Thermodynamics of Competitive Molecular Channel Transport: Application to Artificial Nuclear Pores

In an analytical model channel transport is analyzed as a function of key parameters, determining efficiency and selectivity of particle transport in a competitive molecular environment. These key parameters are the concentration of particles, solvent-channel exchange dynamics, as well as particle-in-channel- and interparticle interaction. These parameters are explicitly related to translocation dynamics and channel occupation probability. Slowing down the exchange dynamics at the channel ends, or elevating the particle concentration reduces the in-channel binding strength necessary to maintain maximum transport. Optimized in-channel interaction may even shift from binding to repulsion. A simple equation gives the interrelation of access dynamics and concentration at this transition point. The model is readily transferred to competitive transport of different species, each of them having their individual in-channel affinity. Combinations of channel affinities are determined which differentially favor selectivity of certain species on the cost of others. Selectivity for a species increases if its in-channel binding enhances the species' translocation probablity when compared to that of the other species. Selectivity increases particularly for a wide binding site, long channels, and fast access dynamics. Recent experiments on competitive transport of in-channel binding and inert molecules through artificial nuclear pores serve as a paradigm for our model. It explains qualitatively and quantitatively how binding molecules are favored for transport at the cost of the transport of inert molecules

A field-theoretic approach to the Wiener Sausage

The Wiener Sausage, the volume traced out by a sphere attached to a Brownian particle, is a classical problem in statistics and mathematical physics. Initially motivated by a range of field-theoretic, technical questions, we present a single loop renormalised perturbation theory of a stochastic process closely related to the Wiener Sausage, which, however, proves to be exact for the exponents and some amplitudes. The field-theoretic approach is particularly elegant and very enjoyable to see at work on such a classic problem. While we recover a number of known, classical results, the field-theoretic techniques deployed provide a particularly versatile framework, which allows easy calculation with different boundary conditions even of higher momenta and more complicated correlation functions. At the same time, we provide a highly instructive, non-trivial example for some of the technical particularities of the field-theoretic description of stochastic processes, such as excluded volume, lack of translational invariance and immobile particles. The aim of the present work is not to improve upon the well-established results for the Wiener Sausage, but to provide a field-theoretic approach to it, in order to gain a better understanding of the field-theoretic obstacles to overcome.Comment: 45 pages, 3 Figures, Springer styl

Enhancement of Transport Selectivity through Nano-Channels by Non-Specific Competition

The functioning of living cells requires efficient and selective transport of materials into and out of the cell, and between different cellular compartments. Much of this transport occurs through nano-scale channels that do not require large scale molecular re-arrangements (such as transition from a ‘closed’ to an ‘open’ state) and do not require a direct input of metabolic energy during transport. Nevertheless, these ‘always open’ channels are highly selective and pass only their cognate molecules, while efficiently excluding all others; indeed, these channels can efficiently transport specific molecules even in the presence of a vast excess of non-specific molecules. Such biological transporters have inspired the creation of artificial nano-channels. These channels can be used as nano-molecular sorters, and can also serve as testbeds for examining modes of biological transport. In this paper, we propose a simple kinetic mechanism that explains how the selectivity of such ‘always open’ channels can be based on the exclusion of non-specific molecules by specific ones, due to the competition for limited space inside the channel. The predictions of the theory account for the behavior of the nuclear pore complex and of artificial nanopores that mimic its function. This theory provides the basis for future work aimed at understanding the selectivity of various biological transport phenomena

The porin and the permeating antibiotic: A selective diffusion barrier in gram-negative bacteria

Gram-negative bacteria are responsible for a large proportion of antibiotic resistant bacterial diseases. These bacteria have a complex cell envelope that comprises an outer membrane and an inner membrane that delimit the periplasm. The outer membrane contains various protein channels, called porins, which are involved in the influx of various compounds, including several classes of antibiotics. Bacterial adaptation to reduce influx through porins is an increasing problem worldwide that contributes, together with efflux systems, to the emergence and dissemination of antibiotic resistance. An exciting challenge is to decipher the genetic and molecular basis of membrane impermeability as a bacterial resistance mechanism. This Review outlines the bacterial response towards antibiotic stress on altered membrane permeability and discusses recent advances in molecular approaches that are improving our knowledge of the physico-chemical parameters that govern the translocation of antibiotics through porin channel

Spatial Distribution of Calcium-Gated Chloride Channels in Olfactory Cilia

Background: In vertebrate olfactory receptor neurons, sensory cilia transduce odor stimuli into changes in neuronal membrane potential. The voltage changes are primarily caused by the sequential openings of two types of channel: a cyclic-nucleotide-gated (CNG) cationic channel and a calcium-gated chloride channel. In frog, the cilia are 25 to 200 mm in length, so the spatial distributions of the channels may be an important determinant of odor sensitivity. Principal Findings: To determine the spatial distribution of the chloride channels, we recorded from single cilia as calcium was allowed to diffuse down the length of the cilium and activate the channels. A computational model of this experiment allowed an estimate of the spatial distribution of the chloride channels. On average, the channels were concentrated in a narrow band centered at a distance of 29 % of the ciliary length, measured from the base of the cilium. This matches the location of the CNG channels determined previously. This non-uniform distribution of transduction proteins is consistent with similar findings in other cilia. Conclusions: On average, the two types of olfactory transduction channel are concentrated in the same region of the cilium

Manipulation of Signaling Thresholds in “Engineered Stem Cell Niches” Identifies Design Criteria for Pluripotent Stem Cell Screens

