Effects of Rhizobium leguminosarum bv. viceae Strains Different in Their Symbiotic Effectiveness on Changes in cAMP and Hydrogen Peroxide Concentrations in Cells of Pea Seedlings

Russian Journal of Plant Physiology - Changes in hydrogen peroxide and cAMP concentrations in pea (Pisum sativum L.) seedlings inoculated with Rhizobium leguminosarum bv. viceae Frank (Rlv) were...     

Potato Cell Plasma Membrane Receptors to Ring Rot Pathogen Extracellular Polysaccharides

The interaction of extracellular polysaccharides (EPS) of the potato ring rot bacterial pathogen Clavibacter michiganensis ssp. sepedonicus (Spieck. et Kott.) Skaptason et Burkh. (Cms) with protoplasts isolated both from leaf cells of plants grown in vitro and microsomal membrane fractions obtained from cell suspension cultures of two potato (Solanum tuberosum L.) cultivars contrasted by their resistance to this pathogen was studied. The EPS intensively bind to protoplast surfaces and microsomal membranes of the susceptible cultivar but not to those of the resistant cultivar. Treatment with protease, excess of unlabelled EPS, and with dextran, did not lead to the binding of fluorochrome‐labelled EPS to protoplasts and microsomal membranes (from both cultivars). It is proposed that (a) a great number of receptors to EPS Cms are present in the plasma membranes of potato cells of susceptible cultivars, (b) these receptors contain proteinaceous sites exposed on the external side of the plasma membrane which participate in EPS binding, and (c) the plasma membranes of cells of resistant cultivars contain a small but sufficient quantity of receptors to EPS able to induce defensive responses in plants.     

Subcellular localization of dehydrins in wheat plant seedlings subjected to low-temperature adaptation

Subcellular localization of dehydrins (dhn) in stem cell tissues of winter wheat seedlings (Triticulum aestivum L., cult. Irkutskaya ozimaya) was studied by immunoelectron microscopy. It was found that cold hardening at 4°C for 7 days resulted in a duplication of the dhn quantity in the cells as compared with control conditions (22°C). The maximum increase of the dhn content was observed in rough endoplasmic reticulum, mitochondria, cell walls, and intercellular spaces (3.8-, 3.0- and 2.8-fold, respectively); minimum increase was found in chloroplasts (1.4-fold). In the membrane compartments (mitochondria, rough endoplasmic reticulum, chloroplasts) low-temperature stress caused an increase of dhn quantity not only near membranes but also in the intermembrane space. A significant accumulation of dhn (2.5-fold) in the nucleus under low- temperature was found. We conclude that cold hardening of the plant induces accumulation and translocation of dhn to the regions of inter- and intracellular compartments that most require protection during the low-temperature stress.     

Plant Adenylate Cyclases

Adenylate cyclase (AC) (ATP diphosphate-lyase cyclizing; EC 4.6.1.1) is a key component of the adenylate cyclase signaling system and catalyzes the generation of cyclic adenosine monophosphate (cAMP) from ATP. This review summarizes data from the literature and the authors' laboratory on the investigation of plant transmembrane (tmAC) and soluble (sAC) adenylate cyclases, in comparison with some key characteristics of adenylate cyclases of animal cells. Plant sAC has been demonstrated to exhibit similarities with animal sAC with respect to certain characteristics. External factors, such as far-red and red light, temperature, exogenous phytohormones, as well as specific triggering compounds of fungal and bacterial origin exert a significant influence on the activity of plant tmAC and sAC.     

