Maintenance of membrane phospholipid asymmetry Lipid-cytoskeletal interactions or lipid pump?

Two models for the mechanism of maintenance of lipid asymmetry in erythrocytes are considered: binding of internal lipids to cytoskeletal proteins, and pumping of internal lipids from the outside to the inside of the cell. Analysis of the kinetics of lipid internalization suggests that the first model is more likely, and that the apparent pumping of lipids represents the activity of an ATP-dependent lipid flip/flop catalyst.     

Biophysics of α-synuclein membrane interactions

Membrane proteins participate in nearly all cellular processes; however, because of experimental limitations, their characterization lags far behind that of soluble proteins. Peripheral membrane proteins are particularly challenging to study because of their inherent propensity to adopt multiple and/or transient conformations in solution and upon membrane association. In this review, we summarize useful biophysical techniques for the study of peripheral membrane proteins and their application in the characterization of the membrane interactions of the natively unfolded and Parkinson's disease (PD) related protein, α-synuclein (α-syn). We give particular focus to studies that have led to the current understanding of membrane-bound α-syn structure and the elucidation of specific membrane properties that affect α-syn-membrane binding. Finally, we discuss biophysical evidence supporting a key role for membranes and α-syn in PD pathogenesis. This article is part of a Special Issue entitled: Membrane protein structure and function.     

Enhanced bactericidal activity of platelet β-lysin forE. coli in the presence of membrane perturbation

Inhibition by sodium chloride of the growth of 19 strains ofLegionella pneumophila and of 10 strains of otherLegionella spp. was studied. Results from growth in buffered -ketoglutarate cysteine yeast extract (BAYE) broth containing 0 to 2.0% sodium chloride indicated that 15/19 laboratory strains ofL. pneumophila were capable of growing in 1.0% to 1.5% sodium chloride, whereas 4 strains ofL. pneumophila and 10 strains of 6 other species were not.L. micdadei andL. longebeachae were the most inhibited in BAYE broth, growing only in concentrations of 0.5% sodium chloride. These in vitro studies indicate thatL. micdadei andL. longbeachae might be differentiated from other species by their low tolerance to salt in BAYE broth, and thatL. pneumophila may be more tolerant to salt concentrations found in brackish water environments.     

The role of membrane thickness in charged protein–lipid interactions

Charged amino acids are known to be important in controlling the actions of integral and peripheral membrane proteins and cell disrupting peptides. Atomistic molecular dynamics studies have shed much light on the mechanisms of membrane binding and translocation of charged protein groups, yet the impact of the full diversity of membrane physico-chemical properties and topologies has yet to be explored. Here we have performed a systematic study of an arginine (Arg) side chain analog moving across saturated phosphatidylcholine (PC) bilayers of variable hydrocarbon tail length from 10 to 18 carbons. For all bilayers we observe similar ion-induced defects, where Arg draws water molecules and lipid head groups into the bilayers to avoid large dehydration energy costs. The free energy profiles all exhibit sharp climbs with increasing penetration into the hydrocarbon core, with predictable shifts between bilayers of different thickness, leading to barrier reduction from 26 kcal/mol for 18 carbons to 6 kcal/mol for 10 carbons. For lipids of 10 and 12 carbons we observe narrow transmembrane pores and corresponding plateaus in the free energy profiles. Allowing for movements of the protein and side chain snorkeling, we argue that the energetic cost for burying Arg inside a thin bilayer will be small, consistent with recent experiments, also leading to a dramatic reduction in pK_a shifts for Arg. We provide evidence that Arg translocation occurs via an ion-induced defect mechanism, except in thick bilayers (of at least 18 carbons) where solubility-diffusion becomes energetically favored. Our findings shed light on the mechanisms of ion movement through membranes of varying composition, with implications for a range of charged protein–lipid interactions and the actions of cell-perturbing peptides. This article is part of a Special Issue entitled: Membrane protein structure and function.     

Disulfide-mediated interactions of the chlamydial major outer membrane protein: role in the differentiation of chlamydiae?

The effects of exogenous reducing agents on a number of biological properties of purified Chlamydia trachomatis LGV-434 and Chlamydia psittaci meningopneumonitis elementary bodies (EBs) have been examined in an attempt to identify in vitro correlates of early events in the differentiation of the infectious EB to the replicative cell type, the reticulate body (RB). Treatment of EBs with dithiothreitol elicited a number of changes normally associated with differentiation to the RB. EBs in the presence of 10 mM dithiothreitol displayed enhanced rates of [14C]glutamate oxidation, reduced infectivity, and decreased osmotic stability, and their Machiavello staining properties changed to those characteristic of the RB. A true differentiation of EB to RB did not take place under these conditions, since EBs treated in this manner and examined by transmission electron microscopy did not demonstrate increased size or decreased electron density as do isolated RBs. Additional studies were initiated to identify the macromolecules involved in this process. With polyacrylamide gel electrophoresis and immunoblotting procedures with monoclonal and polyclonal monospecific antibodies, the chlamydial major outer membrane protein was found to be the predominant component that varied under reducing versus nonreducing conditions. Furthermore, the extent of disulfide-mediated cross-linking of the major outer membrane protein varied between the infective and replicative forms of the C. trachomatis LGV-434 life cycle. Implications of disulfide interactions in the life cycle of chlamydiae are discussed.     