In vivo, stem cell fate is regulated by local microenvironmental parameters. Governing parameters in this stem cell niche include soluble factors, extra-cellular matrix, and cell-cell interactions. The complexity of this in vivo niche limits analyses into how individual niche parameters regulate stem cell fate. Herein we use mouse embryonic stem cells (mESC) and micro-contact printing (µCP) to investigate how niche size controls endogenous signaling thresholds. µCP is used to restrict colony diameter, separation, and degree of clustering. We show, for the first time, spatial control over the activation of the Janus kinase/signal transducer and activator of transcription pathway (Jak-Stat). The functional consequences of this niche-size-dependent signaling control are confirmed by demonstrating that direct and indirect transcriptional targets of Stat3, including members of the Jak-Stat pathway and pluripotency-associated genes, are regulated by colony size. Modeling results and empirical observations demonstrate that colonies less than 100 µm in diameter are too small to maximize endogenous Stat3 activation and that colonies separated by more than 400 µm can be considered independent from each other. These results define parameter boundaries for the use of ESCs in screening studies, demonstrate the importance of context in stem cell responsiveness to exogenous cues, and suggest that niche size is an important parameter in stem cell fate control

How β-Lactam Antibiotics Enter Bacteria: A Dialogue with the Porins

BACKGROUND:Multi-drug resistant (MDR) infections have become a major concern in hospitals worldwide. This study investigates membrane translocation, which is the first step required for drug action on internal bacterial targets. beta-lactams, a major antibiotic class, use porins to pass through the outer membrane barrier of Gram-negative bacteria. Clinical reports have linked the MDR phenotype to altered membrane permeability including porin modification and efflux pump expression. METHODOLOGY/PRINCIPAL FINDINGS: Here influx of beta-lactams through the major Enterobacter aerogenes porin Omp36 is characterized. Conductance measurements through a single Omp36 trimer reconstituted into a planar lipid bilayer allowed us to count the passage of single beta-lactam molecules. Statistical analysis of each transport event yielded the kinetic parameters of antibiotic travel through Omp36 and distinguishable translocation properties of beta-lactams were quantified for ertapenem and cefepime. Expression of Omp36 in an otherwise porin-null bacterial strain is shown to confer increases in the killing rate of these antibiotics and in the corresponding bacterial susceptibility. CONCLUSIONS/SIGNIFICANCE: We propose the idea of a molecular "passport" that allows rapid transport of substrates through porins. Deciphering antibiotic translocation provides new insights for the design of novel drugs that may be highly effective at passing through the porin constriction zone. Such data may hold the key for the next generation of antibiotics capable of rapid intracellular accumulation to circumvent the further development MDR infections

The Drosophila Gap Gene Network Is Composed of Two Parallel Toggle Switches

Drosophila “gap” genes provide the first response to maternal gradients in the early fly embryo. Gap genes are expressed in a series of broad bands across the embryo during first hours of development. The gene network controlling the gap gene expression patterns includes inputs from maternal gradients and mutual repression between the gap genes themselves. In this study we propose a modular design for the gap gene network, involving two relatively independent network domains. The core of each network domain includes a toggle switch corresponding to a pair of mutually repressive gap genes, operated in space by maternal inputs. The toggle switches present in the gap network are evocative of the phage lambda switch, but they are operated positionally (in space) by the maternal gradients, so the synthesis rates for the competing components change along the embryo anterior-posterior axis. Dynamic model, constructed based on the proposed principle, with elements of fractional site occupancy, required 5–7 parameters to fit quantitative spatial expression data for gap gradients. The identified model solutions (parameter combinations) reproduced major dynamic features of the gap gradient system and explained gap expression in a variety of segmentation mutants

Lobe Specific Ca2+-Calmodulin Nano-Domain in Neuronal Spines: A Single Molecule Level Analysis

Calmodulin (CaM) is a ubiquitous Ca2+ buffer and second messenger that affects cellular function as diverse as cardiac excitability, synaptic plasticity, and gene transcription. In CA1 pyramidal neurons, CaM regulates two opposing Ca2+-dependent processes that underlie memory formation: long-term potentiation (LTP) and long-term depression (LTD). Induction of LTP and LTD require activation of Ca2+-CaM-dependent enzymes: Ca2+/CaM-dependent kinase II (CaMKII) and calcineurin, respectively. Yet, it remains unclear as to how Ca2+ and CaM produce these two opposing effects, LTP and LTD. CaM binds 4 Ca2+ ions: two in its N-terminal lobe and two in its C-terminal lobe. Experimental studies have shown that the N- and C-terminal lobes of CaM have different binding kinetics toward Ca2+ and its downstream targets. This may suggest that each lobe of CaM differentially responds to Ca2+ signal patterns. Here, we use a novel event-driven particle-based Monte Carlo simulation and statistical point pattern analysis to explore the spatial and temporal dynamics of lobe-specific Ca2+-CaM interaction at the single molecule level. We show that the N-lobe of CaM, but not the C-lobe, exhibits a nano-scale domain of activation that is highly sensitive to the location of Ca2+ channels, and to the microscopic injection rate of Ca2+ ions. We also demonstrate that Ca2+ saturation takes place via two different pathways depending on the Ca2+ injection rate, one dominated by the N-terminal lobe, and the other one by the C-terminal lobe. Taken together, these results suggest that the two lobes of CaM function as distinct Ca2+ sensors that can differentially transduce Ca2+ influx to downstream targets. We discuss a possible role of the N-terminal lobe-specific Ca2+-CaM nano-domain in CaMKII activation required for the induction of synaptic plasticity