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Sample records for ras alters lens

  1. Altered expression of Bcl-2, c-Myc, H-Ras, K-Ras, and N-Ras does not influence the course of mycosis fungoides

    PubMed Central

    Maj, Joanna; Jankowska-Konsur, Alina; Plomer-Niezgoda, Ewa; Sadakierska-Chudy, Anna

    2013-01-01

    Introduction Data about genetic alterations in mycosis fungoides (MF) are limited and their significance not fully elucidated. The aim of the study was to explore the expression of various oncogenes in MF and to assess their influence on the disease course. Material and methods Skin biopsies from 27 MF patients (14 with early MF and 13 with advanced disease) and 8 healthy volunteers were analyzed by real-time polymerase chain reaction (PCR) to detect Bcl-2, c-Myc, H-Ras, K-Ras and N-Ras expression. All PCR reactions were performed using an Applied Biosystems 7900HT Fast Real-Time PCR System and interpreted using Sequence Detection Systems software which utilizes the comparative delta Ct method. The level of mRNA was normalized to GAPDH expression. All data were analyzed statistically. Results All evaluated oncogenes were found to be expressed in the skin from healthy controls and MF patients. Bcl-2 (–4.2 ±2.2 vs. –2.2 ±1.1; p = 0.01), H-Ras (–3.0 ±3.3 vs. 0.6 ±2.6; p = 0.01) and N-Ras (–3.6 ±2.0 vs. –1.1 ±2.4; p = 0.03) were expressed at significantly lower levels in MF. No relationships between oncogene expression and disease stage, presence of distant metastases and survival were observed (p > 0.05 for all comparisons). Conclusions The pathogenic role and prognostic significance of analyzed oncogenes in MF seem to be limited and further studies are needed to establish better prognostic factors for patients suffering from MF. PMID:24273576

  2. Ras-mediated cell cycle arrest is altered by nuclear oncogenes to induce Schwann cell transformation.

    PubMed Central

    Ridley, A J; Paterson, H F; Noble, M; Land, H

    1988-01-01

    The cellular responses to ras and nuclear oncogenes were investigated in purified populations of rat Schwann cells. v-Ha-ras and SV40 large T cooperate to transform Schwann cells, inducing growth in soft agar and allowing proliferation in the absence of added mitogens. Expression of large T alone reduces their growth factor requirements but is insufficient to induce full transformation. In contrast, expression of v-Ha-ras leads to proliferation arrest in Schwann cells expressing a temperature-sensitive mutant of large T at the restrictive temperature. Cells arrest in either the G1 or G2/M phases of the cell cycle, and can re-enter cell division at the permissive temperature even after prolonged periods at the restrictive conditions. Oncogenic ras proteins also inhibit DNA synthesis when microinjected into Schwann cells. Adenovirus E1a and c-myc oncogenes behave similarly to SV40 large T. They cooperate with Ha-ras oncogenes to transform Schwann cells, and prevent ras-induced growth arrest. Thus nuclear oncogenes fundamentally alter the response of Schwann cells to a ras oncogene from cell cycle arrest to transformation. Images PMID:3049071

  3. Alteration of glycolipids in ras-transfected NIH 3T3 cells

    SciTech Connect

    Matyas, G.R.; Aaronson, S.A.; Brady, R.O.; Fishman, P.H.

    1987-09-01

    Glycosphingolipid alterations upon viral transformation are well documented. Transformation of mouse 3T3 cells with murine sarcoma viruses results in marked decreases in the levels of gangliosides GM1 and GD1a and an increase in gangliotriaosylceramide. The transforming oncogenes of these viruses have been identified as members of the ras gene family. The authors analyzed NIH 3T3 cells transfected with human H-, K- and N-ras oncogenes for their glycolipid composition and expression of cell surface gangliosides. Using conventional thin-layer chromatographic analysis, they found that the level of GM3 was increased and that of GD1a was slightly decreased or unchanged, and GM1 was present but not in quantifiable levels. Cell surface levels of GM1 were determined by /sup 125/I-labeled cholera toxin binding to intact cells. GD1a was determined by cholera toxin binding to cells treated with sialidase prior to toxin binding. All ras-transfected cells had decreased levels of surface GM1 and GD1 as compared to logarithmically growing normal NIH 3T3 cells. Levels of GM1 and, to a lesser extent, GD1a increased as the latter cells became confluent. Using a monoclonal antibody assay, they found that gangliotriaosylceramide was present in all ras-transfected cells studied but not in logarithmically growing untransfected cells. These results indicated that ras oncogenes derived form human tumors are capable of inducing alterations in glycolipid composition.

  4. Alterations in the K-ras and p53 genes in rat lung tumors

    SciTech Connect

    Belinsky, S.A.; Swafford, D.S.; Finch, G.L.; Mitchell, C.E.

    1997-06-01

    Activation of the K-ras protooncogene and inactivation of the p53 tumor suppressor gene are events common to many types of human cancers. Molecular epidemiology studies have associated mutational profiles in these genes with specific exposures. The purpose of this paper is to review investigations that have examined the role of the K-ras and p53 genes in lung tumors induced in the F344 rat by mutagenic and nonmutagenic exposures. Mutation profiles within the K-ras and p53 genes, if present in rat lung tumors, would help to define some of the molecular mechanisms underlying cancer induction by various environmental agents. Pulmonary adenocarcinomas or squamous cell carcinomas were induced by tetranitromethane (TNM), 4-methylnitrosamino-1-(3-pyridyl)-1-butanone (NNK), beryllium metal, plutonium-239, X-ray, diesel exhaust, or carbon black. These agents were chosen because the tumors they produced could arise via different types of DNA damage. Mutation of the K-ras gene was determined by approaches that included DNA transfection, direct sequencing, mismatch hybridization, and restriction fragment length polymorphism analysis. The frequency for mutation of the K-ras gene was exposure dependent. The transition mutations formed could have been derived from deamination of cytosine. Alteration in the p53 gene was assessed by immunohistochemical analysis for p53 protein and single-strand conformation polymorphism (SSCP) analysis of exons 4 to 9. None of the 93 adenocarinomas examined was immunoreactive toward the anti-p53 antibody CM1. In contrast, 14 of 71 squamous cell carcinomas exhibited nuclear p53 immunoreactivity with no correlation to type of exposure. However, SSCP analysis only detected mutations in 2 of 14 squamous cell tumors that were immunoreactive, suggesting that protein stabilization did not stem from mutations within the p53 gene. Thus, the p53 gene does not appear to be involved in the genesis of most rat lung tumors. 2 figs., 2 tabs., 48 refs.

  5. Expression of ras oncogenes in cultured human cells alters the transcriptional and posttranscriptional regulation of cytokine genes.

    PubMed Central

    Demetri, G D; Ernst, T J; Pratt, E S; Zenzie, B W; Rheinwald, J G; Griffin, J D

    1990-01-01

    Autonomous production of cytokines such as the hematopoietic colony-stimulating factors (CSFs), IL-1, or IL-6 has been demonstrated in numerous human and murine neoplasms, and may be involved in the pathogenesis of several paraneoplastic syndromes such as leukocytosis, fever, and hypercalcemia. Because of the high frequency with which mutations in ras protooncogenes have been detected in human tumors, as well as evidence linking ras gene products to activation of certain cellular functions, we investigated whether ras mutations might influence the regulation of cytokine genes. Normal human fibroblasts transfected with a mutant val12 H-ras oncogene expressed increased levels of mRNA transcripts encoding granulocyte-CSF (G-CSF), granulocyte-macrophage-CSF (GM-CSF), and IL-1 beta compared with controls. Human mesothelioma cells transfected with a mutant asp12 N-ras oncogene exhibited similar alterations in cytokine gene expression. Estimates of transcriptional activity by nuclear run-on analysis revealed a selective increase in transcription only for the IL-1 gene. Analysis of mRNA half-life demonstrated a marked increase in the stability of numerous cytokine transcripts, including G-CSF, GM-CSF, IL-1, and IL-6. The addition of anti-IL-1 neutralizing antibody to cultures of cells expressing ras mutants did not block the expression of any of the cytokines examined, suggesting that the baseline expression of GM-CSF, G-CSF, and IL-6 was not a secondary event due to the increased transcription of IL-1. These results indicate that mutations in ras genes may alter expression of several cytokine genes through both transcriptional and posttranscriptional mechanisms. Images PMID:2212010

  6. Molecular kinetics. Ras activation by SOS: allosteric regulation by altered fluctuation dynamics.

    PubMed

    Iversen, Lars; Tu, Hsiung-Lin; Lin, Wan-Chen; Christensen, Sune M; Abel, Steven M; Iwig, Jeff; Wu, Hung-Jen; Gureasko, Jodi; Rhodes, Christopher; Petit, Rebecca S; Hansen, Scott D; Thill, Peter; Yu, Cheng-Han; Stamou, Dimitrios; Chakraborty, Arup K; Kuriyan, John; Groves, Jay T

    2014-07-01

    Activation of the small guanosine triphosphatase H-Ras by the exchange factor Son of Sevenless (SOS) is an important hub for signal transduction. Multiple layers of regulation, through protein and membrane interactions, govern activity of SOS. We characterized the specific activity of individual SOS molecules catalyzing nucleotide exchange in H-Ras. Single-molecule kinetic traces revealed that SOS samples a broad distribution of turnover rates through stochastic fluctuations between distinct, long-lived (more than 100 seconds), functional states. The expected allosteric activation of SOS by Ras-guanosine triphosphate (GTP) was conspicuously absent in the mean rate. However, fluctuations into highly active states were modulated by Ras-GTP. This reveals a mechanism in which functional output may be determined by the dynamical spectrum of rates sampled by a small number of enzymes, rather than the ensemble average.

  7. Classic Ras Proteins Promote Proliferation and Survival Via Distinct Phosphoproteome Alterations in Neurofibromin-Null Malignant Peripheral Nerve Sheath Tumor Cells

    PubMed Central

    Brossier, Nicole M.; Prechtl, Amanda M.; Longo, Jody Fromm; Barnes, Stephen; Wilson, Landon S.; Byer, Stephanie J.; Brosius, Stephanie N.; Carroll, Steven L.

    2015-01-01

    Neurofibromin, the tumor suppressor encoded by the neurofibromatosis type 1 (NF1) gene, potentially suppresses the activation of H-Ras, N-Ras and K-Ras. However, it is not known whether these classic Ras proteins are hyperactivated in NF1-null nerve sheath tumors, how they contribute to tumorigenesis and what signaling pathways mediate their effects. Here we show that H-Ras, N-Ras and K-Ras are coexpressed with their activators, (guanine nucleotide exchange factors), in neurofibromin-null malignant peripheral nerve sheath tumor (MPNST) cells and that all 3 Ras proteins are activated. Dominant negative (DN) H-Ras, a pan-inhibitor of the classic Ras family, inhibited MPNST proliferation and survival, but not migration. However, NF1-null MPNST cells were variably dependent on individual Ras proteins. In some lines, ablation of H-Ras, N-Ras and/or K-Ras inhibited mitogenesis. In others, ablation of a single Ras protein had no effect on proliferation; in these lines, ablation of a single Ras protein resulted in compensatory increases in the activation and/or expression of other Ras proteins. Using mass spectrometry-based phosphoproteomics, we identified 7 signaling networks affecting morphology, proliferation and survival that are regulated by DN H-Ras. Thus, neurofibromin loss activates multiple classic Ras proteins that promote proliferation and survival by regulating several distinct signaling cascades. PMID:25946318

  8. Classic Ras Proteins Promote Proliferation and Survival via Distinct Phosphoproteome Alterations in Neurofibromin-Null Malignant Peripheral Nerve Sheath Tumor Cells.

    PubMed

    Brossier, Nicole M; Prechtl, Amanda M; Longo, Jody Fromm; Barnes, Stephen; Wilson, Landon S; Byer, Stephanie J; Brosius, Stephanie N; Carroll, Steven L

    2015-06-01

    Neurofibromin, the tumor suppressor encoded by the neurofibromatosis type 1 (NF1) gene, potentially suppresses the activation of H-Ras, N-Ras, and K-Ras. However, it is not known whether these classic Ras proteins are hyperactivated in NF1-null nerve sheath tumors, how they contribute to tumorigenesis, and what signaling pathways mediate their effects. Here we show that H-Ras, N-Ras, and K-Ras are coexpressed with their activators (guanine nucleotide exchange factors) in neurofibromin-null malignant peripheral nerve sheath tumor (MPNST) cells, and that all 3 Ras proteins are activated. Dominant negative (DN) H-Ras, a pan-inhibitor of the classic Ras family, inhibited MPNST proliferation and survival, but not migration. However, NF1-null MPNST cells were variably dependent on individual Ras proteins. In some lines, ablation of H-Ras, N-Ras, and/or K-Ras inhibited mitogenesis. In others, ablation of a single Ras protein had no effect on proliferation; in these lines, ablation of a single Ras protein resulted in compensatory increases in the activation and/or expression of other Ras proteins. Using mass spectrometry-based phosphoproteomics, we identified 7 signaling networks affecting morphology, proliferation, and survival that are regulated by DN H-Ras. Thus, neurofibromin loss activates multiple classic Ras proteins that promote proliferation and survival by regulating several distinct signaling cascades.

  9. The structural alteration and aggregation propensity of glycated lens crystallins in the presence of calcium: Importance of lens calcium homeostasis in development of diabetic cataracts.

    PubMed

    Zm, Sara Zafaranchi; Khoshaman, Kazem; Masoudi, Raheleh; Hemmateenejad, Bahram; Yousefi, Reza

    2017-01-01

    The imbalance of the calcium homeostasis in the lenticular tissues of diabetic patients is an important risk factor for development of cataract diseases. In the current study, the impact of elevated levels of calcium ions were investigated on structure and aggregation propensity of glycated lens crystallins using gel electrophoresis and spectroscopic assessments. The glycated proteins indicated significant resistance against calcium-induced structural insults and aggregation. While, glycated crystallins revealed an increased conformational stability; a slight instability was observed for these proteins upon interaction with calcium ions. Also, in the presence of calcium, the proteolytic pattern of native crystallins was altered and that of glycated protein counterparts remained almost unchanged. According to results of this study it is suggested that the structural alteration of lens crystallins upon glycation may significantly reduce their calcium buffering capacity in eye lenses. Therefore, under chronic hyperglycemia accumulation of this cataractogenic metal ion in the lenticular tissues may subsequently culminate in activation of different pathogenic pathways, leading to development of lens opacity and cataract diseases. PMID:27434877

  10. Newborn mouse lens proteome and its alteration by lysine 6 mutant ubiquitin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ubiquitin is a tag that often initiates degradation of proteins by the proteasome in the ubiquitin proteasome system. Targeted expression of K6W mutant ubiquitin (K6W-Ub) in the lens results in defects in lens development and cataract formation, suggesting critical functions for ubiquitin in lens. T...

  11. H-ras-transformed NRK-52E renal epithelial cells have altered growth, morphology, and cytoskeletal structure that correlates with renal cell carcinoma in vivo.

    PubMed

    Best, C J; Tanzer, L R; Phelps, P C; Merriman, R L; Boder, G G; Trump, B F; Elliget, K A

    1999-04-01

    We studied the effect of the ras oncogene on the growth kinetics, morphology, cytoskeletal structure, and tumorigenicity of the widely used NRK-52E rat kidney epithelial cell line and two H-ras oncogene-transformed cell lines, H/1.2-NRK-52E (H/1.2) and H/6.1-NRK-52E (H/6.1). Population doubling times of NRK-52E, H/1.2, and H/6.1 cells were 28, 26, and 24 h, respectively, with the transformed cells reaching higher saturation densities than the parent cells. NRK-52E cells had typical epithelial morphology with growth in colonies. H/1.2 and H/6.1 cell colonies were more closely packed, highly condensed, and had increased plasma membrane ruffling compared to parent cell colonies. NRK-52E cells showed microfilament, microtubule, and intermediate filament networks typical of epithelial cells, while H/1.2 and H/6.1 cells showed altered cytoskeleton architecture, with decreased stress fibers and increased microtubule and intermediate filament staining at the microtubule organizing center. H/1.2 and H/6.1 cells proliferated in an in vitro soft agar transformation assay, indicating anchorage-independence, and rapidly formed tumors in vivo with characteristics of renal cell carcinoma, including mixed populations of sarcomatoid, granular, and clear cells. H/6.1 cells consistently showed more extensive alterations of growth kinetics, morphology, and cytoskeleton than H/1.2 cells, and formed tumors of a more aggressive phenotype. These data suggest that analysis of renal cell characteristics in vitro may have potential in predicting tumor behavior in vivo, and significantly contribute to the utility of these cell lines as in vitro models for examining renal epithelial cell biology and the role of the ras proto-oncogene in signal transduction involving the cytoskeleton.

  12. Ras history

    PubMed Central

    2010-01-01

    Although the roots of Ras sprouted from the rich history of retrovirus research, it was the discovery of mutationally activated RAS genes in human cancer in 1982 that stimulated an intensive research effort to understand Ras protein structure, biochemistry and biology. While the ultimate goal has been developing anti-Ras drugs for cancer treatment, discoveries from Ras have laid the foundation for three broad areas of science. First, they focused studies on the origins of cancer to the molecular level, with the subsequent discovery of genes mutated in cancer that now number in the thousands. Second, elucidation of the biochemical mechanisms by which Ras facilitates signal transduction established many of our fundamental concepts of how a normal cell orchestrates responses to extracellular cues. Third, Ras proteins are also founding members of a large superfamily of small GTPases that regulate all key cellular processes and established the versatile role of small GTP-binding proteins in biology. We highlight some of the key findings of the last 28 years. PMID:21686117

  13. Proton Irradiation Alters Expression of FGF-2 In Human Lens Epithelial Cells

    NASA Technical Reports Server (NTRS)

    Blakely, E. A.; Bjornstad, K. A.; Chang, P. Y.; McNamara, M. P.; Chang, E.

    1999-01-01

    We are investigating a role for proton radiation-induced changes in FGF-2 gene expression as part of the mechanism(s) underlying lens cell injury. Radiation injury to the human lens is associated with the induction of cataract following exposure to protons.

  14. Ras in Cancer and Developmental Diseases

    PubMed Central

    Fernández-Medarde, Alberto; Santos, Eugenio

    2011-01-01

    Somatic, gain-of-function mutations in ras genes were the first specific genetic alterations identified in human cancer about 3 decades ago. Studies during the last quarter century have characterized the Ras proteins as essential components of signaling networks controlling cellular proliferation, differentiation, or survival. The oncogenic mutations of the H-ras, N-ras, or K-ras genes frequently found in human tumors are known to throw off balance the normal outcome of those signaling pathways, thus leading to tumor development. Oncogenic mutations in a number of other upstream or downstream components of Ras signaling pathways (including membrane RTKs or cytosolic kinases) have been detected more recently in association with a variety of cancers. Interestingly, the oncogenic Ras mutations and the mutations in other components of Ras/MAPK signaling pathways appear to be mutually exclusive events in most tumors, indicating that deregulation of Ras-dependent signaling is the essential requirement for tumorigenesis. In contrast to sporadic tumors, separate studies have identified germline mutations in Ras and various other components of Ras signaling pathways that occur in specific association with a number of different familial, developmental syndromes frequently sharing common phenotypic cardiofaciocutaneous features. Finally, even without being a causative force, defective Ras signaling has been cited as a contributing factor to many other human illnesses, including diabetes and immunological and inflammatory disorders. We aim this review at summarizing and updating current knowledge on the contribution of Ras mutations and altered Ras signaling to development of various tumoral and nontumoral pathologies. PMID:21779504

  15. RAS Laboratory Groups

    Cancer.gov

    The RAS Initiative uses multiple technologies to attack RAS-driven cancers. The resources of the Frederick National Lab allocated to the RAS Hub are organized into seven laboratory groups, each contributing to the collaborative effort.

  16. The RAS Initiative

    Cancer.gov

    NCI established the RAS Initiative to explore innovative approaches for attacking the proteins encoded by mutant forms of RAS genes and to ultimately create effective, new therapies for RAS-related cancers.

  17. Regulation of Ras proteins by reactive nitrogen species.

    PubMed

    Davis, Michael F; Vigil, Dom; Campbell, Sharon L

    2011-08-01

    Ras GTPases have been a subject of intense investigation since the early 1980s, when single point mutations in Ras were shown to cause deregulated cell growth control. Subsequently, Ras was identified as the most prevalent oncogene found in human cancer. Ras proteins regulate a host of pathways involved in cell growth, differentiation, and apoptosis by cycling between inactive GDP-bound and active GTP-bound states. Regulation of Ras activity is controlled by cellular factors that alter guanine nucleotide cycling. Oncogenic mutations prevent protein regulatory factors from down-regulating Ras activity, thereby maintaining Ras in a chronically activated state. The central dogma in the field is that protein modulatory factors are the primary regulators of Ras activity. Since the mid-1990s, however, evidence has accumulated that small molecule reactive nitrogen species (RNS) can also influence Ras guanine nucleotide cycling. Herein, we review the basic chemistry behind RNS formation and discuss the mechanism through which various RNS enhance nucleotide exchange in Ras proteins. In addition, we present studies that demonstrate the physiological relevance of RNS-mediated Ras activation within the context of immune system function, brain function, and cancer development. We also highlight future directions and experimental methods that may enhance our ability to detect RNS-mediated activation in cell cultures and in vivo. The development of such methods may ultimately pave new directions for detecting and elucidating how Ras proteins are regulated by redox species, as well as for targeting redox-activated Ras in cancer and other disease states.

  18. Two types of RAS mutants that dominantly interfere with activators of RAS.

    PubMed Central

    Jung, V; Wei, W; Ballester, R; Camonis, J; Mi, S; Van Aelst, L; Wigler, M; Broek, D

    1994-01-01

    In the fission yeast Schizosaccharomyces pombe, ras1 regulates both sexual development (conjugation and sporulation) and cellular morphology. Two types of dominant interfering mutants were isolated in a genetic screen for ras1 mutants that blocked sexual development. The first type of mutation, at Ser-22, analogous to the H-rasAsn-17 mutant (L. A. Feig and G. M. Cooper, Mol. Cell. Biol. 8:3235-3243, 1988), blocked only conjugation, whereas a second type of mutation, at Asp-62, interfered with conjugation, sporulation, and cellular morphology. Analogous mutations at position 64 of Saccharomyces cerevisiae RAS2 or position 57 of human H-ras also resulted in dominant interfering mutants that interfered specifically and more profoundly than mutants of the first type with RAS-associated pathways in both S. pombe or S. cerevisiae. Genetic evidence indicating that both types of interfering mutants function upstream of RAS is provided. Biochemical evidence showing that the mutants are altered in their interaction with the CDC25 class of exchange factors is presented. We show that both H-rasAsn-17 and H-rasTyr-57, compared with wild-type H-ras, are defective in their guanine nucleotide-dependent release from human cdc25 and that this defect is more severe for the H-rasTyr-57 mutant. Such a defect would allow the interfering mutants to remain bound to, thereby sequestering RAS exchange factors. The more severe interference phenotype of this novel interfering mutant suggests that it functions by titrating out other positive regulators of RAS besides those encoded by ste6 and CDC25. Images PMID:8196614

  19. RAS - Screens & Assays

    Cancer.gov

    A primary goal of the RAS Initiative is to develop assays for RAS activity, localization, and signaling and adapt those assays so they can be used for finding new drug candidates. Explore the work leading to highly validated screening protocols.

  20. RAS Initiative - Community Outreach

    Cancer.gov

    Through community and technical collaborations, workshops and symposia, and the distribution of reference reagents, the RAS Initiative seeks to increase the sharing of knowledge and resources essential to defeating cancers caused by mutant RAS genes.

  1. RAS Initiative - Events

    Cancer.gov

    The NCI RAS Initiative has organized multiple events with outside experts to discuss how the latest scientific and technological breakthroughs can be applied to discover vulnerabilities in RAS-driven cancers.

  2. Mechanical Aqueous Alteration Dominates Textures of Gale Crater Rocks: Mars Hand Lens Imager (MAHLI) Results

    NASA Astrophysics Data System (ADS)

    Aileen Yingst, R.; Minitti, Michelle; Edgett, Kenneth; McBride, Marie; Stack, Kathryn

    2015-04-01

    The Mars Hand Lens Imager (MAHLI) acquired sub-mm/pixel scale color images of over 70 individual rocks and outcrops during Curiosity's first year on Mars, permitting the study of textures down to the distinction between silt and very fine sand. We group imaged rock textures into classes based on their grain size, sorting, matrix characteristics, and abundance of pores. Because the recent campaign at Pahrump Hills acquired many more MAHLI images than elsewhere along the rover traverse [6], textural analysis there is more detailed and thus types observed there are sub-divided. Mudstones: These rocks contain framework grains smaller than the highest resolution MAHLI images (16 μm/pixel), and thus are interpreted to consist of grains that are silt-sized or smaller. Some rocks contain nodules, sulfate veins, and Mg-enriched erosionally-resistant ridges. The Pahrump Hills region contains mudstones of at least four different sub-textures: recessive massive, recessive parallel-laminated, resistant laminated-to-massive, and resistant cross-stratified. Recessive mudstones are slope-forming; parallel-laminated recessive mudstones display mm-scale parallel (and in some cases rhythmic) lamination that extends laterally for many meters, and are interbedded with recessive massive mudstones. Coarse cm- to mm-scale laminae appear within resistant mudstones though some portions are more massive; laminae tend to be traceable for cm to meters. Well-sorted sandstones: Rocks in this class are made of gray, fine-to-medium sand and exhibit little to no porosity. Two examples of this class show fine lineations with sub-mm spacing. Aillik, a target in the Shaler outcrop, shows abundant cross-lamination. The Pahrump Hills region contains a sub-texture of well-sorted, very fine to fine-grained cross-stratified sandstone at the dune and ripple-scale. Poorly-sorted sandstones. This class is subdivided into two sub-classes: rounded, coarse-to-very coarse sand grains of variable colors and

  3. The Zonules Selectively Alter the Shape of the Lens During Accommodation Based on the Location of Their Anchorage Points

    PubMed Central

    Nankivil, Derek; Maceo Heilman, Bianca; Durkee, Heather; Manns, Fabrice; Ehrmann, Klaus; Kelly, Shawn; Arrieta-Quintero, Esdras; Parel, Jean-Marie

    2015-01-01

    Purpose. To determine the role of anterior and posterior zonular tension on the optomechanical lens response during accommodation simulation. Methods. Ten eyes from nine hamadryas baboons (4.9 ± 0.7 years) and 20 eyes from 18 cynomolgus monkeys (5.4 ± 0.3 years) were dissected, leaving the lens, zonules, ciliary body, hyaloid membrane, anterior vitreous, and a segmented scleral rim intact. The lens preparation was mounted in a lens stretcher, and the outer scleral shell was displaced radially in a stepwise fashion. The load, lens, and ciliary body diameters, lens power, lens thickness, and the anterior and posterior radius of curvature were measured during stretching. The zonular fibers attached to either the posterior or anterior lens surface were then carefully transected and the experiment was repeated. Zonular transection was confirmed in four eyes via laser scanning confocal microscopy after immunostaining. The effect of zonular transection on the tissue response to stretching was quantified. Results. Without anterior zonules, 48% and 97% of the changes in anterior and posterior radii are retained. Without posterior zonules, 81% and 67% of the changes in anterior and posterior radii are retained. The changes in lens shape were reduced after transecting either the anterior or posterior zonules; however, both surfaces still changed shape. Conclusions. While either the anterior or posterior zonules alone are capable of changing the shape of both lens surfaces, the anterior zonules have a greater effect on the anterior lens surface, and the posterior zonules have a greater effect on the posterior lens surface. PMID:25698707

  4. RAS - Target Identification - Informatics

    Cancer.gov

    The RAS Informatics lab group develops tools to track and analyze “big data” from the RAS Initiative, as well as analyzes data from external projects. By integrating internal and external data, this group helps improve understanding of RAS-driven cancers.

  5. Absolute Quantification of Endogenous Ras Isoform Abundance

    PubMed Central

    Mageean, Craig J.; Griffiths, John R.; Smith, Duncan L.; Clague, Michael J.; Prior, Ian A.

    2015-01-01

    Ras proteins are important signalling hubs situated near the top of networks controlling cell proliferation, differentiation and survival. Three almost identical isoforms, HRAS, KRAS and NRAS, are ubiquitously expressed yet have differing biological and oncogenic properties. In order to help understand the relative biological contributions of each isoform we have optimised a quantitative proteomics method for accurately measuring Ras isoform protein copy number per cell. The use of isotopic protein standards together with selected reaction monitoring for diagnostic peptides is sensitive, robust and suitable for application to sub-milligram quantities of lysates. We find that in a panel of isogenic SW48 colorectal cancer cells, endogenous Ras proteins are highly abundant with ≥260,000 total Ras protein copies per cell and the rank order of isoform abundance is KRAS>NRAS≥HRAS. A subset of oncogenic KRAS mutants exhibit increased total cellular Ras abundance and altered the ratio of mutant versus wild type KRAS protein. These data and methodology are significant because Ras protein copy number is required to parameterise models of signalling networks and informs interpretation of isoform-specific Ras functional data. PMID:26560143

  6. Ras moves to stay in place.

    PubMed

    Schmick, Malte; Kraemer, Astrid; Bastiaens, Philippe I H

    2015-04-01

    Ras is a major intracellular signaling hub. This elevated position comes at a precarious cost: a single point mutation can cause aberrant signaling. The capacity of Ras for signaling is inextricably linked to its enrichment at the plasma membrane (PM). This PM localization is dynamically maintained by three essential elements: alteration of membrane affinities via lipidation and membrane-interaction motifs; trapping on specific membranes coupled with unidirectional vesicular transport to the PM; and regulation of diffusion via interaction with a solubilization factor. This system constitutes a cycle that primarily corrects for the entropic equilibration of Ras to all membranes that dilutes its signaling capacity. We illuminate how this reaction-diffusion system maintains an out-of-equilibrium localization of Ras GTPases and thereby confers signaling functionality to the PM.

  7. Metabolic Dependencies in RAS-Driven Cancers.

    PubMed

    Kimmelman, Alec C

    2015-04-15

    The ability to inhibit the RAS oncogene has been the holy grail of oncology because of the critical role of this gene in a multitude of tumor types. In addition, RAS-mutant tumors are among the most aggressive and refractory to treatment. Although directly targeting the RAS oncogene has proven challenging, an alternative approach for treating RAS-driven cancers is to inhibit critical downstream events that are required for tumor maintenance. Indeed, much focus has been put on inhibiting signaling cascades downstream of RAS. Recent studies have shown that oncogenic RAS promotes a metabolic reprogramming of tumor cells, shifting them toward an anabolic metabolism necessary to produce biomass to support unconstrained proliferation. These cancers also use a diverse set of fuel sources to meet their metabolic needs and have even developed a variety of mechanisms to act as metabolic scavengers to obtain necessary metabolic substrates from both extracellular and intracellular sources. Collectively, these adaptations can create "metabolic bottlenecks" whereby tumor cells rely on particular pathways or rate-limiting metabolites. In this regard, inhibiting individual or combinations of these metabolic pathways can attenuate growth in preclinical models. Because these dependencies are tumor selective and downstream of oncogenic RAS, there is the opportunity for therapeutic intervention. Although targeting tumor metabolism is still in the early days of translation to patients, our continued advances in understanding critical metabolic adaptations in RAS-driven cancers, as well as the ability to study this altered metabolism in relevant tumor models, will accelerate the development of new therapeutic approaches. Clin Cancer Res; 21(8); 1828-34. ©2015 AACR. See all articles in this CCR Focus section, "Targeting RAS-Driven Cancers." PMID:25878364

  8. An orthosteric inhibitor of the RAS-SOS interaction.

    PubMed

    Nickerson, Seth; Joy, Stephen T; Arora, Paramjit S; Bar-Sagi, Dafna

    2013-01-01

    Rat sarcoma (RAS) proteins are signaling nodes that transduce extracellular cues into precise alterations in cellular physiology by engaging effector pathways. RAS signaling thus regulates diverse cell processes including proliferation, migration, differentiation, and survival. Owing to this central role in governing mitogenic signals, RAS pathway components are often dysregulated in human diseases. Targeted therapy of RAS pathways has generally not been successful, largely because of the robust biochemistry of the targets and their multifaceted network of molecular regulators. The rate-limiting step of RAS activation is Son of Sevenless (SOS)-mediated nucleotide exchange involving a single evolutionarily conserved catalytic helix from SOS. Structure function data of this mechanism provided a strong platform to design an SOS-derived, helically constrained peptide mimic as an inhibitor of the RAS-SOS interaction. In this chapter, we review RAS-SOS signaling dynamics and present evidence supporting the novel paradigm of inhibiting their interaction as a therapeutic strategy. We then describe a method of generating helically constrained peptide mimics of protein surfaces, which we have employed to inhibit the RAS-SOS active site interaction. The biochemical and functional properties of this SOS mimic support the premise that inhibition of RAS-nucleotide exchange can effectively block RAS activation and downstream signaling.

  9. CDK4 coexpression with Ras generates malignant human epidermal tumorigenesis.

    PubMed

    Lazarov, Mirella; Kubo, Yoshiaki; Cai, Ti; Dajee, Maya; Tarutani, Masahito; Lin, Qun; Fang, Min; Tao, Shiying; Green, Cheryl L; Khavari, Paul A

    2002-10-01

    Ras acts with other proteins to induce neoplasia. By itself, however, strong Ras signaling can suppress proliferation of normal cells. In primary epidermal cells, we found that oncogenic Ras transiently decreases cyclin-dependent kinase (CDK) 4 expression in association with cell cycle arrest in G1 phase. CDK4 co-expression circumvents Ras growth suppression and induces invasive human neoplasia resembling squamous cell carcinoma. Tumorigenesis is dependent on CDK4 kinase function, with cyclin D1 required but not sufficient for this process. In facilitating escape from G1 growth restraints, Ras and CDK4 alter the composition of cyclin D and cyclin E complexes and promote resistance to growth inhibition by INK4 cyclin-dependent kinase inhibitors. These data identify a new role for oncogenic Ras in CDK4 regulation and highlight the functional importance of CDK4 suppression in preventing uncontrolled growth.

  10. Regulation of Ras Proteins by Reactive Nitrogen Species†

    PubMed Central

    Davis, Michael F.; Vigil, Dom; Campbell, Sharon L.

    2012-01-01

    Ras GTPases have been a subject of intense investigation since the early-80’s, when single point mutations in Ras were shown to cause deregulated cell growth control. Subsequently, Ras was identified as the most prevalent oncogene found in human cancer. Ras proteins regulate a host of pathways involved in cell growth, differentiation, and apoptosis by cycling between inactive GDP-bound and active GTP-bound states. Regulation of Ras activity is controlled by cellular factors that alter guanine nucleotide cycling. Oncogenic mutations prevent protein regulatory factors from down-regulating Ras activity, thereby maintaining Ras in a chronically activated state. The central dogma in the field is that protein modulatory factors are the primary regulators of Ras activity. Since the mid-90’s, however, evidence has accumulated that small molecule reactive nitrogen species (RNS) can also influence Ras guanine nucleotide cycling. Herein, we review the basic chemistry behind RNS formation and discuss the mechanism through which various RNS enhance nucleotide exchange in Ras proteins. In addition, we present studies that demonstrate the physiological relevance of RNS-mediated Ras activation within the context of immune system function, brain function, and cancer development. We also highlight future directions and experimental methods that may enhance our ability to detect RNS-mediated activation in cell cultures and in vivo. The development of such methods may ultimately pave new directions for detecting and elucidating how Ras proteins are regulated by redox species, as well as for targeting redox-activated Ras in cancer and other disease states. PMID:21616138

  11. The RAS Problem

    Cancer.gov

    More than 30% of all human cancers, including a high percentage of lung and colon cancers and 95% of pancreatic cancers are driven by mutations and possibly amplification (increased copies) of RAS genes.

  12. Adenovirus-mediated gene transfer of dominant negative ras(asn17) in 3T3L1 adipocytes does not alter insulin-stimulated P13-kinase activity or glucose transport.

    PubMed

    Gnudi, L; Frevert, E U; Houseknecht, K L; Erhardt, P; Kahn, B B

    1997-01-01

    Recent studies suggest that the ras-map kinase and PI3-kinase cascades converge. We sought to determine whether PI3-kinase is downstream of ras in insulin signaling in a classic insulin target cell. We generated a recombinant adenovirus encoding dominant negative ras by cloning the human H-ras cDNA with a ser to asn substitution at amino acid 17 (ras(asn17)) into the pACCMVpLpA vector and cotransfecting 293 cells with the pJM17 plasmid containing the adenoviral genome. Efficiency of gene transfer was assessed by infecting fully differentiated 3T3L1 adipocytes with a recombinant adenovirus expressing beta-galactosidase (beta-gal); greater than 70% of cells were infected. Infection of adipocytes with ras(asn17) resulted in 10-fold greater expression than endogenous ras. This high efficiency gene transfer allowed biochemical assays. Insulin stimulation of ras-GTP formation was inhibited in ras(asn17)-expressing cells. Map kinase gel mobility shift revealed that insulin (1 UM) or epidermal growth factor (100 ng/ml) resulted in the appearance of a hyperphosphorylated species of p42 map kinase in uninfected cells and those expressing beta-gal but not in cells expressing ras(asn17). In contrast, insulin increased IRS-1-associated PI3-kinase activity approximately 10-fold in control cells and high level overexpression of ras(asn17) did not impair this effect. Similarly, insulin and epidermal growth factor activation of total (no immunoprecipitation) PI3-kinase activity in both cytosol and total cellular membranes and insulin stimulation of glucose transport were not affected by expression of dominant negative ras. Thus, adenovirus-mediated gene transfer is effective for studying insulin signaling in fully differentiated insulin target cells. Inhibition of ras activation abolishes insulin-stimulated phosphorylation of map kinase but does not affect insulin stimulation of PI3-kinase activity. In normal cell physiology, PI3-kinase does not appear to be downstream of ras in

  13. Inhibition of Ras oncogenic activity by Ras protooncogenes.

    PubMed

    Diaz, Roberto; Lue, Jeffrey; Mathews, Jeremy; Yoon, Andrew; Ahn, Daniel; Garcia-España, Antonio; Leonardi, Peter; Vargas, Marcelo P; Pellicer, Angel

    2005-01-10

    Point mutations in ras genes have been found in a large number and wide variety of human tumors. These oncogenic Ras mutants are locked in an active GTP-bound state that leads to a constitutive and deregulated activation of Ras function. The dogma that ras oncogenes are dominant, whereby the mutation of a single allele in a cell will predispose the host cell to transformation regardless of the presence of the normal allele, is being challenged. We have seen that increasing amounts of Ras protooncogenes are able to inhibit the activity of the N-Ras oncogene in the activation of Elk in NIH 3T3 cells and in the formation of foci. We have been able to determine that the inhibitory effect is by competition between Ras protooncogenes and the N-Ras oncogene that occurs first at the effector level at the membranes, then at the processing level and lastly at the effector level in the cytosol. In addition, coexpression of the N-Ras protooncogene in thymic lymphomas induced by the N-Ras oncogene is associated with increased levels of p107, p130 and cyclin A and decreased levels of Rb. In the present report, we have shown that the N-Ras oncogene is not truly dominant over Ras protooncogenes and their competing activities might be depending on cellular context.

