text:
- © 2018 American Society of Plant Biologists. All Rights Reserved.
Transport networks may be defined as sets of connected nodes or hubs where cargo from different origins are sorted to their final destinations. The trans-Golgi network (TGN) is the most discussed and arguably busiest hub operating in the cell. The versatility of the plant TGN distinguishes it from its mammalian counterpart. It is essential for the assembly of cell walls, including the cell plate, and organizes traffic of cargoes not only to but also from the plasma membrane, two pathways that animal cells separately confine to TGN and endosomes, respectively. We stand far from models that integrate the multiple trafficking functions of the plant TGN in physiologically different cellular contexts; however, with current and recent studies, we are gaining insights into the molecular determinants, trafficking routes, and functions of TGN subcompartments.
The trans-Golgi network (TGN) is defined as the membrane compartment on the trans-side of Golgi stacks responsible for the sorting and packaging of cargo molecules for delivery to the plasma membrane and vacuoles (Roth et al., 1985; Griffiths and Simons, 1986; Kang et al., 2011). In plants, the TGN not only provides a final sorting station for Golgi-derived cargoes but also is involved in trafficking/recycling of endosomal material; therefore, the term TGN/early endosome (TGN/EE) better suits its function (Fig. 1; Tanchak et al., 1988; Dettmer et al., 2006; Viotti et al., 2010). The identity of the plant TGN as a distinct organelle and not just a tubular reticulum on the trans-side of the Golgi has been supported by results from a number of experimental approaches in recent years. Cell fractionation and electron microscopy/tomography studies allowed Kang et al. (2011) to propose a model for TGN biogenesis in which the trans-most Golgi cisterna is first transformed into a TGN-type compartment that is still tightly associated with the trans-Golgi stack. Transformation of a trans-most Golgi cisterna into a TGN cisterna is accompanied by cisternae peeling, proliferation of round, secretory-type vesicle (SV) buds, and a reduction of the cisternal membrane area (Kang et al., 2011). These authors elegantly showed that there are two forms of TGN: (1) the Golgi-associated TGN (GA-TGN) cisternae attached to the trans-side of the Golgi; and (2) the detached, free TGN cisternae. The amounts of associated clathrin-coated vesicles (CCVs) and SV buds distinguished the two fractions. Both CCVs and SV buds were more abundant in the free TGN cisternae (Kang et al., 2011).
Superresolution live imaging of the TGN-localized soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein SYP43 was used recently to characterize the dynamic behavior of the GA-TGN and Golgi-independent TGN (GI-TGN) populations (Uemura et al., 2014), corroborating the Golgi cisternal maturation model (Kang et al., 2011). The GA-TGNs localized on the trans-side of the Golgi apparatus, while the GI-TGNs (Golgi-released independent TGNs) were located away from the Golgi apparatus and behaved independently (Uemura et al., 2014). The authors proposed that segregation of GA-TGNs gives rise to the GI-TGNs with a core of the GA-TGN remaining after the process.
It is plausible that GA-TGN represents a population characterized by the TGN-localized COV1 and RAB GTPase-interacting YIP4A/B proteins, since both yip4a/b and cov1 mutants affect the TGN association with Golgi (Gendre et al., 2013; Shirakawa et al., 2014). Further characterization of these proteins could reveal important, unknown elements regarding the functional connections between Golgi and GA-TGN, which appear bidirectional.
Brefeldin A (BFA) is a fungal inhibitor that causes aggregation of TGN, endosomal, and Golgi material in large intracellular bodies (Langhans et al., 2011). Regeneration of Golgi and TGN populations after the removal of BFA from treated cells proceeded independently in tobacco (Nicotiana tabacum) BY2 cells (Ito et al., 2017), supporting the notion that plant TGN is an independent organelle. How GI-TGN and GA-TGN functionally connect and how TGN functions feed back Golgi functions remain poorly understood. Notably, GI-TGNs were found highly abundant in the differentiation zone of the root, in contrast with the meristematic region (Uemura et al., 2014). Thus, it would be interesting to explore the likely reorganization of TGN during physiological responses, such as those evoking a more active secretory function.
The regulation of pH homeostasis within the secretory pathway is crucial (for review, see Schumacher, 2014). An increase of only 0.2 pH units markedly impairs terminal α(2,3)-sialylation of an N-glycosylated reporter protein and induces mislocalization of the corresponding sialyltransferase into the endosomal compartments of mammalian cells (Rivinoja et al., 2009). Genetically encoded pH sensors targeted to specific endomembrane organelles have been engineered in an effort to establish a pH map of the plant endomembrane system. A gradual acidification from pH 7.1 in the endoplasmic reticulum (ER) to pH 5.2 in the vacuole was observed recently, with the pH of TGN at 6.3 (Shen et al., 2013). However, lower TGN pH values of 5.6 and 6.1 have been reported by other groups (Martinière et al., 2013; Luo et al., 2015).
In eukaryotic cells, acidification of the various endomembrane compartments has been proposed to depend largely on the proton pump activity of highly conserved vacuole-type H+-ATPases (V-ATPases; Hager and Helmle, 1981; Nishi and Forgac, 2002; Dettmer et al., 2006). V-ATPases are organized in two multisubunit domains. The peripheral V1 domain, responsible for ATP hydrolysis, is composed of eight different subunits (A–H), while the integral V0 domain forms the proton pore and includes a, c, c′, c′′, d, and e subunits (Nishi and Forgac, 2002; Toei et al., 2010). In plant cells, interfering with the TGN-resident VHA-a1 isoform causes the swelling of Golgi cisternae and inhibits cell expansion and hypocotyl growth (Dettmer et al., 2006; Brüx et al., 2008). Mutants with reduced activity of the cytosolic VHA-c subunit alter the steady-state pH in the TGN/EE, leading to reduced motility of both Golgi and TGN and defects in secretion of the brassinosteroid receptor BRI1 and Cellulose Synthase (Luo et al., 2015). Curiously, the TGN/EE-localized V-ATPase seems to contribute via an undefined mechanism to vacuolar pH (Kriegel et al., 2015). Such results suggest a rather complex interplay between different V-ATPase isoforms in the regulation of pH within each compartment of the plant endomembrane system.
Luminal pH initially determined by the activity of H+ pumps is thought to be fine-tuned by alkalization mechanisms (Orlowski and Grinstein, 2011; Bassil and Blumwald, 2014). Exchange of luminal H+ for Na+ or K+ by NHX antiporters counteracts the acidity generated by the H+ pumps (proton leaks; Orlowski and Grinstein, 2011; Reguera et al., 2015). Two Arabidopsis (Arabidopsis thaliana) NHX isoforms (NHX5 and NHX6) are expressed at the TGN, where they colocalize with the H+ pump VHA-a1 and likely counteract its activity (Bassil et al., 2011, 2012). nhx5 nhx6 double mutants have a more acidic TGN lumen, altered trafficking to the vacuole, and are hypersensitive to salt (Bassil et al., 2012). The latter highlights the relevance of luminal pH in the orchestration of crucial adaptive responses. The transporter AtCLC-d is yet another player thought to adjust TGN's pH by mediating the transport of a counter anion such as Cl− or NO3− into the TGN lumen. As NHX5/6, it colocalizes with VHA-a1 at the TGN, and the deleterious effect of inhibiting VHA-a1 was exaggerated in the clcd-1 mutant, indicating synergistic activities between the two transporters (von der Fecht-Bartenbach et al., 2007).
A role for lipid rafts in the regulation of post-Golgi sorting events has received extensive attention. Lipid rafts are nanoassemblies of specific proteins and lipids that define highly dynamic membrane microdomains and influence the spatiotemporal organization of protein complexes, thereby allowing the regulation of cellular processes (Simons and Sampaio, 2011; Cacas et al., 2012; Malinsky et al., 2013). Rafts might sort proteins and lipids by clustering them together at the TGN into bigger patches, which would then pinch off as vesicles for delivery to the plasma membrane (Schuck and Simons, 2004). Using a shotgun lipidomics approach, Klemm and coworkers first demonstrated that yeast TGN selectively sorts ergosterol and sphingolipid-enriched SVs transporting plasma membrane cargo (Klemm et al., 2009; Surma et al., 2011). C-Laurdan spectrophotometry measurements revealed a higher membrane order in the immunoisolated vesicles, compared with TGN, supporting the hypothesis that lipid rafts play a role in the TGN sorting machinery (Klemm et al., 2009; Surma et al., 2011).
To our knowledge, lipid rafts have not been observed in the plant TGN, but evidence for lipid-based TGN sorting was obtained recently in Arabidopsis, where sphingolipids with α-hydroxylated acyl chains of at least 24 carbon atoms were enriched in SV subdomains of the TGN. Importantly, the authors established a novel link between α-hydroxylated acyl chain enrichment of TGN membranes and secretory trafficking, specifically in polarized transport to the apical membrane of epidermal cells (Wattelet-Boyer et al., 2016).
RAB GTPases
As a trafficking hub, one may conceive the TGN as organized into distinct domains, facilitating cargo sorting into specific vesicles to be dispatched to the next destination. In such a scenario, it is tempting to assume that different RAB GTPases (for review, see Zhen and Stenmark, 2015), a major class of cellular proteins determining membrane identity and trafficking specificity, define separate TGN sorting compartments (Grosshans et al., 2006; Woollard and Moore, 2008; Hutagalung and Novick, 2011; Bhuin and Roy, 2014). Several lines of evidence support such RAB-determined subcompartmentalization. Two subclasses within the large A group of plant RABs, RABA2 and RABA3, identify a distinct TGN/EE membrane domain that functions in cargo delivery to the cell plate during cytokinesis (Chow et al., 2008). Whereas RABA2a defines TGN domains distinct from SYP61 vesicles, evidenced by differing lipid composition, RABA4b preferentially localizes to TGN SV sites together with SYP61, where it is involved in the transport of cell wall components (Preuss et al., 2004; Kang et al., 2011; Wattelet-Boyer et al., 2016; Jonsson et al., 2017). Another TGN domain harboring the secretory SNAREs VAMP721/722 and defined by RABA1 also has been suggested (Asaoka et al., 2013).
As to members of the plant RABB and RABD clades, they have been postulated to operate in traffic between the ER and Golgi (Cheung et al., 2002; Rutherford and Moore, 2002; Zheng et al., 2005; Woollard and Moore, 2008). However, fluorescently tagged RABD1 and RABD2a localized to punctate structures associated with TGN, and both RABD2a and RABD2b were found abundantly in isolated SYP61 vesicles that define a pathway en route from TGN to the plasma membrane (Pinheiro et al., 2009; Drakakaki et al., 2012).
SNAREs are integral membrane proteins required for the fusion of vesicles with their target membrane (for review, see Bombardier and Munson, 2015). SNAREs can be classified based on their site of function: v (vesicle)-SNAREs are localized to the vesicle membrane, and t (target)-SNAREs are localized to the target membrane (Rothman, 1994; Søgaard et al., 1994). Syntaxins are a family of SNAREs involved in recognizing and complexing with other SNAREs on the target membrane to create a t-SNARE complex. The genome of Arabidopsis encodes 24 syntaxins or Syntaxin of Plants (SYPs; Sanderfoot et al., 2000; Bassham and Blatt, 2008).
Distinct types of Arabidopsis syntaxins reside on the various membranes of the secretory pathway. The SYP4-type syntaxins (SYP41–SYP43) and their orthologs from yeast (Tlg2p) and mammals (Syntaxin16) have all been localized to TGN (Holthuis et al., 1998; Simonsen et al., 1998; Bassham et al., 2000). SYP4 members are implicated in the transport of secretory and vacuolar cargo from the TGN. Whether SYP4s function redundantly and localize to the same TGN compartment is to be determined (Bassham et al., 2000; Uemura et al., 2012; Kim and Bassham, 2013). TNO1 (TGN-localized SYP41-interacting protein1) is a TGN-localized coiled-coil protein that interacts with the SYP41 SNARE machinery. TNO1 mutants affect TGN dynamics, vacuolar cargo sorting, and the response to salt stress (Kim and Bassham, 2011).
The Arabidopsis SYP5 family of SNAREs consists of two members, SYP51 and SYP52, which localize to tonoplast and TGN (Sanderfoot et al., 2001; Carter et al., 2004; De Benedictis et al., 2013). Both SYP5s mediate traffic to the vacuole, although they seem to differ in cargo selectivity. Intriguingly, SYP5s appear to inhibit homotypic fusion (SNARE interfering or i-SNARE) when accumulated at the tonoplast. Such an i-SNARE function could provide a control for cargo delivery to the vacuole (De Benedictis et al., 2013).
The SYP6 group is encoded by a single gene, SYP61 (Sanderfoot et al., 2000). SYP61 is proposed to form separate complexes with the SNARE VTI12 and either SYP41 or SYP42 at the TGN (Sanderfoot et al., 2001); however, some lines of evidence suggest that other members of the SYP and VTI families also could take part in these complexes. In the absence of VTI12, prevacuolar compartment-localized VTI11 is able to interact with SYP4s, and proteoliposome fusion assays demonstrated that SYP42 and SYP43 can substitute for SYP41 while VTI11 can substitute for VTI12 in driving lipid mixing (Surpin et al., 2003; Kim and Bassham, 2013). Finally, the ability of SYP41 and SYP61 to independently mediate the fusion of liposomes was shown by Chen et al. (2005), who also identified YKT61 and YKT62, two functionally interchangeable components of the SNARE complexes, required for both SYP41- and SYP61-mediated vesicle fusion
A role for the SYP4/SYP61/VTI12 complex in the recycling of vacuolar sorting receptors to the TGN has been suggested. Vacuolar Protein Sorting45, an interactor of the complex, was proposed to positively regulate this function (Zouhar et al., 2009). The role of the SYP61 compartment in post-Golgi trafficking is discussed later in this review.
Tethering factors are traffic facilitators that function upstream of SNARE proteins in the establishment of an initial connection between an intracellular trafficking vesicle and its target membrane (Barlowe, 1997; Cao et al., 1998; for review, see Dubuke and Munson, 2016). Mechanistically, they have been suggested to mediate vesicle capturing, in virtue of their larger size compared with SNAREs, to accelerate the assembly of SNARE complexes and to provide checkpoints for SNARE specificity (Yu and Hughson, 2010). Plant tethers (for review, see Vukašinović and Žárský, 2016; Ravikumar et al., 2017) relevant to TGN-mediated trafficking include several members of the TRAPP family and the Golgi-associated retrograde protein (GARP) complex. TRAPPs assemble into fairly well-characterized multisubunit complexes in yeasts and mammals (Sacher et al., 2008; Kim et al., 2016). Our current knowledge of the function and organization of plant TRAPP homologs is extremely limited, although some lines of evidence have started to emerge with TGN-associated TRAPP involved in cytokinesis (Ravikumar et al., 2017). Several members of putative Arabidopsis TRAPP complexes were found in the proteome of TGN-associated SYP61 vesicles, suggesting a role for yet uncharacterized plant TRAPPs in the transport and/or delivery of secretory cargo to the plasma membrane (Drakakaki et al., 2012). The role of TGN-associated TRAPPs in cell plate formation and plant development (for review, see Vukašinović and Žárský, 2016; Ravikumar et al., 2017) is discussed at length in the cell plate section below.
In mammals, TGN-localized tetrameric GARP is required for retrograde trafficking from endosomes to the Golgi (Schmitt-John et al., 2005; Pérez-Victoria and Bonifacino, 2009). Mutants of Arabidopsis GARP subunits have implicated the complex in pollen tube elongation, acclimation to heat stress, and vacuolar targeting of the auxin carrier PIN1, necessary for PIN1 polar localization during the establishment of leaf vein patterning (Lobstein et al., 2004; Wang et al., 2011; Pahari et al., 2014). Interestingly, GARP shares three subunits with the Endosome-Associated Recycling Protein (EARP), which resides on recycling endosomes and is required for bringing recycling proteins/cargo to the cell surface (Schindler et al., 2015; Ravikumar et al., 2017). This phenomenon, also characteristic in TRAPP complexes, has been referred to as modularity. Multisubunit tethering complexes can exist in a variety of modular forms as a result of subunit exchange (Desfougères et al., 2015; Ravikumar et al., 2017). The existence of an Arabidopsis homolog also for the EARP-specific subunit, syndetin, hints at GARP modularity conservation in plants.
While the role of CCVs in plant endocytosis is well documented (Bandmann et al., 2012; Bashline et al., 2013; Di Rubbo et al., 2013), their involvement in post-Golgi trafficking remains poorly understood. Adaptor protein complexes (AP1–AP5) consisting of four adaptin subunits select cargo proteins into CCVs (Boehm and Bonifacino, 2001; Nakatsu and Ohno, 2003; Pertl-Obermeyer et al., 2016). The Arabidopsis genome encodes putative subunits of all five AP complexes (Happel et al., 2004; Hirst et al., 2011; Park et al., 2013). Arabidopsis AP1 localizes to the TGN (Park et al., 2013; Wang et al., 2014), and mutants of the AP1M2 subunit are affected in the secretory and vacuolar transport pathways. In addition, trafficking of the essential SNARE KNOLLE from the TGN to the cell plate is impaired in dividing cells of ap1m2 mutants (Park et al., 2013). More recently, TGN-localized AP1 γ-adaptins were found to mediate the targeting of membrane proteins with di-Leu motifs to the tonoplast (Wang et al., 2014). Intriguingly, Arabidopsis AP3 has been implicated in a TGN-bypass pathway that transfers cargo directly from Golgi cisternae to the vacuole and is facilitated by the HOPS-tethering complex (Feraru et al., 2010; Zwiewka et al., 2011; Feng et al., 2017). Perhaps with the exception of AP2, which operates in clathrin-mediated endocytosis at the plasma membrane (Krauss et al., 2006; Di Rubbo et al., 2013), a function in vacuolar transport appears to be the norm among Arabidopsis AP complexes. AP4 was recently found to localize to a TGN subdomain different from that of AP1 and for which mutants of four AP4 adaptins showed defects in the vacuolar sorting of the major storage protein 12S globulin (Fuji et al., 2016).
