@article{BoertleinDraegerSchoenaueretal.2018,
  author    = {B{\"o}rtlein, Charlene and Draeger, Annette and Schoenauer, Roman and Kuhlemann, Alexander and Sauer, Markus and Schneider-Schaulies, Sybille and Avota, Elita},
  title     = {The neutral sphingomyelinase 2 is required to polarize and sustain T Cell receptor signaling},
  series = {Frontiers in Immunology},
  volume    = {9},
  journal   = {Frontiers in Immunology},
  number    = {815},
  doi       = {10.3389/fimmu.2018.00815},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-176572},
  year      = {2018},
  abstract  = {By promoting ceramide release at the cytosolic membrane leaflet, the neutral sphingomyelinase 2 (NSM) is capable of organizing receptor and signalosome segregation. Its role in T cell receptor (TCR) signaling remained so far unknown. We now show that TCR-driven NSM activation is dispensable for TCR clustering and initial phosphorylation, but of crucial importance for further signal amplification. In particular, at low doses of TCR stimulatory antibodies, NSM is required for Ca\(^{2+}\) mobilization and T cell proliferation. NSM-deficient T cells lack sustained CD3ζ and ZAP-70 phosphorylation and are unable to polarize and stabilize their microtubular system. We identified PKCζ as the key NSM downstream effector in this second wave of TCR signaling supporting dynamics of microtubule-organizing center (MTOC). Ceramide supplementation rescued PKCζ membrane recruitment and MTOC translocation in NSM-deficient cells. These findings identify the NSM as essential in TCR signaling when dynamic cytoskeletal reorganization promotes continued lateral and vertical supply of TCR signaling components: CD3ζ, Zap70, and PKCζ, and functional immune synapses are organized and stabilized via MTOC polarization.},
  language  = {en}
}
@article{KuhlemannBeliuJanzenetal.2021,
  author    = {Kuhlemann, Alexander and Beliu, Gerti and Janzen, Dieter and Petrini, Enrica Maria and Taban, Danush and Helmerich, Dominic A. and Doose, S{\"o}ren and Bruno, Martina and Barberis, Andrea and Villmann, Carmen and Sauer, Markus and Werner, Christian},
  title     = {Genetic Code Expansion and Click-Chemistry Labeling to Visualize GABA-A Receptors by Super-Resolution Microscopy},
  series = {Frontiers in Synaptic Neuroscience},
  volume    = {13},
  journal   = {Frontiers in Synaptic Neuroscience},
  issn      = {1663-3563},
  doi       = {10.3389/fnsyn.2021.727406},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-251035},
  year      = {2021},
  abstract  = {Fluorescence labeling of difficult to access protein sites, e.g., in confined compartments, requires small fluorescent labels that can be covalently tethered at well-defined positions with high efficiency. Here, we report site-specific labeling of the extracellular domain of γ-aminobutyric acid type A (GABA-A) receptor subunits by genetic code expansion (GCE) with unnatural amino acids (ncAA) combined with bioorthogonal click-chemistry labeling with tetrazine dyes in HEK-293-T cells and primary cultured neurons. After optimization of GABA-A receptor expression and labeling efficiency, most effective variants were selected for super-resolution microscopy and functionality testing by whole-cell patch clamp. Our results show that GCE with ncAA and bioorthogonal click labeling with small tetrazine dyes represents a versatile method for highly efficient site-specific fluorescence labeling of proteins in a crowded environment, e.g., extracellular protein domains in confined compartments such as the synaptic cleft.},
  language  = {en}
}