The Sphingosine-1-Phosphate (S1P) Lysophospholipid Receptor S1P3 Regulates MAdCAM-1+ Endothelial Cells in Splenic Marginal Sinus Organization

Marginal zones (MZs) are microdomains in the spleen that contain various types of immune cells, including MZ B cells, MOMA1+ metallophilic macrophages, and mucosal addressin cell adhesion molecule 1 (MAdCAM-1)+ endothelial cells. MAdCAM-1+ and MOMA1+ cells line the sinus, that separates MZs from splenic follicles. Here we show that a receptor for the lysophospholipid sphingosine-1-phosphate (S1P), S1P3, is required for normal numbers of splenic immature and MZ B cells, and for S1P-induced chemotaxis of MZ B cells. S1P3 is also essential for proper alignment of MOMA1+ macrophages and MAdCAM-1+ endothelial cells along the marginal sinus. The lack of cohesion of the marginal sinus in S1P3−/− mice affects MZ B cell functions, as wild-type (WT) MZ B cells migrate more into S1P3−/− follicles than into WT follicles after treatment with lipopolysaccharide. Additionally, short-term homing experiments demonstrate that WT MZ B cells home to the S1P3−/− spleen in increased numbers, suggesting a role for the marginal sinus in regulating MZ B cells numbers. Moreover, S1P3−/− mice are defective in mounting immune responses to thymus-independent antigen type 2 due to defects in radiation-resistant cells in the spleen. These data identify lysophospholipids and the S1P3 receptor as essential regulators of the MZ sinus and its role as a barrier to the follicle

Heterodimerization of AML1/ETO with CBFβ is required for leukemogenesis but not for myeloproliferation

The AML1/Runx1 transcription factor and its heterodimerization partner CBFβ are essential regulators of myeloid differentiation. The chromosomal translocation t(8;21), fusing the DNA binding domain of AML1 to the corepressor eight-twenty-one (ETO), is frequently associated with acute myeloid leukemia and generates the AML1/ETO (AE) fusion protein. AE represses target genes usually activated by AML1 and also affects the endogenous repressive function of ETO at Notch target genes. In order to analyze the contribution of CBFβ in AE-mediated leukemogenesis and deregulation of Notch target genes, we introduced two point mutations in a leukemia-initiating version of AE in mice, called AE9a, that disrupt the AML1/CBFβ interaction (AE9aNT). We report that the AE9a/CBFβ interaction is not required for the AE9a-mediated aberrant expression of AML1 target genes, while upregulation/derepression of Notch target genes does require the interaction with CBFβ. Using retroviral transduction to express AE9a in murine adult bone marrow-derived hematopoietic progenitors, we observed that both AE9a and AE9aNT lead to increased myeloproliferation in vivo. However, both development of leukemia and long-term replating capacity are only observed with AE9a but not with AE9aNT. Thus, deregulation of both AML1 and Notch target genes is required for the development of AE9a-driven leukemia

Setting the Stage for Notch: The Drosophila Su(H)-Hairless Repressor Complex.

Notch signaling is iteratively used throughout development to maintain stem cell potential or in other instances allow differentiation. The central transcription factor in Notch signaling is CBF-1/RBP-J, Su(H), Lag-1 (CSL)-Su(H) in Drosophila-which functions as a molecular switch between transcriptional activation and repression. Su(H) represses transcription by forming a complex with the corepressor Hairless (H). The Su(H)-repressor complex not only competes with the Notch intracellular domain (NICD) but also configures the local chromatin landscape. In this issue, Yuan and colleagues determined the structure of the Su(H)/H complex, showing that a major conformational change within Su(H) explains why the binding of NICD and H is mutually exclusive

Association of the Mediator complex with enhancers of active genes

International audienceThe multiprotein Mediator complex has been shown to interact with gene-specific regulatory proteins and RNA polymerase II in vitro. Here, we use chromatin immunoprecipitation to analyze the recruitment of Mediator to GAL genes of yeast in vivo. We find that Mediator associates exclusively with transcriptionally active and not inactive GAL genes. This association maps to the upstream activating sequence, rather than the core promoter, and is independent of RNA polymerase II, general transcription factors, and core promoter sequences. These findings support the idea of Mediator as a primary conduit of regulatory information from enhancers to promoters in eukaryotic cells

Surface views of the CSL coactivator complex (upper) and corepressor complexes (lower).

