Unidirectional Inhibition Research Articles (Page 1)

Amyloid-β (Aβ) fibrillation is a spontaneous, thermodynamic process governed by nucleation and elongation. While many studies have explored the ability of engineered nanomaterials (ENMs) to modulate Aβ fibrillation, such as inhibitors, promoters, and dual-modulators, the key physicochemical property of ENMs that determines this behavior remains unclear. In this study, we developed a comprehensive library of ENMs with well-controlled physicochemical properties, including surface charges, morphologies, and hydrophilicity, to systematically investigate their effects on Aβ40 fibrillation. We identified hydrophilicity as the primary determinant of ENM-mediated modulation, rather than surface charge or morphology. Thioflavin T (ThT) kinetics assays indicated that hydrophilic ENMs exhibited bidirectional modulation, both promoting and inhibiting fibrillation depending on concentration. This bidirectional effect results from a competition between accelerated nucleation and decelerated elongation. While hydrophobic ENMs exhibited only unidirectional inhibition from the initial nucleation phase, two-dimensional-NMR (2D-NMR) mechanism studies indicated that this difference resulted from specific interactions with Aβ40 residues. Hydrophilic ENMs targeted hydrophilic residues involved in elongation, including Arginine R5, Glycine G9, G25, G33, G37, and G38, Lysine K28, and Alanine A30, while hydrophobic ENMs bound to hydrophobic residues critical for nucleation, such as I31. These findings provide mechanistic insight into NP-peptide interactions and lay a foundation for the rational design of nanomaterials to modulate amyloid fibrillation.

Two subclasses of acid-sensing ion channels (ASIC3) and of ATP-sensitive P2X receptors (P2X3Rs) show a partially overlapping expression in sensory neurons. Here we report that both recombinant and native receptors interact with each other in multiple ways. Current measurements with the patch-clamp technique prove that ASIC3 stimulation strongly inhibits the P2X3R current partly by a Ca2+-dependent mechanism. The proton-binding site is critical for this effect and the two receptor channels appear to switch their ionic permeabilities during activation. Co-immunoprecipation proves the close association of the two protein structures. BN-PAGE and SDS-PAGE analysis is also best reconciled with the view that ASIC3 and P2X3Rs form a multiprotein structure. Finally, in vivo measurements in rats reveal the summation of pH and purinergically induced pain. In conclusion, the receptor subunits do not appear to form a heteromeric channel, but tightly associate with each other to form a protein complex, mediating unidirectional inhibition.

Axonal transport is critical for maintaining synaptic transmission. Of interest, anterograde and retrograde axonal transport appear to be interdependent, as perturbing one directional motor often impairs movement in the opposite direction. Here live imaging of Drosophila and hippocampal neuron dense-core vesicles (DCVs) containing a neuropeptide or brain-derived neurotrophic factor shows that the F-actin depolymerizing macrolide toxin mycalolide B (MB) rapidly and selectively abolishes retrograde, but not anterograde, transport in the axon and the nerve terminal. Latrunculin A does not mimic MB, demonstrating that F-actin depolymerization is not responsible for unidirectional transport inhibition. Given that dynactin initiates retrograde transport and that amino acid sequences implicated in macrolide toxin binding are found in the dynactin component actin-related protein 1, we examined dynactin integrity. Remarkably, cell extract and purified protein experiments show that MB induces disassembly of the dynactin complex. Thus imaging selective retrograde transport inhibition led to the discovery of a small-molecule dynactin disruptor. The rapid unidirectional inhibition by MB suggests that dynactin is absolutely required for retrograde DCV transport but does not directly facilitate ongoing anterograde DCV transport in the axon or nerve terminal. More generally, MB's effects bolster the conclusion that anterograde and retrograde axonal transport are not necessarily interdependent.

