Cytoplasmic Ion Concentration Research Articles

Manganese Homeostasis in Bacteria: Interaction of the Small Protein MntS and Manganese Exporter MntP in E. coli

Manganese (Mn) serves as a vital cofactor for bacterial survival, protecting bacteria from reactive oxygen species, radiation, and acidic environments. Bacterial metal ion regulation is associated with pathogenicity, as many virulent strains of bacteria upregulate their ability to obtain metal ions from host species. However, excess metal ions quickly become toxic for most bacteria, resulting in tightly regulated cytoplasmic metal ion concentrations. To better understand regulation of metal levels, our research has focused on the interactions of proteins responsible for Mn homeostasis in Escherichia coli (E. coli), specifically small protein interactions. Small proteins of less than 50 amino acids in length are an emerging group of understudied proteins, yet those that have been studied appear to have important regulatory roles. This research has focused on the small protein called MntS, just 42 amino acids long, which is regulated by the manganese activated transcription factor MntR. In E. coli, MntR represses the production of MntS as well as MntH, a manganese importer, and activates the synthesis of MntP, a manganese exporter. Interestingly, when MntS and MntP are expressed at the same time in the presence of high Mn concentrations, E. coli becomes sensitive to high levels of Mn, suggesting that MntS could interfere with the ability of MntP to export Mn. Indeed, using two‐hybrid assays and co‐purifications we have demonstrated an interaction between MntS and MntP. Additionally, while searching for MntS homologs, we discovered similarity between MntS and the signal peptide (SP) of certain SitA proteins. SitA is a periplasmic protein responsible for cation import, typically Mn. While E. colilacks a SitA protein completely, in the species with the SPs most similar to MntS, the sitAgene is encoded next to mntR. This suggested that the SPs might have similar activity as MntS. We have demonstrated that they produce similar phenotypes in both Mn sensitivity and two‐hybrid assays, suggesting both an evolutionary relationship and a necessary regulatory function. These results provide deeper insight into E. coli’s metal ion regulation, which in the long‐run could allow better treatment of bacterial infections by targeting metal ion regulators.

Adaptation to Varying Salinity in Halomonas elongata: Much More Than Ectoine Accumulation.

The halophilic γ-proteobacterium Halomonas elongata DSM 2581T thrives at salt concentrations well above 10 % NaCl (1.7 M NaCl). A well-known osmoregulatory mechanism is the accumulation of the compatible solute ectoine within the cell in response to osmotic stress. While ectoine accumulation is central to osmoregulation and promotes resistance to high salinity in halophilic bacteria, ectoine has this effect only to a much lesser extent in non-halophiles. We carried out transcriptome analysis of H. elongata grown on two different carbon sources (acetate or glucose), and low (0.17 M NaCl), medium (1 M), and high salinity (2 M) to identify additional mechanisms for adaptation to high saline environments. To avoid a methodological bias, the transcripts were evaluated by applying two methods, DESeq2 and Transcripts Per Million (TPM). The differentially transcribed genes in response to the available carbon sources and salt stress were then compared to the transcriptome profile of Chromohalobacter salexigens, a closely related moderate halophilic bacterium. Transcriptome profiling supports the notion that glucose is degraded via the cytoplasmic Entner-Doudoroff pathway, whereas the Embden-Meyerhoff-Parnas pathway is employed for gluconeogenesis. The machinery of oxidative phosphorylation in H. elongata and C. salexigens differs greatly from that of non-halophilic organisms, and electron flow can occur from quinone to oxygen along four alternative routes. Two of these pathways via cytochrome bo' and cytochrome bd quinol oxidases seem to be upregulated in salt stressed cells. Among the most highly regulated genes in H. elongata and C. salexigens are those encoding chemotaxis and motility proteins, with genes for chemotaxis and flagellar assembly severely downregulated at low salt concentrations. We also compared transcripts at low and high-salt stress (low growth rate) with transcripts at optimal salt concentration and found that the majority of regulated genes were down-regulated in stressed cells, including many genes involved in carbohydrate metabolism, while ribosome synthesis was up-regulated, which is in contrast to what is known from non-halophiles at slow growth. Finally, comparing the acidity of the cytoplasmic proteomes of non-halophiles, extreme halophiles and moderate halophiles suggests adaptation to an increased cytoplasmic ion concentration of H. elongata. Taken together, these results lead us to propose a model for salt tolerance in H. elongata where ion accumulation plays a greater role in salt tolerance than previously assumed.

