Biology subjects

Lira, R. B.

Publications and source records attributed to Lira, R. B..

8 recordsLinked to original sources

bioRxiv2026

Charge and steric hindrance in the glycocalyx govern membrane interactions

The outer leaflet of the plasma membrane - the cells interface with exogenous biomolecules and particles - is largely electrostatically neutral and highly viscous, conditions that are generally unfavorable for fusion of external membrane-bound objects such as liposomes. Yet living cells efficiently interact and fuse with charged liposomes, revealing a discrepancy between synthetic and biological membranes. Here, we identify the glycocalyx, the carbohydrate-rich layer that coats cells, as the component that explains this mismatch. Using synthetic membranes reconstituted with an artificial glycocalyx, we resolve how electrostatics and steric hindrance regulate the interactions of cationic liposomes with synthetic membranes. Charged glycolipids are found to be sufficient to convert non-fusogenic neutral membranes into fusion-competent membranes. In contrast, large polymers in the glycocalyx suppress fusion. Steric hindrance acts primarily at the docking stage, while fusion proceeds through a long-lived hemifusion intermediate. Strikingly, these large differences occur without measurable changes in membrane viscosity or phase state, demonstrating that glycocalyx-mediated control is independent of the membrane mechanics. Selected glycocalyx components can also promote endocytic budding and cargo internalization. We thus show that the glycocalyx is a tunable physicochemical regulator of liposome docking, fusion, and internalization. The findings provide a framework for understanding how biological agents and therapeutics engage cell membranes and they offer key design principles for synthetic cells.

biophysics↗

bioRxiv2026

MEMBRANE PORES ACT AS SELF-RESEALING LIPID SCRAMBLASES

Biological membranes continuously experience leaflet lipid imbalances during growth, lipid synthesis, and vesicle fusion. To alleviate such imbalances and prevent these asymmetries from compromising membrane integrity, cells rely on lipid scramblases - fast and non-specific lipid channel proteins. Here we show that lipid number asymmetry alone is sufficient to drive spontaneous formation of transient hydrophilic pores that function as self-resealing lipid scramblases. Using giant unilamellar vesicles, living cells, and coarse-grained molecular dynamics simulations, we demonstrate that fusion-induced excess lipids in one leaflet lowers membrane edge tension, generating size-selective pores whose size and lifetime scale with the magnitude of asymmetry. Below a critical threshold, these pores reseal spontaneously; above it, membranes collapse. Strikingly, pore opening enables rapid, non-selective lipid translocation between leaflets, dissipating the asymmetry that nucleates the pore and thereby promoting their own closure. Cholesterol buffers moderate imbalances through spontaneous flip flop before pore formation, whereas pore-mediated lipid scrambling relieves the remaining asymmetry and restores cholesterols initial distribution. Our findings identify transient lipid pores as an intrinsic, protein-independent mechanism that couples membrane destabilization to self-repair, providing a universal physical principle for membrane homeostasis during growth, remodelling, and early cellular evolution.

biophysics↗

bioRxiv2025

The Hidden Cost of Fusion: Intrinsic Asymmetry in Vesicle Fusion Restricts Synthetic Cell Growth

Membrane fusion is essential for signaling, cargo delivery, and synthetic cell growth, yet its mechanical consequences remain poorly defined. How fusion-driven membrane growth can be sustained without compromising compartment stability remains an unresolved challenge. Here, we established a minimal reconstituted system where content-loaded small liposomes fuse with single cell-sized giant unilamellar vesicles (GUVs), combining micropipette delivery, electrodeformation, and live imaging. Fusion outcomes were quantified through lipid and content mixing assays, GUV electrodeformation to track area and tension, and phase contrast imaging to monitor leakage. GUVs incorporated lipids and cargo from hundreds of thousands of vesicles at unprecedented efficiency rates, enabling substantial growth. However, accumulation of leaflet asymmetries induced curvature and tension, driving budding, rupture and leakage. Hemifusion amplified these destabilizing effects. Lipid number asymmetries emerge as a dominant mechanical cost of fusion, highlighting how cells may regulate these processes and guiding the design of therapeutic delivery systems and synthetic cells capable of robust and stable growth. Significance StatementMembrane fusion enables essential biological processes from secretion to cell growth. Using a synthetic system, we show that rapid fusion of small vesicles with model cell-like compartments drives dramatic growth but also introduces leaflet asymmetries that build curvature and tension, limit expansion and compromise membrane stability. By uncovering lipid number asymmetry as a key mechanical cost of fusion, our study explains why cells tightly regulate fusion events and provides principles for designing delivery vehicles and synthetic cells capable of robust, sustainable growth.

biophysics↗

bioRxiv2025

Influence of Substrate on Supported Lipid Bilayers: Membrane Pinning, Stretching, Pores, and Remodelling

Model membranes offer a simplified representation of biomembranes, allowing for controlled structural and functional studies without the inherent complexity of biological systems. Due to the confined space and strong interactions with the support, supported lipid bilayers (SLBs) exhibit markedly distinct properties compared to those of their free-standing counterparts. However, the nano and microscale origins of these differences, structural effects and the related biophysical consequences remain incompletely understood. Using a combination of spectroscopy and micromanipulation techniques, we reveal that SLBs are pinned to the support, creating barriers for lipid diffusion. Furthermore, they are intrinsically stretched and under tension, causing membrane rupture and the formation of stable [~]2 nm aqueous pores that allow the permeation of small hydrophilic molecules. Despite strong adhesion, SLBs can undergo localized shape changes associated with membrane fusion, and membrane nanotubes can be extruded under optical forces. The findings underscore the very distinct properties of model membranes, and we anticipate that they will have important consequences in the field of drug-delivery systems and membrane biophysics.

