacity significantly. Interestingly, subsequent co-immunoprecipitation experiments demonstrated that all GCRV viral proteins interacted with several RLR cascades, especially with TANK-binding kinase 1 (TBK1) which was the downstream factor of MAVS. To further illustrate the mechanisms of these interactions between GCRV viral proteins and host RLRs, two of the viral proteins, NS79 (S4) and VP3 (S3), were selected as representative proteins for two distinguished mechanisms. The obtained data demonstrated that NS79 was phosphorylated by gcTBK1, leading to the reduction of host substrate gcIRF3/7 phosphorylation. On the other hand, VP3 degraded gcMAVS and the degradation was significantly reversed by 3-MA. The biological effects of both NS79 and VP3 were consistently found to be related to the suppression of IFN expression and the promotion of viral evasion. Our findings shed light on the special evasion mechanism utilized by fish virus through IFN regulation, which might differ between fish and mammals.

5.
J Immunol ; 205(7): 1819-1829, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32859727

RESUMO

IFN is essential for hosts to defend against viral invasion, whereas it must be tightly regulated to prevent hyperimmune responses. Fish mitochondrial antiviral signaling protein (MAVS) is a vital factor for IFN production, but until now, there have been few studies on the regulation mechanisms of fish MAVS enabling IFN to be properly controlled. In this study, we show that zebrafish RNA-binding motif protein 47 (RBM47) promotes MAVS degradation in a lysosome-dependent manner to suppress IFN production. First, the transcription of IFN activated by polyinosinic/polycytidylic acid (poly I:C), spring viremia of carp virus, or retinoic acid-inducible gene I (RIG-I)-like receptor pathway components were significantly suppressed by RBM47. Second, RBM47 interacted with MAVS and promoted lysosome-dependent degradation of MAVS, changing the cellular location of MAVS from the cytoplasm to the lysosome region. Finally, RBM47 inhibited downstream MITA and IRF3/7 activation, impairing the host antiviral response. Collectively, these data suggest that zebrafish RBM47 negatively regulates IFN production by promoting lysosome-dependent degradation of MAVS, providing insights into the role of RBM47 in the innate antiviral immune response in fish.

6.
Fish Shellfish Immunol ; 102: 449-459, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32408017

RESUMO

Calcium (Ca) is a messenger that regulates a multitude of physiological processes, but its functions in antiviral progress remain undefined. In this study, we found that Ca2+ enhances fish survival to defend against spring viraemia of carp virus (SVCV) infection by reversing the instability of p53 mediated by the viral protein. First, Ca2+ significantly protected cells and fish against SVCV infection by inducing early apoptosis. Additionally, p53 expression, which was inhibited by SVCV N protein, was upregulated by Ca2+ treatment. Then, the mechanism underlying the reduction of K63-linked p53 ubiquitination by SVCV N protein via the K358 site was completely prevented by Ca2+. These findings reveal the role of Ca2+ in lower vertebrates in the antiviral response, which is connected to and corresponds with viral immune evasion, providing a solution to fish diseases caused by pathogens.

7.
J Virol ; 94(15)2020 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-32434890

RESUMO

Spring viremia of carp virus (SVCV) is a highly pathogenic Vesiculovirus in the common carp. The phosphoprotein (P protein) of SVCV is a multifunctional protein that acts as a polymerase cofactor and an antagonist of cellular interferon (IFN) response. Here, we report the 1.5-Å-resolution crystal structure of the P protein central domain (PCD) of SVCV (SVCVPCD). The PCD monomer consists of two ß sheets, an α helix, and another two ß sheets. Two PCD monomers pack together through their hydrophobic surfaces to form a dimer. The mutations of residues on the hydrophobic surfaces of PCD disrupt the dimer formation to different degrees and affect the expression of host IFN consistently. Therefore, the oligomeric state formation of the P protein of SVCV is an important mechanism to negatively regulate host IFN response.IMPORTANCE SVCV can cause spring viremia of carp with up to 90% lethality, and it is the homologous virus of the notorious vesicular stomatitis virus (VSV). There are currently no drugs that effectively cure this disease. P proteins of negative-strand RNA viruses (NSVs) play an essential role in many steps during the replication cycle and an additional role in immunosuppression as a cofactor. All P proteins of NSVs are oligomeric, but the studies on the role of this oligomerization mainly focus on the process of virus transcription or replication, and there are few studies on the role of PCD in immunosuppression. Here, we present the crystal structure of SVCVPCD A new mechanism of immune evasion is clarified by exploring the relationship between SVCVPCD and host IFN response from a structural biology point of view. These findings may provide more accurate target sites for drug design against SVCV and provide new insights into the function of NSVPCD.


