Search

Proprioception of all animals is important in being able to have coordinated locomotion. Stretch activated ion channels (SACs) transduce the mechanical force into electrical signals in the proprioceptive sensory endings. The types of SACs vary among sensory neurons in animals as defined by pharmacological, physiological and molecular identification. The chordotonal organs within insects and crustaceans offer a unique ability to investigate proprioceptive function. The effects of the extracellular environment on neuronal activity, as well as the function of associated SACs are easily accessible and viable in minimal saline for ease in experimentation. The effect of extracellular [Ca2+] on membrane properties which affect voltage-sensitivity of ion channels, threshold of action potentials and SACs can be readily addressed in the chordotonal organ in crab limbs. It is of interest to understand how low extracellular [Ca2+] enhances neural activity considering the SACs in the sensory endings could possibly be Ca2+ channels and that all neural activity is blocked with Mn2+. It is suggested that axonal excitability might be affected independent from the SAC activity due to potential presence of calcium activated potassium channels (K(Ca)) and the ability of Ca2+ to block voltage gated Na+ channels in the axons. Separating the role of Ca2+ on the function of the SACs and the excitability of the axons in the nerves associated with chordotonal organs is addressed. These experiments may aid in understanding the mechanisms of neuronal hyperexcitability during hypocalcemia within mammals.

Purpose North Carolina macular dystrophy (NCMD) is an autosomal dominant, congenital disorder affecting the central retina. Here, we report clinical and genetic findings in three families segregating NCMD and use epigenomic datasets from human tissues to gain insights into the effect of NCMD-implicated variants. Methods Clinical assessment and genetic testing were performed. Publicly available transcriptomic and epigenomic datasets were analyzed and the activity-by-contact method for scoring enhancer elements and linking them to target genes was used. Results A previously described, heterozygous, noncoding variant upstream of the PRDM13 gene was detected in all six affected study participants (chr6:100,040,987G>C [GRCh37/hg19]). Interfamilial and intrafamilial variability were observed; the visual acuity ranged from 0.0 to 1.6 LogMAR and fundoscopic findings ranged from visually insignificant, confluent, drusen-like macular deposits to coloboma-like macular lesions. Variable degrees of peripheral retinal spots (which were easily detected on widefield retinal imaging) were observed in all study subjects. Notably, a 6-year-old patient developed choroidal neovascularization and required treatment with intravitreal bevacizumab injections. Computational analysis of the five single nucleotide variants that have been implicated in NCMD revealed that these noncoding changes lie within two putative enhancer elements; these elements are predicted to interact with PRDM13 in the developing human retina. PRDM13 was found to be expressed in the fetal retina, with greatest expression in the amacrine precursor cell population. Conclusions We provide further evidence supporting the role of PRDM13 dysregulation in the pathogenesis of NCMD and highlight the usefulness of widefield retinal imaging in individuals suspected to have this condition.

PurposeNorth Carolina macular dystrophy (NCMD) is an autosomal dominant, congenital disorder affecting the central retina. Here, we report clinical and genetic findings in three families segregating NCMD and use epigenomic datasets from human tissues to gain insights into the effect of NCMD-implicated variants.MethodsClinical assessment and genetic testing were performed. Publicly-available transcriptomic and epigenomic datasets were analyzed and the ‘Activity-by-Contact’ (ABC) method for scoring enhancer elements and linking them to target genes was used.ResultsA previously-described, heterozygous, non-coding variant upstream of the PRDM13 gene was detected in all six affected study participants (chr6:100,040,987G>C [GRCh37/hg19]). Inter- and intra-familial variability were observed; the visual acuity ranged from 0.0 to 1.6 LogMAR and fundoscopic findings ranged from visually insignificant, confluent, drusen-like macular deposits to coloboma-like macular lesions. Variable degrees of peripheral retinal spots (which were easily detected on widefield retinal imaging) were observed in all study subjects. Notably, a 6-year-old patient developed choroidal neovascularization and required treatment with intravitreal bevacizumab injections. Computational analysis of the five single nucleotide variants that have been implicated in NCMD revealed that these non-coding changes lie within two putative enhancer elements; these elements are predicted to interact with PRDM13 in the developing human retina. PRDM13 was found to be expressed in the fetal retina, with highest expression in the amacrine precursor cell population.ConclusionsWe provide further evidence supporting the role of PRDM13 dysregulation in the pathogenesis of NCMD and highlight the utility of widefield retinal imaging in individuals suspected to have this condition.

This study presents a new framework for extracting single ion diffusion coefficients in ion exchange membranes from experimental ion sorption, salt permeability, and ionic conductivity data. The framework was used to calculate cation and anion diffusion coefficients in a series of commercial ion exchange membranes contacted by aqueous NaCl solutions. Counterion diffusion coefficients were greater than co-ion diffusion coefficients for all membranes after accounting for inherent differences due to ion size. A model for ion diffusion coefficients in ion exchange membranes, incorporating ideas from counterion condensation theory, was proposed to interpret the experimental results. The model predicted co-ion diffusion coefficients reasonably well with no adjustable parameters, while a single adjustable parameter was required to accurately describe counterion diffusion coefficients. The results suggest that for cross-linked ion exchange membranes in which counterion condensation occurs condensed counterions mi...