  14. RAS Mutations and Oncogenesis: Not all RAS Mutations are Created Equally

    PubMed Central

    Miller, Mark Steven; Miller, Lance D.

    2012-01-01

    Mutation in RAS proteins is one of the most common genetic alterations observed in human and experimentally induced rodent cancers. In vivo, oncogenic mutations have been shown to occur at exons 12, 13, and 61, resulting in any 1 of 19 possible point mutations in a given tumor for a specific RAS isoform. While some studies have suggested a possible role of different mutant alleles in determining tumor severity and phenotype, no general consensus has emerged on the oncogenicity of different mutant alleles in tumor formation and progression. Part of this may be due to a lack of a single, signature pathway that shows significant alterations between different mutations. Rather, it is likely that subtle differences in the activation, or lack thereof, of downstream effectors by different RAS mutant alleles may determine the eventual outcome in terms of tumor phenotype. This paper reviews our current understanding of the potential role of different RAS mutations on tumorigenesis, highlights studies in model cell culture and in vivo systems, and discusses the potential of expression array and computational network modeling to dissect out differences in activated RAS genes in conferring a transforming phenotype. PMID:22303394

  15. Tyrosine phosphorylation of RAS by ABL allosterically enhances effector binding

    PubMed Central

    Ting, Pamela Y.; Johnson, Christian W.; Fang, Cong; Cao, Xiaoqing; Graeber, Thomas G.; Mattos, Carla; Colicelli, John

    2015-01-01

    RAS proteins are signal transduction gatekeepers that mediate cell growth, survival, and differentiation through interactions with multiple effector proteins. The RAS effector RAS- and RAB-interacting protein 1 (RIN1) activates its own downstream effectors, the small GTPase RAB5 and the tyrosine kinase Abelson tyrosine-protein kinase (ABL), to modulate endocytosis and cytoskeleton remodeling. To identify ABL substrates downstream of RAS-to-RIN1 signaling, we examined human HEK293T cells overexpressing components of this pathway. Proteomic analysis revealed several novel phosphotyrosine peptides, including Harvey rat sarcoma oncogene (HRAS)-pTyr137. Here we report that ABL phosphorylates tyrosine 137 of H-, K-, and NRAS. Increased RIN1 levels enhanced HRAS-Tyr137 phosphorylation by nearly 5-fold, suggesting that RAS-stimulated RIN1 can drive ABL-mediated RAS modification in a feedback circuit. Tyr137 is well conserved among RAS orthologs and is part of a transprotein H-bond network. Crystal structures of HRASY137F and HRASY137E revealed conformation changes radiating from the mutated residue. Although consistent with Tyr137 participation in allosteric control of HRAS function, the mutations did not alter intrinsic GTP hydrolysis rates in vitro. HRAS-Tyr137 phosphorylation enhanced HRAS signaling capacity in cells, however, as reflected by a 4-fold increase in the association of phosphorylated HRASG12V with its effector protein RAF proto-oncogene serine/threonine protein kinase 1 (RAF1). These data suggest that RAS phosphorylation at Tyr137 allosterically alters protein conformation and effector binding, providing a mechanism for effector-initiated modulation of RAS signaling.—Ting, P. Y., Johnson, C. W., Fang, C., Cao, X., Graeber, T. G., Mattos, C., Colicelli, J. Tyrosine phosphorylation of RAS by ABL allosterically enhances effector binding. PMID:25999467

  16. RAS - Screens & Assays - Drug Discovery

    Cancer.gov

    The RAS Drug Discovery group aims to develop assays that will reveal aspects of RAS biology upon which cancer cells depend. Successful assay formats are made available for high-throughput screening programs to yield potentially effective drug compounds.

  17. RasGRP Ras guanine nucleotide exchange factors in cancer

    PubMed Central

    Ksionda, Olga; Limnander, Andre

    2014-01-01

    Summary RasGRP proteins are activators of Ras and other related small GTPases by the virtue of functioning as guanine nucleotide exchange factors (GEFs). In vertebrates, four RasGRP family members have been described. RasGRP-1 through −4 share many structural domains but there are also subtle differences between each of the different family members. Whereas SOS RasGEFs are ubiquitously expressed, RasGRP proteins are expressed in distinct patterns, such as in different cells of the hematopoietic system and in the brain. Most studies have concentrated on the role of RasGRP proteins in the development and function of immune cell types because of the predominant RasGRP expression profiles in these cells and the immune phenotypes of mice deficient for Rasgrp genes. However, more recent studies demonstrate that RasGRPs also play an important role in tumorigenesis. Examples are skin- and hematological-cancers but also solid malignancies such as melanoma or prostate cancer. These novel studies bring up many new and unanswered questions related to the molecular mechanism of RasGRP-driven oncogenesis, such as new receptor systems that RasGRP appears to respond to as well as regulatory mechanism for RasGRP expression that appear to be perturbed in these cancers. Here we will review some of the known aspects of RasGRP biology in lymphocytes and will discuss the exciting new notion that RasGRP Ras exchange factors play a role in oncogenesis downstream of various growth factor receptors. PMID:24744772

  18. Dietary turmeric modulates DMBA-induced p21{sup ras}, MAP kinases and AP-1/NF-{kappa}B pathway to alter cellular responses during hamster buccal pouch carcinogenesis

    SciTech Connect

    Garg, Rachana; Ingle, Arvind; Maru, Girish

    2008-11-01

    The chemopreventive efficacy of turmeric has been established in experimental systems. However, its mechanism(s) of action are not fully elucidated in vivo. The present study investigates the mechanism of turmeric-mediated chemoprevention in 7,12-dimethylbenz(a)anthracene (DMBA)-induced hamster buccal pouch (HBP) carcinogenesis at 2, 4, 6, 10 and 12 weeks. Dietary turmeric (1%) led to decrease in DMBA-induced tumor burden and multiplicity, and enhanced the latency period in parallel, to its modulatory effects on oncogene products and various cellular responses during HBP tumorigenesis. DMBA-induced expression of ras oncogene product, p21 and downstream target, the mitogen-activated protein kinases were significantly decreased by turmeric during HBP carcinogenesis. Turmeric also diminished the DMBA-induced mRNA expression of proto-oncogenes (c-jun, c-fos) and NF-{kappa}B, leading to decreased protein levels and in further attenuation of DMBA-induced AP-1/NF-{kappa}B DNA-binding in the buccal pouch nuclear extracts. Besides, buccal pouch of hamsters receiving turmeric diet showed significant alterations in DMBA-induced effects: (a) decrease in cell proliferation (diminished PCNA and Bcl2 expression), (b) enhanced apoptosis (increased expression of Bax, caspase-3 and apoptotic index), (c) decrease in inflammation (levels of Cox-2, the downstream target of AP-1/NF-{kappa}B, and PGE2) and (d) aberrant expression of differentiation markers, the cytokeratins (1, 5, 8, and 18). Together, the protective effects of dietary turmeric converge on augmenting apoptosis of the initiated cells and decreasing cell proliferation in DMBA-treated animals, which in turn, is reflected in decreased tumor burden, multiplicity and enhanced latency period. Some of these biomarkers are likely to be helpful in monitoring clinical trials and evaluating drug effect measurements.

  19. Ras enhances Myc protein stability.

    PubMed

    Sears, R; Leone, G; DeGregori, J; Nevins, J R

    1999-02-01

    Various experiments have demonstrated a collaborative action of Myc and Ras, both in normal cell growth control as well as during oncogenesis. We now show that Ras enhances the accumulation of Myc activity by stabilizing the Myc protein. Whereas Myc has a very short half-life when produced in the absence of mitogenic signals, due to degradation by the 26S proteasome, the half-life of Myc increases markedly in growth-stimulated cells. This stabilization is dependent on the Ras/Raf/MAPK pathway and is not augmented by proteasome inhibition, suggesting that Ras inhibits the proteasome-dependent degradation of Myc. We propose that one aspect of Myc-Ras collaboration is an ability of Ras to enhance the accumulation of transcriptionally active Myc protein.

  20. The farnesyltransferase inhibitor, LB42708, inhibits growth and induces apoptosis irreversibly in H-ras and K-ras-transformed rat intestinal epithelial cells

    SciTech Connect

    Kim, Han-Soo; Kim, Ju Won; Gang, Jingu; Wen, Jing; Koh, Sang Seok; Koh, Jong Sung; Chung, Hyun-Ho; Song, Si Young . E-mail: gisong@yumc.yonsei.ac.kr

    2006-09-15

    LB42708 (LB7) and LB42908 (LB9) are pyrrole-based orally active farnesyltransferase inhibitors (FTIs) that have similar structures. The in vitro potencies of these compounds against FTase and GGTase I are remarkably similar, and yet they display different activity in apoptosis induction and morphological reversion of ras-transformed rat intestinal epithelial (RIE) cells. Both FTIs induced cell death despite K-ras prenylation, implying the participation of Ras-independent mechanism(s). Growth inhibition by these two FTIs was accompanied by G1 and G2/M cell cycle arrests in H-ras and K-ras-transformed RIE cells, respectively. We identified three key markers, p21{sup CIP1/WAF1}, RhoB and EGFR, that can explain the differences in the molecular mechanism of action between two FTIs. Only LB7 induced the upregulation of p21{sup CIP1/WAF1} and RhoB above the basal level that led to the cell cycle arrest and to distinct morphological alterations of ras-transformed RIE cells. Both FTIs successfully inhibited the ERK and activated JNK in RIE/K-ras cells. While the addition of conditioned medium from RIE/K-ras reversed the growth inhibition of ras-transformed RIE cells by LB9, it failed to overcome the growth inhibitory effect of LB7 in both H-ras- and K-ras-transformed RIE cells. We found that LB7, but not LB9, decreased the expression of EGFRs that confers the cellular unresponsiveness to EGFR ligands. These results suggest that LB7 causes the induction of p21{sup CIP1/WAF1} and RhoB and downregulation of EGFR that may serve as critical steps in the mechanism by which FTIs trigger irreversible inhibitions on the cell growth and apoptosis in ras-transformed cells.

  1. Inhibition of ras oncogene: a novel approach to antineoplastic therapy.

    PubMed

    Scharovsky, O G; Rozados, V R; Gervasoni, S I; Matar, P

    2000-01-01

    The most frequently detected oncogene alterations, both in animal and human cancers, are the mutations in the ras oncogene family. These oncogenes are mutated or overexpressed in many human tumors, with a high incidence in tumors of the pancreas, thyroid, colon, lung and certain types of leukemia. Ras is a small guanine nucleotide binding protein that transduces biological information from the cell surface to cytoplasmic components within cells. The signal is transduced to the cell nucleus through second messengers, and it ultimately induces cell division. Oncogenic forms of p21(ras) lead to unregulated, sustained signaling through downstream effectors. The ras family of oncogenes is involved in the development of both primary tumors and metastases making it a good therapeutic target. Several therapeutic approaches to cancer have been developed pointing to reducing the altered gene product or to eliminating its biological function: (1) gene therapy with ribozymes, which are able to break down specific RNA sequences, or with antisense oligonucleotides, (2) immunotherapy through passive or active immunization protocols, and (3) inhibition of p21(ras) farnesylation either by inhibition of farnesyl transferase or synthesis inhibition of farnesyl moieties. PMID:10895051

  2. [Role of RAS in prehypertension].

    PubMed

    Inaba, Shinji; Iwai, Masaru; Horiuchi, Masatsugu

    2008-08-01

    Hypertension has long been recognized as a major risk factor of several cardiovascular diseases. It is well known that the renin-angiotensin system(RAS) is involved in the pathogenesis of both hypertension and hypertensive end-organ damage. Untreated hypertension is self-accelerating condition through RAS stimulation. Activation of RAS contributes to the transition from borderline hypertension to established hypertension. Recently, "the Seventh Report of Joint National Committee on Prevention, Detection, Evaluation and Treatment of High Blood Pressure (JNC 7)" proposed a new classification of borderline blood pressure levels, as "prehypertension". The therapeutic focus has begun to shift from the therapy of established hypertension to the prevention of hypertension. This review addressed the relationship between hypertension, prehypertension and the role of RAS. PMID:18700549

  3. Silent assassin: oncogenic ras directs epigenetic inactivation of target genes.

    PubMed

    Cheng, Xiaodong

    2008-01-01

    Oncogenic transformation is associated with genetic changes and epigenetic alterations. A study now shows that oncogenic Ras uses a complex and elaborate epigenetic silencing program to specifically repress the expression of multiple unrelated cancer-suppressing genes through a common pathway. These results suggest that cancer-related epigenetic modifications may arise through a specific and instructive mechanism and that genetic changes and epigenetic alterations are intimately connected and contribute to tumorigenesis cooperatively. PMID:18385037

  4. The Efficacy of Raf Kinase Recruitment to the GTPase H-ras Depends on H-ras Membrane Conformer-specific Nanoclustering*♦

    PubMed Central

    Guzmán, Camilo; Šolman, Maja; Ligabue, Alessio; Blaževitš, Olga; Andrade, Débora M.; Reymond, Luc; Eggeling, Christian; Abankwa, Daniel

    2014-01-01

    Solution structures and biochemical data have provided a wealth of mechanistic insight into Ras GTPases. However, information on how much the membrane organization of these lipid-modified proteins impacts on their signaling is still scarce. Ras proteins are organized into membrane nanoclusters, which are necessary for Ras-MAPK signaling. Using quantitative conventional and super-resolution fluorescence methods, as well as mathematical modeling, we investigated nanoclustering of H-ras helix α4 and hypervariable region mutants that have different bona fide conformations on the membrane. By following the emergence of conformer-specific nanoclusters in the plasma membrane of mammalian cells, we found that conformers impart distinct nanoclustering responses depending on the cytoplasmic levels of the nanocluster scaffold galectin-1. Computational modeling revealed that complexes containing H-ras conformers and galectin-1 affect both the number and lifetime of nanoclusters and thus determine the specific Raf effector recruitment. Our results show that mutations in Ras can affect its nanoclustering response and thus allosterically effector recruitment and downstream signaling. We postulate that cancer- and developmental disease-linked mutations that are associated with the Ras membrane conformation may exhibit so far unrecognized Ras nanoclustering and therefore signaling alterations. PMID:24569991

  5. The efficacy of Raf kinase recruitment to the GTPase H-ras depends on H-ras membrane conformer-specific nanoclustering.

    PubMed

    Guzmán, Camilo; Šolman, Maja; Ligabue, Alessio; Blaževitš, Olga; Andrade, Débora M; Reymond, Luc; Eggeling, Christian; Abankwa, Daniel

    2014-04-01

    Solution structures and biochemical data have provided a wealth of mechanistic insight into Ras GTPases. However, information on how much the membrane organization of these lipid-modified proteins impacts on their signaling is still scarce. Ras proteins are organized into membrane nanoclusters, which are necessary for Ras-MAPK signaling. Using quantitative conventional and super-resolution fluorescence methods, as well as mathematical modeling, we investigated nanoclustering of H-ras helix α4 and hypervariable region mutants that have different bona fide conformations on the membrane. By following the emergence of conformer-specific nanoclusters in the plasma membrane of mammalian cells, we found that conformers impart distinct nanoclustering responses depending on the cytoplasmic levels of the nanocluster scaffold galectin-1. Computational modeling revealed that complexes containing H-ras conformers and galectin-1 affect both the number and lifetime of nanoclusters and thus determine the specific Raf effector recruitment. Our results show that mutations in Ras can affect its nanoclustering response and thus allosterically effector recruitment and downstream signaling. We postulate that cancer- and developmental disease-linked mutations that are associated with the Ras membrane conformation may exhibit so far unrecognized Ras nanoclustering and therefore signaling alterations.

  6. Ras Modifies Proliferation and Invasiveness of Cells Expressing Human Papillomavirus Oncoproteins▿

    PubMed Central

    Yoshida, Satoshi; Kajitani, Naoko; Satsuka, Ayano; Nakamura, Hiroyasu; Sakai, Hiroyuki

    2008-01-01

    Infection by human papillomavirus (HPV) is a major risk factor for human cervical carcinoma. However, the HPV infection alone is not sufficient for cancer formation. Cervical carcinogenesis is considered a multistep process accompanied by genetic alterations of the cell. Ras is activated in approximately 20% of human cancers, and it is related to the metastatic conversion of tumor cells. We investigated how Ras activation was involved in the malignant conversion of HPV-infected lesions. The active form of H-ras was introduced into human primary keratinocytes expressing the HPV type 18 (HPV18) oncoproteins E6 and/or E7. We analyzed the keratinocytes’ growth potentials and found that the activation of the Ras pathway induced senescence-like growth arrest. Senescence could be eliminated by high-risk E7 expression, suggesting that the pRb pathway was important for Ras-induced senescence. Then we analyzed the effect of Ras activation on epidermis development by using an organotypic “raft” culture and found that the E7 and H-ras coexpressions conferred invasive potential on the epidermis. This invasiveness resulted from the upregulation of MT1-MMP and MMP9 by H-ras and E7, respectively, in which the activation of the MEK/extracellular signal-regulated kinase pathway was involved. These results indicated that the activation of Ras or the related signal pathways promoted the malignant conversion of HPV-infected cells. PMID:18579583

  7. Mapping the functional versatility and fragility of Ras GTPase signaling circuits through in vitro network reconstitution.

    PubMed

    Coyle, Scott M; Lim, Wendell A

    2016-01-01

    The Ras-superfamily GTPases are central controllers of cell proliferation and morphology. Ras signaling is mediated by a system of interacting molecules: upstream enzymes (GEF/GAP) regulate Ras's ability to recruit multiple competing downstream effectors. We developed a multiplexed, multi-turnover assay for measuring the dynamic signaling behavior of in vitro reconstituted H-Ras signaling systems. By including both upstream regulators and downstream effectors, we can systematically map how different network configurations shape the dynamic system response. The concentration and identity of both upstream and downstream signaling components strongly impacted the timing, duration, shape, and amplitude of effector outputs. The distorted output of oncogenic alleles of Ras was highly dependent on the balance of positive (GAP) and negative (GEF) regulators in the system. We found that different effectors interpreted the same inputs with distinct output dynamics, enabling a Ras system to encode multiple unique temporal outputs in response to a single input. We also found that different Ras-to-GEF positive feedback mechanisms could reshape output dynamics in distinct ways, such as signal amplification or overshoot minimization. Mapping of the space of output behaviors accessible to Ras provides a design manual for programming Ras circuits, and reveals how these systems are readily adapted to produce an array of dynamic signaling behaviors. Nonetheless, this versatility comes with a trade-off of fragility, as there exist numerous paths to altered signaling behaviors that could cause disease. PMID:26765565

  8. Ras CAAX peptidomimetic FTI-277 selectively blocks oncogenic Ras signaling by inducing cytoplasmic accumulation of inactive Ras-Raf complexes.

    PubMed

    Lerner, E C; Qian, Y; Blaskovich, M A; Fossum, R D; Vogt, A; Sun, J; Cox, A D; Der, C J; Hamilton, A D; Sebti, S M

    1995-11-10

    Ras-induced malignant transformation requires Ras farnesylation, a lipid posttranslational modification catalyzed by farnesyltransferase (FTase). Inhibitors of this enzyme have been shown to block Ras-dependent transformation, but the mechanism by which this occurs remains largely unknown. We have designed FTI-276, a peptide mimetic of the COOH-terminal Cys-Val-Ile-Met of K-Ras4B that inhibited potently FTase in vitro (IC50 = 500 pM) and was highly selective for FTase over geranylgeranyltransferase I (GGTase I) (IC50 = 50 nM). FTI-277, the methyl ester derivative of FTI-276, was extremely potent (IC50 = 100 nM) at inhibiting H-Ras, but not the geranylgeranylated Rap1A processing in whole cells. Treatment of H-Ras oncogene-transformed NIH 3T3 cells with FTI-277 blocked recruitment to the plasma membrane and subsequent activation of the serine/threonine kinase c-Raf-1 in cells transformed by farnesylated Ras (H-RasF), but not geranylgeranylated, Ras (H-RasGG). FTI-277 induced accumulation of cytoplasmic non-farnesylated H-Ras that was able to bind Raf and form cytoplasmic Ras/Raf complexes in which Raf kinase was not activated. Furthermore, FTI-277 blocked constitutive activation of mitogen-activated protein kinase (MAPK) in H-RasF, but not H-RasGG, or Raf-transformed cells. FTI-277 also inhibited oncogenic K-Ras4B processing and constitutive activation of MAPK, but the concentrations required were 100-fold higher than those needed for H-Ras inhibition. The results demonstrate that FTI-277 blocks Ras oncogenic signaling by accumulating inactive Ras/Raf complexes in the cytoplasm, hence preventing constitutive activation of the MAPK cascade.

  9. Activation of intracellular kinases in Xenopus oocytes by p21ras and phospholipases: a comparative study.

    PubMed

    Carnero, A; Lacal, J C

    1995-02-01

    Signal transduction induced by generations of second messengers from membrane phospholipids is a major regulatory mechanism in the control of cell proliferation. Indeed, oncogenic p21ras alters the intracellular levels of phospholipid metabolites in both mammalian cells and Xenopus oocytes. However, it is still controversial whether this alteration it is biologically significant. We have analyzed the ras-induced signal transduction pathway in Xenopus oocytes and have correlated its mechanism of activation with that of the three most relevant phospholipases (PLs). After microinjection, ras-p21 induces a rapid PLD activation followed by a late PLA2 activation. By contrast, phosphatidylcholine-specific PLC was not activated under similar conditions. When each of these PLs was studied for its ability to activate intracellular signalling kinases, all of them were found to activate maturation-promoting factor efficiently. However, only PLD was able to activate MAP kinase and S6 kinase II, a similar pattern to that induced by p21ras proteins. Thus, the comparison of activated enzymes after microinjection of p21ras or PLs indicated that only PLD microinjection mimetized p21ras signalling. Finally, inhibition of the endogenous PLD activity by neomycin substantially reduced the biological activity of p21ras. All these results suggest that PLD activation may constitute a relevant step in ras-induced germinal vesicle breakdown in Xenopus oocytes.

  10. Fendiline Inhibits K-Ras Plasma Membrane Localization and Blocks K-Ras Signal Transmission

    PubMed Central

    van der Hoeven, Dharini; Cho, Kwang-jin; Ma, Xiaoping; Chigurupati, Sravanthi; Parton, Robert G.

    2013-01-01

    Ras proteins regulate signaling pathways important for cell growth, differentiation, and survival. Oncogenic mutant Ras proteins are commonly expressed in human tumors, with mutations of the K-Ras isoform being most prevalent. To be active, K-Ras must undergo posttranslational processing and associate with the plasma membrane. We therefore devised a high-content screening assay to search for inhibitors of K-Ras plasma membrane association. Using this assay, we identified fendiline, an L-type calcium channel blocker, as a specific inhibitor of K-Ras plasma membrane targeting with no detectable effect on the localization of H- and N-Ras. Other classes of L-type calcium channel blockers did not mislocalize K-Ras, suggesting a mechanism that is unrelated to calcium channel blockade. Fendiline did not inhibit K-Ras posttranslational processing but significantly reduced nanoclustering of K-Ras and redistributed K-Ras from the plasma membrane to the endoplasmic reticulum (ER), Golgi apparatus, endosomes, and cytosol. Fendiline significantly inhibited signaling downstream of constitutively active K-Ras and endogenous K-Ras signaling in cells transformed by oncogenic H-Ras. Consistent with these effects, fendiline blocked the proliferation of pancreatic, colon, lung, and endometrial cancer cell lines expressing oncogenic mutant K-Ras. Taken together, these results suggest that inhibitors of K-Ras plasma membrane localization may have utility as novel K-Ras-specific anticancer therapeutics. PMID:23129805

  11. Cardiac remodelling and RAS inhibition.

    PubMed

    Ferrario, Carlos M

    2016-06-01

    Risk factors such as hypertension and diabetes are known to augment the activity and tissue expression of angiotensin II (Ang II), the major effector peptide of the renin-angiotensin system (RAS). Overstimulation of the RAS has been implicated in a chain of events that contribute to the pathogenesis of cardiovascular (CV) disease, including the development of cardiac remodelling. This chain of events has been termed the CV continuum. The concept of CV disease existing as a continuum was first proposed in 1991 and it is believed that intervention at any point within the continuum can modify disease progression. Treatment with antihypertensive agents may result in regression of left ventricular hypertrophy, with different drug classes exhibiting different degrees of efficacy. The greatest decrease in left ventricular mass is observed following treatment with angiotensin converting enzyme inhibitors (ACE-Is), which inhibit Ang II formation. Although ACE-Is and angiotensin receptor blockers (ARBs) provide significant benefits in terms of CV events and stroke, mortality remains high. This is partly due to a failure to completely suppress the RAS, and, as our knowledge has increased, an escape phenomenon has been proposed whereby the human sequence of the 12 amino acid substrate angiotensin-(1-12) is converted to Ang II by the mast cell protease, chymase. Angiotensin-(1-12) is abundant in a wide range of organs and has been shown to increase blood pressure in animal models, an effect abolished by the presence of ACE-Is or ARBs. This review explores the CV continuum, in addition to examining the influence of the RAS. We also consider novel pathways within the RAS and how new therapeutic approaches that target this are required to further reduce Ang II formation, and so provide patients with additional benefits from a more complete blockade of the RAS. PMID:27105891

  12. Adjustable hybrid diffractive/refractive achromatic lens.

    PubMed

    Valley, Pouria; Savidis, Nickolaos; Schwiegerling, Jim; Dodge, Mohammad Reza; Peyman, Gholam; Peyghambarian, N

    2011-04-11

    We demonstrate a variable focal length achromatic lens that consists of a flat liquid crystal diffractive lens and a pressure-controlled fluidic refractive lens. The diffractive lens is composed of a flat binary Fresnel zone structure and a thin liquid crystal layer, producing high efficiency and millisecond switching times while applying a low ac voltage input. The focusing power of the diffractive lens is adjusted by electrically modifying the sub-zones and re-establishing phase wrapping points. The refractive lens includes a fluid chamber with a flat glass surface and an opposing elastic polydimethylsiloxane (PDMS) membrane surface. Inserting fluid volume through a pump system into the clear aperture region alters the membrane curvature and adjusts the refractive lens' focal position. Primary chromatic aberration is remarkably reduced through the coupling of the fluidic and diffractive lenses at selected focal lengths. Potential applications include miniature color imaging systems, medical and ophthalmic devices, or any design that utilizes variable focal length achromats.

  13. Bionic intraocular lens with variable focus and integrated structure

    NASA Astrophysics Data System (ADS)

    Liang, Dan; Wang, Xuan-Yin; Du, Jia-Wei; Xiang, Ke

    2015-10-01

    This paper proposes a bionic accommodating intraocular lens (IOL) for ophthalmic surgery. The designed lens has a solid-liquid mixed integrated structure, which mainly consists of a support ring, elastic membrane, rigid lens, and optical liquid. The lens focus can be adjusted through the deformation of the lens front surface when compressed. The integrated structure of the IOL is presented, as well as a detailed description of the lens materials and fabrication process. Images under different radial pressures are captured, and the lens deformation process, accommodating range, density, and optical property are analyzed. The designed lens achieves a 14.6 D accommodating range under a radial pressure of 51.4 mN and a 0.24 mm alteration of the lens outer radius. The deformation property of the lens matches well with the characteristic of the eye and shows the potential to help patients fully recover their vision accommodation ability after the cataract surgery.

  14. Therapeutic Strategies for Targeting Ras Proteins

    PubMed Central

    Gysin, Stephan; Salt, Megan; Young, Amy; McCormick, Frank

    2011-01-01

    Ras genes are frequently activated in cancer. Attempts to develop drugs that target mutant Ras proteins have, so far, been unsuccessful. Tumors bearing these mutations, therefore, remain among the most difficult to treat. Most efforts to block activated Ras have focused on pathways downstream. Drugs that inhibit Raf kinase have shown clinical benefit in the treatment of malignant melanoma. However, these drugs have failed to show clinical benefit in Ras mutant tumors. It remains unclear to what extent Ras depends on Raf kinase for transforming activity, even though Raf proteins bind directly to Ras and are certainly major effectors of Ras action in normal cells and in development. Furthermore, Raf kinase inhibitors can lead to paradoxical activation of the MAPK pathway. MEK inhibitors block the Ras-MAPK pathway, but often activate the PI3’-kinase, and have shown little clinical benefit as single agents. This activation is mediated by EGF-R and other receptor tyrosine kinases through relief of a negative feedback loop from ERK. Drug combinations that target multiple points within the Ras signaling network are likely to be necessary to achieve substantial clinical benefit. Other effectors may also contribute to Ras signaling and provide a source of targets. In addition, unbiased screens for genes necessary for Ras transformation have revealed new potential targets and have added to our understanding of Ras cancer biology. PMID:21779505

  15. The K-Ras 4A isoform promotes apoptosis but does not affect either lifespan or spontaneous tumor incidence in aging mice

    SciTech Connect

    Plowman, Sarah J.; Arends, Mark J.; Brownstein, David G.; Luo Feijun; Devenney, Paul S.; Rose, Lorraine; Ritchie, Ann-Marie; Berry, Rachel L.; Harrison, David J.; Hooper, Martin L.; Patek, Charles E. . E-mail: Charles.Patek@ed.ac.uk

    2006-01-01

    Ras proteins function as molecular switches in signal transduction pathways, and, here, we examined the effects of the K-ras4A and 4B splice variants on cell function by comparing wild-type embryonic stem (ES) cells with K-ras {sup tm{delta}}{sup 4A/tm{delta}}{sup 4A} (exon 4A knock-out) ES cells which express K-ras4B only and K-ras {sup -/-} (exons 1-3 knock-out) ES cells which express neither splice variant, and intestinal epithelium from wild-type and K-ras {sup tm{delta}}{sup 4A/tm{delta}}{sup 4A} mice. RT-qPCR analysis found that K-ras4B expression was reduced in K-ras {sup tm{delta}}{sup 4A/tm{delta}}{sup 4A} ES cells but unaffected in small intestine. K-Ras deficiency did not affect ES cell growth, and K-Ras4A deficiency did not affect intestinal epithelial proliferation. K-ras {sup tm{delta}}{sup 4A/tm{delta}}{sup 4A} and K-ras {sup -/-} ES cells showed a reduced capacity for differentiation following LIF withdrawal, and K-ras {sup -/-} cells were least differentiated. K-Ras4A deficiency inhibited etoposide-induced apoptosis in ES cells and intestinal epithelial cells. However, K-ras {sup tm{delta}}{sup 4A/tm{delta}}{sup 4A} ES cells were more resistant to etoposide-induced apoptosis than K-ras {sup -/-} cells. The results indicate that (1) K-Ras4A promotes apoptosis while K-Ras4B inhibits it, and (2) K-Ras4B, and possibly K-Ras4A, promotes differentiation. The findings raise the possibility that alteration of the K-Ras4A/4B isoform ratio modulates tumorigenesis by differentially affecting stem cell survival and/or differentiation. However, K-Ras4A deficiency did not affect life expectancy or spontaneous overall tumor incidence in aging mice.

  16. Cloning and characterization of Ras-GRF2, a novel guanine nucleotide exchange factor for Ras.

    PubMed

    Fam, N P; Fan, W T; Wang, Z; Zhang, L J; Chen, H; Moran, M F

    1997-03-01

    Conversion of Ras proteins into an activated GTP-bound state able to bind effector proteins is catalyzed by specific guanine nucleotide exchange factors in response to a large number of extracellular stimuli. Here we report the isolation of mouse cDNAs encoding Ras-GRF2, a multidomain 135-kDa protein containing a COOH-terminal Cdc25-related domain that stimulates release of GDP from Ras but not other GTPases in vitro. Ras-GRF2 bound specifically to immobilized Ras lacking bound nucleotides, suggesting stabilization of the nucleotide-free form of Ras as a mechanism of catalyzing nucleotide exchange. The NH2-terminal region of Ras-GRF2 is predicted to contain features common to various signaling proteins including two pleckstrin homology domains and a Dbl homology region. Ras-GRF2 also contains an IQ motif which was required for its apparent constitutive association with calmodulin in epithelial cells ectopically expressing Ras-GRF2. Transient expression of Ras-GRF2 in kidney epithelial cells stimulated GTP binding by Ras and potentiated calcium ionophore-induced activation of mitogen-activated protein kinase (ERK1) dependent upon the IQ motif. Calcium influx caused Ras-GRF2 subcellular localization to change from cytosolic to peripheral, suggesting a possible mechanism for controlling Ras-GRF2 interactions with Ras at the plasma membrane. Epithelial cells overexpressing Ras-GRF2 are morphologically transformed and grow in a disorganized manner with minimal intercellular contacts. Northern analysis indicated a 9-kb GRF2 transcript in brain and lung, where p135 Ras-GRF2 is known to be expressed, and RNAs of 12 kb and 2.2 kb were detected in several tissues. Thus, Ras-GRF2 proteins with different domain structures may be widely expressed and couple diverse extracellular signals to Ras activation.

  17. Inhibitors of Ras-SOS Interactions.

    PubMed

    Lu, Shaoyong; Jang, Hyunbum; Zhang, Jian; Nussinov, Ruth

    2016-04-19

    Activating Ras mutations are found in about 30 % of human cancers. Ras activation is regulated by guanine nucleotide exchange factors, such as the son of sevenless (SOS), which form protein-protein interactions (PPIs) with Ras and catalyze the exchange of GDP by GTP. This is the rate-limiting step in Ras activation. However, Ras surfaces lack any evident suitable pockets where a molecule might bind tightly, rendering Ras proteins still 'undruggable' for over 30 years. Among the alternative approaches is the design of inhibitors that target the Ras-SOS PPI interface, a strategy that is gaining increasing recognition for treating Ras mutant cancers. Herein we focus on data that has accumulated over the past few years pertaining to the design of small-molecule modulators or peptide mimetics aimed at the interface of the Ras-SOS PPI. We emphasize, however, that even if such Ras-SOS therapeutics are potent, drug resistance may emerge. To counteract this development, we propose "pathway drug cocktails", that is, drug combinations aimed at parallel (or compensatory) pathways. A repertoire of classified cancer, cell/tissue, and pathway/protein combinations would be beneficial toward this goal.

  18. Collection Mode Lens System

    DOEpatents

    Fletcher, Daniel A.; Kino, Gordon S.

    2002-11-05

    A lens system including a collection lens and a microlens spaced from the collection lens adjacent the region to be observed. The diameter of the observablel region depends substantially on the radius of the microlens.

  19. RAS oncogenes: weaving a tumorigenic web

    PubMed Central

    Pylayeva-Gupta, Yuliya; Grabocka, Elda; Bar-Sagi, Dafna

    2013-01-01

    RAS proteins are essential components of signalling pathways that emanate from cell surface receptors. Oncogenic activation of these proteins owing to missense mutations is frequently detected in several types of cancer. A wealth of biochemical and genetic studies indicates that RAS proteins control a complex molecular circuitry that consists of a wide array of interconnecting pathways. In this Review, we describe how RAS oncogenes exploit their extensive signalling reach to affect multiple cellular processes that drive tumorigenesis. PMID:21993244

  20. Ras Regulates Rb via NORE1A.

    PubMed

    Barnoud, Thibaut; Donninger, Howard; Clark, Geoffrey J

    2016-02-01

    Mutations in the Ras oncogene are one of the most frequent events in human cancer. Although Ras regulates numerous growth-promoting pathways to drive transformation, it can paradoxically promote an irreversible cell cycle arrest known as oncogene-induced senescence. Although senescence has clearly been implicated as a major defense mechanism against tumorigenesis, the mechanisms by which Ras can promote such a senescent phenotype remain poorly defined. We have shown recently that the Ras death effector NORE1A plays a critical role in promoting Ras-induced senescence and connects Ras to the regulation of the p53 tumor suppressor. We now show that NORE1A also connects Ras to the regulation of a second major prosenescent tumor suppressor, the retinoblastoma (Rb) protein. We show that Ras induces the formation of a complex between NORE1A and the phosphatase PP1A, promoting the activation of the Rb tumor suppressor by dephosphorylation. Furthermore, suppression of Rb reduces NORE1A senescence activity. These results, together with our previous findings, suggest that NORE1A acts as a critical tumor suppressor node, linking Ras to both the p53 and the Rb pathways to drive senescence.

  1. Binding of calcium ions to Ras promotes Ras guanine nucleotide exchange under emulated physiological conditions.

    PubMed

    Liao, Jun-Ming; Mo, Zhong-Ying; Wu, Ling-Jia; Chen, Jie; Liang, Yi

    2008-11-01

    Both Ras protein and calcium play significant roles in various cellular processes via complex signaling transduction networks. However, it is not well understood whether and how Ca(2+) can directly regulate Ras function. Here we demonstrate by isothermal titration calorimetry that Ca(2+) directly binds to the H-Ras.GDP.Mg(2+) complex with moderate affinity at the first binding site followed by two weak binding events. The results from limited proteinase degradation show that Ca(2+) protects the fragments of H-Ras from being further degraded by trypsin and by proteinase K. HPLC studies together with fluorescence spectroscopic measurements indicate that binding of Ca(2+) to the H-Ras.GDP.Mg(2+) complex remarkably promotes guanine nucleotide exchange on H-Ras under emulated physiological Ca(2+) concentration conditions. Addition of high concentrations of either of two macromolecular crowding agents, Ficoll 70 and dextran 70, dramatically enhances H-Ras guanine nucleotide exchange extent in the presence of Ca(2+) at emulated physiological concentrations, and the nucleotide exchange extent increases significantly with the concentrations of crowding agents. Together, these results indicate that binding of calcium ions to H-Ras remarkably promotes H-Ras guanine nucleotide exchange under emulated physiological conditions. We thus propose that Ca(2+) may activate Ras signaling pathway by interaction with Ras, providing clues to understand the role of calcium in regulating Ras function in physiological environments.

