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.