Retro-transceiver array using monopole antennas

In this paper, we show how a self-tracking antenna array constructed using gimel /4 monopoles can be constructed, which is capable of receiving with gain over an entire 360 degrees azimuthal cut. It is also shown how the self-tacking receive unit can be used in conjunction with a self-phased transmitter so that self-steered spatially selective receive and transmit functions ran be formed simultaneously. The resulting array is capable of maintaining spatially selective receive and transmit functions to a roaming target without prior knowledge of its physical location.

Computationally efficient sequential learning algorithms for direct link resource-allocating networks

Computionally efficient sequential learning algorithms are developed for direct-link resource-allocating networks (DRANs). These are achieved by decomposing existing recursive training algorithms on a layer by layer and neuron by neuron basis. This allows network weights to be updated in an efficient parallel manner and facilitates the implementation of minimal update extensions that yield a significant reduction in computation load per iteration compared to existing sequential learning methods employed in resource-allocation network (RAN) and minimal RAN (MRAN) approaches. The new algorithms, which also incorporate a pruning strategy to control network growth, are evaluated on three different system identification benchmark problems and shown to outperform existing methods both in terms of training error convergence and computational efficiency. (c) 2005 Elsevier B.V. All rights reserved.

The Deubiquitinating Enzyme USP17 Is Highly Expressed in Tumor Biopsies, Is Cell Cycle Regulated, and Is Required for G1-S Progression

Ubiquitination is a reversible posttranslational modification that is essential for cell cycle control, and it is becoming increasingly clear that the removal of ubiquitin from proteins by deubiquitinating enzymes (DUB) is equally important. In this study, we have identified high levels of the DUB USP17 in several tumor-derived cell lines and primary lung, colon, esophagus, and cervix tumor biopsies. We also report that USP17 is tightly regulated during the cell cycle in all the cells examined, being abundantly evident in G1 and absent in S phase. Moreover, regulated USP17 expression was necessary for cell cycle progression because its depletion significantly impaired G1-S transition and blocked cell proliferation. Previously, we have shown that USP17 regulates the intracellular translocation and activation of the GTPase Ras by controlling Ras-converting enzyme 1 (RCE1) activation. RCE1 also regulates the processing of other proteins with a CAAX motif, including Rho family GTPases. We now show that USP17 depletion blocks Ras and RhoA localization and activation. Moreover, our results confirm that USP17-depleted cells have constitutively elevated levels of the cyclin-dependent kinase inhibitors p21cip1 and p27kip1, known downstream targets of Ras and RhoA signaling. These observations clearly show that USP17 is tightly regulated during cell division and that its expression is necessary to coordinate cell cycle progression, and thus, it may be considered a promising novel cancer therapeutic target. Cancer Res; 70(8); 3329–39. ©2010 AACR.

Evidence for a Direct and Functional Interaction between the Regulators of G Protein Signaling-2 and Phosphorylated C Terminus of Cholecystokinin-2 Receptor

Signaling of G protein-coupled receptors (GPCRs) is regulated by different mechanisms. One of these involves regulators of G protein signaling (RGS), which are diverse and multifunctional proteins that bind to active G alpha subunits of G proteins and act as GTPase-activating proteins. Little is known about the molecular mechanisms that govern the selective use of RGS proteins in living cells. We first demonstrated that CCK2R-mediated inositol phosphate production, known to be G(q-)dependent, is more sensitive to RGS2 than to RGS4 and is insensitive to RGS8. Both basal and agonist-stimulated activities of the CCK2R are regulated by RGS2. By combining biochemical functional, and in silico structural approaches, we demonstrate that a direct and functional interaction occurs between RGS2 and agonist-stimulated cholecystokinin receptor-2 (CCK2R) and identified the precise residues involved: phosphorylated Ser434 and Thr439 located in the C-terminal tail of CCK2R and Lys62, Lys63, and Gln67, located in the N-terminal domain of RGS2. These findings confirm previous reports that RGS proteins can interact with GPCRs to modulate their signaling and provide a molecular basis for RGS2 recognition by the CCK2R.

Interactions between the budding yeast IQGAP homologue Iqg1p and its targets revealed by a split-EGFP bimolecular fluorescence complementation assay.