  2. Targeting RAS Membrane Association: Back to the Future for Anti-RAS Drug Discovery?

    PubMed

    Cox, Adrienne D; Der, Channing J; Philips, Mark R

    2015-04-15

    RAS proteins require membrane association for their biologic activity, making this association a logical target for anti-RAS therapeutics. Lipid modification of RAS proteins by a farnesyl isoprenoid is an obligate step in that association, and is an enzymatic process. Accordingly, farnesyltransferase inhibitors (FTI) were developed as potential anti-RAS drugs. The lack of efficacy of FTIs as anticancer drugs was widely seen as indicating that blocking RAS membrane association was a flawed approach to cancer treatment. However, a deeper understanding of RAS modification and trafficking has revealed that this was an erroneous conclusion. In the presence of FTIs, KRAS and NRAS, which are the RAS isoforms most frequently mutated in cancer, become substrates for alternative modification, can still associate with membranes, and can still function. Thus, FTIs failed not because blocking RAS membrane association is an ineffective approach, but because FTIs failed to accomplish that task. Recent findings regarding RAS isoform trafficking and the regulation of RAS subcellular localization have rekindled interest in efforts to target these processes. In particular, improved understanding of the palmitoylation/depalmitoylation cycle that regulates RAS interaction with the plasma membrane, endomembranes, and cytosol, and of the potential importance of RAS chaperones, have led to new approaches. Efforts to validate and target other enzymatically regulated posttranslational modifications are also ongoing. In this review, we revisit lessons learned, describe the current state of the art, and highlight challenging but promising directions to achieve the goal of disrupting RAS membrane association and subcellular localization for anti-RAS drug development. Clin Cancer Res; 21(8); 1819-27. ©2015 AACR. See all articles in this CCR Focus section, "Targeting RAS-Driven Cancers." PMID:25878363

  3. Targeting RAS Membrane Association: Back to the Future for Anti-RAS Drug Discovery?

    PubMed

    Cox, Adrienne D; Der, Channing J; Philips, Mark R

    2015-04-15

    RAS proteins require membrane association for their biologic activity, making this association a logical target for anti-RAS therapeutics. Lipid modification of RAS proteins by a farnesyl isoprenoid is an obligate step in that association, and is an enzymatic process. Accordingly, farnesyltransferase inhibitors (FTI) were developed as potential anti-RAS drugs. The lack of efficacy of FTIs as anticancer drugs was widely seen as indicating that blocking RAS membrane association was a flawed approach to cancer treatment. However, a deeper understanding of RAS modification and trafficking has revealed that this was an erroneous conclusion. In the presence of FTIs, KRAS and NRAS, which are the RAS isoforms most frequently mutated in cancer, become substrates for alternative modification, can still associate with membranes, and can still function. Thus, FTIs failed not because blocking RAS membrane association is an ineffective approach, but because FTIs failed to accomplish that task. Recent findings regarding RAS isoform trafficking and the regulation of RAS subcellular localization have rekindled interest in efforts to target these processes. In particular, improved understanding of the palmitoylation/depalmitoylation cycle that regulates RAS interaction with the plasma membrane, endomembranes, and cytosol, and of the potential importance of RAS chaperones, have led to new approaches. Efforts to validate and target other enzymatically regulated posttranslational modifications are also ongoing. In this review, we revisit lessons learned, describe the current state of the art, and highlight challenging but promising directions to achieve the goal of disrupting RAS membrane association and subcellular localization for anti-RAS drug development. Clin Cancer Res; 21(8); 1819-27. ©2015 AACR. See all articles in this CCR Focus section, "Targeting RAS-Driven Cancers."

  4. Converging or Diverging Lens?

    ERIC Educational Resources Information Center

    Branca, Mario

    2013-01-01

    Why does a lens magnify? Why does it shrink objects? Why does this happen? The activities that we propose here are useful in helping us to understand how lenses work, and they show that the same lens can have different magnification capabilities. A converging lens can also act as a diverging lens. (Contains 4 figures.)

  5. Degradation of Activated K-Ras Orthologue via K-Ras-specific Lysine Residues Is Required for Cytokinesis*

    PubMed Central

    Sumita, Kazutaka; Yoshino, Hirofumi; Sasaki, Mika; Majd, Nazanin; Kahoud, Emily Rose; Takahashi, Hidenori; Takeuchi, Koh; Kuroda, Taruho; Lee, Susan; Charest, Pascale G.; Takeda, Kosuke; Asara, John M.; Firtel, Richard A.; Anastasiou, Dimitrios; Sasaki, Atsuo T.

    2014-01-01

    Mammalian cells encode three closely related Ras proteins, H-Ras, N-Ras, and K-Ras. Oncogenic K-Ras mutations frequently occur in human cancers, which lead to dysregulated cell proliferation and genomic instability. However, mechanistic role of the Ras isoform regulation have remained largely unknown. Furthermore, the dynamics and function of negative regulation of GTP-loaded K-Ras have not been fully investigated. Here, we demonstrate RasG, the Dictyostelium orthologue of K-Ras, is targeted for degradation by polyubiquitination. Both ubiquitination and degradation of RasG were strictly associated with RasG activity. High resolution tandem mass spectrometry (LC-MS/MS) analysis indicated that RasG ubiquitination occurs at C-terminal lysines equivalent to lysines found in human K-Ras but not in H-Ras and N-Ras homologues. Substitution of these lysine residues with arginines (4KR-RasG) diminished RasG ubiquitination and increased RasG protein stability. Cells expressing 4KR-RasG failed to undergo proper cytokinesis and resulted in multinucleated cells. Ectopically expressed human K-Ras undergoes polyubiquitin-mediated degradation in Dictyostelium, whereas human H-Ras and a Dictyostelium H-Ras homologue (RasC) are refractory to ubiquitination. Our results indicate the existence of GTP-loaded K-Ras orthologue-specific degradation system in Dictyostelium, and further identification of the responsible E3-ligase may provide a novel therapeutic approach against K-Ras-mutated cancers. PMID:24338482

  6. Ras does not contribute to the facilitation of hippocampal synaptic plasticity enabled by environmental enrichment.

    PubMed

    Novkovic, T; Heumann, R; Manahan-Vaughan, D

    2015-11-19

    Environmental enrichment (EE), which mimics the wealth of sensory, motor and cognitive stimuli that arise through intense interactions with the ambient environment, results in enhanced hippocampal long-term potentiation (LTP) and spatial learning. A key molecular factor in the mediation of these changes is the brain-derived neurotrophic factor (BDNF). One of the downstream cascades that is activated by BDNF is the cascade linked to the small GTPase, Ras, that triggers mitogen-activated protein kinase (MAPK) activity and is part of the cAMP response element-binding protein (CREB) pathway that can lead to synaptic restructuring to support LTP. Here, we explored whether persistent activation of Ras in neurons further enhances LTP following EE of rodents. Immediately following weaning, transgenic mice that expressed constitutively activated neuronal Ras, or their wildtype (Wt) littermates, underwent 3weeks of constant EE. In the absence of EE, theta burst stimulation (TBS) evoked LTP in the CA1 region of transgenic mice that was not significantly different from LTP in Wts. After 3weeks of EE, hippocampal LTP was improved in Wt mice. Enriched transgenic mice showed an equivalent level of LTP to enriched Wts, but it was not significantly different from non-enriched synRas controls. Western blot analysis performed after a pull-down assay showed that non-enriched transgenic mice expressed higher Ras activity compared to non-enriched Wts. Following EE, Ras activity was reduced in transgenics to levels detected in Wts. These results show that constitutive activation of Ras does not mimic the effects of EE on LTP. In addition, EE results in an equivalent enhancement of LTP transgenics and Wts, coupled with a decrease in Ras activity to Wt levels. This suggests that permanent activation of Ras in neurons of synRas animals following EE results in an altered feedback regulation of endogenous Ras activity that is not a key factor in LTP enhancements. The maintenance of Ras within

  7. Ras does not contribute to the facilitation of hippocampal synaptic plasticity enabled by environmental enrichment.

    PubMed

    Novkovic, T; Heumann, R; Manahan-Vaughan, D

    2015-11-19

    Environmental enrichment (EE), which mimics the wealth of sensory, motor and cognitive stimuli that arise through intense interactions with the ambient environment, results in enhanced hippocampal long-term potentiation (LTP) and spatial learning. A key molecular factor in the mediation of these changes is the brain-derived neurotrophic factor (BDNF). One of the downstream cascades that is activated by BDNF is the cascade linked to the small GTPase, Ras, that triggers mitogen-activated protein kinase (MAPK) activity and is part of the cAMP response element-binding protein (CREB) pathway that can lead to synaptic restructuring to support LTP. Here, we explored whether persistent activation of Ras in neurons further enhances LTP following EE of rodents. Immediately following weaning, transgenic mice that expressed constitutively activated neuronal Ras, or their wildtype (Wt) littermates, underwent 3weeks of constant EE. In the absence of EE, theta burst stimulation (TBS) evoked LTP in the CA1 region of transgenic mice that was not significantly different from LTP in Wts. After 3weeks of EE, hippocampal LTP was improved in Wt mice. Enriched transgenic mice showed an equivalent level of LTP to enriched Wts, but it was not significantly different from non-enriched synRas controls. Western blot analysis performed after a pull-down assay showed that non-enriched transgenic mice expressed higher Ras activity compared to non-enriched Wts. Following EE, Ras activity was reduced in transgenics to levels detected in Wts. These results show that constitutive activation of Ras does not mimic the effects of EE on LTP. In addition, EE results in an equivalent enhancement of LTP transgenics and Wts, coupled with a decrease in Ras activity to Wt levels. This suggests that permanent activation of Ras in neurons of synRas animals following EE results in an altered feedback regulation of endogenous Ras activity that is not a key factor in LTP enhancements. The maintenance of Ras within

  8. Contact lens in keratoconus

    PubMed Central

    Rathi, Varsha M; Mandathara, Preeji S; Dumpati, Srikanth

    2013-01-01

    Contact lenses are required for the visual improvement in patients with keratoconus. Various contact lens options, such as rigid gas permeable (RGP) lenses, soft and soft toric lenses, piggy back contact lenses (PBCL), hybrid lenses and scleral lenses are availble. This article discusses about selection of a lens depending on the type of keratoconus and the fitting philosophies of various contact lenses including the starting trial lens. A Medline search was carried out for articles in the English language with the keywords keratoconus and various contact lenses such as Rose k lens, RGP lens, hybrid lens, scleral lens and PBCL. PMID:23925325

  9. The Lens Capsule

    PubMed Central

    Danysh, Brian P.; Duncan, Melinda K.

    2009-01-01

    The lens capsule is a modified basement membrane that completely surrounds the ocular lens. It is known that this extracellular matrix is important for both the structure and biomechanics of the lens in addition to providing informational cues to maintain lens cell phenotype. This review covers the development and structure of the lens capsule, lens diseases associated with mutations in extracellular matrix genes and the role of the capsule in lens function including those proposed for visual accommodation, selective permeability to infectious agents, and cell signaling. PMID:18773892

  10. Optimizing depuration of salmon in RAS

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Fish cultured within water recirculating aquaculture systems (RAS) can acquire "earthy" or "musty" off-flavors due to bioaccumulation of the compounds geosmin and 2-methylisoborneol (MIB), respectively, which are produced by certain bacterial species present in RAS biosolids and microbial biofilms. ...

  11. Structural analysis of autoinhibition in the Ras-specific exchange factor RasGRP1

    PubMed Central

    Iwig, Jeffrey S; Vercoulen, Yvonne; Das, Rahul; Barros, Tiago; Limnander, Andre; Che, Yan; Pelton, Jeffrey G; Wemmer, David E; Roose, Jeroen P; Kuriyan, John

    2013-01-01

    RasGRP1 and SOS are Ras-specific nucleotide exchange factors that have distinct roles in lymphocyte development. RasGRP1 is important in some cancers and autoimmune diseases but, in contrast to SOS, its regulatory mechanisms are poorly understood. Activating signals lead to the membrane recruitment of RasGRP1 and Ras engagement, but it is unclear how interactions between RasGRP1 and Ras are suppressed in the absence of such signals. We present a crystal structure of a fragment of RasGRP1 in which the Ras-binding site is blocked by an interdomain linker and the membrane-interaction surface of RasGRP1 is hidden within a dimerization interface that may be stabilized by the C-terminal oligomerization domain. NMR data demonstrate that calcium binding to the regulatory module generates substantial conformational changes that are incompatible with the inactive assembly. These features allow RasGRP1 to be maintained in an inactive state that is poised for activation by calcium and membrane-localization signals. DOI: http://dx.doi.org/10.7554/eLife.00813.001 PMID:23908768

  12. A New View of Ras Isoforms in Cancers.

    PubMed

    Nussinov, Ruth; Tsai, Chung-Jung; Chakrabarti, Mayukh; Jang, Hyunbum

    2016-01-01

    Does small GTPase K-Ras4A have a single state or two states, one resembling K-Ras4B and the other N-Ras? A recent study of K-Ras4A made the remarkable observation that even in the absence of the palmitoyl, K-Ras4A can be active at the plasma membrane. Importantly, this suggests that K-Ras4A may exist in two distinct signaling states. In state 1, K-Ras4A is only farnesylated, like K-Ras4B; in state 2, farnesylated and palmitoylated, like N-Ras. The K-Ras4A hypervariable region sequence is positively charged, in between K-Ras4B and N-Ras. Taken together, this raises the possibility that the farnesylated but nonpalmitoylated state 1, like K-Ras4B, binds calmodulin and is associated with colorectal and other adenocarcinomas like lung cancer and pancreatic ductal adenocarcinoma. On the other hand, state 2 may be associated with melanoma and other cancers where N-Ras is a major contributor, such as acute myeloid leukemia. Importantly, H-Ras has two, singly and doubly, palmitoylated states that may also serve distinct functional roles. The multiple signaling states of palmitoylated Ras isoforms question the completeness of small GTPase Ras isoform statistics in different cancer types and call for reevaluation of concepts and protocols. They may also call for reconsideration of oncogenic Ras therapeutics. PMID:26659836

  13. Molecular alterations of Ras-Raf-mitogen-activated protein kinase and phosphatidylinositol 3-kinase-Akt signaling pathways in colorectal cancers from a tertiary hospital at Kuala Lumpur, Malaysia.

    PubMed

    Yip, Wai Kien; Choo, Chee Wei; Leong, Vincent Ching-Shian; Leong, Pooi Pooi; Jabar, Mohd Faisal; Seow, Heng Fong

    2013-10-01

    Molecular alterations in KRAS, BRAF, PIK3CA, and PTEN have been implicated in designing targeted therapy for colorectal cancer (CRC). The present study aimed to determine the status of these molecular alterations in Malaysian CRCs as such data are not available in the literature. We investigated the mutations of KRAS, BRAF, and PTEN, the gene amplification of PIK3CA, and the protein expression of PTEN and phosphatidylinositol 3-kinase (PI3K) catalytic subunit (p110α) by direct DNA sequencing, quantitative real-time PCR, and immunohistochemistry, respectively, in 49 CRC samples. The frequency of KRAS (codons 12, 13, and 61), BRAF (V600E), and PTEN mutations, and PIK3CA amplification was 25.0% (11/44), 2.3% (1/43), 0.0% (0/43), and 76.7% (33/43), respectively. Immunohistochemical staining demonstrated loss of PTEN protein in 54.5% (24/44) of CRCs and no significant difference in PI3K p110α expression between CRCs and the adjacent normal colonic mucosa (p = 0.380). PIK3CA amplification was not associated with PI3K p110α expression level, but associated with male cases (100% of male cases vs 56% of female cases harbored amplified PIK3CA, p = 0.002). PI3K p110α expression was significantly higher (p = 0.041) in poorly/moderately differentiated carcinoma compared with well-differentiated carcinoma. KRAS mutation, PIK3CA amplification, PTEN loss, and PI3K p110α expression did not correlate with Akt phosphorylation or Ki-67 expression. KRAS mutation, PIK3CA amplification, and PTEN loss were not mutually exclusive. This is the first report on CRC in Malaysia showing comparable frequency of KRAS mutation and PTEN loss, lower BRAF mutation rate, higher PIK3CA amplification frequency, and rare PTEN mutation, as compared with published reports.

  14. Targeting RAS Membrane Association: Back to the Future for Anti-RAS Drug Discovery?

    PubMed Central

    Cox, Adrienne D.; Der, Channing J.; Philips, Mark R.

    2015-01-01

    RAS proteins require membrane association for their biological activity, making this association a logical target for anti-RAS therapeutics. Lipid modification of RAS proteins by a farnesyl isoprenoid is an obligate step in that association, and is an enzymatic process. Accordingly, farnesyltransferase inhibitors (FTIs) were developed as potential anti-RAS drugs. The lack of efficacy of FTIs as anti-cancer drugs was widely seen as indicating that blocking RAS membrane association was a flawed approach to cancer treatment. However, a deeper understanding of RAS modification and trafficking has revealed that this was an erroneous conclusion. In the presence of FTIs, KRAS and NRAS, which are the RAS isoforms most frequently mutated in cancer, become substrates for alternative modification, can still associate with membranes, and can still function. Thus, FTIs failed not because blocking RAS membrane association is an ineffective approach, but because FTIs failed to accomplish that task. Recent findings regarding RAS isoform trafficking and the regulation of RAS subcellular localization have rekindled interest in efforts to target these processes. In particular, improved understanding of the palmitoylation/depalmitoylation cycle that regulates RAS interaction with the plasma membrane, endomembranes and cytosol, and of the potential importance of RAS chaperones, have led to new approaches. Efforts to validate and target other enzymatically regulated post-translational modifications are also ongoing. In this review, we revisit lessons learned, describe the current state of the art, and highlight challenging but promising directions to achieve the goal of disrupting RAS membrane association and subcellular localization for anti-RAS drug development. PMID:25878363

  15. The impact of TP53 and RAS mutations on cerebellar glioblastomas.

    PubMed

    Milinkovic, Vedrana P; Skender Gazibara, Milica K; Manojlovic Gacic, Emilija M; Gazibara, Tatjana M; Tanic, Nikola T

    2014-10-01

    Cerebellar glioblastoma (cGBM) is a rare, inadequately characterized disease, without detailed information on its molecular basis. This is the first report analyzing both TP53 and RAS alterations in cGBM. TP53 mutations were detected in more than half of the samples from our cohort, mainly in hotspot codons. There were no activating mutations in hotspot codons 12/13 and 61 of KRAS and HRAS genes in cGBM samples but we detected alterations in other parts of exons 2 and 3 of these genes, including premature induction of STOP codon. This mutation was present in 3 out of 5 patients. High incidence of RAS mutations, as well as significantly longer survival of cGBM patients compared to those with supratentorial GBM suggest that cGBM may have different mechanisms of occurrence. Our results suggest that inactivation of TP53 and RAS may play an important role in the progression of cerebellar GBM. PMID:25036404

  16. Mapping the functional versatility and fragility of Ras GTPase signaling circuits through in vitro network reconstitution

    PubMed Central

    Coyle, Scott M; Lim, Wendell A

    2016-01-01

    The Ras-superfamily GTPases are central controllers of cell proliferation and morphology. Ras signaling is mediated by a system of interacting molecules: upstream enzymes (GEF/GAP) regulate Ras’s ability to recruit multiple competing downstream effectors. We developed a multiplexed, multi-turnover assay for measuring the dynamic signaling behavior of in vitro reconstituted H-Ras signaling systems. By including both upstream regulators and downstream effectors, we can systematically map how different network configurations shape the dynamic system response. The concentration and identity of both upstream and downstream signaling components strongly impacted the timing, duration, shape, and amplitude of effector outputs. The distorted output of oncogenic alleles of Ras was highly dependent on the balance of positive (GAP) and negative (GEF) regulators in the system. We found that different effectors interpreted the same inputs with distinct output dynamics, enabling a Ras system to encode multiple unique temporal outputs in response to a single input. We also found that different Ras-to-GEF positive feedback mechanisms could reshape output dynamics in distinct ways, such as signal amplification or overshoot minimization. Mapping of the space of output behaviors accessible to Ras provides a design manual for programming Ras circuits, and reveals how these systems are readily adapted to produce an array of dynamic signaling behaviors. Nonetheless, this versatility comes with a trade-off of fragility, as there exist numerous paths to altered signaling behaviors that could cause disease. DOI: http://dx.doi.org/10.7554/eLife.12435.001 PMID:26765565

  17. Targeting Ras-RAF-ERK and its Interactive Pathways as a Novel Therapy for Malignant Gliomas

    PubMed Central

    Lo, H.-W.

    2013-01-01

    Malignant gliomas are the most common and the deadliest brain malignancies in adults. Despite the lack of a complete understanding of the biology of these tumors, significant advances have been made in the past decades. One of the key discoveries made in the area of malignant gliomas is that these tumors can be induced and maintained by aberrant signaling networks. In this context, the Ras pathway has been extensively exploited, from both basic and translational perspectives. Although somatic oncogenic mutations of Ras genes are frequent in several cancer types, early investigations on gliomas revealed disappointing facts that the Ras mutations are nearly absent in malignant gliomas and that the BRAF mutations are present in a very small percentage of gliomas. Therefore, the observed deregulation of the Ras-RAF-ERK signaling pathway in gliomas is attributed to its upstream positive regulators, including, EGFR and PDGFR known to be highly active in the majority of malignant gliomas. In contrast to the initial negative results on the somatic mutations of H-Ras, K-Ras and BRAF, recent breakthrough studies on pediatric low-grade astrocytomas uncovered genetic alterations of the BRAF gene involving copy number gains and rearrangements. The 7q34 rearrangements result in a novel in-frame KIAA1549:BRAF fusion gene that possesses constitutive BRAF kinase activity resembling oncogenic BRAF (V600E). In light of the earlier findings and recent breakthroughs, this review summarizes our current understanding of the Ras-RAF-ERK signaling pathway in gliomas and the outcome of preclinical and clinical studies that evaluated the efficacy of Ras-targeted therapy in malignant gliomas. PMID:20718706

  18. Conserved cis-elements bind a protein complex that regulates Drosophila ras2/rop bidirectional expression.

    PubMed Central

    Lightfoot, K.; Maltby, L.; Duarte, R.; Veale, R.; Segev, O.

    1994-01-01

    The Drosophila ras2 promoter region exhibits bidirectional activity, as has been demonstrated for the human c-Ha-ras1 and the mouse c-Ki-ras. Here we address a unique case of ras regulation, as Drosophila ras2 provides the only example to date in which the flanking gene (rop) and its product have been isolated. A linking mechanism of control suggests a mutual interaction between the two gene products. Our studies indicate that the Drosophila ras2 promoter region shares with the human c-Ha-ras1 promoter a CACCC box and an AP-1-like sequence. A 14 bp promoter fragment which holds a CACCC element is demonstrated to interact with a specific transcription factor (factor B). This CACCC promoter element represents a stretch of imperfect palindrome. We present evidence that this factor can form a complex with another specific DNA-binding protein (factor A). The binding sites (A + B) for these protein factors are essential for 95% expression of both genes flanking the promoter (ras2 and rop). Region A consists of four overlapping consensus sequences: a TATA-like element, a DSE-like motif (the core sequence of the serum response element), a DRE octamer, which has been shown to play a role in cell proliferation, and a 5 bp direct repeat representing the GATA consensus sequence. Factor A has a very weak affinity to the full promoter region, but when complexed with factor B binding efficiency is enhanced. We also show that alterations of DNA-protein binding specificities can be achieved by supplementing the growth media with different sera. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:8297724

  19. Mitochondrial dysfunction and organic aciduria in five patients carrying mutations in the Ras-MAPK pathway

    PubMed Central

    Kleefstra, Tjitske; Wortmann, Saskia B; Rodenburg, Richard J T; Bongers, Ernie M H F; Hadzsiev, Kinga; Noordam, Cees; van den Heuvel, Lambert P; Nillesen, Willy M; Hollody, Katalin; Gillessen-Kaesbach, Gabrielle; Lammens, Martin; Smeitink, Jan A M; van der Burgt, Ineke; Morava, Eva

    2011-01-01

    Various syndromes of the Ras-mitogen-activated protein kinase (MAPK) pathway, including the Noonan, Cardio-Facio-Cutaneous, LEOPARD and Costello syndromes, share the common features of craniofacial dysmorphisms, heart defect and short stature. In a subgroup of patients, severe muscle hypotonia, central nervous system involvement and failure to thrive occur as well. In this study we report on five children diagnosed initially with classic metabolic and clinical symptoms of an oxidative phosphorylation disorder. Later in the course of the disease, the children presented with characteristic features of Ras-MAPK pathway-related syndromes, leading to the reevaluation of the initial diagnosis. In the five patients, in addition to the oxidative phosphorylation disorder, disease-causing mutations were detected in the Ras-MAPK pathway. Three of the patients also carried a second, mitochondrial genetic alteration, which was asymptomatically present in their healthy relatives. Did we miss the correct diagnosis in the first place or is mitochondrial dysfunction directly related to Ras-MAPK pathway defects? The Ras-MAPK pathway is known to have various targets, including proteins in the mitochondrial membrane influencing mitochondrial morphology and dynamics. Prospective screening of 18 patients with various Ras-MAPK pathway defects detected biochemical signs of disturbed oxidative phosphorylation in three additional children. We concluded that only a specific, metabolically vulnerable sub-population of patients with Ras-MAPK pathway mutations presents with mitochondrial dysfunction and a more severe, early-onset disease. We postulate that patients with Ras-MAPK mutations have an increased susceptibility, but a second metabolic hit is needed to cause the clinical manifestation of mitochondrial dysfunction. PMID:21063443

  20. MicroRNA-based Therapeutic Strategies for Targeting Mutant and Wild Type RAS in Cancer

    PubMed Central

    Sharma, Sriganesh B.; Ruppert, J. Michael

    2015-01-01

    MicroRNAs (miRs) have been causally implicated in the progression and development of a wide variety of cancers. miRs modulate the activity of key cell signaling networks by regulating the translation of pathway component proteins. Thus, the pharmacological targeting of miRs that regulate cancer cell signaling networks, either by promoting (using miR-supplementation) or by suppressing (using anti-sense oligonucleotide based strategies) miR activity is an area of intense research. The RAS-Extracellular signal regulated kinase (ERK) pathway represents a major miR-regulated signaling network that endows cells with some of the classical hallmarks of cancer, and is often inappropriately activated in malignancies by somatic genetic alteration through point mutation or alteration of gene copy number. In addition, recent progress indicates that many tumors may be deficient in GTPase activating proteins (GAPs) due to the collaborative action of oncogenic microRNAs. Recent studies also suggest that in tumors harboring a mutant RAS allele there is a critical role for wild type RAS proteins in determining overall RAS-ERK pathway activity. Together, these two advances comprise a new opportunity for therapeutic intervention. In this review, we evaluate miR-based therapeutic strategies for modulating RAS-ERK signaling in cancers, in particular for more direct modulation of RAS-GTP levels, with the potential to complement current strategies in order to yield more durable treatment responses. To this end, we discuss the potential for miR-based therapies focused on three prominent miRs including the pan-RAS regulator let-7 and the GAP regulator comprised of miR-206 and miR-21 (miR-206/21). PMID:26284568

  1. Mutations, expression and genomic instability of the H-ras proto-oncogene in squamous cell carcinomas of the head and neck.

    PubMed Central

    Kiaris, H.; Spandidos, D. A.; Jones, A. S.; Vaughan, E. D.; Field, J. K.

    1995-01-01

    Mutation and overexpression are the main activating mechanisms for the ras family of genes in human cancer and the variable tandem repeat (VTR) located at the 3' end of H-ras has been associated with this risk. In the present study, we have analysed the relative levels of expression of H-ras mRNA in 26 samples of squamous cell carcinomas of the head and neck (SCCHN) by competitive reverse transcription-polymerase chain reaction (competitive RT-PCR) and also investigated whether there is an association between ras expression and alterations in the 3'-VTR region. In addition, we have studied the incidence of point mutations in codon 12 of H-ras, codons 12 and 13 of K-ras and codon 61 of N-ras in 120 SCCHN samples. Our results indicate that only two samples carry mutations, both of which are located in codon 12 of K-ras, but that overexpression of the H-ras proto-oncogene is a frequent event in SCCHN [54% (14/26)] and is associated with a favourable prognosis: 3 of 14 patients with H-ras overexpression have died, whereas 9 of 12 patients with low levels of H-ras expression have died. We have also undertaken an analysis of these results together with our previous investigations on microsatellite instability and loss of heterozygosity in SCCHN, but no associations were found. We therefore conclude that ras mutations are an infrequent event in the progression of the SCCHN in the Western world, whereas overexpression of the H-ras proto-oncogene is a common event. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:7599040

  2. Involvement of phosphatidylinositol 3-kinase, but not RalGDS, in TC21/R-Ras2-mediated transformation.

    PubMed

    Murphy, Gretchen A; Graham, Suzanne M; Morita, Staeci; Reks, Sarah E; Rogers-Graham, Kelley; Vojtek, Anne; Kelley, Grant G; Der, Channing J

    2002-03-22

    Oncogenic Ras and activated forms of the Ras-related protein TC21/R-Ras2 share similar abilities to alter cell proliferation. However, in contrast to Ras, we found previously that TC21 fails to activate the Raf-1 serine/threonine kinase. Thus, TC21 must utilize non-Raf effectors to regulate cell function. In this study, we determined that TC21 interacts strongly with some (RalGDS, RGL, RGL2/Rlf, AF6, and the phosphatidylinositol 3-kinase (PI3K) catalytic subunit p110delta), and weakly with other Ras small middle dotGTP-binding proteins. In addition, library screening identified novel TC21-interacting proteins. We also determined that TC21, similar to Ras, mediates activation of phospholipase Cepsilon. We then examined if RalGDS, a RalA guanine nucleotide exchange factor, or PI3K are effectors for TC21-mediated signaling and cell proliferation in murine fibroblasts. We found that overexpression of full-length RalGDS reduced the focus forming activity of activated TC21. Furthermore, expression of activated Ras, but not TC21, enhanced GTP loading on RalA. In fact, TC21 attenuated insulin-stimulated RalA small middle dotGTP formation. In contrast, like Ras, expression of activated TC21 resulted in membrane translocation and an increase in the PI3K-dependent phosphorylation of Akt, and inhibition of PI3K activity interfered with TC21 focus formation. Finally, unlike Ras, TC21 did not activate the Rac small GTPase, indicating that Ras may not activate Rac by PI3K. Taken together, these results suggest that PI3K, but not RalGDS, is an important mediator of cell proliferation by TC21.

  3. Biochemical similarity of Schizosaccharomyces pombe ras1 protein with RAS2 protein of Saccharomyces cervisiae.

    PubMed

    Onozawa, T; Danjoh, I; Fujiyama, A

    1995-07-01

    Schizosaccharomyces pombe contains single ras oncogene homologue, ras1, that functions in the signal transduction pathway conducting the cell's mating processes. To understand the biochemical basis of yeast ras proteins, we have purified the ras1 protein and compared the major biochemical constants with those of RAS2 protein from Saccharomyces cerevisiae and mammalian ras proteins. The purified ras1 protein showed a remarkably high Kd value for GDP binding (178 nM) and for binding with ATP. In contrast, the Kd value for GTP binding and the rate of GTPase activity were 64 nM and 77 x 10(-6) s-1 at 37 degrees C, respectively; both were higher than normal p21ras protein, but at the same level as the RAS2 protein. We directly measured rate of GTP binding and GDP binding which were 3.9 x 10(-3) s-1 and 1.8 x 10(-3) s-1 at 30 degrees C, respectively. On the other hand, exchange rates between bound and free nucleotides remained almost constant throughout the tested combination of GTP and GDP, and were several-fold lower than the binding rate. These results suggest that the release of the guanine nucleotide is the rate-limiting step in the ras-GTP/GDP cycle. As a whole, the biochemical properties of the ras1 protein are close to those of the RAS2 protein, although these two proteins function differently in the signal transduction pathway in the cells. PMID:7483844

  4. Phosphorylation of synaptic GTPase-activating protein (synGAP) by Ca2+/calmodulin-dependent protein kinase II (CaMKII) and cyclin-dependent kinase 5 (CDK5) alters the ratio of its GAP activity toward Ras and Rap GTPases.

    PubMed

    Walkup, Ward G; Washburn, Lorraine; Sweredoski, Michael J; Carlisle, Holly J; Graham, Robert L; Hess, Sonja; Kennedy, Mary B

    2015-02-20

    synGAP is a neuron-specific Ras and Rap GTPase-activating protein (GAP) found in high concentrations in the postsynaptic density (PSD) fraction from the mammalian forebrain. We have previously shown that, in situ in the PSD fraction or in recombinant form in Sf9 cell membranes, synGAP is phosphorylated by Ca(2+)/calmodulin-dependent protein kinase II (CaMKII), another prominent component of the PSD. Here, we show that recombinant synGAP (r-synGAP), lacking 102 residues at the N terminus, can be purified in soluble form and is phosphorylated by cyclin-dependent kinase 5 (CDK5) as well as by CaMKII. Phosphorylation of r-synGAP by CaMKII increases its HRas GAP activity by 25% and its Rap1 GAP activity by 76%. Conversely, phosphorylation by CDK5 increases r-synGAP's HRas GAP activity by 98% and its Rap1 GAP activity by 20%. Thus, phosphorylation by both kinases increases synGAP activity; CaMKII shifts the relative GAP activity toward inactivation of Rap1, and CDK5 shifts the relative activity toward inactivation of HRas. GAP activity toward Rap2 is not altered by phosphorylation by either kinase. CDK5 phosphorylates synGAP primarily at two sites, Ser-773 and Ser-802. Phosphorylation at Ser-773 inhibits r-synGAP activity, and phosphorylation at Ser-802 increases it. However, the net effect of concurrent phosphorylation of both sites, Ser-773 and Ser-802, is an increase in GAP activity. synGAP is phosphorylated at Ser-773 and Ser-802 in the PSD fraction, and its phosphorylation by CDK5 and CaMKII is differentially regulated by activation of NMDA-type glutamate receptors in cultured neurons.

  5. Contact Lens Care

    MedlinePlus

    ... For Consumers Consumer Information by Audience For Women Contact Lens Care Share Tweet Linkedin Pin it More ... 1088, www.fda.gov/medwatch Learn More about Contact Lens Care Other Tips on Contact Lenses Decorative ...

  6. Contact Lens Solution Toxicity

    MedlinePlus

    ... rash and rashes clinical tools newsletter | contact Share | Contact Lens Solution Toxicity Information for adults A A A This image shows a reaction to contact lens solution. The prominent blood vessels and redness ...

  7. A Small Molecule RAS-Mimetic Disrupts RAS Association with Effector Proteins to Block Signaling.

    PubMed

    Athuluri-Divakar, Sai Krishna; Vasquez-Del Carpio, Rodrigo; Dutta, Kaushik; Baker, Stacey J; Cosenza, Stephen C; Basu, Indranil; Gupta, Yogesh K; Reddy, M V Ramana; Ueno, Lynn; Hart, Jonathan R; Vogt, Peter K; Mulholland, David; Guha, Chandan; Aggarwal, Aneel K; Reddy, E Premkumar

    2016-04-21

    Oncogenic activation of RAS genes via point mutations occurs in 20%-30% of human cancers. The development of effective RAS inhibitors has been challenging, necessitating new approaches to inhibit this oncogenic protein. Functional studies have shown that the switch region of RAS interacts with a large number of effector proteins containing a common RAS-binding domain (RBD). Because RBD-mediated interactions are essential for RAS signaling, blocking RBD association with small molecules constitutes an attractive therapeutic approach. Here, we present evidence that rigosertib, a styryl-benzyl sulfone, acts as a RAS-mimetic and interacts with the RBDs of RAF kinases, resulting in their inability to bind to RAS, disruption of RAF activation, and inhibition of the RAS-RAF-MEK pathway. We also find that ribosertib binds to the RBDs of Ral-GDS and PI3Ks. These results suggest that targeting of RBDs across multiple signaling pathways by rigosertib may represent an effective strategy for inactivation of RAS signaling. PMID:27104980

  8. NAM: The 2004 RAS National Astronomy Meeting

    NASA Astrophysics Data System (ADS)

    Jones, Barrie; Norton, Andrew

    2004-06-01

    This year's RAS National Astronomy Meeting was held at the Open University's Milton Keynes campus from 29 March to 2 April. The event was organized by members of the OU Physics & Astronomy Department and Planetary & Space Science Research Institute. Around 450 people attended the meeting, at which more than 220 talks were presented, along with around 90 posters. Co-chairs of RAS NAM04, Barrie Jones and Andrew Norton, summarize.

  9. Inhibition of RAS in diabetic nephropathy

    PubMed Central

    Yacoub, Rabi; Campbell, Kirk N

    2015-01-01

    Diabetic kidney disease (DKD) is a progressive proteinuric renal disorder in patients with type 1 or type 2 diabetes mellitus. It is a common cause of end-stage kidney disease worldwide, particularly in developed countries. Therapeutic targeting of the renin–angiotensin system (RAS) is the most validated clinical strategy for slowing disease progression. DKD is paradoxically a low systematic renin state with an increased intrarenal RAS activity implicated in its pathogenesis. Angiotensin II (AngII), the main peptide of RAS, is not only a vasoactive peptide but functions as a growth factor, activating interstitial fibroblasts and mesangial and tubular cells, while promoting the synthesis of extracellular matrix proteins. AngII also promotes podocyte injury through increased calcium influx and the generation of reactive oxygen species. Blockade of the RAS using either angiotensin converting enzyme inhibitors, or angiotensin receptor blockers can attenuate progressive glomerulosclerosis in animal models, and slows disease progression in humans with DKD. In this review, we summarize the role of intrarenal RAS activation in the pathogenesis and progression of DKD and the rationale for RAS inhibition in this population. PMID:25926752

  10. ASPP1 and ASPP2 bind active RAS, potentiate RAS signalling and enhance p53 activity in cancer cells

    PubMed Central

    Wang, Y; Godin-Heymann, N; Dan Wang, X; Bergamaschi, D; Llanos, S; Lu, X

    2013-01-01

    RAS mutations occur frequently in human cancer and activated RAS signalling contributes to tumour development and progression. Apart from its oncogenic effects on cell growth, active RAS has tumour-suppressive functions via its ability to induce cellular senescence and apoptosis. RAS is known to induce p53-dependent cell cycle arrest, yet its effect on p53-dependent apoptosis remains unclear. We report here that apoptosis-stimulating protein of p53 (ASPP) 1 and 2, two activators of p53, preferentially bind active RAS via their N-terminal RAS-association domains (RAD). Additionally, ASPP2 colocalises with and contributes to RAS cellular membrane localisation and potentiates RAS signalling. In cancer cells, ASPP1 and ASPP2 cooperate with oncogenic RAS to enhance the transcription and apoptotic function of p53. Thus, loss of ASPP1 and ASPP2 in human cancer cells may contribute to the full transforming property of RAS oncogene. PMID:23392125

  11. Small Molecule APY606 Displays Extensive Antitumor Activity in Pancreatic Cancer via Impairing Ras-MAPK Signaling

    PubMed Central

    Guo, Na; Liu, Zuojia; Zhao, Wenjing; Wang, Erkang; Wang, Jin

    2016-01-01

    Pancreatic cancer has been found with abnormal expression or mutation in Ras proteins. Oncogenic Ras activation exploits their extensive signaling reach to affect multiple cellular processes, in which the mitogen-activated protein kinase (MAPK) signaling exerts important roles in tumorigenesis. Therapies targeted Ras are thus of major benefit for pancreatic cancer. Although small molecule APY606 has been successfully picked out by virtual drug screening based on Ras target receptor, its in-depth mechanism remains to be elucidated. We herein assessed the antitumor activity of APY606 against human pancreatic cancer Capan-1 and SW1990 cell lines and explored the effect of Ras-MAPK and apoptosis-related signaling pathway on the activity of APY606. APY606 treatment resulted in a dose- and time-dependent inhibition of cancer cell viability. Additionally, APY606 exhibited strong antitumor activity, as evidenced not only by reduction in tumor cell invasion, migration and mitochondrial membrane potential but also by alteration in several apoptotic indexes. Furthermore, APY606 treatment directly inhibited Ras-GTP and the downstream activation of MAPK, which resulted in the down-regulation of anti-apoptotic protein Bcl-2, leading to the up-regulation of mitochondrial apoptosis pathway-related proteins (Bax, cytosolic Cytochrome c and Caspase 3) and of cyclin-dependent kinase 2 and Cyclin A, E. These data suggest that impairing Ras-MAPK signaling is a novel mechanism of action for APY606 during therapeutic intervention in pancreatic cancer. PMID:27223122

  12. Carcinogen and dietary lipid regulate ras expression and localization in rat colon without affecting farnesylation kinetics.