Assuntos
Fosfoproteínas/química , Rhabdoviridae/química , Proteínas Virais/química , Animais , Cristalografia por Raios X , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta
8.
Fish Shellfish Immunol ; 99: 99-106, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32032764

RESUMO

Grass carp reovirus (GCRV) is an efficient pathogen causing high mortality in grass carp, meanwhile, fish interferon (IFN) is a powerful cytokine enabling host cells to establish an antiviral state; therefore, the strategies used by GCRV to escape the cellular IFN response need to be investigated. Here, we report that GCRV VP56 inhibits host IFN production by degrading the transcription factor IFN regulatory factor 7 (IRF7). First, overexpression of VP56 inhibited the IFN production induced by the polyinosinic-polycytidylic acid (poly I:C) and mitochondrial antiviral signaling protein (MAVS), while the capacity of IRF7 on IFN induction was unaffected. Second, VP56 interacted with RLRs but did not affect the stabilization of the proteins in the normal state, while the phosphorylated IRF7 activated by TBK1 was degraded by VP56 through K48-linked ubiquitination. Finally, overexpression of VP56 remarkably reduced the host cellular ifn transcription and facilitated viral proliferation. Taken together, our results demonstrate that GCRV VP56 suppresses the host IFN response by targeting phosphorylated IRF7 for ubiquitination and degradation.


Assuntos
Carpas/virologia , Fator Regulador 7 de Interferon/metabolismo , Interferons/antagonistas & inibidores , Infecções por Reoviridae/veterinária , Proteínas Virais/genética , Animais , Carpas/imunologia , Feminino , Células HEK293 , Humanos , Imunidade Inata , Fator Regulador 7 de Interferon/imunologia , Interferons/imunologia , Ovário/citologia , Fosforilação , Poli I-C/farmacologia , Reoviridae , Infecções por Reoviridae/imunologia , Ubiquitinação , Proteínas Virais/imunologia
9.
Fish Shellfish Immunol ; 97: 523-530, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31881328

RESUMO

Interferon (IFN) is a vital antiviral factor in host in the early stages after the viral invasion. Meanwhile, viruses have to survive by taking advantage of the cellular machinery and complete their replication. As a result, viruses evolved several immune escape mechanisms to inhibit host IFN expression. However, the mechanisms used to escape the host's IFN system are still unclear for infectious hematopoietic necrosis virus (IHNV). In this study, we report that the N protein of IHNV inhibits IFN1 production in rainbow trout by degrading the MITA. Firstly, the upregulation of IFN1 promoter activity stimulated by poly I:C was suppressed by IHNV infection. Consistent with this result, the overexpression of the N protein of IHNV blocked the IFN1 transcription that was activated by poly I:C and MITA. Secondly, MITA was remarkably decreased by the overexpression of N protein at the protein level. Further analysis demonstrated that the N protein targeted MITA and promoted the ubiquitination of MITA. Taken together, these data suggested that the production of rainbow trout IFN1 could be suppressed by the N protein of IHNV via degrading MITA.


