RESUMO
Paramyxoviruses are negative-polarity RNA viruses of major clinical importance. The dynamic interaction of the RNA-dependent RNA polymerase (RdRP) complex with the encapsidated RNA genome is mechanistically and structurally poorly understood. Having generated recombinant measles (MeV) and canine distemper (CDV) viruses with truncated nucleocapsid (N) protein showing defects in replication kinetics, we have applied a viral evolution approach to the problem. Passaging of recombinants resulted in long-range compensatory mutations that restored RdRP bioactivity in minigenome assays and efficient replication of engineered viruses. Compensatory mutations clustered at an electronically compatible acidic loop in N-core and a basic face of the phosphoprotein X domain (P-XD). Co-affinity precipitations, biolayer interferometry, and molecular docking revealed an electrostatic-driven transiently forming interface between these domains. The compensatory mutations reduced electrostatic compatibility of these microdomains and lowered coprecipitation efficiency, consistent with a molecular checkpoint function that regulates paramyxovirus polymerase mobility through modulation of conformational stability of the P-XD assembly.
Assuntos
Vírus da Cinomose Canina/genética , Vírus do Sarampo/genética , Proteínas do Nucleocapsídeo/química , Fosfoproteínas/química , RNA Polimerase Dependente de RNA/química , Vírus Reordenados/genética , Replicação Viral/genética , Animais , Sítios de Ligação , Linhagem Celular , Chlorocebus aethiops , Clonagem Molecular , Cricetulus , Vírus da Cinomose Canina/metabolismo , Células Epiteliais/metabolismo , Células Epiteliais/virologia , Escherichia coli/genética , Escherichia coli/metabolismo , Expressão Gênica , Vírus do Sarampo/metabolismo , Simulação de Acoplamento Molecular , Mutação , Proteínas do Nucleocapsídeo/genética , Proteínas do Nucleocapsídeo/metabolismo , Fosfoproteínas/genética , Fosfoproteínas/metabolismo , Ligação Proteica , Conformação Proteica em alfa-Hélice , Domínios e Motivos de Interação entre Proteínas , RNA Polimerase Dependente de RNA/genética , RNA Polimerase Dependente de RNA/metabolismo , Vírus Reordenados/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Eletricidade Estática , Células VeroRESUMO
A retrospective immunohistochemical study was designed to investigate the frequency of concomitant traditional infectious disease pathogens in puppies that died suddenly and review the aspects of associated pathogenesis. Fifteen puppies were evaluated; the pathology reports and histopathologic slides of these animals were reviewed to determine the pattern of histopathologic lesions. The intralesional identification of antigens of canine (distemper) morbillivirus (CDV), canine adenovirus-1 and -2 (CAdV-1 and -2), canine parvovirus-2 (CPV-2), Toxoplasma gondii, and Neospora caninum was evaluated by IHC within the histopathologic patterns observed. All puppies contained CDV nucleic acid by molecular testing. The most frequent histopathologic patterns were intestinal crypt necrosis (n = 8), white matter cerebellar demyelination (n = 7), necrohaemorrhagic hepatitis (n = 7), interstitial pneumonia (n = 7), and gallbladder oedema (n = 5). All puppies contained intralesional antigens of CDV in multiple tissues resulting in singular (n = 3), and concomitant dual (n = 3), triple (n = 5) and quadruple (n = 4) infections by CAdV-1, and -2, CPV-2, and N. caninum; T. gondii was not identified. Concomitant infections by CDV was observed with N. caninum (100%; 1/1), CPV-2 (100%; 8/8), CAdV-1 (100%; 8/8), and CAdV-2 (100%; 8/8). Intralesional antigens of CDV and not CAdV-1 were identified in cases of gallbladder oedema. The "blue eye" phenomenon was histologically characterized by corneal oedema and degenerative lesions to the corneal epithelium, without inflammatory reactions.
