Lena Collenburg, Niklas Beyersdorf, Teresa Wiese, Christoph Arenz, Essa M. Saied, Katrin Anne Becker-Flegler, Sibylle Schneider-Schaulies, Elita Avota

• Breakdown of sphingomyelin as catalyzed by the activity of sphingomyelinases profoundly affects biophysical properties of cellular membranes which is particularly important with regard to compartmentalization of surface receptors and their signaling relay. As it is activated both upon TCR ligation and co-stimulation in a spatiotemporally controlled manner, the neutral sphingomyelinase (NSM) has proven to be important in T cell activation, where it appears to play a particularly important role in cytoskeletal reorganization and cellBreakdown of sphingomyelin as catalyzed by the activity of sphingomyelinases profoundly affects biophysical properties of cellular membranes which is particularly important with regard to compartmentalization of surface receptors and their signaling relay. As it is activated both upon TCR ligation and co-stimulation in a spatiotemporally controlled manner, the neutral sphingomyelinase (NSM) has proven to be important in T cell activation, where it appears to play a particularly important role in cytoskeletal reorganization and cell polarization. Because these are important parameters in directional T cell migration and motility in tissues, we analyzed the role of the NSM in these processes. Pharmacological inhibition of NSM interfered with early lymph node homing of T cells in vivo indicating that the enzyme impacts on endothelial adhesion, transendothelial migration, sensing of chemokine gradients or, at a cellular level, acquisition of a polarized phenotype. NSM inhibition reduced adhesion of T cells to TNF-α/IFN-γ activated, but not resting endothelial cells, most likely via inhibiting high-affinity LFA-1 clustering. NSM activity proved to be highly important in directional T cell motility in response to SDF1-α, indicating that their ability to sense and translate chemokine gradients might be NSM dependent. In fact, pharmacological or genetic NSM ablation interfered with T cell polarization both at an overall morphological level and redistribution of CXCR4 and pERM proteins on endothelial cells or fibronectin, as well as with F-actin polymerization in response to SDF1-α stimulation, indicating that efficient directional perception and signaling relay depend on NSM activity. Altogether, these data support a central role of the NSM in T cell recruitment and migration both under homeostatic and inflamed conditions by regulating polarized redistribution of receptors and their coupling to the cytoskeleton.…