Naturally, simultaneous interactions occurred among plants, herbivores, and soil biota, that is, arbuscular mycorrhizal fungi (AMF), nematodes, and fungal pathogens. These multiple interactions play fundamental roles in driving process, structure, and functioning of ecosystems. In this study, we conducted a meta-analysis with 144 papers to investigate the interactions between AMF and plant biotic stressors and their effects on plant growth performance. We found that AMF enhanced plant tolerance to herbivores, nematodes, and fungal pathogens. We also found reciprocal inhibition between AMF and nematodes as well as fungal pathogens, but unidirectional inhibition for AMF on herbivores. Negative effects of AMF on biotic stressors of plants depended on herbivore feeding sites and actioning modes of fungal pathogens. More performance was reduced in root-feeding than in shoot-feeding herbivores and in rotting- than in wilt-fungal pathogens. However, no difference was found for AMF negative effects between migratory and sedentary nematodes. In return, nematodes and fungal pathogens generated more reduction of root colonization in Non-Glomeraceae than in Glomeraceae. Our results suggested that AMF positive effects on plants might be indirectly mediated by competitive inhibition with biotic stressors of plants. These positive and negative interactions make potential contributions to maintaining ecosystem stability and functioning.

Plants and animals use innate immunity as a first defense against pathogens, a costly yet necessary tradeoff between growth and immunity. In Arabidopsis, the regulatory leucine-rich repeat receptor-like kinase (LRR-RLK) BAK1 combines with the LRR-RLKs FLS2 and EFR in pathogen-associated molecular pattern (PAMP)-triggered immunity (PTI) and the LRR-RLK BRI1 in brassinosteroid (BR)-mediated growth. Therefore, a potential tradeoff between these pathways mediated by BAK1 is often postulated. Here, we show a unidirectional inhibition of FLS2-mediated immune signaling by BR perception. Unexpectedly, this effect occurred downstream or independently of complex formation with BAK1 and associated downstream phosphorylation. Thus, BAK1 is not rate-limiting in these pathways. BRs also inhibited signaling triggered by the BAK1-independent recognition of the fungal PAMP chitin. Our results suggest a general mechanism operative in plants in which BR-mediated growth directly antagonizes innate immune signaling.

Physiological knowledge of the neural mechanisms regulating sleep and wakefulness has been advanced by the recent findings concerning sleep/wakefulness-related preoptic/anterior hypothalamic and perifornical (orexin-containing)/posterior hypothalamic neurons. In this paper, we propose a mathematical model of the mechanisms orchestrating a quartet neural system of sleep and wakefulness composed of the following: 1) sleep-active preoptic/anterior hypothalamic neurons (N-R group); 2) wake-active hypothalamic and brain stem neurons exhibiting the highest rate of discharge during wakefulness and the lowest rate of discharge during paradoxical or rapid eye movement (REM) sleep (WA group); 3) brain stem neurons exhibiting the highest rate of discharge during REM sleep (REM group); and 4) basal forebrain, hypothalamic, and brain stem neurons exhibiting a higher rate of discharge during both wakefulness and REM sleep than during nonrapid eye movement (NREM) sleep (W-R group). The WA neurons have mutual inhibitory couplings with the REM and N-R neurons. The W-R neurons have mutual excitatory couplings with the WA and REM neurons. The REM neurons receive unidirectional inhibition from the N-R neurons. In addition, the N-R neurons are activated by two types of sleep-promoting substances (SPS), which play different roles in the homeostatic regulation of sleep and wakefulness. The model well reproduces the actual sleep and wakefulness patterns of rats in addition to the sleep-related neuronal activities across state transitions. In addition, human sleep-wakefulness rhythms can be simulated by manipulating only a few model parameters: inhibitions from the N-R neurons to the REM and WA neurons are enhanced, and circadian regulation of the N-R and WA neurons is exaggerated. Our model could provide a novel framework for the quantitative understanding of the mechanisms regulating sleep and wakefulness.

