Ribonuclease activity of buckwheat plant (Fagopyrum esculentum) cultivars with different sensitivities to buckwheat burn virus.

Ribonucleases (RNases) are present in base-level amounts in intact plants, but this level is able to increase greatly under stress conditions. The possible cause for such an increase is protection against plant RNA-virus attack. Buckwheat burn virus (BBV) is a highly virulent pathogen that belongs to Rhabdoviridae family. In our study, we have analyzed the correlation between RNase activity and resistance of different buckwheat cultivars to BBV infection. Two cultivars, Kara-Dag and Roksolana, with different sensitivities to BBV have been used. Kara-Dag is a cultivar with medium sensitivity to virus and Roksolana is a tolerant cultivar. It has been shown that the base level of RNase activity in Roksolana cultivar was in most cases higher than the corresponding parameter in Kara-Dag cultivar. Both infected and uninfected plants of Roksolana cultivar demonstrated high RNase activity during two weeks. Whereas infected plants of Kara-Dag cultivar demonstrated unstable levels of RNase activity. Significant decline in RNase activity was detected on the 7th day post infection with subsequent gradual increase in RNase activity. Decline of the RNase activity during the first week could promote the virus replication and therefore more successful infection of upper leaves of plants. Unstable levels of RNase activity in infected buckwheat plants may be explained by insufficiency of virus-resistant mechanisms that determines the medium sensitivity of the cultivar to BBV. Thus, plants of buckwheat cultivar having less sensitivity to virus, displayed in general higher RNase activity.

[Matrix activity of mRNP of different cell localization from plants infected by minus-genome curly potato dwarfness virus in protein-synthesizing system in vitro].

Using structures of mRNP of different cell localization isolated from plants infected by minus-genomecurly potato dwarfness virus (CPDV), it was established that synthesis of virus proteins in vitro is mainly realized on membrane-related and free polysomal mRNP capable to direct synthesis of proteins programmed in RNA in cell-free protein-synthesizing systems. The obtained results prove that synthesis of virus proteins is actively realized on membrane-related nonpolysomal mRNP - 23520 imp/min. An analogous distribution of matrix activity is observed also in the case when mRNA isolated from mRNP of different cell localization was introduced in protein-synthesizing system. The least matrix activity was observed in cytoplasmic nonpolysomal mRNPs which are low-active in the translation system in vitro - only 1890 imp/min. The function of free cytoplasmic nonpolysomal mRNP is apparently mainly reduced to the transport and reserve of genetic information in a cell.

[Effect of structural P-protein of potato curly dwarf virus on replicase activity of mRNP-complexes of infected plants].

All representatives of rhabdoviruses contain a nucleocapside phosphoprotein - P-protein which is an essential subunit of the viral RNA-dependent RNA polymerase complex. As a result of studying the effect of nucleocapside protein P(NS) on replicase activity of mRNP isolated from plants infected by potato curly dwarf virus in the system in vitro, it was established that nucleocapside P-protein stimulates considerably the replicase activity of membrane-bound polysomal m-RNP P-protein being available in concentration of 15 microg/ml in the replication system in vitro of membrane-bound polysomal mRNP, the replicase activity increased 11.7 times. This property of nucleocapside P-protein at the same concentration was displayed to a less extent with the presence of free polysomal mRNP, in the system in vitro. Thus the replicase activity mRNP-complexes in the replication system in vitro depends on the presence of nucleocapside viral P-protein in the system. Its concentration being increased or decreased, one can observe the change of the replicase activity.

[Fundamentals of interferon system function in pathology and molecular biological peculiarities of interferon production].

Molecular biological peculiarities of interferon system function in PV-infected persons have been found. It is evident that the interferon production, anti-inflammatory cytokines and their receptors and also defensines play an important role in the mechanism of virus interaction with sensitive cells of macroorganism with development of pathological process. The new conception of expediency for the use of interferons and their inducers as the polyfunctional regulators with a broad spectrum of activity for the treatment of PV-infected patients was suggested. Patents for the method of treatment of PV-infected patients were obtained. New inducers of interferon as well as recombinant IFN-alpha-2b was developed. Our results were introduced in the medical practice.

[Description of bacillus-shaped spot sweetflag virus].

The spot sweetflag virus (SSV) as to its morphology and structural components in comparison with phytorhabdovirus of curly potato dwarf virus (CPDV) and rhabdoviruses of vesicular stomatitis virus (VSV), pathogenic for people and animals, corresponds to the definition of rhabdovirus and belongs to Rhabdoviridae family. Virions of SSV have a bacillus-like form and dimensions 110-130 x 45 nm. SSV contains structural proteins 130, 66, 43-39, 32-30 and 25 kDa. In the virion structure the fatty acids have been identified: palmitic (47 %), linolic (4.2%), oleic (14.9%), stearic (3.94%), holesterol (23%), and also carbohydrates: glucose (25.3%), galactose (18.3%), arabinose (16%), rhamnose (3.1%) and mannose (2.32%). Aminosaccharides: glucosamine and galactosamine, with correlation 1:7.2, were also found out. The paper is presented in Ukrainian.

