@article{CollenburgWalterBurgertetal.2016,
  author    = {Collenburg, Lena and Walter, Tim and Burgert, Anne and Mueller, Nora and Seibel, Juergen and Japtok, Lukasz and Kleuser, Burkhard and Sauer, Markus and Schneider-Schaulies, Sibylle},
  title     = {A Functionalized Sphingolipid Analogue for Studying Redistribution during Activation in Living T Cells},
  series = {The journal of immunology},
  volume    = {196},
  journal   = {The journal of immunology},
  publisher = {American Assoc. of Immunologists},
  address   = {Bethesda},
  issn      = {0022-1767},
  doi       = {10.4049/jimmunol.1502447},
  pages     = {3951 -- 3962},
  year      = {2016},
  abstract  = {Sphingolipids are major components of the plasma membrane. In particular, ceramide serves as an essential building hub for complex sphingolipids, but also as an organizer of membrane domains segregating receptors and signalosomes. Sphingomyelin breakdown as a result of sphingomyelinase activation after ligation of a variety of receptors is the predominant source of ceramides released at the plasma membrane. This especially applies to T lymphocytes where formation of ceramide-enriched membrane microdomains modulates TCR signaling. Because ceramide release and redistribution occur very rapidly in response to receptor ligation, novel tools to further study these processes in living T cells are urgently needed. To meet this demand, we synthesized nontoxic, azido-functionalized ceramides allowing for bio-orthogonal click-reactions to fluorescently label incorporated ceramides, and thus investigate formation of ceramide-enriched domains. Azido-functionalized C-6-ceramides were incorporated into and localized within plasma membrane microdomains and proximal vesicles in T cells. They segregated into clusters after TCR, and especially CD28 ligation, indicating efficient sorting into plasma membrane domains associated with T cell activation; this was abolished upon sphingomyelinase inhibition. Importantly, T cell activation was not abrogated upon incorporation of the compound, which was efficiently excluded from the immune synapse center as has previously been seen in Ab-based studies using fixed cells. Therefore, the functionalized ceramides are novel, highly potent tools to study the subcellular redistribution of ceramides in the course of T cell activation. Moreover, they will certainly also be generally applicable to studies addressing rapid stimulation-mediated ceramide release in living cells.},
  language  = {en}
}
@article{DerakhshaniKurzJaptoketal.2019,
  author    = {Derakhshani, Shaghayegh and Kurz, Andreas and Japtok, Lukasz and Schumacher, Fabian and Pilgram, Lisa and Steinke, Maria and Kleuser, Burkhard and Sauer, Markus and Schneider-Schaulies, Sibylle and Avota, Elita},
  title     = {Measles Virus Infection Fosters Dendritic Cell Motility in a 3D Environment to Enhance Transmission to Target Cells in the Respiratory Epithelium},
  series = {Frontiers in immunology},
  volume    = {10},
  journal   = {Frontiers in immunology},
  publisher = {Frontiers Research Foundation},
  address   = {Lausanne},
  issn      = {1664-3224},
  doi       = {10.3389/fimmu.2019.01294},
  pages     = {14},
  year      = {2019},
  abstract  = {Transmission of measles virus (MV) from dendritic to airway epithelial cells is considered as crucial to viral spread late in infection. Therefore, pathways and effectors governing this process are promising targets for intervention. To identify these, we established a 3D respiratory tract model where MV transmission by infected dendritic cells (DCs) relied on the presence of nectin-4 on H358 lung epithelial cells. Access to recipient cells is an important prerequisite for transmission, and we therefore analyzed migration of MV-exposed DC cultures within the model. Surprisingly, enhanced motility toward the epithelial layer was observed for MV-infected DCs as compared to their uninfected siblings. This occurred independently of factors released from H358 cells indicating that MV infection triggered cytoskeletal remodeling associated with DC polarization enforced velocity. Accordingly, the latter was also observed for MV-infected DCs in collagen matrices and was particularly sensitive to ROCK inhibition indicating infected DCs preferentially employed the amoeboid migration mode. This was also implicated by loss of podosomes and reduced filopodial activity both of which were retained in MV-exposed uninfected DCs. Evidently, sphingosine kinase (SphK) and sphingosine-1-phosphate (S1P) as produced in response to virus-infection in DCs contributed to enhanced velocity because this was abrogated upon inhibition of sphingosine kinase activity. These findings indicate that MV infection promotes a push-and-squeeze fast amoeboid migration mode via the SphK/S1P system characterized by loss of filopodia and podosome dissolution. Consequently, this enables rapid trafficking of virus toward epithelial cells during viral exit.},
  language  = {en}
}
@article{FinkSchumacherSchlegeletal.2021,
  author    = {Fink, Julian and Schumacher, Fabian and Schlegel, Jan and Stenzel, Philipp and Wigger, Dominik and Sauer, Markus and Kleuser, Burkhard and Seibel, J{\"u}rgen},
  title     = {Azidosphinganine enables metabolic labeling and detection of sphingolipid de novo synthesis},
  series = {Organic \& biomolecular chemistry : an international journal of synthetic, physical and biomolecular organic chemistry},
  volume    = {19},
  journal   = {Organic \& biomolecular chemistry : an international journal of synthetic, physical and biomolecular organic chemistry},
  number    = {10},
  publisher = {Royal Society of Chemistry},
  address   = {Cambridge},
  issn      = {1477-0520},
  doi       = {10.1039/d0ob02592e},
  pages     = {2203 -- 2212},
  year      = {2021},
  abstract  = {Here were report the combination of biocompatible click chemistry of omega-azidosphinganine with fluorescence microscopy and mass spectrometry as a powerful tool to elaborate the sphingolipid metabolism. The azide probe was efficiently synthesized over 13 steps starting from l-serine in an overall yield of 20\% and was used for live-cell fluorescence imaging of the endoplasmic reticulum in living cells by bioorthogonal click reaction with a DBCO-labeled fluorophore revealing that the incorporated analogue is mainly localized in the endoplasmic membrane like the endogenous species. A LC-MS(/MS)-based microsomal in vitro assay confirmed that omega-azidosphinganine mimics the natural species enabling the identification and analysis of metabolic breakdown products of sphinganine as a key starting intermediate in the complex sphingolipid biosynthetic pathways. Furthermore, the sphinganine-fluorophore conjugate after click reaction was enzymatically tolerated to form its dihydroceramide and ceramide metabolites. Thus, omega-azidosphinganine represents a useful biofunctional tool for metabolic investigations both by in vivo fluorescence imaging of the sphingolipid subcellular localization in the ER and by in vitro high-resolution mass spectrometry analysis. This should reveal novel insights of the molecular mechanisms sphingolipids and their processing enzymes have e.g. in infection.},
  language  = {en}
}