RESUMO
SP3 (specificity protein 3) is a transcription factor characterized by three conserved Cys2His2 zinc finger motifs that exert a transregulatory effect by binding to GC boxes, either upregulating or downregulating multiple genes or by co-regulating gene expression in coordination with other proteins. SP3 potentially regulates a series of processes, such as the cell cycle, growth, metabolic pathways, and apoptosis, and plays an important role in antiviral effect. The function of sp3 in fish is poorly understood. In this study, the Sp3a open reading frame was cloned from the orange-spotted grouper, Epinephelus coioides. The full-length open reading frame of Sp3a was 2034 bp, encoding 677 amino acids, with a predicted molecular weight of 72.34 kDa and an isoelectric point of 5.05. Phylogenetically, Sp3a in Epinephelus coioides was the most closely related to Sp3a in the Malabar grouper, Epinephelus malabaricus. RT-qPCR revealed ubiquitous expression of Sp3a in all examined grouper tissues, with no significant differences in expression levels among tissues. A eukaryotic expression vector, pEGFP-Sp3a, was constructed and transfected into grouper spleen (GS) cells. Subcellular localization of Sp3a was observed using an inverted fluorescence microscope. When Spa3 was overexpressed in GS cells, the expression of orange-spotted grouper nerve necrosis virus (RGNNV) genes (CP and RdRp) decreased significantly, indicating that Sp3a significantly inhibited RGNNV replication. siRNA inhibition of Sp3a accelerated the intracellular replication of RGNNV, implying the antiviral effect of Sp3a. Conclusively, our findings contribute to further research on the antiviral capabilities of Sp3a in grouper and other fish. Therefore, our research has potential implications on the development of the aquaculture industry.
Assuntos
Bass , Doenças dos Peixes , Proteínas de Peixes , Animais , Doenças dos Peixes/virologia , Doenças dos Peixes/genética , Proteínas de Peixes/genética , Proteínas de Peixes/metabolismo , Bass/genética , Bass/virologia , Fator de Transcrição Sp3/metabolismo , Fator de Transcrição Sp3/genética , Filogenia , Nodaviridae/genética , Clonagem Molecular , Infecções por Vírus de RNA/veterinária , Infecções por Vírus de RNA/virologia , Infecções por Vírus de RNA/genética , Infecções por Vírus de DNA/veterinária , Infecções por Vírus de DNA/virologia , Infecções por Vírus de DNA/genética , Sequência de AminoácidosRESUMO
The inflammasome is a multimeric protein complex that plays a vital role in the defence against pathogens and is therefore considered an essential component of the innate immune system. In this study, the expression patterns of inflammasome genes (NLRC3, ASC, and CAS-1), antiviral genes (IFNγ and MX), and immune genes (IL-1ß and IL-18) were analysed in Oreochromis niloticus liver (ONIL) cells following stimulation with the bacterial ligands peptidoglycan (PGN) and lipopolysaccharide (LPS) and infection with TiLV. The cells were stimulated with PGN and LPS at concentrations of 10, 25, and 50 µg/ml. For viral infection, 106 TCID50 of TiLV per ml was used. After LPS stimulation, all seven genes were found to be expressed at specific time points at each of the three doses tested. However, at even higher doses of LPS, NLRC3 levels decreased. Following TiLV infection, all of the genes showed significant upregulation, especially at early time points. However, the gene expression pattern was found to be unique in PGN-treated cells. For instance, NLRC3 and ASC did not show any response to PGN stimulation, and the expression of IFNγ was downregulated at 25 and 50 µg of PGN per ml. CAS-1 and IL-18 expression was downregulated at 25 µg of PGN per ml. At a higher dose (50 µg/ml), IL-1ß showed downregulation. Overall, our results indicate that these genes are involved in the immune response to viral and bacterial infection and that the degree of response is ligand- and dose-dependent.
