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1.
J Microbiol Biotechnol ; 29(8): 1324-1334, 2019 Aug 28.
Artigo em Inglês | MEDLINE | ID: mdl-31370117

RESUMO

Fish mycobacteriosis is a common bacterial disease in many species of freshwater and marine fish and has caused severe loss of fish production. Mycobacterium marinum has been the most prevalent pathogen observed in several outbreaks of mycobacteriosis of farmed sturgeons in China. However, the immune responses and pathology of sturgeons in mycobacterial infection are rarely studied. Therefore, we used the Illumina RNA-seq method to analyze the transcriptome profile of Acipenser schrenckii challenged with Mycobacterium marinum. To begin, 168,220 non-redundant contigs were acquired from the infection and control groups, and among these, 33,225 contigs have acquired annotations. A total of 4,043 differently expressed (DE) contigs between the two groups were identified, and among these, 2479 were upregulated and 1564 were down-regulated in the infected fish. A total of 1,340 DE contigs with acquired annotations in KEGG were enriched for 124 pathways including the TNF signaling pathway, and the Toll-like receptor signaling pathway. The roles of DE genes involved in significant pathways and other processes were discussed. The 2,209 DE contigs that have yet to acquire proper annotation may represent candidate genes associated with infection in sturgeons and are expected to serve as immunogenetic resources for further study. To our best knowledge, this is the first transcriptome study on sturgeons under bacterial infection.


Assuntos
Doenças dos Peixes/genética , Doenças dos Peixes/imunologia , Proteínas de Peixes/genética , Peixes/genética , Infecções por Micobactéria não Tuberculosa/genética , Infecções por Micobactéria não Tuberculosa/imunologia , Infecções por Micobactéria não Tuberculosa/veterinária , Transcriptoma , Animais , China , Regulação para Baixo , Doenças dos Peixes/microbiologia , Proteínas de Peixes/metabolismo , Perfilação da Expressão Gênica , Imunidade , Anotação de Sequência Molecular , Mycobacterium marinum/patogenicidade , Regulação para Cima
2.
Phys Chem Chem Phys ; 21(35): 19298-19310, 2019 Sep 21.
Artigo em Inglês | MEDLINE | ID: mdl-31451813

RESUMO

The ice/water interface recognition mechanism of antifreeze proteins (AFPs) is highly contentious. Conventionally, protein adsorption on a solid surface is primarily driven by the polar interactions between the hydrophilic residues of the protein and interfacial water of the solid surface. Ice surface recognition by a type III AFP is surprising in this context where the ice binding surface (IBS) is hydrophobic. The present study provides molecular insight into the unusual interface recognition phenomenon of a type III AFP (QAE isoform) from Macrozoarces americanus. Potential of mean force calculations show that the type III AFP adsorbs on the ice surface mediated through a layer of ordered water. Molecular dynamics simulations at lower than ambient temperature reveal that the flat hydrophobic IBS induces ordering of water. The excellent geometrical synergy between the hydration water structure around the IBS and water arrangements on the pyramidal surface favours adsorption on the pyramidal plane. Mutations that interrupt the hydration shell water ordering essentially lead to less efficient adsorption, which greatly reduces the anti-freezing activity of the AFP. Binding free energy calculations of the wild-type and several mutant AFPs reveal that the binding affinity is linearly correlated with the experimentally observed thermal hysteresis activity. Therefore, binding to a specific ice plane with considerable affinity is the dictating factor of the anti-freeze activity for a type III AFP. Mechanistic insights into the ice binding process of the wild-type and different mutant AFPs obtained from this study pave the way for rational design of type III variants with much improved activity, which possesses ample industrial applicability, particularly in cryo-preservation.


Assuntos
Proteínas Anticongelantes/química , Proteínas de Peixes/química , Gelo , Perciformes , Água/química , Animais , Proteínas Anticongelantes/genética , Temperatura Baixa , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Simulação de Dinâmica Molecular , Mutação , Ligação Proteica
3.
DNA Cell Biol ; 38(10): 1100-1111, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31418589

RESUMO

Members of the Sox gene family play crucial roles during reproduction and development, but their genome-wide identification has not yet been performed in large yellow croaker, Larimichthys crocea. In this study, a total of 26 members of the Sox gene family were identified from the genome of large yellow croaker and classified into seven subgroups based on the conserved HMG-box domain they contain. Among the identified Sox gene family members, eight belonged to the SoxB subgroup (five in B1 and three in B2), four belonged to the SoxC subgroup, four belonged to the SoxD subgroup, six belonged to the SoxE subgroup, three belonged to the SoxF subgroup, and one belonged to the SoxK subgroup. During evolution, members of the SoxE subgroup (Sox8, Sox9, Sox10), Sox1, Sox4, Sox6, and Sox11 evolved into two copies, which may be a result of teleost-specific whole-genome duplication. Sox genes were distributed unevenly across 15 chromosomes. The number of introns in large yellow croaker Sox genes varied from 0 to 14. Results of the expression profile during embryogenesis revealed that most of the members of the Sox gene family had lower expression, except several Sox genes, and expression patterns also differed among each Sox gene group and duplicated gene. This study systematically characterized and analyzed the Sox gene family in large yellow croaker and provided new insights into its function during embryogenesis.