    PubMed

    Davidson, L A; Lupton, J R; Jiang, Y H; Chapkin, R S

    1999-05-01

    Epidemiological and experimental data suggest that dietary fiber and fat are major determinants of colorectal cancer. However, the mechanisms by which these dietary constituents alter the incidence of colon cancer have not been elucidated. Evidence indicates that dominant gain-of-function mutations short-circuit protooncogenes and contribute to the pathogenesis of cancer. Therefore, we began to dissect the mechanisms whereby dietary fat and fiber, fed during the initiation, promotion and progression stages of colon tumorigenesis, regulate ras p21 localization, expression and mutation frequency. Male Sprague-Dawley rats (140) were provided with corn oil or fish oil and pectin or cellulose plus or minus the carcinogen azoxymethane (AOM) in a 2 x 2 x 2 factorial design and killed after 34 weeks. We have previously shown adenocarcinoma incidence in these animals to be 70.3% (52/74) for corn oil + AOM and 56.1% (37/66) for fish oil + AOM (P < 0.05). Total ras expression as well as ras membrane:cytosol ratio was 4- to 6-fold higher in colon tumors than in mucosa from AOM- or saline-injected rats. Expression of ras in the mucosal membrane fraction was 13% higher for animals fed corn oil compared with fish oil feeding (P < 0.05), which is noteworthy since ras must be localized at the plasma membrane to function. The elevated ras membrane:cytosol ratio in tumors was not due to increased farnesyl protein transferase activity or prenylation state, as nearly all detectable ras was in the prenylated form. Phosphorylated p42 and p44 mitogen activated protein kinase (ERK) expression was two-fold higher in tumor extracts compared with uninvolved mucosa from AOM- and saline-injected rats (P < 0.05). The frequency of K-ras mutations was not significantly different between the various groups, but there was a trend toward a greater incidence of mutations in tumors from corn oil fed rats (85%) compared with fish oil fed rats (58%). Our results indicate that the carcinogen

  13. Aberration design of zoom lens systems using thick lens modules.

    PubMed

    Zhang, Jinkai; Chen, Xiaobo; Xi, Juntong; Wu, Zhuoqi

    2014-12-20

    A systematic approach for the aberration design of a zoom lens system using a thick lens module is presented. Each component is treated as a thick lens module at the beginning of the design. A thick lens module refers to a thick lens component with a real lens structure, like lens materials, lens curvatures, lens thicknesses, and lens interval distances. All nine third-order aberrations of a thick lens component are considered during the design. The relationship of component aberrations in different zoom positions can be approximated from the aberration shift. After minimizing the aberrations of the zoom lens system, the nine third-order aberrations of every lens component can be determined. Then the thick lens structure of every lens component can be determined after optimization according to their first-order properties and third-order aberration targets. After a third optimization for minimum practical third-order aberrations of a zoom lens system, the aberration design using the thick lens module is complete, which provides a practical zoom lens system with thick lens structures. A double-sided telecentric zoom lens system is designed using the thick lens module in this paper, which shows that this method is practical for zoom lens design.

  14. Role of Aquaporin 0 in lens biomechanics

    SciTech Connect

    Sindhu Kumari, S.; Gupta, Neha; Shiels, Alan; FitzGerald, Paul G.; Menon, Anil G.; Mathias, Richard T.; Varadaraj, Kulandaiappan

    2015-07-10

    Maintenance of proper biomechanics of the eye lens is important for its structural integrity and for the process of accommodation to focus near and far objects. Several studies have shown that specialized cytoskeletal systems such as the beaded filament (BF) and spectrin-actin networks contribute to mammalian lens biomechanics; mutations or deletion in these proteins alters lens biomechanics. Aquaporin 0 (AQP0), which constitutes ∼45% of the total membrane proteins of lens fiber cells, has been shown to function as a water channel and a structural cell-to-cell adhesion (CTCA) protein. Our recent ex vivo study on AQP0 knockout (AQP0 KO) mouse lenses showed the CTCA function of AQP0 could be crucial for establishing the refractive index gradient. However, biomechanical studies on the role of AQP0 are lacking. The present investigation used wild type (WT), AQP5 KO (AQP5{sup −/−}), AQP0 KO (heterozygous KO: AQP0{sup +/−}; homozygous KO: AQP0{sup −/−}; all in C57BL/6J) and WT-FVB/N mouse lenses to learn more about the role of fiber cell AQPs in lens biomechanics. Electron microscopic images exhibited decreases in lens fiber cell compaction and increases in extracellular space due to deletion of even one allele of AQP0. Biomechanical assay revealed that loss of one or both alleles of AQP0 caused a significant reduction in the compressive load-bearing capacity of the lenses compared to WT lenses. Conversely, loss of AQP5 did not alter the lens load-bearing ability. Compressive load-bearing at the suture area of AQP0{sup +/−} lenses showed easy separation while WT lens suture remained intact. These data from KO mouse lenses in conjunction with previous studies on lens-specific BF proteins (CP49 and filensin) suggest that AQP0 and BF proteins could act co-operatively in establishing normal lens biomechanics. We hypothesize that AQP0, with its prolific expression at the fiber cell membrane, could provide anchorage for cytoskeletal structures like BFs and

  15. K-Ras(V14I) -induced Noonan syndrome predisposes to tumour development in mice.

    PubMed

    Hernández-Porras, Isabel; Schuhmacher, Alberto J; Garcia-Medina, Raquel; Jiménez, Beatriz; Cañamero, Marta; de Martino, Alba; Guerra, Carmen

    2016-06-01

    The Noonan syndrome (NS) is an autosomal dominant genetic disorder characterized by short stature, craniofacial dysmorphism, and congenital heart defects. A significant proportion of NS patients may also develop myeloproliferative disorders (MPDs), including juvenile myelomonocytic leukaemia (JMML). Surprisingly, scarce information is available in relation to other tumour types in these patients. We have previously developed and characterized a knock-in mouse model that carries one of the most frequent KRAS-NS-related mutations, the K-Ras(V14I) substitution, which recapitulates most of the alterations described in NS patients, including MPDs. The K-Ras(V14I) mutation is a mild activating K-Ras protein; thus, we have used this model to study tumour susceptibility in comparison with mice expressing the classical K-Ras(G12V) oncogene. Interestingly, our studies have shown that these mice display a generalized tumour predisposition and not just MPDs. In fact, we have observed that the K-Ras(V14I) mutation is capable of cooperating with the p16Ink4a/p19Arf and Trp53 tumour suppressors, as well as with other risk factors such as pancreatitis, thereby leading to a higher cancer incidence. In conclusion, our results illustrate that the K-Ras(V14I) activating protein is able to induce cancer, although at a much lower level than the classical K-Ras(G12V) oncogene, and that it can be significantly modulated by both genetic and non-genetic events. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. PMID:27174785

  16. K-Ras(V14I) -induced Noonan syndrome predisposes to tumour development in mice.

    PubMed

    Hernández-Porras, Isabel; Schuhmacher, Alberto J; Garcia-Medina, Raquel; Jiménez, Beatriz; Cañamero, Marta; de Martino, Alba; Guerra, Carmen

    2016-06-01

    The Noonan syndrome (NS) is an autosomal dominant genetic disorder characterized by short stature, craniofacial dysmorphism, and congenital heart defects. A significant proportion of NS patients may also develop myeloproliferative disorders (MPDs), including juvenile myelomonocytic leukaemia (JMML). Surprisingly, scarce information is available in relation to other tumour types in these patients. We have previously developed and characterized a knock-in mouse model that carries one of the most frequent KRAS-NS-related mutations, the K-Ras(V14I) substitution, which recapitulates most of the alterations described in NS patients, including MPDs. The K-Ras(V14I) mutation is a mild activating K-Ras protein; thus, we have used this model to study tumour susceptibility in comparison with mice expressing the classical K-Ras(G12V) oncogene. Interestingly, our studies have shown that these mice display a generalized tumour predisposition and not just MPDs. In fact, we have observed that the K-Ras(V14I) mutation is capable of cooperating with the p16Ink4a/p19Arf and Trp53 tumour suppressors, as well as with other risk factors such as pancreatitis, thereby leading to a higher cancer incidence. In conclusion, our results illustrate that the K-Ras(V14I) activating protein is able to induce cancer, although at a much lower level than the classical K-Ras(G12V) oncogene, and that it can be significantly modulated by both genetic and non-genetic events. Copyright © 2016 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  17. Signaling through mitogen-activated protein kinase and Rac/Rho does not duplicate the effects of activated Ras on skeletal myogenesis.

    PubMed

    Ramocki, M B; Johnson, S E; White, M A; Ashendel, C L; Konieczny, S F; Taparowsky, E J

    1997-07-01

    The ability of basic helix-loop-helix muscle regulatory factors (MRFs), such as MyoD, to convert nonmuscle cells to a myogenic lineage is regulated by numerous growth factor and oncoprotein signaling pathways. Previous studies have shown that H-Ras 12V inhibits differentiation to a skeletal muscle lineage by disrupting MRF function via a mechanism that is independent of the dimerization, DNA binding, and inherent transcriptional activation properties of the proteins. To investigate the intracellular signaling pathway(s) that mediates the inhibition of MRF-induced myogenesis by oncogenic Ras, we tested two transformation-defective H-Ras 12V effector domain variants for their ability to alter terminal differentiation. H-Ras 12V,35S retains the ability to activate the Raf/MEK/mitogen-activated protein (MAP) kinase cascade, whereas H-Ras 12V,40C is unable to interact directly with Raf-1 yet still influences other signaling intermediates, including Rac and Rho. Expression of each H-Ras 12V variant in C3H10T1/2 cells abrogates MyoD-induced activation of the complete myogenic program, suggesting that MAP kinase-dependent and -independent Ras signaling pathways individually block myogenesis in this model system. However, additional studies with constitutively activated Rac1 and RhoA proteins revealed no negative effects on MyoD-induced myogenesis. Similarly, treatment of Ras-inhibited myoblasts with the MEK1 inhibitor PD98059 revealed that elevated MAP kinase activity is not a significant contributor to the H-Ras 12V effect. These data suggest that an additional Ras pathway, distinct from the well-characterized MAP kinase and Rac/Rho pathways known to be important for the transforming function of activated Ras, is primarily responsible for the inhibition of myogenesis by H-Ras 12V.

  18. VPS35 binds farnesylated N-Ras in the cytosol to regulate N-Ras trafficking.

    PubMed

    Zhou, Mo; Wiener, Heidi; Su, Wenjuan; Zhou, Yong; Liot, Caroline; Ahearn, Ian; Hancock, John F; Philips, Mark R

    2016-08-15

    Ras guanosine triphosphatases (GTPases) regulate signaling pathways only when associated with cellular membranes through their C-terminal prenylated regions. Ras proteins move between membrane compartments in part via diffusion-limited, fluid phase transfer through the cytosol, suggesting that chaperones sequester the polyisoprene lipid from the aqueous environment. In this study, we analyze the nature of the pool of endogenous Ras proteins found in the cytosol. The majority of the pool consists of farnesylated, but not palmitoylated, N-Ras that is associated with a high molecular weight (HMW) complex. Affinity purification and mass spectrographic identification revealed that among the proteins found in the HMW fraction is VPS35, a latent cytosolic component of the retromer coat. VPS35 bound to N-Ras in a farnesyl-dependent, but neither palmitoyl- nor guanosine triphosphate (GTP)-dependent, fashion. Silencing VPS35 increased N-Ras's association with cytoplasmic vesicles, diminished GTP loading of Ras, and inhibited mitogen-activated protein kinase signaling and growth of N-Ras-dependent melanoma cells. PMID:27502489

  19. Introduction of v-Ha-ras oncogene induces differentiation of cultured human medullary thyroid carcinoma cells

    SciTech Connect

    Nakagawa, T.; Mabry, M.; De Bustros, A.; Ihle, J.N.; Nelkin, B.D.; Baylin, S.B.

    1987-08-01

    Medullary thyroid carcinoma (MTC) is an endocrine tumor of the thyroid C cells that expresses high levels of the neuroendocrine peptide hormone calcitonin. During tumor progression in the host, there is an apparent loss of differentiation in MTC cells that involves a consistent decrease in calcitonin content of the tumor cells associated with decreased expression of the calcitonin gene and/or changes in a mRNA alternative-processing pattern away from that characteristic of the parent thyroid C cell. The authors now report that introduction of the viral Harvey ras (v-Ha-ras) oncogene into cultured human MTC cells can reverse such changes in gene expression and can induce endocrine differentiation of the tumor cells. The expression of v-Ha-ras is associated with decreased cellular proliferation and DNA synthesis. There is a marked increase in the number of cytoplasmic secretory granules that are a classic feature of differentiated thyroid C cells. v-Ha-ras expression induces increased expression of the calcitonin gene and the processing of the primary gene transcript is shifted to favor calcitonin mRNA rather than calcitonin-gene-related peptide (CGRP) mRNA production. These studies with cultured human MTC cells provide a model system to study the role of Ha-ras and related genes in neuroendocrine differentiation. The findings suggest an important approach for identifying genes in solid tumors whose altered expression may play a role in the impaired maturational capacity characteristic of cancer cells during tumor progression.

  20. Juvenile myelomonocytic leukemia displays mutations in components of the RAS pathway and the PRC2 network.

    PubMed

    Caye, Aurélie; Strullu, Marion; Guidez, Fabien; Cassinat, Bruno; Gazal, Steven; Fenneteau, Odile; Lainey, Elodie; Nouri, Kazem; Nakhaei-Rad, Saeideh; Dvorsky, Radovan; Lachenaud, Julie; Pereira, Sabrina; Vivent, Jocelyne; Verger, Emmanuelle; Vidaud, Dominique; Galambrun, Claire; Picard, Capucine; Petit, Arnaud; Contet, Audrey; Poirée, Marilyne; Sirvent, Nicolas; Méchinaud, Françoise; Adjaoud, Dalila; Paillard, Catherine; Nelken, Brigitte; Reguerre, Yves; Bertrand, Yves; Häussinger, Dieter; Dalle, Jean-Hugues; Ahmadian, Mohammad Reza; Baruchel, André; Chomienne, Christine; Cavé, Hélène

    2015-11-01

    Juvenile myelomonocytic leukemia (JMML) is a rare and severe myelodysplastic and myeloproliferative neoplasm of early childhood initiated by germline or somatic RAS-activating mutations. Genetic profiling and whole-exome sequencing of a large JMML cohort (118 and 30 cases, respectively) uncovered additional genetic abnormalities in 56 cases (47%). Somatic events were rare (0.38 events/Mb/case) and restricted to sporadic (49/78; 63%) or neurofibromatosis type 1 (NF1)-associated (8/8; 100%) JMML cases. Multiple concomitant genetic hits targeting the RAS pathway were identified in 13 of 78 cases (17%), disproving the concept of mutually exclusive RAS pathway mutations and defining new pathways activated in JMML involving phosphoinositide 3-kinase (PI3K) and the mTORC2 complex through RAC2 mutation. Furthermore, this study highlights PRC2 loss (26/78; 33% of sporadic JMML cases) that switches the methylation/acetylation status of lysine 27 of histone H3 in JMML cases with altered RAS and PRC2 pathways. Finally, the association between JMML outcome and mutational profile suggests a dose-dependent effect for RAS pathway activation, distinguishing very aggressive JMML rapidly progressing to acute myeloid leukemia. PMID:26457648

  1. RasGRP1 and RasGRP3 Are Required for Efficient Generation of Early Thymic Progenitors.

    PubMed

    Golec, Dominic P; Henao Caviedes, Laura M; Baldwin, Troy A

    2016-09-01

    T cell development is dependent on the migration of progenitor cells from the bone marrow to the thymus. Upon reaching the thymus, progenitors undergo a complex developmental program that requires inputs from various highly conserved signaling pathways including the Notch and Wnt pathways. To date, Ras signaling has not been implicated in the very earliest stages of T cell differentiation, but members of a family of Ras activators called RasGRPs have been shown to be involved at multiple stages of T cell development. We examined early T cell development in mice lacking RasGRP1, RasGRP3, and RasGRPs 1 and 3. We report that RasGRP1- and RasGRP3-deficient thymi show significantly reduced numbers of early thymic progenitors (ETPs) relative to wild type thymi. Furthermore, RasGRP1/3 double-deficient thymi show significant reductions in ETP numbers compared with either RasGRP1 or RasGRP3 single-deficient thymi, suggesting that both RasGRP1 and RasGRP3 regulate the generation of ETPs. In addition, competitive bone marrow chimera experiments reveal that RasGRP1/3 double-deficient progenitors intrinsically generate ETPs less efficiently than wild type progenitors. Finally, RasGRP1/3-deficient progenitors show impaired migration toward the CCR9 ligand, CCL25, suggesting that RasGRP1 and RasGRP3 may regulate progenitor entry into the thymus through a CCR9-dependent mechanism. These data demonstrate that, in addition to Notch and Wnt, the highly conserved Ras pathway is critical for the earliest stages of T cell development and further highlight the importance of Ras signaling during thymocyte maturation. PMID:27465532

  2. Cooperative interactions of PTEN deficiency and RAS activation in melanoma metastasis.

    PubMed

    Kim, Minjung

    2010-11-01

    Melanoma displays frequent activation of RAS/RAF/MAPK and PI3K/AKT signaling pathways as well as inactivation of CDKN2A (INK4a/ARF) and PTEN tumor suppressors via genetic and epigenetic alterations. Pathogenetic roles of these melanoma-prone mutations and their genetic interactions have been established in genetically engineered mouse models. Here, we catalog frequent genetic alterations observed in human melanomas and describe mouse models of melanoma initiation and progression, including our recent study that investigated the genetic interactions of RAS activation and PTEN loss in a CDKN2A (INK4a/ARF) null melanoma prone genetic background. We showed that loss of PTEN cooperates with HRAS activation, leading to increased development of melanoma and emergence of metastasis. Moreover, we observed that RNA i-mediated PTEN inactivation in RAS-driven melanomas enhanced migration and invasion with concomitant downregulation of E-cadherin, the major regulator of epithelial and mesenchymal transition, and enhanced AKT2 phosphorylation, which has been previously linked to invasion and metastasis of several cancer types, including breast and ovary. These data show that activated RAS cooperates with PTEN loss in melanoma genesis and progression.

  3. The neurofibromin recruitment factor Spred1 binds to the GAP related domain without affecting Ras inactivation

    PubMed Central

    Dunzendorfer-Matt, Theresia; Mercado, Ellen L.; Maly, Karl; McCormick, Frank; Scheffzek, Klaus

    2016-01-01

    Neurofibromatosis type 1 (NF1) and Legius syndrome are related diseases with partially overlapping symptoms caused by alterations of the tumor suppressor genes NF1 (encoding the protein neurofibromin) and SPRED1 (encoding sprouty-related, EVH1 domain-containing protein 1, Spred1), respectively. Both proteins are negative regulators of Ras/MAPK signaling with neurofibromin functioning as a Ras-specific GTPase activating protein (GAP) and Spred1 acting on hitherto undefined components of the pathway. Importantly, neurofibromin has been identified as a key protein in the development of cancer, as it is genetically altered in a large number of sporadic human malignancies unrelated to NF1. Spred1 has previously been demonstrated to interact with neurofibromin via its N-terminal Ena/VASP Homology 1 (EVH1) domain and to mediate membrane translocation of its target dependent on its C-terminal Sprouty domain. However, the region of neurofibromin required for the interaction with Spred1 has remained unclear. Here we show that the EVH1 domain of Spred1 binds to the noncatalytic (GAPex) portion of the GAP-related domain (GRD) of neurofibromin. Binding is compatible with simultaneous binding of Ras and does not interfere with GAP activity. Our study points to a potential targeting function of the GAPex subdomain of neurofibromin that is present in all known canonical RasGAPs. PMID:27313208

  4. Oncogenic and RASopathy-associated K-RAS mutations relieve membrane-dependent occlusion of the effector-binding site

    PubMed Central

    Mazhab-Jafari, Mohammad T.; Marshall, Christopher B.; Smith, Matthew J.; Gasmi-Seabrook, Geneviève M. C.; Stathopulos, Peter B.; Inagaki, Fuyuhiko; Kay, Lewis E.; Neel, Benjamin G.; Ikura, Mitsuhiko

    2015-01-01

    K-RAS4B (Kirsten rat sarcoma viral oncogene homolog 4B) is a prenylated, membrane-associated GTPase protein that is a critical switch for the propagation of growth factor signaling pathways to diverse effector proteins, including rapidly accelerated fibrosarcoma (RAF) kinases and RAS-related protein guanine nucleotide dissociation stimulator (RALGDS) proteins. Gain-of-function KRAS mutations occur frequently in human cancers and predict poor clinical outcome, whereas germ-line mutations are associated with developmental syndromes. However, it is not known how these mutations affect K-RAS association with biological membranes or whether this impacts signal transduction. Here, we used solution NMR studies of K-RAS4B tethered to nanodiscs to investigate lipid bilayer-anchored K-RAS4B and its interactions with effector protein RAS-binding domains (RBDs). Unexpectedly, we found that the effector-binding region of activated K-RAS4B is occluded by interaction with the membrane in one of the NMR-observable, and thus highly populated, conformational states. Binding of the RAF isoform ARAF and RALGDS RBDs induced marked reorientation of K-RAS4B from the occluded state to RBD-specific effector-bound states. Importantly, we found that two Noonan syndrome-associated mutations, K5N and D153V, which do not affect the GTPase cycle, relieve the occluded orientation by directly altering the electrostatics of two membrane interaction surfaces. Similarly, the most frequent KRAS oncogenic mutation G12D also drives K-RAS4B toward an exposed configuration. Further, the D153V and G12D mutations increase the rate of association of ARAF-RBD with lipid bilayer-tethered K-RAS4B. We revealed a mechanism of K-RAS4B autoinhibition by membrane sequestration of its effector-binding site, which can be disrupted by disease-associated mutations. Stabilizing the autoinhibitory interactions between K-RAS4B and the membrane could be an attractive target for anticancer drug discovery. PMID:25941399

  5. Oncogenic and RASopathy-associated K-RAS mutations relieve membrane-dependent occlusion of the effector-binding site.

    PubMed

    Mazhab-Jafari, Mohammad T; Marshall, Christopher B; Smith, Matthew J; Gasmi-Seabrook, Geneviève M C; Stathopulos, Peter B; Inagaki, Fuyuhiko; Kay, Lewis E; Neel, Benjamin G; Ikura, Mitsuhiko

    2015-05-26

    K-RAS4B (Kirsten rat sarcoma viral oncogene homolog 4B) is a prenylated, membrane-associated GTPase protein that is a critical switch for the propagation of growth factor signaling pathways to diverse effector proteins, including rapidly accelerated fibrosarcoma (RAF) kinases and RAS-related protein guanine nucleotide dissociation stimulator (RALGDS) proteins. Gain-of-function KRAS mutations occur frequently in human cancers and predict poor clinical outcome, whereas germ-line mutations are associated with developmental syndromes. However, it is not known how these mutations affect K-RAS association with biological membranes or whether this impacts signal transduction. Here, we used solution NMR studies of K-RAS4B tethered to nanodiscs to investigate lipid bilayer-anchored K-RAS4B and its interactions with effector protein RAS-binding domains (RBDs). Unexpectedly, we found that the effector-binding region of activated K-RAS4B is occluded by interaction with the membrane in one of the NMR-observable, and thus highly populated, conformational states. Binding of the RAF isoform ARAF and RALGDS RBDs induced marked reorientation of K-RAS4B from the occluded state to RBD-specific effector-bound states. Importantly, we found that two Noonan syndrome-associated mutations, K5N and D153V, which do not affect the GTPase cycle, relieve the occluded orientation by directly altering the electrostatics of two membrane interaction surfaces. Similarly, the most frequent KRAS oncogenic mutation G12D also drives K-RAS4B toward an exposed configuration. Further, the D153V and G12D mutations increase the rate of association of ARAF-RBD with lipid bilayer-tethered K-RAS4B. We revealed a mechanism of K-RAS4B autoinhibition by membrane sequestration of its effector-binding site, which can be disrupted by disease-associated mutations. Stabilizing the autoinhibitory interactions between K-RAS4B and the membrane could be an attractive target for anticancer drug discovery.

  6. Glycation precedes lens crystallin aggregation

    SciTech Connect

    Swamy, M.S.; Perry, R.E.; Abraham, E.C.

    1987-05-01

    Non-enzymatic glycosylation (glycation) seems to have the potential to alter the structure of crystallins and make them susceptible to thiol oxidation leading to disulfide-linked high molecular weight (HMW) aggregate formation. They used streptozotocin diabetic rats during precataract and cataract stages and long-term cell-free glycation of bovine lens crystallins to study the relationship between glycation and lens crystallin aggregation. HMW aggregates and other protein components of the water-soluble (WS) and urea-soluble (US) fractions were separated by molecular sieve high performance liquid chromatography. Glycation was estimated by both (/sup 3/H)NaBH/sub 4/ reduction and phenylboronate agarose affinity chromatography. Levels of total glycated protein (GP) in the US fractions were about 2-fold higher than in the WS fractions and there was a linear increase in GP in both WS and US fractions. This increase was parallelled by a corresponding increase in HMW aggregates. Total GP extracted by the affinity method from the US fraction showed a predominance of HMW aggregates and vice versa. Cell-free glycation studies with bovine crystallins confirmed the results of the animals studies. Increasing glycation caused a corresponding increase in protein insolubilization and the insoluble fraction thus formed also contained more glycated protein. It appears that lens protein glycation, HMW aggregate formation, and protein insolubilization are interrelated.

  7. RAS Symposium Draws Hundreds of Attendees | Poster

    Cancer.gov

    They call themselves “rasologists”: scientists who study the RAS family of genes and the cancers that can arise due to mutations within them. This field of research is at the heart of some sobering numbers. Almost a third of all human cancers, including 95 percent of pancreatic cancers, are driven by mutated RAS genes. The American Cancer Society estimates there were 48,960 new cases of pancreatic cancer in the United States in 2015 and 40,560 deaths from the disease.

  8. RAS Projects at the NCI Frederick National Lab

    Cancer.gov

    The RAS Initiative involves a number of projects focusing on ways to inhibit the proliferation of cancer cells by mutant RAS proteins. Projects are conducted at the FNLCR hub, with collaboration from the research community nationwide.

  9. The RAS Problem: Turning Off a Broken Switch

    Cancer.gov

    The RAS gene is commonly mutated in cancer and researchers are working to better understand how to develop drugs that can target the RAS protein, which for many years has been considered to be “undruggable.”

  10. Calmodulin binds to K-Ras, but not to H- or N-Ras, and modulates its downstream signaling.

    PubMed

    Villalonga, P; López-Alcalá, C; Bosch, M; Chiloeches, A; Rocamora, N; Gil, J; Marais, R; Marshall, C J; Bachs, O; Agell, N

    2001-11-01

    Activation of Ras induces a variety of cellular responses depending on the specific effector activated and the intensity and amplitude of this activation. We have previously shown that calmodulin is an essential molecule in the down-regulation of the Ras/Raf/MEK/extracellularly regulated kinase (ERK) pathway in cultured fibroblasts and that this is due at least in part to an inhibitory effect of calmodulin on Ras activation. Here we show that inhibition of calmodulin synergizes with diverse stimuli (epidermal growth factor, platelet-derived growth factor, bombesin, or fetal bovine serum) to induce ERK activation. Moreover, even in the absence of any added stimuli, activation of Ras by calmodulin inhibition was observed. To identify the calmodulin-binding protein involved in this process, calmodulin affinity chromatography was performed. We show that Ras and Raf from cellular lysates were able to bind to calmodulin. Furthermore, Ras binding to calmodulin was favored in lysates with large amounts of GTP-bound Ras, and it was Raf independent. Interestingly, only one of the Ras isoforms, K-RasB, was able to bind to calmodulin. Furthermore, calmodulin inhibition preferentially activated K-Ras. Interaction between calmodulin and K-RasB is direct and is inhibited by the calmodulin kinase II calmodulin-binding domain. Thus, GTP-bound K-RasB is a calmodulin-binding protein, and we suggest that this binding may be a key element in the modulation of Ras signaling.

  11. ASPP2 Is a Novel Pan-Ras Nanocluster Scaffold

    PubMed Central

    Posada, Itziar M. D.; Serulla, Marc; Zhou, Yong; Oetken-Lindholm, Christina

    2016-01-01

    Ras-induced senescence mediated through ASPP2 represents a barrier to tumour formation. It is initiated by ASPP2’s interaction with Ras at the plasma membrane, which stimulates the Raf/MEK/ERK signaling cascade. Ras to Raf signalling requires Ras to be organized in nanoscale signalling complexes, called nanocluster. We therefore wanted to investigate whether ASPP2 affects Ras nanoclustering. Here we show that ASPP2 increases the nanoscale clustering of all oncogenic Ras isoforms, H-ras, K-ras and N-ras. Structure-function analysis with ASPP2 truncation mutants suggests that the nanocluster scaffolding activity of ASPP2 converges on its α-helical domain. While ASPP2 increased effector recruitment and stimulated ERK and AKT phosphorylation, it did not increase colony formation of RasG12V transformed NIH/3T3 cells. By contrast, ASPP2 was able to suppress the transformation enhancing ability of the nanocluster scaffold Gal-1, by competing with the specific effect of Gal-1 on H-rasG12V- and K-rasG12V-nanoclustering, thus imposing ASPP2’s ERK and AKT signalling signature. Similarly, ASPP2 robustly induced senescence and strongly abrogated mammosphere formation irrespective of whether it was expressed alone or together with Gal-1, which by itself showed the opposite effect in Ras wt or H-ras mutant breast cancer cells. Our results suggest that Gal-1 and ASPP2 functionally compete in nanocluster for active Ras on the plasma membrane. ASPP2 dominates the biological outcome, thus switching from a Gal-1 supported growth-promoting setting to a senescence inducing and stemness suppressive program in cancer cells. Our results support Ras nanocluster as major integrators of tumour fate decision events. PMID:27437940

  12. Prevalence of K-Ras mutations in hepatocellular carcinoma: A Turkish Oncology Group pilot study

    PubMed Central

    TURHAL, NAZIM SERDAR; SAVAŞ, BERNA; ÇOŞKUN, ÖZNUR; BAŞ, EMINE; KARABULUT, BÜLENT; NART, DENIZ; KORKMAZ, TANER; YAVUZER, DILEK; DEMIR, GÖKHAN; DOĞUSOY, GÜLEN; ARTAÇ, MEHMET

    2015-01-01

    Hepatocellular carcinoma (HCC) is the fifth most common male-predominant type of cancer worldwide. There is no effective treatment regimen available for advanced-stage disease and chemotherapy is generally ineffective in these patients. The number of studies on the prevalence of K-Ras mutations in HCC patients is currently limited. A total of 58 patients from 6 comprehensive cancer centers in 4 metropolitan cities of Turkey were enrolled in this study. Each center committed to enroll approximately 10 random patients whose formalin-fixed paraffin-embedded tumor tissues were available for K-Ras, exon 2 genotyping. Two methods were applied based on the availability of adequate amounts of tumor DNA. In the first method, the samples were processed using TheraScreen. The genomic DNA was further used to detect the 7 most frequent somatic mutations (35G>A; 35G>C; 35G>T; 34G>A; 34G>C; 34G>T and 38G>A) in codons 12 and 13 in exon 2 of the K-Ras oncogene by quantitative polymerase chain reaction (PCR). In the second method, the genomic DNA was amplified by PCR using primers specific for K-Ras exon 2 with the GML SeqFinder Sequencing System's KRAS kit. The identified DNA sequence alterations were confirmed by sequencing both DNA strands in two independent experiments with forward and reverse primers. A total of 40 samples had adequate tumor tissue for the mutation analysis. A total of 33 (82.5%) of the investigated samples harbored no mutations in exon 2. All the mutations were identified via a direct sequencing technique, whereas none were identified by TheraScreen. In conclusion, in our patients, HCC exhibited a remarkably low (<20%) K-Ras mutation rate. Patients harboring K-Ras wild-type tumors may be good candidates for treatment with epidermal growth factor inhibitors, such as cetuximab. PMID:26807232

  13. The Ras oncogene signals centrosome amplification in mammary epithelial cells through cyclin D1/Cdk4 and Nek2

    PubMed Central

    Zeng, X; Shaikh, FY; Harrison, MK; Adon, AM; Trimboli, AJ; Carroll, KA; Sharma, N; Timmers, C; Chodosh, LA; Leone, G; Saavedra, HI

    2010-01-01

    Centrosome amplification (CA) contributes to carcinogenesis by generating aneuploidy. Elevated frequencies of CA in most benign breast lesions and primary tumors suggest a causative role for CA in breast cancers. Clearly, identifying which and how altered signal transduction pathways contribute to CA is crucial to breast cancer control. Although a causative and cooperative role for c-Myc and Ras in mammary tumorigenesis is well documented, their ability to generate CA during mammary tumor initiation remains unexplored. To answer that question, K-RasG12D and c-Myc were induced in mouse mammary glands. Although CA was observed in mammary tumors initiated by c-Myc or K-RasG12D, it was detected only in premalignant mammary lesions expressing K-RasG12D. CA, both in vivo and in vitro, was associated with increased expression of the centrosome-regulatory proteins, cyclin D1 and Nek2. Abolishing the expression of cyclin D1, Cdk4 or Nek2 in MCF10A human mammary epithelial cells expressing H-RasG12V abrogated Ras-induced CA, whereas silencing cyclin E1 or B2 had no effect. Thus, we conclude that CA precedes mammary tumorigenesis, and interfering with centrosome-regulatory targets suppresses CA. PMID:20581865

  14. Crystal structure of M-Ras reveals a GTP-bound "off" state conformation of Ras family small GTPases.

    PubMed

    Ye, Min; Shima, Fumi; Muraoka, Shin; Liao, Jingling; Okamoto, Hidetsugu; Yamamoto, Masaki; Tamura, Atsuo; Yagi, Naoto; Ueki, Tatzuo; Kataoka, Tohru

    2005-09-01

    Although some members of Ras family small GTPases, including M-Ras, share the primary structure of their effector regions with Ras, they exhibit vastly different binding properties to Ras effectors such as c-Raf-1. We have solved the crystal structure of M-Ras in the GDP-bound and guanosine 5'-(beta,gamma-imido)triphosphate (Gpp(NH)p)-bound forms. The overall structure of M-Ras resembles those of H-Ras and Rap2A, except that M-Ras-Gpp(NH)p exhibits a distinctive switch I conformation, which is caused by impaired intramolecular interactions between Thr-45 (corresponding to Thr-35 of H-Ras) of the effector region and the gamma-phosphate of Gpp(NH)p. Previous 31P NMR studies showed that H-Ras-Gpp(NH)p exists in two interconverting conformations, states 1 and 2. Whereas state 2 is a predominant form of H-Ras and corresponds to the "on" conformation found in the complex with effectors, state 1 is thought to represent the "off" conformation, whose tertiary structure remains unknown. 31P NMR analysis shows that free M-Ras-Gpp(NH)p predominantly assumes the state 1 conformation, which undergoes conformational transition to state 2 upon association with c-Raf-1. These results indicate that the solved structure of M-Ras-Gp-p(NH)p corresponds to the state 1 conformation. The predominance of state 1 in M-Ras is likely to account for its weak binding ability to the Ras effectors, suggesting the importance of the tertiary structure factor in small GTPase-effector interaction. Further, the first determination of the state 1 structure provides a molecular basis for developing novel anti-cancer drugs as compounds that hold Ras in the state 1 "off" conformation. PMID:15994326

  15. The value of genomics in dissecting the RAS-network and in guiding therapeutics for RAS-driven cancers.

    PubMed

    Shrestha, Gajendra; MacNeil, Shelley M; McQuerry, Jasmine A; Jenkins, David F; Sharma, Sunil; Bild, Andrea H

    2016-10-01

    The rise in genomic knowledge over the past decade has revealed the molecular etiology of many diseases, and has identified intricate signaling network activity in human cancers. Genomics provides the opportunity to determine genome structure and capture the activity of thousands of molecular events concurrently, which is important for deciphering highly complex genetic diseases such as cancer. In this review, we focus on genomic efforts directed towards one of cancer's most frequently mutated networks, the RAS pathway. Genomic tools such as gene expression signatures and assessment of mutations across the RAS network enable the capture of RAS signaling complexity. Due to this high level of interaction and cross-talk within the network, efforts to target RAS signaling in the clinic have generally failed, and we currently lack the ability to directly inhibit the RAS protein with high efficacy. We propose that the use of gene expression data can identify effective treatments that broadly inhibit the RAS network as this approach measures pathway activity independent of mutation status or any single mechanism of activation. Here, we review the genomic studies that map the complexity of the RAS network in cancer, and that show how genomic measurements of RAS pathway activation can identify effective RAS inhibition strategies. We also address the challenges and future directions for treating RAS-driven tumors. In summary, genomic assessment of RAS signaling provides a level of complexity necessary to accurately map the network that matches the intricacy of RAS pathway interactions in cancer.