Assuntos
Proteínas de Peixes/genética , Vírus da Necrose Hematopoética Infecciosa/imunologia , Interferons/imunologia , Proteínas de Membrana/genética , Proteínas do Nucleocapsídeo/imunologia , Oncorhynchus mykiss/imunologia , Animais , Antivirais/farmacologia , Células HEK293 , Interações entre Hospedeiro e Microrganismos/imunologia , Humanos , Vírus da Necrose Hematopoética Infecciosa/genética , Proteínas do Nucleocapsídeo/genética , Oncorhynchus mykiss/virologia , Poli I-C/farmacologia , Infecções por Rhabdoviridae , Ubiquitinação
10.
Fish Shellfish Immunol ; 94: 871-879, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31597087

RESUMO

Mammalian cyclic GMP-AMP synthase (cGAS) senses double-stranded (ds) DNA in the cytosol to activate the innate antiviral response. In the present study, a cGAS-like gene, namely cGASL, was cloned from grass carp Ctenopharyngodon idellus, and its role as a negative regulator of the IFN response was revealed. Phylogenetic analysis indicated that cGASL was evolutionarily closest to cGAS, but was not a true ortholog of cGAS. Overexpression of cGASL inhibited poly I:C-stimulated grass carp (gc)IFN1pro and ISRE activities. In addition, MITA-, but not TBK1-mediated activation of gcIFN1pro was impaired by cGASL. Co-immunoprecipitation and Western blot experiments indicated that cGASL interacted with MITA and TBK1, resulting in a reduction in the phosphorylation of MITA. Lastly, overexpression of cGASL reduced the transcriptional levels of several IFN-stimulated genes activated by MITA. Collectively, these data suggest that cGASL is a negative regulator of IFN response by targeting MITA in fish.


Assuntos
Carpas/genética , Carpas/imunologia , Doenças dos Peixes/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Nucleotidiltransferases/genética , Nucleotidiltransferases/imunologia , Sequência de Aminoácidos , Animais , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica/veterinária , Interferons/metabolismo , Nucleotidiltransferases/química , Filogenia , Poli I-C/farmacologia , Alinhamento de Sequência/veterinária
11.
J Virol ; 93(21)2019 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-31413136

RESUMO

Interferon (IFN) production activated by phosphorylated interferon regulatory factor 7 (IRF7) is a pivotal process during host antiviral infection. For viruses, suppressing the host IFN response is beneficial for viral proliferation; in such cases, evoking host-derived IFN negative regulators would be very useful for viruses. Here, we report that the zebrafish rapunzel 5 (RPZ5) protein which activated by virus degraded phosphorylated IRF7 is activated by TANK-binding kinase 1 (TBK1), leading to a reduction in IFN production. Upon viral infection, zebrafish rpz5 was significantly upregulated, as was ifn, in response to the stimulation. Overexpression of RPZ5 blunted the IFN expression induced by both viral and retinoic acid-inducible gene I (RIG-I) like-receptor (RLR) factors. Subsequently, RPZ5 interacted with RLRs but did not affect the stabilization of the proteins in the normal state. Interestingly, RPZ5 degraded the phosphorylated IRF7 under TBK1 activation through K48-linked ubiquitination. Finally, the overexpression of RPZ5 remarkably reduced the host cell antiviral capacity. These findings suggest that zebrafish RPZ5 is a negative regulator of phosphorylated IRF7 and attenuates IFN expression during viral infection, providing insight into the IFN balance mechanism in fish.IMPORTANCE The phosphorylation of IRF7 is helpful for host IFN production to defend against viral infection; thus, it is a potential target for viruses to mitigate the antiviral response. We report that the fish RPZ5 is an IFN negative regulator induced by fish viruses and degrades the phosphorylated IRF7 activated by TBK1, leading to IFN suppression and promotion of viral proliferation. These findings reveal a novel mechanism for interactions between the host cell and viruses in the lower vertebrate.


Assuntos
Doenças dos Peixes/virologia , Imunidade Inata/imunologia , Interferons/metabolismo , Infecções por Rhabdoviridae/veterinária , Rhabdoviridae/imunologia , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/virologia , Animais , Antivirais/imunologia , Antivirais/metabolismo , Doenças dos Peixes/imunologia , Doenças dos Peixes/metabolismo , Fatores Reguladores de Interferon/genética , Fatores Reguladores de Interferon/metabolismo , Interferons/imunologia , Fosforilação , Infecções por Rhabdoviridae/imunologia , Infecções por Rhabdoviridae/metabolismo , Infecções por Rhabdoviridae/virologia , Ubiquitinação , Replicação Viral , Peixe-Zebra/fisiologia , Proteínas de Peixe-Zebra/genética
12.
Front Immunol ; 10: 1106, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31191518

