Homeocurvature adaptation of phospholipids to pressure in deep-sea invertebrates.

Hydrostatic pressure increases with depth in the ocean, but little is known about the molecular bases of biological pressure tolerance. We describe a mode of pressure adaptation in comb jellies (ctenophores) that also constrains these animals' depth range. Structural analysis of deep-sea ctenophore lipids shows that they form a nonbilayer phase at pressures under which the phase is not typically stable. Lipidomics and all-atom simulations identified phospholipids with strong negative spontaneous curvature, including plasmalogens, as a hallmark of deep-adapted membranes that causes this phase behavior. Synthesis of plasmalogens enhanced pressure tolerance in Escherichia coli, whereas low-curvature lipids had the opposite effect. Imaging of ctenophore tissues indicated that the disintegration of deep-sea animals when decompressed could be driven by a phase transition in their phospholipid membranes.

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.

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.

Membrane mimetic-dependence of GPCR energy landscapes.

We leveraged variable-temperature 19F-NMR spectroscopy to compare the conformational equilibria of the human A2A adenosine receptor (A2AAR), a class A G protein-coupled receptor (GPCR), across a range of temperatures ranging from lower temperatures typically employed in 19F-NMR experiments to physiological temperature. A2AAR complexes with partial agonists and full agonists showed large increases in the population of a fully active conformation with increasing temperature. NMR data measured at physiological temperature were more in line with functional data. This was pronounced for complexes with partial agonists, where the population of active A2AAR was nearly undetectable at lower temperature but became evident at physiological temperature. Temperature-dependent behavior of complexes with either full or partial agonists exhibited a pronounced sensitivity to the specific membrane mimetic employed. Cellular signaling experiments correlated with the temperature-dependent conformational equilibria of A2AAR in lipid nanodiscs but not in some detergents, underscoring the importance of the membrane environment in studies of GPCR function.

Bellwethers of change: population modelling of North Pacific humpback whales from 2002 through 2021 reveals shift from recovery to climate response.

For the 40 years after the end of commercial whaling in 1976, humpback whale populations in the North Pacific Ocean exhibited a prolonged period of recovery. Using mark-recapture methods on the largest individual photo-identification dataset ever assembled for a cetacean, we estimated annual ocean-basin-wide abundance for the species from 2002 through 2021. Trends in annual estimates describe strong post-whaling era population recovery from 16 875 (± 5955) in 2002 to a peak abundance estimate of 33 488 (± 4455) in 2012. An apparent 20% decline from 2012 to 2021, 33 488 (± 4455) to 26 662 (± 4192), suggests the population abruptly reached carrying capacity due to loss of prey resources. This was particularly evident for humpback whales wintering in Hawai'i, where, by 2021, estimated abundance had declined by 34% from a peak in 2013, down to abundance levels previously seen in 2006, and contrasted to an absence of decline in Mainland Mexico breeding humpbacks. The strongest marine heatwave recorded globally to date during the 2014-2016 period appeared to have altered the course of species recovery, with enduring effects. Extending this time series will allow humpback whales to serve as an indicator species for the ecosystem in the face of a changing climate.

Membrane Mimetic-Dependence of GPCR Energy Landscapes.

Protein function strongly depends on temperature, which is related to temperature-dependent changes in the equilibria of protein conformational states. We leveraged variable-temperature 19F-NMR spectroscopy to interrogate the temperature dependence of the conformational landscape of the human A2A adenosine receptor (A2AAR), a class A GPCR. Temperature-induced changes in the conformational equilibria of A2AAR in lipid nanodiscs were markedly dependent on the efficacy of bound drugs. While antagonist complexes displayed only modest changes as the temperature rose, both full and partial agonist complexes exhibited substantial increases in the active state population. Importantly, the temperature-dependent response of complexes with both full and partial agonists exhibited a pronounced sensitivity to the specific membrane mimetic employed. In striking contrast to observations within lipid nanodiscs, in detergent micelles the active state population exhibited different behavior for A2AAR complexes with both full and partial agonists. This underscores the importance of the protein environment in understanding the thermodynamics of GPCR activation.

A collaborative and near-comprehensive North Pacific humpback whale photo-ID dataset.