MtNPF6.5 mediates chloride uptake and nitrate preference in Medicago roots.

The preference for nitrate over chloride through regulation of transporters is a fundamental feature of plant ion homeostasis. We show that Medicago truncatula MtNPF6.5, an ortholog of Arabidopsis thaliana AtNPF6.3/NRT1.1, can mediate nitrate and chloride uptake in Xenopus oocytes but is chloride selective and that its close homologue, MtNPF6.7, can transport nitrate and chloride but is nitrate selective. The MtNPF6.5 mutant showed greatly reduced chloride content relative to wild type, and MtNPF6.5 expression was repressed by high chloride, indicating a primary role for MtNPF6.5 in root chloride uptake. MtNPF6.5 and MtNPF6.7 were repressed and induced by nitrate, respectively, and these responses required the transcription factor MtNLP1. Moreover, loss of MtNLP1 prevented the rapid switch from chloride to nitrate as the main anion in nitrate-starved plants after nitrate provision, providing insight into the underlying mechanism for nitrate preference. Sequence analysis revealed three sub-types of AtNPF6.3 orthologs based on their predicted substrate-binding residues: A (chloride selective), B (nitrate selective), and C (legume specific). The absence of B-type AtNPF6.3 homologues in early diverged plant lineages suggests that they evolved from a chloride-selective MtNPF6.5-like protein.

Ion-Based Cellular Signal Transmission, Principles of Minimum Information Loss, and Evolution by Natural Selection

The Extreme Physical Information EPI principle states that maximum information transmission or, equivalently, a minimum information loss is a fundamental property of nature. Prior work has demonstrated the universal EPI principle allows derivation of nearly all physical laws. Here, we investigate whether EPI can similarly give rise to the fundamental law of life: Evolution. Living systems require information to survive and proliferate. Heritable information in the genome encodes the structure and function of cellular macromolecules but this information remains fixed over time. In contrast, a cell must rapidly and continuously access, analyze, and respond to a wide range of continuously changing spatial and temporal information in the environment. We propose these two information dynamics are linked because the genes encode the structure of the macromolecules that form information conduits necessary for the dynamical interactions with the external environment. However, because the genome does not have the capacity to precisely locate the time and location of external signals, we propose the cell membrane is the site at which most external information is received and processed. In our model, an external signal is detected by gates on transmembrane ion channel and transmitted into the cytoplasm through ions that flow along pre-existing concentration gradients when the gate opens. The resulting cytoplasmic ion “puff” is localized in both time and space, thus producing spatial and temporal information. Small, localized signals in the cytoplasm are “processed” through alterations in the function and location of peripheral membrane proteins. Larger perturbations produce prolonged or spatially extensive changes in cytoplasmic ion concentrations that can be transmitted to other organelles via ion flows along elements of the cytoskeleton. An evolutionary constraint to the ever-increasing acquisition of environmental information is the cost of doing so. One solution to this trade-off is the evolution of information conduits that minimize signal loss during transmission. Since the structures of these conduits are encoded in the genome, evolution of macromolecular conduits that minimize signal loss is linked to and, in fact, governed by a universal principle, termed extreme physical information (EPI). Mathematical analysis of information dynamics based on the flow of ions through membrane channels and along wire-like cytoskeleton macromolecules fulfills the EPI principle. Thus, the empirically derived model of evolution by natural selection, although uniquely applicable to living systems, is theoretically grounded in a universal principle that can also be used to derive the laws of physics. Finally, if minimization of signal loss is a mechanism to overcome energy constraints, the model predicts increasing information and associated complexity are closely linked to increased efficiency of energy production or improved substrate acquisition.

