High-affinity System Research Articles

Aft1 Nuclear Localization and Transcriptional Response to Iron Starvation Rely upon TORC2/Ypk1 Signaling and Sphingolipid Biosynthesis.

Iron scarcity provokes a cellular response consisting of the strong expression of high-affinity systems to optimize iron uptake and mobilization. Aft1 is a primary transcription factor involved in iron homeostasis and controls the expression of high-affinity iron uptake genes in Saccharomyces cerevisiae. Aft1 responds to iron deprivation by translocating from the cytoplasm to the nucleus. Here, we demonstrate that the AGC kinase Ypk1, as well as its upstream regulator TOR Complex 2 (TORC2), are required for proper Aft1 nuclear localization following iron deprivation. We exclude a role for TOR Complex 1 (TORC1) and its downstream effector Sch9, suggesting this response is specific for the TORC2 arm of the TOR pathway. Remarkably, we demonstrate that Aft1 nuclear localization and a robust transcriptional response to iron starvation also require biosynthesis of sphingolipids, including complex sphingolipids such as inositol phosphorylceramide (IPC) and upstream precursors, e.g., long-chain bases (LCBs) and ceramides. Furthermore, we observe the deficiency of Aft1 nuclear localization and impaired transcriptional response in the absence of iron when TORC2-Ypk1 is impaired is partially suppressed by exogenous addition of the LCB dihydrosphingosine (DHS). This latter result is consistent with prior studies linking sphingolipid biosynthesis to TORC2-Ypk1 signaling. Taken together, these results reveal a novel role for sphingolipids, controlled by TORC2-Ypk1, for proper localization and activity of Aft1 in response to iron scarcity.

Kinetic modulation of bacterial hydrolases by microbial community structure in coastal waters.

In this study, we hypothesized that shifts in the kinetic parameters of extracellular hydrolytic enzymes may occur as a consequence of seasonal environmental disturbances and would reflect the level of adaptation of the bacterial community to the organic matter of the ecosystem. We measured the activities of enzymes that play a key role in the bacterial growth (leucine aminopeptidase, β- and α-glucosidases) in surface coastal waters of the Eastern Cantabrian Sea and determined their kinetic parameters by computing kinetic models of distinct complexity. Our results revealed the existence of two clearly distinct enzymatic systems operating at different substrate concentrations: a high-affinity system prevailing at low substrate concentrations and a low-affinity system characteristic of high substrate concentrations. These findings could be the result of distinct functional bacterial assemblages growing concurrently under sharp gradients of high-molecular-weight compounds. We constructed an ecological network based on contemporaneous and time-delayed correlations to explore the associations between the kinetic parameters and the environmental variables. The analysis revealed that the recurring phytoplankton blooms registered throughout the seasonal cycle trigger the wax and wane of those members of the bacterial community able to synthesize and secrete specific enzymes.

Genome-wide identification and in silico analysis of NPF, NRT2, CLC and SLAC1/SLAH nitrate transporters in hexaploid wheat (Triticum aestivum)

Nitrogen transport is one of the most important processes in plants mediated by specialized transmembrane proteins. Plants have two main systems for nitrogen uptake from soil and its transport within the system—a low-affinity transport system and a high-affinity transport system. Nitrate transporters are of special interest in cereal crops because large amount of money is spent on N fertilizers every year to enhance the crop productivity. Till date four gene families of nitrate transporter proteins; NPF (nitrate transporter 1/peptide transporter family), NRT2 (nitrate transporter 2 family), the CLC (chloride channel family), and the SLAC/SLAH (slow anion channel-associated homologues) have been reported in plants. In our study, in silico mining of nitrate transporter genes along with their detailed structure, phylogenetic and expression analysis was carried out. A total of 412 nitrate transporter genes were identified in hexaploid wheat genome using HMMER based homology searches in IWGSC Refseq v2.0. Out of those twenty genes were root specific, 11 leaf/shoot specific and 17 genes were grain/spike specific. The identification of nitrate transporter genes in the close proximity to the previously identified 67 marker-traits associations associated with the nitrogen use efficiency related traits in nested synthetic hexaploid wheat introgression library indicated the robustness of the reported transporter genes. The detailed crosstalk between the genome and proteome and the validation of identified putative candidate genes through expression and gene editing studies may lay down the foundation to improve nitrogen use efficiency of cereal crops.