The purpose of this study was to determine whether Fungizone or amphotericin B lipid complex (ABLC; ABELCET) affects the transfer of cholesteryl ester (CE) by lipid transfer protein I (LTP I; also known as cholesteryl ester transfer protein) between HDL and LDL (bidirectional transfer HDL to LDL and LDL to HDL). Increasing concentrations of either Fungizone or ABELCET (1.25-12.5 microg AmpB/ml) were incubated with HDL and [3H]CE-LDL or [3H]CE-HDL and LDL (the amount of each fraction added was equivalent to 10 microg of cholesterol) and LTP I in delipidated human plasma at 37 degrees C for 90 min. As a positive control, TP2, a monoclonal antibody directed against LTP-1, was added instead of drug. After incubation, manganese and phosphate reagents were then added to precipitate out all of the LDL. The supernatant, consisted of only HDL, was counted for radioactivity to determine the amount of CE transferred from LDL. Similarly, the precipitate consisted of only LDL, was counted for radioactivity to determine the amount of CE transferred from HDL. For Fungizone, the transfer of cholesteryl ester (CE) between HDL and LDL were not significantly different compared to nontreated controls. For ABELCET, CE transfer from HDL to LDL was significantly decreased at 12.5 microg AmpB/ml compared to control. However, transfer from LDL to HDL was not significantly different compared to non-treated controls. Similar results were observed with the major lipid component of ABELCET, dimyristoylphosphatidylcholine. CE transfer from HDL to LDL and LDL to HDL was significantly decreased when using the positive control (TP2). Fungizone does not affect LTP I-mediated transfer of CE between HDL and LDL. ABELCET inhibits transfer from HDL to LDL, but has no effect on CE transfer from LDL to HDL. This uni-directional inhibition may contribute to the high recovery of AmpB in HDL but the very low presence of drug in the LDL fraction following ABELCET incubation.

H(+)-ATP synthase (F(1)F(o) ATPase) catalyzes the synthesis and/or hydrolysis of ATP, and the reactions are strongly affected by all the substrates (products) in a way clearly distinct from that expected of a simple reversibly operating enzyme. Recent studies have revealed the structure of F(1), which is ideally suited for the alternating binding change mechanism, with a rotating gamma-subunit as the energy-driven coupling device. According to this mechanism ATP, ADP, inorganic phosphate (P(i)) and Mg(2+) participate in the forward and reverse overall reactions exclusively as the substrates and products. However, both F(1) and F(1)F(o) demonstrate non-trivial steady-state and pre-steady-state kinetics as a function of variable substrate (product) concentrations. Several effectors cause unidirectional inhibition or activation of the enzyme. When considered separately, the unidirectional effects of ADP, P(i), Mg(2+) and energy supply on ATP synthesis or hydrolysis may possibly be explained by very complex kinetic schemes; taken together, the results suggest that different conformational states of the enzyme operate in the ATP hydrolase and ATP synthase reactions. A possible mechanism for an energy-dependent switch between the two states of F(1)F(o) ATPase is proposed.

Essential lysine residue(s) in the coenzyme non-specific glutamate dehydrogenase from Chlorella pyrenoidosa 82T is responsible for irreversible inactivation by diethyl pyrocarbonate and unspecific inhibition by 4-chloromercuribenzoate

Early intercellular events occurring in cloned T helper (Th) cells following interaction with cloned T suppressor (Ts) cells were studied by stopped-flow fluorometry. It was found that the increase of intracellular Ca2+ ([Ca2+]i) in major histocompatibility complex (MHC)-restricted Th clones induced by the stimulation with antigen and antigen-presenting cells (APC) is inhibited by the incubation with antigen-activated Ts clones. Optimal suppression required that the two cells recognize antigen on the same APC, although the restriction element for recognition could be different. There was an absolute requirement for recognition of the same antigen by these two cell types. The inhibitory effect was unidirectional in that Ts clones could inhibit the increase of [Ca2+]i of Th clones but not vice versa. Ts clones could not suppress the [Ca2+]i response of other Ts clones. If Th and Ts clones do not share the same MHC restriction specificity, a longer co-incubation time for activation of Ts is required for the inhibition of the [Ca2+]i response of the Th clone, suggesting the presence of a non-specific suppressive mediator that selectively acts on Th.

