Aquaglyceroporins: Generalized metalloid channels

Background

Aquaporins (AQPs), members of a superfamily of transmembrane channel proteins, are ubiquitous in all domains of life. They fall into a number of branches that can be functionally categorized into two major sub-groups: i) orthodox aquaporins, which are water-specific channels, and ii) aquaglyceroporins, which allow the transport of water, non-polar solutes, such as urea or glycerol, the reactive oxygen species hydrogen peroxide, and gases such as ammonia, carbon dioxide and nitric oxide and, as described in this review, metalloids.

Scope of review

This review summarizes the key findings that AQP channels conduct bidirectional movement of metalloids into and out of cells.

Major conclusions

As(OH)₃ and Sb(OH)₃ behave as inorganic molecular mimics of glycerol, a property that allows their passage through AQP channels. Plant AQPs also allow the passage of boron and silicon as their hydroxyacids, boric acid (B(OH)₃) and orthosilicic acid (Si(OH)₄), respectively. Genetic analysis suggests that germanic acid (GeO₂) is also a substrate. While As(III), Sb(III) and Ge(IV) are toxic metalloids, borate (B(III)) and silicate (Si(IV)) are essential elements in higher plants.

General significance

The uptake of environmental metalloids by aquaporins provides an understanding of (i) how toxic elements such as arsenic enter the food chain; (ii) the delivery of arsenic and antimony containing drugs in the treatment of certain forms of leukemia and chemotherapy of diseases caused by pathogenic protozoa; and (iii) the possibility that food plants such as rice could be made safer by genetically modifying them to exclude arsenic while still accumulating boron and silicon. This article is part of a Special Issue entitled Aquaporins.     

Assessing Methods for Developing Phosphorus Desorption Isotherms from Soils using Anion Exchange Membranes

Developing desorption isotherms for inorganic phosphorus (P) is a time-consuming and non-standardized procedure. Anion exchange membranes (AEMs) have been successfully used in studies of P desorption kinetics and total membrane-desorbable P, but rarely have they been used for developing P desorption isotherms. Our study had two objectives: (1) to evaluate the suitability of using multiple strips of AEMs (termed the Multiple AEM Method) to develop P desorption isotherms; and (2) to compare the Multiple AEM Method with a sequential-extraction approach using AEMs (termed the Sequential AEM Method) to determine if the manner in which AEMs were used would influence the slope of the desorption isotherm, i.e. the partition coefficient. Both methods yielded well-defined, but numerically different desorption isotherms for all levels of sorbed P. However, estimated K _d values among methods were equivalent in the low and medium levels of P sorbed. The Multiple AEM method was quicker than the Sequential AEM method, but both gave similar K _d values in an agriculturally significant range of soil solution concentrations. These methods should be tested on a range of soil type to determine their suitability in developing P desorption isotherms and to move toward method standardization for desorption isotherms.     

Effects of variations in soil water potential,depth of N placement,and cultivar on postanthesis N uptake by wheat

This study was conducted to investigate the influence of soil water potential, depth of N placement, timing, and cultivar on uptake of a small dose of labeled N applied after anthesis by wheat (Triticum aestivum L.) Understanding postanthesis N accumulation should allow better control of grain protein concentration through proper manipulation of inputs. Two hard, red spring-wheat cultivars were planted in early and late fall each yr of a 2-yr field experiment. Less than 1 kg N ha^–1 as K ¹⁵NO₃ was injected into the soil at two depths: shallow (0.05 to 0.08 m) and deep (0.15 to 0.18 m). In both years an irrigation was applied at anthesis, and injections of labeled N were timed 4, 12, and 20 days after anthesis (DAA). Soil water potential was estimated at the time of injection. Mean recovery of ¹⁵N in grain and straw was 57% of the ¹⁵N applied. Recovery did not differ between the high-protein (Yecora Rojo) and the low-protein (Anza or Yolo) cultivars. Mean recovery from deep placement was 60% versus only 54% from shallow placement (p < 0.01). Delaying the time of injection decreased mean recovery significantly from 58% at 4 DAA to 54% at 20 DAA. This decrease was most pronounced in the shallow placement, where soil drying was most severe. Regressions of recovery on soil water potential of individual cultivar x yr x planting x depth treatments were significant only under the driest conditions. Stepwise regression of ¹⁵N recovery on soil water potential and yield parameters using data from all treatments of both years resulted in an equation including soil water potential and N yield, with a multiple correlation coefficient of 0.64. The translocation of ¹⁵N to grain was higher (0.89) than the nitrogen harvest index (0.69), and showed a highly significant increase with increase in DAA. This experiment indicates that the N uptake capacity of wheat remains reasonably constant between 4 and 20 DAA unless soil drying is severe.     

