Suppression of Th2 cell development by Notch ligands Delta1 and Delta4

Notch signaling plays important roles in Th cell activation. We show that in response to TLR ligation, dendritic cells up-regulate expression of Notch ligands Delta1 and Delta4 via a MyD88-dependent pathway. Expression of Delta1 or Delta4 by dendritic cells enhanced their ability to activate naive Th cells and promote Th1 cell development, and allowed them to strongly inhibit Th2 cell development. Promotion of Th1 cell development was dependent on IFN-gamma and T-bet expression by responding Th cells. However, the inhibition of Th2 cell development occurred independently of IFN-gamma or T-bet, and resulted from a block in IL-4-initiated commitment to the Th2 lineage. The promotion of Th1 cell development by Delta is not a reflection of the delivery of pro-Th1 instructional signal, but rather it is the result of a block in the downstream effects initiated by IL-4 signaling.     

Interactions of hydroxyapatite with proteins and its toxicological effect to zebrafish embryos development

The increased application of nanomaterials has raised the level of public concern regarding possible toxicities caused by exposure to nanostructures. The interactions of nanosized hydroxyapatite (HA) with cytochrome c and hemoglobin were investigated by zeta-potential, UV-vis, fluorescence and circular dichroism. The experimental results indicated that the interactions were formed via charge attraction and hydrogen bond and obeyed Langmuir adsorption isotherm. The two functional proteins bridged between HA particles to aggregate into the coralloid form, where change of the secondary structure of proteins occurred. From effects of nanosized HA, SiO(2) and TiO(2) particles on the zebrafish embryos development, they were adsorbed on the membrane surface confirmed by the electronic scanning microscopy. Nano-HA aggregated into the biggest particles around the membrane protein and then caused a little toxicity to development of zebrafish embryos. The SiO(2) particles were distributed throughout the outer surface and caused jam of membrane passage, delay of the hatching time and axial malformation. Maybe owing to the oxygen free radical activity, TiO(2) caused some serious deformity characters in the cardiovascular system.     

Notch ligands with contrasting functions: Jagged1 and Delta1 in the mouse inner ear

Each of the sensory patches in the epithelium of the inner ear is a mosaic of hair cells and supporting cells. Notch signalling is thought to govern this pattern of differentiation through lateral inhibition. Recent experiments in the chick suggest, however, that Notch signalling also has a prior function - inductive rather than inhibitory - in defining the prosensory patches from which the differentiated cells arise. Several Notch ligands are expressed in each patch, but their individual roles in relation to the two functions of Notch signalling are unclear. We have used a Cre-LoxP approach to knock out two of these ligands, Delta1 (Dll1) and Jagged1 (Jag1), in the mouse ear. In the absence of Dll1, auditory hair cells develop early and in excess, in agreement with the lateral inhibition hypothesis. In the absence of Jag1, by contrast, the total number of these cells is strongly reduced, with complete loss of cochlear outer hair cells and some groups of vestibular hair cells, indicating that Jag1 is required for the prosensory inductive function of Notch. The number of cochlear inner hair cells, however, is almost doubled. This correlates with loss of expression of the cell cycle inhibitor p27(Kip1) (Cdkn1b), suggesting that signalling by Jag1 is also needed to limit proliferation of prosensory cells, and that there is a core part of this population whose prosensory character is established independently of Jag1-Notch signalling. Our findings confirm that Notch signalling in the ear has distinct prosensory and lateral-inhibitory functions, for which different ligands are primarily responsible.     

Cargo of kinesin identified as JIP scaffolding proteins and associated signaling molecules

The cargo that the molecular motor kinesin moves along microtubules has been elusive. We searched for binding partners of the COOH terminus of kinesin light chain, which contains tetratricopeptide repeat (TPR) motifs. Three proteins were found, the c-jun NH(2)-terminal kinase (JNK)-interacting proteins (JIPs) JIP-1, JIP-2, and JIP-3, which are scaffolding proteins for the JNK signaling pathway. Concentration of JIPs in nerve terminals requires kinesin, as evident from the analysis of JIP COOH-terminal mutants and dominant negative kinesin constructs. Coprecipitation experiments suggest that kinesin carries the JIP scaffolds preloaded with cytoplasmic (dual leucine zipper-bearing kinase) and transmembrane signaling molecules (the Reelin receptor, ApoER2). These results demonstrate a direct interaction between conventional kinesin and a cargo, indicate that motor proteins are linked to their membranous cargo via scaffolding proteins, and support a role for motor proteins in spatial regulation of signal transduction pathways.     

