Autophagy is a highly conserved process that degrades damaged macromolecules and organelles. Unlike animals, only scant information is available regarding nitric oxide (NO)-induced autophagy in plants. Such lack of information prompted us to study the roles of the NO donors’ nitrate, nitrite, and sodium nitroprusside in this catabolic process in wheat roots. Furthermore, spermine, a polyamine that is found in all eukaryotic cells, was also tested as a physiological NO donor. Here, we show that in wheat roots, NO donors and spermine can trigger autophagy, with NO and reactive oxygen species (ROS) playing signaling roles based on the visualization of autophagosomes, analyses of the levels of NO, ROS, mitochondrial activity, and the expression of autophagic (ATG) genes. Treatment with nitrite and nitroprusside causes an energy deficit, a typical prerequisite of autophagy, which is indicated by a fall in mitochondrial potential, and the activity of mitochondrial complexes. On the contrary, spermine sustains energy metabolism by upregulating the activity of appropriate genes, including those that encode glyceraldehyde 3-phosphate dehydrogenase GAPDH and SNF1-related protein kinase 1 SnRK1. Taken together, our data suggest that one of the key roles for NO in plants may be to trigger autophagy via diverse mechanisms, thus facilitating the removal of oxidized and damaged cellular constituencies.

Autophagy is a highly conserved catabolic process in eukaryotic cells. Reactive nitrogen species play roles as inductors and signaling molecules of autophagy. A key mechanism of NO-mediated signaling is S-nitrosylation, a post-translational modification (PTM) of proteins at cysteine residues. In the present work, we analyzed the patterns of protein S-nitrosylation during the induction of autophagy in Triticum aestivum roots. The accumulation of S-nitrosylated proteins in the cells during autophagy induced with KNO₂ and antimycin A was visualized using monoclonal antibodies with a Western blot analysis, and proteins were identified using a standard bottom-up proteomics approach. Protein S-nitrosylation is a labile and reversible PTM, and therefore the SNO group can be lost during experimental procedures. A subsequent bioinformatic analysis using predictive algorithms and protein-ligand docking showed that identified proteins possess hypothetical S-nitrosylation sites. Analyzing protein–protein interaction networks enabled us to discover the targets that can directly interact with autophagic proteins, and those that can interact with them indirectly via key multifunctional regulatory proteins. In this study, we show that S-nitrosylation is a key mechanism of NO-mediated regulation of autophagy in wheat roots. A combination of in silico predictive algorithms with a mass spectrometry analysis provides a targeted approach for the identification of S-nitrosylated proteins.

Tungsten belong to heavy metal group, which physiological, biochemical, and molecular action mechanisms are essentially unstudied despite metal wide application in light, heavy, and military industries and the gradual accumulation in the environment. Protein phosphorylation/dephosphorylation (one of the most important posttranslational modifications) is a highly conserved mechanism of intracellular signaling and regulation of many processes of cell activity. Protein tyrosine phosphorylation/dephosphorylation is required for the cell cycle processing, plant growth and differentiation. In this work, the effects of sodium tungstate on pea (Pisum sativum L. cv. Truzhenik) root growth, protein tyrosine phosphorylation, and phosphatase activity in the roots were studied. It was shown that sodium tungstate suppressed growth, changed the mitotic index in the root meristem, and delayed cells at some mitosis phases. Under the influence of tungstate, hydrogen peroxide accumulated in the roots and phosphatase activity was inhibited. It was established by two-dimension electrophoresis and immunoblotting with the highly specific to phosphotyrosine antibody (PY20) that tungstate treatment increased both the number of such proteins and their specific phosphorylation. It is supposed that the inhibition of protein tyrosine phosphatases was one of the reasons for tungstateinduced pea root growth inhibition.

The lipid composition of the mycobint and photobiont symbiotic partners of lichenized ascomycetes varies greatly. The aim of this study was to compare the profile of the major sterols in two closely related lichens from the genus Lobaria with different photobionts. The three-component lichen Lobaria pulmonaria has two photobionts. While the main photobiont is the chlorophycean alga Symbiochloris reticulata, this lichen contains small amounts of the cyanobacterium Nostoc. By contrast, the cyanobacterium Nostoc is the main photobiont in Lobaria retigera. Relatively loosely bound sterols were extracted using a chloroform–methanol mixture, and subsequently, more tightly bound sterols by alkaline saponification. The initial chloroform–methanol extraction step indicated that ergosterol is the principal sterol in both species, with phytosterols constituting a minor fraction. However, the addition of an alkaline saponification step to the standard protocol of sterol extraction greatly increases the release of tightly bound phytosterols, such as campesterol, stigmasterol, and β-sitosterol from L. pulmonaria, but not from L. retigera. Therefore, the mycobionts and Nostoc mainly possess sterols extractable by the standard mixture of chloroform/methanol, while the chlorophycean algal photobiont contains tightly bound sterols. This observation could be important when studying the roles of sterols in the stress tolerance of lichens.