bluuebunny/crossref_metadata_2025 huggingface dataset:
The Water Crisis in the Gaza Strip: Prospects for Resolution
Authors: Weinthal E. , Vengosh A. , Marei A. , Gutierrez A. , Kloppmann W.
Published: September 2005

Israel and the Palestinian Authority share the southern Mediterranean coastal aquifer. Long‐term overexploitation in the Gaza Strip has resulted in a decreasing water table, accompanied by the degradation of its water quality. Due to high levels of salinity and nitrate and boron pollution, most of the ground water is inadequate for both domestic and agricultural consumption. The rapid rate of population growth in the Gaza Strip and dependence upon ground water as a single water source present a serious challenge for future political stability and economic development. Here, we integrate the results of geochemical studies and numerical modeling to postulate different management scenarios for joint management between Israel and the Palestinian Authority. The chemical and isotopic data show that most of the salinity phenomena in the Gaza Strip are derived from the natural flow of saline ground water from Israel toward the Gaza Strip. As a result, the southern coastal aquifer does not resemble a classic “upstream‐downstream” dispute because Israel's pumping of the saline ground water reduces the salinization rates of ground water in the Gaza Strip. Simulation of different pumping scenarios using a monolayer, hydrodynamic, two‐dimensional model (MARTHE) confirms the hypothesis that increasing pumping along the Gaza Strip border combined with a moderate reduction of pumping within the Gaza Strip would improve ground water quality within the Gaza Strip. We find that pumping the saline ground water for a source of reverse‐osmosis desalination and then supplying the desalinated water to the Gaza Strip should be an essential component of a future joint management strategy between Israel and the Palestinian Authority.
A Framework for Resolving the Transboundary Water Allocation Conflict Conundrum
Authors: Rowland Marty
Published: September 2005

This paper describes a methodology for resolving transboundary water disputes that arise when people/states/nations sharing a resource that crosses legal/political jurisdictions disagree about the use of the resource. Laws and treaties written in an attempt to settle disputes are frequently neither enforced nor effective, and disagreements continue. Crises, arising through resource overuse or shortages, worsen the conflict and typically result in further discord, lawsuits, depletion of the resource, and even open‐armed hostility. Many water management experts call for either private/market‐based or state/command‐and‐control resource management systems, but these eventually break down during crisis. The crises therefore necessitate the adoption of a more effective institutional arrangement to address and resolve present and future problems. A better alternative to management by private or state entities and the resolution of conflicts by the mere application of law is a cooperative approach. The Rowland‐Ostrom Framework, introduced in this paper, incorporates Ostrom's eight design principles for sustainable common pool resource management within the context of crisis that involves an urgent threat to the quantity or quality of a resource such as water, as described by the author. This paper demonstrates that although established 15 years ago, Ostrom's design principles remain applicable today for effective, sustainable transboundary water management, and the Rowland‐Ostrom Framework is a model for the equitable use of shared water resources throughout the world.
Ground Water Rights, Spatial Variation, and Transboundary Conflicts
Authors: Matthews Olen Paul
Published: September 2005

The goal for any property rights system is to achieve equity, efficiency, and certainty. Trying to achieve these goals for ground water is difficult because a ground water right is not exclusive. To make matters more complicated, ground water is often under the jurisdiction of more than one political unit. The result is transboundary conflicts. Two critical elements must be included in any system of ground water rights. The system must define how the ground water can be used and define the relationships that each user and each use has with the other users and uses in the system. Unfortunately, these relationships are seldom completely defined and are made more complex by the transboundary scales at which they operate. As ground water moves horizontally across boundaries, different users or different jurisdictions have sequential control, creating conflicts between the first users and subsequent ones. Other problems occur because of vertical relationships, with more than one person or entity having control over ground water at the same time. This simultaneous exercise of authority can create conflicts between an individual who possesses a right to use ground water and a state or federal agency that regulates the same water. Transboundary conflicts occur at different scales and include conflicts between neighboring property owners as well as conflicts between countries. Scale, the property rights structure, and the nature of the relationship between users influence the way transboundary ground water conflicts are resolved.
Securing the Future of Ground Water Resources in the Great Lakes Basin
Authors: Galloway Gerald , Pentland Ralph
Published: September 2005

Ground water is a vital, but underappreciated, natural resource in the Great Lakes basin. It meets many human needs and contributes significantly to the hydrology of the Great Lakes and the health of ecosystems. This paper provides an overview of ground water in the Great Lakes and the institutional and legal setting that governs the use, protection, diversion, and removal of water from the basin and proposes a citizen‐centered vision for management of ground water in the 21st century.
Transboundary Aquifers: A Global Program to Assess, Evaluate, and Develop Policy
Authors: Puri S. , Aureli A.
Published: September 2005