Regulation of Copper Metabolism by Nitrogen Utilization in Saccharomyces cerevisiae.

To understand the relationship between carbon or nitrogen utilization and iron homeostasis, we performed an iron uptake assay with several deletion mutants with partial defects in carbon or nitrogen metabolism. Among them, some deletion mutants defective in carbon metabolism partially and the MEP2 deletion mutant showed lower iron uptake activity than the wild type. Mep2 is known as a high-affinity ammonia transporter in Saccharomyces cerevisiae. Interestingly, we found that nitrogen starvation resulted in lower iron uptake activity than that of wild-type cells without downregulation of the genes involved in the high-affinity iron uptake system FET3/FTR1. However, the gene expression of FRE1 and CTR1 was downregulated by nitrogen starvation. The protein level of Ctr1 was also decreased by nitrogen starvation, and addition of copper to the nitrogen starvation medium partially restored iron uptake activity. However, the expression of MAC1, which is a copper-responsive transcriptional activator, was not downregulated by nitrogen starvation at the transcriptional level but was highly downregulated at the translational level. Mac1 was downregulated dramatically under nitrogen starvation, and treatment with MG132, which is an inhibitor of proteasome-dependent protein degradation, partially attenuated the downregulation of Mac1. Taken together, these results suggest that nitrogen starvation downregulates the high-affinity iron uptake system by degrading Mac1 in a proteasome-dependent manner and eventually downregulates copper metabolism.

Influence of Iron on Physiological Parameters and Intracellular Microcystin in Microcystis Panniformis Strain Isolated from a Reservoir in the Amazon.

In the Amazon, the leaching from soil left unprotected by deforestation increases the entry of iron, among other elements, in aquatic ecosystems, which can cause cyanobacterial blooms. This study aimed to investigate the physiological response of a strain of Microcystis panniformis to iron variation. The strain was isolated from a reservoir located in the Western Amazon and produces microcystin-LR. After a period of iron deprivation, the cultures were submitted to three conditions: control (223 μgFe.L-1), treatment with 23 μgFe.L-1, and absence of iron. At regular intervals for eight days, the cell density, levels of chlorophyll a and microcystins were determined. On the second and fourth day, transcription of genes responsive to iron limitation was quantified. Starting on the fourth day of the experiment, the different iron concentrations affected growth, and on the eighth day in the iron-free condition cell density was 90% lower than in control. Chlorophyll cell quota in 23 μgFe.L-1 and control presented similar values, while without iron the cells became chlorotic as of the fourth day Toxin concentration in cells grow in 0 μgFe.L-1 in relation to the control. Higher transcription levels of the feo and fut genes were observed in the 0 μgFe.L-1 and 23 μgFe.L-1 treatments, indicating that the cells were activating high-affinity capture systems to reestablish an adequate concentration of intracellular iron. The increasing deforestation in the Jamari River Basin (Amazon region), can contribute to the occurrence of toxic cyanobacterial blooms due to the greater entrance of iron in water bodies.

NRT2.1, a major contributor to cadmium uptake controlled by high-affinity nitrate transporters

Management of nitrogen fertilizer is a good strategy for controlling cadmium (Cd) accumulation in plants. Some progress has already been made but much remains to be done. Here, we show that mutants with loss of function of nitrate transporter1.1 (NRT1.1) or nitrate transporter2.1 (NRT2.1) had lower Cd concentrations than wild-type plants under low-nitrate conditions. However, this was eliminated when plants were cultivated in nitrate-free medium or supplied with Cd and nitrate alternately. These findings indicate that inhibition of NRT1.1 or NRT2.1 activity reduces Cd accumulation in plants, and depends on the presence of nitrate. The results showing that nrt2.1–2 mutants had the lowest Cd concentrations compared with Col-0, nrt1.1 and nrt2.4 plants, proves that NRT2.1 is the major contributor to Cd uptake controlled by nitrate high-affinity transporters. NRT2.1 acts as the major contributor to nitrate uptake under Cd stress in low-nitrate conditions, and contributes about 50% to nitrate uptake, while NRT1.1 contributes only 10%, and little is known regarding the role of NRT2.2 and NRT2.4 on nitrate uptake in medium with 200 μM nitrate. Positive correlations between nitrate uptake and Cd concentration in plants were also observed. Collectively, NRT2.1 acts as the major contributor to Cd uptake by controlling nitrate uptake in nitrate high-affinity systems.