With a dual-electrode configuration separable few-unit activity was recorded both on one electrode as well as on two electrodes in the auditory midbrain of the grassfrog to a large variety of stimuli. Activity recorded on one electrode was separated by a pattern recognition technique through the use of features of the action potential waveform. Functional connections between units were established on basis of cross-correlation histograms of pairs of simultaneously recorded units. A hierarchical scheme was adopted to describe the various manifestations of neural correlation. If a peak or trough was observed in the simultaneous cross-correlation histogram, irrespective of stimulus conditions, this was called neural synchrony. If this peak or trough was not equal to its shift predictor estimating the stimulus contribution, neural correlation was considered to be present. About 60% of the pairs exhibited neural synchrony, mostly due to shared stimulus influences, independent of mutual distance of units. About 15% of the unit pairs showed neural correlation indicating a functional neural connection. Neural correlation was observed only in units with a distance smaller than 300 micron. The majority (approximately 85%) of the cases showing neural correlation could be ascribed to neural shared input. Unidirectional excitation was observed only in unit pairs recorded on the same electrode. Unidirectional inhibition could not be demonstrated. The dependency of occurrence of neural correlation on unit distance has implications for models of the functional organization of the auditory midbrain. About half of the neurally correlated pairs showed stimulus dependencies of their functional connections. Together with the observed lack of stimulus invariance of single-unit spectrotemporal sensitivities this indicates a dynamic stimulus dependency of functional neuronal organization in the auditory midbrain of the grassfrog. Neuron pairs with a large overlap of their spectrotemporal sensitivities on average had neurally correlated activities more often than pairs with a smaller amount of overlap. In comparison to single-unit coding, ensemble coding by populations of neurons may show an enhanced selectivity to stimulus characteristics.

Sucrose synthase (UDPglucose:D-fructose 2-alpha-D-glucosyltransferase, EC2.4.1.13), which catalyzes the synthesis and cleavage of sucrose, exhibits differences in some properties between the two reactions. When enzyme previously incubated with oxidized glutathione or oxidized thioredoxin was used, sucrose cleavage was inhibited whereas sucrose synthesis proceeded at a normal rate. Sucrose cleavage activity could be restored by incubation with dithiothreitol or reduced glutathione. The thioredoxin effect was influenced by the presence of cleavage reaction substrates--i.e., sucrose and UDP. Thioredoxin action was rather slow compared with the catalytic reaction. These findings may have important implications for understanding the metabolic role of sucrose synthase and oxidized thioredoxin. Theoretically, the fact that an enzyme catalyzing a reversible reaction is inhibited in one direction only suggests that a modification in the enzyme affinities for its substrates must have occurred.

Parallel tests were conducted utilizing, the capillary tube migration test and the Boyden chamber assay, in order to determine whether the decrease in leukocyte chemotaxis that occurs if overoptimal cytotaxin concentrations are applied is due to migration inhibition. Overoptimal doses of casein produced decreased chemotactic response and migration inhibition for both rabbit macrophages and neutrophils. However, guinea pig neutrophils exhibited no decrease in chemotaxis despite high casein doses. Overoptimal doses of acid-denatured anaphylatoxin produced a decreased chemotactic response and migration inhibition of neutrophils. In both assays, this agent showed no effect upon macrophages. It is concluded that a chemotactic signal at different concentrations can elicit unidirectional migration or migration inhibition. Accordingly, chemotactic leukocyte attraction could be antagonistically regulated not only by serum-derived and lymphocyte-derived migration inhibitory factors but also by high doses of the chemotactic factor itself. Thus, the Boyden chamber technique can measure both chemotactic migration and migration inhibition phenomena.

The kinetic and regulatory properties of partially purified phosphoenolpyruvate (PEP) carboxykinase (EC 4.1.1.3.2) from Rhodospirillum rubrum were studied. The enzyme was active with guanosine- and inosinephosphates and must thus be classified as GTP (ITP): oxaloacetate carboxylase (transphosphorylating.) In the direction of oxaloacetate-foramtion, the enzyme was strongly inhibited by ATP (Ki = 0.03 mM). ITP, UTP, CTP, and GTP were less inhibitory. The inhibition was competitive with respect to GDP or IDP, but not with respect to PEP. In the direction of PEP-synthesis, the enzyme was not inhibited, but rather activated by ATP.

