A mutation resulting in substitution of positively charged Lys53 with negatively charged Glu in the coat protein was introduced in the infectious cDNA copy of the genome of wild-type tobacco mosaic virus strain U1. Kinetic analysis of long-distance virus transport in plants showed that systemic distribution of the mutant virus was delayed by 5-6 days as compared with the wild-type one. On evidence of RNA sequencing in the mutant progeny, Glu50 of the coat protein was substituted with Lys after passage I to compensate for the loss of the positive charge at position 53. Electron microscopy revealed atypical inclusions (rodlike structures, multiple electron-dense globular particles) in the nuclear interchromatin space of leaf mesophyll cells infected with the mutant but not with the wild-type virus.

Amino Acid Sequence↗

PubMedDate not supplied

[Interaction of cysteine-rich protein of Carlavirus with plant defense system].

Viruses of genus Carlavirus encode a small cysteine-rich protein (CRP) of unknown function. To investigate the role of CRP of carlavirus chrysanthemum virus B (CVB), a recombinant potato virus X (PVX) genome was constructed, which carried the CVB CRP gene. Expression of CVB CRP in the PVX genetic background drastically changed the PVX symptom phenotype in N. benthamiana. Instead of symptomless infection and mild mosaic, which are characteristic of PVX in this plant host, the recombinant virus expressing CVB CRP induced formation of necrotic local lesions on inoculated leaves and necrosis of the apical leaves. In N. tabacum, the infection pattern depended on the host genotype: the recombinant PVX was able to spread systemically only in N gene-carrying plants. In agroinfiltration-mediated transient expression assay, CVB CRP did not exhibit the properties of avirulence factor in N. benthamiana and was unable to suppress post-transcriptional gene silencing. Thus, CVB CRP is the viral pathogenicity determinant controlling the virus interaction with plant hosts in a manner which depends on plant defense mediated by resistance genes such as the N gene.

Carlavirus↗

PubMedDate not supplied

[Complete nucleotide sequence of genomic RNA of the potato M-virus].

The complete nucleotide sequence of potato virus M genomic RNA has been determined to be 8534 nucleotides (with the exception of the poly(A) tail at the 3' end). The sequence contains six large open reading frames coding for proteins of mol. wt. 223206, 25438, 11893, 6793, 33906, and 12183 (in 5'----3' direction). According to its primary sequence analysis the 223K protein ORF codes for a virus RNA replicase. The in vitro translation product of 34K protein gene precipitates by the antisera against the RVM indicating that the 34K protein is the virus coat protein. The general aspects of carla- and potexvirus gene organization are discussed.

Amino Acid Sequence↗

PubMedDate not supplied

[Nucleotide sequence and structural organization of the 3'-terminal region of potato virus M].

The sequence of 2630 3'-terminal nucleotides has been determined for the genomic RNA of potato virus M (PVM), a type member of the carlavirus group. Analysis of this nucleotide sequence revealed five open reading frames coding for proteins of mol. wt. 25, 12, 7, 34 and 11 kDa (in 5'----3' direction). The PVM genome organization has been shown to be basically analogous to that of potexviruses, except that the latter at their 3' end lack the gene encoding 11K protein. Amino acid sequence comparison between the PVM 34K protein and the coat proteins of potexviruses has shown a high extent of homology in the C-terminal amino acids. Homology has also been found between the 25, 12 and 7K proteins encoded by PVM RNA and corresponding potexvirus proteins. All this gave grounds for suggestion that carlaviruses and potexviruses belong to the same subgroup of phytoviruses.

Amino Acid Sequence↗

PubMedDate not supplied

[Possible functional role of the DD-domain of RNA-dependent polymerases].

An attempt to study the functional role of one of the most conservative domains found in all RNA-dependent RNA and DNA polymerases of plant and animal viruses (the so called "DD-domain") was made. A structure similar to the "DD-domain" was found in a minor T7 phage tail protein--gpII. Antibodies against this phage protein have been raised and used to probe "DD-domain" in molecules of avian myeloblastose virus reverse transcriptase and E. coli RNA-dependent RNA polymerase. The antibodies are shown to inhibit the activity of these enzymes under certain conditions. At the same time inhibition of the reverse transcriptase reaction causes the decrease in length of the most high molecular cDNA-products as well. The experimental data obtained are discussed in view of the suggested hypothesis on the probable functional role of the "DD-domain" of RNA-dependent polymerases.

Amino Acid Sequence↗

PubMedDate not supplied

[The role of ATP and membrane potential in the penetration of phage T17 DNA into the cell during infection].