Assuntos
Proteínas de Peixes/genética , Regulação da Expressão Gênica no Desenvolvimento , Genoma , Perciformes/genética , Filogenia , Fatores de Transcrição SOXB1/genética , Sequência de Aminoácidos , Animais , Evolução Biológica , Mapeamento Cromossômico , Biologia Computacional , Embrião não Mamífero , Desenvolvimento Embrionário , Éxons , Proteínas de Peixes/classificação , Duplicação Gênica , Íntrons , Família Multigênica , Perciformes/classificação , Isoformas de Proteínas/classificação , Isoformas de Proteínas/genética , Fatores de Transcrição SOXB1/classificação , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos
4.
Artigo em Inglês | MEDLINE | ID: mdl-31255700

RESUMO

Myogenic regulatory factor 4 (MRF4) is a basic helix-loop-helix (bHLH) transcription factor that plays crucial roles in myoblast differentiation and maturation. Here, we report the isolation of the olive flounder (Paralichthys olivaceus) mrf4 gene and the spatiotemporal analysis of its expression patterns. Sequence analysis indicated that flounder mrf4 shared a similar structure with other vertebrate MRF4, including the conserved bHLH domain. Flounder mrf4 contains 3 exons and 2 introns. Sequence alignment and phylogenetic analysis showed that it was highly homologous with Salmo salar, Danio rerio, Takifugu rubripes, and Tetraodon nigroviridis mrf4. Flounder mrf4 was first expressed in the medial region of somites that give rise to slow muscles, and later spread to the lateral region of somites that give rise to fast muscles. Mrf4 transcript levels decreased significantly in mature somites in the trunk region, and expression could only be detected in the caudal somites, consistent with the timing of somite maturation. Transient expression analysis showed that the 506 bp flounder mrf4 promoter was sufficient to direct muscle-specific GFP expression in zebrafish embryos.


Assuntos
Proteínas de Peixes/genética , Linguado/genética , Músculos/metabolismo , Fatores de Regulação Miogênica/genética , Regiões Promotoras Genéticas/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Desenvolvimento Embrionário , Proteínas de Peixes/química , Linguado/embriologia , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Fatores de Regulação Miogênica/química , Especificidade de Órgãos
5.
Gene ; 718: 143989, 2019 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-31326551

RESUMO

Our comparative studies seek to understand the structure and function of ion channels in cartilaginous fish that can detect very low voltage gradients in seawater. The principal channels of the electroreceptor include a calcium activated K channel whose α subunit is Kcnma1, and a voltage-dependent calcium channel, Cacna1d. It has also been suggested based on physiological and pharmacological evidence that a voltage-gated K channel is present in the basal membranes of the receptor cells which modulates synaptic transmitter release. Large conductance calcium-activated K channels (BK) are comprised of four α subunits, encoded by Kcnma1 and modulatory ß subunits of the Kcnmb class. We recently cloned and published the skate Kcnma1 gene and most of Kcnmb4 using purified mRNA of homogenized electroreceptors. Bellono et al. have recently performed RNA sequencing (RNA-seq) on purified mRNA from skate electroreceptors and found several ion channels including Kcnma1. We searched the Bellono et al. RNA-seq repository for additional channels and subunits. Our most significant findings are the presence of two Shaker type voltage dependent K channel sequences which are grouped together as isoforms in the data repository. The larger of these is a skate ortholog of the voltage dependent fast potassium channel Kv1.1, which is expressed at appreciable levels. The second ortholog is similar to Kv1.5 but has fewer N-terminal amino acids than other species. The sequence for Kv1.5 in the skate is very strongly aligned with the recently reported sequence for potassium channels in the electroreceptors of the cat shark, S. retifer, which also modulate synaptic transmission. The latter channel was designated as Kv1.3 in the initial report, but we suggest that these channels are actually orthologs of each other, and that Kv1.5 is the prevailing designation. We also found a beta subunit sequence (Kcnab2) which may co-assemble with one or both of the voltage gated channels. The new channels and subunits were verified by RT-PCR and the Kv1.1 sequence was confirmed by cloning. We also searched the RNA-seq repository for accessory subunits of Kcnma1, and found a computer-generated assembly that contained a complete sequence of its ß subunit, Kcnmb2. Skate Kcnmb2 has a total of 279 amino acids, with 51 novel amino acids at the N-terminus which may play a specific physiological role. This sequence was confirmed by PCR and cloning. However, skate Kcnmb2 is expressed at low levels in the electroreceptor compared to Kcnma1 and skate Kcnmb1 is absent. The evolutionary origin of the newly described K channels and their subunits was studied by alignments with mammalian sequences, including human, and also those in related fish: the whale shark (R. typus), the ghost shark (C.milii), and (S. retifer). There are also orthologous K channels of the lamprey, which has electroreceptors. Tree building and bootstrap programs were used to confirm phylogenetic inferences. Further research should focus on the subcellular locations of these channels, their gating behavior, and the effects of accessory subunits on gating.