Can membrane composition traffic toxins? Mycolactone and preferential membrane interactions

Mycolactone is a cytotoxic and immunosuppressive macrolide produced by Mycobacterium ulcerans and the sole causative agent of the neglected tropical skin disease Buruli ulcer. The toxin acts by invading host cells and interacting with intracellular targets to disrupt multiple fundamental cellular processes. Mycolactone’s amphiphilic nature enables strong interactions with lipophilic environments, including cellular membranes; however, the specificity of these interactions and the role of membranes in the toxin’s pathogenicity remain unknown. It is likely that preferential interactions with lipophilic carriers play a key role in the toxin’s distribution in the host, which, if understood, could provide insights to aid in the development of needed diagnostics for Buruli ulcer disease. In this work, molecular dynamics simulations were combined with enhanced free-energy sampling to characterize mycolactone’s association with and permeation through models of the mammalian endoplasmic reticulum (ER) and plasma membranes (PMs). We find that increased order in the PMs not only leads to a different permeation mechanism compared with that in the ER membrane but also an energetic driving force for ER localization. Increased hydration, membrane deformation, and preferential interactions with unsaturated lipid tails stabilize the toxin in the ER membrane, while disruption of lipid packing is a destabilizing force in the PMs.     

Control and role of pH in peptide–lipid interactions in oriented membrane samples

To understand the molecular mechanisms of amphiphilic membrane-active peptides, one needs to study their interactions with lipid bilayers under ambient conditions. However, it is difficult to control the pH of the sample in biophysical experiments that make use of mechanically aligned multilamellar membrane stacks on solid supports. HPLC-purified peptides tend to be acidic and can change the pH in the sample significantly. Here, we have systematically studied the influence of pH on the lipid interactions of the antimicrobial peptide PGLa embedded in oriented DMPC/DMPG bilayers. Using solid-state NMR (³¹P, ²H, ¹⁹F), both the lipid and peptide components were characterized independently, though in the same oriented samples under typical conditions of maximum hydration. The observed changes in lipid polymorphism were supported by DSC on multilamellar liposome suspensions. On this basis, we can present an optimized sample preparation protocol and discuss the challenges of performing solid-state NMR experiments under controlled pH. DMPC/DMPG bilayers show a significant up-field shift and broadening of the main lipid phase transition temperature when lowering the pH from 10.0 to 2.6. Both, strongly acidic and basic pH, cause a significant degree of lipid hydrolysis, which is exacerbated by the presence of PGLa. The characteristic re-alignment of PGLa from a surface-bound to a tilted state is not affected between pH of 7 to 4 in fluid bilayers. On the other hand, in gel-phase bilayers the peptide remains isotropically mobile under acidic conditions, displays various co-existing orientational states at pH 7, and adopts an unknown structural state at basic pH.     

Emerging roles of cortical microtubule–membrane interactions

Plant cortical microtubules have crucial roles in cell wall development. Cortical microtubules are tightly anchored to the plasma membrane in a highly ordered array, which directs the deposition of cellulose microfibrils by guiding the movement of the cellulose synthase complex. Cortical microtubules also interact with several endomembrane systems to regulate cell wall development and other cellular events. Recent studies have identified new factors that mediate interactions between cortical microtubules and endomembrane systems including the plasma membrane, endosome, exocytic vesicles, and endoplasmic reticulum. These studies revealed that cortical microtubule-membrane interactions are highly dynamic, with specialized roles in developmental and environmental signaling pathways. A recent reconstructive study identified a novel function of the cortical microtubule-plasma membrane interaction, which acts as a lateral fence that defines plasma membrane domains. This review summarizes recent advances in our understanding of the mechanisms and functions of cortical microtubule-membrane interactions.     