New insights into anhydrobiosis using cellular dielectrophoresis-based characterization.

Late embryogenesis abundant (LEA) proteins are found in desiccation-tolerant species from all domains of life. Despite several decades of investigation, the molecular mechanisms by which LEA proteins confer desiccation tolerance are still unclear. In this study, dielectrophoresis (DEP) was used to determine the electrical properties of Drosophila melanogaster (Kc167) cells ectopically expressing LEA proteins from the anhydrobiotic brine shrimp, Artemia franciscana. Dielectrophoresis-based characterization data demonstrate that the expression of two different LEA proteins, AfrLEA3m and AfrLEA6, increases cytoplasmic conductivity of Kc167 cells to a similar extent above control values. The impact on cytoplasmic conductivity was surprising, given that the concentration of cytoplasmic ions is much higher than the concentrations of ectopically expressed proteins. The DEP data also supported previously reported data suggesting that AfrLEA3m can interact directly with membranes during water stress. This hypothesis was strengthened using scanning electron microscopy, where cells expressing AfrLEA3m were found to retain more circular morphology during desiccation, while control cells exhibited a larger variety of shapes in the desiccated state. These data demonstrate that DEP can be a powerful tool to investigate the role of LEA proteins in desiccation tolerance and may allow to characterize protein-membrane interactions in vivo, when direct observations are challenging.

Divalent cations can control a switch-like behavior in heterotypic and homotypic RNA coacervates

Liquid-liquid phase separation (LLPS) of RNA-protein complexes plays a major role in the cellular function of membraneless organelles (MLOs). MLOs are sensitive to changes in cellular conditions, such as fluctuations in cytoplasmic ion concentrations. To investigate the effect of these changes on MLOs, we studied the influence of divalent cations on the physical and chemical properties of RNA coacervates. Using a model system comprised of an arginine-rich peptide and RNA, we predicted and observed that variations in signaling cations exert interaction-dependent effects on RNA LLPS. Changing the ionic environment has opposing effects on the propensity for heterotypic peptide-RNA and homotypic RNA LLPS, which results in a switch between coacervate types. Furthermore, divalent ion variations continuously tune the microenvironments and fluid properties of heterotypic and homotypic droplets. Our results may provide a general mechanism for modulating the biochemical environment of RNA coacervates in a cellular context.

Resveratrol Directly Controls the Activity of Neuronal Ryanodine Receptors at the Single-Channel Level

