BPS2026 – Synergy of acid and oxidative stress on membrane permeability explained by a pH-dependent conformational switch of oxidized phospholipids
(2026)
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- (en) Energetic barriers and intermediates in membrane fission and fusion are typically highly curved. Building a model of these highly curved bilayer regions requires understanding how local lipid composition and interactions adapt to stabilize such a structure. To extract this information, molecular simulations can apply special highly curved, frequently non-lamellar structures that model high-energy sites in intermediates in membrane reshaping. The lipid inverse hexagonal phase, available to both simulation and experiment, allows direct observation of a single highly curved lipidic leaflet. With appropriate boundary conditions (equivalent to external forces), single-bilayer pores and edges can be modeled at equilibrium. Microsecond-scale simulations of toroidal fusion/ fission pores and hemifusion diaphragms allow for observations of how modeled lipids redistribute and change their interactions on structures directly modeling reshaping intermediates. Here, mechanical analysis, dynamic lipid redistribution, and alchemical free-energy perturbation methodology will be applied to extract the model energetics of simulated lipids on these highly curved structures. 2000-Symp BPS2026-pH-dependent mesophase transitions as key determinants of lipid nanoparticle function Lipid nanoparticles (LNPs) are the leading platform for nucleic acid delivery and mRNA vaccination, yet the structural determinants of their activity remain incompletely understood. Here, we show that cationic ionizable lipids (CILs) undergo pH-driven structural transitions-from inverse cubic (Fd3m) to inverse hexagonal (HII) phases-in model systems, and that these transitions correlate with endosomal fusion and the timing of gene delivery. Using high-resolution small-angle X-ray scattering and live-cell imaging on single-cell arrays (LISCA), we directly link these phase changes to the onset and efficiency of mRNA expression. We present an integrated approach combining small-angle X-ray scattering (SAXS) experiments, molecular dynamics (MD) simulations , and a continuum model to elucidate lipid distribution and water content within HII phases. The different approaches consistently yield water contents that seem to correlate with the lipids' transfection efficiencies. Together, our findings identify structural transitions as a key mechanism in LNP-mediated delivery that offer a powerful approach for rational design and optimization of lipid nanoparticles. 2001-SympSelect BPS2026-Synergy of acid and oxidative stress on membrane permeability explained by a pH-dependent conformational switch of oxidized phospholipids The plasma membrane constitutes a selective diffusion barrier that permits effective cell function and communication. During bacterial infections, a defensive measure of the immune system is to phagocytose bacteria and expose them to a combination of acid and oxidative stress within phagolyso-somes. This combination of stresses usually leads to the impairment of the membrane barrier function in bacteria, but the precise mechanisms are not well known. We therefore investigated the effects of acid and oxidative stress on bacterial membranes and membrane models to unravel the precise molecular mechanisms underlying this activity. In Gram-positive bacteria, we observed that acid stress led to increased potassium release and a depleted membrane potential, while oxidative stress increased permeability toward larger molecules. Both effects synergistically inhibited bacterial growth. In Gram-negative bacteria, both stresses acted synergistically on the outer membrane, which correlated with potassium release and growth inhibition , although no large pores were formed in the plasma membrane. To delve into the molecular mechanism behind these synergistic effects, we investigated how acidity might alter membrane permeability induced by oxidized phospholipids in liposomes. We observed a pH-dependent perme-ability change in the presence of oxidized phospholipids that possessed truncated acyl chains with terminal carboxyl groups. This behavior resulted from a pH-dependent conformational switch of the carboxylated acyl chain. At cytosolic pH, the carboxyl was ionized, promoting its presence at the membrane interface, while at low phagolysosomal pH, ionization was reduced, promoting its localization to the membrane interior, which likely led to a compromised hydrophobic barrier function. This mechanism might partly explain the synergy of acid and oxidative stress on oxidized membranes , while truncated carboxylated phospholipids could also be promising candidates for the formulation of pH-dependent liposomal drug-delivery systems. 2002-Symp BPS2026-Engineering structural complexity in lipid nanoparticles to enhance their fusogenicity Lipid nanoparticles (LNPs) are the most successful delivery carriers of biologics to cells and tissues. Expanding LNP-based therapies hinges on efficient cell entry and delivery, a process often hindered by endosomal entrapment. It is well known that depending on molecular properties, lipids self-assemble into different nanostructures. LNPs can recapitulate some of these mesophases dispersed in solution, but how structure imparts interaction with cells and endosomal escape remains unclear. We demonstrate that combining lipid composition with nanostructure synergistically impacts the ability of LNPs to escape endosomes via enhanced fusogenicity. Specifically, LNPs prescribed with bicontinuous cubic and inverse hexagonal internal structures facilitate the topological transition of LNP-endosome fusion-pore formation. In this talk, we show our recent advances in experimental and theoretical approaches that enable the quantification and prediction of lipid fusogenicity. We show that not only spontaneous curvature but also Gaussian modulus plays a critical role in regulating LNP fusogenicity. DNA-points accumulation for imaging in nanoscale topography (DNA-PAINT) has emerged as a powerful super-resolution imaging technique capable of visualizing biological and synthetic structures with nanometer precision. By exploiting the transient binding of short fluorescently labeled DNA ''imager'' strands to complementary ''docking'' strands attached to target molecules, DNA-PAINT enables multiplexed visualization of targets with molecular resolution. We expand the existing repertoire of speed-optimized DNA sequences for DNA-PAINT imaging to drive visualization of up to ten different targets in a sequential manner with molecular resolution. Our speed-optimized imager-docking strand pairs minimize crosstalk and maximize hybridization rates, enabling rapid image acquisition without compromising spatial resolution. We demonstrate this ability by visualizing diverse nuclear targets in single cells and subsequently characterizing their relative spatial distribution. We further outline the ability of the technique to visualize and detect nanoscale reorganization in the chromatin landscape upon transcription inhibition. The ability to image as many as ten targets using this strategy at accelerated speed will drive its further adoption for diverse cellular imaging applications. 2004-Plat BPS2026-Image-based analysis of the genome's fractality during the cell cycle
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Xie, M., Koch, E. H. W., Derks, M. G. N., Bagheri, B., Kovryzhenko, D., Xu, Z., Sobota, A., Karttunen, M., Weingarth, M., Killian, J. A., Breukink, E. J., Sonnen, A. F. P., Lorent, J., & et al. (2026, February). BPS2026 – Synergy of acid and oxidative stress on membrane permeability explained by a pH-dependent conformational switch of oxidized phospholipids. https://doi.org/10.1016/j.bpj.2025.11.2219