Futile plasma membrane cycling of ammonium (NH4+) is characteristic of low-affinity NH4+ transport, and has been proposed to be a critical factor in NH4+ toxicity. Using unidirectional flux analysis with the positron-emitting tracer 13N in intact seedlings of barley (Hordeum vulgare L.), it is shown that rapid, futile NH4+ cycling is alleviated by elevated K+ supply, and that low-affinity NH4+ transport is mediated by a K+-sensitive component, and by a second component that is independent of K+. At low external [K+] (0.1 mM), NH4+ influx (at an external [NH4+] of 10 mM) of 92 micromol g(-1) h(-1) was observed, with an efflux:influx ratio of 0.75, indicative of rapid, futile NH4+ cycling. Elevating K+ supply into the low-affinity K+ transport range (1.5-40 mM) reduced both influx and efflux of NH4+ by as much as 75%, and substantially reduced the efflux:influx ratio. The reduction of NH4+ fluxes was achieved rapidly upon exposure to elevated K+, within 1 min for influx and within 5 min for efflux. The channel inhibitor La3+ decreased high-capacity NH4+ influx only at low K+ concentrations, suggesting that the K+-sensitive component of NH4+ influx may be mediated by non-selective cation channels. Using respiratory measurements and current models of ion flux energetics, the energy cost of concomitant NH4+ and K+ transport at the root plasma membrane, and its consequences for plant growth are discussed. The study presents the first demonstration of the parallel operation of K+-sensitive and -insensitive NH4+ flux mechanisms in plants.
Current models of potassium acquisition and cytochemical processes in plants assume that potassium concentrations in the cytosol ([K + ] cyt ) are maintained homeostatically at approximately 100 mM. Here, we use 42 K radiotracer data in the model plant species Hordeum vulgare L. (barley) to show that this assumption is incorrect. Our study reveals that [K + ] cyt in root cells of intact barley seedlings is held at a minimum of two physiological set points, coinciding with two fundamentally distinct modes of K + transport, each of which is characterized by a unique network of fluxes to and from the cytosol, and reflects variations in mechanisms and energetics of K + transport, cytosolic K + turnover, flux partitioning, and sensitivity to NH þ 4 . Increased external potassium or ammonium concentrations caused a substantial drop in [K + ] cyt , as well as a switch from a transport mode dominated by high-affinity, energydependent, influx to a mode dominated by channelmediated fluxes in both directions across the plasma membrane. Our study provides the first subcellular demonstration of the flexibility, rather than strict homeostasis, of cellular K + maintenance, and of the dynamic interaction between plant membrane fluxes of the two major nutrient cations K + and NH þ 4 .
Using the short-lived radiotracer 42K+, we present a comprehensive subcellular flux analysis of low-affinity K+ transport in plants. We overturn the paradigm of cytosolic K+ pool-size homeostasis and demonstrate that low-affinity K+ transport is characterized by futile cycling of K+ at the plasma membrane. Using two methods of compartmental analysis in intact seedlings of barley (Hordeum vulgare L. cv Klondike), we present data for steady-state unidirectional influx, efflux, net flux, cytosolic pool size, and exchange kinetics, and show that, with increasing external [K+] ([K+]ext), both influx and efflux increase dramatically, and that the ratio of efflux to influx exceeds 70% at [K+]ext ≥ 20 mm. Increasing [K+]ext, furthermore, leads to a shortening of the half-time for cytosolic K+ exchange, to values 2 to 3 times lower than are characteristic of high-affinity transport. Cytosolic K+ concentrations are shown to vary between 40 and 200 mm, depending on [K+]ext, on nitrogen treatment (NO3− or NH4+), and on the dominant mode of transport (high- or low-affinity transport), illustrating the dynamic nature of the cytosolic K+ pool, rather than its homeostatic maintenance. Based on measurements of trans-plasma membrane electrical potential, estimates of cytosolic K+ pool size, and the magnitude of unidirectional K+ fluxes, we describe efflux as the most energetically demanding of the cellular K+ fluxes that constitute low-affinity transport.
