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111.
Structure,function, and expression pattern of a novel sodium-coupled citrate transporter (NaCT) cloned from mammalian brain 总被引:8,自引:0,他引:8
Inoue K Zhuang L Maddox DM Smith SB Ganapathy V 《The Journal of biological chemistry》2002,277(42):39469-39476
Citrate plays a pivotal role not only in the generation of metabolic energy but also in the synthesis of fatty acids, isoprenoids, and cholesterol in mammalian cells. Plasma levels of citrate are the highest ( approximately 135 microm) among the intermediates of the tricarboxylic acid cycle. Here we report on the cloning and functional characterization of a plasma membrane transporter (NaCT for Na+ -coupled citrate transporter) from rat brain that mediates uphill cellular uptake of citrate coupled to an electrochemical Na+ gradient. NaCT consists of 572 amino acids and exhibits structural similarity to the members of the Na+-dicarboxylate cotransporter/Na+ -sulfate cotransporter (NaDC/NaSi) gene family including the recently identified Drosophila Indy. In rat, the expression of NaCT is restricted to liver, testis, and brain. When expressed heterologously in mammalian cells, rat NaCT mediates the transport of citrate with high affinity (Michaelis-Menten constant, approximately 20 microm) and with a Na+:citrate stoichiometry of 4:1. The transporter does interact with other dicarboxylates and tricarboxylates but with considerably lower affinity. In mouse brain, the expression of NaCT mRNA is evident in the cerebral cortex, cerebellum, hippocampus, and olfactory bulb. NaCT represents the first transporter to be identified in mammalian cells that shows preference for citrate over dicarboxylates. This transporter is likely to play an important role in the cellular utilization of citrate in blood for the synthesis of fatty acids and cholesterol (liver) and for the generation of energy (liver and brain). NaCT thus constitutes a potential therapeutic target for the control of body weight, cholesterol levels, and energy homeostasis. 相似文献
112.
Lanni A Mancini FP Sabatino L Silvestri E Franco R De Rosa G Goglia F Colantuoni V 《FEBS letters》2002,517(1-3):7-12
Clavaminate synthase (CAS), a 2-oxoglutarate (2OG) dependent dioxygenase, catalyses three steps in the biosynthesis of clavulanic acid. Crystals of CAS complexed with Fe(II), 2OG and deoxyguanidinoproclavaminate were exposed to nitric oxide (NO) acting as a dioxygen analogue. Prior to exposure with NO, the active site Fe(II) is octahedrally coordinated by a water molecule, the 2-oxo and 1-carboxylate groups of 2OG, and the side-chains of an aspartyl and two histidinyl residues. NO binds to the position previously occupied by the 2OG 1-carboxylate concomitant with rearrangement of the latter to the position previously occupied by the displaced water. 相似文献
113.
Recent studies demonstrate a role for intracellular oxidation in the regulation of neutral sphingomyelinase (N-SMase). Glutathione (GSH) has been shown to regulate N-SMase in vitro and in cells. However, it has not been established whether the effects of GSH in cells are due to direct action on N-SMase. In this study, treatment of human mammary carcinoma MCF-7 cells with diamide, a thiol-depleting agent, caused a decrease in intracellular GSH and degradation of sphingomyelin (SM) to ceramide. The SM pool hydrolyzed in response to diamide belonged to the bacterial SMase-resistant pool of SM. Importantly, pretreatment of MCF-7 cells with GSH, N-acetylcysteine, an antioxidant, or GW69A, a specific N-SMase inhibitor, prevented diamide-induced degradation of SM to ceramide, suggesting that intracellular levels of GSH regulate the extent to which SM is degraded to ceramide and that this probably involves a GW69A-sensitive N-SMase. Unexpectedly, expression of Bcl-xL prevented tumor necrosis factor--induced SM hydrolysis and ceramide accumulation but not the decrease in intracellular GSH. Furthermore, Bcl-xL inhibited diamide-induced SM hydrolysis and ceramide accumulation but not the decrease in intracellular GSH. These results suggest that the site of action of Bcl-xL is downstream of GSH depletion and upstream of ceramide accumulation, and that GSH probably does not exert direct physiologic effects on N-SMase. 相似文献
114.
Aronson MN Complainville A Clérot D Alcalde H Katul L Vetten HJ Gronenborn B Timchenko T 《The Plant journal : for cell and molecular biology》2002,31(6):767-775
The multipartite genome of the nanovirus Faba bean necrotic yellows virus, which consists of one gene on each DNA component, was exploited to construct a series of virus-based episomal vectors designed for transient replication and gene expression in plants. This nanovirus based expression system yields high levels of protein which allows isolation of recombinant protein and protein complexes from plant tissues. As examples, we demonstrated in planta interaction between the nanovirus F-box protein Clink and SKP1, a constituent of the ubiquitin-dependent protein turnover pathway. Thus, replicative nanovirus vectors provide a simple and efficient means for in planta characterization of protein-protein interaction. 相似文献
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The efficiency of gene transfection mediated by nonviral vectors is limited because of nonoptimal intracellular trafficking of transfecting DNA. Most nonviral vectors deliver transfecting DNA into a cell through endocytosis. However, poor escape from endocytic vesicles and inefficient transport of DNA into the nucleus often limits a success of gene transfection. Photochemical transfection is a new method, based on light-induced permeabilisation of endocytic vesicles, liberating transfecting DNA into the cytosol, concurrently increasing the chances for DNA to enter the nucleus.The aim of this study was to investigate the role of the cell cycle for the efficiency of photochemical transfection. It was demonstrated that in asynchronous human colon carcinoma HCT 116 cells photochemical treatment increased the transfection mediated by the nonviral vectors, the cationic polypeptide polylysine and the cationic lipid N-(2-aminoethyl)-N,N-dimethyl-2,3-bis(tetradecyloxy)-1-propanaminium bromide/dioleoylphosphatidylethanolamine (beta AE-DMRIE/DOPE), by 30- and 2.5-fold, respectively. In aphidicolin-synchronised cells, photochemical transfection mediated by polylysine was dependent on the cell cycle: transfection level was 4-fold higher when illumination, inducing photochemical reactions, was performed during the G2/M phase as compared to the G1/early-S phase. The cell cycle influence on photochemical transfection mediated by beta AE-DMRIE/DOPE was very low: only 20% difference between G2/M and the G1/S phase was observed. We suggest that transgenes, photochemically liberated close/during mitosis, perhaps have the highest opportunity to enter the nucleus and be expressed. However, the dependence of photochemical transfection on the cell cycle might be partially disguised by various factors induced by photochemical treatment. 相似文献
118.