  16. The Differential Palmitoylation States of N-Ras and H-Ras Determine Their Distinct Golgi Sub-compartment Localizations

    PubMed Central

    Lynch, Stephen J.; Snitkin, Harriet; Gumper, Iwona; Philips, Mark R.; Sabatini, David; Pellicer, Angel

    2014-01-01

    Despite a high degree of structural homology and shared exchange factors, effectors and GTPase activating proteins, a large body of evidence suggests functional heterogeneity among Ras isoforms. One aspect of Ras biology that may explain this heterogeneity is the differential subcellular localizations driven by the C-terminal hypervariable regions of Ras proteins. Spatial heterogeneity has been documented at the level of organelles: palmitoylated Ras isoforms (H-Ras and N-Ras) localize on the Golgi apparatus whereas K-Ras4B does not. We tested the hypothesis that spatial heterogeneity also exists at the sub-organelle level by studying the localization of differentially palmitoylated Ras isoforms within the Golgi apparatus. Using confocal, live cell fluorescent imaging and immunogold electron microscopy we found that, whereas the doubly palmitoylated H-Ras is distributed throughout the Golgi stacks, the singly palmitoylated N-Ras is polarized with a relative paucity of expression on the trans Golgi. Using palmitoylation mutants we show that the different sub-Golgi distributions of the Ras proteins are a consequence of their differential degree of palmitoylation. Thus, the acylation state of Ras proteins controls not only their distribution between the Golgi apparatus and the plasma membrane but also their distribution within the Golgi stacks. PMID:25158650

  17. The value of genomics in dissecting the RAS-network and in guiding therapeutics for RAS-driven cancers.

    PubMed

    Shrestha, Gajendra; MacNeil, Shelley M; McQuerry, Jasmine A; Jenkins, David F; Sharma, Sunil; Bild, Andrea H

    2016-10-01

    The rise in genomic knowledge over the past decade has revealed the molecular etiology of many diseases, and has identified intricate signaling network activity in human cancers. Genomics provides the opportunity to determine genome structure and capture the activity of thousands of molecular events concurrently, which is important for deciphering highly complex genetic diseases such as cancer. In this review, we focus on genomic efforts directed towards one of cancer's most frequently mutated networks, the RAS pathway. Genomic tools such as gene expression signatures and assessment of mutations across the RAS network enable the capture of RAS signaling complexity. Due to this high level of interaction and cross-talk within the network, efforts to target RAS signaling in the clinic have generally failed, and we currently lack the ability to directly inhibit the RAS protein with high efficacy. We propose that the use of gene expression data can identify effective treatments that broadly inhibit the RAS network as this approach measures pathway activity independent of mutation status or any single mechanism of activation. Here, we review the genomic studies that map the complexity of the RAS network in cancer, and that show how genomic measurements of RAS pathway activation can identify effective RAS inhibition strategies. We also address the challenges and future directions for treating RAS-driven tumors. In summary, genomic assessment of RAS signaling provides a level of complexity necessary to accurately map the network that matches the intricacy of RAS pathway interactions in cancer. PMID:27338857

  18. Drugging Ras GTPase: a comprehensive mechanistic and signaling structural view.

    PubMed

    Lu, Shaoyong; Jang, Hyunbum; Gu, Shuo; Zhang, Jian; Nussinov, Ruth

    2016-09-21

    Ras proteins are small GTPases, cycling between inactive GDP-bound and active GTP-bound states. Through these switches they regulate signaling that controls cell growth and proliferation. Activating Ras mutations are associated with approximately 30% of human cancers, which are frequently resistant to standard therapies. Over the past few years, structural biology and in silico drug design, coupled with improved screening technology, led to a handful of promising inhibitors, raising the possibility of drugging Ras proteins. At the same time, the invariable emergence of drug resistance argues for the critical importance of additionally honing in on signaling pathways which are likely to be involved. Here we overview current advances in Ras structural knowledge, including the conformational dynamic of full-length Ras in solution and at the membrane, therapeutic inhibition of Ras activity by targeting its active site, allosteric sites, and Ras-effector protein-protein interfaces, Ras dimers, the K-Ras4B/calmodulin/PI3Kα trimer, and targeting Ras with siRNA. To mitigate drug resistance, we propose signaling pathways that can be co-targeted along with Ras and explain why. These include pathways leading to the expression (or activation) of YAP1 and c-Myc. We postulate that these and Ras signaling pathways, MAPK/ERK and PI3K/Akt/mTOR, act independently and in corresponding ways in cell cycle control. The structural data are instrumental in the discovery and development of Ras inhibitors for treating RAS-driven cancers. Together with the signaling blueprints through which drug resistance can evolve, this review provides a comprehensive and innovative master plan for tackling mutant Ras proteins.

  19. Drugging Ras GTPase: a comprehensive mechanistic and signaling structural view.

    PubMed

    Lu, Shaoyong; Jang, Hyunbum; Gu, Shuo; Zhang, Jian; Nussinov, Ruth

    2016-09-21

    Ras proteins are small GTPases, cycling between inactive GDP-bound and active GTP-bound states. Through these switches they regulate signaling that controls cell growth and proliferation. Activating Ras mutations are associated with approximately 30% of human cancers, which are frequently resistant to standard therapies. Over the past few years, structural biology and in silico drug design, coupled with improved screening technology, led to a handful of promising inhibitors, raising the possibility of drugging Ras proteins. At the same time, the invariable emergence of drug resistance argues for the critical importance of additionally honing in on signaling pathways which are likely to be involved. Here we overview current advances in Ras structural knowledge, including the conformational dynamic of full-length Ras in solution and at the membrane, therapeutic inhibition of Ras activity by targeting its active site, allosteric sites, and Ras-effector protein-protein interfaces, Ras dimers, the K-Ras4B/calmodulin/PI3Kα trimer, and targeting Ras with siRNA. To mitigate drug resistance, we propose signaling pathways that can be co-targeted along with Ras and explain why. These include pathways leading to the expression (or activation) of YAP1 and c-Myc. We postulate that these and Ras signaling pathways, MAPK/ERK and PI3K/Akt/mTOR, act independently and in corresponding ways in cell cycle control. The structural data are instrumental in the discovery and development of Ras inhibitors for treating RAS-driven cancers. Together with the signaling blueprints through which drug resistance can evolve, this review provides a comprehensive and innovative master plan for tackling mutant Ras proteins. PMID:27396271

  20. Rb and N-ras Function Together To Control Differentiation in the Mouse

    PubMed Central

    Takahashi, Chiaki; Bronson, Roderick T.; Socolovsky, Merav; Contreras, Bernardo; Lee, Kwang Youl; Jacks, Tyler; Noda, Makoto; Kucherlapati, Raju; Ewen, Mark E.

    2003-01-01

    The product of the retinoblastoma tumor suppressor gene (Rb) can control cell proliferation and promote dif-ferentiation. Murine embryos nullizygous for Rb die midgestation with defects in cell cycle regulation, control of apoptosis, and terminal differentiation of several tissues, including skeletal muscle, nervous system, and lens. Previous cell culture-based experiments have suggested that the retinoblastoma protein (pRb) and Ras operate in a common pathway to control cellular differentiation. Here we have tested the hypothesis that the proto-oncogene N-ras participates in Rb-dependent regulation of differentiation by generating and characterizing murine embryos deficient in both N-ras and Rb. We show that deletion of N-ras rescues a unique subset of the developmental defects associated with nullizygosity of Rb, resulting in a significant extension of life span. Rb−/−; N-ras−/− skeletal muscle has normal fiber density, myotube length and thickness, in contrast to Rb-deficient embryos. Additionally, Rb−/−; N-ras−/− muscle shows a restoration in the expression of the late muscle-specific gene MCK, and this correlates with a significant potentiation of MyoD transcriptional activity in Rb−/−; N-ras−/−, compared to Rb−/− myoblasts in culture. The improved differentiation of skeletal muscle in Rb−/−; N-ras−/− embryos occurs despite evidence of deregulated proliferation and apoptosis, as seen in Rb-deficient animals. Our findings suggest that the control of differentiation and proliferation by Rb are genetically separable. PMID:12861012

  1. Novel fabrication technique of hybrid structure lens array for 3D images

    NASA Astrophysics Data System (ADS)

    Lee, Junsik; Kim, Junoh; Kim, Cheoljoong; Shin, Dooseub; Koo, Gyohyun; Won, Yong Hyub

    2016-03-01

    Tunable liquid lens arrays can produce three dimensional images by using electrowetting principle that alters surface tensions by applying voltage. This method has advantages of fast response time and low power consumption. However, it is challenging to fabricate a high fill factor liquid lens array and operate three dimensional images which demand high diopter. This study describes a hybrid structure lens array which has not only a liquid lens array but a solid lens array. A concave-shape lens array is unavoidable when using only the liquid lens array and some voltages are needed to make the lens flat. By placing the solid lens array on the liquid lens array, initial diopter can be positive. To fabricate the hybrid structure lens array, a conventional lithographic process in semiconductor manufacturing is needed. A negative photoresist SU-8 was used as chamber master molds. PDMS and UV adhesive replica molding are done sequentially. Two immiscible liquids, DI water and dodecane, are injected in the fabricated chamber, followed by sealing. The fabricated structure has a 20 by 20 pattern of cylindrical shaped circle array and the aperture size of each lens is 1mm. The thickness of the overall hybrid structure is about 2.8mm. Hybrid structure lens array has many advantages. Solid lens array has almost 100% fill factor and allow high efficiency. Diopter can be increased by more than 200 and negative diopter can be shifted to the positive region. This experiment showed several properties of the hybrid structure and demonstrated its superiority.

  2. EGFR phosphorylates FAM129B to promote Ras activation

    PubMed Central

    Ji, Haitao; Lee, Jong-Ho; Wang, Yugang; Pang, Yilin; Zhang, Tao; Xia, Yan; Zhong, Lianjin; Lyu, Jianxin; Lu, Zhimin

    2016-01-01

    Ras GTPase-activating proteins (GAPs) are important regulators for Ras activation, which is instrumental in tumor development. However, the mechanism underlying this regulation remains elusive. We demonstrate here that activated EGFR phosphorylates the Y593 residue of the protein known as family with sequence similarity 129, member B (FAM129B), which is overexpressed in many types of human cancer. FAM129B phosphorylation increased the interaction between FAM129B and Ras, resulting in reduced binding of p120-RasGAP to Ras. FAM129B phosphorylation promoted Ras activation, increasing ERK1/2- and PKM2-dependent β-catenin transactivation and leading to the enhanced glycolytic gene expression and the Warburg effect; promoting tumor cell proliferation and invasion; and supporting brain tumorigenesis. Our studies unearthed a novel and important mechanism underlying EGFR-mediated Ras activation in tumor development. PMID:26721396

  3. Contact lens hygiene compliance and lens case contamination: A review.

    PubMed

    Wu, Yvonne Tzu-Ying; Willcox, Mark; Zhu, Hua; Stapleton, Fiona

    2015-10-01

    A contaminated contact lens case can act as a reservoir for microorganisms that could potentially compromise contact lens wear and lead to sight threatening adverse events. The rate, level and profile of microbial contamination in lens cases, compliance and other risk factors associated with lens case contamination, and the challenges currently faced in this field are discussed. The rate of lens case contamination is commonly over 50%. Coagulase-negative Staphylococcus, Bacillus spp., Pseudomonas aeruginosa and Serratia marcescens are frequently recovered from lens cases. In addition, we provide suggestions regarding how to clean contact lens cases and improve lens wearers' compliance as well as future lens case design for reducing lens case contamination. This review highlights the challenges in reducing the level of microbial contamination which require an industry wide approach.

  4. Unitary lens semiconductor device

    DOEpatents

    Lear, K.L.

    1997-05-27

    A unitary lens semiconductor device and method are disclosed. The unitary lens semiconductor device is provided with at least one semiconductor layer having a composition varying in the growth direction for unitarily forming one or more lenses in the semiconductor layer. Unitary lens semiconductor devices may be formed as light-processing devices such as microlenses, and as light-active devices such as light-emitting diodes, photodetectors, resonant-cavity light-emitting diodes, vertical-cavity surface-emitting lasers, and resonant cavity photodetectors. 9 figs.

  5. Unitary lens semiconductor device

    DOEpatents

    Lear, Kevin L.

    1997-01-01

    A unitary lens semiconductor device and method. The unitary lens semiconductor device is provided with at least one semiconductor layer having a composition varying in the growth direction for unitarily forming one or more lenses in the semiconductor layer. Unitary lens semiconductor devices may be formed as light-processing devices such as microlenses, and as light-active devices such as light-emitting diodes, photodetectors, resonant-cavity light-emitting diodes, vertical-cavity surface-emitting lasers, and resonant cavity photodetectors.

  6. Intraocular lens fabrication

    DOEpatents

    Salazar, Mike A.; Foreman, Larry R.

    1997-01-01

    This invention describes a method for fabricating an intraocular lens made rom clear Teflon.TM., Mylar.TM., or other thermoplastic material having a thickness of about 0.025 millimeters. These plastic materials are thermoformable and biocompatable with the human eye. The two shaped lenses are bonded together with a variety of procedures which may include thermosetting and solvent based adhesives, laser and impulse welding, and ultrasonic bonding. The fill tube, which is used to inject a refractive filling material is formed with the lens so as not to damage the lens shape. A hypodermic tube may be included inside the fill tube.

  7. Intraocular lens fabrication

    DOEpatents

    Salazar, M.A.; Foreman, L.R.

    1997-07-08

    This invention describes a method for fabricating an intraocular lens made from clear Teflon{trademark}, Mylar{trademark}, or other thermoplastic material having a thickness of about 0.025 millimeters. These plastic materials are thermoformable and biocompatable with the human eye. The two shaped lenses are bonded together with a variety of procedures which may include thermosetting and solvent based adhesives, laser and impulse welding, and ultrasonic bonding. The fill tube, which is used to inject a refractive filling material is formed with the lens so as not to damage the lens shape. A hypodermic tube may be included inside the fill tube. 13 figs.

  8. Ultraviolet spectrograph lens

    SciTech Connect

    Brixner, B.; Winkler, M.A.

    1981-01-01

    A 700-mm f/4.7 spectrograph camera lens was designed for imaging spectral lines in the 200 to 400-nm region on a 120-mm flat image field. Lens elements of fused silica and crystalline calcium fluoride have so little secondary spectrum that raytracing calculations predict a monochromatic resolution limit of 30 lines/mm without refocusing in the 238- to 365-nm region. Light scattering at the polished calcium-fluoride surfaces is avoided by sandwiching the fluoride elements between fused silica and cementing with silicone fluid. The constructed lens makes good spectrograms.

  9. Ultraviolet-spectrograph lens

    SciTech Connect

    Brixner, B.; Winkler, M.A.

    1981-01-01

    A 700-mm f/4.7 spectrograph camera lens was designed for imaging spectral lines in the 200- to 400-nm region on a 120-mm flat image field. Lens elements of fused silica and crystal calcium fluoride give such good achromatization that raytracing calculations predict a resolution limit of 30 lines/mm without refocusing in the 238- to 365-nm region. Light scattering at the polished calcium-fluoride surfaces is avoided by sandwiching the fluoride elements between fused silica and cementing with silicone fluid. The constructed lens makes good spectrograms.

  10. Telescopic vision contact lens

    NASA Astrophysics Data System (ADS)

    Tremblay, Eric J.; Beer, R. Dirk; Arianpour, Ashkan; Ford, Joseph E.

    2011-03-01

    We present the concept, optical design, and first proof of principle experimental results for a telescopic contact lens intended to become a visual aid for age-related macular degeneration (AMD), providing magnification to the user without surgery or external head-mounted optics. Our contact lens optical system can provide a combination of telescopic and non-magnified vision through two independent optical paths through the contact lens. The magnified optical path incorporates a telescopic arrangement of positive and negative annular concentric reflectors to achieve 2.8x - 3x magnification on the eye, while light passing through a central clear aperture provides unmagnified vision.

  11. Design and fabrication of soft zoom lens

    NASA Astrophysics Data System (ADS)

    Lin, Wei-Cheng; Chen, Chao-Chang A.; Huang, Kuo-Cheng

    2008-08-01

    With minimization of optical-electronics devices, conventional optical zoom lens component has been explored with liquid lens and soft polymer membranes. This paper introduces a novel zoom lens system with a soft polymer material, Polydimethylsiloxane (PDMS), its shape and curvature can be controlled by pneumatic pressure. Therefore, effective focal length (EFL) of soft zoom lens (SZL) system can be controlled and altered. According to desired opto-mechanical design, this ZL has been accessed to determine the optical specifications. After the pressure activated from 0 to 0.02MPa, the change of EFL of the SZL system can reach up to 18.77% (33.44mm to 39.717mm). Experimental results show that zoom effects of the developed SZL system are significantly affected by the shape, thickness and curing parameter of PDMS soft lens. The SZL system has been verified with the function of zoom ability. Further research works on the integration of the SZL system with imaging system for mobile devices or robot vision applications.

  12. Reflections From a Fresnel Lens

    ERIC Educational Resources Information Center

    Keeports, David

    2005-01-01

    Reflection of light by a convex Fresnel lens gives rise to two distinct images. A highly convex inverted real reflective image forms on the object side of the lens, while an upright virtual reflective image forms on the opposite side of the lens. I describe here a set of laser experiments performed upon a Fresnel lens. These experiments provide…

  13. The mechanical response of the porcine lens to a spinning test.

    PubMed

    Reilly, Matthew A; Martius, Philipp; Kumar, Saurav; Burd, Harvey J; Stachs, Oliver

    2016-06-01

    The pig lens has been used as a model for presbyopia as pigs lack accommodative ability. Previous studies using microindentation have indicated that the shear modulus distribution is qualitatively similar to that of the aged human lens and that the lens does not alter its refractive power due to equatorial stretching. A lens spinning test was used to determine whether prior lens stiffness data obtained from a sectioned porcine lens were reliable and whether the testing conditions significantly influence the lens' mechanical properties. The elastic modulus distribution determined for fresh lenses closely matched that measured previously using a microindentation test. Confocal scanning laser microscopy was used to evaluate changes to the lens' structure arising from mechanical stress and following storage for up to one week.

  14. Reaction of tyrosine oxidation products with proteins of the lens

    PubMed Central

    Pirie, Antoinette

    1968-01-01

    Oxidation of tyrosine in the presence of bovine lens proteins leads to the formation of brown or black melanoproteins. Both tyrosinase and the oxidizing system of ferrous sulphate–ascorbic acid–EDTA are effective. The fluorescence of the lens proteins is both altered and enhanced by the tyrosine-oxidizing systems. Their fluorescence spectra resemble those of urea-insoluble proteins of human cataractous lens and of 1,2-naphthaquinone–proteins of naphthalene cataract. The lens proteins lose their thiol groups and, in acid hydrolysates of treated β-and γ-crystallins, a substance has been detected chromatographically that behaves similarly to a compound formed when 3,4-dihydroxyphenylalanine (dopa) is oxidized by tyrosinase in the presence of cysteine. Analysis and behaviour of this substance from hydrolysates of lens proteins suggest that it is a compound of cysteine and dopa. PMID:4971287

  15. Contact Lens Risks

    MedlinePlus

    ... Health and Consumer Devices Consumer Products Contact Lenses Contact Lens Risks Share Tweet Linkedin Pin it More ... redness blurred vision swelling pain Serious Hazards of Contact Lenses Symptoms of eye irritation can indicate a ...

  16. IAA RAS Radio Telescope Monitoring System

    NASA Astrophysics Data System (ADS)

    Mikhailov, A.; Lavrov, A.

    2007-07-01

    Institute of Applied Astronomy of the Russian Academy of Sciences (IAA RAS) has three identical radio telescopes, the receiving complex of which consists of five two-channel receivers of different bands, six cryogen systems, and additional devices: four local oscillators, phase calibration generators and IF commutator. The design, hardware and data communication protocol are described. The most convenient way to join the devices of the receiving complex into the common monitoring system is to use the interface which allows to connect numerous devices to the data bus. For the purpose of data communication regulation and to exclude conflicts, a data communication protocol has been designed, which operates with complex formatted data sequences. Formation of such sequences requires considerable data processing capability. That is provided by a microcontroller chip in each slave device. The test version of the software for the central computer has been developed in IAA RAS. We are developing the Mark IV FS software extension modules, which will allow us to control the receiving complex of the radio telescope by special SNAP commands from both operator input and schedule files. We are also developing procedures of automatic measurements of SEFD, system noise temperature and other parameters, available both in VLBI and single-dish modes of operation. The system described has been installed on all IAA RAS radio telescopes at "Svetloe", "Zelenchukskaya" and "Badary" observatories. It has proved to be working quite reliably and to show the perfonmance expected.

  17. Lens auto-centering

    NASA Astrophysics Data System (ADS)

    Lamontagne, Frédéric; Desnoyers, Nichola; Doucet, Michel; Côté, Patrice; Gauvin, Jonny; Anctil, Geneviève; Tremblay, Mathieu

    2015-09-01

    In a typical optical system, optical elements usually need to be precisely positioned and aligned to perform the correct optical function. This positioning and alignment involves securing the optical element in a holder or mount. Proper centering of an optical element with respect to the holder is a delicate operation that generally requires tight manufacturing tolerances or active alignment, resulting in costly optical assemblies. To optimize optical performance and minimize manufacturing cost, there is a need for a lens mounting method that could relax manufacturing tolerance, reduce assembly time and provide high centering accuracy. This paper presents a patent pending lens mounting method developed at INO that can be compared to the drop-in technique for its simplicity while providing the level of accuracy close to that achievable with techniques using a centering machine (usually < 5 μm). This innovative auto-centering method is based on the use of geometrical relationship between the lens diameter, the lens radius of curvature and the thread angle of the retaining ring. The autocentering principle and centering test results performed on real optical assemblies are presented. In addition to the low assembly time, high centering accuracy, and environmental robustness, the INO auto-centering method has the advantage of relaxing lens and barrel bore diameter tolerances as well as lens wedge tolerances. The use of this novel lens mounting method significantly reduces manufacturing and assembly costs for high performance optical systems. Large volume productions would especially benefit from this advancement in precision lens mounting, potentially providing a drastic cost reduction.

  18. Mitogenic signaling mediated by oxidants in Ras-transformed fibroblasts.

    PubMed

    Irani, K; Xia, Y; Zweier, J L; Sollott, S J; Der, C J; Fearon, E R; Sundaresan, M; Finkel, T; Goldschmidt-Clermont, P J

    1997-03-14

    NIH 3T3 fibroblasts stably transformed with a constitutively active isoform of p21(Ras), H-RasV12 (v-H-Ras or EJ-Ras), produced large amounts of the reactive oxygen species superoxide (.O2-). .O2- production was suppressed by the expression of dominant negative isoforms of Ras or Rac1, as well as by treatment with a farnesyltransferase inhibitor or with diphenylene iodonium, a flavoprotein inhibitor. The mitogenic activity of cells expressing H-RasV12 was inhibited by treatment with the chemical antioxidant N-acetyl-L-cysteine. Mitogen-activated protein kinase (MAPK) activity was decreased and c-Jun N-terminal kinase (JNK) was not activated in H-RasV12-transformed cells. Thus, H-RasV12-induced transformation can lead to the production of .O2- through one or more pathways involving a flavoprotein and Rac1. The implication of a reactive oxygen species, probably .O2-, as a mediator of Ras-induced cell cycle progression independent of MAPK and JNK suggests a possible mechanism for the effects of antioxidants against Ras-induced cellular transformation.

  19. Terahertz Artificial Dielectric Lens

    NASA Astrophysics Data System (ADS)

    Mendis, Rajind; Nagai, Masaya; Wang, Yiqiu; Karl, Nicholas; Mittleman, Daniel M.

    2016-03-01

    We have designed, fabricated, and experimentally characterized a lens for the THz regime based on artificial dielectrics. These are man-made media that mimic properties of naturally occurring dielectric media, or even manifest properties that cannot generally occur in nature. For example, the well-known dielectric property, the refractive index, which usually has a value greater than unity, can have a value less than unity in an artificial dielectric. For our lens, the artificial-dielectric medium is made up of a parallel stack of 100 μm thick metal plates that form an array of parallel-plate waveguides. The convergent lens has a plano-concave geometry, in contrast to conventional dielectric lenses. Our results demonstrate that this lens is capable of focusing a 2 cm diameter beam to a spot size of 4 mm, at the design frequency of 0.17 THz. The results further demonstrate that the overall power transmission of the lens can be better than certain conventional dielectric lenses commonly used in the THz regime. Intriguingly, we also observe that under certain conditions, the lens boundary demarcated by the discontinuous plate edges actually resembles a smooth continuous surface. These results highlight the importance of this artificial-dielectric technology for the development of future THz-wave devices.

  20. Terahertz Artificial Dielectric Lens.

    PubMed

    Mendis, Rajind; Nagai, Masaya; Wang, Yiqiu; Karl, Nicholas; Mittleman, Daniel M

    2016-01-01

    We have designed, fabricated, and experimentally characterized a lens for the THz regime based on artificial dielectrics. These are man-made media that mimic properties of naturally occurring dielectric media, or even manifest properties that cannot generally occur in nature. For example, the well-known dielectric property, the refractive index, which usually has a value greater than unity, can have a value less than unity in an artificial dielectric. For our lens, the artificial-dielectric medium is made up of a parallel stack of 100 μm thick metal plates that form an array of parallel-plate waveguides. The convergent lens has a plano-concave geometry, in contrast to conventional dielectric lenses. Our results demonstrate that this lens is capable of focusing a 2 cm diameter beam to a spot size of 4 mm, at the design frequency of 0.17 THz. The results further demonstrate that the overall power transmission of the lens can be better than certain conventional dielectric lenses commonly used in the THz regime. Intriguingly, we also observe that under certain conditions, the lens boundary demarcated by the discontinuous plate edges actually resembles a smooth continuous surface. These results highlight the importance of this artificial-dielectric technology for the development of future THz-wave devices. PMID:26973294

  1. Terahertz Artificial Dielectric Lens.

    PubMed

    Mendis, Rajind; Nagai, Masaya; Wang, Yiqiu; Karl, Nicholas; Mittleman, Daniel M

    2016-03-14

    We have designed, fabricated, and experimentally characterized a lens for the THz regime based on artificial dielectrics. These are man-made media that mimic properties of naturally occurring dielectric media, or even manifest properties that cannot generally occur in nature. For example, the well-known dielectric property, the refractive index, which usually has a value greater than unity, can have a value less than unity in an artificial dielectric. For our lens, the artificial-dielectric medium is made up of a parallel stack of 100 μm thick metal plates that form an array of parallel-plate waveguides. The convergent lens has a plano-concave geometry, in contrast to conventional dielectric lenses. Our results demonstrate that this lens is capable of focusing a 2 cm diameter beam to a spot size of 4 mm, at the design frequency of 0.17 THz. The results further demonstrate that the overall power transmission of the lens can be better than certain conventional dielectric lenses commonly used in the THz regime. Intriguingly, we also observe that under certain conditions, the lens boundary demarcated by the discontinuous plate edges actually resembles a smooth continuous surface. These results highlight the importance of this artificial-dielectric technology for the development of future THz-wave devices.

  2. Terahertz Artificial Dielectric Lens

    PubMed Central

    Mendis, Rajind; Nagai, Masaya; Wang, Yiqiu; Karl, Nicholas; Mittleman, Daniel M.

    2016-01-01

    We have designed, fabricated, and experimentally characterized a lens for the THz regime based on artificial dielectrics. These are man-made media that mimic properties of naturally occurring dielectric media, or even manifest properties that cannot generally occur in nature. For example, the well-known dielectric property, the refractive index, which usually has a value greater than unity, can have a value less than unity in an artificial dielectric. For our lens, the artificial-dielectric medium is made up of a parallel stack of 100 μm thick metal plates that form an array of parallel-plate waveguides. The convergent lens has a plano-concave geometry, in contrast to conventional dielectric lenses. Our results demonstrate that this lens is capable of focusing a 2 cm diameter beam to a spot size of 4 mm, at the design frequency of 0.17 THz. The results further demonstrate that the overall power transmission of the lens can be better than certain conventional dielectric lenses commonly used in the THz regime. Intriguingly, we also observe that under certain conditions, the lens boundary demarcated by the discontinuous plate edges actually resembles a smooth continuous surface. These results highlight the importance of this artificial-dielectric technology for the development of future THz-wave devices. PMID:26973294

  3. Myc and Ras oncogenes engage different energy metabolism programs and evoke distinct patterns of oxidative and DNA replication stress.

    PubMed

    Maya-Mendoza, Apolinar; Ostrakova, Jitka; Kosar, Martin; Hall, Arnaldur; Duskova, Pavlina; Mistrik, Martin; Merchut-Maya, Joanna Maria; Hodny, Zdenek; Bartkova, Jirina; Christensen, Claus; Bartek, Jiri

    2015-03-01

    Both Myc and Ras oncogenes impact cellular metabolism, deregulate redox homeostasis and trigger DNA replication stress (RS) that compromises genomic integrity. However, how are such oncogene-induced effects evoked and temporally related, to what extent are these kinetic parameters shared by Myc and Ras, and how are these cellular changes linked with oncogene-induced cellular senescence in different cell context(s) remain poorly understood. Here, we addressed the above-mentioned open questions by multifaceted comparative analyses of human cellular models with inducible expression of c-Myc and H-RasV12 (Ras), two commonly deregulated oncoproteins operating in a functionally connected signaling network. Our study of DNA replication parameters using the DNA fiber approach and time-course assessment of perturbations in glycolytic flux, oxygen consumption and production of reactive oxygen species (ROS) revealed the following results. First, overabundance of nuclear Myc triggered RS promptly, already after one day of Myc induction, causing slow replication fork progression and fork asymmetry, even before any metabolic changes occurred. In contrast, Ras overexpression initially induced a burst of cell proliferation and increased the speed of replication fork progression. However, after several days of induction Ras caused bioenergetic metabolic changes that correlated with slower DNA replication fork progression and the ensuing cell cycle arrest, gradually leading to senescence. Second, the observed oncogene-induced RS and metabolic alterations were cell-type/context dependent, as shown by comparative analyses of normal human BJ fibroblasts versus U2-OS sarcoma cells. Third, the energy metabolic reprogramming triggered by Ras was more robust compared to impact of Myc. Fourth, the detected oncogene-induced oxidative stress was due to ROS (superoxide) of non-mitochondrial origin and mitochondrial OXPHOS was reduced (Crabtree effect). Overall, our study provides novel

  4. Variable-focal lens using electroactive polymer actuator

    NASA Astrophysics Data System (ADS)

    Vunder, V.; Punning, A.; Aabloo, A.

    2011-03-01

    The paper describes a simple and cost-effective design and fabrication process of a liquid-filled variable-focal lens. The lens was made of soft polymer material, its shape and curvature can be controlled by hydraulic pressure. An electroactive polymer is used as an actuator. A carbon-polymer composite (CPC) was used. The device is composed of elastic membrane upon a circular lens chamber, a reservoir of liquid, and a channel between them. It was made of three layers of polydimethylsiloxane (PDMS), bonded using the partial curing technique. The channels and reservoir were filled with incompressible liquid after curing process. A CPC actuator was mechanically attached to reservoir to compress or decompress the liquid. Squeezing the liquid between the reservoir and the lens chamber will push the membrane inward or outward resulting in the change of the shape of the lens and alteration of its focal length. Depending on the pressure the lens can be plano-convex or plano-concave or even switch between the two configurations. With only a few minor modifications it is possible to fabricate bi-convex and bi-concave lenses. The lens with a 1 mm diameter and the focal length from infinity to 17 mm is reported. The 5x15mm CPC actuator with the working voltage of only up to +/-2.5 V was capable to alter the focal length within the full range of the focal length in 10 seconds.

  5. TLN-4601, a novel anticancer agent, inhibits Ras signaling post Ras prenylation and before MEK activation.

    PubMed

    Boufaied, Nadia; Wioland, My-Anh; Falardeau, Pierre; Gourdeau, Henriette

    2010-06-01

    TLN-4601 is a structurally novel farnesylated dibenzodiazepinone discovered through DECIPHER, Thallion's proprietary drug discovery platform. The compound was shown to have a broad cytotoxic activity (low micromol/l) when tested in the NCI 60 tumor cell line panel and has shown in-vivo antitumor activity in several xenograft models. Related to its farnesylated moiety, the effect of TLN-4601 on Ras mitogen-activated protein kinase signaling was assessed. Downstream Ras signaling events, Raf-1, MEK, and ERK1/2 phosphorylation in MCF7 cells were evaluated by western blot analysis. TLN-4601 prevented epidermal growth factor-induced phosphorylation of Raf-1, MEK, and ERK1/2. This effect was time-dependent and dose-dependent with complete inhibition of protein phosphorylation within 4-6 h at 10 micromol/l. The inhibition of Ras signaling was not mediated by the inhibition of protein prenylation, documented by the lack of effect TLN-4601 on the prenylation of HDJ2 (specific substrate of farnesyltransferase), RAP1A (specific substrate of geranylgeranyl transferase-1), or Ras. As TLN-4601 did not inhibit EGFR, Raf-1, MEK or ERK1/2 kinase activities, the inhibitory effect of TLN-4601 on Ras signaling is not mediated by direct kinase inhibition. Using an Elk-1 trans-activation reporter assay, we found that TLN-4601 inhibits the MEK/ERK pathway at the level of Raf-1. Interestingly, TLN-4601 induces Raf-1 proteasomal-dependent degradation. These data indicate that TLN-4601 may inhibit the Ras-mitogen-activated protein kinase-signaling pathway by depleting the Raf-1 protein.

  6. Overexpressed galectin-3 in pancreatic cancer induces cell proliferation and invasion by binding Ras and activating Ras signaling.

    PubMed

    Song, Shumei; Ji, Baoan; Ramachandran, Vijaya; Wang, Huamin; Hafley, Margarete; Logsdon, Craig; Bresalier, Robert S

    2012-01-01

    Pancreatic cancer (PDAC) is a lethal disease with a five-year survival of 3-5%. Mutations in K-Ras are found in nearly all cases, but K-Ras mutations alone are not sufficient for the development of PDAC. Additional factors contribute to activation of Ras signaling and lead to tumor formation. Galectin-3 (Gal-3), a multifunctional β-galactoside-binding protein, is highly expressed in PDAC. We therefore investigated the functional role of Gal-3 in pancreatic cancer progression and its relationship to Ras signaling. Expression of Gal-3 was determined by immunohistochemistry, Q-PCR and immunoblot. Functional studies were performed using pancreatic cell lines genetically engineered to express high or low levels of Gal-3. Ras activity was examined by Raf pull-down assays. Co-immunoprecipitation and immunofluorescence were used to assess protein-protein interactions. In this study, we demonstrate that Gal-3 was highly up-regulated in human tumors and in a mutant K-Ras mouse model of PDAC. Down-regulation of Gal-3 by lentivirus shRNA decreased PDAC cell proliferation and invasion in vitro and reduced tumor volume and size in an orthotopic mouse model. Gal-3 bound Ras and maintained Ras activity; down-regulation of Gal-3 decreased Ras activity as well as Ras down-stream signaling including phosphorylation of ERK and AKT and Ral A activity. Transfection of Gal-3 cDNA into PDAC cells with low-level Gal-3 augmented Ras activity and its down-stream signaling. These results suggest that Gal-3 contributes to pancreatic cancer progression, in part, by binding Ras and activating Ras signaling. Gal-3 may therefore be a potential novel target for this deadly disease. PMID:22900040

  7. Fermat's least-time principle and the embedded transparent lens

    NASA Astrophysics Data System (ADS)

    Kantowski, R.; Chen, B.; Dai, X.

    2013-10-01

    We present a simplified version of the lowest-order embedded point mass gravitational lens theory and then make the extension of this theory to any embedded transparent lens. Embedding a lens effectively reduces the gravitational potential’s range, i.e., partially shields the lensing potential because the lens mass is made a contributor to the mean mass density of the Universe and not simply superimposed upon it. We give the time-delay function for the embedded point mass lens from which we can derive the simplified lens equation by applying Fermat’s least-time principle. Even though rigorous derivations are only made for the point mass in a flat background, the generalization of the lens equation to lowest order for any distributed lens in any homogeneous background is obvious. We find from this simplified theory that embedding can introduce corrections above the few percent level in weak lensing shears caused by large clusters but only at large impacts. The potential part of the time delay is also affected in strong lensing at the few percent level. Additionally we again confirm that the presence of a cosmological constant alters the gravitational deflection of passing photons.

  8. Dimerize RACK1 upon transformation with oncogenic ras

    SciTech Connect

    Chu, L.-Y.; Chen, Y.-H.; Chuang, N.-N. . E-mail: zonnc@sinica.edu.tw

    2005-05-06

    From our previous studies, we learned that syndecan-2/p120-GAP complex provided docking site for Src to prosecute tyrosine kinase activity upon transformation with oncogenic ras. And, RACK1 protein was reactive with syndecan-2 to keep Src inactivated, but not when Ras was overexpressed. In the present study, we characterized the reaction between RACK1 protein and Ras. RACK1 was isolated from BALB/3T3 cells transfected with plasmids pcDNA3.1-[S-ras(Q{sub 61}K)] of shrimp Penaeus japonicus and RACK1 was revealed to react with GTP-K{sub B}-Ras(Q{sub 61}K), not GDP-K{sub B}-Ras(Q{sub 61}K). This selective interaction between RACK1 and GTP-K{sub B}-Ras(Q{sub 61}K) was further confirmed with RACK1 of human placenta and mouse RACK1-encoded fusion protein. We found that RACK1 was dimerized upon reaction with GTP-K{sub B}-Ras(Q{sub 61}K), as well as with 14-3-3{beta} and geranylgeranyl pyrophosphate, as revealed by phosphorylation with Src tyrosine kinase. We reported the complex of RACK1/GTP-K{sub B}-Ras(Q{sub 61}K) reacted selectively with p120-GAP. This interaction was sufficient to dissemble RACK1 into monomers, a preferred form to compete for the binding of syndecan-2. These data indicate that the reaction of GTP-K{sub B}-Ras(Q{sub 61}K) with RACK1 in dimers may operate a mechanism to deplete RACK1 from reaction with syndecan-2 upon transformation by oncogenic ras and the RACK1/GTP-Ras complex may provide a route to react with p120-GAP and recycle monomeric RACK1 to syndecan-2.

  9. TC21 and Ras share indistinguishable transforming and differentiating activities.

    PubMed

    Graham, S M; Oldham, S M; Martin, C B; Drugan, J K; Zohn, I E; Campbell, S; Der, C J

    1999-03-25

    Constitutively activated mutants of the Ras-related protein TC21/R-Ras2 cause tumorigenic transformation of NIH3T3 cells. However, unlike Ras, TC21 fails to bind to and activate the Raf-1 serine-threonine kinase. Thus, whereas Ras transformation is critically dependent on Raf-1 TC21 activity is promoted by activation of Raf-independent signaling pathways. In the present study, we have further compared the functions of Ras and TC21. First we determined the basis for the inability of TC21 to activate Raf-1. Whereas Ras can interact with the two distinct Ras-binding sequences in NH2-terminus of Raf-1, designated RBS1 and Raf-Cys, TC21 could only bind Raf-Cys. Thus, the inability of TC21 to bind to RBS1 may prevent it from promoting the translocation of Raf-1 to the plasma membrane. Second, we found that TC21 is an activator of the JNK and p38, but not ERK, mitogen-activated protein kinase cascades and that TC21 transforming activity was dependent on Rac function. Thus, like Ras, TC21 may activate a Rac/JNK pathway. Third, we determined if TC21 could cause the same biological consequences as Ras in three distinct cell types. Like Ras, activated TC21 caused transformation of RIE-1 rat intestinal epithelial cells and terminal differentiation of PC12 pheochromocytoma cells. Finally, activated TC21 blocked serum starvation-induced differentiation of C2 myoblasts, whereas dominant negative TC21 greatly accelerated this differentiation process. Therefore, TC21 and Ras share indistinguishable biological activities in all cell types that we have evaluated. These results support the importance of Raf-independent pathways in mediating the actions of Ras and TC21.

