The B Cell Receptor Dictates its Local Lipid Environment

Abstract

The B cell receptor (BCR) is responsible for sensing and responding to intact antigen during the immune response and is hypothesized to nucleate a unique composition of lipids surrounding receptor clusters during antigen binding and signaling. This unique lipid composition, sometimes referred to as a “lipid raft,” is predicted to be enriched in glycosphingolipids and cholesterol. An altered lipid composition surrounding the BCR could influence the partitioning and activity of regulatory proteins and lipids, acting as a mechanism for the B cell to regulate signal transduction following stimulus by antigen. However, without direct observations of this phenomenon, theories surrounding lipid compositional heterogeneity in B cells remain controversial. In this work, we utilize two-color super-resolution localization microscopy (STORM and PALM) to directly observe the lipid composition proximal to B cell receptors, using the photoactivatable fluorescent protein mEos3.2 anchored to the plasma membrane through posttranslational lipid modifications. These probes mimic important regulatory proteins involved in BCR signaling by their membrane anchor but lack the protein interaction domains or biological activity found in the native protein. By analyzing the co-distributions of the B cell receptor and lipid probes using correlation functions, we find that the membrane surrounding B cell receptor clusters is depleted of probes bearing unsaturated and branched geranylgeranyl modifications and is enriched in probes bearing saturated palmitoyl modifications. Quantification of lipid probe distributions reveals that B cell compositional heterogeneity influences the partitioning of these lipid modifications with order 1kBT of effective potential, as expected from predications of composition fluctuations in critical systems. These findings provide definitive evidence of membrane compositional heterogeneity in an important biological signaling system. Our results suggest that compositional fluctuations contribute to cellular responses by influencing the spatial distribution of specific components in the plasma membrane.