We present an ocean-basin-scale dataset that includes tail fluke photographic identification (photo-ID) and encounter data for most living individual humpback whales (Megaptera novaeangliae) in the North Pacific Ocean. The dataset was built through a broad collaboration combining 39 separate curated photo-ID catalogs, supplemented with community science data. Data from throughout the North Pacific were aggregated into 13 regions, including six breeding regions, six feeding regions, and one migratory corridor. All images were compared with minimal pre-processing using a recently developed image recognition algorithm based on machine learning through artificial intelligence; this system is capable of rapidly detecting matches between individuals with an estimated 97-99% accuracy. For the 2001-2021 study period, a total of 27,956 unique individuals were documented in 157,350 encounters. Each individual was encountered, on average, in 5.6 sampling periods (i.e., breeding and feeding seasons), with an annual average of 87% of whales encountered in more than one season. The combined dataset and image recognition tool represents a living and accessible resource for collaborative, basin-wide studies of a keystone marine mammal in a time of rapid ecological change.

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.

Surface viscosities of lipid bilayers determined from equilibrium molecular dynamics simulations.

Lipid membrane viscosity is critical to biological function. Bacterial cells grown in different environments alter their lipid composition in order to maintain a specific viscosity, and membrane viscosity has been linked to the rate of cellular respiration. To understand the factors that determine the viscosity of a membrane, we ran equilibrium all-atom simulations of single component lipid bilayers and calculated their viscosities. The viscosity was calculated via a Green-Kubo relation, with the stress-tensor autocorrelation function modeled by a stretched exponential function. By simulating a series of lipids at different temperatures, we establish the dependence of viscosity on several aspects of lipid chemistry, including hydrocarbon chain length, unsaturation, and backbone structure. Sphingomyelin is found to have a remarkably high viscosity, roughly 20 times that of DPPC. Furthermore, we find that inclusion of the entire range of the dispersion interaction increases viscosity by up to 140%. The simulated viscosities are similar to experimental values obtained from the rotational dynamics of small chromophores and from the diffusion of integral membrane proteins but significantly lower than recent measurements based on the deformation of giant vesicles.

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.

Cholesterol Dependent Activity of the Adenosine A2A Receptor Is Modulated via the Cholesterol Consensus Motif.

BACKGROUND: Membrane cholesterol dysregulation has been shown to alter the activity of the adenosine A2A receptor (A2AR), a G protein-coupled receptor, thereby implicating cholesterol levels in diseases such as Alzheimer's and Parkinson's. A limited number of A2AR crystal structures show the receptor interacting with cholesterol, as such molecular simulations are often used to predict cholesterol interaction sites. METHODS: Here, we use experimental methods to determine whether a specific interaction between amino acid side chains in the cholesterol consensus motif (CCM) of full length, wild-type human A2AR, and cholesterol modulates activity of the receptor by testing the effects of mutational changes on functional consequences, including ligand binding, G protein coupling, and downstream activation of cyclic AMP. RESULTS AND CONCLUSIONS: Our data, taken with previously published studies, support a model of receptor state-dependent binding between cholesterol and the CCM, whereby cholesterol facilitates both G protein coupling and downstream signaling of A2AR.

Humpback whales (Megaptera novaeangliae) attend both Mexico and Hawaii breeding grounds in the same winter: mixing in the northeast Pacific.

Humpback whales that assemble on winter breeding grounds in Mexico and Hawaii have been presumed to be, at least, seasonally isolated. Recently, these assemblies were declared Distinct Population Segments under the US Endangered Species Act. We report two humpback whales attending both breeding grounds in the same season-one moving from Hawaii to Mexico and the other from Mexico to Hawaii. The first was photo-identified in Maui, Hawaii on 23 February 2006 and again, after 53 days and 4545 km, on 17 April 2006 in the Revillagigedo Archipelago, Mexico. The second was photo-identified off Guerrero, Mexico on 16 February 2018 and again, 49 days and 5944 km later, on 6 April 2018 off Maui. The 2006 whale was identified in summer off Kodiak Island, Alaska; the 2018 whale off British Columbia. These Mexico-Hawaii identifications provide definitive evidence that whales in these two winter assemblies may mix during one winter season. This, combined with other lines of evidence on Mexico-Hawaii mixing, including interchange of individuals year to year, long-term similarity of everchanging songs, one earlier same-season travel record, and detection of humpback whales mid-ocean between these locations in winter, suggests reassessment of the 'distinctiveness' of these populations may be warranted.