Determination of cAMP in plant cells by a modified enzyme immunoassay method

Presently, there is no doubt about the functioning of the adenylate cyclase signaling system in plants, but the role of this system in various physiological–biochemical processes has been investigated insufficiently. Cyclic adenosine monophosphate (cAMP), the key component produced by adenylate cyclase, whose concentrations in plant cells vary rather widely, is the indicator of functional activity for this signaling way. In the latter case, in the process of determination of concentrations of this messenger, one encounters difficulties related to insufficient sensitivity of the methods most frequently applied. In this connection, the proposed mechanism is a modification of the method of the enzyme immunoassay (EIA), which is based on immediate measurement of cAMP concentrations in the sample with the use of antibodies. This modification allows us to determine the concentrations of cAMP with the precision of 5 pM, which exceeds the sensitivity of other methods by approximately 10 times. The specificity of the assay has been confirmed by other two independent tests––the capillary electrophoresis and the nuclear magnetic resonance (NMR). It has also been compared to the data obtained with the use of the commercial kit from Sigma–Aldrich. The modification has been tested on such plant objects as in vitro potato plants, and suspension cells of potato and Arabidopsis.     

Activities of Adenylate Cyclase and Changes in cAMP Concentration in Root Cells of Pea Seedlings Infected with Mutualists and Phytopathogens

Work was carried out on pea (Pisum sativum L.) seedling roots to assess the attachment of the nitrogen-fixing symbiotic bacteria Rhizobium leguminosarum bv. vicea (Rlv) and the bacterial phytopathogens—specific Pseudomonas syringae pv. pisi (Psp) and nonspecific Clavibacter michiganensis ssp. sepedonicus (Cms). Different root zones were examined: (I) the meristem, 2 mm from the root tip; (II) the root hair-free zone, 27 mm; (III) the zone of root hair anlages, 712 mm; (IV) the young root hair zone, 1217 mm; and (V) the zone of root hair that completed the growth, 1722 mm. It was found earlier that the zones differed in their susceptibility to Rlv. In the present work, reactions of particular components of the adenylate cyclase signaling system (ACSS) were estimated, i.e., concentration of cAMP and activities of transmembrane adenylate cyclase (tAC) and soluble adenylate cyclase (sAC) in these zones after different times post inoculation (5, 15, 120, and 360 min). It was revealed that the degree of activation of particular components of ACSS did not depend on the sorption rate of differently specialized bacteria. Upon contact with Rlv, the character of changes in tAC and sAC activities was almost the same in different root zones and resembled the dynamics of the cAMP content. Inoculation with Psp changed the cAMP level similarly to that with Rlv, but the dynamics of tAC and sAC was opposite to each other in most cases. Inoculation with Cms, in spite of the absence of its attachment, elevated the cAMP content and activated tAC and sAC. It is suggested that the above-mentioned changes in ACSS is associated with exometabolites of Rlv, Psp, and Cms, which activate the PAMP-induced immunity of the pea seedling cells. The uniform dynamics of cAMP in different root zones upon the exposure to Rlv and Psp seems to reflect the specific reaction and, presumably, fulfills different functions—regulatory with Rlv and defensive with Psp. Upon short-term contact with Cms, the cAMP dynamics in the same root zones displayed a nonspecific character that might be related to the rate of adsorption of exopolysaccharides by the root hair. The systemic response of ACSS was observed in the hypocotyls of the seedlings exposed to any of the three organisms.     

Effect of N-phenyl-2-naphthylamine on activity of adenylate cyclase signal system components and virulence of bacterial phytopathogens and mutualists

The effect of N-phenyl-2-naphthylamine, negative allelochemical isolated from the exudates of roots of pea (Pisum sativum L.), on the growth and activity of the adenylate cyclase signal system and virulence factors of the bacteria Rhizobium leguminosarum bv. viciae and Pseudomonas siringae pv. pisi was studied. It was demonstrated that N-phenyl-2-naphthylamine at a physiological concentration nonspecifically inhibited the growth of these bacteria in both planktonic cultures and biofilms. One of the reasons for this phenomenon is the reduction of intra- and extracellular concentrations of cAMP due to greater activation of phosphodiesterase, which disrupts cAMP, in comparison to soluble adenylyl cyclase, which synthesizes it. At the same time, N-phenyl-2-naphthylamine did not affect activity of either membrane-bound adenylyl cyclase or bacterial virulence factors.