Calcium ion dyshomeostasis contributes to the progression of many neurodegenerative diseases and represents a target for the development of neuroprotective therapies, as reported by Duncan et al. (Molecules 15(3):1168-95, 2010), LaFerla (Nat Rev Neurosci 3(11):862-72, 2002), and Niittykoshi et al. (Invest Ophthalmol Vis Sci 51(12):6387-93, 2010). Dysfunctional ryanodine receptors contribute to calcium ion dyshomeostasis and potentially to the pathogenesis of neurodegenerative diseases by generating abnormal calcium ion release from the endoplasmic reticulum, according to Bruno et al. (Neurobiol Aging 33(5):1001 e1-6, 2012) and Stutzmann et al. (J Neurosci 24(2):508-13, 2004). Since ryanodine receptors share functional and structural similarities with potassium channels, as reported by Lanner et al. (Cold Spring Harb Perspect Biol 2(11):a003996, 2010), and small molecules with anti-oxidant properties, such as resveratrol (3,5,4'-trihydroxy-trans-stilbene), directly control the activity of potassium channels, according to Wang et al. (J Biomed Sci 23(1):47, 2016), McCalley et al. (Molecules 19(6):7327-40, 2014), Novakovic et al. (Mol Hum Reprod 21(6):545-51, 2015), Li et al. (Cardiovasc Res 45(4):1035-45, 2000), Gopalakrishnan et al. (Br J Pharmacol 129(7):1323-32, 2000), and Hambrock et al. (J Biol Chem 282(5):3347-56, 2007), we hypothesized that trans-resveratrol can modulate intracellular calcium signaling through direct binding and functional regulation of ryanodine receptors. The goal of our study was to identify and measure the control of ryanodine receptor activity by trans-resveratrol. Mechanisms of calcium signaling mediated by the direct interaction between trans-resveratrol and ryanodine receptors were identified and measured with single-channel electrophysiology. Addition of trans-resveratrol to the cytoplasmic face of the ryanodine receptor increased single-channel activity at physiological and elevated pathophysiological cytoplasmic calcium ion concentrations. The open probability of the channel increases after interacting with the small molecule in a dose-dependent manner, but remains also dependent on the concentration of its physiological ligand, cytoplasmic-free calcium ions. This study provides the first evidence of a direct functional interaction between trans-resveratrol and ryanodine receptors. Such functional control of ryanodine receptors by trans-resveratrol as a novel mechanism of action could provide additional rationales for the development of novel therapeutic strategies to treat and prevent neurodegenerative diseases.

The Role of Cell Membrane Information Reception, Processing, and Communication in the Structure and Function of Multicellular Tissue.

Investigations of information dynamics in eukaryotic cells focus almost exclusively on heritable information in the genome. Gene networks are modeled as “central processors” that receive, analyze, and respond to intracellular and extracellular signals with the nucleus described as a cell’s control center. Here, we present a model in which cellular information is a distributed system that includes non-genomic information processing in the cell membrane that may quantitatively exceed that of the genome. Within this model, the nucleus largely acts a source of macromolecules and processes information needed to synchronize their production with temporal variations in demand. However, the nucleus cannot produce microsecond responses to acute, life-threatening perturbations and cannot spatially resolve incoming signals or direct macromolecules to the cellular regions where they are needed. In contrast, the cell membrane, as the interface with its environment, can rapidly detect, process, and respond to external threats and opportunities through the large amounts of potential information encoded within the transmembrane ion gradient. Our model proposes environmental information is detected by specialized protein gates within ion-specific transmembrane channels. When the gate receives a specific environmental signal, the ion channel opens and the received information is communicated into the cell via flow of a specific ion species (i.e., K+, Na+, Cl−, Ca2+, Mg2+) along electrochemical gradients. The fluctuation of an ion concentration within the cytoplasm adjacent to the membrane channel can elicit an immediate, local response by altering the location and function of peripheral membrane proteins. Signals that affect a larger surface area of the cell membrane and/or persist over a prolonged time period will produce similarly cytoplasmic changes on larger spatial and time scales. We propose that as the amplitude, spatial extent, and duration of changes in cytoplasmic ion concentrations increase, the information can be communicated to the nucleus and other intracellular structure through ion flows along elements of the cytoskeleton to the centrosome (via microtubules) or proteins in the nuclear membrane (via microfilaments). These dynamics add spatial and temporal context to the more well-recognized information communication from the cell membrane to the nucleus following ligand binding to membrane receptors. Here, the signal is transmitted and amplified through transduction by the canonical molecular (e.g., Mitogen Activated Protein Kinases (MAPK) pathways. Cytoplasmic diffusion allows this information to be broadly distributed to intracellular organelles but at the cost of loss of spatial and temporal information also contained in ligand binding.

