RESUMO
The sequence of the alpha-transinducing factor (alpha-TIF) of canine herpesvirus (CHV-l) was determined. Alignment of the predicted CHV-1 alpha-TIF amino acid sequence with other alpha-TIF homologues reveals a core region of similarity with divergent amino and carboxyl termini. Analysis of the CHV-1 infected cell protein 4 promoter region identified a region containing nine copies of a 52 bp repeat that showed significant up-regulation of transcription by alpha-TIF. This region contained an imperfect 'TAATGARAT' motif, the binding site for herpes simplex virus 1 alpha-TIF, with an imperfect Oct-1 binding site immediately following. The infectious laryngotracheitis virus alpha-TIF was also shown to up-regulate transcription through this region of the promoter. Transfection of CHV-1 genomic DNA failed to yield infectious virus in canine kidney cell lines. Co-transfection of genomic DNA and an alpha-TIF expression plasmid resulted in virus plaques, indicating a potential essential role for alpha-TIF in CHV-1 infection.
Assuntos
Genes Precoces/fisiologia , Herpesvirus Canídeo 1/química , Regiões Promotoras Genéticas/fisiologia , Fatores de Transcrição/fisiologia , Animais , Cães , Regulação Viral da Expressão Gênica , Genes Virais , Dados de Sequência MolecularRESUMO
We report the complete nucleotide (nt) sequence of nine genes of an Australian isolate of canine herpesvirus (CHV). Four of them are located in the unique short (US) region: glycoprotein (g) genes gG, gD and gI, and the protein kinase gene. Five are in the unique long (UL) region: the thymidine kinase gene, gB, gC, gH, and gene homologue UL24. Partial sequence was determined for four genes, two in the UL region (UL21 and virion protein) and two in the US region (US2 and gE). A repeat sequence of 382 nt with unknown function was identified in the 615 nt intergenic region between gH and UL21. A total of 16.93 kb was sequenced and compared with sequences from CHV isolates from the USA, France, Japan and Australia. Only minor nt and/or amino acid (aa) differences were observed.
Assuntos
Genes Virais , Herpesvirus Canídeo 1/genética , Proteínas Quinases/genética , Timidina Quinase/genética , Proteínas do Envelope Viral/genética , Animais , Austrália , Sequência de Bases , Doenças do Cão/virologia , Cães , Glicoproteínas/química , Glicoproteínas/genética , Infecções por Herpesviridae/virologia , Herpesvirus Canídeo 1/química , Dados de Sequência Molecular , Proteínas Quinases/química , Análise de Sequência de DNA , Homologia de Sequência , Timidina Quinase/química , Proteínas do Envelope Viral/química , Proteínas Virais/química , Proteínas Virais/genéticaRESUMO
The canine herpesvirus infected cell protein 0 (CICP0) gene was sequenced. The CICP0 gene was transcribed as a 1.4 kb mRNA from the end of the unique long region nearby the internal repeat during early phase of productive infection of the virus. An open reading frame of the gene encodes a polypeptide of 333 amino acids. The RING finger domain and acidic transcriptional activation domain were found at the N-terminus and within the middle region in the deduced amino acid sequence, respectively, suggesting that the CICP0, like the ICP0 of herpes simplex virus 1, is a transactivating protein.
Assuntos
DNA Viral/genética , Herpesvirus Canídeo 1/genética , Proteínas Imediatamente Precoces/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Northern Blotting , Linhagem Celular , Sequência Consenso , Primers do DNA , Cães , Herpesvirus Canídeo 1/química , Dados de Sequência Molecular , Fases de Leitura Aberta , RNA Mensageiro/análise , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Transativadores/genética , Ubiquitina-Proteína LigasesRESUMO
A recombinant canine herpesvirus (CHV) which expressed glycoprotein B (gB) of pseudorabies virus (PrV) was constructed. The antigenicity of the PrV gB expressed by the recombinant CHV is similar to that of the native PrV. The expressed PrV gB was shown to be transported to the surface of infected cells as judged by an indirected immunofluorescence test. Antibodies raised in mice immunized with the recombinant CHV neutralized the infectivity of PrV in vitro. It is known that the authentic PrV gB exists as a glycoprotein complex, which consists of gBa, gBb and gBc. In MDCK cells, PrV gB expressed by the recombinant CHV was processed like authentic PrV gB, suggesting that the cleavage mechanism of PrV gB depends on a functional cleavage domain from PrV gB gene and protease from infected cells.