Structural characterization of fondaparinux interaction with per-6-amino-beta-cyclodextrin: An NMR and MS study

The highly anionic synthetic pentasaccharide fondaparinux (FDPX) – representing the antithrombin binding sequence of heparin – is in clinical use as a potent anticoagulant. Contrary to the unfractionated heparin, FDPX lacks potent antidote completely reversing its anticoagulant activity, therefore it is of great importance to identify new structures exhibiting strong intermolecular interactions towards FDPX. The polycationic heptakis(6-amino-6-deoxy)-beta-cyclodextrin (NH2-β-CD) can serve as an excellent model compound to mimic these interactions between the oppositely charged oligosaccharides. Herein, extensive NMR spectroscopic and nano-electrospray ionization mass spectrometric (nESI-MS) studies were conducted to understand the molecular-level interactions in the FDPX - NH2-β-CD systems. NMR experiments were performed at pD 7.4 and 2.0. Job’s method of continuous variation and 1H NMR titration experiments suggested the formation of FDPX∙NH2-β-CD complex at pD 7.4, while the presence of multiple complexes was assumed at pD 2.0. Stability constants were determined by separate 1H NMR titrations, yielding log β11=3.65 ± 0.02 at pD 7.4, while log β11 ≥ 4.9 value suggested a high-affinity system at pD 2.0. 2D NOESY NMR studies indicated spatial proximities between the anomeric resonance α-l-iduronic acid residue and the cyclodextrin’s methylene unit in the proximity of the cationic amino function. Acidic degradation of FDPX was investigated by NMR and MS for the first time in detail confirming that desulfation occurs involving one to two sulfate moieties. The desulfation of FDPX was inhibited by the cationic cyclodextrin in the case of equimolar ratio at pD 2.0. This is the first report on the stabilizing effect of cyclodextrin complexation on heparin degradation.

Elevated CO2 differentially regulates root nitrate transporter kinetics in a genotype and nitrate dose-dependent manner

The nitrogen (N) and protein concentration of wheat crop and grain often decline as a result of exposure of the crop to elevated CO2 (EC). In our earlier studies, it was found that the exacerbated production of nitric oxide (NO) represses the transcription of nitrate reductase (NR) and high affinity nitrate transporters (HATS) in EC grown wheat seedlings receiving high N. High N supply under EC also resulted in accumulation of reactive oxygen species (ROS), and reactive nitrogen species (RNS; NO and S- nitrosothiols) ensuing faster senescence and reduced N metabolite concentration in wheat. In this study, the effect of short-term exposure to EC on nitrate uptake kinetics was studied. The impact of EC on constitutive and inducible components of high affinity and low affinity nitrate uptake systems (HATS and LATS) were delineated in two wheat genotypes diverse in terms of nitrate uptake and assimilation capacities. Nitrate dose-response of NR was suppressed by EC in both leaf and root tissues. Plants grown under EC displayed a marked reduction in nitrate uptake kinetic components of LATS. Wheat genotype with high leaf nitrate assimilation capacity was able to maintain considerably higher nitrate uptake rate under EC albeit at a lower rate in comparison to ambient CO2. Wheat leaves exposed to EC displayed a comparatively higher abundance of NO and showed incremental abundance depending on increase in nitrate supply. Exogenous NO supply significantly suppressed the nitrate uptake rate of EC grown plants. Hence, EC-induced production of NO downregulates LATS kinetics in a genotype and nitrate dose-dependent manner.