Natural<Superscript>15</Superscript> N Abundance of Plants and Soil N in a Temperate Coniferous Forest 总被引:3,自引:0,他引:3
Keisuke?Kobakkoba@depe.titech.ac.jp" title="kkoba@i.kyoto-u.ac.jp kkoba@depe.titech.ac.jp" itemprop="email" data-track="click" data-track-action="Email author" data-track-label="">Email author Muneto?Hirobe Lina?Koyama Ayato?Kohzu Naoko?Tokuchi Knute John?Nadelhoffer Eitaro?Wada Hiroshi?Takeda 《Ecosystems》2003,6(5):457-469
Measurement of nitrogen isotopic composition (15N) of plants and soil nitrogen might allow the characteristics of N transformation in an ecosystem to be detected. We tested the measurement of 15N for its ability to provide a picture of N dynamics at the ecosystem level by doing a simple comparison of 15N between soil N pools and plants, and by using an existing model. 15N of plants and soil N was measured together with foliar nitrate reductase activity (NRA) and the foliar NO3– pool at two sites with different nitrification rates in a temperature forest in Japan. 15N of plants was similar to that of soil NO3– in the high-nitrification site. Because of high foliar NRA and the large foliar NO3– pool at this site, we concluded that plant 15N indicated a great reliance of plants on soil NO3– there. However, many 15N of soil N overlapped each other at the other site, and 15N could not provide definitive evidence of the N source. The existing model was verified by measured 15N of soil inorganic N and it explained the variations of plant 15N between the two sites in the context of relative importance of nitrification, but more information about isotopic fractionations during plant N uptake is required for quantitative discussions about the plant N source. The model applied here can provide a basis to compare 15N signatures from different ecosystems and to understand N dynamics. 相似文献
119.
Ruso JM Besada L Martínez-Landeira P Seoane L Prieto G Sarmiento F 《Journal of liposome research》2003,13(2):131-145
An investigation on the dependence of electrophoretic mobilities of unilamellar vesicles of phosphatidylcholine-cholesterol-phosphatidylinositol (PC-Chol-PI) on the concentration of several cations with variations in the relation charge/radius in the range Na+, K+, Cs+, Mg2+, Ca2+, Ba2+, Al3+, and La3+ has been realized. Plots of zeta potential against ion concentration exhibit a maximum for all the cations under study, the position of the maximum is greatly affected by the charge of the ion. From the feature of these plots two phenomenon were observed: an initial binding of cations into the slipping plane for ion concentration below the maximum and a phenomenon of vesicle association for concentration above the maximum. To confirm these observations measurements on dynamic light scattering were performed to obtain the corresponding size distribution of the liposomes at different ion concentrations. Finally the ability of the Stern isotherm to describe the adsorption of the cations to vesicles was tested by two methods. The two main parameters of the theory: the total number of adsorption sites per unit area, N1, and the equilibrium constant, K; (and consequently the free energy of adsorption, deltaG0ads) were calculated for the different ions, showing good agreement. The equilibrium constants of adsorption have been found to obey a linear relationship with ion radius the slope of which decreases with the ion charge. 相似文献
120.
Previously we reported that ultralow concentrations of dynorphins (10(-16) to 10(-12) M) inhibited lipopolysaccharide (LPS)-induced production of nitric oxide (NO) and proinflammatory cytokines in mouse glia without the participation of kappa-opioid receptors. In the current study using mouse cortical neuron-glia cocultures, we examined the possibility that inhibition of glia inflammatory response by dynorphins might be neuroprotective for neurons. LPS, in a concentration-dependent manner, markedly increased the release of lactate dehydrogenase (LDH), an indicator of cellular injury. Ultralow concentrations (10(-14) to 10(-12) M) of dynorphin (dyn) A-(1-8) significantly prevented the LPS-induced release of LDH, loss of neurons, and changes in cell morphology, in addition to inhibition of LPS-induced nitrite production. Meanwhile, ultralow concentrations (10(-15) to 10(-13) M) of des-[Tyr(1)]-dyn A-(2-17), a nonopioid peptide which does not bind to kappa-opioid receptors, exhibited the same inhibitory effect as dyn A-(1-17). These results suggest that dynorphins at ultralow concentrations are capable of reducing LPS-induced neuronal injury and these neuroprotective effects of dynorphins are not mediated by classical opioid receptors. 相似文献