Full scale structural, mechanical and dynamical properties of HIV-1 liposomes.

Enveloped viruses are enclosed by a lipid membrane inside of which are all of the components necessary for the virus life cycle; viral proteins, the viral genome and metabolites. Viral envelopes are lipid bilayers that adopt morphologies ranging from spheres to tubes. The envelope is derived from the host cell during viral replication. Thus, the composition of the bilayer depends on the complex constitution of lipids from the host-cell's organelle(s) where assembly and/or budding of the viral particle occurs. Here, molecular dynamics (MD) simulations of authentic, asymmetric HIV-1 liposomes are used to derive a unique level of resolution of its full-scale structure, mechanics and dynamics. Analysis of the structural properties reveal the distribution of thicknesses of the bilayers over the entire liposome as well as its global fluctuations. Moreover, full-scale mechanical analyses are employed to derive the global bending rigidity of HIV-1 liposomes. Finally, dynamical properties of the lipid molecules reveal important relationships between their 3D diffusion, the location of lipid-rafts and the asymmetrical composition of the envelope. Overall, our simulations reveal complex relationships between the rich lipid composition of the HIV-1 liposome and its structural, mechanical and dynamical properties with critical consequences to different stages of HIV-1's life cycle.

Changes in pairwise correlations during running reshape global network state in the main olfactory bulb.

Neural codes for sensory inputs have been hypothesized to reside in a broader space defined by ongoing patterns of spontaneous activity. To understand the structure of this spontaneous activity in the olfactory system, we performed high-density recordings of neural populations in the main olfactory bulb of awake mice. We observed changes in pairwise correlations of spontaneous activity between mitral and tufted (M/T) cells when animals were running, which resulted in an increase in the entropy of the population. Surprisingly, pairwise maximum entropy models that described the population activity using only assumptions about the firing rates and correlations of neurons were better at predicting the global structure of activity when animals were stationary as compared to when they were running, implying that higher order (3rd, 4th order) interactions governed population activity during locomotion. Taken together, we found that locomotion alters the functional interactions that shape spontaneous population activity at the earliest stages of olfactory processing, one synapse away from the sensory receptors in the nasal epithelium. These data suggest that the coding space available for sensory representations responds adaptively to the animal's behavioral state.NEW & NOTEWORTHY The organization and structure of spontaneous population activity in the olfactory system places constraints of how odor information is represented. Using high-density electrophysiological recordings of mitral and tufted cells, we found that running increases the dimensionality of spontaneous activity, implicating higher order interactions among neurons during locomotion. Behavior, thus, flexibly alters neuronal activity at the earliest stages of sensory processing.

Curvature Energetics Determined by Alchemical Simulation on Four Topologically Distinct Lipid Phases.

The relative curvature energetics of two lipids are tested using thermodynamic integration (TI) on four topologically distinct lipid phases. Simulations use TI to switch between choline headgroup lipids (POPC; that prefers to be flat) and ethanolamine headgroup lipids (POPE; that prefer, for example, the inner monolayer of vesicles). The thermodynamical moving of the lipids between planar, inverse hexagonal (HII), cubic (QII; Pn3m space group), and vesicle topologies reveals differences in material parameters that were previously challenging to access. The methodology allows for predictions of two important lipid material properties: the difference in POPC/POPE monolayer intrinsic curvature (ΔJ0) and the difference in POPC/POPE monolayer Gaussian curvature modulus (Δκ̅m), both of which are connected to the energetics of topological variation. Analysis of the TI data indicates that, consistent with previous experiment and simulation, the J0 of POPE is more negative than POPC (ΔJ0 = -0.018 ± 0.001 Å-1). The theoretical framework extracts significant differences in κ̅m of which POPE is less negative than POPC by 2.0 to 4.0 kcal/mol. The range of these values is determined by considering subsets of the simulations, and disagreement between these subsets suggests separate mechanical parameters at very high curvature. Finally, the fit of the TI data to the model indicates that the position of the pivotal plane of curvature is not constant across topologies at high curvature. Overall, the results offer insights into lipid material properties, the limits of a single HC model, and how to test them using simulation.

