Transmembrane kinase 1-mediated auxin signal regulates membrane-associated clathrin in Arabidopsis roots.

Clathrin-mediated endocytosis (CME) is the major endocytic pathway in eukaryotic cells that directly regulates abundance of plasma membrane proteins. Clathrin triskelia are composed of clathrin heavy chains (CHCs) and light chains (CLCs), and the phytohormone auxin differentially regulates membrane-associated CLCs and CHCs, modulating the endocytosis and therefore the distribution of auxin efflux transporter PIN-FORMED2 (PIN2). However, the molecular mechanisms by which auxin regulates clathrin are still poorly understood. Transmembrane kinase (TMKs) family proteins are considered to contribute to auxin signaling and plant development; it remains unclear whether they are involved in PIN transport by CME. We assessed TMKs involvement in the regulation of clathrin by auxin, using genetic, pharmacological, and cytological approaches including live-cell imaging and immunofluorescence. In tmk1 mutant seedlings, auxin failed to rapidly regulate abundance of both CHC and CLC and to inhibit PIN2 endocytosis, leading to an impaired asymmetric distribution of PIN2 and therefore auxin. Furthermore, TMK3 and TMK4 were shown not to be involved in regulation of clathrin by auxin. In summary, TMK1 is essential for auxin-regulated clathrin recruitment and CME. TMK1 therefore plays a critical role in the establishment of an asymmetric distribution of PIN2 and an auxin gradient during root gravitropism.

Rice OsCIPK17-OsCBL2/3 module enhances shoot Na+ exclusion and plant salt tolerance in transgenic Arabidopsis.

Soil salinity is detrimental to plant growth and remains a major threat to crop productivity of the world. Plants employ various physiological and molecular mechanisms to maintain growth under salt stress. Identification of genes and genetic loci underlying plant salt tolerance holds the key to breeding salt tolerant crops. CIPK-CBL pathways regulate adaptive responses of plants (especially ion transport) to abiotic stresses via fine-tuned Ca2+ signal transduction. In this study, we showed that over-expression of OsCIPK17 in Arabidopsis enhanced primary root elongation under salt stress, which is in a Ca2+ dependent manner. Further investigation revealed that, under salt stress, OsCIPK17 transcript level was significantly induced and its protein moved from the cytosol to the tonoplast. Using both Y2H and BiFC, tonoplast-localised OsCBL2 and OsCBL3 were shown to interact with OsCIPK17. Interestingly, over-expressing salt-induced OsCBL2 or OsCBL3 in Arabidopsis led to enhanced primary root elongation under salt stress. In this process, OsCIPK17 was shown recruited to the tonoplast (similar to the effect of salt stress). Furthermore, transgenic Arabidopsis lines individually over-expressing OsCIPK17, OsCBL2 and OsCBL3 all demonstrated larger biomass and less Na + accumulation in the shoot under salt stress. All data combined suggest that OsCIPK17- OsCBL2/3 module is a major component of shoot Na+ exclusion and therefore plant salt tolerance, which is through enhanced Na + compartmentation into the vacuole in the root. OsCIPK17 and OsCBL2/3 are therefore potential genetic targets that can be used for delivering salt tolerant rice cultivars.

Co-catalysis of trace dissolved Fe(iii) with biochar in hydrogen peroxide activation for enhanced oxidation of pollutants.

Activation of hydrogen peroxide (H2O2) with biochar is a sustainable and low-cost approach for advanced oxidation of organic pollutants, but faces the challenge of a low yield of hydroxyl radical (˙OH). Herein, we hypothesize that the activation efficiency of H2O2 can be enhanced through co-catalysis of trace dissolved iron (Fe) with biochar. Two biochar samples derived from different feedstock, namely LB from liquor-making residue and WB from wood sawdust, were tested in the co-catalytic systems using trace Fe(iii) (0.3 mg L-1). The cumulative ˙OH production in [Fe(iii) + LB]/H2O2 was measured to be 3.28 times that in LB/H2O2, while the cumulative ˙OH production in [Fe(iii) + WB]/H2O2 was 11.9 times that in WB/H2O2. No extra consumption of H2O2 was observed in LB/H2O2 or WB/H2O2 after addition of trace Fe(iii). Consequently, the reaction rate constants (k obs) for oxidation of pollutants (2,4-dichlorophenoxyacetic acid and sulfamethazine) were enhanced by 3.13-9.16 times. Other iron species including dissolved Fe(ii) and iron minerals showed a similar effect on catalyzing 2,4-D oxidation by biochar/H2O2. The interactions involved in adsorption and reduction of Fe(iii) by biochar in which the defects acted as electron donors and oxygen-containing functional groups bridged the electron transfer. The fast regeneration of Fe(ii) in the co-catalytic system resulted in the sustainable ˙OH production, thus the efficient oxidation of pollutants comparable to other advanced oxidation processes was achieved by using dissolved iron at a concentration as low as the concentration that can be found in natural water.