A split-EGFP based bimolecular fluorescence complementation (BiFC) assay has been used to detect interactions between the Saccharomyces cerevisiae cytoskeletal scaffolding protein Iqg1p and three targets: myosin essential light chain (Mlc1p), calmodulin (Cmd1p) and the small GTPase Cdc42p. The format of the BiFC assay used ensures that the proteins are expressed at wild type levels thereby avoiding artefacts due to overexpression. This is the first direct in vivo detection of these interactions; in each case, the complex is localised to discrete regions of the yeast cytoplasm. The labelling with EGFP fragments results in changes in growth kinetics, cell size and budding frequency. This is partly due to the reassembled EGFP locking the complexes into essentially permanent interactions. The consequences of this for Iqg1p interactions and BiFC assays in general are discussed. (c) 2008 International Federation for Cell Biology. Published by Elsevier Ltd. All rights reserved.

Vasodilator-Stimulated Phosphoprotein Regulates Inside-Out Signaling of beta(2) Integrins in Neutrophils

The monomeric GTPase Rap1 controls functional activation of beta2 integrins in leukocytes. In this article, we describe a novel mechanism by which the chemoattractant fMLP activates Rap1 and inside-out signaling of beta2 integrins. We found that fMLP-induced activation of Rap1 in human polymorphonuclear leukocytes or neutrophils and differentiated PLB-985 cells was blocked by inhibitors of the NO/guanosine-3',5'-cyclic monophosphate-dependent protein kinase (cGKI) pathway [N-(3-(aminomethyl)benzyl)acetamidine, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, DT-3 peptide, 8-(4-chlorophenylthio)guanosine 3',5'-cyclic monophosphothioate, Rp-isomer triethylammonium salt-guanosine-3',5'-cyclic monophosphate], indicating that the downstream signaling events in Rap1 activation involve the production of NO and guanosine-3',5'-cyclic monophosphate, as well as the activation of cGKI. Silencing the expression of vasodilator-stimulated phosphoprotein (VASP), a substrate of cGKI, in resting PLB-985 cells or mice neutrophils led to constitutive activation of Rap1. In parallel, silencing VASP in differentiated PLB-985 cells led to recruitment of C3G, a guanine nucleotide exchange factor for Rap1, to the plasma membrane. Expression of murine GFP-tagged phosphodeficient VASP Ser235Ala mutant (murine serine 235 of VASP corresponds to human serine 239) in PLB-985 cells blunted fMLP-induced translocation of C3G to the membrane and activation of Rap1. Thus, bacterial fMLP triggers cGKI-dependent phosphorylation of human VASP on serine 239 and, thereby, controls membrane recruitment of C3G, which is required for activation of Rap1 and beta2 integrin-dependent antibacterial functions of neutrophils.

Exeter-Hall Science and Evangelical Rhetoric in Mid-Victorian Britain

In the early and mid-Victorian period public pronouncements by evangelicals were often described as the antithesis of rational speech. The voice of science, on the other hand, was routinely equated with the voice of reason. This disparity was particularly clear in satirical and critical commentary about the platform rhetoric associated with London’s Exeter Hall, a key meeting place for evangelicals and a metonym for evangelical expressions of Christian belief. It was against this backdrop that the fledgling Young Men’s Christian Association inaugurated a popular series of lectures in 1845. Held in Exeter Hall from 1848, the series ran until 1865 and proved to be immensely popular. By investigating the ways in which the promotion of science was combined with religious exhortation in the YMCA lectures, this paper examines how evangelicals positioned themselves with respect to the growing cultural authority of science. The paper also argues that these efforts were indelibly marked by the Hall and the communicative medium in which they were made. As such, the paper sheds light on the significance of platform culture within and beyond evangelicalism and on the importance of venue and audience in understanding science and religion relations in an age of lecturing.

RhoA protein expression correlates positively with degree of malignancy in astrocytomas

Astrocytic tumors are the most common intracranial neoplasms. Their prognoses correlate with a conventional morphological grading system that suffers from diagnostic subjectivity and hence, inter-observer inconsistency. A molecular marker that provides an objective reference for classification and prognostication of astrocytic tumors would be useful in diagnostic pathology. RhoA, a GTPase protein involved in cell migration and adhesion has been shown to be upregulated in a variety of human cancers. Based on direct analysis of clinical materials, our study demonstrates increased expression of RhoA in high-grade astrocytomas. This observation may be relevant to astrocytoma biology and the development of potential therapeutics against high-grade astrocytomas. Of more immediate consequence, utilization of this marker may aid in the routine pathological grading (and hence prognostication) of astrocytomas. (c) 2006 Elsevier Ireland Ltd. All rights reserved.