Assuntos
Clonagem Molecular , Proteínas de Peixes/genética , Canal de Potássio Kv1.1/genética , Canal de Potássio Kv1.5/genética , Subunidades alfa do Canal de Potássio Ativado por Cálcio de Condutância Alta/genética , Filogenia , Raias/genética , Animais , Proteínas de Peixes/metabolismo , Sequenciamento de Nucleotídeos em Larga Escala , Canal de Potássio Kv1.1/metabolismo , Canal de Potássio Kv1.5/metabolismo , Subunidades alfa do Canal de Potássio Ativado por Cálcio de Condutância Alta/metabolismo , Raias/metabolismo , Especificidade da Espécie
6.
Gene ; 712: 143945, 2019 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-31279712

RESUMO

Reactive oxygen species, generated in all the aerobic organisms, can cause oxidative stress. Excessive ROS may become a source of carcinogen due to DNA damage, lipid peroxidation, cell injury, and cell death. In order to prevent these adverse effects of ROS, antioxidant enzymes have evolved in aerobic organisms. Catalase is a major antioxidant enzyme that breaks down excessive H2O2 and inhibits apoptotic cell death. Here we molecularly characterized catalase from red-lip mullet. The cDNA sequence of LhCAT consists of an ORF of 1545 bp, which encodes a 527 amino acid peptide (~60 kDa). Based on bioinformatics analysis, LhCAT possesses a domain architecture characteristic of catalases, including a catalase proximal active site signature and a catalase proximal heme-ligand signature. It also has heme and NADPH binding sites homologous to previously described catalases. Pairwise alignment with its homologs revealed that LhCAT shares 95.1% identity with Oplegnathus fasciatus catalase and 97.4% similarity with Sparus aurata catalase. An uprooted phylogenetic tree demonstrated that LhCAT resides in a clade with catalases from other teleosts and exhibits a close relationship with Oplegnathus fasciatus catalase. Among twelve tissue types, we observed the highest LhCAT mRNA expression in the liver, followed by blood. Immune challenge by Lactococcus garvieae, or Poly I:C in the blood or spleen resulted in up-regulation at 24 h post injection. We also tested the antioxidant activity of recombinant LhCAT against hydrogen peroxide and found its optimal concentration to be 12.5 µg/mL. Collectively, these data suggested that LhCAT play an important role in antioxidant defense and immune response of red-lip mullet.


Assuntos
Catalase/metabolismo , Proteínas de Peixes/metabolismo , Smegmamorpha , Adjuvantes Imunológicos , Animais , Antioxidantes/metabolismo , Catalase/genética , DNA Complementar/genética , Proteínas de Peixes/genética , Perfilação da Expressão Gênica , Regulação Enzimológica da Expressão Gênica , Heme/química , Peróxido de Hidrogênio/química , Sistema Imunitário , Ligantes , Fígado/enzimologia , Estresse Oxidativo , Filogenia , RNA Mensageiro/metabolismo , Espécies Reativas de Oxigênio , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Regulação para Cima
7.
Dokl Biochem Biophys ; 485(1): 119-122, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31201629

RESUMO

Samples from 11 populations of the Arctic char of the North-European part of Russia belonging to the anadromous and resident forms and two samples from Lake Sobach'e (Taimyr) were studied. The nucleotide sequence of the mitochondrial COI gene was determined in 60 individuals. In the majority of populations, the same COI haplotype was found. In some populations of the resident chars, haplotypes differing from the widespread haplotype in a single nucleotide substitution were found. The obtained genetic data give no reason to distinguish the resident form of the Arctic char from lakes of Karelia and the Kola Peninsula as an independent species, Salvelinus lepechini. The adaptation of the Arctic char to the unstable environmental conditions is ensured primarily by its phenotypic plasticity.


Assuntos
DNA Mitocondrial/genética , Complexo IV da Cadeia de Transporte de Elétrons/genética , Proteínas de Peixes/genética , Proteínas Mitocondriais/genética , Filogenia , Truta/genética , Animais , Europa (Continente)
8.
Artigo em Inglês | MEDLINE | ID: mdl-31176866

RESUMO

Cysteine oxygenase (CDO) is a mononuclear nonhemoglobin enzyme that catalyzes the production of taurine through the cysteine (Cys) pathway and plays a key role in the biosynthesis of taurine in mammals. However, the function of CDOs in bony fish remains poorly understood. In this study, we cloned CDO genes (CaCDO1 and CaCDO2) from Carassius auratus. The cDNA sequences of both CaCDO1 and CaCDO2 encoded putative proteins with 201 amino acids, which included structural features typical of the CDO protein family. Multiple sequence alignment and phylogenetic analysis showed that CaCDO1 and CaCDO2 shared high sequence identities and similarities with C. carpio homologs. Quantitative real-time polymerase chain reaction (qRT-PCR) results revealed that CaCDO1 and CaCDO2 were both broadly expressed in all selected tissues and developmental stages in C. auratus but had differing mRNA levels. In addition, compared to those of the taurine-free group, the in vivo mRNA expression levels of both CaCDO1 and CaCDO2 significantly decreased with increasing dietary taurine levels from 1.0 to 9.0 g/kg. Furthermore, in vitro taurine treatments showed similar inhibitory effects on the expression of CaCDO1 and CaCDO2 in the intestines of C. auratus. Our results also showed that the mRNA expression of CaCDO2 in the intestines was higher than that of CaCDO1 in response to in vivo and in vitro taurine supplementation. Overall, these data may provide new insights into the regulation of fish CDO expression and provide valuable knowledge for improving dietary formulas in aquaculture.