Pathways of glutathione degradation in the yeast Saccharomyces cerevisiae

The degradation of glutathione (GSH) in the yeast Saccharomyces cerevisiae appears to be mediated only by γ-glutamyltranspeptidase and cysteinylglycine dipeptidase. Other enzymes of the γ-glutamyl cycle, γ-glutamyl cyclotransferase and 5-oxo-l-prolinase, are not present in the yeast. In vivo transpeptidation was shown in the presence of a high intracellular level of γ-glutamyltranspeptidase, but only when the de-repressing nitrogen source was a suitable acceptor of the transferase reaction. In contrast, when the de-repressing source was not an acceptor of the transferase reaction (e.g. urea), only glutamate was detected. Intracellular GSH is virtually inert when the level of γ-glutamyltranspeptidase is low. Possible roles for in vivo transpeptidation are discussed.     

Design and in silico modeling of Indoloquinoxaline incorporated keratin nanoparticles for modulation of glucose metabolism in 3T3-L1 adipocytes

The following study was done to assess the glucose utilizing efficiency of Indoloquinoxaline derivative incorporated keratin nanoparticles (NPs) in 3T3-L1 adipocytes. Indoloquinoxaline derivative had wide range of biological activities including antidiabetic activity. In this view, Indoloquinoxaline moiety containing N, N-dimethyl (3-fluoro-6H-indolo [3,2-b] quinoxalin-6-yl) methanamine compound was designed and synthesized, and further it is incorporated into keratin nanoparticles. The formulated NPs, drug entrapment efficiency, releasing capacity, stability, and physicochemical properties were characterized by various spectral analyzer and obtained results of characterizations were confirmed the properties of NPs. The analysis of mechanism underlying the glucose utilization of NPs was examined through molecular docking with identified target, and observed in silico study reports shown strong interaction of NPs in the binding pockets of AMPK and PTP1B. Based on the in silico screening, the formulated NPs was performed for in vitro cellular viability and glucose uptake studies on 3T3-L1 adipocytes. Interestingly, 40 μg of NPs displayed 78.2 ± 2.76% cellular viability, and no cell death was observed at lower concentrations. Further, the concentration dependent glucose utilization was observed at different concentrations of NPs in 3T3-L1 adipocytes. The results of NPs (40 μg) on glucose utilization have revealed eminent result 58.56 ± 4.54% compared to that of Metformin (10 μM) and Insulin (10 μM). The identified results clearly indicated that Indoloquinoxaline derivative incorporated keratin NPs significantly increased glucose utilization efficiency and protect the cells against the insulin resistance.     