Signal transmission through elements of the cytoskeleton form an optimized information network in eukaryotic cells

Multiple prior empirical and theoretical studies have demonstrated wire-like flow of electrons and ions along elements of the cytoskeleton but this has never been linked to a biological function. Here we propose that eukaryotes use this mode of signal transmission to convey spatial and temporal environmental information from the cell membrane to the nucleus. The cell membrane, as the interface between intra- and extra-cellular environments, is the site at which much external information is received. Prior studies have demonstrated that transmembrane ion gradients permit information acquisition when an environmental signal interacts with specialized protein gates in membrane ion channels and producing specific ions to flow into or out of the cell along concentration gradients. The resulting localized change in cytoplasmic ion concentrations and charge density can alter location and enzymatic function of peripheral membrane proteins. This allows the cell to process the information and rapidly deploy a local response. Here we investigate transmission of information received and processed in and around the cell membrane by elements of the cytoskeleton to the nucleus to alter gene expression. We demonstrate signal transmission by ion flow along the cytoskeleton is highly optimized. In particular, microtubules, with diameters of about 30 nm, carry coarse-grained Shannon information to the centrosome adjacent to the nucleus with minimum loss of input source information. And, microfilaments, with diameters of about 4 nm, transmit maximum Fisher (fine-grained) information to protein complexes in the nuclear membrane. These previously unrecognized information dynamics allow continuous integration of spatial and temporal environmental signals with inherited information in the genome.

Exploring the relationship between cytoplasmic ion content variation and multidrug resistance in cancer cells via ion-release based impedance spectroscopy

This study aimed to measure and compare the cytoplasmic ion concentrations of the parental cells and their drug resistant progenies. High-level laboratory and clinically-relevant multidrug resistance (MDR) cell line models were developed and investigated for K562 and CCRF-CEM cells, respectively. Measurements were achieved using a modified version of ion release-based impedance spectroscopy (IRbIS), where the average total ion concentration per cell was determined by measuring the variations in solution conductance after the lysis of a population with a known initial cell count. Results showed that the average of cytoplasmic ion concentration for K562/imaR cells was almost 1.8 times higher than that of K562/wt cells with a standard error of mean (SEM) lower than 7%, having a significant difference at p = 0.05 level. CCRF-CEM/doxR cells had almost a 1.2-fold increase in the ion concentration compared with CCRF-CEM/wt cells with an SEM smaller than 11%. Unlike K562 cells, this was not a significant variation at p = 0.05 level. Having insignificant SEMs proved the validity of this modified implementation of IRbIS measurements. We showed that our new approach enables to identify variations in cytoplasmic ion concentration per cell and to understand the mechanisms driving the toxicity and the development of drug resistance to chemotherapeutic agents.

Induced cell death in Ceratocystis fimbriata by pro‐apoptotic activity of a natural organic compound, perillaldehyde, through Ca2+ overload and accumulation of reactive oxygen species

Black rot caused by the fungus Ceratocystis fimbriata causes notable losses in sweet potato production. Perillaldehyde (PAE), a secondary metabolite in perilla, was studied to determine its antifungal effects on mycelial growth and spore germination of C. fimbriata. The effects of PAE on cell wall integrity and cell membrane permeability were also investigated. To elucidate the possible mechanisms of cell death triggered by PAE, sensitivity of C. fimbriata to PAE toxicity was determined by cytoplasmic and mitochondrial calcium ion concentrations ([Ca2+]c and [Ca2+]m), reactive oxygen species (ROS), mitochondrial membrane potential (MMP), cytochrome c (cyt c) release, metacaspase activation, phosphatidylserine (PS) externalization and DNA fragmentation. The results suggest that mycelial growth and spore germination were inhibited by PAE in a dose‐dependent manner. Ceratocystis fimbriata spores treated with PAE experienced dramatic Ca2+ overload and elevated ROS production. Compared to untreated controls, the proportion of fluorescent cells stained with the ROS indicator DCFH‐DA and treated with a range of PAE concentrations from 0.0625 to 0.50 μL mL−1 increased by 2.9 ± 0.79% to 27.1 ± 0.38%. Ca2+ overload and ROS accumulation induced depolarization of the MMP, contributing to mitochondrial dysfunction. Cyt c was released from the mitochondria to the cytosol, triggering metacaspase activation. The significant antifungal activity of PAE on C. fimbriata was demonstrated by these studies, suggesting that PAE has the potential for wide application to postharvest management of tuber crops, in addition to the application to above‐ground fruit and vegetables that have been previously investigated.