Transboundary aquifers are as important a component of global water resource systems as are transboundary rivers; yet, their recognition in international water policy and legislation is very limited. Existing international conventions and agreements barely address aquifers and their resources. To rectify this deficiency, the International Association of Hydrogeologists and UNESCO's International Hydrological Programme have established the Internationally Shared (transboundary) Aquifer Resource Management (ISARM) Programme. This multiagency cooperative program has launched a number of global and regional initiatives. These are designed to delineate and analyze transboundary aquifer systems and to encourage riparian states to work cooperatively toward mutually beneficial and sustainable aquifer development. The agencies participating in ISARM include international and regional organizations (e.g., Organization of American States, United Nations Environment Programme, United Nations Economic Commission for Europe, Food and Agriculture Organization, and South African Development Community). Using outputs of case studies, the ISARM Programme is building scientific, legal, environmental, socioeconomic, and institutional guidelines and recommendations to aid sharing nations in the management of their transboundary aquifers. Since its start in 2000, the program has completed inventories of transboundary aquifers in the Americas and Africa, and several ISARM case studies have commenced.
Stochastic Study of Solute Transport in a Nonstationary Medium
Authors: Hu Bill X.
Published: September 2005

A Lagrangian stochastic approach is applied to develop a method of moment for solute transport in a physically and chemically nonstationary medium. Stochastic governing equations for mean solute flux and solute covariance are analytically obtained in the first‐order accuracy of log conductivity and/or chemical sorption variances and solved numerically using the finite‐difference method. The developed method, the numerical method of moments (NMM), is used to predict radionuclide solute transport processes in the saturated zone below the Yucca Mountain project area. The mean, variance, and upper bound of the radionuclide mass flux through a control plane 5 km downstream of the footprint of the repository are calculated. According to their chemical sorption capacities, the various radionuclear chemicals are grouped as nonreactive, weakly sorbing, and strongly sorbing chemicals. The NMM method is used to study their transport processes and influence factors. To verify the method of moments, a Monte Carlo simulation is conducted for nonreactive chemical transport. Results indicate the results from the two methods are consistent, but the NMM method is computationally more efficient than the Monte Carlo method. This study adds to the ongoing debate in the literature on the effect of heterogeneity on solute transport prediction, especially on prediction uncertainty, by showing that the standard derivation of solute flux is larger than the mean solute flux even when the hydraulic conductivity within each geological layer is mild. This study provides a method that may become an efficient calculation tool for many environmental projects.
Location and function of vesicle clusters, active zones and Ca2+ channels in the lamprey presynaptic terminal
Authors: Photowala Huzefa , Freed Rachel , Alford Simon
Published: September 2005

Synaptic transmission requires spatial and temporal coordination of a specific sequence of events. The trigger for synaptic vesicle exocytosis is Ca2+ entry into presynaptic terminals, leading to neurotransmitter release at highly specialized sites known as active zones. Ca2+ channel proximity to exocytotic proteins and vesicle clusters at active zones have been inferred from biochemical, histological and ultrastructural data, but direct evidence about functional relationships between these elements in central synapses is absent. We have utilized the lamprey giant reticulospinal synapse to characterize functional colocalization of known synaptic markers in the presynaptic terminal, as well as their reliability during repeated activation. Recycling vesicle clusters, surrounding actin filaments, and physiologically relevant Ca2+ influx all show identical morphological distribution. Ca2+ influx is mediated by clusters of Ca2+ channels that colocalize with the vesicle clusters, defined by imaged sites of vesicle recycling and actin localization. Synaptic transmission is inhibited by block of actin depolymerization, but Ca2+ signalling is unaffected. Functional Ca2+ channels are localized to presynaptic clusters, and Ca2+ transients at these sites account for neurotransmitter release based on their spatial and temporal profiles. Ca2+ transients evoked by single axonal action potentials are mediated solely by voltage‐operated Ca2+ channel activation, and slower Ca2+ rises observed throughout the axon result from Ca2+ diffusion from the synaptic regions. We conclude that at lamprey giant reticulospinal synapses, Ca2+ channels and release sites colocalize, creating a close spatial relationship between active zones and Ca2+ entry sites, which is necessary for site‐specific, Ca2+‐dependent secretion.
Dissociated effects of diazepam and lorazepam on short‐latency afferent inhibition
Authors: Di Lazzaro Vincenzo , Pilato Fabio , Dileone Michele , Tonali Pietro A. , Ziemann Ulf
Published: September 2005

Peripheral nerve inputs have an inhibitory effect on motor cortex excitability at short intervals (short‐latency afferent inhibition, SAI). This can be tested by coupling electrical stimulation of peripheral nerve with transcranial magnetic stimulation (TMS) of the motor cortex. SAI is reduced by the anticholinergic drug scopolamine, and in patients with Alzheimer's disease. Therefore, it is possible that SAI is a marker of central cholinergic activity important for memory function. The benzodiazepine lorazepam also reduces SAI. Since benzodiazepines impair memory formation, but do not do so uniformly, with a maximum amnesic effect after lorazepam but less or no effect after diazepam, we were interested in testing in this non‐behavioural study to what extent the effects of lorazepam and diazepam on circuits involved in SAI could be dissociated. In addition, and for control, we tested the effects of lorazepam and diazepam on short‐interval intracortical inhibition (SICI), a motor cortical inhibition mediated through the GABAA receptor. Lorazepam markedly reduced SAI, whereas diazepam slightly increased it. In contrast, both benzodiazepines uniformly increased SICI. Our findings demonstrate opposite effects of lorazepam and diazepam on SAI, an inhibition modulated by central cholinergic activity, but the same effects on SICI, a marker of neurotransmission through the GABAA receptor. This dissociation suggests, for the first time, that TMS measures of cortical inhibition provide the opportunity to segregate differences of benzodiazepine action in human central nervous system circuits.
Photoreceptor encoding of supersaturating light stimuli in salamander retina
Authors: Xu Jian Wei , Hou Mingli , Slaughter Malcolm M.
Published: September 2005