RESUMO

Expression of major histocompatibility complex class II (MHC II) molecules, which determines both the immune repertoire during development and subsequent triggering of immune responses, is always under the control of a unique (MHC class II) transactivator, CIITA. The IFN-γ-inducible MHC II expression has been extensively and thoroughly studied in humans, but not in bony fish. In this study, the characterization of CIITA was identified and its functional domains were analyzed in grass carp. The absence of GAS and E-box in the promoter region of grass carp CIITA, might imply that the cooperative interaction between STAT1 and USF1 to active the CIITA expression, found in mammals, is not present in bony fish. After the transfection of IFN-γ or IFN-γ rel, only IFN-γ could induce MHC II expression mediated by CIITA. Moreover, interferon regulatory factor (IRF) 2, which cooperates with IRF1 to active the CIITA promoter IV expression in mammals, played an antagonistic role to IRF1 in the activation of grass carp CIITA. These data suggested that grass carp, compared with mammals, has both conservative and unique mechanisms in the regulation of MHC II expression.


Assuntos
Carpas/genética , Carpas/metabolismo , Regulação da Expressão Gênica , Antígenos de Histocompatibilidade Classe II/genética , Fator Regulador 1 de Interferon/metabolismo , Fator Regulador 2 de Interferon/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Carpas/imunologia , Linhagem Celular , DNA Complementar/química , DNA Complementar/genética , Antígenos de Histocompatibilidade Classe II/imunologia , Humanos , Imunomodulação , Interferon gama/metabolismo , Proteínas Nucleares/metabolismo , Fases de Leitura Aberta , Regiões Promotoras Genéticas , Ligação Proteica , Transativadores/metabolismo
13.
Fish Shellfish Immunol ; 92: 224-229, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31200068

RESUMO

Fibroblast growth factor receptor (FGFR) 3 is one of the four distinct membrane-spanning tyrosine kinases required for proper skeletal development. In fish, the role of FGFR3 is still unclear. In this article, we reveal that zebrafish FGFR3 is a negative regulator of interferon (IFN) production in the innate immune response by suppressing the activity of TANK-binding kinase 1 (TBK1) in the process of virus infection. qPCR experiments demonstrate that the transcriptional level of cellular FGFR3 was upregulated by infection with spring viremia of carp virus (SVCV), indicating that FGFR3 might be involved in the process of host cell response to viral infection. Then, overexpression of FGFR3 significantly impeded the IFN promoter activity induced by a stimulator. In addition, the capabilities of a retinoic acid-inducible gene I (RIG-I)-like receptor (RLR) system to activate IFN promoter were decreased during the overexpression of FGFR3. Subsequently, FGFR3 decreased the phosphorylation of interferon regulatory factor 3 (IRF3) and mediator of IRF3 activation (MITA) by TBK1. These findings suggest that zebrafish FGFR3 is a negative regulator of IFN by attenuating the kinase activity of TBK1, leading to the suppression of IFN expression.


Assuntos
Doenças dos Peixes/imunologia , Imunidade Inata/genética , Interferons/genética , Proteínas Serina-Treonina Quinases/genética , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/genética , Proteínas de Peixe-Zebra/genética , Peixe-Zebra/imunologia , Animais , Interferons/metabolismo , Proteínas Serina-Treonina Quinases/imunologia , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/imunologia , Rhabdoviridae/fisiologia , Infecções por Rhabdoviridae/imunologia , Infecções por Rhabdoviridae/veterinária , Transdução de Sinais/imunologia , Peixe-Zebra/genética , Proteínas de Peixe-Zebra/imunologia , Proteínas de Peixe-Zebra/fisiologia
14.
Front Immunol ; 10: 985, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31130963