Assuntos
Cisteína Dioxigenase/genética , Cisteína Dioxigenase/metabolismo , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Carpa Dourada/genética , Carpa Dourada/metabolismo , Taurina/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , DNA Complementar/genética , Regulação da Expressão Gênica no Desenvolvimento , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Carpa Dourada/crescimento & desenvolvimento , Isoenzimas/genética , Isoenzimas/metabolismo , Filogenia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Taurina/farmacologia , Distribuição Tecidual
9.
Artigo em Inglês | MEDLINE | ID: mdl-31176867

RESUMO

In rice field eel (Monopterus albus), germ cell development in the developing gonad has been revealed in detail. However, it is unclear how primordial germ cells (PGCs) migrate to the somatic part of the gonad (genital ridge). This study visualized PGC migration by injecting a chimeric mRNA containing a fluorescent protein fused to the 3' untranslated region (3'UTR) of three different genes, nanos3 of zebrafish (Danio rerio) and dead end (dnd) and vasa of rice field eel. The mRNAs were injected either alone or in pairs into embryos at the one-cell stage. The results showed that mRNAs containing nanos3 and dnd 3'UTRs labeled PGCs over a wider time frame than those containing vasa 3'UTR, suggesting that nanos3 and dnd 3'UTRs are suitable for visualizing PGCs in rice field eel. Using this direct visualization method, the normal migration route of PGCs was observed from the 50%-epiboly stage to hatching stage for the first time, and the ectopic PGCs were also visualized during this period in rice field eel. These findings extend our knowledge of germ cell development, and lay a foundation for further research on the relationship between PGCs and sex differentiation, and on incubation conditions for embryos in rice field eel.


Assuntos
Células Germinativas/metabolismo , Smegmamorpha/embriologia , Regiões 3' não Traduzidas , Sequência de Aminoácidos , Animais , Movimento Celular/genética , Movimento Celular/fisiologia , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/metabolismo , Feminino , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Células Germinativas/citologia , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Masculino , RNA/genética , RNA Mensageiro/genética , Proteínas de Ligação a RNA/genética , Proteínas de Ligação a RNA/metabolismo , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Homologia de Sequência de Aminoácidos , Smegmamorpha/genética , Smegmamorpha/metabolismo , Proteínas de Peixe-Zebra/genética , Proteínas de Peixe-Zebra/metabolismo
10.
Mar Pollut Bull ; 142: 253-262, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31232302

RESUMO

We used manually spawned, field-deployed embryos of a common marine fish species, Pacific herring (Clupea pallasii), to evaluate accumulation of polycyclic aromatic hydrocarbons (PAHs) associated with an incomplete creosote-treated piling (CTP) removal project. Embryos near undisturbed 100-year-old CTPs (before removal) accumulated higher PAHs and exhibited higher cyp1a gene expression than embryos from reference areas. Embryos incubated close to CTP debris after CTP removal showed PAHs 90 times higher than reference areas up to a year after CTP removal. cyp1a fold-induction correlated with total embryo PAHs in all three years. Patterns of individual PAH chemicals differed slightly between embryos, wood sampled from CTPs, and passive samplers. This study illustrates the importance of using appropriate techniques and procedures to remove CTPs in aquatic environments to prevent release of toxic chemicals. Of particular concern is that incomplete CTP removal could expose sensitive life stages of fishes to chemicals that may reduce their survival.


Assuntos
Creosoto , Peixes/embriologia , Hidrocarbonetos Policíclicos Aromáticos/análise , Poluentes Químicos da Água/análise , Animais , Citocromo P-450 CYP1A1/genética , Ecossistema , Ecotoxicologia/métodos , Embrião não Mamífero/efeitos dos fármacos , Embrião não Mamífero/metabolismo , Proteínas de Peixes/genética , Peixes/fisiologia , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Hidrocarbonetos Policíclicos Aromáticos/toxicidade , Washington , Poluentes Químicos da Água/toxicidade , Madeira
11.
Ecotoxicol Environ Saf ; 181: 130-137, 2019 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-31176247

RESUMO

The pikeperch Sander lucioperca is an economically important freshwater species that is currently threatened by higher summer temperatures caused by global warming. To clarify the physiological state of pikeperch reared under relatively high temperatures and to acquire valuable biomarkers to monitor heat stress in this species, 100 fish were subjected to five different temperature treatments, ranging from 23 °C (control) to 36 °C. The physiological and biochemical indexes of liver and blood were determined, and heat-shock cognate 70 kDa protein (Hsc70) mRNA expression profiles were analyzed. The results showed that the activities of superoxide dismutase, catalase, and glutathione peroxidase in heat-stressed pikeperch first increased and then decreased, exhibiting peaks at 34 °C, 28 °C, and 28 °C, respectively. The level of thiobarbituric acid-reactive substances (TBARS) in all experimental groups was significantly higher than that of the control. The numbers of red blood cells, the packed-cell volume, and the contents of hemoglobin were significantly higher in the 34 °C and 36 °C treatment groups. Under heat stress, the albumin, cholesterol, and triglycerides contents decreased with increasing temperatures. Real-time fluorescence-based quantitative RT-PCR showed that Hsc70 mRNA levels increased in all eight of the tested tissues under heat stress. Expression reached maximum levels at 34 °C in the muscle, heart and gill tissues, and at 36 °C in the other five tissues. These results demonstrate that several physiological and biochemical phenotypes, such as oxidative stress, antioxidant enzymes and molecular chaperones, could be important biomarkers of heat stress in pikeperch, and are potentially valuable to uncover the mechanisms of heat-stress responses in fish.