The Golgi apparatus and the TGN fulfill a highly dynamic and distinguishing function in plant cells: to sort and assemble plasma membrane cell wall biosynthetic enzymes, structural proteins, and the cross-linking glycans, pectin, and hemicellulose (Cosgrove, 2005; for review, see Worden et al., 2012; Kim and Brandizzi, 2016; van de Meene et al., 2017). Although extensive studies have led to the identification of the key enzymes involved in the biosynthesis of hemicellulose and pectin (for review, see Atmodjo et al., 2013; Pauly and Keegstra, 2016), comparatively little is known about their transport, deposition, and integration into the cell wall. Polysaccharides originate from distinct locations. Cellulose and callose are synthesized at the plasma membrane, while the Golgi apparatus is the synthesis site of noncellulosic cell wall polysaccharides, hemicellulose, and pectin, which are then transported through the secretory pathway to the apoplast (Driouich et al., 2012; Worden et al., 2012; Kim and Brandizzi, 2016). Our current knowledge of polysaccharide transport in the endomembrane system is derived mainly from immunohistochemical electron microscopy studies (Lynch and Staehelin, 1992; Zhang and Staehelin, 1992).
Golgi and TGN Involvement in the Biosynthesis of Xyloglucans and Pectins
The use of antibodies that recognize a number of xyloglucan (XyG) polymer epitopes identified trans-Golgi cisternae as the exclusive site of synthesis of the XyG backbone in a study conducted in suspension-cultured sycamore (Acer pseudoplatanus) cells. Fucosylated XyG side chains were detected in the trans-cisternae and the TGN, forming the hypothesis of an ordered mechanism of backbone biosynthesis and subsequent substitution in Golgi subcompartments (Zhang and Staehelin, 1992). The TGN involvement in XyG biosynthesis was highlighted further in studies with Arabidopsis roots, which showed labeling of TGN by a fucosylated XyG antibody (Kang et al., 2011). Earlier observations in tobacco BY-2 cells reported a sequence of events during XyG biosynthesis in cis- and medial Golgi cisternae (Chevalier et al., 2010), suggesting differences between species.
The Golgi apparatus also is the site of pectin biosynthesis. Pectins are synthesized and secreted in their methyl esterified form into the cell wall, where they are deesterified by pectin methyl esterases (Caffall and Mohnen, 2009). Studies in sycamore suspension cells showed that the synthesis of the nonesterified Rhamnogalacturonan I (RGI)/Homogalacturonan backbone and the methylesterification of GalUA residues take place in the cis-medial Golgi with subsequent completion in the medial cisternae. More complex pectin oligosaccharides, such as RGI-containing arabinose side chains, are detected only in the TGN (Zhang and Staehelin, 1992). However, the labeling of pectin epitopes differs among cell types of the root tip, suggesting that Golgi function may be altered/reorganized during cell differentiation (Lynch and Staehelin, 1992).
Several enzymes involved in both pectin and XyG biosynthesis have been identified and characterized. Contrasting the pectin biosynthesis assembly line hypothesis is the observation that several glycosyl transferases involved in the biosynthesis of pectin are present in multiprotein complexes at the Golgi. Currently, two pectin biosynthesis models have been proposed: the classical model, which predicts the consecutive addition of sugar residues to the growing polysaccharide; and a recently developed model, in which a block transfer of pectin from one domain onto another domain occurs (Atmodjo et al., 2013). The isolation of TGN SVs that carry polysaccharide cargo will contribute substantially to a more comprehensive understanding of the transport and assembly of cell wall components.
Cellular Determinants of Cell Wall Component Transport
Post-Golgi routes deployed by the plant cell to transport Golgi/TGN-synthesized polysaccharides to the plasma membrane are not fully elucidated. A TGN-resident complex formed by ECHIDNA (ECH) and two members of the YIP family of RAB GTPase-interacting proteins has been implicated in the secretion of pectin and XyG. Mutant plants of YIP4A and YIP4B are defective in cell elongation and the secretion of XyG and RGI pectins, a defect also observed in ech mutants (Gendre et al., 2011, 2013). ECH also seems to be necessary for the proper TGN localization of the ADP-Ribosylation Factor1 (ARF1)-GTPase and the ARF-guanine-exchange factor BIG4. This pathway mediates secretion of the auxin carrier AUX1 from the TGN to the plasma membrane (PM) during hook development (Jonsson et al., 2017). In tobacco BY-2 cells, immunostaining of pectin showed that the Secretory Carrier Membrane Protein2 (SCAMP2) vesicles are involved in pectin transport (Toyooka et al., 2009). A potential interplay between the SCAMP2 and ECH machineries in pectin secretion has not been explored yet.
A pioneering study of plant intracellular vesicle isolation and proteomic analysis revealed that the syntaxin SYP61 defines a TGN compartment carrying cell wall-relevant cargo. Several CSCs were identified in the SYP61 proteome, and electron microscopy immunostaining evidenced the colocalization of CESA6 and SYP61 in TGN vesicles and in close proximity to the PM (Drakakaki et al., 2012). The exogenous administration of CESTRIN, a small molecule that reduces the motility of CSCs at the plasma membrane, increases the association of CESAs with SYP61 vesicles, further implicating the SYP61 compartment in CESA trafficking (Worden et al., 2015). Both ECH and YIP proteins are cargo of SYP61 vesicles, favoring the hypothesis that SYP61-mediated trafficking is involved in the transport of cell wall components. Furthermore, antibodies for fucosylated XyG label a RABA4b TGN compartment, and colocalization of RABA4b and SYP61 was observed, supporting the role of SYP61 in the trafficking of cell wall components (Kang et al., 2011).
Despite the essential roles of glycoproteins in cell wall remodeling, development, and responses to biotic stress, their secretory routes remain poorly characterized. In addition to conventional secretion via TGN, exemplified by SYP61 and SCAMP2 vesicles, evidence pointing to the existence of TGN-independent, unconventional protein secretion pathways is accumulating, which may be implicated in cell wall protein transport (for review, see De Marchis et al., 2013; Davis et al., 2016a; Robinson et al., 2016; van de Meene et al., 2017).
The TGN's key role in organizing and shipping cell wall material is never more obvious than during cytokinesis, in which an entirely new cell wall is built from inside the parent cell. Cytokinesis begins with the delivery of Golgi/TGN-derived vesicles to the plane of division, where they immediately start to fuse and tubulate. The vesicles continue to coalesce into a membrane network (the cell plate) following the tracks laid down by the radially expanding phragmoplast (Samuels et al., 1995; Seguí-Simarro et al., 2004; Smertenko et al., 2017). Deposition of polysaccharides leads to the maturation of the formed structure, which finally joins to the parental cell wall (Drakakaki, 2015).
Chemical inhibition of either endocytosis or secretion in dividing plant cells indicates that both pathways have a role during cell plate formation via the TGN. It is plausible that endocytosed proteins merge into the late secretory pathway and, thus, are delivered to the division plane instead of being recycled to the PM (Dhonukshe et al., 2006; Reichardt et al., 2007; Richter et al., 2014; Müller and Jürgens, 2016).
The TGN is intimately connected to the cell plate, as evidenced by the various proteins shared and shuttled between them, of which a few key components are mentioned below (Fig. 2). Excellent reviews describe in detail the cytoskeletal and membrane dynamics during cytokinesis (McMichael and Bednarek, 2013; Boruc and Van Damme, 2015; Müller and Jürgens, 2016).
A key event in cell plate formation is vesicle fusion mediated by SNARE complexes (El Kasmi et al., 2013), with the complex formed by the Q-SNARE KNOLLE and the R-SNAREs Vesicle-Associated Membrane Protein721 (VAMP721) or VAMP722 playing a preponderant role (Lauber et al., 1997; Zhang et al., 2011). Additional proteins in this complex include the SEC1/Munc18 protein KEULLE, SNAP33, and NPSN11 (Assaad et al., 2001; Heese et al., 2001; Zheng et al., 2002). Interestingly, of the SNAREs involved in cell plate formation, only KNOLLE is specific to the process.
RABA2 and RABA3 preferentially localize to the leading edge of the cell plate, suggesting a function in the delivery and incorporation of new membrane material to the cell plate (Chow et al., 2008). Because overlapping localizations and complementary functions are common among RABAs, a complete roadmap for the multiple pathways and processes in which each is involved is still being drawn. RABA2a and RABA1e vesicles display a different spatiotemporal pattern during cytokinesis, which is exaggerated by the cytokinesis inhibitor Endosidin7 (ES7; Davis et al., 2016b). TGN/EE-resident RABA1d accumulates at early cell plate stages during cytokinesis, and its association with PM proteins suggests a role in the recycling of material from the PM to the growing plate (Berson et al., 2014). Given the fact that the RABA clade is highly elaborated in plants (26 of the 57 total RAB GTPases identified thus far; Woollard and Moore, 2008), it will be interesting to see how/if they are functionally connected to meet the myriad trafficking demands in a plant cell, dividing or otherwise. In addition to RABAs, RABE1s also localize to the cell plate, where they interact with the Stomatal Cytokinesis-Defective (SCD) complex. Inhibition of RABE1 causes cytokinesis defects, emphasizing its role in trafficking to the cell plate (Speth et al., 2009; Ahn et al., 2013; Mayers et al., 2017).
TRAPP tethering complexes are known to act as guanine exchange factors (GEFs) and, thus, activate RAB GTPases (Jones et al., 2000; Wang et al., 2000; Pinar et al., 2015). Colocalization at the TGN and functional studies suggest that TRAPPII acts as a GEF for RABA1c, facilitating RABA1c-mediated trafficking of material from the TGN to the cell plate (Qi et al., 2011). Mutants of the TRAPPII subunit, AtTRS130, exhibit severe cytokinesis defects and irregular aggregations of RABA1c around cell plate-like structures (Jaber et al., 2010; Qi et al., 2011). The expression of constitutively active RABA1c partially rescues attrs130 (Qi et al., 2011), supporting the role of AtTRS130 upstream of RABA1c. During cytokinesis, subunits of the exocyst-tethering complex interact with TRAPP subunits, suggesting synergistic activities of the two complexes (Rybak et al., 2014; for review, see Vukašinović and Žárský, 2016).
In addition to RAB GTPases and their regulators, other protein classes with vesicle formation/budding functions at the TGN participate in cell plate formation. For example, Clathrin Light Chain, Dynamin-Related Proteins, Epsin-like adaptors, and the adaptin-like T-PLATE all have been identified at the cell plate, providing evidence for clathrin-mediated endocytosis in the removal and/or recycling of excess membranes from the cell plate (Fujimoto et al., 2008; Konopka and Bednarek, 2008; Van Damme et al., 2011; Song et al., 2012; McMichael and Bednarek, 2013). Additionally, SCD1 and SCD2 act as a complex to mediate post-Golgi trafficking to the plasma membrane and the cell plate. Multiple lines of evidence point to the interaction between the SCD complex, the exocyst, and RABE1 GTPases in the trafficking of material to the cell plate (Mayers et al., 2017).
The aforementioned AP1, and specifically the µ-subunit (AP1M2), is found in the TGN and is essential for the trafficking of KNOLLE to the cell plate. In an ap1m2 mutant background, KNOLLE is mislocalized around the division plane and cell wall stubs are present, indicating a role of AP1M2 in targeting KNOLLE to the cell plate (Park et al., 2013; Teh et al., 2013).
Exciting discoveries in the nature and composition of cytokinesis vesicles are awaiting, which will form a better picture of membrane and polysaccharide delivery for the buildup of such an essential and dynamic structure.
The plant TGN is unique in that it acts not only in the secretory but also in the endocytic pathway as an EE (Tanchak et al., 1988; Dettmer et al., 2006; Viotti et al., 2010). This distinct, additional function of plant TGN has been associated with the absence of the tubulated endosomes dedicated to recycling, normally found in animal cells (Paez Valencia et al., 2016). The function of TGN as an EE has been demonstrated by live imaging and electron microscopy studies (Dettmer et al., 2006; Lam et al., 2007; Viotti et al., 2010). Immunogold electron microscopy showed an accumulation of the PM-localized brassinosteroid receptor BRI1 to BFA bodies, not prevented by the protein synthesis inhibitor cycloheximide, lending evidence to the exclusively endocytic origin of the material accumulated in the BFA compartment (Viotti et al., 2010).
Recycling of PM proteins through the TGN/EE has proven crucial for the polar distribution of plasma membrane proteins, including auxin carriers like PIN1 (Geldner et al., 2003; Kleine-Vehn et al., 2008, 2011; Luschnig and Vert, 2014). Endosomal recycling in plants largely depends on the activity of small GTPases of the ARF family and their associated GTPase-activating proteins and GEFs (Casanova, 2007; Kleine-Vehn et al., 2008; Paez Valencia et al., 2016). Twelve ARF genes are present in the genome of Arabidopsis (Robinson et al., 2007). The best-characterized ARF1 localizes to endocytic organelles and is implicated in the establishment of apical-basal polarity in epidermal cells (Xu and Scheres, 2005).
Among Arabidopsis ARF-GEFs, GNOM has been arguably the most studied in its function of restricting PM protein localization to basal domains (Geldner et al., 2003). BFA treatments inhibit GNOM, causing the accumulation of the internalized auxin effluxer PIN1 in aggregating GNOM-positive intracellular compartments (Geldner et al., 2003). However, superresolution/electron microscopy studies showed its exclusive localization to Golgi cisternae, thereby challenging its function solely in protein recycling (Naramoto et al., 2014). In support, GNOM was recently implicated in the ER-to-Golgi transport of PIN1 (Doyle et al., 2015). BEN1 is another, BFA-insensitive ARF-GEF that localizes to early endocytic compartments distinct from GNOM-positive endosomes and whose mutants display cell polarity defects (Tanaka et al., 2009). Finally, a role for the BIG subfamily in nonbasal trafficking of plasma membrane proteins was recently supported by pharmacological evidence (Li et al., 2017). Such functions place BIGs distinct from GNOM in the scheme of factors regulating the polarity of PM proteins. Interestingly, and in contrast with GNOM, BIGs do not seem to affect recycling but only protein secretion (Richter et al., 2014). Such functional divergence could lie at the core of the observed stage-dependent, mutually exclusive involvement of GNOM and BIGs in the regulation of the secretion of AUX1 influx carrier to the plasma membrane from the TGN during hook development (Jonsson et al., 2017).
Several lines of evidence support a model for the maturation of TGN/EE into late endosomes or multivesicular bodies (LE/MVB; Scheuring et al., 2011; Singh et al., 2014). Endocytosed plasma membrane proteins that are not recycled back are transferred from EE (TGN) to LE/MVB, where they are internalized into the intraluminal vesicles of the LE. Fusion of the LE with the lysosome/vacuole releases the vesicle cargoes, leading to their degradation. Scheuring et al. (2011) showed that the formation of intraluminal vesicles takes place already at the TGN/EE, while Singh et al. (2014) suggested that endosomal maturation in Arabidopsis originates in a subdomain of the TGN/EE that recruits Rab5-like ARA7 and subsequently transitions into an MVB. The topics of plant EE and LE are covered extensively by other reviews in this issue of Plant Physiology.
The plant TGN comprises an extremely dynamic and diverse vesicle population, which raises many questions for most of which we do not have an answer; meanwhile, technological advances are helping us on the road. Improved protocols for the immunoisolation of vesicles based on specific, vesicle membrane markers have been successfully established, an example of which is the characterization of the SYP61 compartment (Drakakaki et al., 2012; Groen et al., 2014; Heard et al., 2015). Organelle proteomics is allowing the identification of vesicle protein cargoes, and extending glycomic analysis to isolated vesicles will help us to better characterize the secretory routes of cell wall polysaccharides (Obel et al., 2009; Pattathil et al., 2010; Drakakaki et al., 2012; Parsons et al., 2013; Heard et al., 2015; Kračun et al., 2017; Wood et al., 2017). Chemical genomics is enabling the characterization of vesicle-trafficking pathways, recently evidenced by the use of the small molecules ES7 and ES16 to discern the contributions of two RAB GTPases to cell plate formation and to demonstrate the specificity of a trafficking pathway involved in cell polarity, respectively (Davis et al., 2016b; Li et al., 2017). Spatiotemporal image correlation spectroscopy has proven useful to characterize the dynamics of vesicle trafficking to the cell plate (Hebert et al., 2005; van Oostende-Triplet et al., 2017). Finally, lipid profiling, metabolic click labeling, and the use of oligosaccharide-based probes for high-resolution real-time imaging of glycans have recently added to the list of promising avenues to dissect the role of TGN in cargo transport (Anderson et al., 2012; Pattathil et al., 2012; Mravec et al., 2014; Wattelet-Boyer et al., 2016). Such a panoply of tools promises new exciting discoveries about the multiple functions of the plant TGN.
We apologize to our colleagues whose work we were not able to include in this review due to length limitations.
G.D., M.R.R., and D.J.D. wrote the article.
↵1 This work was supported by NSF IOS 1258135 to G.D.
↵[OPEN] Articles can be viewed without a subscription.
- Received August 31, 2017.
- Accepted November 27, 2017.
- Published November 30, 2017.