Myotonic dystrophy type 1 (DM1) is a multisystemic disease caused by an expanded CTG repeat in the 3′ UTR of the dystrophia myotonica protein kinase (DMPK) gene. Short, DNA-based antisense oligonucleotides termed gapmers are a promising strategy to degrade toxic CUG expanded repeat (CUGexp) RNA. Nucleoside analogs are incorporated to increase gapmer affinity and stability; however, some analogs also exhibit toxicity. In this study, we demonstrate that the 2′,4′-BNANC[NMe] (BNANC) modification is a promising nucleoside analog with high potency similar to 2′,4′-LNA (LNA). BNANC gapmers targeting a nonrepetitive region of the DMPK 3′ UTR show allele-specific knockdown of CUGexp RNA and revert characteristic DM1 molecular defects including mis-splicing and accumulation of RNA foci. Notably, the BNANC gapmers tested in this study did not induce caspase activation, in contrast to a sequence matched LNA gapmer. This study indicates that BNANC gapmers warrant further study as a promising RNA targeting therapeutic.

Accurate evaluation of salt diffusion coefficients from transport rate data in ion exchange membranes requires accounting for frame of reference and non-ideal thermodynamic effects. Due to a lack of models and experimental data quantifying membrane ion activity coefficients, it has been impossible to evaluate the impact of non-ideal thermodynamic effects on observed salt diffusion coefficients. Here, a framework is presented that includes both frame of reference (i.e., convection) and non-ideal thermodynamic effects in calculating salt diffusion coefficients in ion exchange membranes. Effective concentration averaged NaCl diffusion coefficients were determined as a function of upstream NaCl concentration in commercial ion exchange membranes from NaCl permeability and sorption measurements via the solution-diffusion model. Frame of reference effects were evaluated using a version of Fick's law that accounts for convection. The factors necessary to account for non-ideal thermodynamic effects were developed using Manning's counter-ion condensation theory. At low upstream NaCl concentrations, frame of reference and non-ideal thermodynamic effects on diffusion coefficients were negligible. However, at higher upstream NaCl concentrations (e.g., >0.1 M), both effects contribute measurably to NaCl diffusion coefficients. Correcting for frame of reference effects increased apparent NaCl diffusion coefficients. However, correcting for thermodynamic non-idealities of the ions sorbed into the membranes reduced apparent NaCl diffusion coefficients. Fortuitously, for the materials considered in this study, frame of reference and non-ideal thermodynamic effects nearly cancel each other.

Heading perception in primates depends heavily on visual optic-flow cues. Yet during self-motion, heading percepts remain stable even though smooth-pursuit eye movements often distort optic flow. Electrophysiological studies have identified visual areas in monkey cortex, including the dorsal medial superior temporal area (MSTd), that signal the true heading direction during pursuit. According to theoretical work, self-motion can be represented accurately by compensating for these distortions in two ways: via retinal mechanisms or via extraretinal efference-copy signals, which predict the sensory consequences of movement. Psychophysical evidence strongly supports the efference-copy hypothesis, but physiological evidence remains inconclusive. Neurons that signal the true heading direction during pursuit are found in visual areas of monkey cortex, including the dorsal medial superior temporal area (MSTd). Here we measured heading tuning in MSTd using a novel stimulus paradigm, in which we stabilize the optic-flow stimulus on the retina during pursuit. This approach isolates the effects on neuronal heading preferences of extraretinal signals, which remain active while the retinal stimulus is prevented from changing. Our results demonstrate a significant but small influence of extraretinal signals on the preferred heading directions of MSTd neurons. Under our stimulus conditions, which are rich in retinal cues, we find that retinal mechanisms dominate physiological corrections for pursuit eye movements, suggesting that extraretinal cues, such as predictive efference-copy mechanisms, have a limited role under naturalistic conditions.Significance StatementSensory systems discount stimulation caused by the animal’s own behavior. For example, eye movements cause irrelevant retinal signals that could interfere with motion perception. The visual system compensates for such self-generated motion, but how this happens is unclear. Two theoretical possibilities are a purely visual calculation or one using an internal signal of eye movements to compensate for their effects. Such a signal can be isolated by experimentally stabilizing the image on a moving retina, but this approach has never been adopted to study motion physiology. Using this method, we find that eye-movement signals have little influence on neural activity in visual cortex, while feed-forward visual calculation has a strong effect and is likely important under real-world conditions.

The dynamic process underlying muscle contraction is the parallel sliding of thin actin filaments along an immobile thick myosin fiber powered by oar-like movements of protruding myosin cross bridges (myosin heads). The free energy for functioning of the myosin nanomotor comes from the hydrolysis of ATP bound to the myosin heads. The unit step of translational movement is based on a mechanical-chemical cycle involving ATP binding to myosin, hydrolysis of the bound ATP with ultimate release of the hydrolysis products, stress-generating conformational changes in the myosin cross bridge, and relief of built-up stress in the myosin power stroke. The cycle is regulated by a transition between weak and strong actin-myosin binding affinities. The dissociation of the weakly bound complex by addition of salt indicates the electrostatic basis for the weak affinity, while structural studies demonstrate that electrostatic interactions among negatively charged amino acid residues of actin and positively charged residues of myosin are involved in the strong binding interface. We therefore conjecture that intermediate states of increasing actin-myosin engagement during the weak-to-strong binding transition also involve electrostatic interactions. Methods of polymer solution physics have shown that the thin actin filament can be regarded in some of its aspects as a net negatively charged polyelectrolyte. Here we employ polyelectrolyte theory to suggest how actin-myosin electrostatic interactions might be of significance in the intermediate stages of binding, ensuring an engaged power stroke of the myosin motor that transmits force to the actin filament, and preventing the motor from getting stuck in a metastable pre-power stroke state. We provide electrostatic force estimates that are in the pN range known to operate in the cycle.

