TY - JOUR AB - T cells develop in the thymus in a highly specialized cellular and extracellular microenvironment. The basement membrane molecule, laminin-5 (LN-5), is predominantly found in the medulla of the human thymic lobules. Using high-resolution light microscopy, we show here that LN-5 is localized in a bi-membranous conduit-like structure, together with other typical basement membrane components including collagen type IV, nidogen and perlecan. Other interstitial matrix components, such as fibrillin-1 or -2, tenascin-C or fibrillar collagen types, were also associated with these structures. Three-dimensional (3D) confocal microscopy suggested a tubular structure, whereas immunoelectron and transmission electron microscopy showed that the core of these tubes contained fibrillar collagens enwrapped by the LN-5-containing membrane. These medullary conduits are surrounded by thymic epithelial cells, which in vitro were found to bind LN-5, but also fibrillin and tenascin-C. Dendritic cells were also detected in close vicinity to the conduits. Both of these stromal cell types express major histocompatibility complex (MHC) class II molecules capable of antigen presentation. The conduits are connected to blood vessels but, with an average diameter of 2 mum, they are too small to transport cells. However, evidence is provided that smaller molecules such as a 10 kDa dextran, but not large molecules (>500 kDa), can be transported in the conduits. These results clearly demonstrate that a conduit system, which is also known from secondary lymphatic organs such as lymph nodes and spleen, is present in the medulla of the human thymus, and that it might serve to transport small blood-borne molecules or chemokines to defined locations within the medulla. AU - Drumea-Mirancea, Mihaela AU - Wessels, Johannes T AU - Müller, Claudia A AU - Essl, Mike AU - Eble, Johannes A AU - Tolosa, Eva AU - Koch, Manuel AU - Reinhardt, Dieter P AU - Michael Sixt AU - Sorokin, Lydia AU - Stierhof, York-Dieter AU - Schwarz, Heinz AU - Klein, Gerd ID - 3934 IS - Pt 7 JF - Journal of Cell Science TI - Characterization of a conduit system containing laminin-5 in the human thymus: a potential transport system for small molecules VL - 119 ER - TY - JOUR AB - Integrins regulate cell behavior through the assembly of multiprotein complexes at the site of cell adhesion. Parvins are components of such a multiprotein complex. They consist of three members (alpha-, beta-, and gamma-parvin), form a functional complex with integrin-linked kinase (ILK) and PINCH, and link integrins to the actin cytoskeleton. Whereas alpha- and beta-parvins are widely expressed, gamma-parvin has been reported to be expressed in hematopoietic organs. In the present study, we report the expression pattern of the parvins in hematopoietic cells and the phenotypic analysis of gamma-parvin-deficient mice. Whereas alpha-parvin is not expressed in hematopoietic cells, beta-parvin is only found in myeloid cells and gamma-parvin is present in both cells of the myeloid and lymphoid lineages, where it binds ILK. Surprisingly, loss of gamma-parvin expression had no effect on blood cell differentiation, proliferation, and survival and no consequence for the T-cell-dependent antibody response and lymphocyte and dendritic cell migration. These data indicate that despite the high expression of gamma-parvin in hematopoietic cells it must play a more subtle role for blood cell homeostasis. AU - Chu, Haiyan AU - Thievessen, Ingo AU - Michael Sixt AU - Lämmermann, Tim AU - Waisman, Ari AU - Braun, Attila AU - Noegel, Angelika A AU - Fässler, Reinhard ID - 3935 IS - 5 JF - Molecular and Cellular Biology TI - γ-Parvin is dispensable for hematopoiesis, leukocyte trafficking, and T-cell-dependent antibody response VL - 26 ER - TY - JOUR AB - At least eight of the twelve known members of the beta1 integrin family are expressed on hematopoietic cells. Among these, the VCAM-1 receptor alpha4beta1 has received most attention as a main factor mediating firm adhesion to the endothelium during blood cell extravasation. Therapeutic trials are ongoing into the use of antibodies and small molecule inhibitors to target this interaction and hence obtain anti-inflammatory effects. However, extravasation is only one possible process that is mediated by beta1 integrins and there is evidence that they also mediate leukocyte retention and positioning in the tissue, lymphocyte activation and possibly migration within the interstitium. Genetic mouse models where integrins are selectively deleted on blood cells have been used to investigate these functions and further studies will be invaluable to critically evaluate therapeutic trials. AU - Michael Sixt AU - Bauer, Martina AU - Lämmermann, Tim AU - Fässler, Reinhard ID - 3936 IS - 5 JF - Current Opinion in Cell Biology TI - β1 integrins: zip codes and signaling relay for blood cells VL - 18 ER - TY - JOUR AB - Wnt11 is a key signal, determining cell polarization and migration during vertebrate gastrulation. It is known that Wnt11 functionally interacts with several signaling components, the homologues of which control planar cell polarity in Drosophila melanogaster. Although in D. melanogaster these components are thought to polarize cells by asymmetrically localizing at the plasma membrane, it is not yet clear whether their subcellular localization plays a similarly important role in vertebrates. We show that in zebrafish embryonic cells, Wnt11 locally functions at the plasma membrane by accumulating its receptor, Frizzled 7, on adjacent sites of cell contacts. Wnt11-induced Frizzled 7 accumulations recruit the intracellular Wnt signaling mediator Dishevelled, as well as Wnt11 itself, and locally increase cell contact persistence. This increase in cell contact persistence is mediated by the local interaction of Wnt11, Frizzled 7, and the atypical cadherin Flamingo at the plasma membrane, and it does not require the activity of further downstream effectors of Wnt11 signaling, such as RhoA and Rok2. We propose that Wnt11, by interacting with Frizzled 7 and Flamingo, modulates local cell contact persistence to coordinate cell movements during gastrulation. AU - Witzel, Sabine AU - Zimyanin, Vitaly AU - Carreira Barbosa, Filipa AU - Tada, Masazumi AU - Heisenberg, Carl-Philipp J ID - 4140 IS - 5 JF - Journal of Cell Biology TI - Wnt11 controls cell contact persistence by local accumulation of Frizzled 7 at the plasma membrane VL - 175 ER - TY - JOUR AB - The detection of microRNAs (miRNAs) at single-cell resolution is important for studying the role of these posttranscriptional regulators. Here, we use a dual-fluorescent green fluorescent protein (GFP)-reporter/monomeric red fluorescent protein (mRFP)-sensor (DFRS) plasmid, injected into zebrafish blastomeres or electroporated into defined tissues of mouse embryos in utero or ex utero, to monitor the dynamics of specific miRNAs in individual live cells. This approach reveals, for example, that in the developing mouse central nervous system,, miR-124a is expressed not only in postmitotic neurons but also in neuronal progenitor cells. Collectively, our results demonstrate that acute administration of DFRS plasmids.offers an alternative to previous in situ hybridization and transgenic approaches and allows the monitoring of miRNA appearance and disappearance in defined cell lineages during vertebrate development. AU - Tonelli, Davide AU - Calegari, Frederico AU - Fei, Ji AU - Nomura, Tadashi AU - Osumi, Noriko AU - Heisenberg, Carl-Philipp J AU - Huttner, Wieland ID - 4145 IS - 6 JF - Biotechniques TI - Single-cell detection of microRNAs in developing vertebrate embryos after acute administration of a dual-fluorescence reporter/sensor plasmid VL - 41 ER -