Eukaryotic cells possess a dynamic network of membranes that vary in lipid composition. To perform numerous biological functions, cells modulate their shape and the lateral organization of proteins associated with membranes. The modulation is generally facilitated by physical cues that recruit proteins to specific regions of the membrane. Analyzing these cues is difficult due to the complexity of the membrane conformations that exist in cells. Here, we examine how different types of membrane proteins respond to changes in curvature and to lipid phases found in the plasma membrane. By using giant plasma membrane vesicles derived from transfected cells, the proteins were positioned in the correct orientation and the analysis was performed in plasma membranes with a biological composition. Nanoscale membrane curvatures were generated by extracting nanotubes from these vesicles with an optical trap. The viral membrane protein neuraminidase was not sensitive to curvature, but it did exhibit strong partitioning (coefficient of K = 0.16) disordered membrane regions. In contrast, the membrane repair protein annexin 5 showed a preference for nanotubes with a density up to 10–15 times higher than that on the more flat vesicle membrane. The investigation of nanoscale effects in isolated plasma membranes provides a quantitative platform for studying peripheral and integral membrane proteins in their natural environment.

The plasma membrane of eukaryotic cells consists of a crowded environment comprised of a high diversity of proteins in a complex lipid matrix. The lateral organization of membrane proteins in the plasma membrane (PM) is closely correlated with biological functions such as endocytosis, membrane budding and other processes which involve protein mediated shaping of the membrane into highly curved structures. Annexin A4 (ANXA4) is a prominent player in a number of biological functions including plasma membrane repair. Its binding to membranes is activated by Ca 2+ influx and it is therefore rapidly recruited to the cell surface near rupture sites where Ca 2+ influx takes place. However, the free edges near rupture sites can easily bend into complex curvatures and hence may accelerate recruitment of curvature sensing proteins to facilitate rapid membrane repair. To analyze the curvature sensing behavior of curvature inducing proteins in crowded membranes, we quantifify the affinity of ANXA4 monomers and trimers for high membrane curvatures by extracting membrane nanotubes from giant plasma membrane vesicles (GPMVs). ANXA4 is found to be a sensor of negative membrane curvatures. Multiscale simulations furthermore predicted that ANXA4 trimers generate membrane curvature upon binding and have an affinity for highly curved membrane regions only within a well defined membrane curvature window. Our results indicate that curvature sensing and mobility of ANXA4 depend on the trimer structure of ANXA4 which could provide new biophysical insight into the role of ANXA4 in membrane repair and other biological processes.

The plasma membrane surrounds every single cell and essentially shapes cell life by separating the interior from the external environment. Thus, maintenance of cell membrane integrity is essential to prevent death caused by disruption of the plasma membrane. To counteract plasma membrane injuries, eukaryotic cells have developed efficient repair tools that depend on Ca 2+ -and phospholipid-binding annexin proteins. Upon membrane damage, annexin family members are activated by a Ca 2+ influx, enabling them to quickly bind at the damaged membrane and facilitate wound healing. Our recent studies, based on interdisciplinary research synergy across molecular cell biology, experimental membrane physics, and computational simulations show that annexins have additional biophysical functions in the repair response besides enabling membrane fusion. Annexins possess different membrane-shaping properties, allowing for a tailored response that involves rapid bending, constriction, and fusion of membrane edges for resealing. Moreover, some annexins have high affinity for highly curved membranes that appear at free edges near rupture sites, a property that might accelerate their recruitment for rapid repair. Here, we discuss the mechanisms of annexin-mediated membrane shaping and curvature sensing in the light of our interdisciplinary approach to study plasma membrane repair.

The plasma membrane of eukaryotic cells consists of a crowded environment comprised of a high diversity of proteins in a complex lipid matrix. The lateral organization of membrane proteins in...

Living cells respond to nanoscopic thermoplasmonic injury by recruiting an annular ring of annexin V.