RESUMO
Orf virus (ORFV) infects sheep and goat tissues, resulting in severe proliferative lesions. To analyze cellular protein expression in ORFV-infected goat skin fibroblast (GSF) cells, we used two-dimensional liquid chromatography-tandem mass spectrometry coupled with isobaric tags for relative and absolute quantification (iTRAQ). The proteomics approach was used along with quantitative reverse transcription polymerase chain reaction (RT-qPCR) to detect differentially expressed proteins in ORFV-infected GSF cells and mock-infected GSF cells. A total of 282 differentially expressed proteins were identified. It was found that 222 host proteins were upregulated and 60 were downregulated following viral infection. We confirmed that these proteins were differentially expressed and found that heat shock 70-kDa protein 1B (HSPA1B) was differentially expressed and localized in the cytoplasm. It was also noted that HSPA1B caused inhibition of viral proliferation, in the middle and late stages of viral infection. The differentially expressed proteins were associated with the biological processes of viral binding, cell structure, signal transduction, cell adhesion, and cell proliferation.
Assuntos
Fibroblastos/metabolismo , Proteínas de Choque Térmico HSP70/fisiologia , Vírus do Orf/fisiologia , Proteoma/genética , Replicação Viral , Animais , Células Cultivadas , Cromatografia Líquida , Fibroblastos/virologia , Cabras , Interações Hospedeiro-Patógeno , Vírus do Orf/genética , Proteômica , Espectrometria de Massas em TandemRESUMO
As a zoonotic disease, ovine contagious pustular dermatitis (Orf) is a serious threat to sheep as well as humans. Orf virus (ORFV) interferon resistance protein (VIR) is the principal virulence protein that encodes a dsRNA-binding protein to inhibit host antiviral response. p53 is one of the key proteins of the host antiviral innate immunity. It not only enhances type I interferon secretion but also induces apoptosis in infected cells, and plays a crucial role in the immune response against various viral infections. However, it remains to be elucidated what role p53 plays in ORFV replication and whether ORFV's own protein VIR regulates p53 expression to promote self-replication. In this study, we showed that p53 has an antiviral effect on ORFV and can inhibit ORFV replication. In addition, ORFV nonstructural protein VIR interacts with p53 and degrades p53, which inhibits p53-mediated positive regulation of downstream antiviral genes. This study provides new insight into the immune evasion mediated by ORFV and identifies VIR as an antagonistic factor for ORFV to evade the antiviral response.
Assuntos
Interações entre Hospedeiro e Microrganismos/genética , Vírus do Orf/genética , Proteína Supressora de Tumor p53/genética , Proteínas Virais/genética , Replicação Viral/genética , Animais , Linhagem Celular , Cricetinae , Ectima Contagioso/virologia , Fibroblastos/imunologia , Fibroblastos/virologia , Regulação Viral da Expressão Gênica , Cabras , Evasão da Resposta Imune/genética , Imunidade Inata , Rim/citologia , Vírus do Orf/fisiologia , Ovinos , Pele/citologia , Proteínas Virais/metabolismoRESUMO
Tumor progression locus 2 (TPL2) is a serine/threonine kinase that belongs to the MAP3K family. The activated TPL2 regulates the innate immune-relevant signaling pathways, such as ERK, JNK, and NF-κB, and the differentiation of immune cells, for example, CD4+ T and NK cells. Therefore, TPL2 plays a critical role in regulating the innate immune response. The present review summarizes the recent advancements in the TPL2-regulated innate immune response.
Assuntos
Linfócitos T CD4-Positivos/imunologia , MAP Quinase Quinase Quinases/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Animais , Diferenciação Celular , Quimiocinas/metabolismo , Humanos , Imunidade Inata , MAP Quinase Quinase Quinases/genética , Sistema de Sinalização das MAP Quinases , Ativação de Macrófagos , Ativação de Neutrófilo , Proteínas Proto-Oncogênicas/genéticaRESUMO
Getah virus (GETV), a mosquito-borne virus that mainly infects horses and pigs, has emerged and spread in China. We developed a highly specific and reproducible TaqMan probe-based quantitative reverse transcription PCR (RT-qPCR) assay targeting the non-structural protein 1 of GETV, whose detection limit is 25.5 copies/µL, which is 100-fold higher than that of conventional RT-PCR. RT-qPCR was used to detect GETV RNA in mosquito and animal clinical samples, showing that the accuracy of RT-qPCR was higher than that of conventional RT-PCR. The newly developed RT-qPCR assay may be a useful alternative tool for rapid, simple and specific diagnosis of GETV infection.
Assuntos
Alphavirus/genética , Culex/virologia , Sondas de DNA/genética , RNA Viral/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Proteínas não Estruturais Virais/genética , Alphavirus/isolamento & purificação , Animais , Sequência de Bases , China , Cavalos , Sus scrofaRESUMO
Purpose To investigate the autophagy characteristics of giant cell tumor of bone and its effect on cell proliferation. Methods Giant cell tumors of bone (GCT-0404 cells) were cultured in vitro and treated with serum-starvation, rapamycin and inhibitor chloroquine. Western blot analysis was performed to detect the autophagic markers LC3 and Beclin 1 expression. Immunofluorescence method was used to show intracellular autophagy formation. Inverted microscope was used to observe the cell morphology. CCK-8 assays were used to detect the cell viability. Results Green fluorescent spots that represented the transformation level of LC3-I to LC3-Ⅱ increased significantly (P＜0.05) by serum-starvation, rapamycin and inhibitor chloroquine respectively, but the expression level of Beclin 1 was not raised. In addition, the cell morphology changed significantly, and cell proliferation was inhibited after serum-starvation, and treatment with rapamycin and inhibitor chloroquine (P＜0.05), Conclusion The changes of autophagy caused by serum starvation, rapamycin and inhibitor chloroquine in GCT-0404 cells may be involved in the down-regulation of cell proliferation.
RESUMO
An outbreak of severe pseudorabies virus (PRV) infection in farmed mink occurred in northern China in late 2014, causing significant economic losses in the local fur industry. Here, we report the first case of a PRV outbreak in mink in northeastern China, caused by feeding farmed mink with raw pork or organs contaminated by PRV. Mink infected with virulent PRV exhibited diarrhea, neurologic signs, and higher mortality, which can be misdiagnosed as highly pathogenic mink enteritis virus (MEV), canine distemper virus (CDV), and food poisoning. However, these were excluded as causative agents by PCR or bacteria isolation. The duration of disease was 3-7 days, and the mortality rate was 80-90%. PRV was characterized using indirect immunofluorescence assays (IFA) and electron microscopy (EM). Phylogenetic analysis based on full-length genome sequences and those of individual genes of this novel virus strain showed that it clustered in an independent branch with several other PRV isolates from China.