In the dark‐adapted salamander retina, spikes could be elicited from rods under normal physiological conditions. Spike activity was observed in rods during the recovery phase of the response to saturating light. These action potentials were calcium spikes, blocked by cadmium and L‐type calcium channel blockers. In response to light stimuli that saturate the rod peak response, calcium action potentials occurred with a delay that depended on light intensity, with stronger light increasing spike latency. Therefore, these spikes encode rod visual information at light intensities beyond rod saturation. Postsynaptic currents of similar time course were observed in second and third order neurones. Since rods exposed to brighter light stimuli produced more delayed spike activity, these signals might contribute to negative afterimages.
Real‐time imaging of Rab3a and Rab5a reveals differential roles in presynaptic function
Authors: Star Erin N. , Newton A. Jamila , Murthy Venkatesh N.
Published: September 2005

We investigated the roles of two Rab‐family proteins, Rab3a and Rab5a, in hippocampal synaptic transmission using real‐time fluorescence imaging. During synaptic activity, Rab3a dissociated from synaptic vesicles and dispersed into neighbouring axonal regions. Dispersion required calcium‐dependent exocytosis and was complete before the entire vesicle pool turned over. In contrast, even prolonged synaptic activity produced limited dispersion of Rab5a. A GTPase‐deficient mutant, Rab3a (Q81L), dispersed more slowly than wild‐type Rab3a, and decreased the rate of exocytosis and the size of the recycling pool of vesicles. While overexpression of Rab3a did not affect vesicle recycling, overexpression of Rab5a reduced the recycling pool size by 50%. We propose that while Rab3a preferentially associates with recycling synaptic vesicles and modulates their trafficking, Rab5a is largely excluded from recycling vesicles.
Tetrahydrobiopterin augments endothelium‐dependent dilatation in sedentary but not in habitually exercising older adults
Authors: Eskurza Iratxe , Myerburgh Laura A. , Kahn Zachary D. , Seals Douglas R.
Published: September 2005

Endothelium‐dependent dilatation (EDD) is impaired with ageing in sedentary, but not in regularly exercising adults. We tested the hypotheses that differences in tetrahydrobiopterin (BH4) bioactivity are key mechanisms explaining the impairment in EDD with sedentary ageing, and the maintenance of EDD with ageing in regularly exercising adults. Brachial artery flow‐mediated dilatation (FMD), normalized for local shear stress, was measured after acute oral placebo or BH4 in young sedentary (YS) (n= 10; 22 ± 1 years, mean ±s.e.m.), older sedentary (OS) (n= 9; 62 ± 2), and older habitually aerobically trained (OT) (n= 12; 66 ± 1) healthy men. At baseline, FMD was ∼50% lower in OS versus YS (1.12 ± 0.09 versus 0.57 ± 0.09 (Δmm (dyn cm−2)) × 10−2, P < 0.001; 1 dyn = 10−5 N), but was preserved in OT (0.93 ± 0.08 (Δmm (dyn cm−2)) × 10−2). BH4 administration improved FMD by ∼45% in OS (1.00 ± 0.10 (Δmm (dyn cm−2)) × 10−2, P < 0.01 versus baseline), but did not affect FMD in YS or OT. Endothelium‐independent dilatation neither differed between groups at baseline nor changed with BH4 administration. These results suggest that BH4 bioactivity may be a key mechanism involved in the impairment of conduit artery EDD with sedentary ageing, and the EDD‐preserving effect of habitual exercise.
Kv4 potassium channel subunits control action potential repolarization and frequency‐dependent broadening in rat hippocampal CA1 pyramidal neurones
Authors: Kim Jinhyun , Wei Dong‐Sheng , Hoffman Dax A.
Published: September 2005

A‐type potassium channels regulate neuronal firing frequency and the back‐propagation of action potentials (APs) into dendrites of hippocampal CA1 pyramidal neurones. Recent molecular cloning studies have found several families of voltage‐gated K+ channel genes expressed in the mammalian brain. At present, information regarding the relationship between the protein products of these genes and the various neuronal functions performed by voltage‐gated K+ channels is lacking. Here we used a combination of molecular, electrophysiological and imaging techniques to show that one such gene, Kv4.2, controls AP half‐width, frequency‐dependent AP broadening and dendritic action potential propagation. Using a modified Sindbis virus, we expressed either the enhanced green fluorescence protein (EGFP)‐tagged Kv4.2 or an EGFP‐tagged dominant negative mutant of Kv4.2 (Kv4.2gW362F) in CA1 pyramidal neurones of organotypic slice cultures. Neurones expressing Kv4.2gW362F displayed broader action potentials with an increase in frequency‐dependent AP broadening during a train compared with control neurones. In addition, Ca2+ imaging of Kv4.2gW362F expressing dendrites revealed enhanced AP back‐propagation compared to control neurones. Conversely, neurones expressing an increased A‐type current through overexpression of Kv4.2 displayed narrower APs with less frequency dependent broadening and decreased dendritic propagation. These results point to Kv4.2 as the major contributor to the A‐current in hippocampal CA1 neurones and suggest a prominent role for Kv4.2 in regulating AP shape and dendritic signalling. As Ca2+ influx occurs primarily during AP repolarization, Kv4.2 activity can regulate cellular processes involving Ca2+‐dependent second messenger cascades such as gene expression and synaptic plasticity.
Parallel reflex pathways from flexor muscle afferents evoking resetting and flexion enhancement during fictive locomotion and scratch in the cat
Authors: Stecina Katinka , Quevedo Jorge , McCrea David A.
Published: September 2005

