@article{PilsKoppPetersonetal.2012, author = {Pils, Stefan and Kopp, Kathrin and Peterson, Lisa and Tascon, Julia Delgado and Nyffenegger-Jann, Naja J. and Hauck, Christof R.}, title = {The Adaptor Molecule Nck Localizes the WAVE Complex to Promote Actin Polymerization during CEACAM3-Mediated Phagocytosis of Bacteria}, series = {PLoS One}, volume = {7}, journal = {PLoS One}, number = {3}, doi = {10.1371/journal.pone.0032808}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-131747}, pages = {e32808}, year = {2012}, abstract = {Background: CEACAM3 is a granulocyte receptor mediating the opsonin-independent recognition and phagocytosis of human-restricted CEACAM-binding bacteria. CEACAM3 function depends on an intracellular immunoreceptor tyrosine-based activation motif (ITAM)-like sequence that is tyrosine phosphorylated by Src family kinases upon receptor engagement. The phosphorylated ITAM-like sequence triggers GTP-loading of Rac by directly associating with the guanine nucleotide exchange factor (GEF) Vav. Rac stimulation in turn is critical for actin cytoskeleton rearrangements that generate lamellipodial protrusions and lead to bacterial uptake. Principal Findings: In our present study we provide biochemical and microscopic evidence that the adaptor proteins Nck1 and Nck2, but not CrkL, Grb2 or SLP-76, bind to tyrosine phosphorylated CEACAM3. The association is phosphorylation-dependent and requires the Nck SH2 domain. Overexpression of the isolated Nck1 SH2 domain, RNAi-mediated knock-down of Nck1, or genetic deletion of Nck1 and Nck2 interfere with CEACAM3-mediated bacterial internalization and with the formation of lamellipodial protrusions. Nck is constitutively associated with WAVE2 and directs the actin nucleation promoting WAVE complex to tyrosine phosphorylated CEACAM3. In turn, dominant-negative WAVE2 as well as shRNA-mediated knock-down of WAVE2 or the WAVE-complex component Nap1 reduce internalization of bacteria. Conclusions: Our results provide novel mechanistic insight into CEACAM3-initiated phagocytosis. We suggest that the CEACAM3 ITAM-like sequence is optimized to co-ordinate a minimal set of cellular factors needed to efficiently trigger actin-based lamellipodial protrusions and rapid pathogen engulfment.}, language = {en} } @article{MayerRabindranathBoerneretal.2013, author = {Mayer, Matthias and Rabindranath, Raman and B{\"o}rner, Juliane and H{\"o}rner, Eva and Bentz, Alexander and Salgado, Josefina and Han, Hong and B{\"o}se, Holger and Probst, J{\"o}rn and Shamonin, Mikhail and Monkman, Gereth J. and Schlunck, G{\"u}nther}, title = {Ultra-Soft PDMS-Based Magnetoactive Elastomers as Dynamic Cell Culture Substrata}, series = {PLOS ONE}, volume = {8}, journal = {PLOS ONE}, number = {10}, issn = {1932-6203}, doi = {10.1371/journal.pone.0076196}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-128246}, pages = {e76196}, year = {2013}, abstract = {Mechanical cues such as extracellular matrix stiffness and movement have a major impact on cell differentiation and function. To replicate these biological features in vitro, soft substrata with tunable elasticity and the possibility for controlled surface translocation are desirable. Here we report on the use of ultra-soft (Young's modulus <100 kPa) PDMS-based magnetoactive elastomers (MAE) as suitable cell culture substrata. Soft non-viscous PDMS (<18 kPa) is produced using a modified extended crosslinker. MAEs are generated by embedding magnetic microparticles into a soft PDMS matrix. Both substrata yield an elasticity-dependent (14 vs. 100 kPa) modulation of alpha-smooth muscle actin expression in primary human fibroblasts. To allow for static or dynamic control of MAE material properties, we devise low magnetic field (approximate to 40 mT) stimulation systems compatible with cell-culture environments. Magnetic field-instigated stiffening (14 to 200 kPa) of soft MAE enhances the spreading of primary human fibroblasts and decreases PAX-7 transcription in human mesenchymal stem cells. Pulsatile MAE movements are generated using oscillating magnetic fields and are well tolerated by adherent human fibroblasts. This MAE system provides spatial and temporal control of substratum material characteristics and permits novel designs when used as dynamic cell culture substrata or cell culture-coated actuator in tissue engineering applications or biomedical devices.}, language = {en} } @article{LeichWeissbachKleinetal.2013, author = {Leich, E. and Weißbach, S. and Klein, H.-U. and Grieb, T. and Pischimarov, J. and St{\"u}hmer, T. and Chatterjee, M. and Steinbrunn, T. and Langer, C. and Eilers, M. and Knop, S. and Einsele, H. and Bargou, R. and Rosenwald, A.