Assuntos
Ciclídeos , Doenças dos Peixes , Inflamassomos , Animais , Ciclídeos/imunologia , Ciclídeos/genética , Inflamassomos/genética , Inflamassomos/imunologia , Inflamassomos/metabolismo , Doenças dos Peixes/imunologia , Doenças dos Peixes/virologia , Doenças dos Peixes/microbiologia , Doenças dos Peixes/genética , Linhagem Celular , Peptidoglicano/farmacologia , Fígado/virologia , Fígado/imunologia , Lipopolissacarídeos/farmacologia , Imunidade Inata , Proteínas de Peixes/genética , Interleucina-18/genética , Interleucina-18/metabolismo , Ligantes , Infecções por Vírus de DNA/imunologia , Infecções por Vírus de DNA/veterinária , Infecções por Vírus de DNA/virologia , Infecções por Vírus de DNA/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/imunologia , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Interleucina-1beta/imunologiaRESUMO
Grouper is one of the most important and valuable mariculture fish in China, with a high economic value. As the production of grouper has increased, massive outbreaks of epidemic diseases have limited the development of the industry. Singapore grouper iridovirus (SGIV) is one of the most serious infectious viral pathogens and has caused huge economic losses to grouper farming worldwide due to its rapid spread and high lethality. To find new strategies for the effective prevention and control of SGIV, we constructed two chimeric DNA vaccines using Lysosome-associated membrane protein 1 (LAMP1) fused with major capsid proteins (MCP) against SGIV. In addition, we evaluated the immune protective effects of vaccines including pcDNA3.1-3HA, pcDNA3.1-MCP, pcDNA3.1-LAMP1, chimeric DNA vaccine pcDNA3.1-MLAMP and pcDNA3.1-LAMCP by intramuscular injection. Our results showed that compared with groups injected with PBS, pcDNA3.1-3HA, pcDNA3.1-LAMP1 or pcDNA3.1-MCP, the antibody titer significantly increased in the chimeric vaccine groups. Moreover, the mRNA levels of immune-related factors in groupers, including IRF3, MHC-I, TNF-α, and CD8, showed the same trend. However, MHC-II and CD4 were significantly increased only in the chimeric vaccine groups. After 28 days of vaccination, groupers were challenged with SGIV, and mortality was documented for each group within 14 days. The data showed that two chimeric DNA vaccines provided 87 % and 91 % immune protection for groupers which were significantly higher than the 52 % protection rate of pcDNA3.1-MCP group, indicating that both forms of LAMP1 chimeric vaccines possessed higher immune protection against SGIV, providing the theoretical foundation for the creation of novel DNA vaccines for fish.
Assuntos
Bass , Infecções por Vírus de DNA , Doenças dos Peixes , Iridovirus , Ranavirus , Vacinas de DNA , Animais , Singapura , Fatores de Transcrição , Infecções por Vírus de DNA/prevenção & controle , Infecções por Vírus de DNA/veterinária , Infecções por Vírus de DNA/genética , Proteínas de Peixes/genéticaRESUMO
PACT (interferon-inducible double-stranded RNA-dependent protein kinase activator A) is a cellular protein which can activate PKR in dsRNA-independent manner. However, the role of PACT in fish virus infection remains largely unknown. In this study, a PACT homologue from grouper (Epinephelus coioides)(EcPACT) was cloned and characterized. The open reading frame of EcPACT has a full length of 924 bp and encodes a protein of 307 amino acids with a predicted molecular weight of 33.29 kDa. Similar to mammals, EcPACT contains three dsRBD domains. EcPACT shares 99.67 % homology with E. lanceolatus. Real-time fluorescence quantitative PCR results showed that EcPACT mRNA was widely expressed in all tissues and abundantly expressed in brain, blood, head kidney and kidney. In addition, SGIV and RGNNV infection significantly upregulated the transcript levels of EcPACT. Subcellular localization analysis showed that EcPACT was mainly distributed in the nucleus. Overexpression of EcPACT inhibited the replication of SGIV and RGNNV in vitro and positively regulated the expression of interferon (IFN) and pro-inflammatory factors. The results provide a better understanding of the relationship between PACT and viral infection in fish.