The equilibrium volume of a polyelectrolyte gel results from a balance between the tendency to swell caused by outbound polymer/counterion diffusion along with Coulomb interactions on the one hand; and, on the other, the elastic resilience of the cross-linked polymer network. Direct Coulomb forces contribute both to non-ideality of the equilibrated Donnan osmotic pressure, but also to stretching of the network. To isolate the effect of polyelectrolyte expansion, we have analyzed a “one-dimensional” version of a gel, a linear chain of charged beads connected by Hooke’s law springs. As in the range of weak Coulomb strengths previously studied, the springs are significantly stretched by the repulsive interactions among the beads even when the Coulomb strength is strong enough to cause condensation of counterions. There is a quasi-abrupt transition from a stretched state to a partially collapsed state in a transition range between weak and strong Coulomb strengths. Fluctuations between stretched and contracted conformations occur within the transition range. As the solvent quality decreases past the transition range, a progressive collapse can result if the condensed counterions strengthen the spring constant.

We examine twist-stretch coupling of unconstrained DNA using polyelectrolyte theory as applied to a line-charge model along with published data on the ionic-strength dependence of the twist angle. We conclude that twist-stretch coupling is negative: environmental changes that stretch free DNA, unconstrained by externally applied pulling or twisting forces, are accompanied by unwinding of the double helix. We also analyze a helical model and conclude that the observed unwinding of the DNA helix when ionic strength is decreased is driven by radial swelling of the helix.

The physical principle underlying the familiar condensation transition from vapor to liquid is the competition between the energetic tendency to condense owing to attractive forces among molecules of the fluid and the entropic tendency to disperse toward the maximum volume available as limited only by the walls of the container. Van der Waals incorporated this principle into his equation of state and was thus able to explain the discontinuous nature of condensation as the result of instability of intermediate states. The volume phase transition of gels, also discontinuous in its sharpest manifestation, can be understood similarly, as a competition between net free energy attraction of polymer segments and purely entropic dissolution into a maximum allowed volume. Viewed in this way, the gel phase transition would require nothing more to describe it than van der Waals’ original equation of state (with osmotic pressure Π replacing pressure P). But the polymer segments in a gel are networked by cross-links, and a consequent restoring force prevents complete dissolution. Like a solid material, and unlike a van der Waals fluid, a fully swollen gel possesses an intrinsic volume of its own. Although all thermodynamic descriptions of gel behavior contain an elastic component, frequently in the form of Flory-style rubber theory, the resulting isotherms usually have the same general appearance as van der Waals isotherms for fluids, so it is not clear whether the solid-like aspect of gels, that is, their intrinsic volume and shape, adds any fundamental physics to the volume phase transition of gels beyond what van der Waals already knew. To address this question, we have constructed a universal chemical potential for gels that captures the volume transition while containing no quantities specific to any particular gel. In this sense, it is analogous to the van der Waals theory of fluids in its universal form, but although it incorporates the van der Waals universal equation of state, it also contains a network elasticity component, not based on Flory theory but instead on a nonlinear Langevin model, that restricts the radius of a fully swollen spherical gel to a solid-like finite universal value of unity, transitioning to a value less than unity when the gel collapses. A new family of isotherms arises, not present in a preponderately van der Waals analysis, namely, profiles of gel density as a function of location in the gel. There is an abrupt onset of large amplitude density fluctuations in the gel at a critical temperature. Then, at a second critical temperature, the entire swollen gel collapses to a high-density phase.

We formulate and analyze a minimal model, based on condensation theory, of the lamellar cationic lipid (CL)-DNA complex of alternately charged lipid bilayers and DNA monolayers in a salt solution. Each lipid bilayer, composed by a random mixture of cationic and neutral lipids, is assumed to be a rigid uniformly charged plane. Each DNA monolayer, located between two lipid bilayers, is formed by the same number of parallel DNAs with a uniform separation distance. For the electrostatic calculation, the model lipoplex is collapsed to a single plane with charge density equal to the net lipid and DNA charge. The free energy difference between the lamellar lipoplex and a reference state of the same number of free lipid bilayers and free DNAs, is calculated as a function of the fraction of CLs, of the ratio of the number of CL charges to the number of negative charges of the DNA phosphates, and of the total number of planes. At the isoelectric point the free energy difference is minimal. The complex formation, already favoured by the decrease of the electrostatic charging free energy, is driven further by the free energy gain due to the release of counterions from the DNAs and from the lipid bilayers, if strongly charged. This minimal model compares well with experiment for lipids having a strong preference for planar geometry and with major features of more detailed models of the lipoplex.

This study presents a framework for predicting salt permeability coefficients in ion exchange membranes in contact with an aqueous salt solution. The model, based on the solution–diffusion mechanism, was tested using experimental salt permeability data for a series of commercial ion exchange membranes. Equilibrium salt partition coefficients were calculated using a thermodynamic framework (i.e., Donnan theory), incorporating Manning’s counterion condensation theory to calculate ion activity coefficients in the membrane phase and the Pitzer model to calculate ion activity coefficients in the solution phase. The model predicted NaCl partition coefficients in a cation exchange membrane and two anion exchange membranes, as well as MgCl2 partition coefficients in a cation exchange membrane, remarkably well at higher external salt concentrations (>0.1 M) and reasonably well at lower external salt concentrations (

A Monte Carlo code applied to the cgDNA coarse-grain rigid-base model of B-form double-stranded DNA is used to predict a sequence-averaged persistence length of lF = 53.5 nm in the sense of Flory, and of lp = 160 bp or 53.5 nm in the sense of apparent tangent–tangent correlation decay. These estimates are slightly higher than the consensus experimental values of 150 bp or 50 nm, but we believe the agreement to be good given that the cgDNA model is itself parametrized from molecular dynamics simulations of short fragments of length 10–20 bp, with no explicit fit to persistence length. Our Monte Carlo simulations further predict that there can be substantial dependence of persistence lengths on the specific sequence S of a fragment. We propose, and confirm the numerical accuracy of, a simple factorization that separates the part of the apparent tangent–tangent correlation decay lp(S) attributable to intrinsic shape, from a part ld(S) attributable purely to stiffness, i.e., a sequence-dependent version of wh...

