“Membrane Sensing and Remodelling during the immune synapse. A spatiotemporal adaptive tale of lipid packing dynamics and collective assembly resolved with Super-resolution quantitative Imaging”

Lymphocyte T cells are responsible for cell-mediated adaptive immune responses, involving transient interactions of the T-cell receptor (TCR) with peptides presented by MHC proteins. A productive interaction triggers the T-cell signaling forming the immunological synapse (IS). Initially, Lck, a membrane-anchored tyrosine kinase, phosphorylates the TCR, ultimately producing basal plasma membrane microclusters and calcium release. For this purpose, the preferred imaging method to unravel nanoclustering at surface-membrane close contact zones during in-vitro T cell activation is TIRF-based super-resolution imaging. Classically, clustering characterisation of the resting (non-activated) state relied on observing T cell onto surfaces coated with Poly-L-lysine; whereas T cells, onto functionalised surfaces coated with anti-CD3, and -CD28 antibodies, resembled the T cell activation during the IS. In the last years, some super-resolution studies suggested protein nanoclustering already at resting state, which contradicts the consensus picture of protein aggregation upon activation. Recently, results from T cells in controlled suspension (immersed in a hydrogel gradient) and later on following its activation highlighted that unnatural cell membrane interactions might hinder our understanding of early T cell activation and the subsequent IS. Whereas these studies focused in the protein clustering and its relation to cortical actin dynamics, they overlooked at the role of the lipids membranes in the early activation and during the IS. Typically, protein clustering is assumed to be coordinated with a higher lateral lipid packing at the membranes. We aim at gaining some knowledge on how lipids would behave in these highly dense localised protein aggregation conditions. For this purpose, to better understand how T cell membranes sense and remodel during the immune synapse, we employ fluorescence imaging correlation spectroscopy based methods and reveal the membrane lipid spatiotemporal localisation, diffusion, and collective motion at the plasma membrane.