Assuntos
Bass , Infecções por Vírus de DNA , Doenças dos Peixes , Viroses , Animais , Sequência de Aminoácidos , Proteínas de Peixes/genética , Proteínas de Peixes/química , Bass/genética , Interferons/genética , Infecções por Vírus de DNA/genética , Imunidade Inata/genética , Filogenia , MamíferosRESUMO
Lymphocystis disease is frequently prevalent and transmissible in various teleost species worldwide due to lymphocystis disease virus (LCDV) infection, causing unsightly growths of benign lymphocystis nodules in fish and resulting in huge economic losses to aquaculture industry. However, the molecular mechanism of lymphocystis formation is unclear. In this study, LCDV was firstly detected in naturally infected flounder (Paralichthys olivaceus) by PCR, histopathological, and immunological techniques. To further understand lymphocystis formation, transcriptome sequencing of skin nodule tissue was performed by using healthy flounder skin as a control. In total, RNA-seq produced 99.36%-99.71% clean reads of raw reads, of which 91.11%-92.89% reads were successfully matched to the flounder genome. The transcriptome data showed good reproducibility between samples, with 3781 up-regulated and 2280 down-regulated differentially expressed genes. GSEA analysis revealed activation of Wnt signaling pathway, Hedgehog signaling pathway, Cell cycle, and Basal cell carcinoma associated with nodule formation. These pathways were analyzed to interact with multiple viral infection and tumor formation pathways. Heat map and protein interaction analysis revealed that these pathways regulated the expression of cell cycle-related genes such as ccnd1 and ccnd2 through key genes including ctnnb1, lef1, tcf3, gli2, and gli3 to promote cell proliferation. Additionally, cGMP-PKG signaling pathway, Calcium signaling pathway, ECM-receptor interaction, and Cytokine-cytokine receptor interaction associated with nodule formation were significantly down-regulated. Among these pathways, tnfsf12, tnfrsf1a, and tnfrsf19, associated with pro-apoptosis, and vdac2, which promotes viral replication by inhibiting apoptosis, were significantly up-regulated. Visual analysis revealed significant down-regulation of cytc, which expresses the pro-apoptotic protein cytochrome C, as well as phb and phb2, which have anti-tumor activity, however, casp3 was significantly up-regulated. Moreover, bcl9, bcl11a, and bcl-xl, which promote cell proliferation and inhibit apoptosis, were significantly upregulated, as were fgfr1, fgfr2, and fgfr3, which are related to tumor formation. Furthermore, RNA-seq data were validated by qRT-PCR, and LCDV copy numbers and expression patterns of focused genes in various tissues were also investigated. These results clarified the pathways and differentially expressed genes associated with lymphocystis nodule development caused by LCDV infection in flounder for the first time, providing a new breakthrough in molecular mechanisms of lymphocystis formation in fish.
Assuntos
Infecções por Vírus de DNA , Linguado , Iridoviridae , Animais , Linguado/genética , Proteínas Hedgehog , Reprodutibilidade dos Testes , Infecções por Vírus de DNA/genética , Infecções por Vírus de DNA/veterinária , Infecções por Vírus de DNA/metabolismo , Perfilação da Expressão Gênica , Iridoviridae/fisiologiaRESUMO
This study investigated the kinetics of red sea bream iridovirus and host gene expression during infection in rock bream (Oplegnathus fasciatus), a species highly sensitive to the virus. After intraperitoneal injection of the viral solution at 104 TCID50/fish, the viral genome copy number in the spleen was 104.7 ± 0.2 and 105.9 ± 0.4 copies/µg DNA at 3 and 5 days post-injection (dpi), respectively. Using transcriptomic analyses via MiSeq, viral gene transcripts were detected at 3 and 5 dpi. Six genes including RING-finger domain-containing protein and laminin-type epidermal growth factor-like domain genes were significantly expressed at 5 dpi. Further, 334 host genes were differentially expressed compared with those before infection. Genes were clustered into four groups based on their expression profiles. Interferon-stimulated genes were more prevalent in groups showing upregulation at 5 dpi and 3 and 5 dpi. In contrast, the group showing downregulation at 3 dpi included inflammation-related genes, such as granzyme and eosinophil peroxidase genes. Downregulation of certain inflammation-related genes may contribute to the susceptibility of this fish to the virus.
Assuntos
Infecções por Vírus de DNA , Doenças dos Peixes , Iridoviridae , Iridovirus , Perciformes , Dourada , Animais , Iridoviridae/fisiologia , Baço , Perciformes/genética , Inflamação , Infecções por Vírus de DNA/genética , Infecções por Vírus de DNA/veterinária , Proteínas de Peixes/genética , FilogeniaRESUMO
BACKGROUND: Megalocytiviruses (MCV) are double-stranded DNA viruses that infect fish. Two species within the genus are epidemiologically important for fish farming: red sea bream iridovirus (RSIV) and infectious spleen and kidney necrosis virus (ISKNV). The objective of this work was to study regions that allow the differentiation and correct diagnosis of RSIV and ISKNV. METHODS: The regions ORF450L, ORF342L, ORF077, and the intergenic region between ORF37 and ORF42R were sequenced and compared with samples from the database. RESULTS: The tree constructed using the sequencing of the PCR product Megalocytivirus. ORF077 separated the three major clades of MCV. RISV genotypes were well divided, but not ISKNV. All qPCRs tests showed acceptable repeatability values, that is, less than 5%. CONCLUSION: Two qPCRs for ISKNV detection and two for RSIV were considered suitable for use in the diagnosis and typing of MCV. The results of this study demonstrate the importance of an accurate evaluation of methodologies for the differentiation of MCV.