Assuntos
Adenovirus Caninos/metabolismo , Coccidiose , Vírus da Cinomose Canina/metabolismo , Doenças do Cão , Neospora/metabolismo , Parvovirus Canino/metabolismo , Viroses , Animais , Coccidiose/metabolismo , Coccidiose/patologia , Coccidiose/veterinária , Doenças do Cão/metabolismo , Doenças do Cão/patologia , Cães , Feminino , Imuno-Histoquímica , Masculino , Estudos Retrospectivos , Viroses/metabolismo , Viroses/patologia , Viroses/veterináriaRESUMO
Oncolytic virotherapy is a novel strategy for treatment of cancer in humans and companion animals as well. Canine distemper virus (CDV), a paramyxovirus, has proven to be oncolytic through induction of apoptosis in canine-derived tumour cells, yet the mechanism behind this inhibitory action is poorly understood. In this study, three human mammary tumour cell lines and one canine-derived adenofibrosarcoma cell line were tested regarding to their susceptibility to CDV infection, cell proliferation, apoptosis, mitochondrial membrane potential and expression of tumour necrosis factor-alpha-induced protein 8 (TNFAIP8). CDV replication-induced cytopathic effect, decrease of cell proliferation rates, and >45% of infected cells were considered death and/or under late apoptosis/necrosis. TNFAIP8 and CDVM gene expression were positively correlated in all cell lines. In addition, mitochondrial membrane depolarization was associated with increase in virus titres (p < 0.005). Thus, these results strongly suggest that both human and canine mammary tumour cells are potential candidates for studies concerning CDV-induced cancer therapy.
Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Vírus da Cinomose Canina/metabolismo , Terapia Viral Oncolítica/veterinária , Adenocarcinoma/terapia , Adenocarcinoma/veterinária , Animais , Apoptose , Neoplasias da Mama/terapia , Morte Celular , Linhagem Celular Tumoral , Proliferação de Células , Doenças do Cão/terapia , Cães , Feminino , Humanos , Neoplasias Mamárias Animais/terapia , Terapia Viral Oncolítica/métodosRESUMO
Histiocytic sarcomas represent rare but fatal neoplasms in humans. Based on the absence of a commercially available human histiocytic sarcoma cell line the frequently affected dog displays a suitable translational model. Canine distemper virus, closely related to measles virus, is a highly promising candidate for oncolytic virotherapy. Therapeutic failures in patients are mostly associated with tumour invasion and metastasis often induced by misdirected cytoskeletal protein activities. Thus, the impact of persistent canine distemper virus infection on the cytoskeletal protein cortactin, which is frequently overexpressed in human cancers with poor prognosis, was investigated in vitro in a canine histiocytic sarcoma cell line (DH82). Though phagocytic activity, proliferation and apoptotic rate were unaltered, a significantly reduced migration activity compared to controls (6 hours and 1 day after seeding) accompanied by a decreased number of cortactin mRNA transcripts (1 day) was detected. Furthermore, persistently canine distemper virus infected DH82 cells showed a predominant diffuse intracytoplasmic cortactin distribution at 6 hours and 1 day compared to controls with a prominent membranous expression pattern (p ≤ 0.05). Summarized, persistent canine distemper virus infection induces reduced tumour cell migration associated with an altered intracellular cortactin distribution, indicating cytoskeletal changes as one of the major pathways of virus-associated inhibition of tumour spread.
Assuntos
Movimento Celular , Cortactina/biossíntese , Vírus da Cinomose Canina/metabolismo , Cinomose/metabolismo , Regulação Neoplásica da Expressão Gênica , Sarcoma Histiocítico/metabolismo , Proteínas de Neoplasias/biossíntese , Animais , Linhagem Celular Tumoral , Cinomose/patologia , Cães , Sarcoma Histiocítico/patologia , Sarcoma Histiocítico/virologia , HumanosRESUMO
Canine distemper virus (CDV) becomes able to use human receptors through a single amino acid substitution in the H protein. In addition, CDV strains possessing an intact C protein replicate well in human epithelial H358 cells. The present study showed that CDV strain 007Lm, which was isolated from lymph node tissue of a dog with distemper, failed to replicate in H358 cells, although it possessed an intact C protein. Sequence analyses suggested that a cysteine-to-tyrosine substitution at position 267 of the V protein caused this growth defect. Analyses using H358 cells constitutively expressing the CDV V protein showed that the V protein with a cysteine, but not that with a tyrosine, at this position effectively blocked the interferon-stimulated signal transduction pathway, and supported virus replication of 007Lm in H358 cells. Thus, the V protein as well as the C protein appears to be functional and essential for CDV replication in human epithelial cells.
