Influence of Plant Growth-Promoting Rhizobacteria on the Formation of Apoplastic Barriers and Uptake of Water and Potassium by Wheat Plants.

The formation of apoplastic barriers is important for controlling the uptake of water and ions by plants, thereby influencing plant growth. However, the effects of plant growth-promoting bacteria on the formation of apoplastic barriers, and the relationship between these effects and the ability of bacteria to influence the content of hormones in plants, have not been sufficiently studied. The content of cytokinins, auxins and potassium, characteristics of water relations, deposition of lignin and suberin and the formation of Casparian bands in the root endodermis of durum wheat (Triticum durum Desf.) plants were evaluated after the introduction of the cytokinin-producing bacterium Bacillus subtilis IB-22 or the auxin-producing bacterium Pseudomonas mandelii IB-Ki14 into their rhizosphere. The experiments were carried out in laboratory conditions in pots with agrochernozem at an optimal level of illumination and watering. Both strains increased shoot biomass, leaf area and chlorophyll content in leaves. Bacteria enhanced the formation of apoplastic barriers, which were most pronounced when plants were treated with P. mandelii IB-Ki14. At the same time, P. mandelii IB-Ki14 caused no decrease in the hydraulic conductivity, while inoculation with B. subtilis IB-22, increased hydraulic conductivity. Cell wall lignification reduced the potassium content in the roots, but did not affect its content in the shoots of plants inoculated with P. mandelii IB-Ki14. Inoculation with B. subtilis IB-22 did not change the potassium content in the roots, but increased it in the shoots.

Effect of ipt Gene Induction in Transgenic Tobacco Plants on Hydraulic Conductance, Formation of Apoplastic Barriers and Aquaporin Activity under Heat Shock.

Cytokinins are known to keep stomata open, which supports gas exchange and correlates with increased photosynthesis. However, keeping the stomata open can be detrimental if the increased transpiration is not compensated for by water supply to the shoots. In this study, we traced the effect of ipt (isopentenyl transferase) gene induction, which increases the concentration of cytokinins in transgenic tobacco plants, on transpiration and hydraulic conductivity. Since water flow depends on the conductivity of the apoplast, the deposition of lignin and suberin in the apoplast was studied by staining with berberine. The effect of an increased concentration of cytokinins on the flow of water through aquaporins (AQPs) was revealed by inhibition of AQPs with HgCl2. It was shown that an elevated concentration of cytokinins in ipt-transgenic plants increases hydraulic conductivity by enhancing the activity of aquaporins and reducing the formation of apoplastic barriers. The simultaneous effect of cytokinins on both stomatal and hydraulic conductivity makes it possible to coordinate the evaporation of water from leaves and its flow from roots to leaves, thereby maintaining the water balance and leaf hydration.

Effects of a Pseudomonas Strain on the Lipid Transfer Proteins, Appoplast Barriers and Activity of Aquaporins Associated with Hydraulic Conductance of Pea Plants.

Lipid transfer proteins (LTPs) are known to be involved in suberin deposition in the Casparian bands of pea roots, thereby reinforcing apoplast barriers. Moreover, the Pseudomonas mandelii IB-Ki14 strain accelerated formation of the Casparian bands in wheat plants, although involvement of LTPs in the process was not studied. Here, we investigated the effects of P. mandelii IB-Ki14 on LTPs, formation of the Casparian bands, hydraulic conductance and activity of aquaporins (AQPs) in pea plants. RT PCR showed a 1.6-1.9-fold up-regulation of the PsLTP-coding genes and an increase in the abundance of LTP proteins in the phloem of pea roots induced by the treatment with P. mandelii IB-Ki14. The treatment was accompanied with increased deposition of suberin in the Casparian bands. Hydraulic conductance did not decrease in association with the bacterial treatment despite strengthening of the apoplast barriers. At the same time, the Fenton reagent, serving as an AQPs inhibitor, decreased hydraulic conductance to a greater extent in treated plants relative to the control group, indicating an increase in the AQP activity by the bacteria. We hypothesize that P. mandelii IB-Ki14 stimulates deposition of suberin, in the biosynthesis of which LTPs are involved, and increases aquaporin activity, which in turn prevents a decrease in hydraulic conductance due to formation of the apoplast barriers in pea roots.

Dynamics of endogenous levels and subcellular localization of ABA and cytokinins during pollen germination in spruce and tobacco.

We used the enzyme-linked immunosorbent assay (ELISA) to assess the level of endogenous hormones in spruce pollen, and immunolocalization and confocal microscopy to study hormone localization in spruce and tobacco pollen. During pollen activation, the levels of ABA, zeatin, and its riboside significantly decreased. After the initiation of polar growth, the levels of all cytokinins increased sharply; ABA level also increased. In dormant spruce pollen grains, zeatin and ABA were localized uniformly throughout the cytoplasm. Zeatin was not detected in the nuclei, and the antheridial cell showed higher levels than the vegetative cell; ABA signal was detected in the cytoplasm and the nuclei. In germinating pollen, both hormones were detected mainly in plastids. The similar pattern was found in growing pollen tubes; signal from ABA also had a noticeable level in the cytosol of the tube cell, and was weaker in the antheridial cell. Zeatin fluorescence, on the other hand, was more pronounced in the antheridial cell. In non-germinated grains of tobacco, zeatin was localized mainly in organelles. ABA in dormant pollen grains demonstrated uniform localization, including the nuclei and cytoplasm of both cells. After germination, zeatin was accumulated in the plasmalemma or cell wall. ABA signal in the cytoplasm decreased; in the nuclei, it remained high. In growing tubes, the strongest zeatin and ABA signals were observed at the plasma membrane. The differences in ABA and cytokinin localization between species and dynamic changes in their level in spruce pollen highlight the key spatial and temporal parameters of hormonal regulation of gymnosperm pollen germination.