  10. Neoplastic transformation of a human prostate epithelial cell line by the v-Ki-ras oncogene.

    PubMed

    Parda, D S; Thraves, P J; Kuettel, M R; Lee, M S; Arnstein, P; Kaighn, M E; Rhim, J S; Dritschilo, A

    1993-01-01

    Investigations of mechanisms of human prostate carcinogenesis are limited by the unavailability of a suitable in vitro model system. We have demonstrated that an immortal, but nontumorigenic, human epithelial cell line (267B1) established from fetal prostate tissue can be malignantly transformed by a biological carcinogen, and can serve as a useful model for investigations of the progression steps of carcinogenesis. Activated Ki-ras was introduced into 267B1 cells by infection with the Kirsten murine sarcoma virus. Morphological alterations and anchorage-independent growth were observed; when cells were injected into nude mice, poorly differentiated adenocarcinomas developed. These findings represent the first evidence of malignant transformation of human prostate epithelial cells in culture, and support a role for Ki-ras activation in a multistep process for prostate neoplastic transformation.

  11. Complex effects of Ras proto-oncogenes in tumorigenesis.

    PubMed

    Diaz, Roberto; Lopez-Barcons, Lluis; Ahn, Daniel; Garcia-Espana, Antonio; Yoon, Andrew; Matthews, Jeremy; Mangues, Ramon; Perez-Soler, Roman; Pellicer, Angel

    2004-04-01

    Ras proteins have been found mutated in about one-third of human tumors. In vitro, Ras has been shown to regulate distinct and contradictory effects, such as cellular proliferation and apoptosis. Nonetheless, the effects that the wild-type protein elicits in tumorigenesis are poorly understood. Depending on the type of tissue, Ras proto-oncogenes appear to either promote or inhibit the tumor phenotype. In this report, we treated wild-type and N-ras knockout mice with 3-methylcholanthrene (MCA) to induce fibrosarcomas and found that MCA is more carcinogenic in wild-type mice than in knockout mice. After injecting different doses of a tumorigenic cell line, the wild-type mice exhibited a shorter latency of tumor development than the knockouts, indicating that there are N-ras-dependent differences in the stromal cells. Likewise, we have analyzed B-cell lymphomas induced by either N-methylnitrosourea or by the N-ras oncogene in mice that over-express the N-ras proto-oncogene and found that the over-expression of wild-type N-ras is able to increase the incidence of these lymphomas. Considered together, our results indicate that Ras proto-oncogenes can enhance or inhibit the malignant phenotype in vivo in different systems.

  12. Activation of ras oncogenes preceding the onset of neoplasia

    SciTech Connect

    Kumar, R.; Barbacid, M. ); Sukumar, S. )

    1990-06-01

    The identification of ras oncogenes in human and animal cancers including precancerous lesions indicates that these genes participate in the early stages of neoplastic development. Yet, these observations do not define the timing of ras oncogene activation in the multistep process of carcinogenesis. To ascertain the timing of ras oncogene activation, an animal model system was devised that involves the induction of mammary carcinomas in rats exposed at birth to the carcinogen nitrosomethylurea. High-resolution restriction fragment length polymorphism analysis of polymerase chain reaction-amplified ras sequences revealed the presence of both H-ras and K-ras oncogenes in normal mammary glands 2 weeks after carcinogen treatment and at least 2 months before the onset of neoplasia. These ras oncogenes can remain latent within the mammary gland until exposure to estrogens, demonstrating that activation of ras oncogenes can precede the onset of neoplasia and suggesting that normal physiological proliferative processes such as estrogen-induced mammary gland development may lead to neoplasia if the targeted cells harbor latent ras oncogenes.

  13. Activation of a human c-K-ras oncogene.

    PubMed Central

    Yamamoto, F; Perucho, M

    1984-01-01

    The human lung carcinomas PR310 and PR371 contain activated c-K-ras oncogenes. The oncogene of PR371 was found to present a mutation at codon 12 of the first coding exon which substitutes cysteine for glycine in the encoded p21 protein. We report here that the transforming gene of PR310 tumor contains a mutation in the second coding exon. An A----T transversion at codon 61 results in the incorporation of histidine instead of glutamine in the c-K-ras gene product. By constructing c-K-ras/c-H-ras chimeric genes we show that this point mutation is sufficient to confer transforming potential to ras genes, and that a hybrid ras gene coding for a protein mutant at both codons 12 and 61 is also capable of transforming NIH3T3 cells. The relative transforming potency of p21 proteins encoded by ras genes mutant at codons 12, 61 or both has been analyzed. Our studies also show that the coding exons of ras genes, including the fourth, can be interchanged and the chimeric p21 ras proteins retain their oncogenic ability in normal rodent established cell lines. PMID:6096811

  14. Deletion of Pim Kinases Elevates the Cellular Levels of Reactive Oxygen Species and Sensitizes to K-Ras-Induced Cell Killing

    PubMed Central

    Song, Jin H.; An, Ningfei; Chatterjee, Shilpak; Kistner-Griffin, Emily; Mahajan, Sandeep; Mehrotra, Shikhar; Kraft, Andrew S.

    2014-01-01

    The Pim protein kinases contribute to transformation by enhancing the activity of oncogenic Myc and Ras, which drives significant metabolic changes during tumorigenesis. In this report, we demonstrate that mouse embryo fibroblasts (MEFs) lacking all three isoforms of Pim protein kinases, triple knockout (TKO), cannot tolerate the expression of activated K-Ras (K-RasG12V) and undergo cell death. Transduction of K-RasG12V into these cells markedly increased the level of cellular reactive oxygen species (ROS). The addition of N-acetyl cysteine attenuates ROS production and reversed the cytotoxic effects of K-RasG12V in the TKO MEFs. The altered cellular redox state caused by the loss of Pim occurred as a result of lower levels of metabolic intermediates in the glycolytic and pentose phosphate pathways as well as abnormal mitochondrial oxidative phosphorylation. TKO MEFs exhibit reduced levels of superoxide dismutase (Sod), glutathione peroxidase 4 (Gpx4) and peroxiredoxin 3 (Prdx3) that render them susceptible to killing by K-RasG12V-mediated ROS production. In contrast, the transduction of c-Myc into TKO cells can overcome the lack of Pim protein kinases by regulating cellular metabolism and Sod2. In the absence of the Pim kinases, c-Myc transduction permitted K-RasG12V-induced cell growth by decreasing Ras-induced cellular ROS levels. These results demonstrate that the Pim protein kinases play an important role in regulating cellular redox, metabolism and K-Ras-stimulated cell growth. PMID:25241892

  15. Effects of mutant human Ki-ras{sup G12C} gene dosage on murine lung tumorigenesis and signaling to its downstream effectors

    SciTech Connect

    Dance-Barnes, Stephanie T.; Kock, Nancy D.; Floyd, Heather S.; Moore, Joseph E.; Mosley, Libyadda J.; D'Agostino, Ralph B.; Pettenati, Mark J.; Miller, Mark Steven

    2008-08-15

    Studies in cell culture have suggested that the level of RAS expression can influence the transformation of cells and the signaling pathways stimulated by mutant RAS expression. However, the levels of RAS expression in vivo appear to be subject to feedback regulation, limiting the total amount of RAS protein that can be expressed. We utilized a bitransgenic mouse lung tumor model that expressed the human Ki-ras{sup G12C} allele in a tetracycline-inducible, lung-specific manner. Treatment for 12 months with 500 {mu}g/ml of doxycycline (DOX) allowed for maximal expression of the human Ki-ras{sup G12C} allele in the lung, and resulted in the development of focal hyperplasia and adenomas. We determined if different levels of mutant RAS expression would influence the phenotype of the lung lesions. Treatment with 25, 100 and 500 {mu}g/ml of DOX resulted in dose-dependent increases in transgene expression and tumor multiplicity. Microscopic analysis of the lungs of mice treated with the 25 {mu}g/ml dose of DOX revealed infrequent foci of hyperplasia, whereas mice treated with the 100 and 500 {mu}g/ml doses exhibited numerous hyperplastic foci and also adenomas. Immunohistochemical and RNA analysis of the downstream effector pathways demonstrated that different levels of mutant RAS transgene expression resulted in differences in the expression and/or phosphorylation of specific signaling molecules. Our results suggest that the molecular alterations driving tumorigenesis may differ at different levels of mutant Ki-ras{sup G12C} expression, and this should be taken into consideration when inducible transgene systems are utilized to promote tumorigenesis in mouse models.

  16. Differential Regulation of N-Myc and c-Myc Synthesis, Degradation, and Transcriptional Activity by the Ras/Mitogen-activated Protein Kinase Pathway*

    PubMed Central

    Kapeli, Katannya; Hurlin, Peter J.

    2011-01-01

    Myc transcription factors are important regulators of proliferation and can promote oncogenesis when deregulated. Deregulated Myc expression in cancers can result from MYC gene amplification and translocation but also from alterations in mitogenic signaling pathways that affect Myc levels through both transcriptional and post-transcription mechanisms. For example, mutations in Ras family GTPase proteins that cause their constitutive activation can increase cellular levels of c-Myc by interfering with its rapid proteasomal degradation. Although enhanced protein stability is generally thought to be applicable to other Myc family members, here we show that c-Myc and its paralog N-Myc respond to oncogenic H-Ras (H-RasG12V) in very different ways. H-RasG12V promotes accumulation of both c-Myc and N-Myc, but although c-Myc accumulation is achieved by enhanced protein stability, N-Myc accumulation is associated with an accelerated rate of translation that overcomes a surprising H-RasG12V-mediated destabilization of N-Myc. We show that H-RasG12V-mediated degradation of N-Myc functions independently of key phosphorylation sites in the highly conserved Myc homology box I region that controls c-Myc protein stability by oncogenic Ras. Finally, we found that N-Myc and c-Myc transcriptional activity is associated with their proteasomal degradation but that N-Myc may be uniquely dependent on Ras-stimulated proteolysis for target gene expression. Taken together, these studies provide mechanistic insight into how oncogenic Ras augments N-Myc levels in cells and suggest that enhanced N-Myc translation and degradation-coupled transactivation may contribute to oncogenesis. PMID:21908617

  17. Rap1 GTPase is required for mouse lens epithelial maintenance and morphogenesis

    PubMed Central

    Maddala, Rupalatha; Nagendran, Tharkika; Lang, Richard A.; Morozov, Alexei; Rao, Ponugoti V.

    2015-01-01

    Rap1, a Ras-like small GTPase, plays a crucial role in cell-matrix adhesive interactions, cell-cell junction formation, cell polarity and migration. The role of Rap1 in vertebrate organ development and tissue architecture, however, remains elusive. We addressed this question in a mouse lens model system using a conditional gene targeting approach. While individual germline deficiency of either Rap1a or Rap1b did not cause overt defects in mouse lens, conditional double deficiency (Rap1 cKO) prior to lens placode formation led to an ocular phenotype including microphthalmia and lens opacification in embryonic mice. The embryonic Rap1 cKO mouse lens exhibited striking defects including loss of E-cadherin- and ZO-1-based cell-cell junctions, disruption of paxillin and β1-integrin-based cell adhesive interactions along with abnormalities in cell shape and apical-basal polarity of epithelium. These epithelial changes were accompanied by increased levels of α-smooth muscle actin, vimentin and N-cadherin, and expression of transcriptional suppressors of E-cadherin (Snai1, Slug and Zeb2), and a mesenchymal metabolic protein (Dihydropyrimidine dehydrogenase). Additionally, while lens differentiation was not overtly affected, increased apoptosis and dysregulated cell cycle progression were noted in epithelium and fibers in Rap1 cKO mice. Collectively these observations uncover a requirement for Rap1 in maintenance of lens epithelial phenotype and morphogenesis. PMID:26212757

  18. FGFR and PTEN signaling interact during lens development to regulate cell survival.

    PubMed

    Chaffee, Blake R; Hoang, Thanh V; Leonard, Melissa R; Bruney, Devin G; Wagner, Brad D; Dowd, Joseph Richard; Leone, Gustavo; Ostrowski, Michael C; Robinson, Michael L

    2016-02-15

    Lens epithelial cells express many receptor tyrosine kinases (RTKs) that stimulate PI3K-AKT and RAS-RAF-MEK-ERK intracellular signaling pathways. These pathways ultimately activate the phosphorylation of key cellular transcription factors and other proteins that control proliferation, survival, metabolism, and differentiation in virtually all cells. Among RTKs in the lens, only stimulation of fibroblast growth factor receptors (FGFRs) elicits a lens epithelial cell to fiber cell differentiation response in mammals. Moreover, although the lens expresses three different Fgfr genes, the isolated removal of Fgfr2 at the lens placode stage inhibits both lens cell survival and fiber cell differentiation. Phosphatase and tensin homolog (PTEN), commonly known as a tumor suppressor, inhibits ERK and AKT activation and initiates both apoptotic pathways, and cell cycle arrest. Here, we show that the combined deletion of Fgfr2 and Pten rescues the cell death phenotype associated with Fgfr2 loss alone. Additionally, Pten removal increased AKT and ERK activation, above the levels of controls, in the presence or absence of Fgfr2. However, isolated deletion of Pten failed to stimulate ectopic fiber cell differentiation, and the combined deletion of Pten and Fgfr2 failed to restore differentiation-specific Aquaporin0 and DnaseIIβ expression in the lens fiber cells.

  19. Tunable optofluidic birefringent lens.

    PubMed

    Wee, D; Hwang, S H; Song, Y S; Youn, J R

    2016-05-01

    An optofluidic birefringent lens is demonstrated using hydrodynamic liquid-liquid (L(2)) interfaces in a microchannel. The L(2) lens comprises a nematic liquid crystal (NLC) phase and an optically isotropic phase for the main stream and the surrounding sub-stream, respectively. When the optofluidic device is subjected to a sufficiently strong electric field perpendicular to the flow direction, NLCs are allowed to orient along the external field rather than the flow direction overcoming fluidic viscous stress. The characteristics of the optofluidic birefringence lens are investigated by experimental and numerical analyses. The difference between the refractive indices of the main stream and the sub-stream changes according to the polarization direction of incident light, which determines the optical behaviour of the lens. The incidence of s-polarized light leads to a short focal point, while p-polarized light has a relatively long focal distance from the same L(2) interface. The curvatures and focal lengths of the lens are successfully evaluated by a hydrodynamic theory of NLCs and a simple ray-tracing model. PMID:27035877

  20. The IAA RAS Correlator First Results

    NASA Technical Reports Server (NTRS)

    Surkis, Igor; Melnikov, Alexey; Shantyr, Violet; Zimovsky, Vladimir

    2010-01-01

    In 2009 the national Russian VLBI observations were processed by the new correlator ARC (Astrometric Radiointerferometric Correlator). The ARC is a VSI-H correlator and equipped with Mark 5B playback terminals. During 2009 ARC was used to process a series of VLBI sessions, observed on stations Svetloe, Zelenchukskaya, and Badary. NGS files were formed, and EOP parameters were obtained by IAA RAS Analysis Center. The accuracies of the pole coordinates and UT1-UTC were 1-2 mas and 0.07-0.1 ms, respectively.

  1. Small molecule binding sites on the Ras:SOS complex can be exploited for inhibition of Ras activation.

    PubMed

    Winter, Jon J G; Anderson, Malcolm; Blades, Kevin; Brassington, Claire; Breeze, Alexander L; Chresta, Christine; Embrey, Kevin; Fairley, Gary; Faulder, Paul; Finlay, M Raymond V; Kettle, Jason G; Nowak, Thorsten; Overman, Ross; Patel, S Joe; Perkins, Paula; Spadola, Loredana; Tart, Jonathan; Tucker, Julie A; Wrigley, Gail

    2015-03-12

    Constitutively active mutant KRas displays a reduced rate of GTP hydrolysis via both intrinsic and GTPase-activating protein-catalyzed mechanisms, resulting in the perpetual activation of Ras pathways. We describe a fragment screening campaign using X-ray crystallography that led to the discovery of three fragment binding sites on the Ras:SOS complex. The identification of tool compounds binding at each of these sites allowed exploration of two new approaches to Ras pathway inhibition by stabilizing or covalently modifying the Ras:SOS complex to prevent the reloading of Ras with GTP. Initially, we identified ligands that bound reversibly to the Ras:SOS complex in two distinct sites, but these compounds were not sufficiently potent inhibitors to validate our stabilization hypothesis. We conclude by demonstrating that covalent modification of Cys118 on Ras leads to a novel mechanism of inhibition of the SOS-mediated interaction between Ras and Raf and is effective at inhibiting the exchange of labeled GDP in both mutant (G12C and G12V) and wild type Ras.

  2. Metamaterial lens design

    NASA Astrophysics Data System (ADS)

    Shepard, Ralph Hamilton, III

    Developments in nanotechnology and material science have produced optical materials with astonishing properties. Theory and experimentation have demonstrated that, among other properties, the law of refraction is reversed at an interface between a naturally occurring material and these so-called metamaterials. As the technology advances metamaterials have the potential to vastly impact the field of optical science. In this study we provide a foundation for future work in the area of geometric optics and lens design with metamaterials. The concept of negative refraction is extended to derive a comprehensive set of first-order imaging principles as well as an exhaustive aberration theory to 4th order. Results demonstrate congruence with the classical theory; however, negative refraction introduces a host of novel properties. In terms of aberration theory, metamaterials present the lens designer with increased flexibility. A singlet can be bent to produce either positive or negative spherical aberration (regardless of its focal length), its contribution to coma can become independent of its conjugate factor, and its field curvature takes on the opposite sign of its focal power. This is shown to be advantageous in some designs such as a finite conjugate relay lens; however, in a wider field of view landscape lens we demonstrate a metamaterial's aberration properties may be detrimental. This study presents the first comprehensive investigation of metamaterial lenses using industry standard lens design software. A formal design study evaluates the performance of doublet and triplet lenses operating at F/5 with a 100 mm focal length, a 20° half field of view, and specific geometric constraints. Computer aided optimization and performance evaluation provide experimental controls to remove designer-induced bias from the results. Positive-index lenses provide benchmarks for comparison to metamaterial systems subjected to identical design constraints. We find that

  3. Microoptical compound lens

    DOEpatents

    Sweatt, William C.; Gill, David D.

    2007-10-23

    An apposition microoptical compound lens comprises a plurality of lenslets arrayed around a segment of a hollow, three-dimensional optical shell. The lenslets collect light from an object and focus the light rays onto the concentric, curved front surface of a coherent fiber bundle. The fiber bundle transports the light rays to a planar detector, forming a plurality of sub-images that can be reconstructed as a full image. The microoptical compound lens can have a small size (millimeters), wide field of view (up to 180.degree.), and adequate resolution for object recognition and tracking.

  4. Zoom optical system using tunable polymer lens

    NASA Astrophysics Data System (ADS)

    Liang, Dan; Wang, Xuan Yin

    2016-07-01

    This paper demonstrated a zoom optical system with variable magnification based on the tunable polymer lens. The designed system mainly consists of two polymer lenses, voice coil motors, a doublet lens and CMOS chip. The zoom magnification can be adjusted by altering the focal length of the two elastic polymer lenses synergistically through controlling the output displacement of the voice coil motor. A static doublet lens in combination with the polymer lenses stabilize the image plane at the CMOS chip. The optical structure of the zoom system is presented, as well as a detailed description including the lens materials and fabrication process. Images with each zoom magnification are captured, and the Spot diagram and MTF are simulated using Zemax software. A change in magnification from 0.13×to 8.44×is demonstrated within the tiny 0.4 mm variation of the displacement load, and produce a 16.1×full range of magnification experimentally. Simulation analyses show that all the radii of the spot diagram under different magnifications are less than 11.3 um, and the modulation transfer function reaches 107 line pairs per mm. The designed optical system shows the potential for developing stable, integrated, and low-cost zoom systems with large magnification range.

  5. Neural induction in the absence of Organizer in salamanders is mediated by Ras/MAPK

    PubMed Central

    Hurtado, Cecilia; De Robertis, E. M.

    2007-01-01

    Research on the mechanisms of embryonic induction had a great setback in the 1940s when Barth discovered and Holtfreter confirmed that ectoderm of Ambystoma maculatum salamander embryos could form brain tissue when cultured in a simple saline solution. We have revisited this classical experiment and found that when cultured animal cap ectoderm attaches to a glass substratum, it can self-organize to form complex organs such as brain vesicles, eyes, lens and olfactory placodes. Only anterior neural organs were generated. Under these culture conditions ERK became diphosphorylated, indicating a sustained activation of the Ras/MAPK pathway. Using sand particles as an heterologous neural inducer similar results were obtained. Addition of U0126, a specific antagonist of MEK, the enzyme that phosphorylates ERK/MAPK, inhibited neural differentiation. The closely related control compound U0124 had no effect. We conclude that neural induction in the absence of organizer in Ambystoma maculatum requires Ras/MAPK activation. These findings provide a molecular explanation for the activity of heterologous neural inducers that dominated thinking in amphibian experimental embryology for many decades. PMID:17540356

  6. A Tribute to Len Barton

    ERIC Educational Resources Information Center

    Tomlinson, Sally

    2010-01-01

    This article constitutes a short personal tribute to Len Barton in honour of his work and our collegial relationship going back over 30 years. It covers how Len saw his intellectual project of providing critical sociological and political perspectives on special education, disability and inclusion, and his own radical political perspectives. Len's…

  7. Ras chaperones: new targets for cancer and immunotherapy.

    PubMed

    Kloog, Yoel; Elad-Sfadia, Galit; Haklai, Roni; Mor, Adam

    2013-01-01

    The Ras inhibitor S-trans,trans-farnesylthiosalicylic acid (FTS, Salirasib®) interferes with Ras membrane interactions that are crucial for Ras-dependent signaling and cellular transformation. FTS had been successfully evaluated in clinical trials of cancer patients. Interestingly, its effect is mediated by targeting Ras chaperones that serve as key coordinators for Ras proper folding and delivery, thus offering a novel target for cancer therapy. The development of new FTS analogs has revealed that the specific modifications to the FTS carboxyl group by esterification and amidation yielded compounds with improved growth inhibitory activity. When FTS was combined with additional therapeutic agents its activity toward Ras was significantly augmented. FTS should be tested not only in cancer but also for genetic diseases associated with abnormal Ras signaling, as well as for various inflammatory and autoimmune disturbances, where Ras plays a major role. We conclude that FTS has a great potential both as a safe anticancer drug and as a promising immune modulator agent. PMID:25033809

  8. H-Ras regulation of TRAIL death receptor mediated apoptosis

    PubMed Central

    Chen, Jun-Jie; Bozza, William P.; Di, Xu; Zhang, Yaqin; Hallett, William; Zhang, Baolin

    2014-01-01

    TNF-related apoptosis-inducing ligand (TRAIL) induces apoptosis through the death receptors (DRs) 4 and/or 5 expressed on the cell surface. Multiple clinical trials are underway to evaluate the antitumor activity of recombinant human TRAIL and agonistic antibodies to DR4 or DR5. However, their therapeutic potential is limited by the high frequency of cancer resistance. Here we provide evidence demonstrating the role of H-Ras in TRAIL receptor mediated apoptosis. By analyzing the genome wide mRNA expression data of the NCI60 cancer cell lines, we found that H-Ras expression was consistently upregulated in TRAIL-resistant cell lines. By contrast, no correlation was found between TRAIL sensitivity and K-Ras expression levels or their mutational profiles. Notably, H-Ras upregulation associated with a surface deficiency of TRAIL death receptors. Selective inhibition of H-Ras activity in TRAIL-resistant cells restored the surface expression of both DR4 and DR5 without changing their total protein levels. The resulting cells became highly susceptible to both TRAIL and agonistic DR5 antibody, whereas K-Ras inhibition had little or no effect on TRAIL-induced apoptosis, indicating H-Ras plays a distinct role in the regulation of TRAIL death receptors. Further studies are warranted to determine the therapeutic potential of H-Ras-specific inhibitors in combination with TRAIL receptor agonists. PMID:25026275

  9. Thin Lens Ray Tracing.

    ERIC Educational Resources Information Center

    Gatland, Ian R.

    2002-01-01

    Proposes a ray tracing approach to thin lens analysis based on a vector form of Snell's law for paraxial rays as an alternative to the usual approach in introductory physics courses. The ray tracing approach accommodates skew rays and thus provides a complete analysis. (Author/KHR)

  10. The Lens of Chemistry

    ERIC Educational Resources Information Center

    Thalos, Mariam

    2013-01-01

    Chemistry possesses a distinctive theoretical lens--a distinctive set of theoretical concerns regarding the dynamics and transformations of a perplexing variety of organic and nonorganic substances--to which it must be faithful. Even if it is true that chemical facts bear a special (reductive) relationship to physical facts, nonetheless it will…

  11. Thermal Lens Microscope

    NASA Astrophysics Data System (ADS)

    Uchiyama, Kenji; Hibara, Akihide; Kimura, Hiroko; Sawada, Tsuguo; Kitamori, Takehiko

    2000-09-01

    We developed a novel laser microscope based on the thermal lens effect induced by a coaxial beam comprised of excitation and probe beams. The signal generation mechanism was confirmed to be an authentic thermal lens effect from the measurement of signal and phase dependences on optical configurations between the sample and the probe beam focus, and therefore, the thermal lens effect theory could be applied. Two-point spatial resolution was determined by the spot size of the excitation beam, not by the thermal diffusion length. Sensitivity was quite high, and the detection ability, evaluated using a submicron microparticle containing dye molecules, was 0.8 zmol/μm2, hence a distribution image of trace chemical species could be obtained quantitatively. In addition, analytes are not restricted to fluorescent species, therefore, the thermal lens microscope is a promising analytical microscope. A two-dimensional image of a histamine molecule distribution, which was produced in mast cells at the femtomole level in a human nasal mucous polyp, was obtained.

  12. Endogenous K-ras signaling in erythroid differentiation.

    PubMed

    Zhang, Jing; Lodish, Harvey F

    2007-08-15

    K-ras is one of the most frequently mutated genes in virtually all types of human cancers. Using mouse fetal liver erythroid progenitors as a model system, we studied the role of endogenous K-ras signaling in erythroid differentiation. When oncogenic K-ras is expressed from its endogenous promoter, it hyperactivates cytokine-dependent signaling pathways and results in a partial block in erythroid differentiation. In erythroid progenitors deficient in K-ras, cytokine-dependent Akt activation is greatly reduced, leading to delays in erythroid differentiation. Thus, both loss- and gain-of-Kras functions affect erythroid differentiation through modulation of cytokine signaling. These results support the notion that in human cancer patients oncogenic Ras signaling might be controlled by antagonizing essential cytokines.

  13. Effect of rigid gas permeable lens flexure on vision.

    PubMed

    Sorbara, L; Fonn, D; MacNeill, K

    1992-12-01

    The flexure of spherical rigid lenses (various materials) and a soft lens was measured using automated over-keratometry on 6 adapted rigid lens wearers (12 eyes) whose corneal toricity ranged from 1.37 to 3.87 D. The results showed: (1) that there was no significant difference in flexure between polymethyl methacrylate (PMMA), silicone acrylate, and the fluorosilicone acrylate lenses (whose Dks ranged from 0 to 115). However, Advent (fluoropolymer) did flex significantly more than the other rigid lenses, and significantly less than the soft lens (Bausch & Lomb U4) and (2) that lens flexure of the rigid lenses did not alter over a 2-h period. We also measured high and low contrast visual acuity (HCVA and LCVA), and the results from subjects wearing Advent and the soft lens were significantly worse than with the other rigid lenses. Finally, the results of this study showed no correlation between rigid lens flexure and permeability and between rigid lens flexure and visual acuity when Advent was excluded from the linear regression analysis.

  14. Ras activation and symmetry breaking during Dictyostelium chemotaxis.

    PubMed

    Kortholt, Arjan; Keizer-Gunnink, Ineke; Kataria, Rama; Van Haastert, Peter J M

    2013-10-01

    Central to chemotaxis is the molecular mechanism by which a shallow spatial gradient of chemoattractant induces symmetry breaking of activated signaling molecules. Previously, we have used Dictyostelium mutants to investigate the minimal requirements for chemotaxis, and identified a basal signaling module providing activation of Ras and F-actin at the leading edge. Here, we show that Ras activation after application of a pipette releasing the chemoattractant cAMP has three phases, each depending on specific guanine-nucleotide-exchange factors (GEFs). Initially a transient activation of Ras occurs at the entire cell boundary, which is proportional to the local cAMP concentrations and therefore slightly stronger at the front than in the rear of the cell. This transient Ras activation is present in gα2 (gpbB)-null cells but not in gβ (gpbA)-null cells, suggesting that Gβγ mediates the initial activation of Ras. The second phase is symmetry breaking: Ras is activated only at the side of the cell closest to the pipette. Symmetry breaking absolutely requires Gα2 and Gβγ, but not the cytoskeleton or four cAMP-induced signaling pathways, those dependent on phosphatidylinositol (3,4,5)-triphosphate [PtdIns(3,4,5)P3], cGMP, TorC2 and PLA2. As cells move in the gradient, the crescent of activated Ras in the front half of the cell becomes confined to a small area at the utmost front of the cell. Confinement of Ras activation leads to cell polarization, and depends on cGMP formation, myosin and F-actin. The experiments show that activation, symmetry breaking and confinement of Ras during Dictyostelium chemotaxis uses different G-protein subunits and a multitude of Ras GEFs and GTPase-activating proteins (GAPs).

  15. RAS Synthetic Lethal Screens Revisited: Still Seeking the Elusive Prize?

    PubMed

    Downward, Julian

    2015-04-15

    The RAS genes are critical oncogenic drivers activated by point mutation in some 20% of human malignancies. However, no pharmacologic approaches to targeting RAS proteins directly have yet succeeded, leading to suggestions that these proteins may be "undruggable." This has led to two alternative indirect approaches to targeting RAS function in cancer. One has been to target RAS signaling pathways downstream at tractable enzymes such as kinases, particularly in combination. The other, which is the focus of this review, has been to seek targets that are essential in cells bearing an activated RAS oncogene, but not those without. This synthetic lethal approach, while rooted in ideas from invertebrate genetics, has been inspired most strongly by the successful use of PARP inhibitors, such as olaparib, in the clinic to treat BRCA defective cancers. Several large-scale screens have been carried out using RNA interference-mediated expression silencing to find genes that are uniquely essential to RAS-mutant but not wild-type cells. These screens have been notable for the low degree of overlap between their results, with the possible exception of proteasome components, and have yet to lead to successful new clinical approaches to the treatment of RAS-mutant cancers. Possible reasons for these disappointing results are discussed here, along with a reevaluation of the approaches taken. On the basis of experience to date, RAS synthetic lethality has so far fallen some way short of its original promise and remains unproven as an approach to finding effective new ways of tackling RAS-mutant cancers. Clin Cancer Res; 21(8); 1802-9. ©2015 AACR. See all articles in this CCR Focus section, "Targeting RAS-Driven Cancers." PMID:25878361

  16. Regulation of collagen I gene expression by ras.

    PubMed Central

    Slack, J L; Parker, M I; Robinson, V R; Bornstein, P

    1992-01-01

    Although transformation of rodent fibroblasts can lead to dramatic changes in expression of extracellular matrix genes, the molecular basis and physiological significance of these changes remain poorly understood. In this study, we have investigated the mechanism(s) by which ras affects expression of the genes encoding type I collagen. Levels of both alpha 1(I) and alpha 2(I) collagen mRNAs were markedly reduced in Rat 1 fibroblasts overexpressing either the N-rasLys-61 or the Ha-rasVal-12 oncogene. In fibroblasts conditionally transformed with N-rasLys-61, alpha 1(I) transcript levels began to decline within 8 h of ras induction and reached 1 to 5% of control levels after 96 h. In contrast, overexpression of normal ras p21 had no effect on alpha 1(I) or alpha 2(I) mRNA levels. Nuclear run-on experiments demonstrated that the transcription rates of both the alpha 1(I) and alpha 2(I) genes were significantly reduced in ras-transformed cells compared with those in parental cells. In addition, the alpha 1(I) transcript was less stable in transformed cells. Chimeric plasmids containing up to 3.6 kb of alpha 1(I) 5'-flanking DNA and up to 2.3 kb of the 3'-flanking region were expressed at equivalent levels in both normal and ras-transformed fibroblasts. However, a cosmid clone containing the entire mouse alpha 1(I) gene, including 3.7 kb of 5'- and 4 kb of 3'-flanking DNA, was expressed at reduced levels in fibroblasts overexpressing oncogenic ras. We conclude that oncogenic ras regulates the type I collagen genes at both transcriptional and posttranscriptional levels and that this effect, at least for the alpha 1(I) gene, may be mediated by sequences located either within the body of the gene itself or in the distal 3'-flanking region. Images PMID:1406656

  17. Down-regulation of let-7 microRNA increased K-ras expression in lung damage induced by radon.

    PubMed

    Chen, Zhihai; Wang, Dapeng; Gu, Chao; Liu, Xing; Pei, Weiwei; Li, Jianxiang; Cao, Yi; Jiao, Yang; Tong, Jian; Nie, Jihua

    2015-09-01

    Radon has long been recognized as a human carcinogen leading to lung cancer, but the underlying mechanisms remain obscure. Recent studies have shown that the let-7 microRNA and K-ras play an important role in the development of various cancers. However, the exact role between let-7 and K-ras in radon induced lung damage has not been explored so far. In the present study, wistar rats and human bronchial epithelial (HBE) cells were long-term exposed to radon, and then alterations in histological pathology of rat lung tissue, ROS, antioxidant enzymes activities and clonogenic formation in HBE cells, as well as changes in let-7 and K-ras expression were determined to observe the adverse effects induced by radon. The results showed that long-term exposure to radon produced severe lung damage in rats, significantly increased ROS production and clonogenic formation ratios and decreased SOD activities in HBE cells. In addition, an obvious down-regulation of let-7 and up-regulation of K-ras were also revealed both in mRNA and in protein level in lung tissue of rats and HBE cells exposed to radon. Furthermore, a significant down-regulation of K-ras was then confirmed in both let-7b-3p and let-7a-2-3p transfected HBE cells. Taken together, the present results propose an involvement of let-7 microRNA and K-ras in radon induced lung damage both in vivo and in vitro, which may thus be of potential value in early diagnosis and therapy of radon-induced lung tumorgenesis.

  18. Germline expression of H-Ras(G12V) causes neurological deficits associated to Costello syndrome.

    PubMed

    Viosca, J; Schuhmacher, A J; Guerra, C; Barco, A

    2009-02-01

    Costello syndrome (CS) is a rare congenital disorder caused by germline activation of H-Ras oncogenes. A mouse model of CS generated by introduction of an oncogenic Gly12Val mutation in the mouse H-Ras locus using homologous recombination in embryonic stem (ES) cells has been recently described. These mice phenocopied some of the abnormalities observed in patients with CS, including facial dysmorphia and cardiomyopathies. We investigated here their neurological and behavioral phenotype. The analysis of H-Ras(G12V) mice revealed phenotypes that resembled the hyperemotivity, hypersensibility and cognitive impairments observed in children with CS. Stronger neurological deficits were found in the analysis of mice homozygous for this mutation than in the analysis of heterozygous mice, suggesting the existence of a gene dose effect. These mice represent the first mouse model for CS, offering an experimental tool to study the molecular and physiological alterations underlying the neurological manifestations of CS and to test new therapies aimed at preventing or ameliorating the cognitive and emotional impairments associated to this condition. PMID:18823404

  19. RAS-RAF-MEK-dependent oxidative cell death involving voltage-dependent anion channels.

    PubMed

    Yagoda, Nicholas; von Rechenberg, Moritz; Zaganjor, Elma; Bauer, Andras J; Yang, Wan Seok; Fridman, Daniel J; Wolpaw, Adam J; Smukste, Inese; Peltier, John M; Boniface, J Jay; Smith, Richard; Lessnick, Stephen L; Sahasrabudhe, Sudhir; Stockwell, Brent R

    2007-06-14

    Therapeutics that discriminate between the genetic makeup of normal cells and tumour cells are valuable for treating and understanding cancer. Small molecules with oncogene-selective lethality may reveal novel functions of oncoproteins and enable the creation of more selective drugs. Here we describe the mechanism of action of the selective anti-tumour agent erastin, involving the RAS-RAF-MEK signalling pathway functioning in cell proliferation, differentiation and survival. Erastin exhibits greater lethality in human tumour cells harbouring mutations in the oncogenes HRAS, KRAS or BRAF. Using affinity purification and mass spectrometry, we discovered that erastin acts through mitochondrial voltage-dependent anion channels (VDACs)--a novel target for anti-cancer drugs. We show that erastin treatment of cells harbouring oncogenic RAS causes the appearance of oxidative species and subsequent death through an oxidative, non-apoptotic mechanism. RNA-interference-mediated knockdown of VDAC2 or VDAC3 caused resistance to erastin, implicating these two VDAC isoforms in the mechanism of action of erastin. Moreover, using purified mitochondria expressing a single VDAC isoform, we found that erastin alters the permeability of the outer mitochondrial membrane. Finally, using a radiolabelled analogue and a filter-binding assay, we show that erastin binds directly to VDAC2. These results demonstrate that ligands to VDAC proteins can induce non-apoptotic cell death selectively in some tumour cells harbouring activating mutations in the RAS-RAF-MEK pathway.

  20. Nonlinear ionizing radiation-induced changes in eye lens cell proliferation, cyclin D1 expression and lens shape

    PubMed Central

    Markiewicz, Ewa; Barnard, Stephen; Haines, Jackie; Coster, Margaret; van Geel, Orry; Wu, Weiju; Richards, Shane; Ainsbury, Elizabeth; Rothkamm, Kai; Bouffler, Simon; Quinlan, Roy A.