Friend or Foe? Implication of the autophagy-lysosome pathway in SARS-CoV-2 infection and COVID-19

There is increasing amount of evidence indicating the close interplays between the replication cycle of SARS-CoV-2 and the autophagy-lysosome pathway in the host cells. While autophagy machinery is known to either assist or inhibit the viral replication process, the reciprocal effects of the SARS-CoV-2 on the autophagy-lysosome pathway have also been increasingly appreciated. More importantly, despite the disappointing results from the clinical trials of chloroquine and hydroxychloroquine in treatment of COVID-19, there is still ongoing effort in discovering new therapeutics targeting the autophagy-lysosome pathway. In this review, we provide an update-to-date summary of the interplays between the autophagy-lysosome pathway in the host cells and the pathogen SARS-CoV-2 at the molecular level, to highlight the prognostic value of autophagy markers in COVID-19 patients and to discuss the potential of developing novel therapeutic strategies for COVID-19 by targeting the autophagy-lysosome pathway. Thus, understanding the nature of such interactions between SARS-CoV-2 and the autophagy-lysosome pathway in the host cells is expected to provide novel strategies in battling against this global pandemic.     

Climate or diet? The importance of biotic interactions in determining species range size

Aim

Species geographical range sizes play a crucial role in determining species vulnerability to extinction. Although several mechanisms affect range sizes, the number of biotic interactions and species climatic tolerance are often thought to play discernible roles, defining two dimensions of the Hutchinsonian niche. Yet, the relative importance of the trophic and the climatic niche for determining species range sizes is largely unknown.

Location

Central and northern Europe.

Time period

Present.

Major taxa studied

Gall-inducing sawflies and their parasitoids.

Methods

We use data documenting the spatial distributions and biotic interactions of 96 herbivore species, and their 125 parasitoids, across Europe and analyse the relationship between species range size and the climatic and trophic dimensions of the niche. We then compare the observed relationships with null expectations based on species occupancy to understand whether the relationships observed are an inevitable consequence of species range size or if they contain information about the importance of each dimension of the niche on species range size.

Results

We find that both niche dimensions are positively correlated with species range size, with larger ranges being associated with wider climatic tolerances and larger numbers of interactions. However, diet breadth appears to more strongly limit species range size. Species with larger ranges have more interactions locally and they are also able to interact with a larger diversity of species across sites (i.e. higher β-diversity), resulting in a larger number of interactions at continental scales.

Main conclusions

We show for the first time how different aspects of species diet niches are related to their range size. Our study offers new insight into the importance of biotic interactions in determining species spatial distributions, which is critical for improving understanding and predictions of species vulnerability to extinction under the current rates of global environmental change.     

Gene-environment interactions in human disease: nuisance or opportunity?

Many environmental risk factors for common, complex human diseases have been revealed by epidemiologic studies, but how genotypes at specific loci modulate individual responses to environmental risk factors is largely unknown. Gene-environment interactions will be missed in genome-wide association studies and could account for some of the 'missing heritability' for these diseases. In this review, we focus on asthma as a model disease for studying gene-environment interactions because of relatively large numbers of candidate gene-environment interactions with asthma risk in the literature. Identifying these interactions using genome-wide approaches poses formidable methodological problems, and elucidating molecular mechanisms for these interactions has been challenging. We suggest that studying gene-environment interactions in animal models, although more tractable, might not be sufficient to shed light on the genetic architecture of human diseases. Lastly, we propose avenues for future studies to find gene-environment interactions.     

The effects and mechanism of flavonoid–rePON1 interactions. Structure–activity relationship study

Flavonoids are plant phenolic secondary metabolites that are widely distributed in the human diet. These antioxidants have received much attention because of their neuroprotective, cardioprotective, and chemopreventive actions. While a major focus has been on the flavonoids’ antioxidant properties, there is an emerging view that many of the potential health benefits of flavonoids and their in vivo metabolites are due to modulatory actions in cells through direct interactions with proteins, and not necessarily due to their antioxidant function. This view relies on the observations that flavonoids are present in the circulation at very low concentrations, which are not sufficient to exert effective antioxidant effects. The enzyme paraoxonase 1 (PON1) is associated with high-density lipoprotein (HDL), and is responsible for many of HDLs’ antiatherogenic properties. We previously showed that the flavonoid glabridin binds to rePON1 and affects the enzyme’s 3D structure. This interaction protects the enzyme from inhibition by an atherogenic component of the human carotid plaque. Here, we broadened our study to an investigation of the structure–activity relationships (SARs) of 12 flavonoids from different subclasses with rePON1 using Trp-fluorescence quenching, modeling calculations and Cu²⁺-induced low-density lipoprotein (LDL) oxidation methods. Our findings emphasize the ‘protein-binding’ mechanism by which flavonoids exert their beneficial biological role toward rePON1. Flavonoids’ capacity to interact with the enzyme’s rePON1 hydrophobic groove mostly dictates their pro/antioxidant behavior.     

Do Polyamines Participate in the Long-Distance Translocation of Stress Signals in Plants?