Modulating the mechanokinetics of spontaneous contractions of the myometrium of rats using calix[4]arene C-90 – plasma membrane calcium ATPase inhibitor

Introduction. Plasma membrane calcium ATPase is a constitutive structure of cells that functions as a high affinity system of releasing Са2+ ions from the cytoplasm and ensures a long-term maintenance of the basal concentration of these cations in the state of dormancy. Currently, there are no satisfactory means for the pharmacological correction of plasma membrane calcium ATPase function. Thus, elaboration, synthesis, and study of substances with the targeted impact on plasma membrane calcium ATPase are topical issues. Previously, we determined the ability of this calix[4]arene in the concentration of 10 µM to inhibit the contractive activity and to slow down the relaxation of smooth muscle of the myometrium in the NO-dependent way, which considerably decreases the normalized maximal velocity of the relaxation phase. Materials and Methods. The tenzometric methods and mechanokinetic analysis were used to investigate the impact of the cumulative increase in the concentration of calix[4]arene С-90 (10 nM – 100 µM) on the spontaneous contractive activity of the myometrium of rats. The complete profile of spontaneous cycles of contractions-relaxa­tions was studied by the empirical multiparameter method of complex mechanokinetic analysis, elaborated by us (with the consideration of the parameters of time (τ0, τC and τR), force (Fmax, FC and FR), velocity (VC and VR), and impulse (Іmax, ІC and ІR). Results. Calix[4]arene C-90 evoked the dose-dependent inhibition of spontaneous contractive activity of the myometrium preparations. Its high concentrations caused a change in the structure of the contraction act, such as an increase in the duration of the contraction phase, while the duration of the relaxation phase did not show any changes. The multiparameter method of the complex mechanokinetic analysis demonstrated that in the whole range of the investigated concentrations, substance С-90 considerably decreases the indices of force parameters (Fmax, FC and FR) and the values of impulses of force Іmax, IC and IR of the spontaneous contractions of the myometrium. On the background of all the applied concentrations (10-7–10-4 М), calix[4]arene С-90 conditioned the slowing down of the relaxation of spontaneous contractions in uterine muscle prepa­rations of rats, which was reflected in a reliable decrease in the parameter for the maxi­mal velocity of the relaxation phase (VR). Conclusions. The results of the study demonstrate that calix[4]arene С-90 inhibits the processes of Са2+ extrusion from smooth muscle cells myoplasm, probably, impacting plasma membrane calcium ATPase molecules directly. It is noteworthy that C-90 is also likely to inhibit the processes of the intake of these cations to cells from the extracellular medium, causing a decrease in the velocity of force intensification during the contraction phase, and reducing the frequency and force of the spontaneous contractions in the myometrium.

Co-Overexpression of OsNAR2.1 and OsNRT2.3a Increased Agronomic Nitrogen Use Efficiency in Transgenic Rice Plants.

The NO3 - transporter plays an important role in rice nitrogen acquisition and nitrogen-use efficiency. Our previous studies have shown that the high affinity systems for nitrate uptake in rice is mediated by a two-component NRT2/NAR2 transport system. In this study, transgenic plants were successful developed by overexpression of OsNAR2.1 alone, OsNRT2.3a alone and co-overexpression of OsNAR2.1 and OsNRT2.3a. Our field experiments indicated that transgenic lines expressing p35S:OsNAR2.1 or p35S:OsNAR2.1-p35S:OsNRT2.3a constructs exhibited increased grain yields of approximately 14.1% and 24.6% compared with wild-type (cv. Wuyunjing 7, WT) plants, and the agricultural nitrogen use efficiency increased by 15.8% and 28.6%, respectively. Compared with WT, the 15N influx in roots of p35S:OsNAR2.1 and p35S: OsNAR2.1-p35S:OsNRT2.3a lines increased 18.9%‑27.8% in response to 0.2 mM, 2.5 mM 15NO3 –, and 1.25 mM 15NH4 15NO3, while there was no significant difference between p35S:OsNAR2.1 and p35S:OsNAR2.1-p35S:OsNRT2.3a lines; only the 15N distribution ratio of shoot to root for p35S:OsNAR2.1-p35S:OsNRT2.3a lines increased significantly. However, there were no significant differences in nitrogen use efficiency, 15N influx in roots and the yield between the p35S:NRT2.3a transgenic lines and WT. This study indicated that co-overexpression of OsNAR2.1 and OsNRT2.3a could increase rice yield and nitrogen use efficiency.