We give approximate solutions for some problems in polyelectrolyte theory involving nonuniform charge distributions. Distributions of condensed counterions and electrostatic free energies are found for line-charge models of oligomers, end segments, junctions separating regions of different charge densities, and two semi-infinite line charges approaching each other end-on. The end-on potential of mean force in the latter case is screened by the Debye length. With that exception, we find little correlation between the Debye screening length and the much smaller distances marking the onset of significant end effects. We suggest a biological application to DNA unwinding by helicases.

Equilibrium partitioning of ions between a membrane and a contiguous external solution strongly influences transport properties of polymeric membranes used for water purification and energy generation applications. This study presents a theoretical framework to quantitatively predict ion sorption from aqueous electrolytes (e.g., NaCl, MgCl2) into charged (i.e., ion exchange) polymers. The model was compared with experimental NaCl, MgCl2, and CaCl2 sorption data in commercial cation and anion exchange membranes. Ion sorption in charged polymers was modeled using a thermodynamic approach based on Donnan theory coupled with Manning's counter-ion condensation theory to describe non-ideal behavior of ions in the membrane. Ion activity coefficients in solution were calculated using the Pitzer model. The resulting model, with no adjustable parameters, provides remarkably good agreement with experimental values of membrane mobile salt concentration. The generality of the model was further demonstrated using literature data for ion sorption of various electrolytes in charged polymers, including HCl sorption in Nafion.

In a previous paper, we have shown that counterions condense on charged spheres and planes as well as on the linear polyelectrolyte models of charged lines and cylinders, provided that the surface charge densities exceed a critical value. Here we give a more complete calculation resulting in the electrostatic free energies of all of these geometric shapes. We consider a macroion immersed in salt solution and also a macroion in the presence only of its own counterions. For a sphere, cylinder, and plane with equal surface charge densities, the sphere has the lowest electrostatic free energy and the plane the highest. We apply the results to the coil-to-globule transition and to the growth of spheres by fusion of two smaller spheres to form a larger one.

We give a contemporary and direct derivation of a classical, but insufficiently familiar, result in the theory of linear elasticity-a representation for the energy of a stressed elastic rod with central axis that intrinsically takes the shape of a general space curve. We show that the geometric torsion of the space curve, while playing a crucial role in the bending energy, is physically unrelated to the elastic twist. We prove that the twist energy vanishes in the lowest-energy states of a rod subject to constraints that do not restrict the twist. The stretching and contraction energies of a free helical spring are computed. There are local high-energy minima. We show the possibility of using the spring to model the chirality of DNA. We then compare our results with an available atomic level energy simulation that was performed on DNA unconstrained in the same sense as the free spring. We find some possible reflections of springlike behavior in the mechanics of DNA, but, unsurprisingly, the base pairs lend a material substance to the core of DNA that a spring does not capture.

X-ray diffraction structures of the nucleosome core particle along with a variety of experiments are consistent with the idea that an important source of the free energy holding DNA to the superhelical ramp on the histone octamer surface is obtained from a relatively small amount of electrostatic neutralization of the DNA phosphate charge by positively charged histone groups, especially arginine residues. Here we present a theoretical analysis of a simple model that emphasizes the competition between the high degree of bending of the stiff DNA molecule required for its tight curvature on the histone octamer and the neutralization of the DNA phosphate charge by basic histone residues. Our calculation accounts for the strong influence of condensed counterions on the electrostatic interactions. We find that the minimum amount of free energy required to bend DNA into axial conformity with the superhelical ramp at physiological salt concentration can be provided by a scant 6% neutralization of the phosphate charge, in close correspondence to the stoichiometric neutralization of phosphate charge by the arginine side chain that intrudes into the inward-facing minor groove of each DNA double helical turn.

We present a simple model for the binding of persistence-length segments of a polyelectrolyte to an oppositely charged spherical macroion with radius of the same order as the polymer persistence length. The polyelectrolyte is taken as a rod with elastic bending capacity. The macroion is abstracted as an absolutely rigid circular arc. In the bound state, the polyelectrolyte bends onto the macroion, and some of the opposite charges are mutually neutralized. The curvature of the macroion imposes stability limits on the bound state. The model predicts transitions to bound-state stability in the same conditions (polyelectrolyte charge density and bare persistence length, macroion radius, salt concentration) under which they are observed in experimental studies of the binding of polyelectrolytes to oppositely charged micelles. The calculations also suggest why macroscopic phase separation occurs in experiments below a critical salt concentration but not above.

We describe how the stability properties of DNA minicircles can be directly read from plots of various biologically intuitive quantities along families of equilibrium configurations. Our conclusions follow from extensions of the mathematical theory of distinguished bifurcation diagrams that are applied within the specific context of an elastic rod model of minicircles. Families of equilibria arise as a twisting angle alpha is varied. This angle is intimately related to the continuously varying linking number Lk for nicked DNA configurations that is defined as the sum of Twist and Writhe. We present several examples of such distinguished bifurcation diagrams involving plots of the energy E, linking number Lk, and a twist moment m3, along families of cyclized equilibria of both intrinsically straight and intrinsically curved DNA fragments.

