Varying the position of phospholipid acyl chain unsaturation modulates hopanoid and sterol ordering.

The cell membrane must balance mechanical stability with fluidity to function as both a barrier and an organizational platform. Key to this balance is the ordering of hydrocarbon chains and the packing of lipids. Many eukaryotes synthesize sterols, which are uniquely capable of modulating the lipid order to decouple membrane stability from fluidity. Ancient sterol analogs known as hopanoids are found in many bacteria and proposed as ancestral ordering lipids. The juxtaposition of sterols and hopanoids in extant organisms prompts us to ask why both pathways persist, especially in light of their convergent ability to order lipids. In this work, simulations, monolayer experiments, and cellular assays show that hopanoids and sterols order unsaturated phospholipids differently based on the position of double bonds in the phospholipid acyl chain. We find that cholesterol and diplopterol's methyl group distributions lead to distinct effects on unsaturated lipids. In Mesoplasma florum, diplopterol's constrained ordering capacity reduces membrane resistance to osmotic stress, unlike cholesterol. These findings suggest that cholesterol's broader lipid-ordering ability may have facilitated the exploration of a more diverse lipidomic landscape in eukaryotic membranes.

A cross-sectional, observational study of nutritional status and eating behaviours in people living with dementia in acute care settings.

AIMS AND OBJECTIVES: To understand the nutritional status, observing eating difficulties during mealtimes for people living with dementia in acute care settings. BACKGROUND: Changed eating behaviours caused by declining cognitive function is common in people living with dementia which can lead to malnutrition. Malnutrition is associated with prolonged hospitalisation and increased mortality. People living with dementia in acute care settings are at high risk of malnutrition. This highlights the importance of better understanding the nutritional intake and eating behaviours of people living with dementia in acute care settings. DESIGN: This study is a cross-sectional, observational study. METHODS: Data of mealtime difficulties and nutritional status of people living with dementia were collected in four geriatric care wards (in acute or sub-acute hospitals) by using Feeding Difficulty Index and Mini Nutritional Assessment Short-Form. The STROBE checklist was used throughout this study. RESULTS: The study included 94 people living with dementia. The median age of the participants was 85.86 years old, with a Feeding Difficulty Index of 8.27 and had stayed in hospitals for average 14.46 days, with an average total feeding time of 24.61 min. Only 1.2% of participants were considered to be in normal nutritional status, whereas 72.1% were malnourished. All participants required partial or full assistance during mealtime. Participants with higher scores on the Feeding Difficulty Index have longer total feeding times, compared to those with lower scores. CONCLUSIONS: Malnutrition is prevalent in people living with dementia. People living with dementia demonstrate varying mealtime difficulties depending on the level of dependence. Mealtime assistance training programs are warranted and are beneficial for nursing staff and family members to improve their feeding skills and knowledge. NO PATIENT OR PUBLIC CONTRIBUTION: This study did not involve patients, service users, caregivers or members of the public. RELEVANCE TO CLINICAL PRACTICE: The study is relevant to clinical practice by identifying changed eating behaviours or mealtime difficulties in people living with dementia in acute care settings can significantly decrease the risk of malnutrition.

Spontaneous lipid binding to the nicotinic acetylcholine receptor in a native membrane.

The nicotinic acetylcholine receptor (nAChR) and other pentameric ligand-gated ion channels are native to neuronal membranes with an unusual lipid composition. While it is well-established that these receptors can be significantly modulated by lipids, the underlying mechanisms have been primarily studied in model membranes with few lipid species. Here, we use coarse-grained molecular dynamics simulation to probe specific binding of lipids in a complex quasi-neuronal membrane. We ran a total of 50 µs of simulations of a single nAChR in a membrane composed of 36 species of lipids. Competition between multiple lipid species produces a complex distribution. We find that overall, cholesterol selects for concave inter-subunit sites and polyunsaturated fatty acids select for convex M4 sites, while monounsaturated and saturated lipids are unenriched in the nAChR boundary. We propose the "density-threshold affinity" as a metric calculated from continuous density distributions, which reduces to a standard affinity in two-state binding. We find that the density-threshold affinity for M4 weakens with chain rigidity, which suggests that flexible chains may help relax packing defects caused by the conical protein shape. For any site, PE headgroups have the strongest affinity of all phospholipid headgroups, but anionic lipids still yield moderately high affinities for the M4 sites as expected. We observe cooperative effects between anionic headgroups and saturated chains at the M4 site in the inner leaflet. We also analyze affinities for individual anionic headgroups. When combined, these insights may reconcile several apparently contradictory experiments on the role of anionic phospholipids in modulating nAChR.