Assuntos
Infecções por Vírus de DNA , Doenças dos Peixes , Iridoviridae , Iridovirus , Animais , Iridoviridae/genética , Reação em Cadeia da Polimerase em Tempo Real , Infecções por Vírus de DNA/genética , Infecções por Vírus de DNA/veterinária , FilogeniaRESUMO
Protein kinase C (PKC) constitutes the main signal transduction pathway, and participates in the signal pathway of cell proliferation and movement in mammals. In this study, PKC-É was obtained from Epinephelus coioides, an important marine fish cultivated in the coastal areas of southern China and Southeast Asia. The full length cDNA of PKC-É was 3362 bp in length containing a 23 bp 5'UTR, a 1719 bp 3'UTR, and a 1620 bp open reading frame encoding 539 amino acids. It contains three conservative domains including protein kinase C conserved region 2 (C2), Serine/Threonine protein kinases, catalytic domain (S_TKc) and ser/thr-type protein kinases (S_TK_X). Its mRNA can be detected in all 11 tissues examined of E. coioides, and the expression was significantly upregulated response to Singapore grouper iridovirus (SGIV) infection, one of the important pathogens of marine fish. Upregulated E. coioides PKC-É significantly inhibited the activation of nuclear factor kappa-B (NF-κB) and activator protein-1 (AP-1), and SGIV-induced cell apoptosis. The results indicated that the PKC-É may play an important role in pathogenic stimulation.
Assuntos
Bass , Infecções por Vírus de DNA , Doenças dos Peixes , Iridovirus , Ranavirus , Animais , Bass/genética , Bass/metabolismo , Iridovirus/fisiologia , Singapura , Infecções por Vírus de DNA/genética , Proteínas de Peixes/metabolismo , Ranavirus/fisiologia , Proteína Quinase C/genética , Clonagem Molecular , Filogenia , Mamíferos/genéticaRESUMO
Iridoviruses are large DNA viruses which cause great economic losses to the aquaculture industry and serious threats to ecological diversity worldwide. Singapore grouper iridovirus (SGIV), a novel member of the genus Ranavirus, causes high mortality in grouper aquaculture. Previous work on genome annotation demonstrated that SGIV contained numerous uncharacterized or hypothetical open reading frames (ORFs), whose functions remained largely unknown. Here, we reported that the protein encoded by SGIV ORF131R (VP131) was localized predominantly within the endoplasmic reticulum (ER). Ectopic expression of GFP-VP131 significantly enhanced SGIV replication, while VP131 knockdown decreased viral infection in vitro, suggesting that VP131 functioned as a proviral factor during SGIV infection. Overexpression of GFP-VP131 inhibited the interferon (IFN)-1 promoter activity and mRNA level of IFN-related genes induced by poly(I:C), Epinephelus coioides cyclic GMP/AMP synthase (EccGAS)/stimulator of IFN genes (EcSTING), TANK-binding kinase 1 (EcTBK1), or melanoma differentiation-associated gene 5 (EcMDA5), whereas such activation induced by mitochondrial antiviral signaling protein (EcMAVS) was not affected. Moreover, VP131 interacted with EcSTING and degraded EcSTING through both the autophagy-lysosome pathway and ubiquitin-proteasome pathway, and targeted for the K63-linked ubiquitination. Of note, we also found that EcSTING significantly accelerated the formation of GFP-VP131 aggregates in co-transfected cells. Finally, GFP-VP131 inhibited EcSTING- or EcTBK1-induced antiviral activity upon red-spotted grouper nervous necrosis virus (RGNNV) infection. Together, our results demonstrated that the SGIV VP131 negatively regulated the IFN response by inhibiting EcSTING-EcTBK1 signaling for viral evasion. IMPORTANCE STING has been identified as a critical factor participating in the innate immune response which recruits and phosphorylates TBK1 and IFN regulatory factor 3 (IRF3) to induce IFN production and defend against viral infection. However, viruses also distort the STING-TBK1 pathway to negatively regulate the IFN response and facilitate viral replication. Here, we reported that SGIV VP131 interacted with EcSTING within the ER and degraded EcSTING, leading to the suppression of IFN production and the promotion of SGIV infection. These results for the first time demonstrated that fish iridovirus evaded the host antiviral response via abrogating the STING-TBK1 signaling pathway.