Constitutive activation of the Raf-MAPK pathway causes negative feedback inhibition of Ras-PI3K-AKT and cellular arrest through the EphA(2) receptor

The Raf-mitogen-activated protein kinase (MAPK) and phosphatidylinositide 3-kinase (PI3K)-AKT pathways are two downstream effectors of the small GTPase Ras. Although both pathways are positively regulated by Ras, the Raf-MAPK and PI3K-AKT pathways have been shown to control opposing functions within the cell, suggesting a need for cross-talk regulation. The PI3K -AKT pathway can inhibit the Raf-MAPK pathway directly during processes such as muscle differentiation. Here we describe the ability of the Raf-MAPK pathway to negatively regulate the PI3K-AKT pathway during cellular arrest. Constitutive activation of Raf or methyl ethyl ketone 1 (MEK1) leads to inhibition of AKT and cellular arrest. Furthermore, we show that activation of Raf-MEK1 signaling causes negative feedback inhibition of Ras through the ephrin receptor EphA(2). EphA(2)-mediated negative feedback inhibition is required for Raf-induced AKT inhibition and cell cycle arrest, therefore establishing the inhibition of the Ras-PI3K-AKT pathway as a necessary event for the Raf-MEK1-regulated cellular arrest.

C3G overexpression in glomerular epithelial cells during anti-GBM-induced glomerulonephritis

The guanine nucleotide exchange factor C3G, along with the CrkII adaptor protein, mediates GTP activation of the small GTPase proteins Rap1 and R-Ras, facilitating their activation of downstream signaling pathways, which had been found to be important in the pathogenesis of glomerulonephritis. We found that expression of C3G protein was upregulated in glomerular epithelial cells in an experimental model of accelerated anti-GBM antibody-induced glomerulonephritis expression. To determine the consequence of its increased expression, we transfected C3G (using adenoviral constructs) into cultured glomerular epithelial cells and measured the activated forms (i.e., GTP-bound) forms of Rap1 and R-Ras. Activation of Rap1 was not affected by C3G; however, the basal level of GTP-bound R-Ras was decreased. Further, C3G over-expression enhanced the activation of R-Ras in response to endothelin. Overexpression of C3G also led to a significant reduction in glomerular epithelial cell spreading and decreased the cells' E-cadherin expression and augmented their migration. We found that C3G was overexpressed in accelerated anti-GBM antibody-induced glomerulonephritis and suggest that this modulates glomerular epithelial cell morphology and behavior.

PTEN regulates colorectal epithelial apoptosis through Cdc42 signalling.

BACKGROUND:
Phosphatase and tensin homologue deleted on chromosome 10 (PTEN) regulation of the Rho-like GTPase Cdc42 has a central role in epithelial polarised growth, but effects of this molecular network on apoptosis remain unclear.

METHODS:
To investigate the role of Cdc42 in PTEN-dependent cell death, we used flow cytometry, in vitro pull-down assays, poly(ADP ribose) polymerase (PARP) cleavage and other immunoblots in isogenic PTEN-expressing and -deficient colorectal cells (HCT116PTEN(+/+), HCT116PTEN(-/-), Caco2 and Caco2 ShPTEN cells) after transfection or treatment strategies.

RESULTS:
The PTEN knockout or suppression by short hairpin RNA or small interfering RNA (siRNA) inhibited Cdc42 activity, PARP cleavage and/or apoptosis in flow cytometry assays. Transfection of cells with wild-type or constitutively active Cdc42 enhanced PARP cleavage, whereas siRNA silencing of Cdc42 inhibited PARP cleavage and/or apoptosis. Pharmacological upregulation of PTEN by sodium butyrate (NaBt) treatment enhanced Cdc42 activity, PARP cleavage and apoptosis, whereas Cdc42 siRNA suppressed NaBt-induced PARP cleavage. Cdc42-dependent signals can suppress glycogen synthase kinase-ß (GSK3ß) activity. Pharmacological inhibition of GSK3ß by lithium chloride treatment mimicked effects of Cdc42 in promotion of PARP cleavage and/or apoptosis.

CONCLUSION:
Phosphatase and tensin homologue deleted on chromosome 10 may influence apoptosis in colorectal epithelium through Cdc42 signalling, thus providing a regulatory framework for both polarised growth and programmed cell death.