Ironing out a survival strategy

Cancer Metastatic cells display a notable ability to adapt to—and even thrive in—harsh microenvironments. One extreme example is leptomeningeal metastases (LM), cancer cells that enter a region of the central nervous system called the subarachnoid space, which is filled with cerebral spinal fluid (CSF). This anatomic site has a limited supply of micronutrients such as iron, and it harbors immune cells. Chi et al. used single-cell RNA sequencing to study CSF samples from cancer patients with LM (see the Perspective by Garzia and Taylor). They found that LM cells express and use components of a high-affinity iron-capturing system. Through this mechanism, the LM cells avoid the adverse effects of iron deprivation and potentially escape immune attack by limiting the supply of iron to macrophages. Science this issue p. [276][1]; see also p. [250][2] [1]: /lookup/doi/10.1126/science.aaz2193 [2]: /lookup/doi/10.1126/science.abb7041

Carbon metabolism snapshot by ddPCR during the early step of Candida albicans phagocytosis by macrophages.

During Candida macrophage interactions, phagocytosed yeast cells feed in order to grow, develop hyphae and escape. Through numerous proteomic and transcriptomic studies, two metabolic phases have been described. A shift to a starvation mode is generally identified as early as one-hour post phagocytosis, followed by a glycolytic growth mode after C. albicans escaped from the macrophage. Healthy macrophages contain low amounts of glucose. To determine if this carbon source was sensed and metabolized by the pathogen, we explored the transcription level of a delimited set of key genes expressed in C. albicans cells during phagocytosis by macrophages, at an early stage of the interaction. This analysis was performed using a technical digital droplet PCR approach to quantify reliably the expression of carbon metabolic genes after 30min of phagocytosis. Our data confirm the technique of digital droplet PCR for the detection of C. albicans transcripts using cells recovered after a short period of phagocytosis. At this stage, carbon metabolism is clearly oriented towards the use of alternative sources. However, the activation of high-affinity glucose transport system suggests that the low amount of glucose initially present in the macrophages is detected by the pathogen.

Generating a High Valency Biotin Binder by Selecting Uniform Protein Assemblies via Crystallization

Crystallization is a common practice in the purification process in small molecule synthesis while selecting the wanted product. For proteins it is rarely applied due to the methodological predicaments in obtaining crystals. Our observation of the stabilized octamers in the crystal structure of hoefavidin, a novel dimeric member of the avidin family, led to the notion of developing a novel biotechnological tool via covalent crosslinking. The avidin–biotin system has been exploited for decades utilizing the ultra-high affinity between avidin and biotin as a basis for numerous applications. Optimizing the system led to the discovery of a novel group of dimeric avidins including hoefavidin. Hoefavidin has a dynamic quaternary structure, where a dimer is the basis for generating the octamer via crystallographic symmetry operation. Upon biotin binding in solution hoefavidin dissociates solely into dimers. In order to stabilize the octamer, we designed the P61C mutant to form a disulfide bridge stabilizing the octamer and preventing dissociation upon biotin binding. The process of selecting P61C hoefavidin uniform octamers includes crystallization followed by dissolving the crystals. The P61C modified hoefavidin octamer can have a substantial added value to the various biotechnological applications and advances of the biotin based high affinity systems.

SAR11 bacteria have a high affinity and multifunctional glycine betaine transporter.