Recent experimental, theoretical, and computational developments in the field of nucleic acid electrostatics have brought interesting concepts to the fore. The phosphate charge on the double helix apparently influences its structure. When the charge is neutralized asymmetrically, the resulting force imbalance drives bending toward the neutralized side. When the charge is uniformly neutralized, the force imbalance acts to buckle the helical axis, resulting in a compact tertiary conformation. Sharing of condensed counterions by single strands is a stabilizing factor for formation of the double helix. Sharing of condensed counterions by two double helices causes clustering of DNA and may be a factor in RNA folding. Support for these statements is reviewed.

Dibutyltin (DBT) is used in the plastics polymerization process as a catalyst in polyvinyl chloride (PVC) products and is the primary degradation product of tributyltin (TBT), an antifoulant in marine paint. DBT and other organotin compounds make their way into the environment through antifoulants, PVC processing plants, and PVC products maintained in water and water-handling systems. A flow-through saltwater life-cycle toxicity test was conducted to determine the chronic effect of DBT to the sheepshead minnow (Cyprinodon variegatus Lacepede), an estuarine species. Embryos were monitored through hatch, maturation, growth, and reproduction in DBT concentrations of 158, 286, 453, 887, and 1510 µg l−1. Progeny were monitored for survival as embryos and fry/juveniles, and growth for 30 days post-isolation. Mean length of parental generation fish was significantly reduced on day 30 at DBT concentrations ≥887 µg l−1, setting the lowest observable effect concentration (LOEC) at 887 µg l−1 and the no observable effect concentration (NOEC) at 453 µg l−1. Fecundity, as egg viability, was significantly reduced at the LOEC. Survival of parental and progeny generation embryos and mean length, wet weight and dry weight of progeny generation juveniles were not significantly affected at concentrations ≤LOEC. TBT, a toxic impurity in DBT reversibly produced in DBT by the process of comproportionation, was also monitored throughout this study. Comparing measured levels of TBT in this study with levels exerting toxic effects in an earlier TBT life-cycle study with C. variegatus suggests biological responses in this study were likely due to the TBT impurity and not to DBT alone. Results indicate that TBT impurity as low as 0.1% may have a significant influence on the perceived toxicity of DBT and that spontaneous production of TBT in DBT may be the major source of biological toxicity of DBT. Copyright © 2003 John Wiley & Sons, Ltd.

The toxic effects of nonsteroidal anti-inflammatory drugs (NSAIDs) on the small bowel have been reported extensively. A growing number of reports of toxic effects of NSAIDs on the colon have appeared recently. The clinical presentation, endoscopic appearances and histological findings of so-called NSAID colopathy are quite varied, as illustrated by a series of four patients described in this report. Presenting symptoms and signs in this series include iron-deficiency anaemia and crampy abdominal pain, but alteration of bowel habit, weight loss, and even nausea and vomiting have also been described. One patient in this series has large-bowel diaphragms, considered by some to be pathognomonic of NSAID effects. Each of the four patients had right-sided colonic lesions only, possibly supporting a direct toxic effect of NSAIDs. Management usually involves simply stopping the offending NSAID. A review of the literature on this under-recognized entity is presented.

DNA dodecamers of the alternating d(CG).d(CG) sequence with six phosphate groups either charge-neutralized or substituted by neutral methylphosphonates across the major or minor groove have been subjected to energy minimization to determine the conformational effect of the asymmetric elimination of phosphate charge. We report bending angles, directions of bending, and detailed structural characteristics such as groove widths and local base-pair parameters. Our principal results are that charge neutralization on one face of the DNA induces significant bending toward the neutralized face, in agreement with theoretical predictions on a simplified model and experimental data on a similar base-pair sequence, and that the DNA conformation averaged over all stereospecific methylphosphonate substitutions is nearly the same as the conformation produced by charge neutralization of the phosphates. Individual isomers, however, cover a wide range of structures, with the magnitude and direction of overall bending sensitive to the precise stereochemical pattern of neutralization. Our simulation does not explicitly contain counterions, and the results therefore suggest that counterions can influence DNA structure by neutralizing the phosphate charge. These data provide new hints into the molecular mechanisms which underlie the deformations of DNA structure induced by the binding of positively charged proteins and other tightly associated cationic species.

We have developed explicit formulas for the excess number of counterions condensed on kinked and intersecting charged lines caused by the more intense electric field in the neighborhood of the kink or intersection. As expected, the number of additionally bound counterions is greater for more pronounced kinks, and also increases with the number of lines that intersect at a common point. We have also analyzed the electrostatic interaction potential as a function of distance between two charged lines in skewed orientation. Our finding in this case is that in a range of close distances the lines must cross a free energy barrier in order to separate.