Assuntos
Vírus da Cinomose Canina/metabolismo , Cinomose/virologia , Células Epiteliais/virologia , Replicação Viral/fisiologia , Animais , Linhagem Celular , Cães , Células Epiteliais/metabolismo , HumanosRESUMO
Canine distemper virus (CDV) causes highly contagious respiratory, gastrointestinal, and neurological diseases in wild and domestic animal species. Despite a broad vaccination campaign, the disease is still a serious problem worldwide. In this study, six field CDV strains were isolated from three dogs, two raccoon dogs, and one badger in Korea. The full sequence of the genes encoding fusion (F) and hemagglutinin (H) proteins were compared with those of other CDVs including field and vaccine strains. The phylogenetic analysis for the F and H genes indicated that the two CDV strains isolated from dogs were most closely related to Chinese strains in the Asia-1 genotype. Another four strains were closely related to Japanese strains in the Asia-2 genotype. The six currently isolated strains shared 90.2-92.1% and 88.2-91.8% identities with eight commercial vaccine strains in their nucleotide and amino acid sequences of the F protein, respectively. They also showed 90.1-91.4% and 87.8-90.7% identities with the same vaccine strains in their nucleotide and deduced amino acid sequences of the H protein, respectively. Different N-linked glycosylation sites were identified in the F and H genes of the six isolates from the prototype vaccine strain Onderstepoort. Collectively, these results demonstrate that at least two different CDV genotypes currently exist in Korea. The considerable genetic differences between the vaccine strains and wild-type isolates would be a major factor of the incomplete protection of dogs from CDV infections.
Assuntos
Vírus da Cinomose Canina/genética , Cinomose/virologia , Hemaglutininas Virais/genética , Mustelidae/virologia , Cães Guaxinins/virologia , Proteínas Virais de Fusão/genética , Sequência de Aminoácidos , Animais , Animais Selvagens/virologia , Vírus da Cinomose Canina/classificação , Vírus da Cinomose Canina/isolamento & purificação , Vírus da Cinomose Canina/metabolismo , Cães , Hemaglutininas Virais/metabolismo , Dados de Sequência Molecular , Filogenia , República da Coreia , Homologia de Sequência de Aminoácidos , Proteínas Virais de Fusão/metabolismoRESUMO
A canine distemper virus (CDV) strain, CYN07-dV, associated with a lethal outbreak in monkeys, used human signaling lymphocyte activation molecule as a receptor only poorly but readily adapted to use it following a P541S substitution in the hemagglutinin protein. Since CYN07-dV had an intrinsic ability to use human nectin-4, the adapted virus became able to use both human immune and epithelial cell receptors, as well as monkey and canine ones, suggesting that CDV can potentially infect humans.