Grafting has been used in agriculture for over 2000 years. Disease resistance and environmental tolerance are highly beneficial traits that can be provided through use of grafting, although the mechanisms, in particular for resistance, have frequently been unknown. As information emerges that describes plant disease resistance mechanisms, the proteins, and nucleic acids that play a critical role in disease management can be expressed in genetically engineered (GE) plant lines. Utilizing transgrafting, the combination of a GE rootstock with a wild-type (WT) scion, or the reverse, has the potential to provide pest and pathogen resistance, impart biotic and abiotic stress tolerance, or increase plant vigor and productivity. Of central importance to these potential benefits is the question of to what extent nucleic acids and proteins are transmitted across a graft junction and whether the movement of these molecules will affect the efficacy of the transgrafting approach. Using a variety of specific examples, this review will report on the movement of organellar DNA, RNAs, and proteins across graft unions. Attention will be specifically drawn to the use of small RNAs and gene silencing within transgrafted plants, with a particular focus on pathogen resistance. The use of GE rootstocks or scions has the potential to extend the horticultural utility of grafting by combining this ancient technique with the molecular strategies of the modern era.
The disruption of K+ transport and accumulation is symptomatic of NH4+ toxicity in plants. In this study, the influence of K+ supply (0.02–40 mM) and nitrogen source (10 mM NH4+ or NO3–) on root plasma membrane K+ fluxes and cytosolic K+ pools, plant growth, and whole-plant K+ distribution in the NH4+-tolerant plant species rice (Oryza sativa L.) was examined. Using the radiotracer 42K+, tissue mineral analysis, and growth data, it is shown that rice is affected by NH4+ toxicity under high-affinity K+ transport conditions. Substantial recovery of growth was seen as [K+]ext was increased from 0.02 mM to 0.1 mM, and, at 1.5 mM, growth was superior on NH4+. Growth recovery at these concentrations was accompanied by greater influx of K+ into root cells, translocation of K+ to the shoot, and tissue K+. Elevating the K+ supply also resulted in a significant reduction of NH4+ influx, as measured by 13N radiotracing. In the low-affinity K+ transport range, NH4+ stimulated K+ influx relative to NO3– controls. It is concluded that rice, despite its well-known tolerance to NH4+, nevertheless displays considerable growth suppression and disruption of K+ homeostasis under this N regime at low [K+]ext, but displays efficient recovery from NH4+ inhibition, and indeed a stimulation of K+ acquisition, when [K+]ext is increased in the presence of NH4+.
ABSTRACT, from a similar baseline, and to a similar extent, at both low and high K + supply. We compare our results to those of other groups, and conclude that the maintenance of the cytosolic Na + : K + ratio is not central to plant survival under NaCl stress. We offer alternative explanations for sodium sensitivity in relation to the primary acquisition mechanisms of Na + and K + .
The interaction of sodium and potassium ions in the context of the primary entry of Na+ into plant cells, and the subsequent development of sodium toxicity, has been the subject of much recent attention. In the present study, the technique of compartmental analysis with the radiotracers 42K+ and 24Na+ was applied in intact seedlings of barley (Hordeum vulgare L.) to test the hypothesis that elevated levels of K+ in the growth medium will reduce both rapid, futile Na+ cycling at the plasma membrane, and Na+ build-up in the cytosol of root cells, under saline conditions (100 mM NaCl). We reject this hypothesis, showing that, over a wide (400-fold) range of K+ supply, K+ neither reduces the primary fluxes of Na+ at the root plasma membrane nor suppresses Na+ accumulation in the cytosol. By contrast, 100 mM NaCl suppressed the cytosolic K+ pool by 47–73%, and also substantially decreased low-affinity K+ transport across the plasma membrane. We confirm that the cytosolic [K+]:[Na+] ratio is a poor predictor of growth performance under saline conditions, while a good correlation is seen between growth and the tissue ratios of the two ions. The data provide insight into the mechanisms that mediate the toxic influx of sodium across the root plasma membrane under salinity stress, demonstrating that, in the glycophyte barley, K+ and Na+ are unlikely to share a common low-affinity pathway for entry into the plant cell.
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