    2015-01-01

    Elevated cataract risk after radiation exposure was established soon after the discovery of X-rays in 1895. Today, increased cataract incidence among medical imaging practitioners and after nuclear incidents has highlighted how little is still understood about the biological responses of the lens to low-dose ionizing radiation (IR). Here, we show for the first time that in mice, lens epithelial cells (LECs) in the peripheral region repair DNA double strand breaks (DSB) after exposure to 20 and 100 mGy more slowly compared with circulating blood lymphocytes, as demonstrated by counts of γH2AX foci in cell nuclei. LECs in the central region repaired DSBs faster than either LECs in the lens periphery or lymphocytes. Although DSB markers (γH2AX, 53BP1 and RAD51) in both lens regions showed linear dose responses at the 1 h timepoint, nonlinear responses were observed in lenses for EdU (5-ethynyl-2′-deoxy-uridine) incorporation, cyclin D1 staining and cell density after 24 h at 100 and 250 mGy. After 10 months, the lens aspect ratio was also altered, an indicator of the consequences of the altered cell proliferation and cell density changes. A best-fit model demonstrated a dose-response peak at 500 mGy. These data identify specific nonlinear biological responses to low (less than 1000 mGy) dose IR-induced DNA damage in the lens epithelium. PMID:25924630

  1. Elemental profiles in Emory mouse lens

    SciTech Connect

    Bagchi, M.; Emanuel, K. )

    1991-01-01

    Energy dispersive x-ray microprobe analysis was used to determine the distribution of chloride, potassium, phosphorus and sulfur in the epithelial cells of the lenses obtained from 3 to 7 month old Emory mice and 7 month old cataract resistant strain of Emory mice. Rapidly frozen lenses were fractured in the frozen state and lyophilized. The anterior epithelial cells were analyzed from equator to equator. The results show that the epithelial cells of the 7 month old Emory mouse lens have considerably higher amounts of chloride, sulfur, potassium and phosphorus. Presence of increased amount of potassium in the epithelial cells is intriguing. The data obtained from these experiments show that the changes in the elemental levels of epithelial cells are similar to observed alteration found in the lens fiber mass of 7 month old Emory mouse.

  2. Uhrf1 and Dnmt1 are required for development and maintenance of the zebrafish lens

    PubMed Central

    Tittle, Rachel K.; Sze, Ryan; Ng, Anthony; Nuckels, Richard J.; Swartz, Mary E.; Anderson, Ryan M.; Bosch, Justin; Stainier, Didier Y.R.; Eberhart, Johann K.; Gross, Jeffrey M.

    2010-01-01

    SUMMARY DNA methylation is one of the key mechanisms underlying the epigenetic regulation of gene expression. During DNA replication, the methylation pattern of the parent strand is maintained on the replicated strand through the action of Dnmt1 (DNA Methyltransferase 1). In mammals, Dnmt1 is recruited to hemimethylated replication foci by Uhrf1 (Ubiquitin-like, Containing PHD and RING Finger Domains 1). Here we show that Uhrf1 is required for DNA methylation in vivo during zebrafish embryogenesis. Due in part to the early embryonic lethality of Dnmt1 and Uhrf1 knockout mice, roles for these proteins during lens development have yet to be reported. We show that zebrafish mutants in uhrf1 and dnmt1 have defects in lens development and maintenance. uhrf1 and dnmt1 are expressed in the lens epithelium, and in the absence of Uhrf1 or of catalytically active Dnmt1, lens epithelial cells have altered gene expression and reduced proliferation in both mutant backgrounds. This is correlated with a wave of apoptosis in the epithelial layer, which is followed by apoptosis and unraveling of secondary lens fibers. Despite these disruptions in the lens fiber region, lens fibers express appropriate differentiation markers. The results of lens transplant experiments demonstrate that Uhrf1 and Dnmt1 functions are required lens-autonomously, but perhaps not cell-autonomously, during lens development in zebrafish. These data provide the first evidence that Uhrf1 and Dnmt1 function is required for vertebrate lens development and maintenance. PMID:21126517

  3. The small G protein H-Ras in the mesolimbic system is a molecular gateway to alcohol-seeking and excessive drinking behaviors.

    PubMed

    Ben Hamida, Sami; Neasta, Jeremie; Lasek, Amy W; Kharazia, Viktor; Zou, Mimi; Carnicella, Sebastien; Janak, Patricia H; Ron, Dorit

    2012-11-01

    Uncontrolled consumption of alcohol is a hallmark of alcohol abuse disorders; however, the central molecular mechanisms underlying excessive alcohol consumption are still unclear. Here, we report that the GTP binding protein, H-Ras in the nucleus accumbens (NAc) plays a key role in neuroadaptations that underlie excessive alcohol-drinking behaviors. Specifically, acute (15 min) systemic administration of alcohol (2.5 g/kg) leads to the activation of H-Ras in the NAc of mice, which is observed even 24 h later. Similarly, rat operant self-administration of alcohol (20%) also results in the activation of H-Ras in the NAc. Using the same procedures, we provide evidence suggesting that the exchange factor GRF1 is upstream of H-Ras activation by alcohol. Importantly, we show that infection of mice NAc with lentivirus expressing a short hairpin RNA that targets the H-Ras gene produces a significant reduction of voluntary consumption of 20% alcohol. In contrast, knockdown of H-Ras in the NAc of mice did not alter water, quinine, and saccharin intake. Furthermore, using two-bottle choice and operant self-administration procedures, we show that inhibiting H-Ras activity by intra-NAc infusion of the farnesyltransferase inhibitor, FTI-276, produced a robust decrease of rats' alcohol drinking; however, sucrose consumption was unaltered. Finally, intra-NAc infusion of FTI-276 also resulted in an attenuation of seeking for alcohol. Together, these results position H-Ras as a central molecular mediator of alcohol's actions within the mesolimbic system and put forward the potential value of the enzyme as a novel target to treat alcohol use disorders.

  4. K-Ras mutation detection in liquid biopsy and tumor tissue as prognostic biomarker in patients with pancreatic cancer: a systematic review with meta-analysis.

    PubMed

    Li, Tao; Zheng, Yuanting; Sun, Hong; Zhuang, Rongyuan; Liu, Jing; Liu, Tianshu; Cai, Weimin

    2016-07-01

    K-Ras gene mutations have been found in most pancreatic cancers; however, conflicting data on the prognostic value of K-Ras mutations in pancreatic cancer have been published. We conducted a meta-analysis to assess its prognostic significance. Literature searches of PubMed, EMBASE, Cochrane Library, Web of Science and Google Scholar were performed through December 2015 to identify publications exploring the association of K-Ras mutation with overall survival. Forty eligible studies involving 3427 patients with pancreatic cancer were included in the present meta-analysis. Our analysis showed a hazard ratio (HR) of negative association with survival of 1.61 [95 % confidence interval (CI) 1.36-1.90; p < 0.01] in K-Ras mutant pancreatic cancer patients. In subgroup analyses, K-Ras mutations detected in tumor tissues and in liquid biopsies had HRs of 1.37 (95 % CI 1.20-1.57; p < 0.01) and 3.16 (95 % CI 2.1-4.71; p < 0.01), respectively. In addition, the HR was higher when K-Ras mutations were detected in fresh frozen samples (HR = 2.01, 95 % CI 1.28-3.16, p = 0.002) than in formalin-fixed, paraffin-embedded (FFPE) samples (HR = 1.29, 95 % CI 1.12-1.49, p < 0.01). Though K-Ras alterations are more frequent among non-East Asian individuals than East Asian individuals, there were no significant differences in HRs of survival between the two ethnic subgroups. In conclusion, this meta-analysis suggests that K-Ras mutations are associated with a worse overall survival in pancreatic cancer patients, especially when mutations are detected in liquid biopsies or fresh frozen tumor tissue samples.

  5. K-Ras mutation detection in liquid biopsy and tumor tissue as prognostic biomarker in patients with pancreatic cancer: a systematic review with meta-analysis.

    PubMed

    Li, Tao; Zheng, Yuanting; Sun, Hong; Zhuang, Rongyuan; Liu, Jing; Liu, Tianshu; Cai, Weimin

    2016-07-01

    K-Ras gene mutations have been found in most pancreatic cancers; however, conflicting data on the prognostic value of K-Ras mutations in pancreatic cancer have been published. We conducted a meta-analysis to assess its prognostic significance. Literature searches of PubMed, EMBASE, Cochrane Library, Web of Science and Google Scholar were performed through December 2015 to identify publications exploring the association of K-Ras mutation with overall survival. Forty eligible studies involving 3427 patients with pancreatic cancer were included in the present meta-analysis. Our analysis showed a hazard ratio (HR) of negative association with survival of 1.61 [95 % confidence interval (CI) 1.36-1.90; p < 0.01] in K-Ras mutant pancreatic cancer patients. In subgroup analyses, K-Ras mutations detected in tumor tissues and in liquid biopsies had HRs of 1.37 (95 % CI 1.20-1.57; p < 0.01) and 3.16 (95 % CI 2.1-4.71; p < 0.01), respectively. In addition, the HR was higher when K-Ras mutations were detected in fresh frozen samples (HR = 2.01, 95 % CI 1.28-3.16, p = 0.002) than in formalin-fixed, paraffin-embedded (FFPE) samples (HR = 1.29, 95 % CI 1.12-1.49, p < 0.01). Though K-Ras alterations are more frequent among non-East Asian individuals than East Asian individuals, there were no significant differences in HRs of survival between the two ethnic subgroups. In conclusion, this meta-analysis suggests that K-Ras mutations are associated with a worse overall survival in pancreatic cancer patients, especially when mutations are detected in liquid biopsies or fresh frozen tumor tissue samples. PMID:27225938

  6. RasGRP3 regulates the migration of glioma cells via interaction with Arp3

    PubMed Central

    Lee, Hae Kyung; Finniss, Susan; Cazacu, Simona; Xiang, Cunli; Poisson, Laila M.; Blumberg, Peter M.; Brodie, Chaya

    2015-01-01

    Glioblastoma (GBM), the most aggressive primary brain tumors, are highly infiltrative. Although GBM express high Ras activity and Ras proteins have been implicated in gliomagenesis, Ras-activating mutations are not frequent in these tumors. RasGRP3, an important signaling protein responsive to diacylglycerol (DAG), increases Ras activation. Here, we examined the expression and functions of RasGRP3 in GBM and glioma cells. RasGRP3 expression was upregulated in GBM specimens and glioma stem cells compared with normal brains and neural stem cells, respectively. RasGRP3 activated Ras and Rap1 in glioma cells and increased cell migration and invasion partially via Ras activation. Using pull-down assay and mass spectroscopy we identified the actin-related protein, Arp3, as a novel interacting protein of RasGRP3. The interaction of RasGRP3 and Arp3 was validated by immunofluorescence staining and co-immunoprecipitation, and PMA, which activates RasGRP3 and induces its translocation to the peri-nuclear region, increased the association of Arp3 and RasGRP3. Arp3 was upregulated in GBM, regulated cell spreading and migration and its silencing partially decreased these effects of RasGRP3 in glioma cells. In summary, RasGRP3 acts as an important integrating signaling protein of the DAG and Ras signaling pathways and actin polymerization and represents an important therapeutic target in GBM. PMID:25682201

  7. Studies of gravitational lens systems discovered in the Cosmic Lens All-Sky Survey

    NASA Astrophysics Data System (ADS)

    Rusin, David Joseph

    2001-11-01

    This thesis describes research conducted on and inspired by the Cosmic Lens All-Sky Survey (CLASS), which searches for new cases of gravitational lensing among compact radio sources. CLASS aims to provide the largest and best-studied sample of lens systems for use in constraining the properties of galaxy mass distributions, determining the Hubble parameter and placing limits on the cosmological constant. The goal of this thesis was to complete observations of the CLASS sample, discover and thoroughly investigate new lenses, and apply them to interesting astrophysical problems. We begin with a detailed overview of the CLASS project, including scientific goals, the radio source sample, survey observations, candidate selection and follow-ups. Results are then presented from the third phase of the CLASS survey (CLASS-3), which yielded three new gravitational lens systems. 130850+054 and 131152+199 both consist of a pair of lensed images. 131359+154 features six images of a single source, and is the first arcsecond-scale system in which a source is lensed into more than four images. We also present observations and modeling of the CLASS-2 gravitational lens B2319+051. We use the absence of detectable central images in deep radio maps of CLASS lens systems to place powerful constraints on the inner mass profiles of leasing galaxies. These analyses imply that the profile slopes cannot be much shallower than isothermal. Finally, we consider the relative frequency of two and four-image lens systems, and demonstrate that there is a statistically significant overdensity of quads in the CLASS sample. We investigate a range of factors that may be increasing the frequency of radio quads, including external shear fields, mass distributions flatter than the light, shallow leasing mass profiles, finite core radii, satellite galaxies, and alterations to the luminosity function for faint flat-spectrum radio sources. Surprisingly, none of these mechanisms provide a particularly

  8. Inhibition of SHP2-mediated dephosphorylation of Ras suppresses oncogenesis

    PubMed Central

    Bunda, Severa; Burrell, Kelly; Heir, Pardeep; Zeng, Lifan; Alamsahebpour, Amir; Kano, Yoshihito; Raught, Brian; Zhang, Zhong-Yin; Zadeh, Gelareh; Ohh, Michael

    2015-01-01

    Ras is phosphorylated on a conserved tyrosine at position 32 within the switch I region via Src kinase. This phosphorylation inhibits the binding of effector Raf while promoting the engagement of GTPase-activating protein (GAP) and GTP hydrolysis. Here we identify SHP2 as the ubiquitously expressed tyrosine phosphatase that preferentially binds to and dephosphorylates Ras to increase its association with Raf and activate downstream proliferative Ras/ERK/MAPK signalling. In comparison to normal astrocytes, SHP2 activity is elevated in astrocytes isolated from glioblastoma multiforme (GBM)-prone H-Ras(12V) knock-in mice as well as in glioma cell lines and patient-derived GBM specimens exhibiting hyperactive Ras. Pharmacologic inhibition of SHP2 activity attenuates cell proliferation, soft-agar colony formation and orthotopic GBM growth in NOD/SCID mice and decelerates the progression of low-grade astrocytoma to GBM in a spontaneous transgenic glioma mouse model. These results identify SHP2 as a direct activator of Ras and a potential therapeutic target for cancers driven by a previously ‘undruggable' oncogenic or hyperactive Ras. PMID:26617336

  9. RAS Synthetic Lethal Screens Revisited: Still Seeking the Elusive Prize?

    PubMed Central

    Downward, Julian

    2015-01-01

    The RAS genes are critical oncogenic drivers activated by point mutation in some 20% of human malignancies. However, no pharmacological approaches to targeting RAS proteins directly have yet succeeded, leading to suggestions that these proteins may be “undruggable.” This has led to two alternative indirect approaches to targeting RAS function in cancer. One has been to target RAS signaling pathways downstream at tractable enzymes such as kinases, particularly in combination. The other, which is the focus of this review, has been to seek targets that are essential in cells bearing an activated RAS oncogene, but not those without. This synthetic lethal approach, while rooted in ideas from invertebrate genetics, has been inspired most strongly by the successful use of PARP inhibitors, such as olaparib, in the clinic to treat BRCA defective cancers. Several large-scale screens have been carried out using RNA interference-mediated expression silencing to find genes that are uniquely essential to RAS mutant but not wild type cells. These screens have been notable for the low degree of overlap between their results, with the possible exception of proteasome components, and have yet to lead to successful new clinical approaches to the treatment of RAS mutant cancers. Possible reasons for these disappointing results are discussed here, along with a re-evaluation of the approaches taken. Based on experience to date, RAS synthetic lethality has so far fallen some way short of its original promise and remains unproven as an approach to finding effective new ways of tackling RAS mutant cancers. PMID:25878361

  10. GTP-dependent association of Raf-1 with Ha-Ras: identification of Raf as a target downstream of Ras in mammalian cells.

    PubMed Central

    Koide, H; Satoh, T; Nakafuku, M; Kaziro, Y

    1993-01-01

    Ras is involved in signal transduction of various factors for growth, differentiation, and oncogenesis. Recent studies have revealed several proteins that function upstream and downstream of the Ras signaling pathway. However, its immediate downstream target molecular has not yet been identified. In an effort to identify the Ras-associated downstream proteins, we added recombinant Ha-Ras in a GTP-bound form to cell-free lysates and used several antibodies against Ras to immunoprecipitate Ras complexes. We found that a serine/threonine kinase, Raf-1, was coimmunoprecipitated with Ha-Ras by two anti-Ras antibodies (LA069 and Y13-238), whereas a neutralizing antibody against Ras (Y13-259) could not precipitate Raf-1. The coimmunoprecipitation was observed with a complex of Ras and guanosine 5'-[gamma- thio]triphosphate but not with a complex of Ras and guanosine 5'-[beta-thio]diphosphate. The GTP-dependent association of Ha-Ras with Raf-1 was observed with lysates of various types of cultured cells, including NIH 3T3, pheochromocytoma (PC) 12, Ba/F3, and Jurkat T cells, and also with crude extracts from rat brain. Furthermore, Raf-1 was precipitated with a transforming Ha-Ras mutant ([Val12]Ras) and wild-type Ha-Ras but not with an effector-region mutant ([Leu35,ARg37]Ras) that lacks transforming activity. These results indicate that Ras.GTP physically associates with Raf either directly or through other component(s) and strongly suggest that Raf functions in close downstream proximity to Ras in mammalian cells. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 PMID:8378348

  11. Serum-dependent transcriptional networks identify distinct functional roles for H-Ras and N-Ras during initial stages of the cell cycle

    PubMed Central

    2009-01-01

    Background Using oligonucleotide microarrays, we compared transcriptional profiles corresponding to the initial cell cycle stages of mouse fibroblasts lacking the small GTPases H-Ras and/or N-Ras with those of matching, wild-type controls. Results Serum-starved wild-type and knockout ras fibroblasts had very similar transcriptional profiles, indicating that H-Ras and N-Ras do not significantly control transcriptional responses to serum deprivation stress. In contrast, genomic disruption of H-ras or N-ras, individually or in combination, determined specific differential gene expression profiles in response to post-starvation stimulation with serum for 1 hour (G0/G1 transition) or 8 hours (mid-G1 progression). The absence of N-Ras caused significantly higher changes than the absence of H-Ras in the wave of transcriptional activation linked to G0/G1 transition. In contrast, the absence of H-Ras affected the profile of the transcriptional wave detected during G1 progression more strongly than did the absence of N-Ras. H-Ras was predominantly functionally associated with growth and proliferation, whereas N-Ras had a closer link to the regulation of development, the cell cycle, immunomodulation and apoptosis. Mechanistic analysis indicated that extracellular signal-regulated kinase (ERK)-dependent activation of signal transducer and activator of transcription 1 (Stat1) mediates the regulatory effect of N-Ras on defense and immunity, whereas the pro-apoptotic effects of N-Ras are mediated through ERK and p38 mitogen-activated protein kinase signaling. Conclusions Our observations confirm the notion of an absolute requirement for different peaks of Ras activity during the initial stages of the cell cycle and document the functional specificity of H-Ras and N-Ras during those processes. PMID:19895680

  12. Functional modular contact lens

    NASA Astrophysics Data System (ADS)

    Shum, Angela J.; Cowan, Melissa; Lähdesmäki, Ilkka; Lingley, Andrew; Otis, Brian; Parviz, Babak A.

    2009-08-01

    Tear fluid offers a potential route for non-invasive sensing of physiological parameters. Utilization of this potential depends on the ability to manufacture sensors that can be placed on the surface of the eye. A contact lens makes a natural platform for such sensors, but contact lens polymers present a challenge for sensor fabrication. This paper describes a microfabrication process for constructing sensors that can be integrated into the structure of a functional contact lens in the future. To demonstrate the capabilities of the process, an amperometric glucose sensor was fabricated on a polymer substrate. The sensor consists of platinum working and counter electrodes, as well as a region of indium-tin oxide (ITO) for glucose oxidase immobilization. An external silver-silver chloride electrode was used as the reference electrode during the characterization experiments. Sensor operation was validated by hydrogen peroxide measurements in the 10- 20 μM range and glucose measurements in the 0.125-20 mM range.

  13. Analysis of the K-ras and p53 pathways in x-ray-induced lung tumors in the rat

    SciTech Connect

    Belinsky, S.A.; Middleton, S.K.; Hahn, F.F.; Nikula, K.J.; Picksley, S.M.

    1996-04-01

    The risk from exposure to low-dose radiation in conjunction with cigarette smoking has not been estimated due in part to lmited knowledge surrounding the molecular mechanisms underlying radiation-induced cancers. The purpose of this investigation was to determine the frequency for alterations in genes within the K-ras and p53 signal and cell cycle regulatory pathways, respectively, in X-ray-induced lung tumors in the F344/N rat. These tumors were examined for genetic alterations in the K-ras, c-raf-1, p53, mdm2 and cip1 genes. No K-ras mutations were detected by sequencing in 18 squamous cell carcinomas (SCCs) or 17 adenocarcinomas. However, using a K-ras codon 12 mutation selection assay, a codon 12 GGT {r_arrow} GAT mutation was detected in one SCC, suggesting that activation of the K-ras proto-oncogene is both a rare and late event. Single-strand conformation polymorphism (SSCP) analysis of the kinase-binding domain of the c-raf-1 gene did not detect any polymorphisms. Three of 18 SCCs but none of the adenocarcinomas showed p53 nuclear immunoreactivity. Single-strand conformation polymorphism analysis of exons 4-9 of the p53 gene detected only an exon 9 mutation in one SCC. Mutations were not detected in the three SCCs with immunoreactive p53 protein. No amplification of the mdm2 gene was detected; however, nuclear mdm2 immunoreactivity was present in one of the three SCCs that stained positive for the p53 protein. The complete cDNA of the rat cip1 gene comprising 810 bases was cloned and sequenced. The frequency of somatic mutations in exon 2 of the cip1 gene was determined by SSCP analysis. No alterations in electrophoretic mobility were detected. The results of this investigation indicate that alterations in the K-ras and p53 pathways do not play a major role in the genesis of X-ray-induced lung tumors in the rat. 49 refs., 5 figs.

  14. Dominant inhibition of lens placode formation in mice

    PubMed Central

    Zhang, Yan; Burgess, Daniel; Overbeek, Paul A.; Govindarajan, Venkatesh

    2008-01-01

    The lens in the vertebrate eye has been shown to be critical for proper differentiation of the surrounding ocular tissues including the cornea, iris and ciliary body. In mice, previous investigators have assayed the consequences of molecular ablation of the lens. However, in these studies, lens ablation was initiated (and completed) after the cornea, retina, iris and ciliary body had initiated their differentiation programs thereby precluding analysis of the early role of the lens in fate determination of these tissues. In the present study, we have ablated the lens precursor cells of the surface ectoderm by generation of transgenic mice that express an attenuated version of diphtheria toxin (Tox176) linked to a modified Pax6 promoter that is active in the lens ectodermal precursors. In these mice, lens precursor cells fail to express Sox2, Prox1 and αA-crystallin and die before the formation of a lens placode. The Tox176 mice also showed profound alterations in the corneal differentiation program. The corneal epithelium displayed histological features of the skin, and expressed markers of skin differentiation such as Keratin 1 and 10 instead of Keratin 12, a marker of corneal epithelial differentiation. In the Tox176 mice, in the absence of the lens, extensive folding of the retina was seen. However, differentiation of the major cell types in the retina including the ganglion, amacrine, bipolar and horizontal cells was not affected. Unexpectedly, ectopic placement of the retinal pigmented epithelium was seen between the folds of the retina. Initial specification of the presumptive ciliary body and iris at the anterior margins of the retina was not altered in the Tox176 mice but their subsequent differentiation was blocked. Lacrimal and Harderian glands, which are derived from the Pax6-expressing surface ectodermal precursors, also failed to differentiate. These results suggest that, in mice, specification of the retina, ciliary body and iris occurs at the very

  15. Canonical RTK-Ras-ERK signaling and related alternative pathways

    PubMed Central

    Sundaram, Meera V.

    2013-01-01

    Receptor Tyrosine Kinase (RTK)-Ras-Extracellular signal-regulated kinase (ERK) signaling pathways control many aspects of C. elegans development and behavior. Studies in C. elegans helped elucidate the basic framework of the RTK-Ras-ERK pathway and continue to provide insights into its complex regulation, its biological roles, how it elicits cell-type appropriate responses, and how it interacts with other signaling pathways to do so. C. elegans studies have also revealed biological contexts in which alternative RTK- or Ras-dependent pathways are used instead of the canonical pathway. PMID:23908058

  16. PGA1-induced apoptosis involves specific activation of H-Ras and N-Ras in cellular endomembranes.

    PubMed

    Anta, B; Pérez-Rodríguez, A; Castro, J; García-Domínguez, C A; Ibiza, S; Martínez, N; Durá, L M; Hernández, S; Gragera, T; Peña-Jiménez, D; Yunta, M; Zarich, N; Crespo, P; Serrador, J M; Santos, E; Muñoz, A; Oliva, J L; Rojas-Cabañeros, J M

    2016-01-01

    The cyclopentenone prostaglandin A1 (PGA1) is an inducer of cell death in cancer cells. However, the mechanism that initiates this cytotoxic response remains elusive. Here we report that PGA1 triggers apoptosis by a process that entails the specific activation of H- and N-Ras isoforms, leading to caspase activation. Cells without H- and N-Ras did not undergo apoptosis upon PGA1 treatment; in these cells, the cellular demise was rescued by overexpression of either H-Ras or N-Ras. Consistently, the mutant H-Ras-C118S, defective for binding PGA1, did not produce cell death. Molecular analysis revealed a key role for the RAF-MEK-ERK signaling pathway in the apoptotic process through the induction of calpain activity and caspase-12 cleavage. We propose that PGA1 evokes a specific physiological cell death program, through H- and N-Ras, but not K-Ras, activation at endomembranes. Our results highlight a novel mechanism that may be of potential interest for tumor treatment. PMID:27468687

  17. PGA1-induced apoptosis involves specific activation of H-Ras and N-Ras in cellular endomembranes

    PubMed Central

    Anta, B; Pérez-Rodríguez, A; Castro, J; García- Domínguez, C A; Ibiza, S; Martínez, N; Durá, L M; Hernández, S; Gragera, T; Peña-Jiménez, D; Yunta, M; Zarich, N; Crespo, P; Serrador, J M; Santos, E; Muñoz, A; Oliva, J L; Rojas-Cabañeros, J M

    2016-01-01

    The cyclopentenone prostaglandin A1 (PGA1) is an inducer of cell death in cancer cells. However, the mechanism that initiates this cytotoxic response remains elusive. Here we report that PGA1 triggers apoptosis by a process that entails the specific activation of H- and N-Ras isoforms, leading to caspase activation. Cells without H- and N-Ras did not undergo apoptosis upon PGA1 treatment; in these cells, the cellular demise was rescued by overexpression of either H-Ras or N-Ras. Consistently, the mutant H-Ras-C118S, defective for binding PGA1, did not produce cell death. Molecular analysis revealed a key role for the RAF-MEK-ERK signaling pathway in the apoptotic process through the induction of calpain activity and caspase-12 cleavage. We propose that PGA1 evokes a specific physiological cell death program, through H- and N-Ras, but not K-Ras, activation at endomembranes. Our results highlight a novel mechanism that may be of potential interest for tumor treatment. PMID:27468687

  18. PGA1-induced apoptosis involves specific activation of H-Ras and N-Ras in cellular endomembranes.

    PubMed

    Anta, B; Pérez-Rodríguez, A; Castro, J; García-Domínguez, C A; Ibiza, S; Martínez, N; Durá, L M; Hernández, S; Gragera, T; Peña-Jiménez, D; Yunta, M; Zarich, N; Crespo, P; Serrador, J M; Santos, E; Muñoz, A; Oliva, J L; Rojas-Cabañeros, J M

    2016-07-28

    The cyclopentenone prostaglandin A1 (PGA1) is an inducer of cell death in cancer cells. However, the mechanism that initiates this cytotoxic response remains elusive. Here we report that PGA1 triggers apoptosis by a process that entails the specific activation of H- and N-Ras isoforms, leading to caspase activation. Cells without H- and N-Ras did not undergo apoptosis upon PGA1 treatment; in these cells, the cellular demise was rescued by overexpression of either H-Ras or N-Ras. Consistently, the mutant H-Ras-C118S, defective for binding PGA1, did not produce cell death. Molecular analysis revealed a key role for the RAF-MEK-ERK signaling pathway in the apoptotic process through the induction of calpain activity and caspase-12 cleavage. We propose that PGA1 evokes a specific physiological cell death program, through H- and N-Ras, but not K-Ras, activation at endomembranes. Our results highlight a novel mechanism that may be of potential interest for tumor treatment.

  19. Dominant negative Ras attenuates pathological ventricular remodeling in pressure overload cardiac hypertrophy

    PubMed Central

    Ramos-Kuri, Manuel; Rapti, Kleopatra; Mehel, Hind; Zhang, Shihong; Dhandapany, Perundurai S.; Liang, Lifan; García-Carrancá, Alejandro; Bobe, Regis; Fischmeister, Rodolphe; Adnot, Serge; Lebeche, Djamel; Hajjar, Roger J.; Lipskaia, Larissa; Chemaly, Elie R.

    2015-01-01

    The importance of the oncogene Ras in cardiac hypertrophy is well appreciated. The hypertrophic effects of the constitutively active mutant Ras-Val12 are revealed by clinical syndromes due to the Ras mutations and experimental studies. We examined the possible anti-hypertrophic effect of Ras inhibition in vitro using rat neonatal cardiomyocytes (NRCM) and in vivo in the setting of pressure-overload left ventricular (LV) hypertrophy (POH) in rats. Ras functions were modulated via adenovirus directed gene transfer of active mutant Ras-Val12 or dominant negative mutant N17-DN-Ras (DN-Ras). Ras-Val12 expression in vitro activates NFAT resulting in pro-hypertrophic and cardio-toxic effects on NRCM beating and Z-line organization. In contrast, the DN-Ras was antihypertrophic on NRCM, inhibited NFAT and exerted cardio-protective effects attested by preserved NRCM beating and Z line structure. Additional experiments with silencing H-Ras gene strategy corroborated the antihypertrophic effects of siRNA-H-Ras on NRCM. In vivo, with the POH model, both Ras mutants were associated with similar hypertrophy two weeks after simultaneous induction of POH and Ras-mutant gene transfer. However, LV diameters were higher and LV fractional shortening lower in the Ras-Val12 group compared to control and DN-Ras. Moreover, DN-Ras reduced the cross-sectional area of cardiomyocytes in vivo, and decreased the expression of markers of pathologic cardiac hypertrophy. In isolated adult cardiomyocytes after 2 weeks of POH and Ras-mutant gene transfer, DN-Ras improved sarcomere shortening and calcium transients compared to Ras-Val12. Overall, DN-Ras promotes a more physiological form of hypertrophy, suggesting an interesting therapeutic target for pathological cardiac hypertrophy. PMID:26260012

  20. Dominant negative Ras attenuates pathological ventricular remodeling in pressure overload cardiac hypertrophy.

    PubMed

    Ramos-Kuri, Manuel; Rapti, Kleopatra; Mehel, Hind; Zhang, Shihong; Dhandapany, Perundurai S; Liang, Lifan; García-Carrancá, Alejandro; Bobe, Regis; Fischmeister, Rodolphe; Adnot, Serge; Lebeche, Djamel; Hajjar, Roger J; Lipskaia, Larissa; Chemaly, Elie R

    2015-11-01

    The importance of the oncogene Ras in cardiac hypertrophy is well appreciated. The hypertrophic effects of the constitutively active mutant Ras-Val12 are revealed by clinical syndromes due to the Ras mutations and experimental studies. We examined the possible anti-hypertrophic effect of Ras inhibition in vitro using rat neonatal cardiomyocytes (NRCM) and in vivo in the setting of pressure-overload left ventricular (LV) hypertrophy (POH) in rats. Ras functions were modulated via adenovirus directed gene transfer of active mutant Ras-Val12 or dominant negative mutant N17-DN-Ras (DN-Ras). Ras-Val12 expression in vitro activates NFAT resulting in pro-hypertrophic and cardio-toxic effects on NRCM beating and Z-line organization. In contrast, the DN-Ras was antihypertrophic on NRCM, inhibited NFAT and exerted cardio-protective effects attested by preserved NRCM beating and Z line structure. Additional experiments with silencing H-Ras gene strategy corroborated the antihypertrophic effects of siRNA-H-Ras on NRCM. In vivo, with the POH model, both Ras mutants were associated with similar hypertrophy two weeks after simultaneous induction of POH and Ras-mutant gene transfer. However, LV diameters were higher and LV fractional shortening lower in the Ras-Val12 group compared to control and DN-Ras. Moreover, DN-Ras reduced the cross-sectional area of cardiomyocytes in vivo, and decreased the expression of markers of pathologic cardiac hypertrophy. In isolated adult cardiomyocytes after 2 weeks of POH and Ras-mutant gene transfer, DN-Ras improved sarcomere shortening and calcium transients compared to Ras-Val12. Overall, DN-Ras promotes a more physiological form of hypertrophy, suggesting an interesting therapeutic target for pathological cardiac hypertrophy.

  1. Photochemical Modulation of Ras-Mediated Signal Transduction using Caged Farnesyltransferase Inhibitors: Activation via One- and Two-Photon Excitation

    PubMed Central

    Abate-Pella, Daniel; Zeliadt, Nicholette A.; Ochocki, Joshua D.; Warmka, Janel K.; Dore, Timothy M.; Blank, David A.; Wattenberg, Elizabeth V.; Distefano, Mark D.

    2012-01-01

    The creation of caged molecules involves the attachment of protecting groups to biologically active compounds such as ligands, substrates, and drugs that can be removed under specific conditions. Photoremovable caging groups are the most common due to their ability to be removed with high spatial and temporal resolution. Here, the synthesis and photochemistry of a caged inhibitor of protein farnesyltransferase, Bhc-FTI, is described. The inhibitor was caged by alkylation of a critical thiol functional group with a Bhc moiety; while Bhc is well established as a protecting group for carboxylates and phosphates, it has not been extensively used to cage sulfhydryls. The resulting caged molecule, Bhc-FTI, can be photolyzed with UV light to release the inhibitor (FTI) that prevents Ras farnesylation, Ras membrane localization and downstream signaling. Finally, it is shown that Bhc-FTI can be uncaged by two-photon excitation to produce FTI at levels sufficient to inhibit Ras localization and alter cell morphology. Given the widespread involvement of Ras proteins in signal transduction pathways, this caged inhibitor should be useful in a plethora of studies. PMID:22492666

  2. STAT3 supports experimental K-RasG12D–induced murine myeloproliferative neoplasms dependent on serine phosphorylation

    PubMed Central

    Gough, Daniel J.; Marié, Isabelle J.; Lobry, Camille; Aifantis, Iannis

    2014-01-01

    Juvenile myelomonocytic leukemia, acute myeloid leukemia (AML), and other myeloproliferative neoplasms (MPNs) are genetically heterogeneous but frequently display activating mutations in Ras GTPases and activation of signal transducer and activator of transcription 3 (STAT3). Altered STAT3 activity is observed in up to 50% of AML correlating with poor prognosis. Activated STAT proteins, classically associated with tyrosine phosphorylation, support tumor development as transcription factors, but alternative STAT functions independent of tyrosine phosphorylation have been documented, including roles for serine-phosphorylated STAT3 in mitochondria supporting transformation by oncogenic Ras. We examined requirements for STAT3 in experimental murine K-Ras–dependent hematopoietic neoplasia. We show that STAT3 is phosphorylated on S727 but not Y705 in diseased animals. Moreover, a mouse with a point mutation abrogating STAT3 S727 phosphorylation displayed delayed onset and decreased disease severity with significantly extended survival. Activated K-Ras required STAT3 for cytokine-independent growth of myeloid progenitors in vitro, and mitochondrially restricted STAT3 and STAT3-Y705F, both transcriptionally inert mutants, supported factor-independent growth. STAT3 was dispensable for growth of normal or K-Ras–mutant myeloid progenitors in response to cytokines. However, abrogation of STAT3-S727 phosphorylation impaired factor-independent malignant growth. These data document that serine-phosphorylated mitochondrial STAT3 supports neoplastic hematopoietic cell growth induced by K-Ras. PMID:25150294

  3. Modulation of the Ras/MAPK signalling pathway by the redox function of selenoproteins in Drosophila melanogaster.

    PubMed

    Morey, M; Serras, F; Baguñà, J; Hafen, E; Corominas, M

    2001-10-01

    Modulation of reactive oxygen species (ROS) plays a key role in signal transduction pathways. Selenoproteins act controlling the redox balance of the cell. We have studied how the alteration of the redox balance caused by patufet (selD(ptuf)), a null mutation in the Drosophila melanogaster selenophosphate synthetase 1 (sps1) gene, which codes for the SelD enzyme of the selenoprotein biosynthesis, affects the Ras/MAPK signalling pathway. The selD(ptuf) mutation dominantly suppresses the phenotypes in the eye and the wing caused by hyperactivation of the Ras/MAPK cassette and the activated forms of the Drosophila EGF receptor (DER) and Sevenless (Sev) receptor tyrosine kinases (RTKs), which signal in the eye and wing, respectively. No dominant interaction is observed with sensitized conditions in the Wnt, Notch, Insulin-Pi3K, and DPP signalling pathways. Our current hypothesis is that selenoproteins selectively modulate the Ras/MAPK signalling pathway through their antioxidant function. This is further supported by the fact that a selenoprotein-independent increase in ROS caused by the catalase amorphic Cat(n1) allele also reduces Ras/MAPK signalling. Here, we present the first evidence for the role of intracellular redox environment in signalling pathways in Drosophila as a whole organism.

  4. Identification and characterization of rain, a novel Ras-interacting protein with a unique subcellular localization.

    PubMed

    Mitin, Natalia Y; Ramocki, Melissa B; Zullo, Alfred J; Der, Channing J; Konieczny, Stephen F; Taparowsky, Elizabeth J

    2004-05-21

    The Ras small GTPase functions as a signaling node and is activated by extracellular stimuli. Upon activation, Ras interacts with a spectrum of functionally diverse downstream effectors and stimulates multiple cytoplasmic signaling cascades that regulate cellular proliferation, differentiation, and apoptosis. In addition to the association of Ras with the plasma membrane, recent studies have established an association of Ras with Golgi membranes. Whereas the effectors of signal transduction by activated, plasma membrane-localized Ras are well characterized, very little is known about the effectors used by Golgi-localized Ras. In this study, we report the identification of a novel Ras-interacting protein, Rain, that may serve as an effector for endomembrane-associated Ras. Rain does not share significant sequence similarity with any known mammalian proteins, but contains a Ras-associating domain that is found in RalGDS, AF-6, and other characterized Ras effectors. Rain interacts with Ras in a GTP-dependent manner in vitro and in vivo, requires an intact Ras core effector-binding domain for this interaction, and thus fits the definition of a Ras effector. Unlike other Ras effectors, however, Rain is localized to perinuclear, juxta-Golgi vesicles in intact cells and is recruited to the Golgi by activated Ras. Finally, we found that Rain cooperates with activated Raf and causes synergistic transformation of NIH3T3 cells. Taken together, these observations support a role for Rain as a novel protein that can serve as an effector of endomembrane-localized Ras.

  5. THE OPTIMAL GRAVITATIONAL LENS TELESCOPE

    SciTech Connect

    Surdej, J.; Hanot, C.; Sadibekova, T.; Delacroix, C.; Habraken, S.; Coleman, P.; Dominik, M.; Le Coroller, H.; Mawet, D.; Quintana, H.; Sluse, D.

