The Biophysics of Living Membranes: Protein Partitioning and Functional Differentiation in Ordered Plasma Membrane Domains

Abstract

The lipid raft hypothesis posits that ordered, lateral membrane domains are important for the regulation of plasma membrane (PM) structure and function in eukaryotes. Giant Plasma Membrane Vesicles (GPMVs) are isolated plasma membranes that microscopically separate into coexisting ordered and disordered phases, facilitating experimental analysis of the composition and physical properties of ordered raft domains in biological membranes. In these studies, we analyze the biophysical and lipidomic differentiation of the plasma membrane in Mesenchymal Stem Cells (MSC) as the cells undergo differentiation into adipocytes and osteoblasts. During differentiation, dramatic remodeling the PM lipidome, including lipid acyl chain length and unsaturation, results inchanges to membrane order and phase separation. These observations elucidate the compositional determinants of biophysical properties in biological membranes, as well as identify lineage-specific PM features. These results facilitate rational remodeling of membrane phenotypes to direct differentiation, as supplementation with ꙍ-3 docosahexaneoic acid (DHA) promoted the osteoblastic PM phenotype and potentiated osteogenic differentiation. The differences in the physical properties of the coexisting domains lead to preferential protein partitioning between them. We evaluate the structural determinants of raft partitioning of a model transmembrane protein, and find that raft phase partitioning is related to features of the protein's transmembrane domain (TMD), namely the hydrophobic length and surface area of the TMD. Longer TMDs impart greater raft association while TMDs with larger, bulkier amino acid side chains prefer the non-raft phase. We present a simple physical model wherein raft partitioning is driven by phase-dependent differences in interfacial energy between the TMD and its surrounding lipid matrix, and find excellent quantitative agreement with observations that provide the first predictions of protein-lipid surface tension. These results point the way to a general rule for raft partitioning of transmembrane proteins.