Plasma Membrane Calcium ATPase-Neuroplastin Complexes Are Selectively Stabilized in GM1-Containing Lipid Rafts

The recent identification of plasma membrane (Ca2+)-ATPase (PMCA)-Neuroplastin (Np) complexes has renewed attention on cell regulation of cytosolic calcium extrusion, which is of particular relevance in neurons. Here, we tested the hypothesis that PMCA-Neuroplastin complexes exist in specific ganglioside-containing rafts, which could affect calcium homeostasis. We analyzed the abundance of all four PMCA paralogs (PMCA1-4) and Neuroplastin isoforms (Np65 and Np55) in lipid rafts and bulk membrane fractions from GM2/GD2 synthase-deficient mouse brains. In these fractions, we found altered distribution of Np65/Np55 and selected PMCA isoforms, namely PMCA1 and 2. Cell surface staining and confocal microscopy identified GM1 as the main complex ganglioside co-localizing with Neuroplastin in cultured hippocampal neurons. Furthermore, blocking GM1 with a specific antibody resulted in delayed calcium restoration of electrically evoked calcium transients in the soma of hippocampal neurons. The content and composition of all ganglioside species were unchanged in Neuroplastin-deficient mouse brains. Therefore, we conclude that altered composition or disorganization of ganglioside-containing rafts results in changed regulation of calcium signals in neurons. We propose that GM1 could be a key sphingolipid for ensuring proper location of the PMCA-Neuroplastin complexes into rafts in order to participate in the regulation of neuronal calcium homeostasis.

Pattern forming systems coupling linear bulk diffusion to dynamically active membranes or cells

Some analytical and numerical results are presented for pattern formation properties associated with novel types of reaction–diffusion (RD) systems that involve the coupling of bulk diffusion in the interior of a multi-dimensional spatial domain to nonlinear processes that occur either on the domain boundary or within localized compartments that are confined within the domain. The class of bulk-membrane system considered herein is derived from an asymptotic analysis in the limit of small thickness of a thin domain that surrounds the bulk medium. When the bulk domain is a two-dimensional disk, a weakly nonlinear analysis is used to characterize Turing and Hopf bifurcations that can arise from the linearization around a radially symmetric, but spatially non-uniform, steady-state of the bulk-membrane system. In a singularly perturbed limit, the existence and linear stability of localized membrane-bound spike patterns is analysed for a Gierer–Meinhardt activator-inhibitor model that includes bulk coupling. Finally, the emergence of collective intracellular oscillations is studied for a class of PDE-ODE bulk-cell model in a bounded two-dimensional domain that contains spatially localized, but dynamically active, circular cells that are coupled through a linear bulk diffusion field. Applications of such coupled bulk-membrane or bulk-cell systems to some biological systems are outlined, and some open problems in this area are discussed. This article is part of the theme issue ‘Recent progress and open frontiers in Turing’s theory of morphogenesis’.

Cell patterning by secretion-induced plasma membrane flows

Cells self-organize using reaction-diffusion and fluid-flow principles. Whether bulk membrane flows contribute to cell patterning has not been established. Here, using mathematical modelling, optogenetics and synthetic probes, we show that polarized exocytosis causes lateral membrane flows away from regions of membrane insertion. Plasma membrane-associated proteins with sufficiently low diffusion and/or detachment rates couple to the flows and deplete from areas of exocytosis. In rod-shaped fission yeast cells, zones of Cdc42 GTPase activity driving polarized exocytosis are limited by GTPase activating proteins (GAPs). We show that membrane flows pattern the GAP Rga4 distribution and coupling of a synthetic GAP to membrane flows is sufficient to establish the rod shape. Thus, membrane flows induced by Cdc42-dependent exocytosis form a negative feedback restricting the zone of Cdc42 activity.One Sentence SummaryExocytosis causes bulk membrane flows that drag associated proteins and form a negative feedback restricting the exocytic site.

Activation of G-protein coupled receptors is thermodynamically linked to lipid solvation

Preferential lipid solvation of the G-protein coupled A2A adenosine receptor (A2AR) is evaluated from 35 μsec of all-atom molecular dynamics simulation. A coarse-grained transition matrix algorithm is developed to overcome slow equilibration of the first solvation shell, obtaining statistically robust estimates of the free energy of solvation by different lipids for the receptor in different activation states. Results indicate preference for solvation by unsaturated chains, which favors the active receptor. A model for lipid-dependent GPCR activity is proposed in which the chemical potential of lipids in the bulk membrane modulates receptor activity. The enthalpy and entropy associated with moving saturated vs. unsaturated lipids from bulk to A2AR’s first solvation shell are compared. In the simulated mixture, saturated chains are disordered (i.e., obtain a favorable entropic contribution) when partitioning to the receptor surface, but this is outweighed by a favorable enthalpic contribution for unsaturated chains to occupy the first solvation shell.

Retromer forms low order oligomers on supported lipid bilayers

Retromer is a protein sorting device that orchestrates the selection and export of integral membrane proteins from the endosome via retrograde and plasma membrane recycling pathways. Long standing hypotheses regarding the Retromer sorting mechanism posit that oligomeric interactions between Retromer and associated accessory factors on the endosome membrane drives clustering of Retromer-bound integral membrane cargo prior to its packaging into a nascent transport carrier. To test this hypothesis, we examined interactions between the components of the SNX3-Retromer sorting pathway using quantitative single particle fluorescence microscopy of a reconstituted system comprising a supported bilayer, Retromer, a model cargo protein, the accessory proteins SNX3, RAB7, and the Retromer-binding segment of the WASHC2C subunit of the WASH complex. The predominant species of membrane associated Retromer are low order: monomers (∼18%), dimers (∼35%), trimers (∼24%) and tetramers (∼24%). Unexpectedly, neither cargo nor accessory factors promote Retromer oligomerization on a supported bilayer. The results indicate that Retromer has an intrinsic propensity to form low order oligomers and that neither membrane association nor accessory factors potentiate oligomerization. Hence, Retromer is a minimally concentrative sorting device adapted to bulk membrane trafficking from the endosomal system.

Transport Properties of Film and Bulk Sr0.98Zr0.95Y0.05O3−δ Membranes

In electrode-supported solid oxide fuel cells (SOFCs) with a thin electrolyte, the electrolyte performance can be affected by its interaction with the electrode, therefore, it is particularly important to study the charge transport properties of thin electrode-supported electrolytes. The transport numbers of charged species in Ni-cermet supported Sr0.98Zr0.95Y0.05O3−δ (SZY) membranes were studied and compared to those of the bulk membrane. SZY films of 2.5 μm thickness were fabricated by the chemical solution deposition technique. It was shown that the surface layer of the films contained 1.5–2 at.% Ni due to Ni diffusion from the substrate. The Ni-cermet supported 2.5 μm-thick membrane operating in the fuel cell mode was found to possess the effective transport number of oxygen ions of 0.97 at 550 °C, close to that for the bulk SZY membrane (0.99). The high ionic transport numbers indicate that diffusional interaction between SZY films and Ni-cermet supporting electrodes does not entail electrolyte degradation. The relationship between SZY conductivity and oxygen partial pressure was derived from the data on effective conductivity and ionic transport numbers for the membrane operating under two different oxygen partial pressure gradients—in air/argon and air/hydrogen concentration cells.

Surface modification of Nafion membranes by ion implantation to reduce methanol crossover in direct methanol fuel cells

The surface of Nafion membranes was modified by light ion implantation to reduce methanol crossover without affecting bulk membrane properties.