RESUMO

In East Asia, the dark sleeper, Odontobutis obscura (O. obscura) is a crucial commercial species of freshwater fish; however, its molecular biology research is still undeveloped, including its innate immune system, which is pivotal to antiviral responses. In this study, we cloned and identified the characterization and kinase function of dark sleeper TANK-binding kinase 1 (TBK1), supplementing the evidence of the conservation of this classical factor in fish. First, the ORF of Odontobutis obscurus (O. obscura) TBK1 (OdTBK1) was cloned from liver tissue by RACE-PCR. Subsequent nucleic acid and amino acid sequence analysis suggested that OdTBK1 is homologous with other fish TBK1, and the N-terminal Serine/Threonine protein kinases catalytic domain (S_TKc) and C-terminal coiled coil domain (CCD) are conserved. Subsequently, the cellular distribution demonstrated that OdTBK1 was located in the cytoplasm region. With regard to the identification of functions, OdTBK1 activated several interferon (IFN) promoters' activity and induced downstream IFN-stimulated genes (ISGs) expression. In a canonical manner, wild-type OdTBK1 significantly phosphorylated interferon regulatory factor 3 (IRF3) but failed when the N-terminal region was truncated. Furthermore, overexpression of OdTBK1 decreased viral proliferation remarkably. Collectively, these data systematically analyzed the characterization and function of OdTBK1, initiating the study of the innate antiviral response of dark sleeper.


Assuntos
Doenças dos Peixes/imunologia , Proteínas de Peixes/imunologia , Peixes/imunologia , Fator Regulador 3 de Interferon/imunologia , Proteínas Serina-Treonina Quinases/imunologia , Infecções por Vírus de RNA/imunologia , Animais , Linhagem Celular , Proteínas de Peixes/genética , Peixes/virologia , Humanos , Imunidade Inata , Interferons/imunologia , Proteínas Serina-Treonina Quinases/genética
15.
Fish Shellfish Immunol ; 89: 301-308, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30965085

RESUMO

Indoleamine 2,3-dioxygenase (IDO) is a kind of dioxygenase that can catalyze the degradation of levo-tryptophan (L-Trp) and plays key roles in immune tolerance. In this study, the IDO gene was cloned and functionally characterized from grass carp (gcIDO). The results showed that gcIDO overexpressed in GCO cells could catalyze the degradation of L-Trp through the L-Trp - kynurenine pathway, and this activity could be promoted by δ-aminolevulinic acid (ALA) while inhibited by levo-1-methyl tryptophan (L-1MT). Moreover, gcIDO was constitutively expressed in various tissues, and its expression could be significantly up-regulated by LPS and Poly (I:C) in peripheral blood leukocytes (PBLs). Furthermore, recombinant TGF-ß1 of grass carp could up-regulate the expression of IDO, TGF-ß1, CD25, and Foxp3 in PBLs, indicating that the TGF-ß1/IDO pathway is present in fish. In the soybean meal induced enteritis (SBMIE) model, the expression of gcIDO in the intestine was up-regulated significantly, demonstrating that gcIDO may play an immunoregulatory role in SBMIE. Taken together, these data suggest that the IDO plays multiple roles in the immunity of fish.


Assuntos
Carpas/genética , Enterite/veterinária , Doenças dos Peixes/genética , Regulação da Expressão Gênica/imunologia , Indolamina-Pirrol 2,3,-Dioxigenase/genética , Indolamina-Pirrol 2,3,-Dioxigenase/imunologia , Animais , Carpas/imunologia , Enterite/induzido quimicamente , Enterite/genética , Enterite/imunologia , Doenças dos Peixes/induzido quimicamente , Doenças dos Peixes/imunologia , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Lipopolissacarídeos/farmacologia , Poli I-C/farmacologia , Soja/química , Fator de Crescimento Transformador beta1/genética
16.
PLoS Pathog ; 15(3): e1007695, 2019 03.
Artigo em Inglês | MEDLINE | ID: mdl-30925159