text:
IBM Working with Google, Lyft on Microservices Management
IBM is partnering with Google and Lyft on developing microservice management infrastructure for applications built on top of Kubernetes. At the same time, IBM is working on extending a framework for managing clusters in a way that can be used across multiple types of, for example, Kubernetes and Hadoop clusters.
Shriram Rajagopalan, a staff member within IBM Research, says the work involving Google and Lyft is intended to significantly simplify tasks such as A/B testing, resilience testing, monitoring and policy enforcement using design principles gleaned from years of operational container experience at Google, IBM and Lyft.
A core element of this effort involves Project Gremlin, a systematic approach to resilience testing that IBM has been developing. Gremlin is designed to enable IT organizations to independently test the failure-recovery logic embedded within a microservice regardless of the programming language or business logic involved. Gremlin intercepts application programming calls being made by the microservice to insert a fake failure created using Python-based templates, which enable the IT organization to observe how that microservice responds. If the microservice hangs, then it becomes apparent that the IT organization needs to rework the failure-recovery logic embedded in the microservice. Rajagoplan says Gremlin is not a replacement for other approaches to testing microservices resiliency. Rather, IBM views it as another tool to be deployed as complement to, for example, a Chaos Monkey test.
Given IBM's historic focus on IT management software, the work being done around Kubernetes should not come as a surprise. But IBM views Kubernetes as only one type of cluster that needs to be managed using a comprehensive software-defined framework. IBM recently unveiled a Spectrum Computing portfolio of software that makes it easier to manage multiple types of applications running on the same cluster.
Bernie Spang, vice president of Software-Defined Infrastructure for IBM, says that approach will serve as a new model for how IBM will more broadly approach the management of IT. Instead of thinking in terms of individual virtual and physical machines, IBM is developing a software-defined architecture that will make it possible for IT organizations to comprehensively manage multiple types of clusters, including Hadoop and Kubernetes, comprehensively alongside one another. Obviously, each class of cluster will have its own management framework. But as the application programming interfaces (APIs) surrounding those clusters become more open, it then becomes feasible to consider managing the entire IT environment at a higher level of abstraction, says Spang.
Obviously, the rise of containers and microservices represents a major new IT management challenge. IT organizations must find a means to manage not only lots of individual microservices when necessary, but also entire clusters as a single logical entity. At the same time, the IT management framework must be extensible enough to incorporate multiple types of container architectures and clusters as advances in these areas continue to evolve rapidly. From an enterprise IT perspective, it's still early days as far as containers and clusters are concerned. But in terms of IT management frameworks, the writing is already on the wall concerning the continued viability of existing approaches.

