[The participation of salicylic and jasmonic acids in genetic and induced resistance of tomato to Meloidogyne incognita (Kofoid and White, 1919)].

Salicylic (SA) and jasmonic (JA) acids are the best known mediators of signal systems in plants. In this investigation the participation and character of interactions between SA- and JA-signals under the induced and genetic resistance of plants to nematodes was investigated on the model system tomato (Lycopersicon esculentum) and the root-knot nematode Meloidogyne incognita. This study demonstrates that application of JA and SA to tomato foliage induces systemic effects that suppress root-knot nematode infestation, inhibition of nematode reproduction, and also increased activity of LOX and PAL, the enzymes of biosynthesis of JA and SA. JA treatment did not inhibit Mz-mediated resistance, which suggests a lack of signaling conflicts between these two forms of defense.

[Elicitor activity of chitosan and arachidonic acid: their similarity and distinction].

Two elicitors-chitosan and arachidonic acid-induced the same defense responses in potatoes, stimulating the processes of wound reparation and inducing the formation of phytoalexins, inhibitors of proteinase, and active forms of oxygen. However, chitosan induced the defense potential of plant tissues at concentrations higher than those of arachidonic acid. The protective action of chitosan was defined by two parameters, i.e., the ability to induce the immune responses in plant tissues and to exhibit a toxic effect on the pathogen development, causing late blight and seedling blight, whereas the elicitor effect of arachidonic acid depended on its ability to induce the defense potential of plant tissues only.

[Involvement of salicylic acid in induction of nematode resistance in plants].

The role of salicylic acid (SA) as a possible signaling component in the case of the infection of plants with nematodes has been studied using a model system consisting of the tomato (Lycopersicon esculentum (Mill.) and race 1 of the gall eelworm Meloidogyne incognita (Kofoid and White, 1919; Chitwood, 1949). The pre-planting SA treatment of tomato seeds results in an increased nematode resistance of susceptible tomato cultivars; the protective effect is higher in the case of SA combined with chitosan, a biogenic elicitor of plant resistance. The studied preparations stimulate the growth and development of the plants. The increase in the resistance of tomato plants is related to the increased activity of phenylalanine ammonia-lyase and an increased SA content in plant tissues infected with nematodes; both these factors significantly influence nematode development.

[Wound repair in plant tissues (Review)].

Signaling systems responsible for repair processes in plants and manifestation of defensive effects in plant tissues were analyzed. Special attention was given to jasmonic acid, a mobile systemic repair signal, as well as to jasmonate biosynthesis and signal transport to the areas where protective responses of plants are induced. The main defense responses of potato tubers induced by wounding were considered.

[Chymotrypsin and trypsin inhibitor isolated from potato tubers].

Potato Kunitz-type chymotrypsin inhibitor (PKCI-23) was isolated from potato tubers (Solanum tuberosum L., Zhukov's Jubilee breed) and purified to a homogenous state. The protein was purified by gel-filtration chromatography and ion-exchange chromatography using Sephadex G-75 and CM-Sepharose CL-6B, respectively. PKCI-23 protein has been shown to inhibit both chymotrypsin and trypsin with equal efficacy, forming equimolar complexes with these enzymes. However, much weaker inhibitory effect of PKCI-23 has been observed for Carlsberg subtilisin. The N-terminal 20 amino acid sequence of PKCI-23 has been sequenced. PKCI-23 has been shown to suppress, with different efficacy, the growth and development of pathogenic microorganisms Fusarium culmorum (Wm. G. Sm.) Sacc. and Phytophtora infestans (Mont.) de Bary that infect potato.

[Immunomodulating activity of chitosan derivatives with salicylic acid and its fragments].

A study of biological activity of the derivatives of the chitin-chitosan oligomer with salicylic acid and its fragments showed that chitosan salicylate actively protected potato tubers against Phytophthora infestans but sharply inhibited reparation of potato tissues. N-(2-Hydroxybenzyl)chitosan exhibited good protective properties but did not influence wound reparation. N-(2-Hydroxy-3-methoxybenzyl)-N-pyridox-chitosan, which contained the pyridoxal and 2-hydroxy-3-methoxy fragments, was the most efficient, stimulating both defense against late blight and wound reparation in potato tissues.

Protein trypsin inhibitor from potato tubers.

A protein of 22 kDa designated as PKTI-22 was isolated from potato tubers (Solanum tuberosum L., cv. Istrinskii) and purified to homogeneity using CM-Sepharose CL-6B ion-exchange chromatography. The protein efficiently suppressed the activity of trypsin, affected chymotrypsin less, and did not affect subtilisin Carlsberg. The N-terminal sequence of PKTI-22 (20 amino acid residues) was found to be highly homologous with the amino acid sequences of the potato Kunitz-type proteinase inhibitors of group B (PKPI-B) that were aligned from the corresponding gene sequences and was identical to the sequence (from the 2nd to the 20th residue) of the recombinant protein PKPI-B10. These data together with the observed similarity of the properties of two proteins indicate that the PKTI-22 protein is encoded by the PKPI-B10 gene.