RESUMO

p53, which regulates cell-cycle arrest and apoptosis, is a crucial target for viruses to release cells from cell-cycle checkpoints or to protect cells from apoptosis for their own benefit. Viral evasion mechanisms of aquatic viruses remain mysterious. Here, we report the spring viremia of carp virus (SVCV) degrading and stabilizing p53 in the ubiquitin-proteasome pathway by the N and P proteins, respectively. Early in an SVCV infection, significant induction was observed in the S phase and p53 was decreased in the protein level. Further experiments demonstrated that p53 interacted with SVCV N protein and was degraded by suppressing the K63-linked ubiquitination. However, the increase of p53 was observed late in the infection and experiments suggested that p53 was bound to SVCV P protein and stabilized by enhancing the K63-linked ubiquitination. Finally, lysine residue 358 was the key site for p53 K63-linked ubiquitination by the N and P proteins. Thus, our findings suggest that fish p53 is modulated by SVCV N and P protein in two distinct mechanisms, which uncovers the strategy for the subversion of p53-mediated host innate immune responses by aquatic viruses.


Assuntos
Rhabdoviridae/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismo , Animais , Pontos de Checagem do Ciclo Celular/fisiologia , Vírus de DNA , Doenças dos Peixes , Regulação Viral da Expressão Gênica/genética , Células HEK293 , Humanos , Imunidade Inata , Rhabdoviridae/patogenicidade , Ubiquitinação , Viremia , Peixe-Zebra , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
17.
J Immunol ; 202(2): 559-566, 2019 01 15.
Artigo em Inglês | MEDLINE | ID: mdl-30530482

RESUMO

IFN production is crucial for hosts to defend against viral infection, yet it must be tightly controlled to maintain immune homeostasis. TANK-binding kinase 1 (TBK1) is a pivotal kinase in the IFN induction signaling pathway, but it is negatively regulated by multiple molecules to avoid the excessive expression of IFN in mammals. However, the identified TBK1 suppressors and the mechanisms are rare in fish. In this study, we show that zebrafish major vault protein (MVP) recruits and degrades TBK1 in a lysosome-dependent manner to inhibit IFN production. Through viral infection, polyinosinic:polycytidylic acid and RIG-I-like receptor factor stimulation upregulated IFN expression, but overexpression of MVP significantly subverted these inductions. On the protein level, MVP interacted with TBK1, and interestingly, MVP recruited TBK1 from a uniformly distributed state in the cytoplasm to an aggregated state. Finally, MVP mediated the lysosome-dependent degradation of TBK1 and decreased the IFN response and IFN-stimulated genes expression. Our findings reveal that zebrafish MVP is a negative regulator of IFN production by restricting the activation of TBK1, supplying evidence of the balanced mechanisms of IFN expression in lower vertebrates.


Assuntos
Proteínas de Peixes/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Partículas de Ribonucleoproteínas em Forma de Abóbada/metabolismo , Viroses/imunologia , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/imunologia , Animais , Animais Geneticamente Modificados , Proteínas de Peixes/genética , Regulação da Expressão Gênica , Células HEK293 , Humanos , Interferons/genética , Lisossomos/metabolismo , Poli I-C/imunologia , Agregação Patológica de Proteínas , Ligação Proteica , Proteólise , Transdução de Sinais , Partículas de Ribonucleoproteínas em Forma de Abóbada/genética
18.
Dev Comp Immunol ; 93: 78-88, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30590066

RESUMO

Interleukin-35 (IL-35) is a member of the IL-12 cytokine family and a heterodimeric protein formed by Epstein-Barr virus-induced gene 3 (EBI3) and IL-12p35. Emerging evidence showed that IL-35 is a key player in the regulation of cellular communication, differentiation, and inflammation. To date, no studies on fish IL-35 have been documented. In this work, we first identify two splicing isoforms of EBI3, EBI3a and EBI3b, from grass carp (Ctenopharyngodon idella). EBI3a is composed of 299 amino acid residues and possesses an immunoglobulin-like (Ig-like) domain and a fibronectin type 3 (FN3) domain that is a conservative domain in vertebrate EBI3. However, the EBI3b is composed of 177 amino acid residues and only contains an Ig-like domain. The result of Co-immunoprecipitation suggests that only EBI3a can associate with IL-12p35 to form IL-35 in grass carp. Like the function of IL-35 in human and mouse, recombinant grass carp IL-35 protein could induce the expression of genes EBI3a, IL-12p35, and CD25-like and downregulate the expression of genes CD4-1, CD4-2, IL-17A/F1, and RORγ2. Taken together, these results indicate for the first time that a teleost IL-35 may also have the ability to induce regulatory T (Treg) cells, inhibit effector T (Teff) cell proliferation and restrict the differentiation and function of T helper 17 (Th17) cells in teleost.