Novel method to differentiate 3T3 L1 cells in vitro to produce highly sensitive adipocytes for a GLUT4 mediated glucose uptake using fluorescent glucose analog

Adipocytes play a vital role in glucose metabolism. 3T3 L1 pre adipocytes after differentiation to adipocytes serve as excellent in vitro models and are useful tools in understanding the glucose metabolism. The traditional approaches adopted in pre adipocyte differentiation are lengthy exercises involving the usage of IBMX and Dexamethasone. Any effort to shorten the time of differentiation and quality expression of functional differentiation in 3T3 L1 cells in terms of enhanced Insulin sensitivity has an advantage in the drug discovery process. Thus, there is a need to develop a new effective method of differentiating the pre adipocytes to adipocytes and to use such methods for developing efficacious therapeutic molecules. We observed that a combination of Dexamethasone and Troglitazone generated differentiated adipocytes over fewer days as compared to the combination of IBMX and Dexamethasone which constitutes the standard protocol followed in our laboratory. The experiments conducted to compare the quality of differentiation yielded by various differentiating agents indicated that the lipid droplet accumulation increased by 112 % and the GLUT4 mediated glucose uptake by 137 % in cells differentiated with Troglitazone and Dexamethasone than in cells differentiated traditionally. The comparative studies conducted for evaluating efficient measurable glucose uptake by GOPOD assay, radioactive ³H-2-deoxy-D-glucose assay and by non-radioactive 6-NBDG (fluorescent analog of glucose) indicated that the non-radioactive method using 6-NBDG showed a higher signal to noise ratio than the conventional indirect glucose uptake method (GOPOD assay) and the radioactive ³H-2-deoxy-D-glucose uptake method. Differentiated 3T3 L1 cells when triggered with 2.5 ng/mL of Insulin showed 3.3 fold more glucose uptake in non-radioactive method over the radioactive ³H-2-deoxy-D-glucose uptake method. The results of this study have suggested that a combination of Dexamethasone and Troglitazone for 3T3 L1 cell differentiation helps in better quality differentiation over a short period of time with increased sensitivity to Insulin. The application of these findings for developing new methods of screening novel Insulin mimetics and for evaluating the immunological responses has been discussed.     

Responses of Two Coastal Algae (Skeletonema costatum and Chlorella vulgaris) to Changes in Light and Iron Levels1

Iron (Fe) is essential for phytoplankton growth and photosynthesis, and is proposed to be an important factor regulating algal blooms under replete major nutrients in coastal environments. Here, Skeletonema costatum, a typical red-tide diatom species, and Chlorella vulgaris, a widely distributed Chlorella, were chosen to examine carbon fixation and Fe uptake by coastal algae under dark and light conditions with different Fe levels. The cellular carbon fixation and intracellular Fe uptake were measured via ¹⁴C and ⁵⁵Fe tracer assay, respectively. Cell growth, cell size, and chlorophyll-α concentration were measured to investigate the algal physiological variation in different treatments. Our results showed that cellular Fe uptake proceeds under dark and the uptake rates were comparable to or even higher than those in the light for both algal species. Fe requirements per unit carbon fixation were also higher in the dark resulting in higher Fe: C ratios. During the experimental period, high Fe addition significantly enhanced cellular carbon fixation and Fe uptake. Compared to C. vulgaris, S. costatum was the common dominant bloom species because of its lower Fe demand but higher Fe uptake rate. This study provides some of the first measurements of Fe quotas in coastal phytoplankton cells, and implies that light and Fe concentrations may influence the phytoplankton community succession when blooms occur in coastal ecosystems.     

Enantioselective Uptake and Translocation of a Novel Chiral Neonicotinoid Insecticide Cycloxaprid in Youdonger (Brassica campestris subsp. Chinensis)