Assuntos
Proteínas de Peixes/metabolismo , Proteínas de Choque Térmico HSC70/metabolismo , Resposta ao Choque Térmico , Percas/metabolismo , Animais , Antioxidantes/metabolismo , Catalase/metabolismo , Contagem de Eritrócitos , Proteínas de Peixes/genética , Glutationa Peroxidase/metabolismo , Proteínas de Choque Térmico HSC70/genética , Resposta ao Choque Térmico/genética , Hemoglobinas/análise , Fígado/enzimologia , Fígado/metabolismo , Estresse Oxidativo , Percas/genética , Superóxido Dismutase/metabolismo , Substâncias Reativas com Ácido Tiobarbitúrico/metabolismo
12.
Cytogenet Genome Res ; 158(2): 106-113, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31203273

RESUMO

The karyotypes of the family Parodontidae consist of 2n = 54 chromosomes. The main chromosomal evolutionary changes of its species are attributed to chromosome rearrangements in repetitive DNA regions in their genomes. Physical mapping of the H1 and H4 histones was performed in 7 Parodontidae species to analyze the chromosome rearrangements involved in karyotype diversification in the group. In parallel, the observation of a partial sequence of an endogenous retrovirus (ERV) retrotransposon in the H1 histone sequence was evaluated to verify molecular co-option of the transposable elements (TEs) and to assess paralogous sequence dispersion in the karyotypes. Six of the studied species had an interstitial histone gene cluster in the short arm of the autosomal pair 13. Besides this interstitial cluster, in Apareiodon davisi, a probable further site was detected in the terminal region of the long arm in the same chromosome pair. The H1/H4 clusters in Parodon cf. pongoensis were located in the smallest chromosomes (pair 20). In addition, scattered H1 signals were observed on the chromosomes in all species. The H1 sequence showed an ERV in the open reading frame (ORF), and the scattered H1 signals on the chromosomes were attributed to the ERV's location. The H4 sequence had no similarity to the TEs and displayed no dispersed signals. Furthermore, the degeneration of the inner ERV in the H1 sequence (which overlapped a stretch of the H1 ORF) was discussed regarding the likelihood of molecular co-option of this retroelement in histone gene function in Parodontidae.


Assuntos
Caraciformes/genética , Mapeamento Cromossômico/veterinária , Histonas/genética , Animais , Caraciformes/metabolismo , Feminino , Proteínas de Peixes/genética , Hibridização in Situ Fluorescente , Cariotipagem , Masculino , Retroelementos , Análise de Sequência de DNA/veterinária
13.
Fish Shellfish Immunol ; 91: 78-86, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31039439

RESUMO

Type I interferons, as a class of multipotent cytokines, play a key role in host antiviral immune responses. In this study, a type I IFN coding gene of gibel carp, Carassius auratus gibelio, CagIFNa was cloned and sequenced. The full-length cDNA sequence of CagIFNa consists of 724 nucleotides that encode a predicted protein of 183 amino acids. CagIFNa has two highly conserved cysteine residues in the deduced protein, which is mostly conserved in the fish group I type I IFNs. CagIFNa was identified as a member of the IFNa subgroup of group I type I IFNs by phylogenetic analysis. CagIFNa transcripts were detected in all investigated tissues with higher levels in the liver, intestine, spleen and head kidney of gibel carp. Following injection with Cyprinid herpesvirus 2 (CyHV-2), CagIFNa gene expression was significantly inhibited in the spleen but delayed and then increased in head kidneys. Similarly, while CagIFNa expression was rapidly induced in gibel carp brain (GiCB) cells by poly I:C stimulation and its high induction level was delayed following CyHV-2 infection. CagIFNa overexpression in GiCB cells drastically reduced virus CPE and titer. Furthermore, several genes associated with type I IFN signaling pathway including IRF3, IRF7, IRF9, STAT1, Mx1 and PKR were induced in GiCB cells overexpressing CagIFNa upon CyHV-2 infection. These results show that CagIFNa plays a role in antiviral immune system in gibel carp.