Reflex actions of muscle afferents in hindlimb flexor nerves were examined on ipsilateral motoneurone activity recorded in peripheral nerves during midbrain stimulation‐evoked fictive locomotion and during fictive scratch in decerebrate cats. Trains of stimuli (15–30 shocks at 200 Hz) were delivered during the flexion phase at intensities sufficient to activate both group I and II afferents (5 times threshold, T). In many preparations tibialis anterior (TA) nerve stimulation terminated ongoing flexion and reset the locomotor cycle to extension (19/31 experiments) while extensor digitorum longus (EDL) stimulation increased and prolonged the ongoing flexor phase activity (20/33 preparations). The effects of sartorius, iliopsoas and peroneus longus muscle afferent stimulation were qualitatively similar to those of EDL nerve. Resetting to extension was seen only with higher intensity stimulation (5T) while ongoing flexor activity was often enhanced at group I intensity (2T) stimulation. The effects of flexor nerve stimulation were qualitatively similar during fictive scratch. Reflex reversals were consistently observed in some fictive locomotor preparations. In those cases, EDL stimulation produced a resetting to extension and TA stimulation prolonged the ongoing flexion phase. Occasionally reflex reversals occurred spontaneously during only one of several stimulus presentations. The variable and opposite actions of flexor afferents on the locomotor step cycle indicate the existence of parallel spinal reflex pathways. A hypothetical organization of reflex pathways from flexor muscle afferents to the spinal pattern generator networks with competing actions of group I and group II afferents on the flexor and extensor portions of this central circuitry is proposed.
Functional NR2B‐ and NR2D‐containing NMDA receptor channels in rat substantia nigra dopaminergic neurones
Authors: Jones Susan , Gibb Alasdair J.
Published: September 2005

NMDA receptors regulate burst firing of dopaminergic neurones in the substantia nigra pars compacta (SNc) and may contribute to excitotoxic cell death in Parkinson's disease (PD). In order to investigate the subunit composition of functional NMDA receptors in identified rat SNc dopaminergic neurones, we have analysed the properties of individual NMDA receptor channels in outside‐out patches. NMDA (100 nm) activated channels corresponding to four chord conductances of 18, 30, 41 and 54 pS. Direct transitions were observed between all conductance levels. Between 18 pS and 41 pS conductance levels, direct transitions were asymmetric, consistent with the presence of NR2D‐containing NMDA receptors. Channel activity in response to 100 nm or 200 μm NMDA was not affected by zinc or TPEN (N,N,N′,N′‐tetrakis‐[2‐pyridylmethyl]‐ethylenediamine), indicating that SNc dopaminergic neurones do not contain functional NR2A subunits. The effect of the NR2B antagonist ifenprodil was complex: 1 μm ifenprodil reduced open probability, while 10 μm reduced channel open time but had no effect on open probability of channels activated by 100 nm NMDA. When the concentration of NMDA was increased to 200 μm, ifenprodil (10 μm) produced the expected reduction in open probability. These results indicate that NR2B subunits are present in SNc dopaminergic neurones. Taken together, these findings indicate that NR2D and NR2B subunits form functional NMDA receptor channels in SNc dopaminergic neurones, and suggest that they may form a triheteromeric NMDA receptor composed of NR1/NR2B/NR2D subunits.
Activation of AMPK in rat hypothalamus participates in cold‐induced resistance to nutrient‐dependent anorexigenic signals
Authors: Roman Erika A. , Cesquini Maristela , Stoppa Graziela R. , Carvalheira José B. , Torsoni Márcio A. , Velloso Lício A.
Published: September 2005

The exposure of homeothermic animals to a cold environment leads to a powerful activation of orexigenic signalling which is accompanied by molecular and functional resistance to insulin‐induced inhibition of feeding. Recent evidence suggests that AMPK participates in nutrient‐dependent control of satiety and adiposity. The objective of the present study was to evaluate the effect of cold exposure upon the molecular activation of AMPK signalling in the hypothalamus of rats. Immunoblotting demonstrated that cold exposure per se is sufficient for inducing, on a time‐dependent basis, the molecular activation of the serine/threonine kinase AMP‐activated protein kinase (AMPK) and inactivation of the acetyl‐CoA carboxylase (ACC). These molecular phenomena were accompanied by resistance to nutrient‐induced inactivation of AMPK and activation of ACC. Moreover, cold‐exposure led to a partial inhibition of a feeding‐induced anorexigenic response, which was paralleled by resistance to insulin‐induced suppression of feeding. Finally, cold exposure significantly impaired insulin‐induced inhibition of AMPK through a mechanism dependent on the molecular cross‐talk between phosphatidylinositol‐3(PI3)‐kinase/Akt and AMPK. In conclusion, increased feeding during cold exposure results, at least in part, from resistance to insulin‐ and nutrient‐dependent anorexigenic signalling in the hypothalamus.
On the peripheral and central chemoreception and control of breathing: an emerging role of ATP
Authors: Gourine Alexander V.
Published: September 2005