Assuntos
Adaptação Fisiológica/genética , Antígenos CD/metabolismo , Moléculas de Adesão Celular/metabolismo , Surtos de Doenças , Vírus da Cinomose Canina/metabolismo , Macaca/virologia , Doenças dos Macacos/virologia , Receptores de Superfície Celular/metabolismo , Substituição de Aminoácidos , Animais , Chlorocebus aethiops , Cinomose/epidemiologia , Cinomose/virologia , Vírus da Cinomose Canina/genética , Vírus da Cinomose Canina/patogenicidade , Cães , Células Epiteliais/metabolismo , Células Epiteliais/virologia , Hemaglutininas Virais/genética , Humanos , Doenças dos Macacos/epidemiologia , Doenças dos Macacos/mortalidade , Receptores Virais/metabolismo , Membro 1 da Família de Moléculas de Sinalização da Ativação Linfocitária , Células VeroRESUMO
Recent outbreaks in monkeys have proven that canine distemper virus (CDV) causes diseases in a wide range of mammals. CDV uses SLAM and nectin4 as receptors to replicate in susceptible animals. Here, we show that human nectin4, but not human SLAM, is fully functional as a CDV receptor. The CDV Ac96I strain hardly replicated in nectin4-expressing human epithelial NCI-H358 cells, but readily adapted to grow in them. Unsurprisingly, no amino acid change in the H protein was required for the adaptation. The original Ac96I strain possessed a truncated C protein, and a subpopulation possessing the intact C protein was selected after growth in NCI-H358 cells. Other CDV strains possessing the intact C protein showed significantly higher growth abilities in NCI-H358 cells than the Ac96I strain with the truncated C protein. These findings suggest that the C protein is functional in human epithelial cells and critical for CDV replication in them.
Assuntos
Moléculas de Adesão Celular/metabolismo , Vírus da Cinomose Canina/fisiologia , Células Epiteliais/virologia , Receptores Virais/metabolismo , Replicação Viral , Sequência de Aminoácidos , Animais , Antígenos CD/genética , Antígenos CD/metabolismo , Moléculas de Adesão Celular/genética , Chlorocebus aethiops , Cinomose/virologia , Vírus da Cinomose Canina/genética , Vírus da Cinomose Canina/metabolismo , Cães , Humanos , Dados de Sequência Molecular , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/metabolismo , Receptores Virais/genética , Análise de Sequência de DNA , Membro 1 da Família de Moléculas de Sinalização da Ativação Linfocitária , Células VeroRESUMO
We evaluated the expression of the Fas receptor gene in Vero cells infected with the Lederle vaccine strain of canine distemper virus using RT-PCR. Vero cells were plated, and after being grown for 24 h in MEM with 5% FBS, 80-90% confluent monolayer cultures were infected with the virus. The cells were harvested at 3, 6, 9, and 15 h post-infection. Uninfected Vero cells were used as a control. Total RNA was isolated from Vero cells using 1 mL Trizol(®) LS, and RT was performed using 2 µg total RNA. Primer pairs for RT-PCR amplification for the canine distemper virus nucleocapsid gene, the S26 reference gene, and the Vero rFas gene were used to analyze expression in Vero cells. RT-PCR results revealed virus activity at 3, 6, 9, and 15 h in the virus-infected Vero cells. The S26 housekeeping gene was amplified in virus infected and control samples. However, expression of the cell death receptor Fas was detected in Vero cells only at 15 h post-infection. We suggest that the Lederle vaccine induces apoptosis by Fas receptor signaling, possibly through caspase-8 signaling rather than through mitochondrial signaling in the infected cells.