Untangling Direct and Domain-Mediated Interactions Between Nicotinic Acetylcholine Receptors in DHA-Rich Membranes.

At the neuromuscular junction (NMJ), the nicotinic acetylcholine receptor (nAChR) self-associates to give rise to rapid muscle movement. While lipid domains have maintained nAChR aggregates in vitro, their specific roles in nAChR clustering are currently unknown. In the present study, we carried out coarse-grained molecular dynamics simulations (CG-MD) of 1-4 nAChR molecules in two membrane environments: one mixture containing domain-forming, homoacidic lipids, and a second mixture consisting of heteroacidic lipids. Spontaneous dimerization of nAChRs was up to ten times more likely in domain-forming membranes; however, the effect was not significant in four-protein systems, suggesting that lipid domains are less critical to nAChR oligomerization when protein concentration is higher. With regard to lipid preferences, nAChRs consistently partitioned into liquid-disordered domains occupied by the omega-3 ([Formula: see text]-3) fatty acid, docosahexaenoic acid (DHA); enrichment of DHA boundary lipids increased with protein concentration, particularly in homoacidic membranes. This result suggests dimer formation blocks access of saturated chains and cholesterol, but not polyunsaturated chains, to boundary lipid sites.

Varying the position of phospholipid acyl chain unsaturation modulates hopanoid and sterol ordering.

The cell membrane must balance mechanical stability with fluidity to function as both a barrier and an organizational platform. Key to this balance is the thermodynamic ordering of lipids. Most Eukaryotes employ sterols, which are uniquely capable of modulating lipid order to decouple membrane stability from fluidity. Ancient sterol analogues known as hopanoids are found in many bacteria and are proposed as ancestral ordering lipids. The juxtaposition of sterols and hopanoids in extant organisms prompts us to ask why both pathways persist, especially in light of their convergent ability to order lipids. We reveal that both hopanoids and sterols order unsaturated phospholipids differently based on the position of double bonds in the phospholipid's acyl chain. We find that cholesterol and diplopterol's methyl group distributions lead to distinct effects on unsaturated lipids. In Mesoplasma florum, diplopterol's constrained ordering capacity reduces membrane resistance to osmotic stress, unlike cholesterol. These findings suggest cholesterol's broader lipid ordering ability may have facilitated the exploration of a more diverse lipidomic landscape in eukaryotic membranes.

Anionic Phospholipids Control Mechanisms of GPCR-G Protein Recognition.

G protein-coupled receptors (GPCRs) are embedded in phospholipids that strongly influence drug-stimulated signaling. Anionic lipids are particularly important for GPCR signaling complex formation, but a mechanism for this role is not understood. Using NMR spectroscopy, we visualized the impact of anionic lipids on the function-related conformational equilibria of the human A 2A adenosine receptor (A 2A AR) in bilayers containing defined mixtures of zwitterionic and anionic phospholipids. Anionic lipids primed the receptor to form complexes with G proteins through a conformational selection process. Without anionic lipids, signaling complex formation proceeded through a less favorable induced fit mechanism. In computational models, anionic lipids mimicked interactions between a G protein and positively charged residues in A 2A AR at the receptor intracellular surface, stabilizing a pre-activated receptor conformation. Replacing these residues strikingly altered the receptor response to anionic lipids in experiments. High sequence conservation of the same residues among all GPCRs supports a general role for lipid-receptor charge complementarity in signaling.