Peripheral and central respiratory chemoreceptors are ultimately responsible for maintenance of constant levels of arterialP,Pand [H+], protecting the brain from hypoxia and ensuring that the breathing is always appropriate for metabolism. The aim of this discussion is to shed some light on the potential mechanisms ofchemosensory transduction– the process which links chemosensory mechanisms to the central nervous mechanisms controlling breathing. Recent experimental data suggest that the purine nucleotide ATP acts as a common mediator of peripheral and central chemosensory transduction (within the carotid body and the medulla oblongata, respectively). In response to a decrease inP(hypoxia) oxygen‐sensitive glomus cells of the carotid body release ATP to activate chemoafferent fibres of the carotid sinus nerve which transmit this information to the brainstem respiratory centres. In response to an increase inP/[H+](hypercapnia) chemosensitive structures located on the ventral surface of the medulla oblongata rapidly release ATP, which acts locally within the medullary respiratory network. The functional role of ATP released at both sites is similar – to evoke adaptive enhancement in breathing. Understanding the mechanisms of ATP release in response to chemosensory stimulation may prove to be essential for further detailed analysis of cellular and molecular mechanisms underlying respiratory chemosensitivity.
Paired‐pulse potentiation of α7‐containing nAChRs in rat hippocampal CA1 stratum radiatum interneurones
Authors: Klein Rebecca C. , Yakel Jerrel L.
Published: September 2005

Diverse subtypes of nicotinic acetylcholine receptors (nAChRs), including fast‐desensitizing α7‐containing receptors, are expressed in the CNS. While nAChRs appear to regulate cognitive processing and synaptic plasticity, little is known to date about how this regulation occurs, particularly in brain regions known to be important for cognition. By combining patch‐clamp electrophysiology with local photolysis of caged carbachol to rapidly activate the α7‐containing nAChRs in rat hippocampal CA1 stratum radiatum interneurones in slices, we describe a novel transient up‐regulation of channel function. The nAChRs were activated using a paired‐pulse uncaging protocol, where the duration of the UV laser pulses (5–25 ms) and the interval between pulses (200 ms to 30 s) were varied. At relatively long interpulse intervals, we observed a strong (> 75%) decrease in the amplitude of the second response due to desensitization. However, when two pulses were applied at a 200 ms interval, a > 3‐fold increase in the amplitude of the second response was observed, a phenomenon referred to here as paired‐pulse potentiation. Interestingly, this potentiation appeared to be regulated by [Ca2+]i, and/or Ca2+‐dependent processes, as it was significantly enhanced by dialysing cells with either the Ca2+chelator BAPTA, or with peptide inhibitors of either calcineurin or PKC, and was attenuated by dialysing cells with the CaMKII inhibitor KN‐93. No potentiation was observed using caged GABA or glutamate, indicating some specificity for nAChRs. Thus, rat hippocampal α7‐containing nAChRs possess a newly described phenomenon of paired‐pulse potentiation that may be involved in regulating synaptic plasticity in the hippocampus.
Parallel Patterns of Evolution in the Genomes and Transcriptomes of Humans and Chimpanzees
Authors: Khaitovich Philipp , Hellmann Ines , Enard Wolfgang , Nowick Katja , Leinweber Marcus , Franz Henriette , Weiss Gunter , Lachmann Michael , Pääbo Svante
Published: September 2005

The determination of the chimpanzee genome sequence provides a means to study both structural and functional aspects of the evolution of the human genome. Here we compare humans and chimpanzees with respect to differences in expression levels and protein-coding sequences for genes active in brain, heart, liver, kidney, and testis. We find that the patterns of differences in gene expression and gene sequences are markedly similar. In particular, there is a gradation of selective constraints among the tissues so that the brain shows the least differences between the species whereas liver shows the most. Furthermore, expression levels as well as amino acid sequences of genes active in more tissues have diverged less between the species than have genes active in fewer tissues. In general, these patterns are consistent with a model of neutral evolution with negative selection. However, for X-chromosomal genes expressed in testis, patterns suggestive of positive selection on sequence changes as well as expression changes are seen. Furthermore, although genes expressed in the brain have changed less than have genes expressed in other tissues, in agreement with previous work we find that genes active in brain have accumulated more changes on the human than on the chimpanzee lineage.
Direct Isolation of Satellite Cells for Skeletal Muscle Regeneration
Authors: Montarras Didier , Morgan Jennifer , Collins Charlotte , Relaix Frédéric , Zaffran Stéphane , Cumano Ana , Partridge Terence , Buckingham Margaret
Published: September 2005

Muscle satellite cells contribute to muscle regeneration. We have used a
Pax3
GFP/+
mouse line to directly isolate (Pax3)(green fluorescent protein)–expressing muscle satellite cells, by flow cytometry from adult skeletal muscles, as a homogeneous population of small, nongranular, Pax7+, CD34+, CD45–, Sca1– cells. The flow cytometry parameters thus established enabled us to isolate satellite cells from wild-type muscles. Such cells, grafted into muscles of
mdx nu/nu
mice, contributed both to fiber repair and to the muscle satellite cell compartment. Expansion of these cells in culture before engraftment reduced their regenerative capacity.
Movement of Eukaryotic mRNAs Between Polysomes and Cytoplasmic Processing Bodies
Authors: Brengues Muriel , Teixeira Daniela , Parker Roy
Published: September 2005