Assuntos
Bass , Infecções por Vírus de DNA , Doenças dos Peixes , Iridovirus , Ranavirus , Animais , Antivirais , Bass/genética , Bass/metabolismo , Infecções por Vírus de DNA/veterinária , Infecções por Vírus de DNA/genética , Proteínas de Peixes , Imunidade Inata/genética , Fator Regulador 3 de Interferon/metabolismo , Interferons/metabolismo , Iridovirus/genética , Iridovirus/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Ranavirus/genética , RNA Mensageiro/genética , Singapura , Ubiquitinas/metabolismoRESUMO
Persistent virus infections are achieved when the intricate balance of virus replication, host-cell division and successful immune evasion is met. The genomes of persistent DNA viruses are either maintained as extrachromosomal episomes or can integrate into the host genome. Common to both these strategies of persistence is the chromatinisation of viral DNA by cellular histones which, like host DNA, are subject to epigenetic modification. Epigenetic repression of viral genes required for lytic replication occurs, while genes required for latent or persistent infection are maintained in an active chromatin state. Viruses utilise host-cell chromatin insulators, which function to maintain epigenetic boundaries and enforce this strict transcriptional programme. Here, we review insulator protein function in virus transcription control, focussing on CCCTC-binding factor (CTCF) and cofactors. We describe CTCF-dependent activities in virus transcription regulation through epigenetic and promoter-enhancer insulation, three-dimensional chromatin looping and manipulation of transcript splicing.
Assuntos
Cromatina , Infecções por Vírus de DNA , Infecções por Vírus de DNA/genética , DNA Viral/genética , Epigênese Genética , Humanos , Latência Viral/genética , Replicação ViralRESUMO
Approximately 170 RNA modifications have been identified and these are critical for determining the fate and function of cellular RNAs. Similar to human transcripts, viral RNAs possess an extensive RNA modification landscape. While initial efforts largely focused on investigating the RNA modification landscape in the context of RNA virus infection, a growing body of work has explored the impact of RNA modifications on DNA virus biology. These studies have revealed roles for RNA modifications in DNA virus infection, including gene regulation and viral pathogenesis. In this review, we will discuss the current knowledge on how RNA modifications impact DNA virus biology.
Assuntos
Infecções por Vírus de DNA , Replicação Viral , Biologia , Infecções por Vírus de DNA/genética , Regulação da Expressão Gênica , Humanos , RNA Viral/genéticaRESUMO
The DNA sensor cGAS detects cytosolic DNA and instigates type I interferon (IFN) expression. Recent studies find that cGAS also localizes in the nucleus and binds the chromatin. Despite the mechanism controlling nuclear cGAS activation is well elucidated, whether nuclear cGAS participates in DNA sensing is unclear. Here, we report that herpes simplex virus 1 (HSV-1) infection caused the release of cGAS from the chromatin into the nuclear soluble fraction. Like its cytosolic counterpart, the leaked nuclear soluble cGAS also sensed viral DNA, produced cGAMP, and induced mRNA expression of type I IFN and interferon-stimulated genes. Consistently, the nuclear soluble cGAS limited HSV-1 infection. Furthermore, enzyme-deficient mutation (D307A) or cGAS inhibitor RU.251 abolished nuclear cGAS-mediated innate immune responses, suggesting that enzymatic activity is also required for nuclear soluble cGAS. Taken all together, our study demonstrates that nuclear soluble cGAS acts as a nuclear DNA sensor detecting nuclear-replicating DNA viruses.
Assuntos
Infecções por Vírus de DNA , Vírus de DNA , Nucleotidiltransferases , Cromatina , DNA/genética , DNA/metabolismo , Infecções por Vírus de DNA/genética , Infecções por Vírus de DNA/metabolismo , Infecções por Vírus de DNA/virologia , Vírus de DNA/genética , Vírus de DNA/metabolismo , Herpes Simples/genética , Humanos , Nucleotidiltransferases/genética , Nucleotidiltransferases/metabolismoRESUMO
Type I interferons (IFNs) are the first frontline of the host innate immune response against invading pathogens. Herein, we characterized an unknown protein encoded by phospholipase A2 inhibitor and LY6/PLAUR domain-containing (PINLYP) gene that interacted with TBK1 and induced type I IFN in a TBK1- and IRF3-dependent manner. Loss of PINLYP impaired the activation of IRF3 and production of IFN-ß induced by DNA virus, RNA virus, and various Toll-like receptor ligands in multiple cell types. Because PINLYP deficiency in mice engendered an early embryonic lethality in mice, we generated a conditional mouse in which PINLYP was depleted in dendritic cells. Mice lacking PINLYP in dendritic cells were defective in type I IFN induction and more susceptible to lethal virus infection. Thus, PINLYP is a positive regulator of type I IFN innate immunity and important for effective host defense against viral infection.