subset: commoncrawl


text: Editing National Democratic Party of Germany 0 You do not have permission to edit this page, for the following reasons: The action you have requested is limited to users in one of the groups: Users, Autoconfirmed users, FANDOM Staff, FANDOM Helpers, Wiki Managers, Content Team Members. Please log in to edit this wiki.Joining Logopedia is free, and it only takes a minute.We hope that you sign in, and become a member of the community!Click here to log in or create an account You must confirm your e-mail address before editing pages. Please set and validate your e-mail address through your user preferences. more + You can view and copy the source of this page: [[File:800px-NPD-Logo-2013.svg.png|left|400px]] Return to National Democratic Party of Germany. Retrieved from "https://logos.fandom.com/wiki/National_Democratic_Party_of_Germany" Edit summary Preview Mobile Desktop Show changes


text:
ST. LOUIS -- Writers and readers – save the dates June 16-18 2017 for the first annual Gateway to Publishing Conference and Convention at the Renaissance Hotel near Lambert International Airport.
Hosted by the St. Louis Writers Guild, this is a unique opportunity for beginning and seasoned writers to hone their craft and advance their writing to the next level. Established novelists can also sell their books at the book fair, which is free to the public, and meet with their fans at other reader-focused events.
Register for Gateway to Publishing and get access to all the workshops and writers' panels, as well as:
* Dinner on Saturday night
* Networking events
* Two pitch sessions with the literary agent, editor, or publisher of your choice (so long as spaces are available).
A few of the outstanding faculty members include:
* New York Times Bestselling and award-winning author Eileen Dreyer (known as Kathleen Korbel to her Silhouette readers!)
* Stephanie Hansen, Literary Agent at Metamorphosis Literary Agency
* Jennifer Geist, publisher at Pen & Publish, Inc. and its imprints: Brick Mantel Books, Open Books Press, and Transformation Media Books
* New York Times Bestselling Author Angie Fox
* Cynthia Graham, the award-winning Brick Mantel Books author
* Ann Leckie, Hugo, Nebula, and Arthur C. Clarke Award-winning author
Literary organization workshops at Gateway to Publishing include:
* St. Louis Press Club
* St. Louis Publishers Association
* Missouri Chapter of Romance Writers of America
* Greater St. Louis Chapter of Sisters in Crime
Cost: General Registration: $250, SLWG Members: $210, Students/Affiliated Organizations: $230, Gateway Con Masterclass: $50 per class
Visit www.stlwritersguild.org/gatewaycon.html to register and for more information.


text:
Maja Pawinska Sims 29 May 2019 // 8:00AM GMT
LONDON — Zeno Group's managing director of Europe, Steve Earl, is leaving the firm after six years to become London managing director of APCO Worldwide.
He replaces James Acheson-Gray, who left APCO at the end of last year and is one of the founding partners of new corporate consultancy Apella.
Earl founded Zeno's operations in Europe and played a leading role in the acquisition of 3 Monkeys to create 3 Monkeys Zeno in January 2016. He starts work for APCO on 1 July, reporting to Claire Boussagol, president of APCO in Europe.
He started his career as a regional news reporter before joining the agency world, including a stint at Weber Shandwick, and then founding Rainier PR with Stephen Waddington (now MD of Metia) in 1998.
The business was sold to marketing agency Loewy in 2006 and the group's operations were merged in 2009 as Speed Communications, with Earl and Waddington as joint managing directors. They both left Speed in 2012, with Earl going to Zeno and Waddington joining Ketchum.
Earl told the Holmes Report: "Zeno was a massive ride but it's time to take on a different type of challenge. APCO being independent was a big consideration for me as it allows you to do things you can't do in a network – it was irresistible."
He said he would initially be focused on accelerating growth, and making the UK a bigger component of the global APCO network: "There are great people and great clients, but it can do even more: it's not yet punching at the weight it needs to. It also needs to be a more prominent player in the London market, and to be more assertive about getting more UK-based companies on the roster."
Boussagol added: "I am delighted to welcome Steve to lead APCO's London office during such a critical period for the UK, Europe and the global marketplace, especially given the projected instability and uncertainty in a post-Brexit world. Steve and the excellent existing team, currently led by joint deputy MDs Alex Bigland and Polly Kennedy, are ideally placed to provide counsel and expertise to help clients understand how to succeed in the market during these times of extreme complexity."
Earl joins the firm during APCO's 35th anniversary since it was founded by executive chairman Margery Kraus. The agency is still independent, with nearly 800 employees in more than 30 markets.
At Zeno, CEO Barby Siegel told the Holmes Report: "Steve has been a driving force in helping Zeno to establish and grow our presence in Europe. We thank Steve for all that he has done, and wish him the very best as he begins his next chapter."
She said Zeno did not discuss specific talent or hiring plans, but added: "We have a strong senior team in place that will continue to move our business forward."


text:
This wine comes from the grapes of the Heredad de Vistahermosa estate.
It is one of the largest vineyards in Rioja, with 163 Ha, located between 550 m. and 750 m. altitude, which makes it also one of the highest.
The vineyard is located in a privileged and historic area in Rioja for the Garnacha variety. The altitude, the climate, which is colder than expected in this area, and the peculiar soil, with a large number of pebbles and low pH, are perfect for growing this variety, which is 46 years old.
That's why, in 2005, it was decided to try and make a wine that was 100% Garnacha, something quite rare and peculiar in Rioja, where 90% of the wines are Tempranillo. The result was so good that it was bottled under the brand Conversa.
The production of this year was very limited, about 2,000 bottles, "from 2,006 the production is about 8,000 bottles" although we do not Conversa every year.
The grape is selected in a very particular way, practically bunch by bunch among the best vines of the plots chosen each year and it is harvested by hand in boxes of 15 Kg.
It has been aged for 12 months in oak barrels, which gives it an indication of Reserva.