For a novel potential commercial chiral pesticide, an independent study on the fate characteristics and residues of each stereoisomer is essential if the application rates for the pesticide and human exposure are to be reduced. The absorption and translocation behavior of a chiral insecticide, cycloxaprid, in plants treated by root immersion and blade smearing was studied using ¹⁴C‐labeling tracer techniques. With the root treatment, total absorption of (1R;8S)‐cycloxaprid (RS) (12.39%) was much greater than that of (1S;8R)‐cycloxaprid (SR) (3.31%) at 192 h after treatment (HAT). The mass concentrations ( RS / SR ) of cycloxaprid in the roots, cotyledons, leaf 1, leaf 2, and leaf 3 were 37.0/16.8, 8.3/2.8, 11.7/6.5, 5.1/4.8, and 8.0/4.7 mg kg^‐1 (fresh weight), respectively, at 192 HAT at an initial concentration 1.6 mg kg^‐1. With the foliar application treatment, no significant difference was observed between the total absorption of RS (3.11%) and SR (4.03%) at the end of the treatment. Both acropetal and basipetal transport of absorbed ¹⁴C occurred and more than 71.83% of absorbed RS and 82.42% of SR remained in the treated leaf. Stereoselective absorption was observed during root uptake but not during foliar absorption. Chirality 25:686–691, 2013. © 2013 Wiley Periodicals, Inc.     

Rhizospheric NO affects N uptake and metabolism in Scots pine (Pinus sylvestris L.) seedlings depending on soil N availability and N source

We investigated the interaction of rhizospheric nitric oxide (NO) concentration (i.e. low, ambient or high) and soil nitrogen (N) availability (i.e. low or high) with organic and inorganic N uptake by fine roots of Pinus sylvestris L. seedlings by ¹⁵N feeding experiments under controlled conditions. N metabolites in fine roots were analysed to link N uptake to N nutrition. NO affected N uptake depending on N source and soil N availability. The suppression of nitrate uptake in the presence of ammonium and glutamine was overruled by high NO. The effects of NO on N uptake with increasing N availability showed different patterns: (1) increasing N uptake regardless of NO concentration (i.e. ammonium); (2) increasing N uptake only with high NO concentration (i.e. nitrate and arginine); and (3) decreasing N uptake (i.e. glutamine). At low N availability and high NO nitrate accumulated in the roots indicating insufficient substrates for nitrate reduction or its storage in root vacuoles. Individual amino acid concentrations were negatively affected with increasing NO (i.e. asparagine and glutamine with low N availability, serine and proline with high N availability). In conclusion, this study provides first evidence that NO affects N uptake and metabolism in a conifer.     

Mechanisms for L-channel-mediated increase in [Ca]i and its reduction by anti-bipolar drugs in cultured astrocytes combined with its mRNA expression in freshly isolated cells support the importance of astrocytic L-channels

The importance of Ca²⁺ signaling in astrocytes is undisputed but a potential role of Ca²⁺ influx via L-channels in the brain in vivo is disputed, although expression of these channels in cultured astrocytes is recognized. This study shows that an increase in free cytosolic Ca²⁺ concentration ([Ca²⁺]_i) in astrocytes in primary cultures in response to an increased extracellular K⁺ concentration (45 mM) is inhibited not only by nifedipine (confirming previous observations) but also to a very large extent by ryanodine, inhibiting ryanodine receptor-mediated release of Ca²⁺, known to occur in response to an elevation in [Ca²⁺]_i. This means that the actual influx of Ca²⁺ is modest, which may contribute to the difficulty in demonstrating L-channel-mediated Ca²⁺ currents in astrocytes in intact brain tissue. Chronic treatment with any of the 3 conventional anti-bipolar drugs lithium, carbamazepine or valproic acid similarly causes a pronounced inhibition of K⁺-mediated increase in [Ca²⁺]_i. This is shown to be due to an inhibition of capacitative Ca²⁺ influx, reflected by decreased mRNA and protein expression of the ‘transient receptor potential channel’ (TRPC1), a constituent of store-operated channels (SOCEs). Literature data are cited (i) showing that depolarization-mediated Ca²⁺ influx in response to an elevated extracellular K⁺ concentration is important for generation of Ca²⁺ oscillations and for the stimulatory effect of elevated K⁺ concentrations in intact, non-cultured brain tissue, and (ii) that Ca²⁺ channel activity is dependent upon availability of metabolic substrates, including glycogen. Finally, expression of mRNA for Ca_v1.3 is demonstrated in freshly separated astrocytes from normal brain.