Assuntos
Células Dendríticas/imunologia , Inibidores Enzimáticos/imunologia , Imunidade Inata , Interferon beta/imunologia , Animais , Linhagem Celular , Infecções por Vírus de DNA/genética , Infecções por Vírus de DNA/imunologia , Vírus de DNA/genética , Vírus de DNA/imunologia , Humanos , Interferon beta/genética , Camundongos , Camundongos Knockout , Infecções por Vírus de RNA/genética , Infecções por Vírus de RNA/imunologia , Vírus de RNA/genética , Vírus de RNA/imunologiaRESUMO
Cell survival is based on the stability of intracellular state. It was well known that biochemical reactions in cells require specific intracellular environments, such as pH and calcium concentration. While the mechanism of stabilizing the intracellular environment is complex and far from clear. In this study, a Sma and Mad related protein 5 gene (LvSmad5) of Litopenaeus vannamei was cloned. LvSmad5 was located to both cytoplasm and nucleus. And subcellular localization of LvSmad5 was responsed to the changing of cells internal and external environment. Besides, it was found that subcellular localization of LvSmad5 was also regulated by unfolded protein response. Moreover, it was proved that nucleic localization of LvSmad5 could significantly increase the white spot syndrome virus (WSSV) infection in shrimp, and knockdown expression of LvSmad5 decreased the cumulative mortality of WSSV infection shrimp. Further investigation revealed that cytoplasm LvSmad5 could interplay with shrimp hexokinase 1, and contribute to glycolysis. These results indicated that LvSmad5 played a role in L. vannamei environmental stress response, and was used by WSSV for its replication.
Assuntos
Infecções por Vírus de DNA/genética , Doenças dos Peixes/genética , Proteínas de Peixes/genética , Penaeidae/genética , Proteína Smad5/genética , Estresse Fisiológico/genética , Vírus da Síndrome da Mancha Branca 1/fisiologia , Sequência de Aminoácidos , Animais , Núcleo Celular , Clonagem Molecular , Citoplasma , Infecções por Vírus de DNA/mortalidade , Infecções por Vírus de DNA/veterinária , Infecções por Vírus de DNA/virologia , Doenças dos Peixes/mortalidade , Doenças dos Peixes/virologia , Penaeidae/virologia , Resposta a Proteínas não Dobradas/genética , Replicação ViralRESUMO
Background: Frog Virus 3 (FV3) is a large dsDNA virus belonging to Ranaviruses of family Iridoviridae. Ranaviruses infect cold-blood vertebrates including amphibians, fish and reptiles, and contribute to catastrophic amphibian declines. FV3 has a genome at ~105 kb that contains nearly 100 coding genes and 50 intergenic regions as annotated in its reference genome. Previous studies have mainly focused on coding genes and rarely addressed potential non-coding regulatory role of intergenic regions. Results: Using a whole transcriptomic analysis of total RNA samples containing both the viral and cellular transcripts from FV3-infected frog tissues, we detected virus-specific reads mapping in non-coding intergenic regions, in addition to reads from coding genes. Further analyses identified multiple cis-regulatory elements (CREs) in intergenic regions neighboring highly transcribed coding genes. These CREs include not only a virus TATA-Box present in FV3 core promoters as in eukaryotic genes, but also viral mimics of CREs interacting with several transcription factors including CEBPs, CREBs, IRFs, NF-κB, and STATs, which are critical for regulation of cellular immunity and cytokine responses. Our study suggests that intergenic regions immediately upstream of highly expressed FV3 genes have evolved to bind IRFs, NF-κB, and STATs more efficiently. Moreover, we found an enrichment of putative microRNA (miRNA) sequences in more than five intergenic regions of the FV3 genome. Our sequence analysis indicates that a fraction of these viral miRNAs is targeting the 3'-UTR regions of Xenopus genes involved in interferon (IFN)-dependent responses, including particularly those encoding IFN receptor subunits and IFN-regulatory factors (IRFs). Conclusions: Using the FV3 model, this study provides a first genome-wide analysis of non-coding regulatory mechanisms adopted by ranaviruses to epigenetically regulate both viral and host gene expressions, which have co-evolved to interact especially with the host IFN response.