Assuntos
Apoptose , Vírus da Cinomose Canina/metabolismo , Cinomose/metabolismo , Regulação da Expressão Gênica , Transdução de Sinais , Receptor fas/biossíntese , Animais , Caspase 8/metabolismo , Chlorocebus aethiops , Cães , Fatores de Tempo , Células Vero , Vacinas Virais/metabolismoRESUMO
Paramyxovirinae envelope glycoproteins constitute a premier model to dissect how specific and dynamic interactions in multisubunit membrane protein complexes can control deep-seated conformational rearrangements. However, individual residues that determine reciprocal specificity of the viral attachment and fusion (F) proteins have not been identified. We have developed an assay based on a pair of canine distemper virus (CDV) F proteins (strains Onderstepoort (ODP) and Lederle) that share approximately 95% identity but differ in their ability to form functional complexes with the measles virus (MV) attachment protein (H). Characterization of CDV F chimeras and mutagenesis reveals four residues in CDV F-ODP (positions 164, 219, 233, and 317) required for productive interaction with MV H. Mutating these residues to the Lederle type disrupts triggering of F-ODP by MV H without affecting functionality when co-expressed with CDV H. Co-immunoprecipitation shows a stronger physical interaction of F-ODP than F-Lederle with MV H. Mutagenesis of MV F highlights the MV residues homologous to CDV F residues 233 and 317 as determinants for physical glycoprotein interaction and fusion activity under homotypic conditions. In assay reversal, the introduction of sections of the CDV H stalk into MV H shows a five-residue fragment (residues 110-114) to mediate specificity for CDV F-Lederle. All of the MV H stalk chimeras are surface-expressed, show hemadsorption activity, and trigger MV F. Combining the five-residue H chimera with the CDV F-ODP quadruple mutant partially restores activity, indicating that the residues identified in either glycoprotein contribute interdependently to the formation of functional complexes. Their localization in structural models of F and H suggests that placement in particular of F residue 233 in close proximity to the 110-114 region of H is structurally conceivable.
Assuntos
Vírus da Cinomose Canina/metabolismo , Paramyxoviridae/metabolismo , Proteínas Virais de Fusão/química , Sequência de Aminoácidos , Animais , Chlorocebus aethiops , Cricetinae , Cães , Modelos Moleculares , Dados de Sequência Molecular , Mutação , Ligação Proteica , Proteínas Recombinantes de Fusão/química , Propriedades de Superfície , Células VeroRESUMO
Canine distemper virus (CDV) causes a chronic, demyelinating, progressive or relapsing neurological disease in dogs, because CDV persists in the CNS. Persistence of virulent CDV, such as the A75/17 strain has been reproduced in cell cultures where it is associated with a non-cytolytic infection with very limited cell-cell fusion. This is in sharp contrast to attenuated CDV infection in cell cultures, such as the Onderstepoort (OP) CDV strain, which produces extensive fusion activity and cytolysis. Fusion efficiency may be determined by the structure of the viral fusion protein per se but also by its interaction with other structural proteins of CDV. This was studied by combining genes derived from persistent and non-persistent CDV strains in transient transfection experiments. It was found that fusion efficiency was markedly attenuated by the structure of the fusion protein of the neurovirulent A75/17-CDV. Moreover, we showed that the interaction of the surface glycoproteins with the M protein of the persistent strain greatly influenced fusion activity. Site directed mutagenesis showed that the c-terminus of the M protein is of particular importance in this respect. Interestingly, although the nucleocapsid protein alone did not affect F/H-induced cell-cell fusion, maximal inhibition occurred when the latter was added to combined glycoproteins with matrix protein. Thus, the present study suggests that very limited fusogenicity in virulent CDV infection, which favours persistence by limiting cell destruction involves complex interactions between all viral structural proteins.
Assuntos
Fusão de Membrana , Proteínas do Nucleocapsídeo/metabolismo , Proteínas da Matriz Viral/metabolismo , Animais , Fusão Celular , Chlorocebus aethiops , Vírus da Cinomose Canina/genética , Vírus da Cinomose Canina/metabolismo , Cães , Células Vero , Proteínas Virais de Fusão/metabolismo , Proteínas da Matriz Viral/química , Proteínas Estruturais Virais/metabolismoRESUMO
Canine distemper virus (CDV), a mobillivirus related to measles virus causes a chronic progressive demyelinating disease, associated with persistence of the virus in the central nervous system (CNS). CNS persistence of morbilliviruses has been associated with cell-to-cell spread, thereby limiting immune detection. The mechanism of cell-to-cell spread remains uncertain. In the present study we studied viral spread comparing a cytolytic (non-persistent) and a persistent CDV strain in cell cultures. Cytolytic CDV spread in a compact concentric manner with extensive cell fusion and destruction of the monolayer. Persistent CDV exhibited a heterogeneous cell-to-cell pattern of spread without cell fusion and 100-fold reduction of infectious viral titers in supernatants as compared to the cytolytic strain. Ultrastructurally, low infectious titers correlated with limited budding of persistent CDV as compared to the cytolytic strain, which shed large numbers of viral particles. The pattern of heterogeneous cell-to-cell viral spread can be explained by low production of infectious viral particles in only few areas of the cell membrane. In this way persistent CDV only spreads to a small proportion of the cells surrounding an infected one. Our studies suggest that both cell-to-cell spread and limited production of infectious virus are related to reduced expression of fusogenic complexes in the cell membrane. Such complexes consist of a synergistic configuration of the attachment (H) and fusion (F) proteins on the cell surface. F und H proteins exhibited a marked degree of colocalization in cytolytic CDV infection but not in persistent CDV as seen by confocal laser microscopy. In addition, analysis of CDV F protein expression using vaccinia constructs of both strains revealed an additional large fraction of uncleaved fusion protein in the persistent strain. This suggests that the paucity of active fusion complexes is due to restricted intracellular processing of the viral fusion protein.