[The inhibition of potato tubers wound reparation].

The multiple washing of the wound surface of potato tubers by water adversely affected the protective properties of wound periderm. Immune inhibitor beta-1,3-beta-1,6 glucan had a property of local effect and inhibited the process of wound healing. The pentasaccharide of xyloglucan caused necrosis of potato tuber tissue and prevented the wound reparation process.

[Wound healing and induced resistance in potato tubers].

It was demonstrated that biogenic elicitors, arachidonic acid and chitosan, locally and systemically stimulated wound healing in potato tuber tissues by increasing the number of wound periderm layers, accelerating the development of cork cambium (phellogen), and inducing proteinase inhibitors. The signal molecules, jasmonic and salicylic acids, had different effects on the development of wound periderm: jasmonic acid locally and systemically stimulated potato wound healing and elevated the level of proteinase inhibitors, whereas salicylic acid did not have any effect on wound healing and even blocked the formation of proteinase inhibitors.

[Obtaining and study of monosaccharide derivatives of low-molecular-weight chitosan].

The possibility of obtaining monosaccharide derivatives of low-molecular-weight chitosan with the use of the Maillard reaction was studied. Chitosan derivatives (molecular weight, 24 and 5 kDa) obtained with glucosamine, N-acetyl galactosamine, galactose, and mannose with a substitution degree of 4-14% and a yield of 60-80% were obtained. Some physicochemical and biological properties of these derivatives were studied. We showed that monosaccharide derivatives of low-molecular-weight chitosan exhibited antibacterial activity. Chitosan at a concentration of 0.01% caused 100% death of bacteria B. subtilis and E. coil. The strongest antibacterial effect was exhibited by 24-kDa derivatives: only 0.02-0.08% of cells survived. These derivatives were two orders of magnitude more effective than the 5-kDa chitosan modified with galactose.

[Activation of elicitor defensive properties by systemic signal molecules during the interaction between potato and Phytophthora].

The elicitor arachidonic acid in combination with jasmonic acid (JA) induced a higher level of defense against the late blight agent in potato (Solanum tuberosum L.) tissues than in combination with salicylic acid (SA). On the contrary, the elicitor chitosan displayed a higher inductive effect in combination with SA as compared with JA. The optimal concentrations of tested compounds were selected for designing the compositions activating wound repair, induction of proteinase inhibitors, and resistance to the biotrophic pathogen Phytophthora infestans (Mont.) de Bary. It was demonstrated that the compositions of elicitor and systemic signal molecules provided a faster spreading of an inducing effect in the potato tissues.

[Effect of proteinaceous proteinase inhibitors from potato tubers on the growth and development of phytopathogenic microorganisms].

We studied the effect of two proteins, PSPI-21 and PKSI, on the growth and development of phytopathogenic microorganisms (Phytophthora infestans oomycete and Fusarium culmorum fungus). Both proteins were isolated from potato tubers (Solanum tuberosum L., cv. Istrinskii) and served as inhibitors of serine proteinases. These proteins differed in the ability to inhibit growth of Phytophthora infestans oomycete and Fusarium culmorum fungus. PSPI-21 was the most potent in modulating the growth of oomycete mycelium. PKSI primarily affected the growth of the fungal mycelium. The proteins under study induced complete destruction of oomycete zoospores and partial destruction of fungal macroconidia. Our results suggest that these proteins are involved in the protection of potato plants from phytopathogenic microorganisms.

Heterologous expression, purification, and properties of a potato protein inhibitor of serine proteinases.

The gene PKPI-B10 [AF536175] encoding in potato (Solanum tuberosum L., cv. Istrinskii) a Kunitz-type protein inhibitor of proteinases (PKPI) has been cloned into the pET23a vector and then expressed in Escherichia coli. The recombinant protein PKPI-B10 obtained as inclusion bodies was denatured, separated from admixtures by ion-exchange fast protein liquid chromatography (FPLC) on MonoQ under denaturing conditions, and renatured. The native protein was additionally purified by ion-exchange FPLC on DEAE-Toyopearl. The PKPI-B10 protein effectively inhibits the activity of trypsin, significantly weaker suppresses the activity of chymotrypsin, and has no effect on other serine proteinases: human leukocyte elastase, subtilisin Carlsberg, and proteinase K, and also the plant cysteine proteinase papain.