Protein Charge and Ion Concentration Affect Protein Diffusion in the Prokaryotic Cytoplasm

Contrary to eukaryotic cells, both bacteria and archaea (mostly) lack active cytoplasmic transport. The encounter rate between proteins in the cytoplasm thus depends (almost) exclusively on lateral diffusion. Previously, it was found that the cytoplasmic protein diffusion coefficient drops with an increase of (1) its molecular weight and (2) cytoplasmic excluded volume. Here we show the dependence of the protein diffusion coefficient on the strength of binding interactions. We have done FRAP measurements to obtain diffusion coefficients of surface-modified GFPs in Escherichia coli, Lactococcus lactis and Haloferax volcanii. The tested variants of GFP vary in net charge from −30 to +25. The examined organisms differ in their cytoplasmic ion concentration. The GFP diffusion coefficient drops with increasing positive charge, up to two orders of magnitude. This is caused by binding of the GFPs to ribosomes. The drop in the diffusion coefficient decreases with increased cytoplasmic ion concentration. Our findings suggest a limit to what the surface charge of a protein can be in the cytoplasm of a given cell.

CLIC1 Inhibition Attenuates Vascular Inflammation, Oxidative Stress, and Endothelial Injury.

Endothelial dysfunction, which includes endothelial oxidative damage and vascular inflammation, is a key initiating step in the pathogenesis of atherosclerosis (AS) and an independent risk factor for this disorder. Intracellular chloride channel 1 (CLIC1), a novel metamorphic protein, acts as a sensor of cell oxidation and is involved in inflammation. In this study, we hypothesize that CLIC1 plays an important role in AS. Apolipoprotein E-deficient mice were supplied with a normal diet or a high-fat and high-cholesterol diet for 8 weeks. Overexpressed CLIC1 was associated with the accelerated atherosclerotic plaque development, amplified oxidative stress, and in vivo release of inflammatory cytokines. We subsequently examined the underlying molecular mechanisms through in vitro experiments. Treatment of cultured human umbilical vein endothelial cells (HUVECs) with H2O2 induced endothelial oxidative damage and enhanced CLIC1 expression. Suppressing CLIC1 expression through gene knocked-out (CLIC1−/−) or using the specific inhibitor indanyloxyacetic acid-94 (IAA94) reduced ROS production, increased SOD enzyme activity, and significantly decreased MDA level. CLIC1−/− HUVECs exhibited significantly reduced expression of TNF-α and IL-1β as well as ICAM-1 and VCAM-1 at the protein levels. In addition, H2O2 promoted CLIC1 translocation to the cell membrane and insertion into lipid membranes, whereas IAA94 inhibited CLIC1 membrane translocation induced by H2O2. By contrast, the majority of CLIC1 did not aggregate on the cell membrane in normal HUVECs, and this finding is consistent with the changes in cytoplasmic chloride ion concentration. This study demonstrates for the first time that CLIC1 is overexpressed during AS development both in vitro and in vivo and can regulate the accumulation of inflammatory cytokines and production of oxidative stress. Our results also highlight that deregulation of endothelial functions may be associated with the membrane translocation of CLIC1 and active chloride-selective ion channels in endothelial cells.

Comparative chemical genomics reveal that the spiroindolone antimalarial KAE609 (Cipargamin) is a P-type ATPase inhibitor.

