Membrane lipid order of sub‐synaptic T cell vesicles correlates with their dynamics and function

• Published in: Traffic (Copenhagen, Denmark) Vol. 19; no. 1; pp. 29 - 35
• Main Authors: Ashdown, George W., Williamson, David J., Soh, Gary H.M., Day, Nathan, Burn, Garth L., Owen, Dylan M.
• Format: Journal Article
• Language: English
• Published: Former Munksgaard John Wiley & Sons A/S 01.01.2018; Wiley Subscription Services, Inc
• Subjects: Actin; Actin Cytoskeleton - metabolism; Adaptor Proteins, Signal Transducing - metabolism; Antigen-presenting cells; Cell activation; Cells, Cultured; Cytoplasmic Vesicles - metabolism; Cytoskeleton; Dynamics; Humans; Immunological synapses; Immunological Synapses - chemistry; Immunological Synapses - metabolism; Immunological Synapses - ultrastructure; Immunology; Immunosuppressive agents; Jurkat Cells; Lipids; Live Cell; Lymphocytes T; Membrane Lipids - chemistry; Membrane Lipids - metabolism; Membrane order; Membrane Proteins - metabolism; Synapse; Synaptic vesicles; T cell; T-Lymphocytes - metabolism; T-Lymphocytes - ultrastructure; Tubulin; Vesicle; Vesicle; Synapse; Dynamics; T cell; Cytoskeleton; Membrane order; Live Cell
• ISSN: 1398-9219; 1600-0854
• DOI: 10.1111/tra.12532

During an immune response, T cells survey antigen presenting cells for antigenic peptides via the formation of an interface known as an immunological synapse. Among the complex and dynamic biophysical phenomena occurring at this interface is the trafficking of sub‐synaptic vesicles carrying a variety of proximal signalling molecules. Here, we show that rather than being a homogeneous population, these vesicles display a diversity of membrane lipid order profiles, as measured using the environmentally sensitive dye di‐4‐ANEPPDHQ and multi‐spectral TIRF microscopy. Using live‐cell imaging, vesicle tracking and a variety of small molecule drugs to manipulate components of the actin and tubulin cytoskeleton, we show that the membrane lipid order of these vesicles correlate with their dynamics. Furthermore, we show that the key proximal signalling molecule Linker for Activation of T cells (LAT) is enriched in specific vesicle populations as defined by their higher membrane order. These results imply that vesicle lipid order may represent a novel regulatory mechanism for the sorting and trafficking of signalling molecules at the immunological synapse, and, potentially, other cellular structures. Proteins segregate based on their environment, including lipid content of plasma membranes, which display order heterogeneity. Here we show sub‐synaptic vesicles in T cells forming an immunological synapse also exhibit lipid heterogeneity. Their composition correlates with their cargo (demonstrated with Linker for Activation of T cells (LAT) and myelin and lymphocyte (MAL)), potentially demonstrating a lipid‐based sorting mechanism. Finally, vesicle dynamics are shown to be modulated by the cytoskeleton adjacent to the synapse, with vesicle order favouring different cytoskeletal populations.