Assuntos
Herpesvirus Canídeo 1/química , Herpesvirus Suídeo 1/química , Pseudorraiva/imunologia , Proteínas do Envelope Viral/química , Animais , Anticorpos Monoclonais , Anticorpos Antivirais/imunologia , Antígenos Virais/análise , Western Blotting/veterinária , Técnica Indireta de Fluorescência para Anticorpo/veterinária , Herpesvirus Canídeo 1/crescimento & desenvolvimento , Herpesvirus Suídeo 1/imunologia , Camundongos , Testes de Neutralização/veterinária , Proteínas Recombinantes , Transfecção , Proteínas do Envelope Viral/imunologiaRESUMO
The nucleotide sequence of the immediate early (IE) gene of canine herpesvirus was determined. This gene was located in the inverted repeat regions, encoding a polypeptide of 1,383 amino acids. The predicted amino acid sequence was most closely related to that of the feline herpesvirus 1 IE protein among those of other alphaherpesviruses. DNA binding and transcriptional activation domains were found in the IE protein. A spliced region of the IE gene transcript was determined in its 5' non-coding region.
Assuntos
Genes Precoces/genética , Herpesvirus Canídeo 1/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Linhagem Celular , DNA Viral/química , DNA Viral/genética , Cães , Genoma Viral , Herpesvirus Canídeo 1/química , Proteínas Imediatamente Precoces/genética , Dados de Sequência Molecular , RNA Mensageiro/genética , Análise de Sequência de DNARESUMO
The genes encoding the canine herpesvirus (CHV) glycoprotein B (gB), gC and gD homologues have been reported already. However, products of these genes have not been identified yet. Previously, we have identified three CHV glycoproteins, gp 145/112, gp80 and gp47 using a panel of monoclonal antibodies (MAbs). To determine which CHV glycoprotein corresponds to gB, gC or gD, the putative genes of gB, gC, and gD of CHV were inserted into the thymidine kinase gene of vaccinia virus LC16mO strain under the control of the early-late promoter for the vaccinia virus 7.5-kilodalton polypeptide. We demonstrated here that gp145/112, gp80 and gp47 were the translation products of the CHV gB, gC and gD genes, respectively. The antigenic authenticity of recombinant gB, gC and gD were confirmed by a panel of MAbs specific for each glycoprotein produced in CHV-infected cells. Immunization of mice with these recombinants produced high titers of neutralizing antibodies against CHV. These results suggest that recombinant vaccinia viruses expressing CHV gB, gC and gD may be useful to develop a vaccine to control CHV infection.
Assuntos
Herpesvirus Canídeo 1/química , Vaccinia virus/genética , Proteínas do Envelope Viral/análise , Animais , Herpesvirus Canídeo 1/genética , Camundongos , Recombinação Genética , Proteínas do Envelope Viral/genéticaRESUMO
The YP11mu strain of a plaque-selected canine herpesvirus (CHV) encoded a smaller molecular weight (MW) of gD than those of other strains including YP2 strain (Xuan et al., 1990). When nucleotide sequence of the mutated gD of YP11mu strain (gD(YP11mu)) was compared with that of gDs of other CHV strains, gD(YP11mu) lacked 12 nucleotides encoding 4 amino acids, NKTI, including one predicted potential N-linked glycosylation site and no other change was found in other regions. When the gD(YP11mu) and gD of YP2 strain (gD(YP2)) expressed in COS-7 and insect (Spodoptera frugiperda; Sf9) cells were compared each other, both gDs reacted with a panel of monoclonal antibodies (MAbs) against CHV gD by indirect immunofluorescence analysis and the gD(YP11mu) possessed an MW of approximately 47-51 and 39-44 kDa in COS-7 and Sf9 cells, respectively, which were smaller than the expressed gD(YP2) (approximately 51-55 and 41-46 kDa, respectively) by immunoblot analysis. After treatment with tunicamycin, the MW of both gDs in Sf9 cells became approximately 37 kDa. When hemagglutination (HA) test using canine red blood cells (RBC) were carried out, lysates of Sf9 cells expressing CHV gDs agglutinated canine RBC. Serum from mice inoculated with lysates of Sf9 cells expressing the gDs possessed a high titer of virus-neutralizing (VN) activities against CHV. These results indicated that the deletion of 4 amino acids possessing approximately 4 kDa of glyco-chain from gD of CHV in mammalian cells does not affect HA activity and VN antibody-inducing activity and that this deletion of gD(YP11mu) might be a good selective marker for development of recombinant viruses as a live vaccine.
