RESUMO
Usurpation of the host's signalling pathways is a common strategy employed by viruses to promote their successful replication. Here we show that infection with the orthopoxvirus vaccinia virus (VACV) leads to sustained stimulation of c-Jun activity during the entire infective cycle. This stimulation is temporally regulated through MEK/ERK or MKK/JNK pathways, i.e. during the early/mid phase (1 to 6 hpi) and in the late phase (9 to 24 hpi) of the infective cycle, respectively. As a transcriptional regulator, upon infection with VACV, c-Jun is translocated from the cytoplasm to the nucleus, where it binds to the AP-1 DNA sequence found at the promoter region of its target genes. To investigate the role played by c-Jun during VACV replication cycle, we generated cell lines that stably express a c-Jun-dominant negative (DNc-Jun) mutation. Our data revealed that c-Jun is required during early infection to assist with viral DNA replication, as demonstrated by the decreased amount of viral DNA found in the DNc-Jun cells. We also demonstrated that c-Jun regulates the expression of the early growth response gene (egr-1), a gene previously shown to affect VACV replication mediated by MEK/ERK signalling. VACV-induced stimulation of the MKK/JNK/JUN pathway impacts viral dissemination, as we observed a significant reduction in both viral yield, during late stages of infection, and virus plaque size. Collectively, our data suggest that, by modulating the host's signalling pathways through a common target such as c-Jun, VACV temporally regulates its infective cycle in order to successfully replicate and subsequently spread.
Assuntos
MAP Quinases Reguladas por Sinal Extracelular/metabolismo , MAP Quinase Quinase 4/metabolismo , MAP Quinase Quinase Quinases/metabolismo , Quinases de Proteína Quinase Ativadas por Mitógeno/metabolismo , Proteínas Proto-Oncogênicas c-jun/metabolismo , Vaccinia virus/fisiologia , Animais , Linhagem Celular , DNA Viral , Proteína 1 de Resposta de Crescimento Precoce/genética , Proteína 1 de Resposta de Crescimento Precoce/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/genética , Fibroblastos/virologia , Regulação Enzimológica da Expressão Gênica/fisiologia , Regulação Viral da Expressão Gênica/fisiologia , MAP Quinase Quinase 4/genética , MAP Quinase Quinase Quinases/genética , Camundongos , Quinases de Proteína Quinase Ativadas por Mitógeno/genética , Fosforilação , Proteínas Proto-Oncogênicas c-jun/genética , Replicação ViralRESUMO
BACKGROUND: The identification of novel giant viruses from the nucleocytoplasmic large DNA viruses group and their virophages has increased in the last decade and has helped to shed light on viral evolution. This study describe the discovery, isolation and characterization of Samba virus (SMBV), a novel giant virus belonging to the Mimivirus genus, which was isolated from the Negro River in the Brazilian Amazon. We also report the isolation of an SMBV-associated virophage named Rio Negro (RNV), which is the first Mimivirus virophage to be isolated in the Americas. METHODS/RESULTS: Based on a phylogenetic analysis, SMBV belongs to group A of the putative Megavirales order, possibly a new virus related to Acanthamoeba polyphaga mimivirus (APMV). SMBV is the largest virus isolated in Brazil, with an average particle diameter about 574 nm. The SMBV genome contains 938 ORFs, of which nine are ORFans. The 1,213.6 kb SMBV genome is one of the largest genome of any group A Mimivirus described to date. Electron microscopy showed RNV particle accumulation near SMBV and APMV factories resulting in the production of defective SMBV and APMV particles and decreasing the infectivity of these two viruses by several logs. CONCLUSION: This discovery expands our knowledge of Mimiviridae evolution and ecology.
Assuntos
Mimiviridae/isolamento & purificação , Filogenia , Rios/virologia , Brasil , DNA Viral/química , DNA Viral/genética , Microscopia Eletrônica de Transmissão , Mimiviridae/classificação , Mimiviridae/genética , Mimiviridae/ultraestrutura , Dados de Sequência Molecular , Fases de Leitura Aberta , Floresta Úmida , Análise de Sequência de DNA , Vírion/ultraestruturaRESUMO
To investigate circulation of mimiviruses in the Amazon Region of Brazil, we surveyed 513 serum samples from domestic and wild mammals. Neutralizing antibodies were detected in 15 sample pools, and mimivirus DNA was detected in 9 pools of serum from capuchin monkeys and in 16 pools of serum from cattle.
Assuntos
Doenças dos Animais/epidemiologia , Doenças dos Animais/virologia , Viroses/veterinária , Sequência de Aminoácidos , Animais , Animais Domésticos , Animais Selvagens , Brasil/epidemiologia , DNA Viral , Geografia , Mamíferos , Mimiviridae , Dados de Sequência Molecular , Filogenia , Alinhamento de Sequência , Carga ViralRESUMO
To detect orthopoxvirus in the Brazilian Amazon, we conducted a serosurvey of 344 wild animals. Neutralizing antibodies against orthopoxvirus were detected by plaque-reduction neutralizing tests in 84 serum samples. Amplicons from 6 monkey samples were sequenced. These amplicons identified vaccinia virus genetically similar to strains from bovine vaccinia outbreaks in Brazil.
Assuntos
Doenças dos Macacos/epidemiologia , Vaccinia virus/isolamento & purificação , Vacínia/veterinária , Alouatta , Animais , Tatus , Brasil/epidemiologia , Cebus , Chlorocebus aethiops , DNA Viral/análise , DNA Viral/genética , Raposas , Hemaglutininas Virais/análise , Hemaglutininas Virais/genética , Humanos , Peptídeos e Proteínas de Sinalização Intercelular , Dados de Sequência Molecular , Doenças dos Macacos/imunologia , Doenças dos Macacos/virologia , Testes de Neutralização , Gambás , Peptídeos/análise , Peptídeos/genética , Filogenia , Prevalência , Procyonidae , Roedores , Análise de Sequência de DNA , Vacínia/epidemiologia , Vacínia/imunologia , Vacínia/virologia , Vaccinia virus/genética , Vaccinia virus/imunologia , Células VeroRESUMO
The SPAn232 virus (SPAnv) was isolated from sentinel mice in the forest of Cotia, São Paulo, Brazil. It was grouped originally as a Cotia virus (CV) sample due to serological cross-reaction with the latter. However, SPAnv presented genetic characteristics that differed from CV and indicated that SPAnv is a member of the vaccinia virus (VV) subgroup. SPAnv showed a HindIII-digested DNA pattern similar to those of the WR and Lister strains of VV. Also, SPAnv presented genes homologous to the vaccinia growth factor, thymidine kinase and A-type inclusion (ATI) genes from VV. RFLP analysis of the SPAnv ATI homologous gene indicated that the virus belongs to the VV group. Nucleotide sequences from SPAnv genes showed up to 99% similarity with the same genes from VV. Such a relationship was confirmed visually through the drawing of phylogenetic trees. The results point out the occurrence of a VV strain that is possibly in active circulation in the forests of Southeast Brazil.