Physical interaction of Delta1, Jagged1, and Jagged2 with Notch1 and Notch3 receptors

The Delta/Serrate/LAG-2 (DSL) domain-containing proteins, Delta1, Jagged1, and Jagged2, are considered to be ligands for Notch receptors. However, the physical interaction between the three DSL proteins and respective Notch receptors remained largely unknown. In this study, we investigated this issue through the targeting of Notch1 and Notch3 in two experimental systems using fusion proteins comprising their extracellular portions. Cell-binding assays showed that soluble forms of Notch1 and Notch3 proteins physically bound to the three DSL proteins on the cell surface. In solid-phase binding assays using immobilized soluble Notch1 and Notch3 proteins, it was revealed that each DSL protein directly bound to the soluble Notch proteins with different affinities. All interactions between the DSL proteins and soluble Notch proteins were dependent on Ca(2+). Taken together, these results suggest that Delta1, Jagged1, and Jagged2 are ligands for Notch1 and Notch3 receptors.     

Structural basis for interaction of DivIVA/GpsB proteins with their ligands

DivIVA proteins and their GpsB homologues are late cell division proteins found in Gram‐positive bacteria. DivIVA/GpsB proteins associate with the inner leaflet of the cytosolic membrane and act as scaffolds for other proteins required for cell growth and division. DivIVA/GpsB proteins comprise an N‐terminal lipid‐binding domain for membrane association fused to C‐terminal domains supporting oligomerization. Despite sharing the same domain organization, DivIVA and GpsB serve different cellular functions: DivIVA plays diverse roles in division site selection, chromosome segregation and controlling peptidoglycan homeostasis, whereas GpsB contributes to the spatiotemporal control of penicillin‐binding protein activity. The crystal structures of the lipid‐binding domains of DivIVA from Bacillus subtilis and GpsB from several species share a fold unique to this group of proteins, whereas the C‐terminal domains of DivIVA and GpsB are radically different. A number of pivotal features identified from the crystal structures explain the functional differences between the proteins. Herein we discuss these structural and functional relationships and recent advances in our understanding of how DivIVA/GpsB proteins bind and recruit their interaction partners, knowledge that might be useful for future structure‐based DivIVA/GpsB inhibitor design.     

The intracellular domain of the human protocadherin hFat1 interacts with Homer signalling scaffolding proteins

The cadherin superfamily protein Fat1 is known to interact with the EVH1 domain of mammalian Ena/VASP. Here we demonstrate that: (i) the scaffolding proteins Homer-3 and Homer-1 also interact with the EVH1 binding site of hFat1 in vitro, and (ii) binding of Homer-3 and Mena to hFat1 is mutually competitive. Endogenous Fat1 binds to immobilised Homer-3 and endogenous Homer-3 binds to immobilised Fat1. Both, endogenous and over-expressed Fat1 exhibit co-localisation with Homer-3 in cellular protrusions and at the plasma membrane of HeLa cells. As Homer proteins and Fat1 have been both linked to psychic disorders, their interaction may be of patho-physiological importance.     

Inhibition of Jagged-mediated Notch signaling disrupts zebrafish biliary development and generates multi-organ defects compatible with an Alagille syndrome phenocopy

The Alagille Syndrome (AGS) is a heritable disorder affecting the liver and other organs. Causative dominant mutations in human Jagged 1 have been identified in most AGS patients. Related organ defects occur in mice that carry jagged 1 and notch 2 mutations. Multiple jagged and notch genes are expressed in the developing zebrafish liver. Compound jagged and notch gene knockdowns alter zebrafish biliary, kidney, pancreatic, cardiac and craniofacial development in a manner compatible with an AGS phenocopy. These data confirm an evolutionarily conserved role for Notch signaling in vertebrate liver development, and support the zebrafish as a model system for diseases of the human biliary system.     

