Singlet oxygen (¹O₂) refers to the lowest excited electronic state of molecular oxygen. It easily oxidizes biological molecules and, therefore, is cytotoxic. In plant cells, ¹O₂ is formed mostly in the light in thylakoid membranes by reaction centers of photosystem II. In high concentrations, ¹O₂ destroys membranes, proteins and DNA, inhibits protein synthesis in chloroplasts leading to photoinhibition of photosynthesis, and can result in cell death. However, ¹O₂ also acts as a signal relaying information from chloroplasts to the nucleus, regulating expression of nuclear genes. In spite of its extremely short lifetime, ¹O₂ can diffuse from the chloroplasts into the cytoplasm and the apoplast. As shown by recent studies, ¹O₂-activated signaling pathways depend not only on the levels but also on the sites of ¹O₂ production in chloroplasts, and can activate two types of responses, either acclimation to high light or programmed cell death. ¹O₂ can be produced in high amounts also in root cells during drought stress. This review summarizes recent advances in research on mechanisms and sites of ¹O₂ generation in plants, on ¹O₂-activated pathways of retrograde- and cellular signaling, and on the methods to study ¹O₂ production in plants.

In plant cells the homeostatic control of energy balance involves the production and recycling of adenylates with macroergic bonds, ATP and ADP. The maintenance of anabolic processes requires the relative saturation of the adenylate pool with high energy phosphoanhydride bonds. The bulk of ATP synthesis is carried out both in mitochondria and in chloroplasts while optimal ATP levels within other cell compartments are maintained by adenylate kinases (AK). AK activity was recently found in cytosol, mitochondria, plastids and the nucleus. ATP synthesis in energy-producing organelles, as well as redistribution of nutrients among cellular compartments, requires fine-tuned regulation of ion homeostasis. A special role in energy metabolism is played by autophagy, a process of active degradation of unwanted and/or damaged cell components and macromolecules within the central lytic vacuole. So-called constitutive autophagy controls the quality of cellular contents under favorable conditions, i.e., when the cellular energy status is high. Energy depletion can lead to the activation of the pro-survival process of autophagic removal and utilization of damaged structures; the breakdown products are then used for ATP regeneration and de novo synthesis of macromolecules. Mitophagy and chlorophagy maintain the populations of healthy and functional energy-producing “stations”, preventing accumulation of defective mitochondria and chloroplasts as potential sources of dangerous reactive oxygen species. However, the increase of autophagic flux above a threshold level can lead to the execution of the vacuolar type of programmed cell death (PCD). In this case autophagy also contributes to preservation of energy through support of the outflow of nutrients from dying cells to healthy neighboring tissues. In plants, two central protein kinases, SnRK1 (Snf1-related protein kinase 1) and TOR (target of rapamycin), are responsible for the regulation of the metabolic switch between anabolic and catabolic pathways. TOR promotes the energy-demanding metabolic reactions in response to nutrient availability and simultaneously suppresses catabolism including autophagy. SnRK1, the antagonist of TOR, senses a decline in cellular energy supply and reacts by inducing autophagy through several independent pathways. Here, we provide an overview of the recent knowledge about the interplay between SnRK1 and TOR, autophagy and PCD in course of the regulation of energy balance in plants.

The microalga Haematococcus lacustris (formerly H. pluvialis) is able to accumulate high amounts of the carotenoid astaxanthin in the course of adaptation to stresses like salinity. Technologies aimed at production of natural astaxanthin for commercial purposes often involve salinity stress; however, after a switch to stressful conditions, H. lacustris experiences massive cell death which negatively influences astaxanthin yield. This study addressed the possibility to improve cell survival in H. lacustris subjected to salinity via manipulation of the levels of autophagy using AZD8055, a known inhibitor of TOR kinase previously shown to accelerate autophagy in several microalgae. Addition of NaCl in concentrations of 0.2% or 0.8% to the growth medium induced formation of autophagosomes in H. lacustris, while simultaneous addition of AZD8055 up to a final concentration of 0.2 µM further stimulated this process. AZD8055 significantly improved the yield of H. lacustris cells after 5 days of exposure to 0.2% NaCl. Strikingly, this occurred by acceleration of cell growth, and not by acceleration of aplanospore formation. The level of astaxanthin synthesis was not affected by AZD8055. However, cytological data suggested a role of autophagosomes, lysosomes and Golgi cisternae in cell remodeling during high salt stress.

Contrarily to chronic stresses, acute (i.e., fast and dramatic) changes in environmental factors like temperature, radiation, concentration of toxic substances, or pathogen attack often lead to DNA damage. Some of the stress factors are genotoxic, i.e., they damage the DNA via physical interactions or via interference with DNA replication/repair machinery. However, cytotoxic factors, i.e., those that do not directly damage the DNA, can lead to secondary genotoxic effects either via the induction of the production of reactive oxygen, carbon, or nitrogen species, or via the activation of programmed cell death and related endonucleases. The extent of this damage, as well as the ability of the cell to repair it, represent a significant part of plant stress responses. Information about DNA damage is important for physiological studies as it helps to understand the complex adaptive responses of plants and even to predict the outcome of the plant’s exposure to acute stress. Single cell gel electrophoresis (Comet assay) provides a convenient and relatively inexpensive tool to evaluate DNA strand breaks in the different organs of higher plants, as well as in unicellular algae. Comet assays are widely used in ecotoxicology and biomonitoring applications; however, they are still relatively rarely used in physiological studies. In this review, we provide an overview of the basic principles and of useful variations of the protocols of Comet assays, as well as of their use in plant studies, in order to encourage plant physiologists to include this tool in the analysis of plant stress responses.

Responses of plants deficient in chlorophyll b (Chl b) to a long-term (lasting for 7 days) decrease in illumination and subsequent restoration of normal illumination were investigated in chlorina 3613 mutant of barley (Hordeum vulgare L.). Efficiency of acclimation was estimated by productivity. Throughout the entire vegetation period, control chlorina 3613 plants growing under full natural illumination (PAR photon flux density of 2000–2200 μmol/(m²s)) were notable for a low content of chlorophyll a (Chl a), slow growth, and low productivity as compared with Donaria parent genotype (wild type). In the experiments, mature chlorina 3613 plants were shaded for 1 week, so that radiant flux density of PAR came to 60 or 40% of that in full sunlight. In experimental chlorina 3613 plants subjected to shading for 7 days, accumulation of Chl a and the lack of Chl b were accompanied by activation of growth processes and rise in total biomass; in contrast, in Donaria 7-day-long shading negatively influenced the accumulation of biomass by the plants. After restoration of full natural illumination, growth and productivity characteristics of chlorina 3613 plants, which for 7 days received only 40% of full natural illumination, became close to the characteristics of wild-type plants. Thus, the lack of Chl b in chlorina 3613 plants did not affect growth and productivity after a 7-day-long shading (to 40% of full illumination).