Eukaryotic cells contain nontranslating messenger RNA concentrated in P-bodies, which are sites where the mRNA can be decapped and degraded. We present evidence that mRNA molecules within yeast P-bodies can also return to translation. First, inhibiting delivery of new mRNAs to P-bodies leads to their disassembly independent of mRNA decay. Second, P-bodies decline in a translation initiation–dependent manner during stress recovery. Third, reporter mRNAs concentrate in P-bodies when translation initiation is blocked and resume translation and exit P-bodies when translation is restored. Fourth, stationary phase yeast have large P-bodies containing mRNAs that reenter translation when growth resumes. The reciprocal movement of mRNAs between polysomes and P-bodies is likely to be important in the control of mRNA translation and degradation. Moreover, the presence of related proteins in P-bodies and maternal mRNA storage granules suggests this mechanism is widely adapted for mRNA storage.
Coherent Manipulation of Coupled Electron Spins in Semiconductor Quantum Dots
Authors: Petta J. R. , Johnson A. C. , Taylor J. M. , Laird E. A. , Yacoby A. , Lukin M. D. , Marcus C. M. , Hanson M. P. , Gossard A. C.
Published: September 2005

We demonstrated coherent control of a quantum two-level system based on two-electron spin states in a double quantum dot, allowing state preparation, coherent manipulation, and projective readout. These techniques are based on rapid electrical control of the exchange interaction. Separating and later recombining a singlet spin state provided a measurement of the spin dephasing time,
T
2
*, of ∼10 nanoseconds, limited by hyperfine interactions with the gallium arsenide host nuclei. Rabi oscillations of two-electron spin states were demonstrated, and spin-echo pulse sequences were used to suppress hyperfine-induced dephasing. Using these quantum control techniques, a coherence time for two-electron spin states exceeding 1 microsecond was observed.
Towards a Genome-Based Taxonomy for Prokaryotes
Authors: Konstantinidis Konstantinos T. , Tiedje James M.
Published: September 2005

The ranks higher than the species in the prokaryotic taxonomy are primarily designated based on phylogenetic analysis of the 16S rRNA gene sequences, but no definite standards exist for the absolute relatedness (measured by 16S rRNA or other means) between the ranks. Accordingly, it remains unknown how comparable the ranks are between different organisms. To gain insights into this question, we studied the relationship between shared gene content and genetic relatedness for 175 fully sequenced strains, using as a robust measure of relatedness the average amino acid identity (AAI) of the shared genes. Our results reveal that adjacent ranks (e.g., phylum versus class) frequently show extensive overlap in terms of genetic and gene content relatedness of the grouped organisms, and hence, the current system is of limited predictive power in this respect. The overlap between nonadjacent ranks (e.g., phylum versus family) is generally limited and attributable to clear inconsistencies of the taxonomy. In addition to providing means for standardizing taxonomy, our AAI-based approach provides a means to evaluate the robustness of alternative genetic markers for phylogenetic purposes. For instance, the 23S rRNA gene was found to be as good a marker as the 16S rRNA gene, while several of the widely distributed protein-coding genes, such as the RNA polymerase and gyrase subunits, show a strong phylogenetic signal, albeit less strong than the rRNA genes (0.78 >
R
2
> 0.69 for the protein-coding genes versus
R
2
= 0.84 for the rRNA genes). The AAI approach outlined here could contribute significantly to a genome-based taxonomy for all microbial organisms.
Differential Roles of the Universal Stress Proteins of
Escherichia coli
in Oxidative Stress Resistance, Adhesion, and Motility
Authors: Nachin Laurence , Nannmark Ulf , Nyström Thomas
Published: September 2005

The universal stress protein (UspA) superfamily encompasses a conserved group of proteins that are found in bacteria, archaea, and eukaryotes.
Escherichia coli
harbors six
usp
genes—
uspA
, -
C
, -
D
, -
E
, -
F
, and -
G
—the expression of which is triggered by a large variety of environmental insults. The
uspA
gene is important for survival during cellular growth arrest, but the exact physiological role of the Usp proteins is not known. In this work we have performed phenotypic characterization of mutants with deletions of the six different
usp
genes. We report on hitherto unknown functions of these genes linked to motility, adhesion, and oxidative stress resistance, and we show that
usp
functions are both overlapping and distinct. Both UspA and UspD are required in the defense against superoxide-generating agents, and UspD appears also important in controlling intracellular levels of iron. In contrast, UspC is not involved in stress resistance or iron metabolism but is essential, like UspE, for cellular motility. Electron microscopy demonstrates that
uspC
and
uspE
mutants are devoid of flagella. In addition, the function of the
uspC
and
uspE
genes is linked to cell adhesion, measured as FimH-mediated agglutination of yeast cells. While the UspC and UspE proteins promote motility at the expense of adhesion, the UspF and UspG proteins exhibit the exact opposite effects. We suggest that the Usp proteins have evolved different physiological functions that reprogram the cell towards defense and escape during cellular stress.
Modulation of the Sensitivity of FimB Recombination to Branched-Chain Amino Acids and Alanine in
Escherichia coli
K-12
Authors: Lahooti Maryam , Roesch Paula L. , Blomfield Ian C.
Published: September 2005