The melting temperature of the poly(dA)·poly(dT) double helix is exquisitely sensitive to salt concentration, and the helix-to-coil transition is sharp. Modern calorimetric instrumentation allows this transition to be detected and characterized with high precision at extremely low duplex concentrations. We have taken advantage of these properties to show that this duplex can be used as a sensitive probe to detect and to characterize the influence of other solutes on solution properties. We demonstrate how the temperature associated with poly(dA)·poly(dT) melting can be used to define the change in bulk solution cation concentration imparted by the presence of other duplex and triplex solutes, in both their native and denatured states. We use this information to critically evaluate features of counterion condensation theory, as well as to illustrate “crosstalk” between different, non-contacting solute molecules. Specifically, we probe the melting of a synthetic homopolymer, poly(dA)·poly(dT), in the presence of excess genomic salmon sperm DNA, or in the presence of one of two synthetic RNA polymers (the poly(rA)·poly(rU) duplex or the poly(rU)·poly(rA)·poly(rU) triplex). We find that these additions cause a shift in the melting temperature of poly(dA)·poly(dT), which is proportional to the concentration of the added polymer and dependent on its conformational state (B versus A, native versus denatured, and triplex versus duplex). To a first approximation, the magnitude of the observed tm shift does not depend significantly on whether the added polymer is RNA or DNA, but it does depend on the number of strands making up the helix of the added polymer. We ascribe the observed changes in melting temperature of poly(dA)·poly(dT) to the increase in ionic strength of the bulk solution brought about by the presence of the added nucleic acid and its associated counterions. We refer to this communication between non-contacting biopolymers in solution as solvent-mediated crosstalk. By comparison with a known standard curve of tmversus log[Na+] for poly(dA)·poly(dT), we estimate the magnitude of the apparent change in ionic strength resulting from the presence of the bulk nucleic acid, and we compare these results with predictions from theory. We find that current theoretical considerations correctly predict the direction of the tm shift (the melting temperature increases), while overestimating its magnitude. Specifically, we observe an apparent increase in ionic strength equal to 5 % of the concentration of the added duplex DNA or RNA (in mol phosphate), and an additional apparent increase of about 9.5 % of the nucleic acid concentration (mol phosphate) upon denaturation of the added DNA or RNA, yielding a total apparent increase of 14.5 %. For the poly(rU)·poly(rA)·poly(rU) triplex, the total apparent increase in ionic strength corresponds to about 13.6 % of the amount of added triplex (moles phosphate). The effect we observe is due to coupled equilibria between the solute molecules mediated by modulations in cation concentration induced by the presence and/or the transition of one of the solute molecules. We note that our results are general, so one can use a different solute probe sensitive to proton binding to characterize subtle changes in solution pH induced by the presence of another solute in solution. We discuss some of the broader implications of these measurements/results in terms of nucleic acid melting in multicomponent systems, in terms of probing counterion environments, and in terms of potential regulatory mechanisms.

We extend the theory of counterion condensation from the standard representation of a locally stiff rodlike polyion as a line of discrete charges to a helical lattice of charges. The number of counterions condensing on a helix with given axial charge density is the same as on a line of the same charge density, but the electrostatic free energy is substantially less for the helix than for the line. In fact, the free energy of assembling the helical charge lattice becomes negative at higher salt concentrations, indicating electrostatic stabilization of the helix due to the mixing entropy of condensed counterions. The electrostatic persistence lengths of the line and helix subjected to locally elastic thermal bending fluctuations differ only slightly. They have the same κ -2 dependence as the Odijk-Skolnick-Fixman persistence length but a different prefactor. It is commonly believed that elastic bending models for the persistence length are applicable to double-helical DNA, but we point out that DNA apparently has unique mechanical properties that do not conform to the elastic model of bending. On the other hand, the helical model and its subsequent generalization to a double helix will serve as a basis for the electrostatic free energy of DNA conformational transitions.

Within the context of DNA rings, we analyze the relationship between intrinsic shape and the existence of multiple stable equilibria, either nicked or cyclized with the same link. A simple test, based on a perturbation expansion of symmetry breaking within a continuum elastic rod model, provides good predictions of the occurrence of such multiple equilibria. The reliability of these predictions is verified by direct computation of nicked and cyclized equilibria for several thousand DNA minicircles with lengths of 200 and 900bp. Furthermore, our computations of equilibria for nicked rings predict properties of the equilibrium distribution of link, as calculated by much more computationally intensive Monte Carlo simulations.

Monte Carlo simulations of the interactions of Na+ and Cl- ions with λ repressor protein and its DNA operator site have revealed an interesting effect [Jayaram, B.; DiCapua, F. M.; Beveridge, D. L. J. Am. Chem. Soc. 1991, 113, 5211−5215]. When the protein is close to its binding site on DNA, the presence of the small ions strengthens the electrostatic net attractive force between the formally charged protein residues and the phosphate groups on DNA. The effect has been interpreted as a manifestation of the release of counterions condensed on DNA. We show that although counterions are indeed released, the enhancement of the attractive force at short distances has a different origin. There is a direct attraction between DNA phosphates and positively charged protein residues, and a direct repulsion between DNA phosphates and negatively charged protein residues. Close to the DNA, the net direct force is attractive. By weakening the direct repulsion between DNA phosphates and the negatively charged glutamate a...

We report further development of a pair potential for two identically charged rodlike polyions oriented in parallel. An attractive force mediated by condensed counterions of any valence, including univalent, is identified for separation distances between a Debye screening length and about a third of a Debye length. The characteristics of the potential are explored as a function of ionic strength, dielectric constant, counterion valence, and charge density of the polyions. The work required to assemble more than two polyions in parallel array is shown not to be pairwise additive. The free energy of assembling many parallel polyions in a hexagonally ordered cluster is calculated as a function of the number N of polyions in the cluster. The free energy has a minimum at a finite cluster size. Discrepancies between the theoretical structure of the stable cluster and experimental observations on polyelectrolyte clusters (with univalent counterions) are discussed, and it is tentatively concluded that the apparen...

We have computed the fraction of DNA phosphate charge neutralized by condensed counterions for parallel pairs of linear DNA segments, DNA circles, and representative closed-circular supercoiled DNA configurations ranging in length from 42 to 3000 base pairs. We find significant but small uptake of condensed counterions for the more compact structures relative to an isolated linear DNA segment.

It has been inferred from electric birefringence and electric dichroism data that the field-induced dipole moment on charged rodlike particles saturates at higher field strengths. We give theoretical justification for this interpretation of the data. We calculate the threshold field marking the onset of saturation and compare the result with measured values.