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 µs of all-atom molecular dynamics simulation. A coarse-grained transition matrix algorithm is developed to overcome slow equilibration of the first solvation shell, obtaining 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 G-protein-coupled receptor activity is proposed in which the chemical potential of lipids in the bulk membrane modulates receptor activity. The entropies associated with moving saturated and unsaturated lipids from bulk to A2AR's first solvation shell are evaluated. Overall, the acyl chains are more disordered (i.e., obtain a favorable entropic contribution) when partitioning to the receptor surface, and this effect is augmented for the saturated chains, which are relatively more ordered in bulk.

Composition dependence of cholesterol flip-flop rates in physiological mixtures.

Unbiased, all-atom simulations of mixtures representative of the inner and outer leaflets of a mammalian red blood cell and a synaptic vesicle reveal many cholesterol flip-flop events over the 5 µsec duration of the simulations. Enough events are observed for a direct estimate of the flip-flop rate. Slower rates are found in more ordered membranes, and faster rates in more disordered membranes, consistent with earlier reports in the literature. However, the rates found here are neither as fast as the fastest nor as slow as the slowest rates obtained by previous simulations. The difference likely stems from the compositions studied here, which unlike previous work include exclusively lipids with differing acyl chains, as observed in mammalian lipidomes.

Laterally Resolved Small-Angle Scattering Intensity from Lipid Bilayer Simulations: An Exact and a Limited-Range Treatment.

When combined, molecular simulations and small-angle scattering experiments are able to provide molecular-scale resolution of structure. Separately, scattering experiments provide only intermingled pair correlations between atoms, while molecular simulations are limited by model quality and the relatively short time scales that they can access. Their combined strength relies on agreement between the experimental spectra and those computed by simulation. To date, computing the neutron spectra from a molecular simulation of a lipid bilayer is straightforward only if the structure is approximated by laterally averaging the in-plane bilayer structure. However, this neglects all information about lateral heterogeneity, e.g., clustering of components in a lipid mixture. This paper presents two methods for computing the scattering intensity of simulated bilayers with in-plane heterogeneity, enabling a full treatment of both the transverse and lateral bilayer structure for the first time. The first method, termed the Dirac Brush, computes the exact spectra including spurious artifacts resulting from using information from neighboring periodic cells to account for the long-range structure of the bilayer. The second method, termed PFFT, applies a mean-field treatment in the field far from a scattering element, resulting in a correlation range that can be tuned (eliminating correlations with neighboring periodic images), but with computational cost that prohibits obtaining the exact (Dirac Brush) spectra. Following their derivation, the two methods are applied to a coarse-grained molecular simulation of a bilayer inhomogeneity, demonstrating the contributions of lateral correlations to the resulting spectra.

Ultrafast Formation of the Charge Transfer State of Prodan Reveals Unique Aspects of the Chromophore Environment.

Lipophilic dyes such as laurdan and prodan are widely used in membrane biology due to a strong bathochromic shift in emission that reports the structural parameters of the membrane such as area per molecule. Disentangling of the factors which control the spectral shift is complicated by the stabilization of a charge-transfer-like excitation of the dye in polar environments. Predicting the emission therefore requires modeling both the relaxation of the environment and the corresponding evolution of the excited state. Here, an approach is presented in which (i) the local environment is sampled by a classical molecular dynamics (MD) simulation of the dye and solvent, (ii) the electronically excited state of prodan upon light absorption is predicted by numerical quantum mechanics (QM), (iii) the iterative relaxation of the environment around the excited dye by MD coupled with the evolution of the excited state is performed, and (iv) the emission properties are predicted by QM. The QM steps are computed using the many-body Green's function in the GW approximation and the Bethe-Salpeter equation with the environment modeled as fixed point charges, sampled in the MD simulation steps. The comparison to ultrafast time-resolved transient absorption measurements demonstrates that the iterative molecular mechanics (MM)/QM approach agrees quantitatively with both the polarity-dependent shift in emission and the time scale over which the charge transfer state is stabilized. Together the simulations and experimental measurements suggest that the evolution into the charge transfer state is slower in amphiphilic solvents.