A kinetic study of "malic' enzyme (EC 1.1.1.40) from potato suggests that the mechanism is Ordered Bi Ter with NADP+ binding before malate, and NADPH binding before pyruvate and HCO3-. The analysis is complicated by the non-linearity that occurs in some of the plots. meso-Tartrate is shown to inhibit the oxidative decarboxylation of malate but to activate the reductive carboxylation of pyruvate. To explain these unidirectional effects it is suggested that the control site of "malic' enzyme binds organic acids (including meso-tartrate) which activate the enzyme. meso-Tartrate, however, competes with malate for the active site and thus inhibits the oxidative decarboxylation of malate. Because meso-tartrate does not compete effectively with pyruvate for enzyme-NADPH, its binding at the control site leads to a stimulation of the carboxylation of pyruvate. A similar explanation is advanced for the observation that malic acid stimulates its own synthesis.

1. The receptive field properties of single units in the cat superior colliculus have been analyzed quantitatively from average response histograms of the activity elicited in these units by moving light slits. 2. The majority of receptive fields, when tested with narrow (less than 0.5° wide) moving stimuli, have a single excitatory region. In the minority of the receptive fields, two or more spatially separated excitatory regions could be differentiated. Each excitatory region usually had an elliptical or rectangular shape with its major axis parallel or nearly parallel to the horizontal meridian of the visual field. A small proportion (1%) of the receptive fields had only inhibitory regions. 3. A majority of collicular units showed preferences for a particular speed of stimulus motion. Cells with preferences for slow ( 100°/sec). There was also a positive correlation between the speed preference of a unit and the size of the excitatory region of its receptive field. 4. A high proportion of cells had little (<1 spike/sec) spontaneous activity. Of units with high spontaneous activity, a majority had large excitatory regions in their receptive fields and did not show direction selectivity. 5. In addition to the excitatory regions in most receptive fields, there were also inhibitory regions. Stimulation of the latter suppressed both the excitatory response and background firing of the cell. The inhibition and excitation had the same latency, but inhibition lasted at least 150 msec after discontinuation of stimulation, thus reducing responsiveness of the cell after the stimulation. 6. It was found that the majority of collicular units possess a preference for stimulus movements away from the area centralis. 7. Evidence is presented supporting the view that direction selectivity in collicular units is principally determined by unidirectional inhibition. 8. Implications of these results for the mechanism determining receptive field organization in the colliculus are discussed.

The unidirectional inhibition of glutamate dehydrogenase from Blastocladiella emersonii

Abstract A variety of metabolites and chelating agents have been found to elicit a unique form of inhibition on an NAD-specific glutamate dehydrogenase from Blastocladiella emersonii. When measuring the initial rate of the reaction, the oxidative deamination of glutamate was completely inhibited by the metabolites citrate, isocitrate, fructose 1,6-diphosphate, and fumarate and by the nonmetabolites ATP and ethylenediaminetetraacetate. (ATP may be regarded as a metabolite in some reactions involving its transformation to ADP and AMP.) The rate of the reductive amination of α-ketoglutarate reaction was not affected by any of the inhibitors except ATP. The phenomenon of unidirectional inhibition in these initial velocity studies does not consider the microscopic reversibility of the reaction. Divalent cations, AMP, and ADP reversed this inhibition. AMP and ADP, in addition, served as allosteric effectors of this enzyme, activating the reaction in both directions of assay. ATP, on the other hand, participated as a negative allosteric effector. The inhibitory effects of the metabolites and EDTA were dependent on pH. All of the metabolic inhibitors are important intermediates which are involved in the production of energy in the tricarboxylic acid cycle and in glycolysis. A scheme that incorporates these findings into a regulatory network connecting glycolysis and the tricarboxylic acid cycle is formulated.