The effects of tributyltin (TBT) to the life cycle of the estuarine fish Cyprinodon variegatus were examined in a 180-day flow-through exposure. The study was initiated with embryos less than 24 h postfertilization and monitored through hatch, maturation, growth, and reproduction under continuous exposure to mean measured TBT concentrations of 5.4, 3.2, 1.3, 0.66, and 0.41 microg TBT/L. Progeny isolated at the onset of reproduction by the parental generation (F0) were monitored for survival as embryos, survival as fry/juveniles, and growth 30 days postisolation. TBT, at a concentration of 5.4 microg/L, significantly reduced embryo survival of the F0 generation sheepshead minnows. By day 145 of the exposure, significant effects to the survival of this generation increased and included all test concentrations/=0.66 microg/L. Survival of 0.66 microg/L was reduced 59% relative to control survival by termination of the F0 generation. Growth of F0 generation organisms as measured by standard length was significantly reduced only on day 90 at 3.2 microg/L, however no significant reductions of wet or dry weight related to treatment concentration were detected. Due to complete mortality of organisms exposed to 5.4 microg/L by study day 7, effects to fecundity and progeny were monitored at measured concentrations of 3.2, 1.3, 0.66, and 0.41 microg TBT/L. Fecundity, as measured by the production of viable eggs produced per female per day, was unaffected by any of the test treatments. All F1 generation embryos isolated from treatment chambers into 3.2 microg/L died. Survival, standard length, wet and dry weight of the F1 generation at the remaining treatment concentrations were unaffected. The results of this study indicate that exposure to TBT reduced survival of the F0 generation sheepshead minnow and establishes the lowest observed effect concentration (LOEC) as 0.66 microg TBT/L, and the no observed effect concentration (NOEC) as 0.41 microg TBT/L for this species.http://link.springer-ny. com/link/service/journals/00244/bibs/37n2p258.html

We develop formulas for the counterion−polyion and coion−polyion potentials of mean force in the context of counterion condensation theory. For separation distances on the scale of a Debye screening length, both potentials conform to Debye−Huckel screening behavior. Close to the polyion, in the region defining the layer of condensed counterions, the counterion potential is attractive and unscreened. A coion in this region acts like a charged group on the polyion; its potential is repulsive, and it augments the ability of the polymer to condense counterions. There is an interfacial region between condensed layer and diffuse cloud from which counterions are expelled back into the cloud but which accumulates coions. The associated radial distribution functions exhibit two peaks for the counterion and a single peak for the coion.

Boron toxicity has been recognised as an important problem limiting production in the low-rainfall regions of southern Australia, West Asia and North Africa. Genetic variation for boron toxicity tolerance in barley has been characterised but the mode of inheritance and the location of genes controlling tolerance were not previously known. A population of 150 doubled-haploid lines from a cross between a boron toxicity tolerant Algerian landrace, Sahara 3771, and the intolerant Australian cultivar Clipper was screened in four tolerance assays. An RFLP linkage map of the Clipper×Sahara population was used to identify chromosomal regions associated with boron tolerance in barley. Interval regression-mapping allowed the detection of four chromosomal regions involved in the boron tolerance traits measured. A region on chromosome 2H was associated with leaf-symptom expression, a region on chromosome 3H was associated with a reduction of the affect of boron toxicity on root growth suppression, a region on chromosome 6H was associated with reduced boron uptake, and a region on chromosome 4H was also associated with the control of boron uptake as well as being associated with root-length response, dry matter production and symptom expression. The benefits and potential of marker-assisted selection for boron toxicity tolerance are discussed.

High-power substrates for hybrid circuits that combine aluminium with an anodic coating are highly efficient as the heat generated from chips mounted on the coated substrates can be dissipated rapidly. However, due to the difference between the thermal expansion coefficients of the two materials, a large thermal tensile stress can be built up during heating of the substrate, which causes cracking in the coating and subsequently the loss of electrical integrity. In this investigation a simple method of measuring the thermal expansion coefficient for an anodic substrate is described. Measurements were taken from an aluminium/coating composite and the thermal expansion coefficient of the coating was determined using the principles of elementary mechanics.

Complex formation between small carboxyl-terminated dendritic polymers and oppositely charged poly(dimethyldiallylammonium chloride) was monitored by turbidimetry, dynamic light scattering, viscometry and potentiometric titration. All techniques reveal a discontinuity at a well-defined pH, corresponding to the point of incipient complex formation. Conversion of this critical pH to a surface charge density σcrit leads to the observation that σcrit, to a first approximation, varies linearly with the Debye−Huckel parameter κ and that binding occurs more easily (i.e., at lower σcrit) for the larger dendrimer. A model is presented in which the distortion of the polyelectrolyte backbone in the complex is described in terms of the elastic resistance to bending around the contour of the spherical macroion, acting in opposition to attractive Coulombic forces. The elastic resistance is treated as a Hookean effect obtained from the measured persistence length. This simple theory produces the principal features of th...

Anthropogenic environmental stress is a likely contributor to outbreaks of disease due to immunosuppression or increased host vulnerability. Estuarine organisms are exposed to variable concentrations of marine antifouling agents, such as tributyltin (TBT), with higher exposures existing near ports and marinas. Eastern oysters ( Crassostrea virginica ), inhabiting the northern Gulf of Mexico and the Atlantic coast of North America, are susceptible to a protozoan pathogen, Perkinsus marinus , which has devastated oyster populations and depleted oyster fisheries throughout its range. This study examined the effects of exposure to environmental levels of TBT on susceptibility and survival of oysters when subsequently challenged with P. marinus . Oysters were exposed to TBT (0, 30 and 80 ng/L) in flow-through aquaria for 9 weeks, then moved to static aquaria, where they were challenged with parasites and monitored for an additional 8 weeks for mortality and disease. Results indicated increased infection intensity and oyster mortality in the TBT-exposed treatments relative to unexposed controls. It is hypothesized that TBT exacerbates the infectious disease process and that exposed oysters succumb at lower levels of infection.