Assuntos
Doenças dos Peixes/imunologia , Regulação da Expressão Gênica/imunologia , Carpa Dourada/genética , Carpa Dourada/imunologia , Imunidade Inata/genética , Interferon Tipo I/genética , Interferon Tipo I/imunologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica/veterinária , Herpesviridae , Infecções por Herpesviridae/imunologia , Interferon Tipo I/química , Filogenia , Poli I-C/farmacologia , Distribuição Aleatória , Alinhamento de Sequência/veterinária
14.
Fish Shellfish Immunol ; 91: 87-98, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31082517

RESUMO

Excessive perfluorooctane sulfonate (PFOS) in natural water ecosystem has the potential to detrimentally affect immune system, but little is known of such effects or underlying mechanisms in fish. In the present study, we evaluated the effects of PFOS on growth performance, organizational microstructure, activities of immune-related enzymes and expressions of immune-related genes in male zebrafish (Danio rerio) exposed to different concentrations of 0, 0.02, 0.04 and 0.08 mg/L of PFOS for 7, 14, and 21 days or cotreatment with PFOS and PDTC to investigate the effects of PFOS on immune system and the potential toxic mechanisms caused by PFOS. The results indicated that PFOS accumulated in livers after exposure, and remarkably elevations were found in three exposure groups compared with the control group at three stages. The growth of the adult zebrafish in the experiments was significantly inhibited, the microstructures of liver were serious damaged. The ROS levels were remarkably increased. The activities of ACP, AKP, and lysozyme were obviously decreased, while the activities of MPO and NF-κB were significantly increased. The expressions of immune-related mRNA were significantly affected. After co-treatment with PFOS and PDTC, the growth inhibition, the morphological damage, the ROS induction, and the expressions of immune-related mRNA were reversed. Taken together, the results indicated that PFOS can significantly inhibit the growth, disturb the immune system by changing the normal structure of liver, the activities of immune-related enzymes, and a series of gene transcriptions involved in immune regulation in liver of male zebrafish. PFOS-induced pro-inflammatory effect of hepatocytes was observed, and the involvement of NF-κB signaling pathway was participated in its action mechanism. These findings provide further evidence that PFOS interferes with the immune regulation of liver of male zebrafish under in vivo conditions.


Assuntos
Ácidos Alcanossulfônicos/toxicidade , Proteínas de Peixes/imunologia , Fluorcarbonetos/toxicidade , Fígado/efeitos dos fármacos , NF-kappa B/imunologia , Poluentes Químicos da Água/toxicidade , Peixe-Zebra/fisiologia , Animais , Relação Dose-Resposta a Droga , Proteínas de Peixes/genética , Fígado/metabolismo , Masculino , NF-kappa B/genética , Distribuição Aleatória , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Tiocarbamatos/toxicidade , Peixe-Zebra/genética , Peixe-Zebra/crescimento & desenvolvimento , Peixe-Zebra/imunologia
15.
Fish Shellfish Immunol ; 91: 40-49, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31082519

RESUMO

DEAD (Asp-Glu-Ala-Asp)-box polypeptide 41 (DDX41) is a member of the DEXDc family of helicases, that has recently been identified to be a crucial intracellular DNA sensor that triggers multiple signaling molecules to activate the type I interferon response. However, the precise function of DDX41 in fish during a viral infection remains unknown. In the present study, the DDX41 homolog from orange spotted grouper, Epinephelus coioides (EcDDX41), was cloned and its potential role in the immune response to a fish viral infection were investigated. EcDDX41 encodes a putative protein of 614 amino acid residues that contained two conserved domains: 1) DEADc domain; and 2) HELICc domain. The sequence analysis indicated that EcDDX41 shared 99%, 94%, and 86% identity with Asian seabass (Lates calcarifer), zebrafish (Danio rerio), and humans (Homo sapiens), respectively. EcDDX41 mRNA was present in all of the detected tissues, with the highest level of expression in the gills. The level of EcDDX41 expression was up-regulated following infection with Singapore grouper iridovirus (SGIV) or red-spotted grouper nervous necrosis virus (RGNNV) in grouper spleen (GS) cell cultures, suggesting that EcDDX41 may be involved in fish virus infection. Furthermore, EcDDX41 overexpression in GS cells significantly inhibited SGIV and RGNNV replication. EcDDX41 overexpression significantly increased the expression of antiviral and inflammatory cytokine genes, including interferon regulatory factor genes (e.g., IRF1, IRF2, IRF3, and IRF7), interferon induced genes (e.g., ISG15, ISG56, IFP35, Viperin, and MXI), and pro-inflammatory cytokine genes (e.g., TNFα, IL-1ß, and IL-8). Moreover, EcDDX41 positively regulated the mitochondrial antiviral-signaling protein (MAVS) and TANK-binding kinase 1 (TBK1)-induced interferon immune response, but did mediate IRF3 activation (MITA) to evoke an interferon immune response in unstimulated cells. Together, our results provide novel insight into the role of fish DDX41 in the antiviral innate immune response.


Assuntos
Bass/genética , Bass/imunologia , RNA Helicases DEAD-box/genética , RNA Helicases DEAD-box/imunologia , Doenças dos Peixes/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Sequência de Aminoácidos , Animais , RNA Helicases DEAD-box/química , Infecções por Vírus de DNA/imunologia , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica/veterinária , Nodaviridae/fisiologia , Filogenia , Infecções por Vírus de RNA/imunologia , Ranavirus/fisiologia , Alinhamento de Sequência/veterinária
16.
Fish Shellfish Immunol ; 91: 148-158, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31082520