IQ-motif selectivity in human IQGAP2 and IQGAP3: binding of calmodulin and myosin essential light chain

The IQGAP [IQ-motif-containing GAP (GTPase-activating protein)] family members are eukaryotic proteins that act at the interface between cellular signalling and the cytoskeleton. As such they collect numerous inputs from a variety of signalling pathways. A key binding partner is the calcium-sensing protein CaM (calmodulin). This protein binds mainly through a series of IQ-motifs which are located towards the middle of the primary sequence of the IQGAPs. In some IQGAPs, these motifs also provide binding sites for CaM-like proteins such as myosin essential light chain and S100B. Using synthetic peptides and native gel electrophoresis, the binding properties of the IQ-motifs from human IQGAP2 and IQGAP3 have been mapped. The second and third IQ-motifs in IQGAP2 and all four of the IQ-motifs of IQGAP3 interacted with CaM in the presence of calcium ions. However, there were differences in the type of interaction: while some IQ-motifs were able to form complexes with CaM which were stable under the conditions of the experiment, others formed more transient interactions. The first IQ-motifs from IQGAP2 and IQGAP3 formed transient interactions with CaM in the absence of calcium and the first motif from IQGAP3 formed a transient interaction with the myosin essential light chain MIc1sa. None of these IQ-motifs interacted with S100B. Molecular modelling suggested that all of the IQ-motifs, except the first one from IQGAP2 formed alpha-helices in solution. These results extend our knowledge of the selectivity of IQ-motifs for CaM and related proteins.

Rescue of glandular dysmorphogenesis in PTEN-deficient colorectal cancer epithelium by PPARγ-targeted therapy

Disruption of glandular architecture associates with poor clinical outcome in high-grade colorectal cancer (CRC). Phosphatase and tensin homolog deleted on chromosome ten (PTEN) regulates morphogenic growth of benign MDCK (Madin Darby Canine Kidney) cells through effects on the Rho-like GTPase cdc42 (cell division cycle 42). This study investigates PTEN-dependent morphogenesis in a CRC model. Stable short hairpin RNA knockdown of PTEN in Caco-2 cells influenced expression or localization of cdc42 guanine nucleotide exchange factors and inhibited cdc42 activation. Parental Caco-2 cells formed regular hollow gland-like structures (glands) with a single central lumen, in three-dimensional (3D) cultures. Conversely, PTEN-deficient Caco-2 ShPTEN cells formed irregular glands with multiple abnormal lumens as well as intra- and/or intercellular vacuoles evocative of the high-grade CRC phenotype. Effects of targeted treatment were investigated. Phosphatidinylinositol 3-kinase (PI3K) modulating treatment did not affect gland morphogenesis but did influence gland number, gland size and/or cell size within glands. As PTEN may be regulated by the nuclear receptor peroxisome proliferator-activated receptor-? (PPAR?), cultures were treated with the PPAR? ligand rosiglitazone. This treatment enhanced PTEN expression, cdc42 activation and rescued dysmorphogenesis by restoring single lumen formation in Caco-2 ShPTEN glands. Rosiglitazone effects on cdc42 activation and Caco-2 ShPTEN gland development were attenuated by cotreatment with GW9662, a PPAR? antagonist. Taken together, these studies show PTEN-cdc42 regulation of lumen formation in a 3D model of human CRC glandular morphogenesis. Treatment by the PPAR? ligand rosiglitazone, but not PI3K modulators, rescued colorectal glandular dysmorphogenesis of PTEN deficiency.

Long-term genomic instability in human lymphocytes induced by single-particle irradiation

Recent evidence suggests that genomic instability, which is an important step in carcinogenesis, may be important in the effectiveness of radiation as a carcinogen, particularly for high-LET radiations. Understanding the biological effects underpinning the risks associated with low doses of densely ionizing radiations is complicated in experimental systems by the Poisson distribution of particles that ran be delivered, In this study, we report an approach to determine the effect of the lowest possible cellular radiation dose of densely ionizing at particles, that of a single particle traversal. Using microbeam technology and an approach for immobilizing human T-lymphocytes, we have measured the effects of single alpha -particle traversals on the surviving progeny of cells. A significant increase in the proportion of aberrant cells is observed 12-13 population doublings after exposure, with a high level of chromatid-type aberrations, indicative of an instability phenotype, These data suggest that instability may be important in situations where even a single particle traverses human cells. (C) 2001 by Radiation Research Society.