[Functional properties of prosomal protein 39 kappaDa from Datura stramonium leaves infected with Potato Virus X].

Free cytoplasmic informosomes isolated from Datura stramonium plants infected by PVX contain a low-molecular ribonucleoprotein complex (RNP). This complex as to its main physico-chemical parameters (sedimentation coefficient 10S, buoyant density in CsSO4 1.31 g/cm3, stability to 1% lauroylsarcosinate-Na) corresponds to the prosome (inhibitory RNP). Prosomes isolated from free mRNP of D. stramonium plants infected by PVX contain the protein of 39 kDa. This protein was shown to be capable to phosphorylate in vitro in the composition of informosomes and prosomes. It is possible that this protein can be the protein-repressor, since it is absent in the translated polysome-associated form of mRNP. The label incorporation has shown that the protein of 39 kDa is able to reduce in vitro the template activity of genomic RNA PVX to 40% and RNA TMIV--to 30%. Moreover, the protein 39 of kDa has the protease activity. It affects substrate-case in like trypsin. It is supposed that it can participate in splitting the intracellular proteins as well as in the expression of the virus genome, it can also influence the template activity of cell RNAs.

[Antigenic relationship between nucleocapsid proteins of phyto- and zoorhabdoviruses].

The methods of electrophoresis in PAAG and immunological method were used for comparative analysis of structural proteins of phytorhabdovirus of potato curly dwarf (PCDV) and zoorhabdoviruses-vesicular stomatitis virus (VSV) and fixed rabies Virus (RV). Molecular weight of viral proteins was determined by the method of polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The proteins with molecular weight 45-51 kD, are probably, the major component of the viral nucleocapsid. Nucleocapsid protein 45 kD RV virus was isolated by the method of preparative electrophoresis and then the monospecific serum was obtained. The Ouchterlony and immunoblotting method were used to show, that nucleocapsid proteins with molecular weights 51 and 45 kD both of phytorhabdovirus PCDV and zoorhabdoviruses VSV and RV are serologically related. The obtained data may be used in biotechnology as the basis for creation of a new class of diagnostic preparations with the purpose to detect RV virus using proteins of curly potato dwarf virus and may be also used in serological tests to reveal viruses of Rhabdoviridae family in various eukaryotic objects.

[Immunospecific detection of the potato virus X by the surface plasmon resonance].

Highly specific and proximal method of laboratory diagnostics of the viral disease, has been developed using the structural protein of the potato virus X, as a model, and monospecific antibodies to it. The immunospecific determination of the potato virus X was carried out by the surface plasmon resonance method using specific IgG-antigen complexes, immobilized on the sensor surface modified by rodanide and protein A of Staphylococcus aureus.

[Physico-chemical properties and functional peculiarities of mRNP of different subcellular location of Nicotiana rustica plants affected by potato curly dwarf virus]. / Fiziko-khimicheskie svoistva i funktsional'nye osobennosti mRNP razlichnoi subkletochnoi lokalizatsii rastenii Nicotiana rustica, porazhennykh virusom kurchavoi karlikovosti kartofelia.

mRNP of different subcellular location of N. rustica plants affected by PCDV contain virus-specific proteins. RNA-replicase, poly (A)-polymerase and protein kinase have been found in mRNP under study. The most expressed replicase, poly(A)-polymerase and protein kinase activity has been noticed in the systems containing membrane-bound non-polysomal mRNP.

[Enzyme activity of nucleocapsid proteins of the curly potato dwarf virus]. / Fermentativnaia aktivnost' nukleokapsidnykh belkov fitorabdovirusa kurchavoi karlikovosti kartofelia.

The individual nucleocapsid proteins of phytorhabdovirus of curly potato dwarf were isolated: N-protein (m. w. 56 kDa), NS-protein (m. w. 49 kDa), L-protein (m. w. 128 kDa). It was establish that L and NS proteins displayed enzyme activity in the replication, phosphorylation and adenylation systems in vitro. Major N protein (m. w. 56 kDa) did not show the enzyme activity in these systems. The revelation of RNA-polymerase, poly(A)-polymerase and proteinkinase activities of nucleocapsid proteins are characteristic of the viruses belonging to the Rhabdoviridae family.

[Isolation and purification of virus damaging sunflower]. / Vydilennia ta ochystka virusu, shcho soniashnyk.

A procedure has been developed for purifying intact virus's isolate particles evoking yellow spot mosaic disease in sunflower. Purification of pathogen in 0.1 M sodium phosphate buffer, pH 8.0 containing 0.05 M Na3SO3 and 0.2% 2-mercaptoethanol is used. After first clarification extract was exposed to two cycles of high-speed centrifugation and fractionated in linear 10-40% (wt vol-1) sucrose density gradient. Virus was recovered from appropriate fractions after dialysis against 0.01 M Na2SO3.