Anionic phospholipids control mechanisms of GPCR-G protein recognition.

G protein-coupled receptors (GPCRs) are embedded in phospholipids that strongly influence drug-stimulated signaling. Anionic lipids are particularly important for GPCR signaling complex formation, but a mechanism for this role is not understood. Using NMR spectroscopy, we explore the impact of anionic lipids on the function-related conformational equilibria of the human A2A adenosine receptor (A2AAR) in bilayers containing defined mixtures of zwitterionic and anionic phospholipids. Anionic lipids prime the receptor to form complexes with G proteins through a conformational selection process. Without anionic lipids, signaling complex formation proceeds through a less favorable induced fit mechanism. In computational models, anionic lipids mimic interactions between a G protein and positively charged residues in A2AAR at the receptor intracellular surface, stabilizing a pre-activated receptor conformation. Replacing these residues strikingly alters the receptor response to anionic lipids in experiments. High sequence conservation of the same residues among all GPCRs supports a general role for lipid-receptor charge complementarity in signaling.

Boundary lipids of the nicotinic acetylcholine receptor: Spontaneous partitioning via coarse-grained molecular dynamics simulation.

Reconstituted nicotinic acetylcholine receptors (nAChRs) exhibit significant gain-of-function upon addition of cholesterol to reconstitution mixtures, and cholesterol affects the organization of nAChRs within domain-forming membranes, but whether nAChR partitions to cholesterol-rich liquid-ordered ("raft" or lo) domains or cholesterol-poor liquid-disordered (ldo) domains is unknown. We use coarse-grained molecular dynamics simulations to observe spontaneous interactions of cholesterol, saturated lipids, and polyunsaturated (PUFA) lipids with nAChRs. In binary Dipalmitoylphosphatidylcholine:Cholesterol (DPPC:CHOL) mixtures, both CHOL and DPPC acyl chains were observed spontaneously entering deep "non-annular" cavities in the nAChR TMD, particularly at the subunit interface and the ß subunit center, facilitated by the low amino acid density in the cryo-EM structure of nAChR in a native membrane. Cholesterol was highly enriched in the annulus around the TMD, but this effect extended over (at most) 5-10 Å. In domain-forming ternary mixtures containing PUFAs, the presence of a single receptor did not significantly affect the likelihood of domain formation. nAChR partitioned to any cholesterol-poor ldo domain that was present, regardless of whether the ldo or lo domain lipids had PC or PE headgroups. Enrichment of PUFAs among boundary lipids was positively correlated with their propensity for demixing from cholesterol-rich phases. Long n-3 chains (tested here with Docosahexaenoic Acid, DHA) were highly enriched in annular and non-annular embedded sites, partially displacing cholesterol and completely displacing DPPC, and occupying sites even deeper within the bundle. Shorter n-6 chains were far less effective at displacing cholesterol from non-annular sites.

Direct binding of phosphatidylglycerol at specific sites modulates desensitization of a ligand-gated ion channel.

Pentameric ligand-gated ion channels (pLGICs) are essential determinants of synaptic transmission, and are modulated by specific lipids including anionic phospholipids. The exact modulatory effect of anionic phospholipids in pLGICs and the mechanism of this effect are not well understood. Using native mass spectrometry, coarse-grained molecular dynamics simulations and functional assays, we show that the anionic phospholipid, 1-palmitoyl-2-oleoyl phosphatidylglycerol (POPG), preferentially binds to and stabilizes the pLGIC, Erwinia ligand-gated ion channel (ELIC), and decreases ELIC desensitization. Mutations of five arginines located in the interfacial regions of the transmembrane domain (TMD) reduce POPG binding, and a subset of these mutations increase ELIC desensitization. In contrast, a mutation that decreases ELIC desensitization, increases POPG binding. The results support a mechanism by which POPG stabilizes the open state of ELIC relative to the desensitized state by direct binding at specific sites.