Intracellular cell-autonomous association of Notch and its ligands: a novel mechanism of Notch signal modification.

Notch (N) and its ligands, Delta (Dl) and Serrate (Ser), are membrane-spanning proteins with EGF repeats. They play an essential role in mediating proliferation and segregated differentiation of stem cells. One of the prominent features of N signal system is that its ligands are anchored to the plasma membrane, which allows the ligand/receptor association only between the neighboring cells. Various lines of evidences have verified this intercellular signal transmission, but there also have been implications that expression of Dl or Ser interferes cell-autonomously with the ability of the cell to receive N signal, implying that N and its ligands may interact in the same cell. Here, we demonstrate that N, Dl, and Ser cell-autonomously form homomeric or heteromeric complexes. The cell-autonomous heteromeric complexes are not present on the cell surface, implying that the association occurs in the endoreticulum or Golgi apparatus. Expression of Dl or Ser cell-autonomously reduces the N-mediated HES-5 promoter activity, indicating that the cell-autonomous association alters the N signal receptivity. Intracellular deletion of Dl shows elevated activity of this dominant-negative effect. In vivo overexpression study suggests that the cell-autonomous function of Dl and Ser is independent of the ligand specificity and may be modulated by Fringe (Fg), which inhibits the formation of the cell-autonomous Dl/N or Ser/N complex.     

Three novel Notch genes in zebrafish: implications for vertebrate Notch gene evolution and function

 Notch genes encode transmembrane receptors that interact with numerous signal transduction pathways and are essential for animal development. To facilitate analysis of vertebrate Notch gene function, we isolated cDNA fragments of three novel Notch genes from zebrafish (Danio rerio), Notch1b, Notch5 and Notch6. Notch1b is a second zebrafish Notch1 gene. From analysis of the Notch1b sequence we argue that the various vertebrate Notch gene subfamilies encode receptors with different signalling specificities. Notch5 and Notch6 represent novel vertebrate Notch gene subfamilies. Remarkably, Notch1b lacks expression in presomitic mesoderm, Notch5 is expressed in a metameric pattern within the presomitic mesoderm whilst Notch6 expression is excluded from the nervous system. The expression patterns of these genes suggest important roles in gastrulation, somitogenesis, tail bud extension, myogenesis, heart development and neurogenesis. We discuss the implications of our observations for Notch gene evolution and function. Received: 20 January 1997 / Accepted: 12 February 1997     

The adhesion force of Notch with Delta and the rate of Notch signaling

Notch signaling is repeatedly used during animal development to specify cell fates. Using atomic force microscopy on live cells, chemical inhibitors, and conventional analyses, we show that the rate of Notch signaling is linked to the adhesion force between cells expressing Notch receptors and Delta ligand. Both the Notch extracellular and intracellular domains are required for the high adhesion force with Delta. This high adhesion force is lost within minutes, primarily due to the action of Presenilin on Notch. Reduced turnover or Delta pulling accelerate this loss. These data suggest that strong adhesion between Notch and Delta might serve as a booster for initiating Notch signaling at a high rate.     

Dynamin isoform-specific interaction with the shank/ProSAP scaffolding proteins of the postsynaptic density and actin cytoskeleton

Dynamin is a GTPase involved in endocytosis and other aspects of membrane trafficking. A critical function in the presynaptic compartment attributed to the brain-specific dynamin isoform, dynamin-1, is in synaptic vesicle recycling. We report that dynamin-2 specifically interacts with members of the Shank/ProSAP family of postsynaptic density scaffolding proteins and present evidence that dynamin-2 is specifically associated with the postsynaptic density. These data are consistent with a role for this otherwise broadly distributed form of dynamin in glutamate receptor down-regulation and other aspects of postsynaptic membrane turnover.     

Locus Delta in the Notch signaling system: organization and pleiotropic function during Drosophila development

Delta locus is the important component of the Delta-Notch signaling system implicating in a general mechanism of local cell signaling. Delta and Notch encode the evolutionary conserved cell surface proteins that interact and function as ligand (DELTA) and receptor (NOTCH) in a wide variety of cell fate specification events during oogenesis, embryogenesis and metamorphosis.