RESUMEN
Helianthus annuus L. is an important oilseed crop, which exhibits moderate salt tolerance and can be cultivated in areas affected by salinity. Using patch-clamp electrophysiology, we have characterized Na+ influx and K+ efflux conductances in protoplasts of salt-tolerant H. annuus L. hybrid KBSH-53 under high salinity. This work demonstrates that the plasma membrane of sunflower root cells has a classic set of ionic conductances dominated by K+ outwardly rectifying channels (KORs) and non-selective cation channels (NSCCs). KORs in sunflower show extreme Na+ sensitivity at high extracellular [Ca2+] that can potentially have a positive adaptive effect under salt stress (decreasing K+ loss). Na+ influx currents in sunflower roots demonstrate voltage-independent activation, lack time-dependent component, and are sensitive to Gd3+. Sunflower Na+-permeable NSCCs mediate a much weaker Na+ influx currents on the background of physiological levels of Ca2+ as compared to other species. This suggests that sunflower NSCCs have greater Ca2+ sensitivity. The responses of Na+ influx to Ca2+ correlates well with protection of sunflower growth by external Ca2+ in seedlings treated with NaCl. It can be, thus, hypothesized that NaCl tolerance in sunflower seedling roots is programmed at the ion channel level via their sensitivity to Ca2+ and Na+.
RESUMEN
Silver nanoparticles (Ag NPs) are the world's most important nanomaterial and nanotoxicant. The aim of this study was to determine the early stages of interactions between Ag NPs and plant cells, and to investigate their physiological roles. We have shown that the addition of Ag NPs to cultivation medium, at levels above 300 mg L(-1) , inhibited Arabidopsis thaliana root elongation and leaf expansion. This also resulted in decreased photosynthetic efficiency and the extreme accumulation of Ag in tissues. Acute application of Ag NPs induced a transient elevation of [Ca(2+) ]cyt and the accumulation of reactive oxygen species (ROS; partially generated by NADPH oxidase). Whole-cell patch-clamp measurements on root cell protoplasts demonstrated that Ag NPs slightly inhibited plasma membrane K(+) efflux and Ca(2+) influx currents, or caused membrane breakdown; however, in excised outside-out patches, Ag NPs activated Gd(3+) -sensitive Ca(2+) influx channels with unitary conductance of approximately 56 pS. Bulk particles did not modify the plasma membrane currents. Tests with electron paramagnetic resonance spectroscopy showed that Ag NPs were not able to catalyse hydroxyl radical generation, but that they directly oxidized the major plant antioxidant, l-ascorbic acid. Overall, the data presented shed light on mechanisms of the impact of nanosilver on plant cells, and show that these include the induction of classical stress signalling reactions (mediated by [Ca(2+) ]cyt and ROS) and a specific effect on the plasma membrane conductance and the reduced ascorbate.
Asunto(s)
Arabidopsis/metabolismo , Membrana Celular/metabolismo , Nanopartículas del Metal/química , Plata/química , Ácido Ascórbico/metabolismo , Calcio/metabolismo , Canales Iónicos/metabolismo , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Brassinosteroids (BRs) are an important class of plant hormones with a multitude of functions. They have been intensively investigated for their biosynthesis, distribution and physiological functions. The aim of this study was to examine possible effects of BRs on the plant plasma membrane cation conductances and Ca(2+) signalling. The wheat root protoplasts (tested by patch-clamping) and excised arabidopsis roots (analysed by Ca(2+)-aequorin chemiluminometry), were used. In the whole-cell plasma membrane patches, 24-epibrassinolide, 28-homobrassionolide or 24-epicastasterone (1 µM) were applied exogenously. 24-Epicastasterone increased the activity of the K(+) efflux conductance in 50% of tested protoplasts while 24-epibrassonolide and 28-homobrassionolide did not modify the plasma membrane currents. Addition of 24-epicastasterone at the cytosolic side (to the pipette solution) resulted in dramatic stimulation of a time-dependent K(+) efflux current (in 30% of protoplasts) and an activation of Ca(2+) influx currents (in 30% of protoplasts). Gadolinium ions, which are blockers of cation channels, inhibited the 24-epicastasterone-induced cation channel activities. In Arabidopsis thaliana plants constitutively expressing aequorin, exogenous 24-epibrassonolide, 28-homobrassionolide and 24-epicastasterone induced a transient elevation of the cytosolic free Ca(2+), which was inhibited by Gd(3+) and mediated by Ca(2+) influx from the bathing solution. In Ca(2+)-aequorin tests, 10 µM of exogenous BRs was the minimal concentration at which statistically significant changes of the cytosolic Ca(2+) were observed. In conclusion, the obtained results suggest that the plasma membrane of root cells contains the brassinosteroid-activated cation-permeable channels, which can probably be involved in rapid regulation of the K(+) homeostasis and Ca(2+) signalling.
