Nitric Oxide Induces Autophagy in Triticum aestivum Roots.

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.

Mdivi-1 Induced Mitochondrial Fusion as a Potential Mechanism to Enhance Stress Tolerance in Wheat.

Mitochondria play a key role in providing energy to cells. These organelles are constantly undergoing dynamic processes of fusion and fission that change in stressful conditions. The role of mitochondrial fusion in wheat root cells was studied using Mdivi-1, an inhibitor of the mitochondrial fragmentation protein Drp1. The effect of the inhibitor was studied on mitochondrial dynamics in the roots of wheat seedlings subjected to a wounding stress, simulated by excision. Treatment of the stressed roots with the inhibitor increased the size of the mitochondria, enhanced their functional activity, and elevated their membrane potentials. Mitochondrial fusion was accompanied by a decrease in ROS formation and associated cell damage. Exposure to Mdivi-1 also upregulated genes encoding the glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and an energy sensor AMP-dependent protein sucrose non-fermenting-related kinase (SnRK1), suggesting that mitochondrial fusion is associated with a general activation of energy metabolism. Controlling mitochondrial fusion rates could change the physiology of wheat plants by altering the energy status of the cell and helping to mitigate the effects of stress.

Effect of respiratory inhibitors on mitochondrial complexes and ADP/ATP translocators in the Triticum aestivum roots.

Effective functioning of the mitochondrial complexes of the oxidative phosphorylation (OXPHOS) system is necessary for ATP synthesis. The OXPHOS complexes exist both as individual forms and supercomplexes, whose formation and stability are supported by specific protein and lipid factors. In this paper, we report on the types and activities of OXPHOS complexes and supercomplexes from wheat (Triticum aestivum L.) root mitochondria analyzed by blue native polyacrylamide gel electrophoresis (BN-PAGE). The activity of OXPHOS complexes decreased when a mixture of rotenone, an inhibitor of complex I, and antimycin A, an inhibitor of complex III (R + AA) was applied to the BN-PAGE gels. By contrast, the types and activities of the OXPHOS complexes and supercomplexes did not change when they were isolated from the R + AA treated roots. However, the amount of the mitochondrial membrane-bound low molecular mass proteins in these roots markedly increased. The proteins were identified as ANT1 and ANT2 (ADP/ATP translocators) and ABA 8'-hydroxylase. We suggest that these low molecular mass proteins contribute to fine control mechanisms that stabilize mitochondrial supercomplexes and help to overcome an inhibitor-induced energy deficit by enhancing ADP/ATP transfer and ultimately improving the supply of ATP.

Mitochondrial morphology and dynamics in Triticum aestivum roots in response to rotenone and antimycin A.

Mitochondria are dynamic organelles, capable of fusion and fission as a part of cellular responses to various signals, such as the shifts in the redox status of a cell. The mitochondrial electron transport chain (ETC.) is involved in the generation of reactive oxygen species (ROS), with complexes I and III contributing the most to this process. Disruptions of ETC. can lead to increased ROS generation. Here, we demonstrate the appearance of giant mitochondria in wheat roots in response to simultaneous application of the respiratory inhibitors rotenone (complex I of mitochondrial ETC.) and antimycin A (complex III of mitochondrial ETC.). The existence of such megamitochondria was temporary, and following longer treatment with inhibitors mitochondria resumed their conventional size and oval shape. Changes in mitochondrial morphology were accompanied with a decrease in mitochondrial potential and an unexpected increase in oxygen consumption. Changes in mitochondrial morphology and activity may result from the fusion and fission of mitochondria induced by the disruption of mitochondrial ETC. Results from experiments with the inhibitor of mitochondrial fission Mdivi-1 suggest that the retarded fission may facilitate plant mitochondria to appear in a fused shape. The processes of mitochondrial fusion and fission are involved in the regulation of the efficacy of the functions of the respiratory chain complexes and ROS metabolism during stresses. The changes in morphology of mitochondria, along with the changes in their functional activity, can be a part of the strategy of the plant adaptation to stresses.