Characterization and expression of Ailuropoda melanoleuca leptin (ob gene).

Leptin, an adipocyte-derived hormone, plays important roles in metabolism and reproduction. In this article, we report the cloning, expression, and identification of the giant panda leptin (gLeptin) gene and its variants. The gLeptin cDNA was 504 bp long, encoding a precursor peptide of 167 amino acids including 21 residues of signal peptide. A short variant of gLeptin was 501 bp long, encoding a 166-aa peptide and also including a 21-aa signal peptide. Giant panda leptin was 99.4%, 94.6%, and 92.8% identical to that of black bear, dog, and cat, respectively, but was only 81.4% and 80.8% identical to that of human and rat. The cloned gLeptin gene was expressed in Escherichia coli, with expression confirmed by Western blotting and MALDI-TOF-TOF MS PMF. After purification, renaturation, and condensation, the gLeptin protein was injected into Kunming mice. The recombinant gLeptin significantly inhibited food intake by 41.8% and reduced body weight by 5.1% in the mice.

Effect of surfactants on the removal of nitrobenzene by Fe-bearing montmorillonite/Fe(II).

Nonionic and anionic surfactants often occur in anaerobic environments, but their roles in the removal of organic contaminants by Fe-bearing mineral/Fe(II) have not been determined. In this study, batch experiments were performed to investigate the effects of a nonionic surfactant (TX-100) and an anionic surfactant (SDBS) on the removal of nitrobenzene (NB) by Fe-bearing montmorillonite (FM)/Fe(II). Mössbauer spectrum and XPS were applied to analyze the edge surface bound Fe(II) and secondary minerals formed on FM. The contribution of surfactant to the enrichment of NB on FM was studied. The results showed that TX-100 and SDBS had opposite effects on the removal of NB by FM/Fe(II) at neutral pH. The presence of TX-100 improved the removal efficiency of NB from 36.4% to 70.0%, and increased the initial removal rate by 1.7 times. This enhancement effect was mainly attributed to the formation of more active edge surface bound Fe(II) that can reduce more NB to aniline. Formation of more magnetite on FM and selective enrichment of NB on the reactive surface also contributed to the removal of NB. In contrast, the presence of SDBS reduced the amount of edge surface bound Fe(II) via formation of SDBS-Fe(II) complex, which decreased the removal efficiency of NB.

Effects of surfactants on the removal of nitrobenzene by Fe(II) sorbed on goethite.

Surfactants are easily accumulated in groundwater, sediment, and aquifers, due to their excessive use in household, industrial, and agricultural processes. These residual surfactants are expected to influence the transformation and fate of organic contaminants by Fe(II) sorbed on iron oxides in anaerobic environments. Here, we investigated the effects of various surfactants including nonionic TX-100, anionic SDBS and bio-surfactant saponin on the removal of nitrobenzene (NB) by Fe(II) sorbed on goethite (goethite/Fe(II)) through batch experiments. We also elucidated the mechanism behind the effects by XPS, XRD, and determination of the amounts of sorbed Fe(II) on goethite. The results showed that the presence of TX-100 improved NB removal from 77.2% in the absence of surfactant, to 93.8% within 6 h, and improved the removal rate by about 1.3 times. In contrast, the presence of SDBS decreased the removal efficiency to 45.5%, and the presence of saponin nearly inhibited the removal of NB completely. The removed NB was finally nearly reduced to aniline in the absence or presence of surfactants, except in the case of saponin. The amounts of sorbed Fe(II) listed in the sequence as goethite/Fe(II)+TX-100 > goethite/Fe(II) > goethite/Fe(II)+SDBS > goethite/Fe(II)+saponin, and this order negatively correlated with that in the redox potential of these systems. These results confirmed that the presence of surfactants influenced the sorption of Fe(II) on goethite and changed the reactivity of goethite/Fe(II) for NB removal. This finding will promote a clear understanding of the impact of coexisting surfactants on the transformation and fate of organic contaminants in real anaerobic environments.