biophysics↗

bioRxiv2025

Lipid flip flop regulates the shape of growing and dividing synthetic cells

Cells grow their boundaries by incorporating newly synthesized lipids into their membranes as well as through fusion of intracellular vesicles. As these processes yield trans-bilayer imbalances in lipid numbers, cells must actively flip lipid molecules across the bilayer to enable growth. Using giant and small unilamellar vesicles (GUVs and SUVs, respectively), we here recapitulate cellular growth and division under various conditions of transmembrane flip flop of lipids. By dynamically monitoring the changes in reduced volume and spontaneous curvature of GUVs that grow by fusion of many small SUVs, the morphology of these growing synthetic cells is quantified. We demonstrate that lipid flip flop relaxes curvature stresses and yields more symmetrically sized buds. Further increasing the neck curvature is shown to lead to bud scission. The mechanisms presented here offer fundamental insights into cell growth and division, which are important for understanding early protocells and designing synthetic cells that are able to grow and divide.

synthetic biology↗

bioRxiv2024

Membrane fusion-based drug delivery liposomes transiently modify the material properties of synthetic and biological membranes

Many drug targets are located in intracellular compartments of cells but they often remain inaccessible to standard imaging and therapeutic agents. To aid intracellular delivery, drug carrier nanoparticles have been used to overcome the barrier imposed by the plasma membrane. The carrier must entrap large amounts of cargo, efficiently and quickly deliver the cargo in the cytosol or other intracellular compartments, and must be as inert as possible. In other words, they should not induce cellular responses or alter the cell state in the course of delivery. Here, we show that cationic liposomes with high charge density efficiently fuse with synthetic membranes and the plasma membrane of living cells. Direct fusion efficiently delivers large amounts of cargo to cells and cell-like vesicles within seconds, bypassing slow and often inefficient internalization-based pathways. These effects depend on liposome charge density and, to some extent, liposome concentration and the helper lipid. However, fusion-mediated cargo delivery results in the incorporation of large amounts of foreign lipids that leads to changes in the material properties of these membranes, namely modifications in membrane packing and fluidity, induction of membrane curvature, decrease in surface tension and the formation of (short-lived) pores. Importantly, these effects are transient and liposome removal allows cells to recover their state prior to liposome interaction.

biophysics↗

bioRxiv2022

GM1 asymmetry in the membrane stabilizes pores

Cell membranes are highly asymmetric and their stability against poration is crucial for survival. We investigated the influence of membrane asymmetry on electroporation of giant unilamellar vesicles with membranes doped with GM1, a ganglioside asymmetrically enriched in the outer leaflet of neuronal cell membranes. Compared to symmetric membranes, the lifetimes of micronsized pores are about an order of magnitude longer suggesting that pores are stabilized by GM1. Internal membrane nanotubes caused by the GM1 asymmetry, obstruct and additionally slow down pore closure, effectively reducing pore edge tension and leading to leaky membranes. Our results point to the drastic effects this ganglioside can have on pore resealing in biotechnology applications based on poration as well as on membrane repair processes. SIGNIFICANCEMembrane pore closure is crucial for cell survival and is important for biotechnological and medicine applications based on transfer of material, e.g. drugs, genes, through pores. Electroporation is widely used as means to perforate the membrane but factors governing membrane resealing are still a matter of debate, in particular the large variations of pore lifetimes. Here, we probed the effect of bilayer asymmetry on pore dynamics employing cell-sized giant unilamellar vesicles doped with the ganglioside GM1 (asymmetrically enriched in neurons). We find that the presence of GM1 and its asymmetric distribution in the bilayer dramatically slows down pore resealing, not only by mere molecular stabilization of the pore rim, but also by generating membrane nanotubes.

biophysics↗

bioRxiv2020

To close or to collapse: the role of charges on membrane stability upon pore formation

Resealing of membrane pores is crucial for cell survival. We study membrane surface charge and medium composition as defining regulators of membrane stability. Pores are generated by electric field or detergents. Giant vesicles composed of zwitterionic and negatively charged lipids mixed at varying ratios are subjected to a single strong electric pulse. Charged vesicles are prone to catastrophic collapse transforming them into tubular structures. The spectrum of destabilization responses includes the generation of long-living submicroscopic pores and partial vesicle bursting. The origin of these phenomena is related to the membrane edge tension, which governs pore closure. This edge tension significantly decreases as a function of the fraction of charged lipids. Destabilization of charged vesicles upon pore formation is universal - it is also observed with other poration stimuli. Disruption propensity is enhanced for membranes made of lipids with higher degree of unsaturation. It can be reversed by screening membrane charge in the presence of calcium ions. We interpret the observed findings in light of theories of stability and curvature generation and discuss mechanisms acting in cells to prevent total membrane collapse upon poration. Enhanced membrane stability is crucial for the success of electroporation-based technologies for cancer treatment and gene transfer.

biophysics↗