Assuntos
Comunicação Celular , Vírus da Cinomose Canina/metabolismo , Cinomose/patologia , Animais , Western Blotting , Chlorocebus aethiops , Cinomose/virologia , Imunofluorescência/métodos , Regulação Viral da Expressão Gênica , Glicoproteínas/metabolismo , Microscopia Confocal , Microscopia Eletrônica/métodos , RNA Mensageiro/biossíntese , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo , Tripsina/farmacologia , Células Vero/ultraestrutura , Células Vero/virologia , Proteínas Virais de Fusão/metabolismo , Proteínas Virais/metabolismoRESUMO
The fusion (F) proteins of most paramyxoviruses are classical type I glycoproteins with a short hydrophobic leader sequence closely following the translation initiation codon. The predicted reading frame of the canine distemper virus (CDV) F protein is more complex, with a short hydrophobic sequence beginning 115 codons downstream of the first AUG. To verify if the sequence between the first AUG and the hydrophobic region is translated, we produced a specific antiserum that indeed detected a short-lived F protein precursor that we named PreF(0). A peptide resulting from PreF(0) cleavage was identified and named Pre, and its half-life was measured to be about 30 min. PreF(0) cleavage was completed before proteolytic activation of F(0) into its F(1) and F(2) subunits by furin. To test the hypothesis that the Pre peptide may influence protein activity, we compared the function of F proteins synthesized with that peptide to that of F proteins synthesized with a shorter amino-terminal signal sequence. F proteins synthesized with the Pre peptide were more stable and less active. Thus, the Pre peptide modulates the function of the CDV F protein. Interestingly, a distinct two-hit activation process has been recently described for human respiratory syncytial virus, another paramyxovirus.
Assuntos
Sequência de Aminoácidos , Vírus da Cinomose Canina/patogenicidade , Regulação Viral da Expressão Gênica , Precursores de Proteínas/genética , Proteínas Virais de Fusão/metabolismo , Animais , Chlorocebus aethiops , Vírus da Cinomose Canina/genética , Vírus da Cinomose Canina/metabolismo , Cães , Dados de Sequência Molecular , Mutação , Biossíntese de Proteínas , Precursores de Proteínas/metabolismo , Recombinação Genética , Células Vero , Proteínas Virais de Fusão/genéticaRESUMO
Sixteen dogs with naturally occurring acute and chronic canine distemper virus (CDV) encephalitis were examined immunohistochemically for the presence of the five major CDV-specific proteins in the central nervous system. Monoclonal antibodies (mAbs) directed against two, three, four and five epitopes of the nucleo- (N), phospho- (P), fusion (F), and hemagglutinin (H) protein, respectively, and a polyclonal monospecific antibody recognizing the matrix (M) protein were used. Both core proteins and their epitopes, three F protein epitopes and the M protein were demonstrated in all animals examined. A fourth F protein epitope was found only in 13 animals. The H-2 and H-3 epitope of the H protein were detected in 15, the H-1 and H-5 epitope in 14, and the H-4 epitope in 3 animals. All viral proteins were observed in the same types of brain cells including neurons and astrocytes. The N and P protein were demonstrated in nucleus, cytoplasm and cell processes, whereas M, H and F protein were observed in the cytoplasm only and rarely in cell processes. In addition, the M protein was detected occasionally in the nucleus of neurons and reactive astrocytes. Intralesional distribution of CDV-specific proteins varied between core and surface proteins. In acute and subacute lesions without associated inflammation, expression of the M, H and F protein was only slightly diminished compared to the N and P protein. However, plaques with severe inflammation were either devoid of viral antigen or exhibited N- and P protein-specific immunoreactivity exclusively at the periphery, whereas expression of surface proteins was severely reduced or absent. These results are suggestive of restricted synthesis of CDV envelope proteins in acute, and more prominent in chronic, distemper encephalitis.