text:
COVENTRY and Warwickshire has one of the worst postal services in England and Wales.
About a fifth of all first-class mail across the CV postcode area is not being delivered to households and businesses on time.
Latest performance figures from Royal Mail show that just 75.6 per cent of first-class post reached its destination the next day between October and December 2007.
This brought the figure so far for the year - which will end this April - to 80.1 per cent of first-class post arriving on time.
Royal Mail blamed the poor performance on a string of national and local disputes, including a series of one-day strikes in Coventry against the proposed move of the sorting office in Bishop Street to Northampton and the closure of the crown post office in Hertford Street.
It means residents and traders in the CV postcode area are getting the eighth worst service in England and Wales out of 111 postcode areas.
The performance is level with London East, closely behind London South and North, Chelmsford, Colchester, Oxford, Liverpool and Stoke-on-Trent.
Najma Hafeez, chairman of Postwatch Midlands, said: "Even after the strike was over the company failed to meet customer expectations at Christmas.
"This is the busiest posting time for households and with more and more deliveries from internet shopping, an opportunity to rebuild a tarnished brand was squandered."
Royal Mail is likely to miss 10 of its 12 annual licence targets when the year ends - the worst annual performance since 2003/04, when it missed all its licence targets.
Disgruntled Coventry resident Katie Rowlatt, 36, of St Lawrence's Road, Foleshill, said: "The postal service has not been great over the past few months. We have had problems with bills arriving late."
A spokesman for Royal Mail said: "With the strike behind us and a wide-ranging agreement on modernisation in place, we are now focused on delivering once again a consistent, high quality of service to all our customers. Delivering the best possible service is Royal Mail's number one priority."


text:
City to grant no more State capital for construction projects this year
City to grant no more State capital for construction projects this year
HCM City People�s Committee will not adjust or grant additional State
capital for construction and investment projects in the remaining months of this
year in order to stabilize the macro economy and ensure social welfare and
sustainable development.
Instead, the city will allocate capital for projects and fields targeted to
control inflation and price rises, and store necessary consumer goods to serve
people in the last six months and the coming occassion of Tet (lunar new year).
The city government assigned relevant agencies and district
authorities to accelerate the implementation of projects that were allocated capital
earlier and ensure their quality, especially key projects which affect many
households and the city�s environment.
The city government also asked its agencies, district authorities
and State companies to reduce their public spending by 10%, increase savings and
limit overseas trips for study and experience-learning purposes.
(Translated by MT � July 17, 2008)


text: Giovanni Sollima was born in Palermo into a family of musicians. Still a teenager, he embarked on a brilliant international career of cellist but, at the same time, his creative curiosity led him to explore the field of composition. His unmistakable style is characterized by contaminations between different genres: minimalism, rock, electronic and ethnic music, with echoes of ancient and baroque music. His music has been played by most renowned classical performers. As a soloist, or with different instrumental groups, he performed his compositions all over the world. Remarkable is the cello performance in the Sahara Desert and the one underwater in a sicilian' gebbia (a tank for irrigation' water). In 2012 he had been the main creator and the artistic director of the explosive musical ensemble of the "100 Cellos". Giovanni Sollima plays a cello Francesco Ruggeri (Cremona, 1679). Moreover he uses, in his creations, western and eastern acoustic instruments and electrical and electronic tools, mixed with others of his own invention, such as the aquilarco, and others especially made for him, like the ice-cello that in Winter 2007 he played at 3200 meters above sea level, in an igloo' theatre built in a glacier of the Dolomites.


text:
Plant Pot Stand, 2 Shelves
Product Code:
£109.99 £91.66
Out of stock
Present your ornamental flowers, plants or herbs in an arrangement to do them justice with the help of these tiered pot stands.
The solid steel construction means these pedestals are extremely sturdy and weather resistant, making them suitable for both indoor and outdoor use. Perfect for the greenhouse or the patio.
Designed and manufactured in the UK
NB: This product is supplied direct from the supplier. Please allow up to 14 days for delivery.
H: 77.5cm x W: 82.5cm x D: 43cm


text:
The erectile dysfunction medications are those drugs that are used to treat this condition. Either you've got this health condition as a result of anabolic steroid use (which is a known side effect) either you've got it as a result of natural occurrences, the erectile dysfunction treating medicines are amazing for such needs. When a man is suffering from erectile dysfunction it can affect him both physically and psychologically, that's why is so important to treat this condition. Read more
The medications that are helping with erectile dysfunction treatment are also known as PDE 5 inhibitors. There may be others, but by far the most famous ones are PDE 5 inhibitor types of drugs and the most famous PDE5 inhibitor medications treating erectile dysfunction include: Viagra containing Sildenafil Citrate, Cialis containing Tadalafil and Levitra containing Vardenafil.
Yet, there are many other types of medications too. Plus, if talking about other sexual health drugs then there are other types too. Nonetheless, the most famous sexual health drugs are erectile dysfunction ones and below you can find the most famous drugs.
The erectile dysfunction drugs are coming in various different methods of administration. Most common and widely used ED drugs are used orally by mouth. But they can come as tablets, capsules, pills, sublingual compounds, oral jelly and many others.
Erectile Dysfunction is by far one of the most embarrassing condition for a man, so it must be treated, and the best way to treat the condition is to choose one of the products which you can find in the list below as these are the best and safest medications for treating this health condition.
Regardless if you've received it naturally or because of using anabolic steroids. Depending on what exactly you want, you may choose one medication, which is better for your needs.
Among those medications, you would find the genuine products only, which are best in terms of quality vs price ratio. Viagra for sale, Cialis for sale, Levitra for sale too and you can be sure the quality is the highest you can find anywhere online.
- Sale!
- Erectile Dysfunction, General Pharmacy
- Sildenafil
- Sildenafil is the active substance found in the extremely famous medication – Viagra. The full name of the substance is Sildenafil Citrate and Viagra is just the brand name for this medicine. However, there are a lot of different generics for Viagra, also containing high quality Sildenafil Citrate offered in many different names.
- Buy product
- Sale!
- Erectile Dysfunction, General Pharmacy
- Tadalafil
- Tadalafil is a medication that is most often being sold as the brand name Cialis but you may find Tadalafil being sold as a lot of other different generic names except for Cialis. Tadalafil is a medication that is mostly being used for treating erectile dysfunction.
- Buy product
- Sale!
- Erectile Dysfunction, General Pharmacy
- Vardenafil
- Vardenafil is the active substance of the brand name Levitra but it can be found being sold as various other generic names. Vardenafil is a medication that is being used for treating erectile dysfunction. This is a condition which is consisting of a man having issues to get and to maintain an erection.
- Add to cart


text:
TapImmune Announces Phase 2 Ovarian Cancer Trial Study with AstraZeneca/MedImmune and Sloan Kettering Cancer Institute
April 21, 2016
JACKSONVILLE, Florida, April 21, 2016 /PRNewswire/ –
Study to commence Q2 2016 at Sloan Kettering Institute
Cancer vaccine and checkpoint inhibitor combination to be evaluated in platinum-resistant ovarian cancer patients
TapImmune,Inc. (TPIV), a clinical-stage immunology-oncology company specializing in the development of innovative peptide and gene-based immunotherapeutics and vaccines for the treatment of cancer & metastatic disease, announced today plans to initiate a Phase 2 trial of its cancer vaccine, TPIV 200, a multi-epitope anti-folate receptor vaccine (FR�), in combination with Astra Zeneca (NYSE: AZN) durvalumab (MEDI4736), an anti-PD-L1 antibody, in patients with platinum-resistant ovarian cancer. The study will commence in the second quarter of 2016 at Memorial Sloan Kettering Cancer Center in New York and will be led by Jason Konner, M.D. as Principal Investigator.
The two Companies will share clinical costs, while TapImmune will supply TPIV 200 and MedImmune will supply Durvalumab (MED14736) for the trials. TapImmune recently obtained Orphan Drug Designation for TPIV 200 in ovarian cancer from the U.S. Food and Drug Administration.
This single arm Phase 2 trial will include 40 women with high-grade ovarian, tubal, or primary peritoneal carcinomas, who have progressed within 6 months of their most recent platinum chemotherapy. The primary objective of the study is to determine the effectiveness of the combination by measuring Overall Response Rate [ORR = Complete Response (CR) + Partial Response (PR)] by RECIST and Progression Free Survival (PFS) rate at 6 months. Secondary endpoints will be safety and immune and correlation of FR�-specific immune responses with clinical efficacy.
"This collaboration is a significant event for TapImmune," stated Dr. Glynn Wilson, Chairman and CEO of TapImmune. "We are delighted to bring a leading T-cell vaccine platform into this combination study and to work with AstraZeneca/Medimmune and Sloan Kettering in a patient population that is in dire need of an effective treatment."
"This study is part of a larger Phase 2 strategy for TPIV 200 that is designed to greatly increase our understanding of the vaccine while providing clinical evidence of efficacy," Dr. Wilson added.
TPIV 200 is a multi-epitope peptide vaccine that targets Folate Receptor Alpha, which is overexpressed in multiple cancers including over 90% of ovarian cancer cells. In Phase I clinical studies conducted at the Mayo Clinic in patients with breast and ovarian cancer, this vaccine was shown to be safe and well tolerated and to give robust cellular immune responses in 20 out of 21 evaluable patients.
About Durvalumab (MEDI4736)
Durvalumab is an investigational human monoclonal antibody directed against PD-L1 developed at MedImmune LLC. Signals from PD-L1 help tumors avoid detection by the immune system. Durvalumab blocks these signals, countering the tumor's immune-evading tactics. Durvalumab is being developed, alongside other immunotherapies, to empower the patient's immune system and attack the cancer. Durvalumab is being investigated in an extensive clinical trial program, as monotherapy or in combination with tremelimumab, in NSCLC, head and neck, gastric, pancreatic, bladder and blood cancers.
About Medimmune/AstraZeneca
MedImmune, LLC, is the global biologics research and development arm of AstraZeneca (NYSE: AZN) and is located in Gaithersburg, Md., one of AstraZeneca's three global R&D centers. For more information, please visit http://www.medimmune.com.
AstraZeneca is a global, innovation-driven biopharmaceutical business that focuses on the discovery, development and commercialization of prescription medicines, primarily for the treatment of diseases in three main therapy areas – respiratory, inflammation, autoimmune disease (RIA), cardiovascular and metabolic disease (CVMD) and oncology – as well as in infection and neuroscience. For more information please visit http://www.astrazeneca.com.
About TapImmune Inc.
TapImmune Inc. is an immune-oncology company specializing in the development of innovative technologies for the treatment of cancer, including metastasis, and infectious disease. The Company's peptide or nucleic acid-based immunotherapeutics, comprise one or multiple naturally processed epitopes (NPEs) designed to comprehensively stimulate a patients' killer T-cells, helper T-cells and to restore or further augment antigen presentation by using proprietary nucleic acid-based expression systems. The Company's technologies may be used as stand-alone medications or in combination with current treatment modalities. For more information please visit https://www.markertherapeutics.com.
Forward-Looking Statement Disclaimer
This release contains forward-looking information within the meaning of the Private Securities Litigation Reform Act of 1995. Statements in this news release concerning the Company's expectations, plans, business outlook or future performance, and any other statements concerning assumptions made or expectations as to any future events, conditions, performance or other matters, are "forward-looking statements". Forward-looking statements are by their nature subject to risks, uncertainties and other factors which could cause actual results to differ materially from those stored in such statements. Such risks, uncertainties and factors include, but are not limited to the risks set forth in the Company's most recent Form 10-K, 10-Q and other SEC filings which are available through EDGAR at http://www.sec.gov. The Company assumes no obligation to update the forward-looking statements.
TapImmune Inc.,
Glynn Wilson, Ph.D.
Chairman & CEO
SOURCE TapImmune Inc.