Effect of Salinity on Stomatal Conductance, Leaf Hydraulic Conductance, HvPIP2 Aquaporin, and Abscisic Acid Abundance in Barley Leaf Cells.

The stomatal closure of salt-stressed plants reduces transpiration bringing about the maintenance of plant tissue hydration. The aim of this work was to test for any involvement of aquaporins (AQPs) in stomatal closure under salinity. The changes in the level of aquaporins in the cells were detected with the help of an immunohistochemical technique using antibodies against HvPIP2;2. In parallel, leaf sections were stained for abscisic acid (ABA). The effects of salinity were compared to those of exogenously applied ABA on leaf HvPIP2;2 levels and the stomatal and leaf hydraulic conductance of barley plants. Salinity reduced the abundance of HvPIP2;2 in the cells of the mestome sheath due to it being the more likely hydraulic barrier due to the deposition of lignin, accompanied by a decline in the hydraulic conductivity, transpiration, and ABA accumulation. The effects of exogenous ABA differed from those of salinity. This hormone decreased transpiration but increased the shoot hydraulic conductivity and PIP2;2 abundance. The difference in the action of the exogenous hormone and salinity may be related to the difference in the ABA distribution between leaf cells, with the hormone accumulating mainly in the mesophyll of salt-stressed plants and in the cells of the bundle sheaths of ABA-treated plants. The obtained results suggest the following succession of events: salinity decreases water flow into the shoots due to the decreased abundance of PIP2;2 and hydraulic conductance, while the decline in leaf hydration leads to the production of ABA in the leaves and stomatal closure.

The Effects of Rhizosphere Inoculation with Pseudomonas mandelii on Formation of Apoplast Barriers, HvPIP2 Aquaporins and Hydraulic Conductance of Barley.

Pseudomonas mandelii strain IB-Ki14 has recently been shown to strengthen the apoplastic barriers of salt-stressed plants, which prevents the entry of toxic sodium. It was of interest to find out whether the same effect manifests itself in the absence of salinity and how this affects the hydraulic conductivity of barley plants. Berberine staining confirmed that the bacterial treatment enhanced the deposition of lignin and suberin and formation of Casparian bands in the roots of barley plants. The calculation of hydraulic conductance by relating transpiration to leaf water potential showed that it did not decrease in bacteria-treated plants. We hypothesized that reduced apoplastic conductivity could be compensated by the higher conductivity of the water pathway across the membranes. This assumption was confirmed by the results of the immunolocalization of HvPIP2;5 aquaporins with specific antibodies, showing their increased abundance around the areas of the endodermis and exodermis of bacteria-treated plants. The immunolocalization with antibodies against auxins and abscisic acid revealed elevated levels of these hormones in the roots of plants treated with bacteria. This root accumulation of hormones is likely to be associated with the ability of Pseudomonas mandelii IB-Ki14 to synthesize these hormones. The involvement of abscisic acid in the control of aquaporin abundance and auxins-in the regulation of and formation of apoplast barriers-is discussed.

Effects of Phytohormone-Producing Rhizobacteria on Casparian Band Formation, Ion Homeostasis and Salt Tolerance of Durum Wheat.

Inoculation with plant growth-promoting rhizobacteria can increase plant salt resistance. We aimed to reveal bacterial effects on the formation of apoplastic barriers and hormone concentration in relation to maintaining ion homeostasis and growth of salt-stressed plants. The rhizosphere of a durum wheat variety was inoculated with cytokinin-producing Bacillus subtilis and auxin-producing Pseudomonas mandelii strains. Plant growth, deposition of lignin and suberin and concentrations of sodium, potassium, phosphorus and hormones were studied in the plants exposed to salinity. Accumulation of sodium inhibited plant growth accompanied by a decline in potassium in roots and phosphorus in shoots of the salt-stressed plants. Inoculation with both bacterial strains resulted in faster appearance of Casparian bands in root endodermis and an increased growth of salt-stressed plants. B. subtilis prevented the decline in both potassium and phosphorus concentrations and increased concentration of cytokinins in salt-stressed plants. P. mandelii decreased the level of sodium accumulation and increased the concentration of auxin. Growth promotion was greater in plants inoculated with B. subtilis. Increased ion homeostasis may be related to the capacity of bacteria to accelerate the formation of Casparian bands preventing uncontrolled diffusion of solutes through the apoplast. We discuss the relative impacts of the decline in Na accumulation and maintenance of K and P content for growth improvement of salt-stressed plants and their possible relation to the changes in hormone concentration in plants.