Assuntos
Infecções por Vírus de DNA/veterinária , DNA Intergênico/genética , Regulação Viral da Expressão Gênica , Interações Hospedeiro-Patógeno/genética , MicroRNAs/genética , RNA Viral/biossíntese , Ranavirus/genética , Xenopus laevis/virologia , Regiões 3' não Traduzidas , Animais , Infecções por Vírus de DNA/genética , Genoma Viral , Fatores Reguladores de Interferon/biossíntese , Fatores Reguladores de Interferon/genética , Interferência de RNA , RNA Viral/genética , Distribuição Aleatória , Receptores de Interferon/biossíntese , Receptores de Interferon/genética , Organismos Livres de Patógenos Específicos , Transcriptoma , Xenopus laevis/genética , Xenopus laevis/metabolismoRESUMO
The genus Megalocytivirus includes viruses known to cause significant disease in aquacultured fish stocks. Herein, we report the complete genome sequences of two megalocytiviruses (MCVs) isolated from diseased albino rainbow sharks Epalzeorhynchos frenatum reared on farms in the United States in 2018 and 2019. Histopathological examination revealed typical megalocytivirus microscopic lesions (i.e., basophilic cytoplasmic inclusions) that were most commonly observed in the spleen and kidney. Transmission electron microscopic examination of spleen and kidney tissues from specimens of the 2018 case revealed hexagonally shaped virus particles with a mean diameter of 153 ± 6 nm (n = 20) from opposite vertices and 131 ± 5 nm (n = 20) from opposite faces. Two MCV-specific conventional PCR assays confirmed the presence of MCV DNA in the collected samples. Full genome sequencing of both 2018 and 2019 Epalzeorhynchos frenatus iridoviruses (EFIV) was accomplished using a next-generation sequencing approach. Phylogenomic analyses revealed that both EFIV isolates belong to the infectious spleen and kidney necrosis virus (ISKNV) genotype within the genus Megalocytivirus. This study is the first report of ISKNV in albino rainbow sharks.
Assuntos
Infecções por Vírus de DNA/genética , Genoma Viral/genética , Iridoviridae/genética , Tubarões/virologia , Animais , Infecções por Vírus de DNA/virologia , Fazendas , Doenças dos Peixes/genética , Doenças dos Peixes/virologia , Peixes/genética , Peixes/virologia , Humanos , Filogenia , Tubarões/genética , Estados Unidos , Sequenciamento Completo do GenomaRESUMO
Multiple novel circular replication-associated protein (Rep)-encoding single stranded (CRESS) DNA viruses have been extensively identified in the feces of humans and animals. Here, we first detected CRESS DNA virus (named Horse-CRESS DNA-like virus, HCLV) in two fecal samples from 10 imported thoroughbred (TB) horses in the customs quarantine station in North Xinjiang province, China. Additionally, we found that this virus was not detected in local breeds (LBs) (0/41) and was found only in imported TB horses (2/73). We obtained the whole-genome sequences of four viruses (HCLV ALSK-3-4, ALSK-13-10, CJ-1-2, and CJ-13-1). Unlike Circovirus and Cyclovirus, whose genome sequences have 1700 to 2100 nucleotides (nt), these HCLVs have circular genome with 3503, 3504, 3485, 3491 nt, respectively and five major ORFs. The ORF1 gene encodes the Rep protein in HCLVs. Furthermore, the Rep protein of the four HCLVs share 23.3-84.8%, 21.6-27.4%, 23.7-27.2% amino acid identity with the corresponding reference viruses of Kirkoviruses, genus Circovirus, and genus Cyclovirus, respectively. Moreover, RCR domain, P-loop NTPase domains, and nonanucleotide motif (TAGTATTAC) of the HCLVs are similar to Circovirus and Cyclovirus. Phylogenetic analysis showed that the virus was grouped together with members in Kirkoviruses. These results suggest the HCLV probably entered Xinjiang province via the international trade of horses.
Assuntos
Infecções por Vírus de DNA/genética , Vírus de DNA/genética , Genoma Viral/genética , Genômica , Animais , China/epidemiologia , Infecções por Vírus de DNA/veterinária , Infecções por Vírus de DNA/virologia , Doenças dos Cavalos/genética , Doenças dos Cavalos/virologia , Cavalos/genética , Cavalos/virologia , Sequenciamento Completo do GenomaRESUMO
OBJECTIVES: An association between costimulatory molecule gene polymorphisms and viral infection after hematopoietic stem cell transplantation may be related to clinical outcomes, especially acute graft-versus-host disease. Cytotoxic T-lymphocyte antigen 4 has been suggested as a crucial negative regulator of the immune system. In this study, our objective was to investigate the association between cytotoxic T-lymphocyte antigen-4 gene polymorphisms (including -1722 T/C, -1661 A/G, -318 C/T, and +49 A/G) and torque teno virus infection after hematopoietic stem cell transplantation in patients with and without acute graft-versus-host disease. MATERIALS AND METHODS: Our study included 71 recipients. We evaluated cytotoxic T-lymphocyte antigen 4 gene polymorphisms using the polymerase chain reaction-restriction fragment length polymorphism method. RESULTS: Our results showed that the GG genotype of the cytotoxic T-lymphocyte antigen 4 +49 A/G was significantly more frequent in transplanted patients infected with torque teno virus, whereas the AG genotype was more common in transplanted patients who did not have this infection. In addition, the -1661 AA and GA genotypes and -318 TC genotypes were significantly more frequent in transplanted patients infected with the virus and who had low-grade (grades I and II) acute graft-versus-host disease. Among those with grade I graft-versus-host disease, the GG genotype of the cytotoxic T-lymphocyte antigen 4+49 A/G was more frequent in transplanted patients with torque teno virus infection, whereas the AG genotype was higher in transplanted patients who did not have this infection. CONCLUSIONS: This is the first report indicating that cytotoxic T-lymphocyte antigen 4 gene polymorphism may be implicated in prevalence of torque teno virus infection after stem cell transplant. Further larger studies and evaluation of other costimulatory molecules are suggested.