text:
(Recorded 01/21/2018)
Author: Kim Clements
Ep 15: Navigating The Blend
(Recorded 9/10/2017)
Ep 14: Rahab's Story
(Recorded 07/09/2017)
Middle School or Jr. High Whichever?!
Next week the daughter of one of my dearest heart friends is starting Middle School. I have had the privilege of walking along the road with my friend during infertility, the choice to adopt and finally the wonderful day when her beautiful China Doll came home forever. Continue reading
The Strong Willed Child (A Parent's Reflections Part II)
The musicians on stage were seated as they quietly played while the pastor was finishing his final thoughts for the day. All I could see were Mychal's hands as he played the bass, so like his Dad's Continue reading
A Crib, A Dresser and A Divine Appointment (Summer Reflections Part I)
Sometimes and idea is just an idea and sometimes it an idea with a bullet. A bullet speeds from a gun with intention Continue reading
It takes your breath away the first time you look into that tiny face that belongs to the child you brought home from the hospital all those years ago almost 25 now. Continue reading
Ep 13: The Personal and Powerful Work of the Holy Spirit
(Recorded 5-14-2017)
Ep 12: Blessed are those who hunger and thirst for righteousness.
Recorded 04/09/2017
Ep 11: Blessed are the Merciful
Recorded 03/05/2017
Pajama's and my Mom's Slippers
I am wearing my pajama's again and my Mom's slippers and I feel justifiably grateful that it is storming and cold outside so that I can stay in my pajamas for the rest of the day because I was going to anyways. I have been in them all weekend except church this morning and I have been in them every day after school. When I am at school all I want to do is get through the day so I can go home and put on my pajama's. I have been so grateful for the cold and rain it fits my mood and I can wrap up in my grey robe-wrap as I sit and watch TV or just sit. When I do get up I shuffle through the house in my Mom's purple slippers, the last new pair she ever wore. Continue reading
Ep 10: Building a Strong Family Bond Moment by Moment
(Recorded – 12/04/2016)
I was sitting in the 2nd row after the service waiting until the men stacking chairs would get to me and then I would move. I was scrolling on my phone and I was waiting for my husband as I have done on any given Sunday in any number of places. He had preached that morning in this church we were starting to call home and while I was comfortable with that idea opening my heart to engaging people was still not something that felt safe. I loved that Tim was loved and respected and that his preaching ministry was welcomed and wanted and that was good enough for me. I was basically minding my own business until a cute little woman sat down next to me and asked to talk to me. I was a bit surprised she approached me at all since I know exactly how intimidating I can look when I am scrolling on my phone daring anyone to approach me, so I was intrigued by what she had to say. Lois Pittman is the women's ministry director at our church (for you Bethanites she is Eunice Bruegman's sister) and she started her conversation by describing a women's event that occurred on a Saturday in the spring where women from the church were asked to speak instead of bringing in outside speakers. Continue reading
Ep 9: Rooted
(Recorded at Neighborhood Church 10/16/2016)
Buster and Eddie Buster too…
I had my feet up on the dash and I was immersed in the current summer novel of choice and Tim was lost in his thoughts as he sometimes likes to be when he is driving, companionable silence in our little Honda Fit as we buzzed across highway 152 on our way to Morgan Hill. We were meeting my college roommate and her husband to see Michael McDonald and Boz Scaggs in Saratoga at the Paul Masson Mountain Winery (amazing venue) when you are older trips back to your younger days with friends and music of those days is always a wonderful way to end the summer. My cell phone rang breaking the quiet and I looked down to see Mychal's name on the screen and immediately answered the phone, "Hey bud, what's up? My phone was connected to the Bluetooth so we could both hear him and he answered "I have a fun story to tell you…" Continue reading


text:
This article may be accessed by subscribers and e-wallet holders (EUR 6)
Log in to read more
Not reader yet?
Create an account to access our FREE SECTION
Khalifa Haftar has no Russian base but gets an airlift
Khalifa Haftar's inability to conclude his offensive on Tripoli has incited his Russian ally to strengthen its logistical and military support for him. The Watiya air base has in no way come under Russian control, contrary to claims from some [. [...]


text:
Meet The Affordable & Stylish Hyundai Accent
The Hyundai Accent is the budget-friendly, yet stylish sedan that we know our Harrisonburg, VA friends will love. Check out the Accent at Harrisonburg Hyundai today.
Many of our customers want a new car, but they don't think that they can afford them. What they don't realize is that we do have budget-friendly cars that won't break the bank. The bonus? They are stylish too. You don't have to compromise when you choose the Hyundai Accent. We welcome you to visit us at Harrisonburg Hyundai and see how much money you can save.
We understand that many people that are shopping for a car may be on a tight budget. This could be because they recently graduated from college, yet need a reliable vehicle to get to work, need a second family car, or maybe a teen driver needs a reliable ride. Whatever your situation is, the Hyundai Accent is budget-friendly. With a starting MSRP of just $14,745, it is one of the most affordable sedans on the market. This low price means a lower monthly payment that almost anyone can afford so you keep more cash in your account. Guess what? You will even save money at the pump because the Accent gets an EPA estimated 27 miles per gallon in the city and 37 miles per gallon on the highway. That means you can go further for less and stop less frequently to fill up too. Let us show you how much money the Accent can save you when you talk to one of our product specialists at Harrisonburg Hyundai today.
Comfort & Convenience
Just because you purchase a budget-friendly sedan doesn't mean you have to sacrifice your comfort and convenience features. The Hyundai Accent gives you more bang for your buck. You don't have to compromise features just because you want to save money. The Accent features steering wheel mounted audio and cruise controls that put everything at the tips of your fingers so you won't have to take your eyes off of the road to find your favorite radio station. The six-speaker audio system lets you pound out tunes from radio or SiriusXM Satellite Radio that offers over 150 channels of talk, news, sports, comedy, and music stations for hours of endless entertainment options while on the road. The Accent puts everything within your reach so you won't have to stretch to adjust the dials. Let us show you the comfort and convenience features on the Accent at Harrisonburg Hyundai.
Changing the Temp
Hiding the Digital Speed
Front Seat Heater
Smart Trunk
Safety Abounds
When you drive a smaller sedan, safety can be a top priority. There are so many distracted drivers on the road driving their gas guzzling trucks and SUVs to watch out for that anything that makes you safe is helpful. The Hyundai Accent is one of the safest sedans in its class. The Accent features six airbags that surround you and acts as a cushion in case of an accident to lessen the impact of the collision. The Accent also has the Vehicle Stability Management System that has active safety features. These include the Motor-Driven Power Steering, Electronic Stability Control, and Traction Control System. This powerful combination of features keeps you stable as you drive down the road for a safer drive. It also comes with anti-lock brakes and Brake Assist to help you stop faster. Talk to one of our product specialists at Harrisonburg Hyundai today.
Harrisonburg Hyundai – The Home Of Buying Confidence
At Harrisonburg Hyundai, we put our customers first because we believe you matter. We want you to feel confident when you buy your Hyundai Accent from us. That is why we take all the time you need to help you feel comfortable about your decision. We proudly list all of the information you need to help you educate yourself about the vehicle you are shopping for in one convenient place to make it easier for you. We have a team of professional, friendly, courteous, and knowledgeable people standing by to serve you and answer any questions you may have by phone, email, online, or in person when you stop by and see us. Get started today by browsing our diverse selection of Hyundai Accent models, and then come see us for a test drive.
Total Satisfaction Mile After Mile, at Harrisonburg Hyundai
The Harrisonburg Hyundai team is passionate about Hyundai vehicles; but as much as we love Hyundai, we're even more passionate about taking care of YOU. We're in business to serve the people of this community, and we know that our loyal customers all over Harrisonburg, VA and beyond are the reason we're still in business today. We work hard every day to repay that loyalty with an incredible selection of the best new Hyundai vehicles and a diverse array of pre-owned cars, unbeatable prices and unparalleled value, as well as a personalized experience that's always focused on YOU and your total satisfaction. Discover the difference when you buy the Harrisonburg Hyundai way today.


text:
Prints & Graphics
16" x 20" - $25
16" x 20" - $25
Decorate Your Dojo
High Quality prints suitable for framing. Perfect in your dojo, office, or home.
Great gifts! These large prints are top quality, printed on heavy weight paper.
Designed for framing in standard size frames.
Sample Watermarks Do Not Appear On Final Print


text:
The new Porsche 911 GT2 expresses the power and performance potential of the fastest-ever 911 even at a standstill. Distinguished by its striking design, with large air intakes at the front and the rear wing so characteristic of this very special model, the latest GT2 maintains the reputation of this specialised edition.
Available exclusively with rear-wheel drive and a manual six-speed gearbox, this high-performance athlete accelerates from 0 to 62mph in just 3.7 seconds and reaches a top speed of 204mph. This is combined with an average fuel consumption of 22.6mpg (12.5 litres/100km) in the New European Driving Cycle (NEDC), thus offering a level of fuel economy regarded as quite impossible in this performance class just a few years ago.
The power unit of the new 911 GT2 is based on the six-cylinder 'boxer' engine of the 911 Turbo, with air supply by two exhaust gas turbochargers featuring variable turbine geometry (VTG). The 3.6-litre power unit achieves its maximum output of 530bhp (390kW) at 6,500rpm, with maximum torque of 680 Newton-metres (501 lb-ft) maintained consistently between 2,200 and 4,500rpm. With its unladen weight of 1,440kg (3,175lb), the 911 GT2 offers a power-to-weight ratio of just 2.72kg/6.00lb per horsepower.
The increase in engine power by 50 bhp over the 'regular' 911 Turbo model is provided in the new 911 GT2 by fitting the two turbochargers with a larger compressor wheel and a flow-optimised turbine housing, raising turbocharger pressure to an even higher level. And for the first time, the engineers at Porsche have combined the turbocharged engine with an expansion-type intake manifold – a genuine revolution on turbocharged power units. The expansion intake manifold uses the principle of oscillating air in the intake manifold during the cooler expansion phase, keeping the temperature of the fuel/air mixture lower than is the case with the 911 Turbo. This, in turn, means a significant increase in all-round efficiency, with fuel consumption down by up to 15 per cent under full load despite the increase in engine output.
The 911 GT2 offers new achievements also in terms of emissions management and control. This is the first Porsche homologated for the road to feature a rear silencer and tailpipes made of titanium as standard equipment. This expensive material reduces weight by approximately 50 per cent versus a comparable component made of stainless steel to just 9kg.
The new 911 GT2 comes as standard with Porsche Ceramic Composite Brakes (PCCB), featuring brake discs made of a composite carbon fibre/ceramic compound ensuring maximum stopping power maintained with an extremely high level of consistency. A further advantage is that PCCB reduces the weight of the unsprung masses versus comparable grey cast-iron discs by approximately 20kg (44lb).
Yet another feature fitted as standard is the electronically controlled Porsche Active Suspension Management (PASM). The GT2 rides on 19-inch diameter light-alloy wheels, running at the front on 235/35 ZR 19 tyres and at the rear on 325/30 ZR 19 tyres.
The new 911 GT2 will cost from £131,070, including a Porsche Vehicle Tracking System (VTS), a sophisticated vehicle security package approved to Thatcham Category 5 standard, and a Porsche Driving Experience course.
The 911 GT2 will make its debut at the International Auto Show in Frankfurt, Germany, in September and arrives in Porsche Centres across the UK and Ireland during November 2007.