Marine bacterioplankton face stiff competition for limited nutrient resources. SAR11, a ubiquitous clade of very small and highly abundant Alphaproteobacteria, are known to devote much of their energy to synthesizing ATP-binding cassette periplasmic proteins that bind substrates. We hypothesized that their small size and relatively large periplasmic space might enable them to outcompete other bacterioplankton for nutrients. Using uptake experiments with 14 C-glycine betaine, we discovered that two strains of SAR11, Candidatus Pelagibacter sp. HTCC7211 and Cand. P. ubique HTCC1062, have extraordinarily high affinity for glycine betaine (GBT), with half-saturation (K s ) values around 1 nM and specific affinity values between 8 and 14 L mg cell-1 h-1 . Competitive inhibition studies indicated that the GBT transporters in these strains are multifunctional, transporting multiple substrates in addition to GBT. Both strains could use most of the transported compounds for metabolism and ATP production. Our findings indicate that Pelagibacter cells are primarily responsible for the high affinity and multifunctional GBT uptake systems observed in seawater. Maximization of whole-cell affinities may enable these organisms to compete effectively for nutrients during periods when the gross transport capacity of the heterotrophic plankton community exceeds the supply, depressing ambient concentrations.

The regulation of iron homeostasis in the fungal human pathogen Candida glabrata.

Iron is an essential element to most microorganisms, yet an excess of iron is toxic. Hence, living cells have to maintain a tight balance between iron uptake and iron consumption and storage. The control of intracellular iron concentrations is particularly challenging for pathogens because mammalian organisms have evolved sophisticated high-affinity systems to sequester iron from microbes and because iron availability fluctuates among the different host niches. In this review, we present the current understanding of iron homeostasis and its regulation in the fungal pathogen Candida glabrata. This yeast is an emerging pathogen which has become the second leading cause of candidemia, a life-threatening invasive mycosis. C. glabrata is relatively poorly studied compared to the closely related model yeast Saccharomyces cerevisiae or to the pathogenic yeast Candida albicans. Still, several research groups have started to identify the actors of C. glabrata iron homeostasis and its transcriptional and post-transcriptional regulation. These studies have revealed interesting particularities of C. glabrata and have shed new light on the evolution of fungal iron homeostasis.

Sugar transport systems in Kluyveromyces marxianus CCT 7735.

The pattern of glucose repression in most Kluyveromyces marxianus strains does not correlate with fermentative behaviour; however, glucose repression and fermentative metabolism appear to be linked to the kinetics of sugar uptake. In this work, we show that lactose transport in K. marxianus CCT 7735 by lactose-grown cells is mediated by a low-affinity H+-sugar symporter. This system is glucose repressed and able to transport galactose with low affinity. We also observed the activity of a distinct lactose transporter in response to raffinose. Regarding glucose uptake, specificities of at least three low-affinity systems rely on the carbon source available in a given growth medium. Interestingly, it was observed only one high-affinity system is able to transport both glucose and galactose. We also showed that K. marxianus CCT 7735 regulates the expression of sugar transport systems in response to glucose availability.

Humic acid differentially improves nitrate kinetics under low- and high-affinity systems and alters the expression of plasma membrane H+-ATPases and nitrate transporters in rice

Humic acids (HAs) have a major effect on nutrient uptake, metabolism, growth and development in plants. Here, we evaluated the effect of HA pretreatment applied with a nutrient solution on the uptake kinetics of nitrate nitrogen (N-NO3−) and the metabolism of nitrogen (N) in rice under conditions of high and low NO3− supply. In addition, the kinetic parameters of NO3− uptake, N metabolites, and nitrate transporters (NRTs) and the plasma membrane (PM) H+-ATPase gene expression were examined. The plants were grown in a growth chamber with modified Hoagland and Arnon solution until 21 days after germination (DAG), and they were then transferred to a solution without N for 48 h and then to another solution without N and with and without the addition of HAs for another 48 h. After this period of N deprivation, the plants received new nutrient solutions containing 0.2 and 2.0 mM N-NO3−. Treatment of rice plants with HA promoted the induction of the genes OsNRT2.1-2.2/OsNAR2.1 and some isoforms PM H+-ATPase in roots. The application of HAs differentially modified the parameters of the uptake kinetics of NO3− under both concentrations. When grown with 0.2 mM NO3−, the plants pretreated with HA had lower Km and Cmin values as well as a higher Vmax/Km ratio. When grown with 2 mM NO3−, the plants pretreated with HA had a higher Vmax value, a greater root and shoot mass, and a lower root/shoot ratio. The N fractions were also altered by pretreatment with HA, and a greater accumulation of NO3− and N-amino was observed in the roots and shoots, respectively, of plants pretreated with HA. The results suggest that pretreatment with HA modifies root morphology and gene expression of PM H+-ATPases and NO3− transporters, resulting in a greater efficiency of NO3− acquisition by high- and low-affinity systems.