Assuntos
Antígeno CTLA-4/genética , Infecções por Vírus de DNA/genética , Doença Enxerto-Hospedeiro , Transplante de Células-Tronco Hematopoéticas , Torque teno virus , Doença Enxerto-Hospedeiro/diagnóstico , Doença Enxerto-Hospedeiro/genética , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Humanos , Polimorfismo GenéticoRESUMO
Torque teno virus (TTV) is a commensal human virus observed as a circular single-negative-strand DNA molecule in various tissues and biological samples, notably in blood serum and lymphocytes. TTV has no apparent clinical significance, although it might be very useful as a prospective tool for gene delivery or as an epidemiological marker. Human populations are ubiquitously infected with TTV; the prevalence may reach 100%. The majority of babies become spontaneously infected with TTV, so that by the end of the first year of life, the prevalence reaches 'adult' values. TTV positivity in healthy early infancy and the presence of TTV in umbilical cord blood samples have been reported. The mechanism of infection and the dynamics of TTV prevalence in infants with age remain understudied. Meanwhile, the potential diagnostic and prognostic value of TTV as a marker deserves special attention and study, along with the possibility, causes and consequences of placental transmission of TTV under normal or pathological conditions.
Assuntos
Infecções por Vírus de DNA/virologia , Torque teno virus/fisiologia , Fatores Etários , Infecções por Vírus de DNA/sangue , Infecções por Vírus de DNA/genética , DNA Viral/genética , Feminino , Humanos , Lactente , Recém-Nascido , Gravidez , Prevalência , Reação em Cadeia da Polimerase em Tempo Real , Carga ViralRESUMO
Decapod iridescent virus 1 (DIV1) results in severe economic losses in shrimp aquaculture. However, little is known about the physiological effect of DIV1 infection on the host. In this study, we found that the lethal dose 50 of DIV1-infected Litopenaeus vannamei after 48, 72, 96, and 156 h were 4.86 × 106, 5.07 × 105, 2.13 × 105, and 2.38 × 104 copies/µg DNA, respectively. In order to investigate the mechanisms of DIV1 infection, a comparative transcriptome analysis of hemocytes from L. vannamei, infected or not with DIV1, was conducted. The BUSCO analysis showed that the transcriptome was with high completeness (complete single-copy BUSCOs: 57.3%, complete duplicated BUSCOs: 41.1%, fragmentation: 0.8%, missing: 0.8%). A total of 168,854 unigenes were assembled, with an average length of 601 bp. Based on homology searches, Kyoto Encyclopedia of Genes and Genomes (KEGG), gene ontology (GO), and cluster of orthologous groups of proteins (KOG) analysis, 62,270 (36.88%) unigenes were annotated. Among them, 1,112 differentially expressed genes (DEGs) were identified, of which 889 genes were up-regulated and 223 genes were down-regulated after DIV1 infection. These genes were mainly annotated to the major metabolic processes such as fructose and mannose metabolism, carbon metabolism, and inositol phosphate metabolism. Among these metabolic pathways, the triosephosphate isomerase (TPI) family was the most eye-catching DEG as it participates in several metabolic processes. Three types of TPI, LvTPI-like, LvTPI-Blike, and LvTPI-Blike1, were obtained for gene silencing by RNA interference. The results showed that LvTPI-like and LvTPI-Blike1 silencing caused a high mortality rate among L. vannamei. However, LvTPI-like and LvTPI-Blike silencing reduced DIV1 replication in DIV1-infected L. vannamei. All the results indicated that TPI-like genes play an important role during DIV1 infection, which provides valuable insight into the infection mechanism of DIV1 in shrimp and may aid in preventing viral diseases in shrimp culture.