text:
Courteney Cox and Matthew Perry are not dating, despite a report.
The story claimed that the former "Friends" co-stars had grown close after Courteney's split from Snow Patrol guitarist Johnny McDaid, but Gossip Cop has learned it's bogus.
The article claims they are taking it slow and excited to see what the future holds, but sadly for Monica and Chandler fans, they remain strictly friends.
The story was further fueled when a 2005 photo of Cox and Perry out to lunch resurfaced, and fans believed it was a new pic.
Meanwhile, a source close to Cox told Gossip Cop there is no truth to the dating rumors.
For more rumors busts, visit GossipCop.com.


text:
Since our beginnings in 2004, Aeris has established risk management standards for private community loan funds (CDFIs) and has helped drive the development of impact measurement and management practices in the CDFI industry—a learning curve now being replicated in the broader impact investment world. In 2013, the Aeris Cloud became the first online platform to help CDFI investors understand the impact and financial performance of their investments. Today, the broader investing field seeks a reliable, high-quality tool to collect, manage, and analyze impact data. Aeris' unique experience and expertise can help accelerate the pace of impact investing.
Giving Strong is a social impact consulting firm that advises foundations, corporations and other social enterprises on strategy and communications to reach their impact goals.
The Edward W. Hazen Foundation, a private foundation established in 1925, is committed to supporting organizing and leadership of young people and communities of color in dismantling structural inequity based on race and class.
The Weissberg Foundation ("Foundation") envisions a just world that recognizes inequities and builds access, opportunity, and power so that all can thrive. Through funding, amplification, capacity building, and collaboration, the Foundation advances organizations and efforts that give voice and opportunity to historically marginalized populations. The Foundation supports this mission and its operations with the assets in its Endowment Portfolio ("Endowment"). All capital has impact (positive, negative, and neutral) and the Foundation will evaluate this impact through the lens of our mission, economic opportunity, and with attention to the broader public good. We acknowledge that capital markets perpetuate structures of unequal access to opportunities, which reinforces and grows the social and economic inequities of our society. Only through intentional allocation of capital can we influence, expose, and change these existing conditions. Achieving social equity and sustainability requires alignment of our Endowment with the Foundation's mission and vision. A fiduciary responsibility of the Foundation is to invest capital to maximize a rate of return in a way that that does not harm or detract from its mission or vision. When applicable, the Foundation will prioritize investments in traditionally marginalized populations, reduction of inequalities, sustainable cities and communities, and peace justice and strong institutions.
The vision of the Wilson Foundation is to act as a catalyst for change.
The change in the nature, role and strength of the family and the community has alienated individuals and citizens from each other and their institutions resulting in a significant decline in the quality of life for all of us. Our mission is to improve the quality of life through initiating and supporting projects that measurably demonstrate a means of creating a sense of belonging within the family and the community.
Have a question, website feedback, or idea to make our services better?
Please contact [email protected] if you have trouble logging in.


text:
Macclesfield Town Under 18s launched another devastating performance this afternoon, as five second half goals helped to secure a 7 - 1 victory against Northwich Victoria.
The visitors began the game in the ascendancy, with Jake Hall unleashing a thunderous strike at goal in the 7th minute which was tipped onto the bar by the Northwich 'keeper.
The game then settled down somewhat as the contest largely became a midfield battle, with both sides unable to create an real clear cut chances.
The next opportunity of note came in the 25th minute, when Hussein Mohammed burst through the home defence - yet he was thwarted by the home 'keeper who managed to tip the ball around the far post.
Manager Jim Hall then introduced Freddie Sass into the fray and that decision proved to be an inspired one, as three minutes after his introduction the youngster hammered the ball home from eighteen yards to finally break the deadlock.
Two minutes later our lead was doubled, as Jake Hall rifled home a twenty yard strike to firmly put The Silkmen in control.
The visitors emerged from the half-time interval looking to end the game as a contest in the least amount of time possible and just three minutes into the second period, Mohammed's cross from the left was easily converted by Sass for number three.
Northwich did try to get back into the game with a number of neatly crafted moves, yet Mohammed made it four on 55 minutes - as he tapped the ball home from close range.
It was to become five on the hour mark, when Max Phillips hammered home a twenty-five yard screamer into the bottom corner and ten minutes from time, James Madeira seized on a loose ball in the Northwich penalty area to fire home the sixth.
Shaun Holden then made it seven with just six minutes left on the clock, before Ben Hickey claimed a consolation for the hosts as he converted from close range.
The lads return to the Moss Rose next Sunday, as they contest their League Cup Quarter Final tie with Davenham.
After the game, we caught up with Freddie Sass and Dan Dolling -
Macclesfield Town: Reynolds, Jordan, Box, Midgley, Dolling, Kerr, Hall, Phillips, Mohammed, Richardson, Holden.
Subs: Sass, Samms, Madeira, Nolan, Arkian


text:
Bristol Farms
Bristol Farms is a chain of high-end grocery stores located in the Southern California area. As the sole chalk sign artist of the Westchester store, I wrote, illustrated, and maintained all handwritten signs. This included the price tags for the deli, produce, hot bar, meat, and front end departments. My illustrative work combined with brand consistency was a reason the store had its highest gross profit margin compared to previous fiscal years.


text:
SHARP QW-GD52I472X Full-size Integrated Dishwasher
Product code: 179399
Product features
- 82 x 59.6 x 55 cm (H x W x D)
- Ideal for larger households (15 place settings)
- Energy rating: A++
- Quick wash: 30 minutes
- Ideal for open plan living
Product features
- 82 x 59.6 x 55 cm (H x W x D)
- Ideal for larger households (15 place settings)
- Energy rating: A++
- Quick wash: 30 minutes
- Ideal for open plan living
- Reduce your energy bill with an A++ energy efficiency rating
-IntelliWash sensor uses the right amount of heat and waster for economical cleaning
- Enjoy a sparkling clean wash in just 50 minutes with the Express wash cycle
- Perfect for families and entertaining with 15 place settings
Reduce your energy bill
Save money on your energy bills with an A++ energy efficiency rating. This Sharp QW-GD52I472X Full Size Dishwasher only uses nine litres of water per cycle, so you can reduce your carbon footprint and help to make savings on your utilities whilst enjoying a consistently great wash.
IntelliWash sensor
Even if the dishwasher is filled up with dirty dishes, the IntelliWash sensor will ensure that the correct amount of water, time and energy is used to achieve a perfect clean.
When the dishwasher is running quietly, it's not always possible to hear whether it's on or not. That's why the ILED program running indicator lets you know when it's time to unload by emitting a gentle LED glow.
Enjoy a sparkling clean wash
Great for when you need dishes cleaned in a hurry, this super-speedy Express wash cycle can perform a 65% wash in just 50 minutes, so your crockery and cutlery will always look perfectly clean.
For an even quicker clean this dishwasher uses Turbo Drying technology, which helps to get your dishes super-dry and shiny in less time.
Perfect for families
With 15 place settings, this dishwasher is perfect for busy households. For added convenience, the internal third tray can be removed to help you accommodate long-handled utensils and larger pots and pans.
Technical specifications for SHARP QW-GD52I472X Full-size Integrated Dishwasher
|Full-size integrated dishwasher
|Manufacturer's guarantee
|2 years
|Number of programs
|Number of wash temperatures
|- Auto Delicate 30-50
- Auto Intensive 60-70
- Auto Normal 50-60
- Eco 50
- Express 50 min
- Hygiene 60
- Quick 30 min
- Quick Wash 18 min
|Quick wash time
|18 minutes
|Time remaining indicator
|Delay timer
|Height-adjustable top basket - while loaded
|Third rack
|Cutlery basket / cutlery tray
|Adjustable racking
|Adjustable prongs
|Interior material
|Stainless steel
|Overflow protection
|Additional features
|- IntelliWash
- Turbo Drying
- Microban coated filter
- ILED program running indicator
|Digital display
|LED screen
|Rinse aid indicator
|Salt indicator
|Wash efficiency rating
|Energy efficiency rating
|Place settings
|Annual energy consumption
|267 kWh
|Annual water consumption
|2520 litres
|Energy consumption per cycle
|0.94 kWh
|Water consumption per cycle
|9 litres
|Drying efficiency rating
|Noise level
|47 dB(A)
|Please note this product should be installed by a qualified engineer, such as one of our Knowhow experts.
|- 820 x 596 x 550 (H x W x D)
- 32.28 x 23.46 x 21.65 inches (H x W x D)
|Recess dimensions
|820-870 x 600 x 600 mm (H x W x D)
|Boxed dimensions
|850 x 660 x 640 mm (H x W x D)
|38 kg
|Boxed weight
|40 kg
|Box contents
|- Sharp QW-GD52I472X Full-size Integrated Dishwasher
- Instructions
- Water hose
- Water testing strip
Delivering your online order
Get this delivered when it suits you
We'll show you your current delivery options for this item in checkout. Subject to the time you order, and where you want delivery, these options will include:
- Standard delivery within 5 working days - €24.99
Availability CheckerHow and when can you get hold of your selections? Finding out should be quick and easy, so with just a few clicks our Availability Checker shows you:
- Delivery availability in your area
- Local stores with stock you can collect from now
- Enter a town or eircode in the box on the product page for an item you've selected
- Click on the magnifying glass
- Review the delivery and collection options local to you – just click on any one to add it to your basket
Stock updatesFind something you want but it's not in stock? On the item's product page, simply click on 'Email me when back in stock', and let us have your email address. We'll let you know as soon as it's available again.
Order online and collect in store
- Enter a town or eircode in the box on the product page for an item you've selected
- Click on the magnifying glass
- Review the collection options local to you – just click on any one to add it to your basket
Reserve and Collect
If your item is in stock locally, you can reserve it online for free, to collect from store minutes later. So you'll get it even quicker than we can bring it to you, and with no delivery charges. Once reserved, your item's held till closing time the next day, to be paid for when you collect.
Let's install and recycle
Integrated Appliance Installation
We can also install your new appliance on the day we deliver, for most of Ireland. Just add installation at the 'Confirm basket' page at the checkout,charge applies.Here's what we'll do:
- Check existing electrical and plumbing connections are within 1.5 metres of the appliance.
- Disconnect the existing appliance, remove the plinth and panel door, then un-box and check the new product.
- Connect your new appliance to the electricity, mains water and waste.
- Test there are no leaks and that the appliance is level and fully operational.
- Re-fit the plinth and panel door to the new appliance.
- Take the packaging from your new appliance away. If you add recycling to your order we'll also take your old appliance away for recycling.
Note: Before we arrive, you will need to make sure access is clear, that the appliance is an identical size to the one it is replacing, that it will fit into the existing space and to the door panels. Plumbing and electric connections must be within 1.5 metres.


