Machine learning–driven multiscale modeling reveals lipid-dependent dynamics of RAS signaling proteins

RAS is a signaling protein associated with the cell membrane that is mutated in up to 30% of human cancers. RAS signaling has been proposed to be regulated by dynamic heterogeneity of the cell membrane. Investigating such a mechanism requires near-atomistic detail at macroscopic temporal and spatial scales, which is not possible with conventional computational or experimental techniques. We demonstrate here a multiscale simulation infrastructure that uses machine learning to create a scale-bridging ensemble of over 100,000 simulations of active wild-type KRAS on a complex, asymmetric membrane. Initialized and validated with experimental data (including a new structure of active wild-type KRAS), these simulations represent a substantial advance in the ability to characterize RAS-membrane biology. We report distinctive patterns of local lipid composition that correlate with interfacially promiscuous RAS multimerization. These lipid fingerprints are coupled to RAS dynamics, predicted to influence effector binding, and therefore may be a mechanism for regulating cell signaling cascades.

Electrically Controlling and Optically Observing the Membrane Potential of Supported Lipid Bilayers

Supported lipid bilayers are a well-developed model system for the study of membranes and their associated proteins, such as membrane channels, enzymes, and receptors. These versatile model membranes can be made from various components, ranging from simple synthetic phospholipids to complex mixtures of constituents, mimicking the cell membrane with its relevant physiochemical and molecular phenomena. In addition, the high stability of supported lipids bilayers allows for their study via a wide array of experimental probes. In this work, we describe a platform for supported lipid bilayers that is accessible both electrically and optically. We show that the polarization of the supported membrane can be electrically controlled and optically probed using voltage-sensitive dyes. Membrane polarization dynamics is understood through electrochemical impedance spectroscopy and the analysis of the equivalent electrical circuit. We also describe the effect of the conducting electrode layer on the fluorescence of the optical probe through metal-induced energy transfer. We conclude with a discussion on possible applications of this platform for the study of voltage-dependent membrane proteins and other processes in membrane biology and surface science.

Deciphering a hexameric protein complex with Angstrom optical resolution

Cryogenic optical localization in three dimensions (COLD) was recently shown to resolve up to four binding sites on a single protein. However, because COLD relies on intensity fluctuations that result from the blinking behavior of fluorophores, it is limited to cases, where individual emitters show different brightness. This significantly lowers the measurement yield. To extend the number of resolved sites as well as the measurement yield, we employ partial labeling and combine it with polarization encoding in order to identify single fluorophores during their stochastic blinking. We then use a particle classification scheme to identify and resolve heterogenous subsets and combine them to reconstruct the three-dimensional arrangement of large molecular complexes. We showcase this method (polarCOLD) by resolving the trimer arrangement of proliferating cell nuclear antigen (PCNA) and the hexamer geometry of Caseinolytic Peptidase B (ClpB) of Thermus thermophilus in its quaternary structure, both with Angstrom resolution. The combination of polarCOLD and single-particle cryogenic electron microscopy (cryoEM) promises to provide crucial insight into intrinsic, environmental and dynamic heterogeneities of biomolecular structures. Furthermore, our approach is fully compatible with fluorescent protein labeling and can, thus, be used in a wide range of studies in cell and membrane biology.

A Comprehensive Review on the Interplay between Neisseria spp. and Host Sphingolipid Metabolites

Sphingolipids represent a class of structural related lipids involved in membrane biology and various cellular processes including cell growth, apoptosis, inflammation and migration. Over the past decade, sphingolipids have become the focus of intensive studies regarding their involvement in infectious diseases. Pathogens can manipulate the sphingolipid metabolism resulting in cell membrane reorganization and receptor recruitment to facilitate their entry. They may recruit specific host sphingolipid metabolites to establish a favorable niche for intracellular survival and proliferation. In contrast, some sphingolipid metabolites can also act as a first line defense against bacteria based on their antimicrobial activity. In this review, we will focus on the strategies employed by pathogenic Neisseria spp. to modulate the sphingolipid metabolism and hijack the sphingolipid balance in the host to promote cellular colonization, invasion and intracellular survival. Novel techniques and innovative approaches will be highlighted that allow imaging of sphingolipid derivatives in the host cell as well as in the pathogen.