RESUMO

The Rho family GTPase Rac1 acts as a molecular switch for signal transduction to regulate various cellular functions. Here, a Rac1 homolog (LcRac1) was identified in large yellow croaker (Larimichthys crocea), one of the most economically important marine fishes. The LcRac1 protein was expressed in Escherichia coli and purified. Subsequently the specific antibody was raised using the purified fusion protein (GST-LcRac1). LcRac1 was ubiquitously expressed in all 12 tissues we examined, with the highest expression in heart and blood and the weakest expression in head-kidney and spleen. Moreover, time course analysis revealed that LcRac1 expression was obviously up-regulated in liver, spleen and head-kidney after immunization with Poly I:C, LPS and Vibrio parahemolyticus. On the other hand, on the basis of protein interaction, it was found that the LcRac1 interacted with Tropomyosin, a crucial protein in the process of phagocytosis. Furthermore, RNAi assays indicated that the phagocytic percentage and phagocytic index were significantly decreased when the LcRac1 gene was silenced by sequence-specific siRNA. Fluorescence microscopy assays revealed FITC-labeled V. parahemolyticus were remarkably decreased after LcRac1 was silenced by sequence-specific siRNA at 24 h. These findings implicate the vital role of LcRac1 in innate immunity in the large yellow croaker.


Assuntos
Doenças dos Peixes/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Perciformes/genética , Perciformes/imunologia , Proteínas rac1 de Ligação ao GTP/genética , Proteínas rac1 de Ligação ao GTP/imunologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica/veterinária , Lipopolissacarídeos/farmacologia , Fagocitose/genética , Filogenia , Poli I-C/farmacologia , Alinhamento de Sequência/veterinária , Tropomiosina , Vibrioses/imunologia , Vibrioses/veterinária , Vibrio parahaemolyticus/fisiologia , Proteínas rac1 de Ligação ao GTP/química
17.
Fish Shellfish Immunol ; 91: 1-11, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31085326

RESUMO

The immune mechanism elicited in pufferfish (Takifugu obscurus) against the invasion of Aeromonas hydrophila is still poorly understood. We examined the spleen of pufferfish at the transcriptome and proteome levels by using Illumina-seq and TMT coupled mass spectrometry after 12 h infection by A. hydrophila, respectively. A total of 2,339 genes (1,512 up-regulated and 827 down-regulated) and 537 (237 up-regulated and 300 down-regulated) proteins were identified. GO and KEGG analyses revealed that the responses to stimulus were the main biological processes, intestinal immune network for IgT production and calcium signaling pathway. Fourteen genes (8 up-regulated and 6 down-regulated) and proteins (5 up-regulated and 9 down-regulated) involved immune responses or signal transduction were validated by qRT-PCR and parallel reaction monitoring to confirm the reliability of the transcriptomic and proteomic analyses, respectively. Moreover, qRT-PCR and flow cytometry were used to detect dynamics of the genes in calcium signaling pathway and changes of concentration of cytoplasm Ca2+ in spleen cells within a 72 h challenge. This study provides the findings regarding immune response, especially intestinal immune network for IgT production pathway and calcium signaling pathway at the molecular, protein and cellular in pufferfish after infection by A. hydrophila. These results would provide a new insight and molecular targets into the response to pathogenic infection in pufferfish.


Assuntos
Doenças dos Peixes/imunologia , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Regulação da Expressão Gênica/imunologia , Baço/imunologia , Takifugu/genética , Takifugu/imunologia , Aeromonas hydrophila/fisiologia , Animais , Regulação para Baixo , Doenças dos Peixes/microbiologia , Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Negativas/microbiologia , Infecções por Bactérias Gram-Negativas/veterinária , Proteoma/genética , Proteoma/imunologia , Transcriptoma , Regulação para Cima
18.
Fish Shellfish Immunol ; 91: 108-121, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31091461

RESUMO

As an adaptor in Toll-like receptor (TLR) signaling pathway, Toll/interleukin-1 receptor (TIR) domain containing adaptor inducing interferon-ß (TRIF) mediates downstream signaling cascades and plays important roles in host innate immune responses. In the present study, a TRIF ortholog named Lc-TRIF was identified in large yellow croaker (Larimichthys crocea). Sequence comparison analysis revealed that Lc-TRIF has a conserved TIR domain but without TRAF6 binding motif. The genome structure of Lc-TRIF is conserved, with two exons and one intron. Syntenic comparison showed that the loci of fish TRIF was different from that in mammals or birds, and TRAM was absent in the genomes of fish, amphibians, and birds, but present in mammals and reptiles. Expression analysis revealed that Lc-TRIF was broadly expressed in examined organs/tissues, with the highest expression level in gill and weakest in brain, and could be up-regulated under poly I:C, LPS, PGN, and Pseudomonas plecoglossicida stimulation. Fluorescence microscopy results showed that Lc-TRIF exhibited a global localization throughout the entire cell including the nucleus in HEK 293T cells. Additionally, luciferase assays demonstrated that Lc-TRIF expression could significantly induce NF-κB, type I IFN, IRF3 as well as IRF7 promoter activation. These results collectively indicated that Lc-TRIF was function in host antiviral and antibacterial responses via NF-κB and IRF3/7 related signaling pathway.


