Membrane phase separation drives organization at B cell receptor clusters

Abstract

Isolated plasma membranes separate into two coexisting liquid phases with distinct lipid and protein compositions but live cell plasma membranes do not macroscopically phase separate, leading to questions of whether and how the membrane phase transition contributes to functional heterogeneity in cells. Using quantitative super resolution microscopy we show that B cell receptor signaling platforms are nanoscale domains that quantitatively enrich membrane probes based on probe phase partitioning in isolated plasma membrane vesicles. Phase partitioning in vesicles also predicts relative probe mobility and retention at receptor clusters. The convergence between measurements in live cells and isolated membranes establishes a clear role for the membrane phase transition as an organizing principle in cells. We propose that physical properties fundamental to the membrane phase transition give rise to a plasma membrane that is a highly responsive medium, capable of compartmentalizing cellular processes in response to diverse stimuli.

Membrane Cargo Density-Dependent Interaction between Protein and Lipid Domains on the Giant Unilamellar Vesicles

“…Lipid bilayer at different compositions shows various phase behaviors, such as gel-fluidic phase and miscibility transitions. , Fluidic domain separation is of particular importance when it comes to understanding the membrane organization of the plasma membrane. , Ternary mixture model lipid bilayers with unsaturated acyl chain lipids, saturated acyl chain lipids, and cholesterol can form distinct fluidic binary domains, namely, liquid ordered (lo) and liquid disordered (ld) domains. , Giant plasma membrane vesicles directly detached from the living cell plasma membranes were reported to exhibit a similar domain separation behavior. − However, in living cells, such domain separation exists at a much smaller scale and is more dynamic − with some notable exception cases. , The physiological lipid domains are often referred to as lipid rafts , that are involved in various cellular activities. − As different molecules will have different partitioning behaviors in the raft domains, − lipid rafts can play the functional role as a molecular sorting platform. More recently, another biological domain formation that has become an important topic is liquid–liquid phase separation (LLPS) of proteins.…”

Low membrane fluidity triggers lipid phase separation and protein segregation in vivo

“…This principal mechanism of lipid domain formation is best studied in the context of lipid rafts (Lingwood & Simons, 2010; Shaw et al , 2021). Here the co-existence of fluid liquid-disordered and liquid-ordered phases, and the associated protein segregation, has been demonstrated in membranes of living eukaryotic cells (Toulmay & Prinz, 2013; Shelby et al , 2021). In contrast, comprehensive in vivo studies on gel-fluid lipid phase separation in live cells have been challenging due to the tendency of cholesterol/hopanoids to suppress gel-fluid phase transitions (Heberle & Feigenson, 2011), and due to the difficulty to modify the membrane fatty acid composition and, thus, fluidity without inducing lipotoxicity (Shen et al , 2017; Budin et al , 2018).…”

Measuring the co-localization and dynamics of mobile proteins in live cells undergoing signaling responses

“…One advantage of this approach is that it provides a means to exclude systematic contributions from membrane topography that are present over the entire measurement but are not removed by the ROI. We found this to be important when quantifying small signals [45].…”

“…Single and multi-color SMLM has been used to image components of the immune response in a wide variety of contexts [12][13][14][15][16][17][18][19][20][21]. Our own work has largely focused on probing early events in B cell receptor signaling [22][23][24].…”