Accumulation and ethylene-dependent translocation of free polyamines was studied in various organs, the phloem and xylem exudates of common ice plants (Mesembryanthemum crystallinum L.). Under normal conditions (23–25°C), spermidine predominated among polyamines. Cadaverine was found in old leaves, stems, and, in large quantities, in roots. The heat shock treatment (HS; 47°C, 2 h) of intact plant shoots induced intense evolution of ethylene from leaves but reduced the leaf content of polyamines. Under these conditions, the concentration of polyamines in roots, particularly of cadaverine, increased many times. The HS treatment of roots (40°C, 2 h) induced translocation of cadaverine to stems and putrescine to leaves. An enhanced polyamine content after HS treatment was also found in the xylem and phloem exudates. The exposure of detached leaves to ethylene led to a reduction in their putrescine and spermidine and accumulation of cadaverine, which implies the ethylene-dependent formation of cadaverine and a possible relation between the HS-induced translocation of this diamine to roots and the transient ethylene evolution by leaves. To validate this hypothesis, we compared the ethylene evolution rate and interorgan partitioning of cadaverine and other polyamines for two lines of Arabidopsis thaliana: the wild type (Col-0) and ein4 mutant with impaired ethylene reception. In plants grown in light at 20–21°C, the rate of ethylene evolution by rosetted leaves was higher in the mutant than in the wild type. The content of putrescine and spermidine was reduced in mutant leaves, whereas cadaverine concentration increased almost threefold compared with the wild type. In roots, cadaverine was found only in the wild type and not in the mutant line. Our data indicate the ethylene-dependent formation of cadaverine in leaves and possible involvement of cadaverine and ethylene in the long-distance translocation of stress (HS) signal in plants.     

Misfolding of membrane proteins in health and disease: the lady or the tiger?

Protein misfolding is increasingly recognized as a factor in many diseases, including cystic fibrosis, Parkinson's, Alzheimer's and atherosclerosis. Many proteins involved in misfolding-based pathologies are membrane-associated, such that the bilayer may play roles in normal and aberrant folding. It can be argued that the in vivo partitioning of eukaryotic membrane proteins between folding and misfolding pathways is under kinetic control. Moreover, the balance between these pathways can be surprisingly delicate.     

Is inward calcium current in crayfish muscle membrane constituted of one or two components?

Two inward currents were observed in crayfish muscle membrane during depolarization steps by the method described by Adrian et al. (1970). Under voltage clamp conditions, hyperpolarization steps elicited a large current (leak current If), associated with an inward voltage dependent current. This inward current was inhibited by niflumic acid (NA), a drug known to block Cl---HCO-3 exchange (Cousin et Motais 1982; Br?lè et al. 1983b). Dynamic outward currents triggered by depolarizing steps were inhibited to a great extent by TEA, the not inhibited portion disappearing when procaine (2 mmol/l) was added to external solution. In the presence of TEA, procaine and NA, it was thus possible to dissect the regenerative calcium current (ICa) into two components: a "fast component" (ICa1) and a "slow component" (ICa2). The reversal potential of ICa was 65 mV (for [Ca]0 = 2.8 mmol/l), and [Ca]i could be calculated to be 1.6 X 10(-5) mol/l. This value of [Ca]i is the same as calculated from values reported by Hencek and Zachar (1977). ICa1 was triggered at a threshold membrane potential of -45 mV and ICa2 at -30 mV. Moreover, the inactivation kinetics for ICa1 was faster than that for ICa2. Our results are in perfect agreement with those obtained by Zahradník and Zachar (1982) who postulated two populations of calcium channels.     

Impairment of the activity of the plasma membrane Ca²⁺-ATPase in Alzheimer's disease

AD (Alzheimer's disease) is an age-associated neurodegenerative disorder where the accumulation of neurotoxic Aβ (amyloid β-peptide) in senile plaques is a typical feature. Recent studies point out a relationship between Aβ neurotoxicity and Ca2+ dyshomoeostasis, but the molecular mechanisms involved are still under discussion. The PMCAs (plasma membrane Ca2+-ATPases) are a multi-isoform family of proteins highly expressed in brain that is implicated in the maintenance of low intraneural Ca2+ concentration. Therefore the malfunction of this pump may also be responsible for Ca2+ homoeostasis failure in AD. We have found that the Ca2+-dependence of PMCA activity is affected in human brains diagnosed with AD, being related to the enrichment of Aβ. The peptide produces an inhibitory effect on the activity of PMCA which is isoform-specific, with the greatest inhibition of PMCA4. Besides, cholesterol blocked the inhibitory effect of Aβ, which is consistent with the lack of any Aβ effect on PMCA4 found in cholesterol-enriched lipid rafts isolated from pig brain. These observations suggest that PMCAs are a functional component of the machinery that leads to Ca2+ dysregulation in AD and propose cholesterol enrichment in rafts as a protector of the Aβ-mediated inhibition on PMCA.