Asunto(s)
Brasinoesteroides/metabolismo , Canales de Calcio/metabolismo , Canales de Potasio/metabolismo , Transducción de Señal , Triticum/metabolismoRESUMEN
Electrolyte leakage accompanies plant response to stresses, such as salinity, pathogen attack, drought, heavy metals, hyperthermia, and hypothermia; however, the mechanism and physiological role of this phenomenon have only recently been clarified. Accumulating evidence shows that electrolyte leakage is mainly related to K(+) efflux from plant cells, which is mediated by plasma membrane cation conductances. Recent studies have demonstrated that these conductances include components with different kinetics of activation and cation selectivity. Most probably they are encoded by GORK, SKOR, and annexin genes. Hypothetically, cyclic nucleotide-gated channels and ionotropic glutamate receptors can also be involved. The stress-induced electrolyte leakage is usually accompanied by accumulation of reactive oxygen species (ROS) and often results in programmed cell death (PCD). Recent data strongly suggest that these reactions are linked to each other. ROS have been shown to activate GORK, SKOR, and annexins. ROS-activated K(+) efflux through GORK channels results in dramatic K(+) loss from plant cells, which stimulates proteases and endonucleases, and promotes PCD. This mechanism is likely to trigger plant PCD under severe stress. However, in moderate stress conditions, K(+) efflux could play an essential role as a 'metabolic switch' in anabolic reactions, stimulating catabolic processes and saving 'metabolic' energy for adaptation and repair needs.
Asunto(s)
Apoptosis , Fenómenos Fisiológicos de las Plantas , Plantas/metabolismo , Canales de Potasio/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Equilibrio Hidroelectrolítico , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Plantas/genética , Potasio/metabolismo , Canales de Potasio/genéticaRESUMEN
Reactive oxygen species (ROS) are central to plant stress response, signalling, development and a multitude of other processes. In this study, the plasma-membrane hydroxyl radical (HR)-activated K(+) channel responsible for K(+) efflux from root cells during stress accompanied by ROS generation is characterised. The channel showed 16-pS unitary conductance and was sensitive to Ca(2+), tetraethylammonium, Ba(2+), Cs(+) and free-radical scavengers. The channel was not found in the gork1-1 mutant, which lacks a major plasma-membrane outwardly rectifying K(+) channel. In intact Arabidopsis roots, both HRs and stress induced a dramatic K(+) efflux that was much smaller in gork1-1 plants. Tests with electron paramagnetic resonance spectroscopy showed that NaCl can stimulate HR generation in roots and this might lead to K(+)-channel activation. In animals, activation of K(+)-efflux channels by HRs can trigger programmed cell death (PCD). PCD symptoms in Arabidopsis roots developed much more slowly in gork1-1 and wild-type plants treated with K(+)-channel blockers or HR scavengers. Therefore, similar to animal counterparts, plant HR-activated K(+) channels are also involved in PCD. Overall, this study provides new insight into the regulation of plant cation transport by ROS and demonstrates possible physiological properties of plant HR-activated K(+) channels.