The role of ATP and membrane potential in phage T7 DNA injection into E. coli during infection has been studied. Entrance of phage T7 genes of class II and III was shown to be prevented by arsenate, indicating the requirement for phosphorylated macroergs in the phage DNA injection. The injection process was also inhibited by exposition of the cells to the uncoupler of oxidative phosphorylation. Dependence of the injection efficiency on the membrane-potential value has been shown to be sigmoidal, which suggests a regulatory role of the membrane potential in phage T7 DNA injection from the virion into the host cell.

Adenosine Triphosphate↗

PubMedDate not supplied

[Bacteriophages T3 and T7: transcription-dependent mechanism of phage DNA transport into the cell during infection].

The mechanism by which bacteriophage T3 DNA is transported into the E. coli cell during infection was studied. The data obtained testify that bacteriophage T3, similarly to what we have earlier found for bacteriophage T7, introduces its DNA into the infected cell is a transcription-dependent way. A detailed discussion is presented on the occurrence of the transcription-coupled transport of viral DNA into the infected cell and on a number of general issues concerning the transport functions of template-directed enzymes.

Biological Transport↗

PubMedDate not supplied

[Conformational transition of DNA within the B-family as revealed by anisotropy circular dichroism].

The native T2 DNA in solutions with different LiCl concentrations was oriented by pumping through a capillary device similar to that described in rf. 1. The change of the parallel, deltaepsilon parallel, and perpendicular, 2 delta epsilon perpendicular, components of circular dichroism tensor was studied as a function of salt concentration. Positive (delta epsilon parallel 280) and negative (2 delta epsilon perpendicular 280) components were shown to increase in absolute magnitude, so that there was a monotonous drop in the longwave band magnitude of the isotropic delta epsilon 280 as a result of compensation; the shortwave band magnitude of the isotopic delta 245 and its anisotropic components were constant. It is proposed that while the helix is winding as a result of LiCl rise progressive tilting of base pairs takes place. It was also shown that the glucosylated T2 DNA in solution is wound to a greater extent than the non-glucosylated DNA of the same GC-content.

Circular Dichroism↗

PubMedDate not supplied

[Transcription initiation: influence of ionic strength and actinomycin D on the kinetics of the open promoter complex formation between Escherichia coli RNA-polymerase and T7 DNA].

A membrane filter assay has been devised to study the influence of ionic strength (0--150 mM NaCl) and actinomycin D on the kinetics of the open promoter complex formation between E. coli RNA-polymerase and [3H]DNA of T7 phage. The dependence of the complex formation rate upon the ionic strength is non-monotoneus with a maximum at 75--100 mM. The addition of one actinomycin molecule per 200 base pairs decreases the open promoter complex formation rate at ionic strength less than 100 mM. A promoter site selection model including liner diffusional selection is proposed which is in a good agreement with the experimental results obtained.

DNA, Viral↗

PubMedDate not supplied

[Structural transitions in spin-labeled DNA].

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Animals↗

PubMedDate not supplied

[Conformational transitions within the limits of the 2-stranded state of spin-labeled DNA].

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Aziridines↗

PubMedDate not supplied

[Coupling of bacteriophage T7 DNA penetration with its transcription, during infection].

Further research on bacteriophage T7 DNA penetration mechanism into E. coli cells during the infection was carried out. The DNA-RNA-hybridization on nitrocellulose filters revealed that in the presence of chloramphenicol the T7 DNA penetration from the virion into a host cell was coupled with its transcription by the bacterial RNA polymerase. The data obtained indicate that in the absence of antibiotics the penetration of a part of T7 genome which correspondes to class II and III genes is coupled with its transcription by a phage-specific RNA polymerase. Along with this the host restriction-modification system when its activity is not inhibited by the phage-induced proteins will be able to cleave the penetrated T7 DNA just after its transcription was accomplished. Considering these data along with our conception on direct involvement of transcription in T7 DNA penetration process during the infection one can suggest that E. coli RNA polymerase molecules which provide the phage DNA transport, are localized at the inner surface of cytoplasmic membrane.

Chloramphenicol↗

PubMedDate not supplied

[Gradual penetration of T7 phage DNA molecules into Escherichia coli cells during infection in the presence of chloramphenicol].

The cell-attached T7 phage DNA has been analyzed when E. coli was infected in the presence of chloramphenicol or rifampicin followed by sonication to provide phage desorption. Sucrose gradient sedimentation of the cell T7-DNA followed by HpaI digestion and agarose gel electrophoresis were performed. The results obtained suggest a gradual entrance of the T7-DNA molecule into the host cell starting on its left end bearing early genes. These data support the conception that the T7-DNA entrance into host cell is directly coupled with its transcription by RNA polymerase. At the same time one more HpaI fragment was found even in the cells, infected with the phage in the process of rifampicin. It may be that this fragment corresponds to the right end of the T7 chromosome, thus suggesting that short fragment of the T7-DNA right end can also enter the host cell early in infection.

Chloramphenicol↗