Assuntos
Vírus da Cinomose Canina/metabolismo , Cinomose/metabolismo , Encefalite/metabolismo , Proteínas de Membrana/biossíntese , Proteínas Virais/biossíntese , Animais , Anticorpos Monoclonais/imunologia , Encéfalo/patologia , Movimento Celular/fisiologia , Cinomose/imunologia , Cães , Encefalite/imunologia , Encefalite/veterinária , Epitopos/imunologia , Feminino , Imuno-Histoquímica , Masculino , Proteínas de Membrana/imunologia , Proteínas do Core Viral/biossíntese , Proteínas do Core Viral/imunologia , Proteínas Virais/imunologiaRESUMO
The replication of canine distemper virus (CDV) in Vero cells was found to require certain amino acids such as arginine, methionine and valine. The deprivation of methionine caused the most marked reduction in virus yield. In cells cultured in medium deprived of methionine, the early processes of viral replication such as adsorption, penetration and uncoating of virus occurred at normal rates, but the syntheses of viral RNA and protein were markedly reduced. The addition of S-adenosylmethionine to methionine-free medium resulted in the growth of CDV to the level obtained in cells with complete medium. Moreover, cycloleucine, which is known to reduce the methylation of mRNA by inhibiting the synthesis of S-adenosylmethionine, also inhibited the growth of CDV, and the addition of methionine or S-adenosylmethionine reversed the inhibitory effect of cycloleucine. The possibility of an inhibition of methylation of mRNA in methionine-deprived cells is discussed.
Assuntos
Vírus da Cinomose Canina/metabolismo , Metionina/metabolismo , Replicação Viral/efeitos dos fármacos , Animais , Linhagem Celular , Cicloleucina/metabolismo , Vírus da Cinomose Canina/crescimento & desenvolvimento , Vírus da Cinomose Canina/fisiologia , Metionina/farmacologia , RNA Viral/biossíntese , S-Adenosilmetionina/metabolismo , Proteínas Virais/biossínteseAssuntos
Vírus da Cinomose Canina/metabolismo , Cinomose/metabolismo , Doenças do Cão/metabolismo , Encefalite/veterinária , Proteínas Virais/biossíntese , Animais , Anticorpos Antivirais/análise , Vírus da Cinomose Canina/crescimento & desenvolvimento , Vírus da Cinomose Canina/imunologia , Cães , Encefalite/metabolismo , Humanos , Biossíntese Peptídica , Panencefalite Esclerosante Subaguda/metabolismo , Proteínas Virais/imunologiaRESUMO
Gas chromatograms of sertim extracts of dogs inoculated with canine infectious hepatitis virus showed two metabolites not observed in uninoculated animals. Chromatograms of extracts of tissue cultures of dog kidney, inoculated with viruses causing canine hepatitis, herpes, and distemper, and a parainfluenza virus similar to simian virus-5, each showed two or more different metabolites. Two of the distinguishing products from cultures inoculated with hepatitis virus were chromatographically indistinguishable from those found in serums of the animals.