The spiroindolones, a new class of antimalarial medicines discovered in a cellular screen, are rendered less active by mutations in a parasite P-type ATPase, PfATP4. We show here that S. cerevisiae also acquires mutations in a gene encoding a P-type ATPase (ScPMA1) after exposure to spiroindolones and that these mutations are sufficient for resistance. KAE609 resistance mutations in ScPMA1 do not confer resistance to unrelated antimicrobials, but do confer cross sensitivity to the alkyl-lysophospholipid edelfosine, which is known to displace ScPma1p from the plasma membrane. Using an in vitro cell-free assay, we demonstrate that KAE609 directly inhibits ScPma1p ATPase activity. KAE609 also increases cytoplasmic hydrogen ion concentrations in yeast cells. Computer docking into a ScPma1p homology model identifies a binding mode that supports genetic resistance determinants and in vitro experimental structure-activity relationships in both P. falciparum and S. cerevisiae. This model also suggests a shared binding site with the dihydroisoquinolones antimalarials. Our data support a model in which KAE609 exerts its antimalarial activity by directly interfering with P-type ATPase activity.

Research progress on ion channels in human glioma

Glioma is a serious threat to human health. Increasing evidences indicate that abnormal expression and activities of a number of ion channels, e.g. voltage-gated K+, Ca2+, Cl- channels, are involved in the growth, proliferation, migration and invasion of glioma cells. Ion channels may affect the occurrence and development of glioma cells through regulating the membrane potential, adjusting the cytoplasmic volume, ion concentration, and pH value. In-depth understanding the role of ion channels in formation, development and transfer process of glioma may provide new targets for glioma prevention and treatment. In this paper, the advanced studies of ion channels in glioma were briefly reviewed. Key words: Glioma; Ion channels; Abnormal expression

Calcium signals and oocyte maturation in marine invertebrates.

In various oocytes and eggs of animals, transient elevations in cytoplasmic calcium ion concentrations are known to regulate key processes during fertilization and the completion of meiosis. However, whether or not calcium transients also help to reinitiate meiotic progression at the onset of oocyte maturation remains controversial. This article summarizes reports of calcium signals playing essential roles during maturation onset (=germinal vesicle breakdown, GVBD) in several kinds of marine invertebrate oocytes. Conversely, other data from the literature, as well as previously unpublished findings for jellyfish oocytes, fail to support the view that calcium signals are required for GVBD. In addition to assessing the effects of calcium transients on GVBD in marine invertebrate oocytes, the ability of maturing oocytes to enhance their calcium-releasing capabilities after GVBD is also reviewed. Furthermore, possible explanations are proposed for the contradictory results that have been obtained regarding calcium signals during oocyte maturation in marine invertebrates.

Distribution pattern of Caragana species under the influence of climate gradient in the Inner Mongolia region, China

There is a strong climate gradient in the Inner Mongolia region, China, with solar radiation and air temperature increasing but precipitation decreasing gradually from the northeast to the southwest. Sixteen Caragana species exist in the Inner Mongolia region. These Caragana species exhibit a distribution pattern across moisture zones and form a geographical replacement series. In order to examine the mechanisms responsible for Caragana species distribution pattern, we selected 12 Caragana species that exhibit a distinct distribution pattern across multiple moisture zones in the Inner Mongolia region, and determined the relationships between the leaf ecological and physiological traits of these Caragana species and the aridity index and solar radiation. Along with the climatic drought gradient and the solar radiation intensification from the northeast to the southwest, leaf ecological characteristics of Caragana species changed drastically, i.e. the leaf shape gradually turned from flat into tegular or tubbish; the leaf hair became denser, longer and lighter in color; the leaf area, leaf biomass and specific leaf area (SLA) decreased significantly; the leaf thickness and the ratio of leaf thickness to leaf area increased significantly; and the leaf chlorophyll content decreased significantly. As the climatic drought stress increased, osmotic potentials of the main osmotic adjustment substances and the cytoplasmic ion concentration of Caragana species increased significantly. Meanwhile, the total and free water contents and water potential of leaves decreased significantly; the ratio of bound to free water increased significantly; the stomatal conductance and transpiration rate reduced significantly; and the water use efficiency (WUE) increased significantly. In addition, with the intensification of climatic drought stress, peroxidase (POD) and superoxide dismutase (SOD) activities in leaves increased significantly. As a result, the malondialdehyde (MDA) content increased while the oxygen free radical content decreased. Our results showed that most of the leaf ecological and physiological traits of the 12 Caragana species varied in accordance with the climatic drought gradient in the Inner Mongolia region, which reflected the adaptation of the Caragana species to the local climate conditions. With relatively more active metabolism and faster growth, the Caragana species in the northeast had strong competitive abilities; on the other hand, with stronger resistance to climatic drought stress, the Caragana species in the southwest could survive in harsh environments. Based on our results, we concluded that both the environmental gradients and the adaptive responses of Caragana species to their environments played important roles in the formation of the Caragana species distribution pattern across the Inner Mongolia region.