Free energies of membrane stalk formation from a lipidomics perspective

Many biological membranes are asymmetric and exhibit complex lipid composition, comprising hundreds of distinct chemical species. Identifying the biological function and advantage of this complexity is a central goal of membrane biology. Here, we study how membrane complexity controls the energetics of the first steps of membrane fusions, that is, the formation of a stalk. We first present a computationally efficient method for simulating thermodynamically reversible pathways of stalk formation at coarse-grained resolution. The method reveals that the inner leaflet of a typical plasma membrane is far more fusogenic than the outer leaflet, which is likely an adaptation to evolutionary pressure. To rationalize these findings by the distinct lipid compositions, we computed ~200 free energies of stalk formation in membranes with different lipid head groups, tail lengths, tail unsaturations, and sterol content. In summary, the simulations reveal a drastic influence of the lipid composition on stalk formation and a comprehensive fusogenicity map of many biologically relevant lipid classes.

Negative-sense RNA viruses: An underexplored platform for examining virus–host lipid interactions

Viruses are pathogenic agents that can infect all varieties of organisms, including plants, animals, and humans. These microscopic particles are genetically simple as they encode a limited number of proteins that undertake a wide range of functions. While structurally distinct, viruses often share common characteristics that have evolved to aid in their infectious life cycles. A commonly underappreciated characteristic of many deadly viruses is a lipid envelope that surrounds their protein and genetic contents. Notably, the lipid envelope is formed from the host cell the virus infects. Lipid-enveloped viruses comprise a diverse range of pathogenic viruses, which often lead to high fatality rates and many lack effective therapeutics and/or vaccines. This perspective primarily focuses on the negative-sense RNA viruses from the order Mononegavirales, which obtain their lipid envelope from the host plasma membrane. Specifically, the perspective highlights the common themes of host cell lipid and membrane biology necessary for virus replication, assembly, and budding.

Cholera Toxin as a Probe for Membrane Biology

Cholera toxin B-subunit (CTxB) has emerged as one of the most widely utilized tools in membrane biology and biophysics. CTxB is a homopentameric stable protein that binds tightly to up to five GM1 glycosphingolipids. This provides a robust and tractable model for exploring membrane structure and its dynamics including vesicular trafficking and nanodomain assembly. Here, we review important advances in these fields enabled by use of CTxB and its lipid receptor GM1.

Bitter Taste and Olfactory Receptors: Beyond Chemical Sensing in the Tongue and the Nose

The Up-and-Coming-Scientist section of the current issue of the Journal of Membrane Biology features the invited essay by Dr. Mercedes Alfonso-Prieto, Assistant Professor at the Forschungszentrum Jülich (FZJ), Germany, and the Heinrich-Heine University Düsseldorf, Vogt Institute for Brain Research. Dr. Alfonso-Prieto completed her doctoral degree in chemistry at the Barcelona Science Park, Spain, in 2009, pursued post-doctoral research in computational molecular sciences at Temple University, USA, and then, as a Marie Curie post-doctoral fellow at the University of Barcelona, worked on computations of enzyme reactions and modeling of photoswitchable ligands targeting neuronal receptors. In 2016, she joined the Institute for Advanced Science and the Institute for Computational Biomedicine at the FZJ, where she pursues research on modeling and simulation of chemical senses. The invited essay by Dr. Alfonso-Prieto discusses state-of-the-art modeling of molecular receptors involved in chemical sensing – the senses of taste and smell. These receptors, and computational methods to study them, are the focus of Dr. Alfonso-Prieto’s research. Recently, Dr. Alfonso-Prieto and colleagues have presented a new methodology to predict ligand binding poses for GPCRs, and extensive computations that deciphered the ligand selectivity determinants of bitter taste receptors. These developments inform our current understanding of how taste occurs at the molecular level. Graphic Abstract