Assuntos
Carpas/imunologia , Interleucinas/genética , Interleucinas/imunologia , Isoformas de Proteínas/genética , Linfócitos T Reguladores/imunologia , Células Th17/imunologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Proliferação de Células , Domínio de Fibronectina Tipo III/genética , Regulação da Expressão Gênica/imunologia , Imunoglobulinas/genética , Subunidade p35 da Interleucina-12/metabolismo , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia
19.
J Immunol ; 202(1): 119-130, 2019 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-30504422

RESUMO

Viral infection activates the transcription factor IFN regulatory factor 7 (IRF7), which plays a critical role in the induction of IFNs and innate antiviral immune response. How virus-induced IFN signaling is controlled in fish is not fully understood. In this study, we demonstrate that N-myc downstream-regulated gene 1a (NDRG1a) in zebrafish plays a role as a negative regulator for virus-triggered IFN induction. First, the activation of the IFN promoter stimulated by the polyinosinic-polycytidylic acid or spring viremia of carp virus was decreased by the overexpression of NDRG1a. Second, NDRG1a interacted with IRF7 and blocked the IFN transcription activated by IRF7. Furthermore, NDRG1a was phosphorylated by TANK-binding kinase 1 (TBK1) and promoted the K48-linked ubiquitination and degradation of IRF7. Finally, the overexpression of NDRG1a blunted the transcription of several IFN-stimulated genes, resulting in the host cells becoming susceptible to spring viremia of carp virus infection. Our findings suggest that fish NDRG1a negatively regulates the cellular antiviral response by targeting IRF7 for ubiquitination and degradation, providing insights into the novel role of NDRG1a on the innate antiviral immune response in fish.


Assuntos
Doenças dos Peixes/imunologia , Doenças dos Peixes/virologia , Fatores Reguladores de Interferon/metabolismo , Interferons/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Infecções por Rhabdoviridae/imunologia , Infecções por Rhabdoviridae/veterinária , Rhabdoviridae/fisiologia , Proteínas Supressoras de Tumor/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/imunologia , Animais , Células Cultivadas , Suscetibilidade a Doenças , Imunidade Inata , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteólise , Proteínas Proto-Oncogênicas/metabolismo , Transdução de Sinais , Proteínas Supressoras de Tumor/genética , Ubiquitinação , Proteínas de Peixe-Zebra/genética
20.
Fish Shellfish Immunol ; 81: 304-308, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-30030114

RESUMO

In bony fish, CD40 and CD154 are two very important costimulatory molecules involved in T and B cell cooperation in thymus-dependent antibody production. In the current study, we identified the cDNAs of CD40 and CD154 and analyzed their genomic structures in grass carp. Quantitative real-time PCR indicated that the CD40 and CD154 were mainly expressed in immune organs. After challenge with grass carp reovirus (GCRV), these two genes were up-regulated at 72 h in head kidney and spleen. Moreover, seven and five single nucleotide polymorphisms (SNPs) were identified in the CD40 and CD154 respectively. Statistical analysis indicated that three SNPs in the coding region of the CD40 were significantly associated with the resistance of grass carp against GCRV. These results indicated that CD40 and CD154 play important roles in the responses to GCRV in grass carp. The SNP markers in the CD40 associated with the resistance to GCRV may facilitate the disease-resistant breeding of grass carp.


Assuntos
Antígenos CD40/genética , Ligante de CD40/genética , Carpas/genética , Doenças dos Peixes/genética , Infecções por Reoviridae/genética , Animais , DNA Complementar/genética , Resistência à Doença/genética , Polimorfismo de Nucleotídeo Único , Infecções por Reoviridae/veterinária
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