Phase variation of type 1 fimbriae of
Escherichia coli
requires the site-specific recombination of a short invertible element. Inversion is catalyzed by FimB (switching in either direction) or FimE (inversion mainly from on to off) and is influenced by auxiliary factors integration host factor (IHF) and leucine-responsive regulatory protein (Lrp). These proteins bind to sites (IHF site II and Lrp sites 1 and 2) within the invertible element to stimulate recombination, presumably by bending the DNA to enhance synapses. Interaction of Lrp with a third site (site 3) cooperatively with sites 1 and 2 (termed complex 1) impedes recombination. Inversion is stimulated by the branched-chain amino acids (particularly leucine) and alanine, and according to a current model, the amino acids promote the selective loss of Lrp from site 3 (complex 2). Here we show that the central portion of the
fim
invertible element, situated between Lrp site 3 and IHF site II, is dispensable for FimB recombination but that this region is also required for full amino acid stimulation of inversion. Further work reveals that the region is likely to contain multiple regulatory elements. Lrp site 3 is shown to bind the regulatory protein with low affinity, and a mutation that enhances binding to this element is found both to diminish the stimulatory effects of IVLA on FimB recombination and to inhibit recombination in the absence of the amino acids. The results obtained emphasize the importance of Lrp site 3 as a control element but also highlight the complexity of the regulatory system that affects this site.
Mutant Analysis Reveals a Specific Requirement for Protein P30 inMycoplasma pneumoniaeGliding Motility
Authors: Hasselbring Benjamin M. , Jordan Jarrat L. , Krause Duncan C.
Published: September 2005

The cell-wall-less prokaryoteMycoplasma pneumoniae, long considered among the smallest and simplest cells capable of self-replication, has a distinct cellular polarity characterized by the presence of a differentiated terminal organelle which functions in adherence to human respiratory epithelium, gliding motility, and cell division. Characterization of hemadsorption (HA)-negative mutants has resulted in identification of several terminal organelle proteins, including P30, the loss of which results in developmental defects and decreased adherence to host cells, but their impact onM. pneumoniaegliding has not been investigated. Here we examined the contribution of P30 to gliding motility on the basis of satellite growth and cell gliding velocity and frequency.M. pneumoniaeHA mutant II-3 lacking P30 was nonmotile, but HA mutant II-7 producing a truncated P30 was motile, albeit at a velocity 50-fold less than that of the wild type. HA-positive revertant II-3R producing an altered P30 was unexpectedly not fully wild type with respect to gliding. Complementation of mutant II-3 with recombinant wild-type and mutant alleles confirmed the correlation between gliding defect and loss or alteration in P30. Surprisingly, fusion of yellow fluorescent protein to the C terminus of P30 had little impact on cell gliding velocity and significantly enhanced HA. Finally, while quantitative examination of HA revealed clear distinctions among these mutant strains, gliding defects did not correlate strictly with the HA phenotype, and all strains attached to glass at wild-type levels. Taken together, these findings suggest a role for P30 in gliding motility that is distinct from its requirement in adherence.
Function of the Cytochrome
bc
1
-
aa
3
Branch of the Respiratory Network in Mycobacteria and Network Adaptation Occurring in Response to Its Disruption
Authors: Matsoso Limenako G. , Kana Bavesh D. , Crellin Paul K. , Lea-Smith David J. , Pelosi Assunta , Powell David , Dawes Stephanie S. , Rubin Harvey , Coppel Ross L. , Mizrahi Valerie
Published: September 2005

The aerobic electron transport chain in
Mycobacterium smegmatis
can terminate in one of three possible terminal oxidase complexes. The structure and function of the electron transport pathway leading from the menaquinol-menaquinone pool to the cytochrome
bc
1
complex and terminating in the
aa
3
-type cytochrome
c
oxidase was characterized.
M. smegmatis
strains with mutations in the
bc
1
complex and in subunit II of cyctochome
c
oxidase were found to be profoundly growth impaired, confirming the importance of this respiratory pathway for mycobacterial growth under aerobic conditions. Disruption of this pathway resulted in an adaptation of the respiratory network that is characterized by a marked up-regulation of
cydAB
, which encodes the bioenergetically less efficient and microaerobically induced cytochrome
bd
-type menaquinol oxidase that is required for the growth of
M. smegmatis
under O
2
-limiting conditions. Further insights into the adaptation of this organism to rerouting of the electron flux through the branch terminating in the
bd
-type oxidase were revealed by expression profiling of the
bc
1
-deficient mutant strain using a partial-genome microarray of
M. smegmatis
that is enriched in essential genes. Although the expression profile was indicative of an increase in the reduced state of the respiratory chain, blockage of the
bc
1
-
aa
3
pathway did not induce the sentinel genes of
M. smegmatis
that are induced by oxygen starvation and are regulated by the DosR two-component regulator.
Roles of the Regulatory Proteins FlhF and FlhG in the
Vibrio cholerae
Flagellar Transcription Hierarchy
Authors: Correa Nidia E. , Peng Fen , Klose Karl E.
Published: September 2005