text:
Toggle navigation
The Blissful Herbs Story
Herbs for Pregnancy
Herbs for Birth
Herbs for the Postpartum & Breastfeeding
Herbs for Little Ones
Herbs for Fertility & Libido
Herbs for Pregnancy Loss
Herbs for the Moon Cycle
The Family Remedy Chest
Herbs for Strength and Calm
Herbs for Men
Bath Herbs
Herbal Salves
Herbal Tinctures
Herbal Syrups, Mels & Cordials
Accessories, Books & Misc
Umbilical Cord Ties & Sibling Wrist Bands
Package Deals & Gift Packs
Bespoke Teas & Singles
Herbalist Consultations
Sales and Specials
Artisan Tea Sets by Rohan
Need Help With Your Order? Tips & FAQs
Blissful Herbs on Social Media
Essential Oils by Young Living
Beating the Dreaded Lurgy - give that virus the vamoose
Blissful Herbs Gift Boutique
Herbalist Consultations
Birth Services
Julie's Blog
view cart
sign in
Returning Customers
Forgot Password?
sign up
Accessories, Books & Misc
Pretty Petals
view details
Bamboo tea strainer
view details
Bamboo long-handled spoon
view details
Bamboo long-handled spoon and Bamboo tea strainer
view details
Re-usable unbleached muslin drawstring bag
view details
Cleansing bottle
view details
Paper Filter tea bags
view details
Sassy Soap
view details
Henna Tubes
view details
Glass Tea Infuser Bottle
view details
Bone Broth Medicinal Herbs
view details
Anointing Oil
view details
3.30 Boost Nutritional Powder
view details
Heavenly Peace Sleep Sachet - out of stock, sorry!
view details
Spice of Life Organic Herb Sprinkle
view details
Blissful Herbs Gift Certificate
view details
Mummy's Having a Homebirth picture book
view details
'Welcome Home' picture book
view details
Hand Sanitizer
view details
Nature Deva Perfume Balms
view details
E-Book: Best Birth Resources
view details
E-Book: A Doula's Guide to Looking After Your Perineum
view details
Organic Aloe Vera Gel 500 ml
view details
Essential Oil Vial Necklace
view details
Ear Savers - for use with face masks
view details


text:
Who Should Be Trump's VP?
First for some news you definitely weren't expecting: Despite getting dropped by Univision, NBC, and Macy's this week, it seems Donald Trump is actually doing well in the polls. In fact, despite his disparaging remarks against Mexican immigrants last week, Trump recently placed second behind Jeb Bush in a New Hampshire poll. (Second!) The first question that this brought to my mind: Who in the world would seriously vote for him? And then there was this: Now that he's apparently a viable contender, who will Trump choose as his Vice President? (You know, in this fantasy world where the host of The Apprentice could one day become the leader of the free world.) As the race begins to heat up, many of Trump's fellow Republicans have come forward to comment on Trump's recent remarks about Mexican immigrants — and have left me wondering which ones might be vying for a spot beside him on the ticket.
If you're just catching up, much of the buzz around Trump's presidential bid has to do with his announcement speech June 16, in which he said Mexican immigrants are "bringing drugs, they're bringing crime, they're rapists." (Exactly the kind of sweeping racist comments you'd expect for a future Prez!) And while that wasn't the only shocking thing Trump said in his announcement, it's definitely the one that's received the most attention, for obvious reasons. While some of his fellow Republicans have hurled harsh criticism at the Donald following his remarks, others have both laughed off Trump's comments and actually lent their support.
So which ones (if any) might be angling to be his VP one day? Let's take a quick look at what other GOP-ers are saying about him...
1. Ted Cruz
Yep, Ted Cruz is apparently a big fan of The Donald, and showed his support for the would-be president just this week. After Trump was called out for those racist comments about Mexican immigrants and was dropped by Univision, NBC, and Macy's, Cruz stepped in and defended Trump, calling him "terrific." (Hmmm... does it sound like somebody's sucking up, or is it just me?)
2. Rick Perry
Rick Perry also wasn't deterred by some of Trump's more cringe-worthy and straight-up racist comments last week. In fact, he was pretty amused by them. On Wednesday, Perry shrugged off criticism over Trump's comments about Mexico, and instead simply called it "good TV." But that's not to say he supports him — according to Mediaite, the former governor of Texas was quick to say we should all "let [Trump] run on that," adding that instead, "I will run on actual experience."
3. George Pataki
Despite being a fellow New Yorker and fellow Republican, Governor George Pataki was not cool with Trump's disparaging remarks made last week. In fact, Pataki dismissed Trump's comments as "disrespectful," and told Business Insider that nobody should ever "attack the character of those who are trying to build a better future." (So yeah, I'm going to go ahead and rule out a Trump-Pataki ticket for 2016.)
4. Mike Huckabee
One guy Trump does have in his corner? Mike Huckabee. And if I'm being honest here, his praise of Donald Trump is hinting at a straight-up brofest. When Trump announced that he was running for president earlier this month, Huckabee stated that Trump "has the courage to put his name on the ballot and take the incoming attacks." He also added, "I personally like him." Geez, just kiss him already.
So there you have it, America. Whether you love him, hate him, or just find his hair simultaneously hilarious and terrifying, we all have an opinion about Donald Trump. (And if that recent Republican nominee poll is to believed, it looks like it's not all bad!) While I am not exactly confident that Trump has enough momentum to ultimately overtake Bush as the front-runner for the Republican ticket, I really can't help but wonder about what on Earth a Trump-Huckabee (Trumpabee?) White House would look like... But hey, maybe we should just go straight to the source himself: On June 17, while speaking with ABC News, Trump said, "I think Oprah would be great [as Vice President]. I'd love to have Oprah." He also thinks it'd be a slam dunk. "I think we'd win easily, actually," The Donald added. (Something tells me that Oprah, who apparently has not responded to ABC News' calls, might not exactly join a Republican ticket. Just a hunch...)
No matter who's in that spot, though, one thing's for sure: it's definitely going to be an interesting race come 2016.
Images: Getty Images (4)


text: finally, the wait is over!! "i'm calling" featuring M C Collinz is a song that runs through Ur brains, pierces through your bones, sinks down to your heart and brings your spirit alive!! trust me, u haven't heard what sounds like this before.. soon after ' blessings ' naija on fire' mujo ' by 2YUNG the african famous prolific rap singer dropped and became wide spread ,hit the air waves and was well received by his ever supporting fans across the globe.This time around he features MC COLLINZ a fellow music legend,an afro high life music sensation , the 'NEK UNEK' crooner on this new sound tittled 'I'M CALLING'.the song is an R and B pop sound..this is a potential anthem' a good recommendation for your playlist.you can connect with 2yung on these social media platform..


text:
Time: November 14, 2018 from 11:30am to 1pm
Location: City Fish Market
Street: 7940 Glades Rd.
City/Town: Boca Raton
Phone: 5616208888
Event Type: politics/networking
Organized By: Howard Appell
Latest Activity: Nov 10, 2018
Export to Outlook or iCal (.ics)
Colonel Robert L. Allen with the Palm Beach County Sheriff's Department will speak to the Gold Coast Tiger Bay Club on November 14th at the City Fish Market in Boca Raton. He will be talking about Homeland Security in Palm Beach County. Tickets for this speech and luncheon are only $35 for first time guests. Join us for this informative event at one of Boca's best clubs for networking, The Gold Coast Tiger Bay Club. Don't forget to bring business cards. Get your tickets here. www.goldcoasttigerbayclub.com
© 2020 Created by Mary Kate Leming. Powered by


text:
Taste at Hotel Jen Tanglin Singapore by Shangri-La
Jen's Kitchen On-the-Go
Jen's Kitchen On-the-Go serves some really great coffee and light bites from the kitchen, as well as some of Singapore's finest artisanal products from some great young up and coming brands. Fast Wi-Fi and charging stations are also available.
Check out our menu here.
For all reservations or enquiries, please call (65) 6831 4374 or email email@example.com


text:
Cottage Cheese Spinach Salad
Total TimePrep/Total Time: 10 min.
Makes10 servings
- 1 package (10 ounces) fresh spinach, torn
- 1-1/2 cups Daisy 4% cottage cheese
- 1/2 cup chopped pecans, toasted
- 1/2 cup sugar
- 3 tablespoons white vinegar
- 2 teaspoons prepared horseradish
- 1/2 teaspoon salt
- 1/2 teaspoon ground mustard
- In a large serving bowl, layer half of the spinach, cottage cheese and pecans. Repeat layers. In a small bowl, combine the remaining ingredients. Drizzle over salad; toss to coat.
Nutrition Facts1 cup: 121 calories, 6g fat (1g saturated fat), 7mg cholesterol, 257mg sodium, 13g carbohydrate (11g sugars, 1g fiber), 5g protein.
Recommended Video
Aug 9, 2016
My family loves spinach, but hates the texture of cottage cheese-and we all love this salad. I cut the sugar in half and used walnuts because we had them on hand, was so easy and delicious! Will be making for all the summers cookouts!
Aug 17, 2015
My husband and I cut the recipe in half and ate it all! Very good and unusual!
Aug 8, 2014
Excellent! I've been making this salad for some time now, and it's delicious!
May 3, 2011
No comment left
Nov 18, 2009
GREAT spinach salad, wonderful flavour!!!
May 24, 2009
No comment left
Apr 12, 2009
WOW! This is one great salad. I am not a huge fan of spinach, but the flavor the dressing adds to the spinach is to die for. A little heat from the horseradish makes all the difference!I always make enough for left overs, to have as a quick, healthy lunch. After storing the dressing in a canning jar, I shake to mix it, and store leftovers in the fridge. When eating leftover, I just warm the jar in the microwave a few seconds, to get the sugar mixed again, and in seconds I have a wonderful lunch! Cant stress how great this salad is.
Aug 24, 2008
No comment left
Feb 11, 2007
No comment left
Sep 12, 2006
No comment left