}, title = {Multiple myeloma is affected by multiple and heterogeneous somatic mutations in adhesion- and receptor tyrosine kinase signaling molecules}, series = {Blood Cancer Journal}, volume = {3}, journal = {Blood Cancer Journal}, number = {e102}, doi = {10.1038/bcj.2012.47}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-128663}, year = {2013}, abstract = {Multiple myeloma (MM) is a largely incurable plasma cell malignancy with a poorly understood and heterogeneous clinical course. To identify potential, functionally relevant somatic mutations in MM, we performed whole-exome sequencing of five primary MM, corresponding germline DNA and six MM cell lines, and developed a bioinformatics strategy that also integrated published mutational data of 38 MM patients. Our analysis confirms that identical, recurrent mutations of single genes are infrequent in MM, but highlights that mutations cluster in important cellular pathways. Specifically, we show enrichment of mutations in adhesion molecules of MM cells, emphasizing the important role for the interaction of the MM cells with their microenvironment. We describe an increased rate of mutations in receptor tyrosine kinases (RTKs) and associated signaling effectors, for example, in EGFR, ERBB3, KRAS and MAP2K2, pointing to a role of aberrant RTK signaling in the development or progression of MM. The diversity of mutations affecting different nodes of a particular signaling network appears to be an intrinsic feature of individual MM samples, and the elucidation of intra- as well as interindividual redundancy in mutations that affect survival pathways will help to better tailor targeted therapeutic strategies to the specific needs of the MM patient.}, language = {en} } @article{EmmertWitzelHeinrich2016, author = {Emmert, M. and Witzel, P. and Heinrich, D.}, title = {Challenges in tissue engineering - towards cell control inside artificial scaffolds}, series = {Soft Matter}, volume = {12}, journal = {Soft Matter}, number = {19}, doi = {10.1039/c5sm02844b}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-191341}, pages = {4287-4294}, year = {2016}, abstract = {Control of living cells is vital for the survival of organisms. Each cell inside an organism is exposed to diverse external mechano-chemical cues, all coordinated in a spatio-temporal pattern triggering individual cell functions. This complex interplay between external chemical cues and mechanical 3D environments is translated into intracellular signaling loops. Here, we describe how external mechano-chemical cues control cell functions, especially cell migration, and influence intracellular information transport. In particular, this work focuses on the quantitative analysis of (1) intracellular vesicle transport to understand intracellular state changes in response to external cues, (2) cellular sensing of external chemotactic cues, and (3) the cells' ability to migrate in 3D structured environments, artificially fabricated to mimic the 3D environment of tissue in the human body.}, language = {en} } @article{CollenburgBeyersdorfWieseetal.2017, author = {Collenburg, Lena and Beyersdorf, Niklas and Wiese, Teresa and Arenz, Christoph and Saied, Essa M. and Becker-Flegler, Katrin Anne and Schneider-Schaulies, Sibylle and Avota, Elita}, title = {The activity of the neutral sphingomyelinase is important in T cell recruitment and directional migration}, series = {Frontiers in Immunology}, volume = {8}, journal = {Frontiers in Immunology}, number = {1007}, doi = {10.3389/fimmu.2017.01007}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-158944}, year = {2017}, abstract = {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 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.}, language = {en} } @article{AlzheimerSvenssonKoenigetal.2020, author = {Alzheimer, Mona and Svensson, Sarah L. and K{\"o}nig, Fabian and Schweinlin, Matthias and Metzger, Marco and Walles, Heike and Sharma, Cynthia M.}, title = {A three-dimensional intestinal tissue model reveals factors and small regulatory RNAs important for colonization with Campylobacter jejuni}, series = {PLoS Pathogens}, volume = {16}, journal = {PLoS Pathogens}, number = {2}, doi = {10.1371/journal.ppat.1008304}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-229454}, year = {2020}, abstract = {The Gram-negative Epsilonproteobacterium Campylobacter jejuni is currently the most prevalent bacterial foodborne pathogen. Like for many other human pathogens, infection studies with C. jejuni mainly employ artificial animal or cell culture models that can be limited in their ability to reflect the in-vivo environment within the human host. Here, we report the development and application of a human three-dimensional (3D) infection model based on tissue engineering to study host-pathogen interactions. Our intestinal 3D tissue model is built on a decellularized extracellular matrix scaffold, which is reseeded with human Caco-2 cells. Dynamic culture conditions enable the formation of a polarized mucosal epithelial barrier reminiscent of the 3D microarchitecture of the human small intestine. Infection with C. jejuni demonstrates that the 3D tissue model can reveal isolate-dependent colonization and barrier disruption phenotypes accompanied by perturbed localization of cell-cell junctions. Pathogenesis-related phenotypes of C. jejuni mutant strains in the 3D model deviated from those obtained with 2D-monolayers, but recapitulated phenotypes previously observed in animal models. Moreover, we demonstrate the involvement of a small regulatory RNA pair, CJnc180/190, during infections and observe different phenotypes of CJnc180/190 mutant strains in 2D vs. 3D infection models. Hereby, the CJnc190 sRNA exerts its pathogenic influence, at least in part, via repression of PtmG, which is involved in flagellin modification. Our results suggest that the Caco-2 cell-based 3D tissue model is a valuable and biologically relevant tool between in-vitro and in-vivo infection models to study virulence of C. jejuni and other gastrointestinal pathogens.}, language = {en} }