<p>(A) The DNA-bound CSL activator complex consists of CSL (green), NICD (RAM domain, red; ankyrin repeats, yellow), and mastermind (MAM, orange). (PDB-ID: 1TTU). (B) KyoT2 (red) interacts with the BTD of CSL, similar to the NICD RAM domain (RAM-type). (PDB-ID: 4J2X). (C) Hairless interacts with the CTD of Su(H), resulting in a dramatic change of CTD conformation (H-type). (PDB-ID: 5E24). (D) The crystal structure of the SMRT/HDAC1 associated repressor protein (SHARP)-CSL corepressor complex and the CSL-RBPJ interacting and tubulin associated (RITA) corepressor complex is unknown at the moment (PDB-ID, RBPJ bound to DNA: 3BRG).</p

Available CSL complex structure data (protein data bank [PDB] database).

<p>Available CSL complex structure data (protein data bank [PDB] database).</p

Chromatin regulator spen/sharp in x inactivation and disease

Enzymes, such as histone methyltransferases and demethylases, histone acetyltransferases and deacetylases, and DNA methyltransferases are known as epigenetic modifiers that are often implicated in tumorigenesis and disease. One of the best‐studied chromatin-based mechanism is X chromosome inactivation (XCI), a process that establishes facultative heterochromatin on only one X chromosome in females and establishes the right dosage of gene expression. The specificity factor for this process is the long non‐coding RNA X inactive specific transcript (Xist), which is upregulated from one X chromosome in female cells. Subsequently, Xist is bound by the corepressor SHARP/SPEN, recruiting and/or activating histone deacetylases (HDACs), leading to the loss of active chromatin marks such as H3K27ac. In addition, polycomb complexes PRC1 and PRC2 establish wide‐spread accumulation of H3K27me3 and H2AK119ub1 chromatin marks. The lack of active marks and establishment of repressive marks set the stage for DNA methyltransferases (DNMTs) to stably silence the X chromosome. Here, we will review the recent advances in understanding the molecular mechanisms of how heterochromatin formation is established and put this into the context of carcinogenesis and disease

The Notch intracellular domain integrates signals from Wnt, Hedgehog, TGFβ/BMP and hypoxia pathways

Notch signaling is a highly conserved signal transduction pathway that regulates stem cell maintenance and differentiation in several organ systems. Upon activation, the Notch receptor is proteolytically processed, its intracellular domain (NICD) translocates into the nucleus and activates expression of target genes. Output, strength and duration of the signal are tightly regulated by post-translational modifications. Here we review the intracellular post-translational regulation of Notch that fine-tunes the outcome of the Notch response. We also describe how crosstalk with other conserved signaling pathways like the Wnt, Hedgehog, hypoxia and TGFβ/BMP pathways can affect Notch signaling output. This regulation can happen by regulation of ligand, receptor or transcription factor expression, regulation of protein stability of intracellular key components, usage of the same cofactors or coregulation of the same key target genes. Since carcinogenesis is often dependent on at least two of these pathways, a better understanding of their molecular crosstalk is pivotal.publisher: Elsevier articletitle: The Notch intracellular domain integrates signals from Wnt, Hedgehog, TGFβ/BMP and hypoxia pathways journaltitle: Biochimica et Biophysica Acta (BBA) - Molecular Cell Research articlelink: http://dx.doi.org/10.1016/j.bbamcr.2015.11.020 content_type: article copyright: Copyright © 2015 The Authors. Published by Elsevier B.V.status: publishe

Fibrin sheaths in central venous port catheters: treatment with low-dose, single injection of urokinase on an outpatient basis

Purpose: Evaluation of the efficacy of single-shot, low-dose urokinase administration for the treatment of port catheter-associated fibrin sheaths. Methods: Forty-six patients were retrospectively evaluated for 54 episodes of port catheter dysfunction. The presence of a fibrin sheath was detected by angiographic contrast examinations. On an outpatient basis, patients subsequently received thrombolysis consisting of a single injection of urokinase (15.000 IU in 1.5 mL normal saline) through the port system. A second attempt was made in cases of treatment failure. Patients were followed up for technical success, complications and long-term outcome. Results: Port dysfunction occurred at a median of 117 days after implantation (range: 7-825 days). The technical success after first port dysfunction by thrombolysis was 87% (40/46); thereof, initial thrombolysis was effective in 78% (36/46). Nine patients (20%) received a second dose of urokinase after previous treatment failure. Follow-up was available for 26 of 40 patients after successful thrombolysis. In 8 of these, rethrombosis occurred after a median of 98 days (range: 21-354 days), whereby rethrombolysis was effective in 5 of 7 (63%) patients. The overall success of all thrombolyses performed was 70% (45/64). No procedure-related technical or clinical complications occurred. After first favorable thrombolysis, a Kaplan-Meier analysis yielded a 30-, 90- and 180-day probability of patency of 96%, 87% and 81%. Conclusion: Thrombolytic therapy on an outpatient basis appears to be a safe and efficient. Three-month patency rates are comparable to more invasive treatment options, including catheter exchange over a guide wire and percutaneous fibrin sheath stripping