Membrane Criticality Enriches Components at Sites of Adhesion

Abstract

Giant plasma membrane vesicles isolated from living cells undergo large and dynamic critical fluctuations near room temperature. These results have led us to hypothesize that fluctuations persist to growth temperatures of 37°C. In this study, we demonstrate that criticality can greatly impact the organization of components at sites of adhesion between vesicles and a planar supported membrane. Adhesion is accomplished through biotin- streptavidin binding by either directly incorporating trace quantities of biotin-tagged lipids, or indirectly using a biotin conjugated Cholera Toxin B subunit that binds to the ganglioside GM1 incorporated into vesicles. Consistent with previous studies, we observe finite adhesion domains rich in the adhesion molecule (streptavidin) when a PEG lipid is incorporated into vesicles. When vesicle membranes contain coexisting liquid phases, the presence of the adhesion domain affects the localization of phase separated domains based on the partitioning of the adhesion molecules. For example, we observe enrichment of the disordered phase marker DiIC12 at the adhesion domain when the adhesion molecule is a lipid that prefers liquid-disordered phase regions. When vesicles have critical membrane compositions, DiI remains enriched at sites of adhesion at temperatures >15°C above the transition temperature. This occurs even though DiI is not directly involved in adhesion. Instead, DiI partitioning occurs likely because adhesion molecules gather fluctuations that are rich in the lipid probe. At these same temperatures, free-floating vesicles appear uniform on the micron-scale. We also observe DiI enrichment or depletion at the adhesion domain when giant plasma membrane vesicles are adhered to a supported bilayer at 37°C. This occurs even though measured critical temperatures are close to room temperature. Our results demonstrate that membrane criticality can significantly alter the organization of membrane components even when critical temperatures are well below growth temperatures.