Label-Free FimH Protein Interaction Analysis Using Silicon Nanoribbon BioFETs

The detection of biomarkers at very low concentration and low cost is increasingly important for clinical diagnosis. Moreover, monitoring affinities for receptor-antagonist interactions by time-resolved measurements is crucial for drug discovery and development. Biosensors based on ion-sensitive field-effect transistors (BioFETs) are promising candidates for being integrated into CMOS structures and cost-effective production. The detection of DNA and proteins with silicon nanowires has been successfully demonstrated using high affinity systems such as the biotin–streptavidin interaction. Here, we show the time-resolved label-free detection of the interaction of the bacterial FimH lectin with an immobilized mannose ligand on gold-coated silicon nanoribbon BioFETs. By comparing our results with a commercial state of the art surface plasmon resonance system, additional surface effects become visible when using this charge based detection method. Furthermore, we demonstrate the effect of sensor area on signal...

Uptake Kinetics of Arsenic in Upland Rice Cultivar Zhonghan 221 Inoculated with Arbuscular Mycorrhizal Fungi

Arbuscular mycorrhizal fungi (AMF) appear to be highly associated with arsenic (As) uptake in host plants because arsenate (As(V)) and phosphorus (P) share the same transporter, whereby AMF can enhance P uptake. A short-term experiment was conducted for low- (0 to 0.05 mM As) and high-affinity (0 to 2.5 mM As) uptake systems, to investigate the AMF role on As uptake mechanism in plants, which may explain As uptake kinetics in upland rice cultivar: Zhonghan 221. When concentration of As ranged from 0 to 0.05 mM, Funneliformis geosporum (Fg) significantly decreased arsenite (As(III)) and monomethylarsonicacid (MMA) uptake when (p < 0.05) compared to non-mycorrhizal (NM) treatment, since the major route for (As(III)) in rice roots—rice silicon transporter Lsi1 would be influenced by Fg inoculation at high As concentrations. Fg can also reduce As(V) uptake significantly (p < 0.05) under both uptake systems relative to NM treatment, whereas, Funneliformis mosseae (Fm) increased As(V) and MMA uptake in rice roots, with MMA uptake rate generally lower than As(III) and As(V). Using suitable AMF species inoculation with rice, As uptake and accumulation in rice grains can be reduced and the risk to human health, once consumed, can be minimized.

Glucose-PTS Involvement in Maltose Metabolism by Streptococcus mutans.

Streptococcus mutans grows with starch-derived maltose in the presence of saliva. Maltose transported into the cells is mediated by the MalQ protein (4-alpha-glucanotransferase) to produce glucose and maltooligosaccharides. Glucose can be phosphorylated to glucose 6-phosphate, which can enter the glycolysis pathway. The MalQ enzyme is essential in the catabolism of maltose when it is the sole carbon source, suggesting the presence of a downstream glucokinase of the MalQ enzyme reaction. However, a glucokinase gene-inactivated mutant (glk mutant) grew with maltose as the sole carbon source, with no residual glucokinase activity. This left a phosphoenolpyruvate-dependent phosphotransferase system (PTS) as the only candidate pathway for the phosphorylation of glucose in its transport as a substrate. Our hypothesis was that intracellular glucose derived from maltose mediated by the MalQ protein was released into the extracellular environment, and that such glucose was transported back into the cells by a PTS. The mannose PTS encoded by the manL, manM, and manN genes transports glucose into cells as a high affinity system with concomitant phosphorylation. The purpose of this study was to investigate extracellular glucose by using an enzyme-linked photometrical method, monitoring absorbance changes at 340 nm in supernatant of S. mutans cells. A significant amount of glucose was detected in the extracellular fluid of a glk, manLM double mutant. These results suggest that the glk and manLMN genes participate in maltose catabolism in this organism. The significance of multiple metabolic pathways for important energy sources, including maltose, in the oral environment is discussed.