“Trilongins” Offer Insight into Mold Toxicity

“Trilongins” Offer Insight into Mold Toxicity

The FlatwormMacrostomum lignanoIs a Powerful Model Organism for Ion Channel and Stem Cell Research

Bioelectrical signals generated by ion channels play crucial roles in many cellular processes in both excitable and nonexcitable cells. Some ion channels are directly implemented in chemical signaling pathways, the others are involved in regulation of cytoplasmic or vesicular ion concentrations, pH, cell volume, and membrane potentials. Together with ion transporters and gap junction complexes, ion channels form steady-state voltage gradients across the cell membranes in nonexcitable cells. These membrane potentials are involved in regulation of such processes as migration guidance, cell proliferation, and body axis patterning during development and regeneration. While the importance of membrane potential in stem cell maintenance, proliferation, and differentiation is evident, the mechanisms of this bioelectric control of stem cell activity are still not well understood, and the role of specific ion channels in these processes remains unclear. Here we introduce the flatworm Macrostomum lignano as a versatile model organism for addressing these topics. We discuss biological and experimental properties of M. lignano, provide an overview of the recently developed experimental tools for this animal model, and demonstrate how manipulation of membrane potential influences regeneration in M. lignano.

Temporary Sequestration of Potassium by Mitochondria in Astrocytes

Increases in extracellular potassium concentration ([K(+)](o)), which can occur during neuronal activity and under pathological conditions such as ischemia, lead to a variety of potentially detrimental effects on neuronal function. Although astrocytes are known to contribute to the clearance of excess K(+)(o), the mechanisms are not fully understood. We examined the potential role of mitochondria in sequestering K(+) in astrocytes. Astrocytes were loaded with the fluorescent K(+) indicator PBFI and release of K(+) from mitochondria into the cytoplasm was examined after uncoupling the mitochondrial membrane potential with carbonyl cyanide m-chlorophenylhydrazone (CCCP). Under the experimental conditions employed, transient applications of elevated [K(+)](o) led to increases in K(+) within mitochondria, as assessed by increases in the magnitudes of cytoplasmic [K(+)] ([K(+)](i)) transients evoked by brief exposures to CCCP. When mitochondrial K(+) sequestration was impaired by prolonged application of CCCP, there was a robust increase in [K(+)](i) upon exposure to elevated [K(+)](o). Blockade of plasmalemmal K(+) uptake routes by ouabain, Ba(2+), or a mixture of voltage-activated K(+) channel inhibitors reduced K(+) uptake into mitochondria. Also, reductions in mitochondrial K(+) uptake occurred in the presence of mito-K(ATP) channel inhibitors. Rises in [K(+)](i) evoked by brief applications of CCCP following exposure to high [K(+)](o) were also reduced by gap junction blockers and in astrocytes isolated from connexin43-null mice, suggesting that connexins also play a role in K(+) uptake into astrocyte mitochondria. We conclude that mitochondria play a key role in K(+)(o) handling by astrocytes.