We review some of the characteristic properties of the structure of polyelectrolyte solutions: the condensed layer of counterions that forms abruptly at a critical threshold charge density on the polymer chain; the more diffuse Debye-Hückel cloud, which is spatially distinct from the condensed layer; and the entropic release of counterions from the condensed layer as a driving force for the binding of oppositely charged ligands. We present a reminder of the basis of our current understanding in a variety of experiments, simulations, and theories; and we attempt as well to clarify some misunderstandings. We present a new analysis of a lattice model that suggests why the limiting laws for polyelectrolyte thermodynamics have proved to be accurate despite the neglect of polymer-polymer interactions in their original derivation. We sketch recent progress in constructing a potential between counterion and polyion. A counterion located in the interface between condensed layer and Debye cloud is repelled from the polyion, creating a sharp boundary between the two counterion populations.

The Ramanathan-Woodbury formulas representing the charge density critical for the onset of counterion condensation on finite-length polymers are derived by three alternate methods, an extension of Debye-Huckel theory, a theory of end effects, and by density functional theory. For charged oligomers with length of the same order as the Debye length, the threshold for condensation is the same as for polymers of length much greater than the Debye lenght. However, the threshold depends both on length and salt concentration if the oligomer is shorter than the Debye length, in such a way as to recede to infinity as the ratio of oligomer length to Debye length tends to zero (i.e., condensation vanishes in this limit). The extended Debye-Huckel theory additionally provides a new result for the partition function of the condensed layer, showing that the free energy of the condensed counterions is different on an oligomer and a polymer, even when the fractional extent of condensation is the same. The end effect theory discloses a hitherto unnoticed connection between the number of counterions condensed at the ends of a long polymer and the number condensed on a short oligomer.

A theory of attractive forces between identical highly charged polyions in a 1:1 supporting electrolyte, generated by the interaction of the layers of condensed counterions, is extended to a Z:Z‘ electrolyte and to rodlike polymer segments of any charge density. If the combined charge density of a polyion pair does not exceed the charge density critical for counterion condensation, then there is no attraction. But an attraction arises if the combined charge density does exceed the condensation threshold, even if the individual polymer charge densities lie below the critical value. The intensity of the attraction at a fixed ionic strength decreases with higher values of counterion valence. The effect of increased ionic strength is to decrease both the range and the intensity of the attractive force.

The primitive model and the line of charges model for ionized polymers both predict the emergence of a condensed counterion layer at a critical polymer charge density, but the two models describe the condensed layer in different ways. The Zimm-Le Bret and Ramanathan distributions for the condensed layer in the primitive model are reviewed. The extended Debye-Huckel theory of the line of charges model is recast in a more general form in an attempt to make its underlying assumptions transparent. We show that the counterion-polyion potential of mean force at great distances behaves differently for the two models.

A survey of experimental and theoretical investigations into the condensation of counterions on polyelectrolyte chains above a critical threshold of polymer charge density is presented. Current models contain critical formation of a condensed layer that cannot be diluted off the polymer but differ qualitatively in their description of the structure of the layer. A documented position is taken that the model of a line of discrete charges is more realistic than a uniformly charged cylinder. The question of attractive electrostatic interactions between identically charged polymers is addressed.

The effects of two new non-peptide angiotensin receptor antagonists, irbesartan (SR 47436/BMS-186295, (2-n-butyl-4-spirocyclopentane-1-[((2'-tetrazol-5-yl)bipheny l-4-yl)methyl]2 - imidazolin-5-one) and SR 47155A (2-n-butyl-4-spirocyclopentane-1-[((2'-carboxy)biphenyl-4-yl)methy l]2- imidazolin-5-one, trifluoroacetate), on angiotensin II-induced pressor responses were studied in the pithed rat in comparison to losartan, EXP 3174 and [Sar1,Val5,Ala8]angiotensin II. SR 47155A (1-10 mg/kg i.v.) and losartan (1-10 mg/kg i.v.) shifted dose dependently the dose-response curve of angiotensin II to the right without affecting the maximal response. SR 47436 (0.3-10 mg/kg i.v.), EXP 3174 (0.03-1 mg/kg i.v.) and [Sar1,Val5,Ala8]angiotensin II (0.03-1 mg/kg i.v.) induced, at least at high doses, a non-parallel shift to the right of the angiotensin II dose-response curve and this was associated with a reduction of the maximal response. During a 70 min period, the effect of [Sar1,Val5,Ala8]angiotensin II (1 mg/kg i.v.) on the angiotensin II (0.3 microgram/kg i.v.)-induced pressor response was shown to be reversible, the effect of SR 47155A (10 mg/kg i.v.) was partially reversible and the effect of SR 47436 (3 mg/kg i.v.), EXP 3174 (1 mg/kg i.v.) or losartan (6 mg/kg i.v.) was not reversed at the end of this 70 min period. Administration of SR 47155A (10 mg/kg i.v.) before SR 47436 (1-10 mg/kg i.v.) reversed the reduced angiotensin II-maximal response induced by SR 47436. Administration of SR 47436 (10 mg/kg i.v.) before SR 47155A (1-10 mg/kg i.v.) prevented the full development of the pressor response as observed in the absence of SR 47436. In the pithed rat, SR 47436 (30 mg/kg i.v.) and losartan (30 mg/kg i.v.) reduced the change in diastolic blood pressure induced by electrical stimulation of the spinal cord only at low stimulation rates. Taken together these results indicate that SR 47436, under in vivo conditions, is a potent non-peptide angiotensin receptor antagonist. The type of antagonism (partially insurmountable but selective) can be explained by different theoretical models which are discussed.