Assuntos
Proteínas Adaptadoras de Transporte Vesicular/genética , Proteínas Adaptadoras de Transporte Vesicular/imunologia , Doenças dos Peixes/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Perciformes/genética , Perciformes/imunologia , Proteínas Adaptadoras de Transporte Vesicular/química , Sequência de Aminoácidos , Animais , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica/veterinária , Filogenia , Alinhamento de Sequência/veterinária
19.
Fish Shellfish Immunol ; 91: 68-77, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31096060

RESUMO

Mannose-binding lectin-associated serine protease-1 (MASP-1), a multifunctional serine protease, plays an important role in innate immunity which is capable of activating the lectin pathway of the complement system and also triggering coagulation cascade system. In this study, a MASP-1 homolog (OnMASP-1) was identified from Nile tilapia (Oreochromis niloticus) and characterized at expression and inflammation functional levels. The open reading frame (ORF) of OnMASP-1 is 2187 bp of nucleotide sequence encoding a polypeptide of 728 amino acids. The deduced amino acid sequence has 6 characteristic structures, including two C1r/C1s-Uegf-BMP domains (CUB), one epidermal growth factor domain (EGF), two complement control protein domains (CCP) and a catalytic serine protease domain (SP). Expression analysis revealed that the OnMASP-1 was highly expressed in the liver, and widely exhibited in other tissues containing intestine, spleen and kidney. In addition, the OnMASP-1 expression was significantly up-regulated in spleen and head kidney following challenges with Streptococcus agalactiae and Aeromonas hydrophila. The up-regulations of OnMASP-1 mRNA and protein expression were also demonstrated in hepatocytes and monocytes/macrophages in vitro stimulation with S. agalactiae and A. hydrophila. Recombinant OnMASP-1 protein was likely to participate in the regulation of inflammatory and migration reaction by monocytes/macrophages. These results indicated that OnMASP-1, playing an important role in innate immunity, was likely to involve in host defense against bacterial infection in Nile tilapia.


Assuntos
Ciclídeos/genética , Ciclídeos/imunologia , Doenças dos Peixes/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Serina Proteases Associadas a Proteína de Ligação a Manose/genética , Serina Proteases Associadas a Proteína de Ligação a Manose/imunologia , Aeromonas hydrophila/fisiologia , Sequência de Aminoácidos , Animais , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica/veterinária , Infecções por Bactérias Gram-Negativas/imunologia , Serina Proteases Associadas a Proteína de Ligação a Manose/química , Filogenia , Alinhamento de Sequência/veterinária , Infecções Estreptocócicas/imunologia , Streptococcus agalactiae/fisiologia
20.
Fish Shellfish Immunol ; 91: 29-39, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31100439

RESUMO

The present study aim to investigate the effects of dietary Gelsemium elegans alkaloids supplementation in Megalobrama amblycephala. A basal diet supplemented with 0, 5, 10, 20 and 40 mg/kg G. elegans alkaloids were fed to M. amblycephala for 12 weeks. The study indicated that dietary 20 mg/kg and 40 mg/kg G. elegans alkaloids supplementation could significantly improve final body weight (FBW), weight gain rate (WGR), specific growth rate (SGR), feed conversion ratio (FCR) and protein efficiency ratio (PER) (P < 0.05). The 20 mg/kg and 40 mg/kg G. elegans alkaloids groups showed significantly higher whole body and muscle crude protein and crude lipid contents compared to the control group (P < 0.05). The amino acid contents in muscle were also significantly increased in 20 mg/kg and 40 mg/kg groups (P < 0.05). Dietary 40 mg/kg G. elegans alkaloids had a significant effect on the contents of LDH, AST, ALT, ALP, TG, TC, LDL-C, HDL-C, ALB and TP in M. amblycephala (P < 0.05). Fish fed 20 mg/kg and 40 mg/kg dietary G. elegans alkaloids showed significant increase in complement 3, complement 4 and immunoglobulin M contents. The liver antioxidant enzymes (SOD, CAT and T-AOC) and MDA content significantly increased at 20 mg/kg and 40 mg/kg G. elegans alkaloids supplement (P < 0.05). The mRNA levels of immune-related genes IL-1ß, IL8, TNF-α and IFN-α were significantly up-regulated, whereas TGF-ß and IL10 genes were significantly down-regulated in the liver, spleen and head kidney of fish fed dietary supplementation with 20 mg/kg and 40 mg/kg G. elegans alkaloids. After challenge with Aeromonas hydrophila, significant higher survival rate was observed at 20 mg/kg and 40 mg/kg G. elegans alkaloids supplement (P < 0.05). Therefore, these results indicated that M. amblycephala fed a diet supplemented with 20 mg/kg and 40 mg/kg G. elegans alkaloids could significantly promote its growth performance, lipids and amino acids deposition, immune ability and resistance to Aeromonas hydrophila.


Assuntos
Cyprinidae/imunologia , Resistência à Doença/imunologia , Doenças dos Peixes/prevenção & controle , Gelsemium/química , Extratos Vegetais/farmacologia , Adjuvantes Imunológicos/farmacologia , Aeromonas hydrophila/fisiologia , Alcaloides/química , Alcaloides/farmacologia , Ração Animal/análise , Animais , Cyprinidae/crescimento & desenvolvimento , Dieta/veterinária , Suplementos Nutricionais/análise , Relação Dose-Resposta a Droga , Doenças dos Peixes/imunologia , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Negativas/prevenção & controle , Infecções por Bactérias Gram-Negativas/veterinária , Extratos Vegetais/química
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