High-level secretory expression, purification and characterization of Ailuropoda melanoleuca growth hormone in Pichia pastoris.

Growth hormone is one of the most important hormones, which is involved in many reproductive processes of giant panda Ailuropoda melanoleuca. In this study, the mature peptide of A. melanoleuca growth hormone (AmGH) was successfully expressed and secreted in Pichia pastoris under the control of AOX1 promoter. The expression condition for AmGH in P. pastoris, such as the expression time, pH value and methanol concentration in the BMMY were optimized and the AmGH expression level is about 100 mg/L using GS115 recombinant under optimized condition (96 h of 1.5% methanol induction). The secreted nascent AmGH were purified using ammonium sulfate fractionation. The mature AmGH protein exhibited a molecular mass of approximately 22 kDa on SDS-PAGE. This study would provide a new opportunity for large-scale expression and purification of AmGH, which might facilitate studies on the biological activity of AmGH.

Overexpression of the Tibetan Plateau annual wild barley (Hordeum spontaneum) HsCIPKs enhances rice tolerance to heavy metal toxicities and other abiotic stresses.

BACKGROUND: The calcineurin B-like protein (CBL) and CBL-interacting protein kinase (CIPK) signaling system plays a key regulatory role in plant stress signaling. The roles of plant-specific CIPKs, essential for CBL-CIPK functions, in the response to various abiotic stresses have been extensively studied so far. However, until now, the possible roles of the CIPKs in the plant response to heavy metal toxicities are largely unknown. RESULTS: In this study, we used bioinformatic and molecular strategies to isolate 12 HsCIPK genes in Tibetan Plateau annual wild barley (Hordeum spontaneum C. Koch) and subsequently identified their functional roles in the response to heavy metal toxicities. The results showed that multiple HsCIPKs were transcriptionally regulated by heavy metal toxicities (e.g., Hg, Cd, Cr, Pb, and Cu) and other abiotic stresses (e.g., salt, drought, aluminum, low and high temperature, and abscisic acid). Furthermore, the ectopic overexpression of each HsCIPK in rice (Oryza sativa L. cv Nipponbare) showed that transgenic plants of multiple HsCIPKs displayed enhanced tolerance of root growth to heavy metal toxicities (Hg, Cd, Cr, and Cu), salt and drought stresses. These results suggest that HsCIPKs are involved in the response to heavy metal toxicities and other abiotic stresses. CONCLUSIONS: Tibetan Plateau annual wild barley HsCIPKs possess broad applications in genetically engineering of rice with tolerance to heavy metal toxicities and other abiotic stresses.

Influence of Al-oxide on pesticide sorption to woody biochars with different surface areas.

Biochars' properties will change after application in soil due to the interactions with soil constituents, which would then impact the performance of biochars as soil amendment. For a better understanding on these interactions, two woody biochars of different surface areas (SA) were physically treated with aluminum oxide (Al-oxide) to investigate its potential influence on biochars' sorption property. Both the micropore area and mesopore (17∼500 Å in diameter) area of the low-SA biochar were enhanced by at least 1.5 times after treatment with Al-oxide, whereas the same treatment did not change the surface characteristics of the high-SA biochar due partly to its well-developed porosity. The enhanced sorption of the pesticide isoproturon to the Al-oxide-treated low-SA biochar was observed and is positively related to the increased mesopore area. The desorption hysteresis of pesticide from the low-SA biochar was strengthened because of more pesticide molecules entrapped in the expanded pores by Al-oxide. However, no obvious change of pesticide sorption to the high-SA biochar after Al-oxide treatment was observed, corresponding to its unchanged porosity. The results suggest that the influence of Al-oxide on the biochars' sorption property is dependent on their porosity. This study will provide valuable information on the use of biochars for reducing the bioavailability of pesticides.