Lipid-Dependent Interaction of Human N-BAR Domain Proteins with Sarcolemma Mono- and Bilayers.

The N-BAR domain of the human Bin1 protein is indispensable for T-tubule biogenesis in skeletal muscles. It binds to lipid mono- and bilayers that mimic the sarcolemma membrane composition, and it transforms vesicles into uniform tubules by generating a decorating protein scaffold. We found that Δ(1-33)BAR, lacking the N-terminal amphipathic helix (H0), and H0 alone bind to sarcolemma monolayers, although both proteins are not able to tubulate sarcolemma vesicles. By variation of the lipid composition, we elucidated the role of PI(4,5)P2, cholesterol, and an asymmetric sarcolemma composition for Bin1-N-BAR binding and sarcolemma tubulation. Our results indicate that Bin1-N-BAR binding is low in the absence of PI(4,5)P2 and it is affected by additional changes in the negative headgroup charge and lipid acyl chain composition. However, it is not dependent on the cholesterol content. The results from Langmuir monolayer experiments are complementary to lipid bilayer studies using electron microscopy that provides information on membrane curvature generation.

Supramolecular organization of the human N-BAR domain in shaping the sarcolemma membrane.

The 30kDa N-BAR domain of the human Bin1 protein is essential for the generation of skeletal muscle T-tubules. By electron cryo-microscopy and electron cryo-tomography with a direct electron detector, we found that Bin1-N-BAR domains assemble into scaffolds of low long-range order that form flexible membrane tubules. The diameter of the tubules closely matches the curved shape of the N-BAR domain, which depends on the composition of the target membrane. These insights are fundamental to our understanding of T-tubule formation and function in human skeletal muscle.