Ion Current Rectification,Limiting and Overlimiting Conductances in Nanopores

Previous reports on Poisson-Nernst-Planck (PNP) simulations of solid-state nanopores have focused on steady state behaviour under simplified boundary conditions. These are Neumann boundary conditions for the voltage at the pore walls, and in some cases also Donnan equilibrium boundary conditions for concentrations and voltages at both entrances of the nanopore. In this paper, we report time-dependent and steady state PNP simulations under less restrictive boundary conditions, including Neumann boundary conditions applied throughout the membrane relatively far away from the nanopore. We simulated ion currents through cylindrical and conical nanopores with several surface charge configurations, studying the spatial and temporal dependence of the currents contributed by each ion species. This revealed that, due to slow co-diffusion of oppositely charged ions, steady state is generally not reached in simulations or in practice. Furthermore, it is shown that ion concentration polarization is responsible for the observed limiting conductances and ion current rectification in nanopores with asymmetric surface charges or shapes. Hence, after more than a decade of collective research attempting to understand the nature of ion current rectification in solid-state nanopores, a relatively intuitive model is retrieved. Moreover, we measured and simulated current-voltage characteristics of rectifying silicon nitride nanopores presenting overlimiting conductances. The similarity between measurement and simulation shows that overlimiting conductances can result from the increased conductance of the electric double-layer at the membrane surface at the depletion side due to voltage-induced polarization charges. The MATLAB source code of the simulation software is available via the website http://micr.vub.ac.be.     

Induction of Neuronal Morphology in the 661W Cone Photoreceptor Cell Line with Staurosporine

Purpose

RGC-5 cells undergo differentiation into a neuronal phenotype with low concentrations of staurosporine. Although the RGC-5 cell line was initially thought to be of retinal ganglion cell origin, recent evidence suggests that the RGC-5 line could have been the result of contamination with 661W mouse cone photoreceptor cells. This raised the possibility that a cone photoreceptor cell line could be multipotent and could be differentiated to a neuronal phenotype.

Methods

661W and RGC-5 cells, non-neuronal retinal astrocytes, retinal endothelial cells, retinal pericytes, M21 melanoma cells, K562 chronic myelogenous leukemia cells, and Daudi Burkitt lymphoma cells, were differentiated with staurosporine. The resulting morphology was quantitated using NeuronJ with respect to neurite counts and topology.

Results

Treatment with staurosporine induced similar-appearing morphological differentiation in both 661W and RGC-5 cells. The following measures were not significantly different between 661W and RGC-5 cells: number of neurites per cell, total neurite field length, number of neurite branch points, and cell viability. Neuronal-like differentiation was not observed in the other cell lines tested.

Conclusions

661W and RGC-5 cells have virtually identical and distinctive morphology when differentiated with low concentrations of staurosporine. This result demonstrates that a retinal neuronal precursor cell with cone photoreceptor lineage can be differentiated to express a neuronal morphology.     

Structural insights into the intertwined dimer of fyn SH2

Src homology 2 domains are interaction modules dedicated to the recognition of phosphotyrosine sites incorporated in numerous proteins found in intracellular signaling pathways. Here we provide for the first time structural insight into the dimerization of Fyn SH2 both in solution and in crystalline conditions, providing novel crystal structures of both the dimer and peptide‐bound structures of Fyn SH2. Using nuclear magnetic resonance chemical shift analysis, we show how the peptide is able to eradicate the dimerization, leading to monomeric SH2 in its bound state. Furthermore, we show that Fyn SH2's dimer form differs from other SH2 dimers reported earlier. Interestingly, the Fyn dimer can be used to construct a completed dimer model of Fyn without any steric clashes. Together these results extend our understanding of SH2 dimerization, giving structural details, on one hand, and suggesting a possible physiological relevance of such behavior, on the other hand.     

1H, 13C,and 15N backbone and side-chain chemical shift assignments of the free and bound forms of the human PTPN11 second SH2 domain

Src homology 2 (SH2) domains have an important role in the regulation of protein activity and intracellular signaling processes. They are geared to bind to specific phosphotyrosine (pY) motifs, with a substrate sequence specificity depending on the three amino acids immediately C-terminal to the pY. Here we report for the first time the ¹H, ¹⁵N and ¹³C backbone and side-chain chemical shift assignments for the C-terminal SH2 domain of the human protein tyrosine phosphatase PTPN11, both in its free and bound forms, where the ligand in the latter corresponds to a specific sequence of the human erythropoietin receptor.     

Impact of exacerbations of cystic fibrosis on muscle strength

Background

Adult patients with cystic fibrosis have peripheral muscle weakness, which is related to exercise intolerance and poor prognosis. The influence of acute exacerbations on muscle strength has been poorly studied. This study aimed to investigate whether quadriceps force (QF), as assessed with an involuntary technique, changes during intravenous antibiotics therapy (IVAT) for an exacerbation.

Methods

QF was measured in 20 patients using twitch stimulation of the femoral nerve at the day of hospitalization (day 1) and at termination (day 14) of the IVAT. Physical activity was monitored during IVAT using a SenseWear armband. Ten stable patients served as control subjects.

Results

QF did not change during exacerbation (potentiated twitch force at day 1: 140 ± 42 N, at day 14: 140 ± 47 N), but a decrease was observed in individual patients. Changes in twitch force during exacerbation were correlated with time spent in activities of at least moderate intensity (r = 0.61, p = 0.007).

Conclusions

QF does not systematically decrease during exacerbations of cystic fibrosis. Individual changes in QF are well correlated with daily time spent in activities of at least moderate intensity.     

Molecular control and function of endoreplication in development and physiology

Endoreplication, also called endoreduplication, is a cell cycle variant of multicellular eukaryotes in which mitosis is skipped and cells repeatedly replicate their DNA, resulting in cellular polyploidy. In recent years, research results have shed light on the molecular mechanism of endoreplication control, but the function of this cell-cycle variant has remained elusive. However, new evidence is at last providing insight into the biological relevance of cellular polyploidy, demonstrating that endoreplication is essential for developmental processes, such as cell fate maintenance, and is a prominent response to physiological conditions, such as pathogen attack or DNA damage. Thus, endoreplication is being revealed as an important module in plant growth that contributes to the robustness of plant life.