COPI coat assembly occurs on liquid-disordered domains and the associated membrane deformations are limited by membrane tension

Abstract

Cytoplasmic coat proteins are required for cargo selection and budding of tubulovesicular transport intermediates that shuttle between intracellular compartments. To better understand the physical parameters governing coat assembly and coat-induced membrane deformation, we have reconstituted the Arf1-dependent assembly of the COPI coat on giant unilamellar vesicles by using fluorescently labeled Arf1 and coatomer. Membrane recruitment of Arf1-GTP occurs exclusively on disordered lipid domains and does not induce optically visible membrane deformation. In the presence of Arf1-GTP, coatomer self-assembles into weakly curved coats on membranes under high tension, while it induces extensive membrane deformation at low membrane tension. These deformations appear to have a composition different from the parental membrane because they are protected from phase transition. These findings suggest that the COPI coat is adapted to liquid disordered membrane domains where it could promote lipid sorting and that its mechanical effects can be tuned by membrane tension.budding ͉ giant unilamellar vesicle ͉ Golgi ͉ intracellular transport ͉ lipid sorting

Regulation of the Golgi complex by phospholipid remodeling enzymes

“…Curved membranes tend to exclude sphingolipids and are enriched in phospholipids. For instance, in vitro reconstitution of COPI budding has shown that phase transition of phospholipid enriched domains is prevented at bud sites [49]. Phospholipid transfer proteins, flippases, PIPs, and, DAG also play important roles in the dynamic organization of the Golgi complex but are beyond the scope of this review [55, 78, 81–87].…”

Single Lipid Molecule Dynamics on Supported Lipid Bilayers with Membrane Curvature

“…Although accumulation of proteins and lipids at curvature is observed in a variety of experiments [23,29,30,31,32,33], single molecules have demonstrably different behavior [33]. So far, single proteins have not been observed accumulating at curvature, suggesting that a cluster of molecules is needed for accumulation or that the affinity for curvature is quite low and hard to observe in single molecule experiments, where the concentration is inherently low.…”

Formation of COPI-coated vesicles at a glance

“…Lipids that form liquidordered phases, such as sphingomyelin and cholesterol, and are abundant at the plasma membrane, are excluded from COPI-coated vesicles (Brügger et al, 2000). In in vitro reconstituted systems, Arf1-GTP and COPI assemble on liquid-disordered domains, and protect them from undergoing phase separation, suggesting that COPI assists lipid sorting during coat polymerization (Manneville et al, 2008).…”