Vibrio cholerae
, the causative agent of the human diarrheal disease cholera, is a motile bacterium with a single polar flagellum, and motility has been inferred to be an important aspect of virulence. The
V. cholerae
flagellar hierarchy is organized into four classes of genes. The expression of each class of genes within a flagellar hierarchy is generally tightly regulated in other bacteria by both positive and negative regulatory elements. To further elucidate flagellar biogenesis in
V. cholerae
, we characterized the roles of the three putative regulatory genes,
flhF
,
flhG
, and VC2061.
V. cholerae flhF
and
flhG
mutants appeared nonmotile in a soft agar assay. Electron microscopy revealed that the
flhF
mutant lacked a polar flagellum, while interestingly, the
flhG
mutant possessed multiple (8 to 10) polar flagella per cell. The transcriptional activity of class III and class IV gene promoters in the
flhF
mutant was decreased, suggesting that FlhF acts as a positive regulator of class III gene transcription. The transcription of all four classes of flagellar promoters was increased in the
flhG
mutant, suggesting that FlhG acts as a negative regulator of class I gene transcription. Additionally, the ability to colonize the infant mouse intestine was reduced for the
flhG
mutant (∼10-fold), indicating that the negative regulation of class I flagellar genes enhances virulence. The
V. cholerae
VC2061 mutant was motile and produced a polar flagellum indistinguishable from that of the wild type, and the transcriptional activities of the four classes of flagellar promoters were similar to that of the wild type. Our results indicate that FlhG and FlhF regulate class I and class III flagellar transcription, respectively, while VC2061 plays no detectable role in
V. cholerae
flagellar biogenesis.
Transcriptional Response ofEscherichia colito External Zinc
Authors: Yamamoto Kaneyoshi , Ishihama Akira
Published: September 2005

Transcriptional response ofEscherichia colito extracellular zinc was studied using DNA microarray and S1 mapping assays. Addition of external zinc induced the expression of zinc exporter ZntA and inhibited the expression of zinc importer ZnuC. In the continuous presence of zinc, ZnuC repression took place at lower zinc concentrations than ZntA induction. The microarray assay indicated that the addition of excess external zinc induces the expression of many genes that are organized in the regulon for cysteine biosynthesis, implying that cysteine plays a role in transient trapping of free zinc for maintenance of zinc homeostasis. Besides the RpoE regulon, other genes were also induced by zinc, suggesting that periplasmic proteins denatured by zinc induce the genes for protein repair. The microarray data of the newly identified zinc-responsive promoters were confirmed by S1 mapping.
Three Chymotrypsin Genes Are Members of the AdpA Regulon in the A-Factor Regulatory Cascade in
Streptomyces griseus
Authors: Tomono Ayami , Tsai Yisan , Ohnishi Yasuo , Horinouchi Sueharu
Published: September 2005

AdpA is a key transcriptional activator in the A-factor regulatory cascade in
Streptomyces griseus
, activating a number of genes required for secondary metabolism and morphological differentiation. Of the five chymotrypsin-type serine protease genes,
sprA
,
sprB
, and
sprD
were transcribed in response to AdpA, showing that these protease genes are members of the AdpA regulon. These proteases were predicted to play the same physiological role, since these protease genes were transcribed in a similar time course during growth and the matured enzymes showed high end-to-end similarity to one another. AdpA bound two sites upstream of the
sprA
promoter approximately at positions −375 and −50 with respect to the transcriptional start point of
sprA
. Mutational analysis of the AdpA-binding sites showed that both AdpA-binding sites were essential for transcriptional activation. AdpA bound a single site at position −50 in front of the
sprB
promoter and greatly enhanced the transcription of
sprB
. The AdpA-binding site at position −40 was essential for transcription of
sprD
, although there was an additional AdpA-binding site at position −180. Most chymotrypsin activity excreted by
S. griseus
was attributed to SprA and SprB, because mutant Δ
sprAB
, having a deletion in both
sprA
and
sprB
, lost almost all chymotrypsin activity, as did mutant Δ
adpA
. Even the double mutant Δ
sprAB
and triple mutant Δ
sprABD
grew normally and developed aerial hyphae and spores over the same time course as the wild-type strain.
Independent and Interchangeable Multimerization Domains of the AbrB, Abh, and SpoVT Global Regulatory Proteins
Authors: Yao Fude , Strauch Mark A.
Published: September 2005

The global regulators AbrB, Abh, and SpoVT are paralogous proteins showing their most extensive sequence homologies in the DNA-binding amino-terminal regions (about 50 residues). The carboxyl-terminal portion of AbrB has been hypothesized to be a multimerization domain with little if any role in DNA-binding recognition or specificity. To investigate the multimerization potentials of the carboxyl-terminal portions of AbrB, Abh, and SpoVT we utilized an in vivo multimerization assay system based upon fusion of the domains to the DNA binding domain of the λ cI repressor protein. The results indicate that the N and C domains of all three paralogues are independent dimerization modules and that the intact Abh and SpoVT proteins are most probably tetramers. Chimeric proteins consisting of the AbrB N-terminal DNA-binding domain fused to the C domain of either Abh or SpoVT are indistinguishable from wild-type AbrB in their ability to regulate an AbrB target promoter in vivo.