    2010-05-15

    Given an observed gravitational lens mirage produced by a foreground deflector (cf. galaxy, quasar, cluster, ...), it is possible via numerical lens inversion to retrieve the real source image, taking full advantage of the magnifying power of the cosmic lens. This has been achieved in the past for several remarkable gravitational lens systems. Instead, we propose here to invert an observed multiply imaged source directly at the telescope using an ad hoc optical instrument which is described in the present paper. Compared to the previous method, this should allow one to detect fainter source features as well as to use such an optimal gravitational lens telescope to explore even fainter objects located behind and near the lens. Laboratory and numerical experiments illustrate this new approach.

  6. Miniature hybrid optical imaging lens

    DOEpatents

    Sitter, D.N. Jr.; Simpson, M.L.

    1997-10-21

    A miniature lens system that corrects for imaging and chromatic aberrations is disclosed, the lens system being fabricated from primarily commercially-available components. A first element at the input to a lens housing is an aperture stop. A second optical element is a refractive element with a diffractive element closely coupled to, or formed a part of, the rear surface of the refractive element. Spaced closely to the diffractive element is a baffle to limit the area of the image, and this is closely followed by a second refractive lens element to provide the final correction. The image, corrected for aberrations exits the last lens element to impinge upon a detector plane were is positioned any desired detector array. The diffractive element is fabricated according to an equation that includes, as variables, the design wavelength, the index of refraction and the radius from an optical axis of the lens system components. 2 figs.

  7. Miniature hybrid optical imaging lens

    DOEpatents

    Sitter, Jr., David N.; Simpson, Marc L.

    1997-01-01

    A miniature lens system that corrects for imaging and chromatic aberrations, the lens system being fabricated from primarily commercially-available components. A first element at the input to a lens housing is an aperture stop. A second optical element is a refractive element with a diffractive element closely coupled to, or formed a part of, the rear surface of the refractive element. Spaced closely to the diffractive element is a baffle to limit the area of the image, and this is closely followed by a second refractive lens element to provide the final correction. The image, corrected for aberrations exits the last lens element to impinge upon a detector plane were is positioned any desired detector array. The diffractive element is fabricated according to an equation that includes, as variables, the design wavelength, the index of refraction and the radius from an optical axis of the lens system components.

  8. Retinoic acid regulation by CYP26 in vertebrate lens regeneration

    PubMed Central

    Thomas, Alvin G; Henry, Jonathan J

    2014-01-01

    Xenopus laevis is among the few species that are capable of fully regenerating a lost lens de novo. This occurs upon removal of the lens, when secreted factors from the retina are permitted to reach the cornea epithelium and trigger it to form a new lens. Although many studies have investigated the retinal factors that initiate lens regeneration, relatively little is known about what factors support this process and make the cornea competent to form a lens. We presently investigate the role of Retinoic acid (RA) signaling in lens regeneration in Xenopus. RA is a highly important morphogen during vertebrate development, including the development of various eye tissues, and has been previously implicated in several regenerative processes as well. For instance, Wolffian lens regeneration in the newt requires active RA signaling. In contrast, we provide evidence here that lens regeneration in Xenopus actually depends on the attenuation of RA signaling, which is regulated by the RA-degrading enzyme CYP26. Using RTPCR we examined the expression of RA synthesis and metabolism related genes within ocular tissues. We found expression of aldh1a1, aldh1a2, and aldh1a3, as well as cyp26a1 and cyp26b1 in both normal and regenerating corneal tissue. On the other hand, cyp26c1 does not appear to be expressed in either control or regenerating corneas, but it is expressed in the lens. Additionally in the lens, we found expression of aldh1a1 and aldh1a2, but not aldh1a3. Using an inhibitor of CYP26, and separately using exogenous retinoids, as well as RA signaling inhibitors, we demonstrate that CYP26 activity is necessary for lens regeneration to occur. We also find using phosphorylated Histone H3 labeling that CYP26 antagonism reduces cell proliferation in the cornea, and using qPCR we find that exogenous retinoids alter the expression of putative corneal stem cell markers. Furthermore, the Xenopus cornea is composed of an outer layer and inner basal epithelium, as well as a

  9. High Dk piggyback contact lens system for contact lens-intolerant keratoconus patients

    PubMed Central

    Sengor, Tomris; Kurna, Sevda Aydin; Aki, Suat; Özkurt, Yelda

    2011-01-01

    Background: The aim of the study was to examine the clinical success of high Dk (oxygen permeability) piggyback contact lens (PBCL) systems for the correction of contact lens intolerant keratoconus patients. Methods: Sixteen patients (29 eyes) who were not able to wear gas-permeable rigid lenses were included in this study. Hyper Dk silicone hydrogel (oxygen transmissibility or Dk/t = 150 units) and fluorosilicone methacrylate copolymer (Dk/t = 100 units) lenses were chosen as the PBCL systems. The clinical examinations included visual acuity and corneal observation by biomicroscopy, keratometer reading, and fluorescein staining before and after fitting the PBCL system. Results: Indications for using PBCL system were: lens stabilization and comfort, improving comfort, and adding protection to the cone. Visual acuities increased significantly in all of the patients compared with spectacles (P = 0). Improvement in visual acuity compared with rigid lenses alone was recorded in 89.7% of eyes and no alteration of the visual acuity was observed in 10.3% of the eyes. Wearing time of PBCL systems for most of the patients was limited time (mean 6 months, range 3–12 months); thereafter they tolerated rigid lenses alone except for 2 patients. Conclusion: The PBCL system is a safe and effective method to provide centering and corneal protection against mechanical trauma by the rigid lenses for keratoconus patients and may increase contact lens tolerance. PMID:21468342

  10. Chemical biology tools for regulating RAS signaling complexity in space and time.

    PubMed

    van Hattum, Hilde; Waldmann, Herbert

    2014-09-18

    Rat sarcoma (RAS) family members are small GTPases that control a number of signaling pathways important for normal cellular proliferation. Therefore, it is no surprise that a significant portion of human tumors express constitutively active mutated RAS proteins, which leads to deregulation of RAS signaling pathways, resulting in pathological perturbations of cell growth and death. Although the molecular details of RAS signaling cascades are well understood, there is still a largely unmet need for small molecule probes to control RAS signaling in space and time. More broadly, given the prevalence of mutated RAS in cancer, the need to translate the insights obtained from using small molecule probes into clinically useful drugs is also significant. In this review, we introduce RAS proteins and the signaling pathways they are involved in, and discuss some of the innovative chemical biology approaches to regulate RAS signaling, which include the exploitation of newly identified binding pockets, covalent inhibitors for mutated RAS, and RAS localization impairment.

  11. Women and the RAS: 100 years of Fellowship

    NASA Astrophysics Data System (ADS)

    Bailey, Mandy

    2016-02-01

    In January 1916, the RAS elected its first women Fellows. Mandy Bailey looks back at the social and scientific circumstances of this step towards equality, introducing a year of articles celebrating the centenary.

  12. Chaperone-mediated specificity in Ras and Rap signaling.

    PubMed

    Azoulay-Alfaguter, Inbar; Strazza, Marianne; Mor, Adam

    2015-01-01

    Ras and Rap proteins are closely related small guanosine triphosphatase (GTPases) that share similar effector-binding domains but operate in a very different signaling networks; Ras has a dominant role in cell proliferation, while Rap mediates cell adhesion. Ras and Rap proteins are regulated by several shared processes such as post-translational modification, phosphorylation, activation by guanine exchange factors and inhibition by GTPase-activating proteins. Sub-cellular localization and trafficking of these proteins to and from the plasma membrane are additional important regulatory features that impact small GTPases function. Despite its importance, the trafficking mechanisms of Ras and Rap proteins are not completely understood. Chaperone proteins play a critical role in trafficking of GTPases and will be the focus of the discussion in this work. We will review several aspects of chaperone biology focusing on specificity toward particular members of the small GTPase family. Understanding this specificity should provide key insights into drug development targeting individual small GTPases.

  13. Interferon-β Signaling Contributes to Ras Transformation

    PubMed Central

    Tsai, Yu-Chen; Pestka, Sidney; Wang, Lu-Hai; Runnels, Loren W.; Wan, Shan; Lyu, Yi Lisa; Liu, Leroy F.

    2011-01-01

    Increasing evidence has pointed to activated type I interferon signaling in tumors. However, the molecular basis for such activation and its role in tumorigenesis remain unclear. In the current studies, we report that activation of type I interferon (IFN) signaling in tumor cells is primarily due to elevated secretion of the type I interferon, IFN-β. Studies in oncogene-transformed cells suggest that oncogenes such as Ras and Src can activate IFN-β signaling. Significantly, elevated IFN-β signaling in Ras-transformed mammary epithelial MCF-10A cells was shown to contribute to Ras transformation as evidenced by morphological changes, anchorage-independent growth, and migratory properties. Our results demonstrate for the first time that the type I IFN, IFN-β, contributes to Ras transformation and support the notion that oncogene-induced cytokines play important roles in oncogene transformation. PMID:21897875

  14. The RASopathies: Developmental syndromes of Ras/MAPK pathway dysregulation

    PubMed Central

    Tidyman, William E.; Rauen, Katherine A.

    2009-01-01

    The Ras/mitogen activated protein kinase (MAPK) pathway is essential in the regulation of the cell cycle, differentiation, growth and cell senescence, all of which are critical to normal development. It is therefore not surprising that its dysregulation has profound effects on development. A class of developmental disorders, the “RASopathies”, is caused by germline mutations in genes that encode protein components of the Ras/MAPK pathway. The vast majority of these mutations result in increased signal transduction down the Ras/MAPK pathway, but usually to a lesser extent than somatic mutations associated with oncogenesis. Each syndrome exhibits unique phenotypic features, however, since they all cause dysregulation of the Ras/MAPK pathway, there are numerous overlapping phenotypic features between the syndromes, including characteristic facial features, cardiac defects, cutaneous abnormalities, neurocognitive delay and a predisposition to malignancies. Here we review the clinical and underlying molecular basis for each of these syndromes. PMID:19467855

  15. Ageing of glutathione reductase in the lens.

    PubMed

    Zhang, W Z; Augusteyn, R C

    1994-07-01

    The distribution of glutathione reductase activity in concentric layers from the lens has been determined as a function of age for 16 species. Primate lenses have almost ten times the level of glutathione reductase found in other species. Comparison with the activity of hexokinase revealed that this is not due to a higher overall rate of metabolism in these lenses. By contrast, the higher activity found in bird and fish lenses reflects a higher metabolic activity in these tissues. In all species, a gradient of activity was observed with the highest specific activity in the outermost cortical fibres, decreasing to virtually no activity in the inner parts of the tissue. No alterations were found in this gradient with increasing age, other than an increase in the amount of nuclear tissue essentially devoid of activity. The maximum activity in the outer cortical fibres was the same, regardless of the age of the lens. The time taken, in different species, for the specific activity to decrease by half, was estimated from the rate of protein accumulation. This time was found to vary from a few days to several years, indicating that the decrease in activity is not due to ageing but rather, it is related to the maturation of fibre cells. These observations are discussed in terms of current concepts of lens ageing and cataract formation. PMID:7835401

  16. Dispersion-compensated Fresnel lens

    DOEpatents

    Johnson, K.C.

    1992-11-03

    A transmission grating is used to reduce chromatic aberration in a Fresnel lens, wherein the lens chromatic dispersion is offset and substantially canceled by the grating's diffraction-induced dispersion. The grating comprises a Fresnel-type pattern of microscopic facets molded directly into the lens surface. The facets would typically have a profile height of around 4[times]10[sup [minus]5] inch and a profile width of at least 10[sup [minus]3] inch. In its primary intended application, the invention would function to improve the optical performance of a Fresnel lens used to concentrate direct sunlight. 10 figs.

  17. Dispersion-compensated fresnel lens

    DOEpatents

    Johnson, Kenneth C.

    1992-01-01

    A transmission grating is used to reduce chromatic aberration in a Fresnel lens, wherein the lens chromatic dispersion is offset and substantially canceled by the grating's diffraction-induced dispersion. The grating comprises a Fresnel-type pattern of microscopic facets molded directly into the lens surface. The facets would typically have a profile height of around 4.multidot.10.sup.-5 inch and a profile width of at least 10.sup.-3 inch. In its primary intended application, the invention would function to improve the optical performance of a Fresnel lens used to concentrate direct sunlight.

  18. PRG3 induces Ras-dependent oncogenic cooperation in gliomas

    PubMed Central

    Yakubov, Eduard; Chen, Daishi; Broggini, Thomas; Sehm, Tina; Majernik, Gökce Hatipoglu; Hock, Stefan W.; Schwarz, Marc; Engelhorn, Tobias; Doerfler, Arnd; Buchfelder, Michael; Eyupoglu, Ilker Y.; Savaskan, Nicolai E.

    2016-01-01

    Malignant gliomas are one of the most devastating cancers in humans. One characteristic hallmark of malignant gliomas is their cellular heterogeneity with frequent genetic lesions and disturbed gene expression levels conferring selective growth advantage. Here, we report on the neuronal-associated growth promoting gene PRG3 executing oncogenic cooperation in gliomas. We have identified perturbed PRG3 levels in human malignant brain tumors displaying either elevated or down-regulated PRG3 levels compared to non-transformed specimens. Further, imbalanced PRG3 levels in gliomas foster Ras-driven oncogenic amplification with increased proliferation and cell migration although angiogenesis was unaffected. Hence, PRG3 interacts with RasGEF1 (RasGRF1/CDC25), undergoes Ras-induced challenges, whereas deletion of the C-terminal domain of PRG3 (PRG3ΔCT) inhibits Ras. Moreover PRG3 silencing makes gliomas resistant to Ras inhibition. In vivo disequilibrated PRG3 gliomas show aggravated proliferation, invasion, and deteriorate clinical outcome. Thus, our data show that the interference with PRG3 homeostasis amplifies oncogenic properties and foster the malignancy potential in gliomas. PMID:27058420

  19. Calmodulin modulates H-Ras mediated Raf-1 activation.

    PubMed

    Moretó, Jemina; Lladó, Anna; Vidal-Quadras, Maite; Calvo, Maria; Pol, Albert; Enrich, Carlos; Tebar, Francesc

    2008-06-01

    We have previously demonstrated that, in COS-1 cells, inhibition of calmodulin increases Ras-GTP levels although it decreases Raf-1 activity and consequently MAPK. The present study analyzes the role of calmodulin in the regulation of Raf-1. First we show, using FRET microscopy, that inhibition of Raf-1 was not a consequence of a decreased interaction between H-Ras and Raf-1. Besides, the analysis of the phosphorylation state of Raf-1 showed that calmodulin, through downstream PI3K, is essential to ensure the Ser338-Raf-1 phosphorylation, critical for Raf-1 activation. We also show that the expression of a dominant negative mutant of PI3K impairs the calmodulin-mediated Raf-1 activation; in addition, both calmodulin and PI3K inhibitors decrease phospho-Ser338 and Raf-1 activity from upstream active H-Ras (H-RasG12V) and this effect is dependent on endocytosis. Importantly, in H-Ras depleted COS-1 cells, calmodulin does not modulate MAPK activation. Altogether, the results suggest that calmodulin regulation of MAPK in COS-1 cells relies upon H-Ras control of Raf-1 activity and involves PI3K.

  20. Plk2 Raps up Ras to subdue synapses

    PubMed Central

    Lee, Kea Joo; Hoe, Hyang-Sook

    2011-01-01

    We recently identified the activity-inducible protein kinase Plk2 as a novel overseer of the balance between Ras and Rap small GTPases. Plk2 achieves a profound level of regulatory control by interacting with and phosphorylating at least four Ras and Rap guanine nucleotide exchange factors (GEFs) and GTPase activating proteins (GAPs). Combined, these actions result in synergistic suppression of Ras and hyperstimulation of Rap signaling. Perturbation of Plk2 function abolished homeostatic adaptation of synapses to enhanced activity and impaired behavioral adaptation in various learning tasks, indicating that this regulation was critical for maintaining appropriate Ras/Rap levels. These studies provide insights into the highly cooperative nature of Ras and Rap regulation in neurons. However, different GEF and GAP substrates of Plk2 also controlled specific aspects of dendritic spine morphology, illustrating the ability of individual GAPs/GEFs to assemble microdomains of Ras and Rap signaling that respond to different stimuli and couple to distinct output pathways. PMID:21776418

  1. Lens of Eye Dosimetry

    SciTech Connect

    Mallett, Michael Wesley

    2015-03-23

    An analysis of LANL occupational dose measurements was made with respect to lens of eye dose (LOE), in particular, for plutonium workers. Table 1 shows the reported LOE as a ratio of the “deep” (photon only) and “deep+neutron” dose for routine monitored workers at LANL for the past ten years. The data compares the mean and range of these values for plutonium workers* and non-routine plutonium workers. All doses were reported based on measurements with the LANL Model 8823 TLD.

  2. Physics of electrostatic lens

    NASA Astrophysics Data System (ADS)

    1981-09-01

    The purpose of this program was to study the physics of the ion-energy boosting electrostatic lens for collective ion acceleration in the Luce diode. Extensive work was done in preparation for experiments on the PI Pulserad 1150. Analytic work was done on the orbit of protons in a mass spectrometer and a copper stack for nuclear activation analysis of proton energy spectrum has been designed. Unfortunately, a parallel program which would provide the Luce diode for the collective ion acceleration experiment never materialized. As a result no experiments were actually performed on the Pulserad 1150.

  3. The 2014 IODC lens design problem: the Cinderella lens

    NASA Astrophysics Data System (ADS)

    Juergens, Richard C.

    2014-12-01

    The lens design problem for the 2014 IODC is to design a 100 mm focal length lens in which all the components of the lens can be manufactured from ten Schott N-BK7 lens blanks 100 mm in diameter x 30 mm thick. The lens is used monochromatically at 587.56 nm. The goal of the problem is to maximize the product of the entrance pupil diameter and the semi-field of view while holding the RMS wavefront error to <= 0.070 wave within the field of view. There were 45 entries from 13 different countries. Four different commercial lens design programs were used, along with six custom, in-house programs. The number of lens elements in the entries ranged from 10 to 52. The winning entry from Jon Ehrmann had 25 lens elements, and had an entrance pupil diameter of 33.9 mm and a semi-field of view of 62.5° for a merit function product of 2,119.

  4. Resistance of R-Ras knockout mice to skin tumour induction

    PubMed Central

    May, Ulrike; Prince, Stuart; Vähätupa, Maria; Laitinen, Anni M.; Nieminen, Katriina; Uusitalo-Järvinen, Hannele; Järvinen, Tero A. H.

    2015-01-01

    The R-ras gene encodes a small GTPase that is a member of the Ras family. Despite close sequence similarities, R-Ras is functionally distinct from the prototypic Ras proteins; no transformative activity and no activating mutations of R-Ras in human malignancies have been reported for it. R-Ras activity appears inhibitory towards tumour proliferation and invasion, and to promote cellular quiescence. Contrary to this, using mice with a deletion of the R-ras gene, we found that R-Ras facilitates DMBA/TPA-induced skin tumour induction. The tumours appeared in wild-type (WT) mice on average 6 weeks earlier than in R-Ras knockout (R-Ras KO) mice. WT mice developed almost 6 times more tumours than R-Ras KO mice. Despite strong R-Ras protein expression in the dermal blood vessels, no R-Ras could be detected in the epidermis from where the tumours arose. The DMBA/TPA skin tumourigenesis-model is highly dependent upon inflammation, and we found a greatly attenuated skin inflammatory response to DMBA/TPA-treatment in the R-Ras KO mice in the context of leukocyte infiltration and proinflammatory cytokine expression. Thus, these data suggest that despite its characterised role in promoting cellular quiescence, R-Ras is pro-tumourigenic in the DMBA/TPA tumour model and important for the inflammatory response to DMBA/TPA treatment. PMID:26133397

  5. K-Ras4B phosphorylation at Ser181 is inhibited by calmodulin and modulates K-Ras activity and function.

    PubMed

    Alvarez-Moya, B; López-Alcalá, C; Drosten, M; Bachs, O; Agell, N

    2010-11-01

    Fine tuning of Ras activity is widely known as a mechanism to induce different cellular responses. Recently, we have shown that calmodulin (CaM) binds to K-Ras and that K-Ras phosphorylation inhibits its interaction with CaM. In this study we report that CaM inhibits K-Ras phosphorylation at Ser181 by protein kinase C (PKC) in vivo, and this is a mechanism to modulate K-Ras activity and signaling. Although CaM inhibition increased the activation of endogenous K-Ras, PKC inhibition decreased its activation status. We demonstrate that K-Ras phosphorylation decreased susceptibility to p120GAP activity. Accordingly, we also observed that non-phosphorylable K-Ras mutant exhibits a less sustained activation profile and do not efficiently activate AKT at low growth factor doses compared with wild-type K-Ras. It is interesting that the physiological responses induced by K-Ras are affected by this phosphorylation; when K-Ras cannot be phosphorylated it exhibits a remarkably decreased ability to stimulate proliferation in non-saturated serum conditions. Finally, we demonstrate that phosphorylation also regulates oncogenic K-Ras functions, as focus formation capacity, mobility and apoptosis resistance upon adriamycin treatment of cells expressing oncogenic K-Ras that cannot be phosphorylated are highly compromised. Moreover, at low serum concentration proliferation and survival is practically inhibited when cells cannot phosphorylate oncogenic K-Ras. In this condition, K-Ras phosphorylation is essential to ensure a proper activation of mitogen-activated protein kinase and PI3K/AKT pathways. In summary, our findings suggest that the interplay between CaM interaction and PKC phosphorylation is essential to regulate non-oncogenic and oncogenic K-Ras activity and functionality.

  6. Human iPS Cell-Derived Neurons Uncover the Impact of Increased Ras Signaling in Costello Syndrome

    PubMed Central

    Rooney, Gemma E.; Goodwin, Alice F.; Depeille, Philippe; Sharir, Amnon; Schofield, Claude M.; Yeh, Erika; Roose, Jeroen P.; Klein, Ophir D.; Rauen, Katherine A.; Weiss, Lauren A.

    2016-01-01

    Increasing evidence implicates abnormal Ras signaling as a major contributor in neurodevelopmental disorders, yet how such signaling causes cortical pathogenesis is unknown. We examined the consequences of aberrant Ras signaling in the developing mouse brain and uncovered several critical phenotypes, including increased production of cortical neurons and morphological deficits. To determine whether these phenotypes are recapitulated in humans, we generated induced pluripotent stem (iPS) cell lines from patients with Costello syndrome (CS), a developmental disorder caused by abnormal Ras signaling and characterized by neurodevelopmental abnormalities, such as cognitive impairment and autism. Directed differentiation toward a neuroectodermal fate revealed an extended progenitor phase and subsequent increased production of cortical neurons. Morphological analysis of mature neurons revealed significantly altered neurite length and soma size in CS patients. This study demonstrates the synergy between mouse and human models and validates the use of iPS cells as a platform to study the underlying cellular pathologies resulting from signaling deficits. SIGNIFICANCE STATEMENT Increasing evidence implicates Ras signaling dysfunction as a major contributor in psychiatric and neurodevelopmental disorders, such as cognitive impairment and autism, but the underlying cortical cellular pathogenesis remains unclear. This study is the first to reveal human neuronal pathogenesis resulting from abnormal Ras signaling and provides insights into how these phenotypic abnormalities likely contribute to neurodevelopmental disorders. We also demonstrate the synergy between mouse and human models, thereby validating the use of iPS cells as a platform to study underlying cellular pathologies resulting from signaling deficits. Recapitulating human cellular pathologies in vitro facilitates the future high throughput screening of potential therapeutic agents that may reverse phenotypic and

  7. KLF4 regulates adult lung tumor-initiating cells and represses K-Ras-mediated lung cancer.

    PubMed

    Yu, T; Chen, X; Zhang, W; Liu, J; Avdiushko, R; Napier, D L; Liu, A X; Neltner, J M; Wang, C; Cohen, D; Liu, C

    2016-02-01

    Lung cancer is the leading cause of cancer-related mortality in both men and women worldwide. To identify novel factors that contribute to lung cancer pathogenesis, we analyzed a lung cancer database from The Cancer Genome Atlas and found that Krüppel-like Factor 4 (KLF4) expression is significantly lower in patients' lung cancer tissue than in normal lung tissue. In addition, we identified seven missense mutations in the KLF4 gene. KLF4 is a transcription factor that regulates cell proliferation and differentiation as well as the self-renewal of stem cells. To understand the role of KLF4 in the lung, we generated a tamoxifen-induced Klf4 knockout mouse model. We found that KLF4 inhibits lung cancer cell growth and that depletion of Klf4 altered the differentiation pattern in the developing lung. To understand how KLF4 functions during lung tumorigenesis, we generated the K-ras(LSL-G12D/+);Klf4(fl/fl) mouse model, and we used adenovirus-expressed Cre to induce K-ras activation and Klf4 depletion in the lung. Although Klf4 deletion alone or K-ras mutation alone can trigger lung tumor formation, Klf4 deletion combined with K-ras mutation significantly enhanced lung tumor formation. We also found that Klf4 deletion in conjunction with K-ras activation caused lung inflammation. To understand the mechanism whereby KLF4 is regulated during lung tumorigenesis, we analyzed KLF4 promoter methylation and the profiles of epigenetic factors. We found that Class I histone deacetylases (HDACs) are overexpressed in lung cancer and that HDAC inhibitors induced expression of KLF4 and inhibited proliferation of lung cancer cells, suggesting that KLF4 is probably repressed by histone acetylation and that HDACs are valuable drug targets for lung cancer treatment.

  8. Electrostatic Focusing Lens

    NASA Astrophysics Data System (ADS)

    Thomas, Eric; Hopkins, Demitri

    2011-10-01

    We developed an electrostatic focusing lens capable of generating DD reactions, by focusing deuterium ions generated from a pointed emitter at a frozen heavy water target. Due to difficulty with the pointed emitter, we later switched to a hollow cathode design. To model the lenses, chamber, and calculate the dimensions for the design that would maximize ion energy and density, the program SIMION was used. During stable operation, vacuum was hand adjusted around 10-13 mTorr. To keep stable beam, DC voltage generator was varied between 15-25 kV. Hand adjusting was necessary, because at points in the operation the frozen heavy water would release vapor at an increased rate. This caused the pressure to rise and the beam current to spike, creating instabilities and an arc to the lens. Three methods were used to determine successful DD production. (1) Two differently shielded Geiger counters (unshielded and UHMW-PE insulated tube), (2) Spectrophotometer comparing control peaks with heavy water tests, and (3) a calibrated bubble dosimeter specific to neutrons. Analysis of the results suggest the neutrons flux varied from 532 to 1.4 × 106 neutrons/sec, and require further tests to plot and narrow results.

  9. Delineation of the Roles Played by RasG and RasC in cAMP-dependent Signal Transduction during the Early Development of Dictyostelium discoideum

    PubMed Central

    Bolourani, Parvin; Spiegelman, George B.

    2006-01-01

    On starvation, the cellular slime mold Dictyostelium discoideum initiates a program of development leading to formation of multicellular structures. The initial cell aggregation requires chemotaxis to cyclic AMP (cAMP) and relay of the cAMP signal by the activation of adenylyl cyclase (ACA), and it has been shown previously that the Ras protein RasC is involved in both processes. Insertional inactivation of the rasG gene resulted in delayed aggregation and a partial inhibition of early gene expression, suggesting that RasG also has a role in early development. Both chemotaxis and ACA activation were reduced in the rasG− cells, but the effect on chemotaxis was more pronounced. When the responses of rasG− cells to cAMP were compared with the responses of rasC− and rasC−rasG− strains, generated in otherwise isogenic backgrounds, these studies revealed that signal transduction through RasG is more important in chemotaxis and early gene expression, but that signal transduction through RasC is more important in ACA activation. Because the loss of either of the two Ras proteins alone did not result in a total loss of signal output down either of the branches of the cAMP signal-response pathway, there appears to be some overlap of function. PMID:16885420

  10. The Significance of Ras Activity in Pancreatic Cancer Initiation

    PubMed Central

    Logsdon, Craig D.; Lu, Weiqin

    2016-01-01

    The genetic landscape of pancreatic cancer shows nearly ubiquitous mutations of K-RAS. However, oncogenic K-Rasmt alone is not sufficient to lead to pancreatic ductal adenocarcinoma (PDAC) in either human or in genetically modified adult mouse models. Many stimulants, such as high fat diet, CCK, LPS, PGE2 and others, have physiological effects at low concentrations that are mediated in part through modest increases in K-Ras activity. However, at high concentrations, they induce inflammation that, in the presence of oncogenic K-Ras expression, substantially accelerates PDAC formation. The mechanism involves increased activity of oncogenic K-Rasmt. Unlike what has been proposed in the standard paradigm for the role of Ras in oncogenesis, oncogenic K-Rasmt is now known to not be constitutively active. Rather, it can be activated by standard mechanisms similar to wild-type K-Ras, but its activity is sustained for a prolonged period. Furthermore, if the level of K-Ras activity exceeds a threshold at which it begins to generate its own activators, then a feed-forward loop is formed between K-Ras activity and inflammation and pathological processes including oncogenesis are initiated. Oncogenic K-Rasmt activation, a key event in PDAC initiation and development, is subject to complex regulatory mechanisms. Reagents which inhibit inflammation, such as the Cox2 inhibitor celecoxib, block the feed-forward loop and prevent induction of PDAC in models with endogenous oncogenic K-Rasmt. Increased understanding of the role of activating and inhibitory mechanisms on oncogenic K-Rasmt activity is of paramount importance for the development of preventive and therapeutic strategies to fight against this lethal disease. PMID:26929740

  11. H-ras, but not N-ras, induces an invasive phenotype in human breast epithelial cells: a role for MMP-2 in the H-ras-induced invasive phenotype.

    PubMed

    Moon, A; Kim, M S; Kim, T G; Kim, S H; Kim, H E; Chen, Y Q; Kim, H R

    2000-01-15

    Elevated p21ras expression is associated with tumor aggressiveness in breast cancer including the extent of invasion into fat tissues, infiltration into lymphatic vessels and tumor recurrence. In the present study, we have examined the roles of H-ras and N-ras, members of the human ras gene family, in the pathogenesis of breast cancer. We show that H-ras, but not N-ras, induces an invasive phenotype in human breast epithelial cells (MCF10A) as determined by the Matrigel invasion assay, whereas both H-ras and N-ras induce anchorage-independent growth, as shown by soft agar assay. We examined the effects of H-ras and N-ras activation on the expression of MMP-2 and MMP-9, which can degrade type IV collagen, the major structural collagen of the basement membrane. We show that MMP-2 is efficiently induced by H-ras, whereas MMP-9 induction is more prominent in N-ras-activated MCF10A cells. We also show that H-ras-mediated invasiveness is significantly inhibited when the expression of MMP-2 is down-regulated, using an oligodeoxyribonucleotide complementary to the MMP-2 mRNA, or when MMP-2 activity is blocked by its inhibitor TIMP-2 (tissue inhibitors of matrix metalloproteinase-2). Our results show that the H-ras-induced invasive phenotype is associated more closely with the expression of MMP-2 in human breast epithelial cells, rather than the induction of MMP-9 expression, as shown previously for rat embryonic fibroblasts.

  12. [Research advances of K-ras mutation in the prognosis and targeted therapy of gastric cancer].

    PubMed

    Huang, Y; Wei, J; Liu, B R

    2016-02-01

    K-ras mutations have been described in 30% of human cancers with significantly different mutation frequencies. High K-ras mutation frequency is found in many cancers such as pancreas and lung cancers, whereas, gastric cancer has a relatively low K-ras mutation frequency. In recent years, numerous researches have focused on the K-ras mutation in gastric cancer. This review summarizes the K-ras mutation frequency in gastric cancer, the relationship of K-ras mutation with clinicopathologic features and prognosis of gastric cancer patients, targeted therapy for K-ras mutated gastric cancer, some small-molecular inhibitors of K-ras, and development of targeted therapy drugs for K-ras signaling pathway in gastric cancer.

  13. A role for RalGDS and a novel Ras effector in the Ras-mediated inhibition of skeletal myogenesis.

    PubMed

    Ramocki, M B; White, M A; Konieczny, S F; Taparowsky, E J

    1998-07-10

    Oncogenic Ras inhibits the differentiation of skeletal muscle cells through the activation of multiple downstream signaling pathways, including a Raf-dependent, mitogen-activated or extracellular signal-regulated kinase kinase/mitogen-activated protein kinase (MEK/MAPK)-independent pathway. Here we report that a non-Raf binding Ras effector-loop variant (H-Ras G12V,E37G), which retains interaction with the Ral guanine nucleotide dissociation stimulator (RalGDS), inhibits the conversion of MyoD-expressing C3H10T1/2 mouse fibroblasts to skeletal muscle. We show that H-Ras G12V,E37G, RalGDS, and the membrane-localized RalGDS CAAX protein inhibit the activity of alpha-actin-Luc, a muscle-specific reporter gene containing a necessary E-box and serum response factor (SRF) binding site, while a RalGDS protein defective for Ras interaction has no effect on alpha-actin-Luc transcription. H-Ras G12V,E37G does not activate endogenous MAPK, but does increase SRF-dependent transcription. Interestingly, RalGDS, RalGDS CAAX, and RalA G23V inhibit H-Ras G12V, E37G-induced expression of an SRF-regulated reporter gene, demonstrating that signaling through RalGDS does not duplicate the action of H-Ras G12V,E37G in this system. As additional evidence for this, we show that H-Ras G12V,E37G inhibits the expression of troponin I-Luc, an SRF-independent muscle-specific reporter gene, whereas RalGDS and RalGDS CAAX do not. Although our studies show that signaling through RalGDS can interfere with the expression of reporter genes dependent on SRF activity (including alpha-actin-Luc), our studies also provide strong evidence that an additional signaling molecule(s) activated by H-Ras G12V,E37G is required to achieve the complete inhibition of the myogenic differentiation program.

  14. The relationship between microvessel count and the expression of vascular endothelial growth factor, p53, and K-ras in non-small cell lung cancer.

    PubMed Central

    Kang, Y. H.; Kim, K. S.; Yu, Y. K.; Lim, S. C.; Kim, Y. C.; Park, K. O.

    2001-01-01

    Using immunohistochemical staining, we studied the relationship between the microvessel count (MC) and the expression of K-ras, mutant p53 protein, and vascular endothelial growth factor (VEGF) in 61 surgically resected non-small cell lung cancers (NSCLC) (42 squamous cell carcinoma, 14 adenocarcinoma, 2 large cell carcinoma, 2 adenosquamous carcinoma, and 1 mucoepidermoid carcinoma). MC of the tumors with lymph node (LN) metastasis was significantly higher than that of tumors without LN metastasis (66.1+/-23.1 vs. 33.8+/-13.1, p<0.05). VEGF was positive in 54 patients (88.5%). MC was 58.1+/-25.2 (mean+/-S.D.) in a x200 field, and it was significantly higher in VEGF(+) tumors than in VEGF(-) tumors (61.4+/-23.7 vs. 32.9+/-23.8, p<0.05). VEGF expression was higher in K-ras-positive or mutant p53-positive tumors than in negative tumors (p<0.05). MC was significantly higher in K-ras(+) tumors than in K-ras(-) tumors, although it did not differ according to the level of mutant p53 protein expression. Survival did not differ with VEGF, mutant p53, or K-ras expression, or the level of MC. In conclusion, there is a flow of molecular alterations from K-ras and p53, to VEGF expression, leading to angiogenesis and ultimately lymph node metastasis. Correlations between variables in close approximation and the lack of prognostic significance of individual molecular alterations suggest that tumorigenesis and metastasis are multifactorial processes. PMID:11511786

  15. 1α, 25-Dihydroxyvitamin D regulates hypoxia-inducible factor-1α in untransformed and Harvey-ras transfected breast epithelial cells.

    PubMed

    Jiang, Yan; Zheng, Wei; Teegarden, Dorothy

    2010-12-01

    The purpose of this study was to determine the mechanism by which 1α, 25-dihydroxyvitamin D (1,25(OH)(2)D) alters hypoxia-inducible factor-1α (HIF-1α) protein in untransformed and Harvey-ras (H-ras) oncogene transfected MCF10A breast epithelial cells. Treatment with 1,25(OH)(2)D (10nM) increased both mRNA (2.55±0.6-fold vs. vehicle, p=0.03) and protein levels (2.37±0.3-fold vs. vehicle, p<0.0001) of HIF-1α in MCF10A cells in 12h, which remained elevated at 24h. However, in H-ras transfected MCF10A cells, 1,25(OH)(2)D treatment increased HIF-1α protein level (2.08±0.38-fold vs. vehicle, p=0.05) at 12h, with no change in mRNA level and HIF-1α protein level returned to baseline after 24h. A transcription inhibitor prevented the 1,25(OH)(2)D induction of HIF-1α protein and mRNA levels in MCF10A cells, but failed to alter the induction of HIF-1α protein level in H-ras transfected MCF10A cells. On the other hand, inhibition of proteasomal degradation prevented the 1,25(OH)(2)D-induced HIF-1α protein level in H-ras transfected MCF10A but not in MCF10A cells. These results support that 1,25(OH)(2)D regulates HIF-1α protein level via transcriptional regulation in MCF10A cells in contrast to through proteosomal degradation with the presence of H-ras oncogene in MCF10A cells.

  16. Cellular and subcellular localization of Ras guanyl nucleotide-releasing protein in the rat hippocampus.

    PubMed

    Pierret, P; Vallée, A; Mechawar, N; Dower, N A; Stone, J C; Richardson, P M; Dunn, R J

    2001-01-01

    Ras guanyl nucleotide-releasing protein (RasGRP) is a recently discovered Ras guanyl nucleotide exchange factor that is expressed in selected regions of the rodent CNS, with high levels of expression in the hippocampus. Biochemical studies suggest that RasGRP can activate the Ras signal pathway in response to changes in diacylglycerol and possibly calcium. To investigate potential sites for RasGRP signaling, we have determined the cellular and subcellular localization of RasGRP protein in adult rat hippocampus, and have also examined the appearance of RasGRP mRNA and protein during hippocampal development. RasGRP immunoreactivity is predominately localized to those neurons participating in the direct cortico-hippocampo-cortical loop. In both hippocampal and entorhinal neurons, RasGRP protein appeared to be localized to both dendrites and somata, but not to axons. Electron microscopy of hippocampal pyramidal cells confirmed RasGRP immunoreactivity in neuronal cell bodies and dendrites, where it appeared to be associated with microtubules. The localization of RasGRP to dendrites suggests a role for this pathway in the regulation of dendritic function. Examination of developing hippocampal structures indicated that RasGRP mRNA and protein appear synchronously during the first 2 weeks of postnatal development as these neurons become fully mature. This result indicates that the RasGRP signal transduction pathway is not required during early hippocampal development, but is a feature of mature neurons during the later stages of development.

  17. Signal integration by lipid-mediated spatial cross talk between Ras nanoclusters.

    PubMed

    Zhou, Yong; Liang, Hong; Rodkey, Travis; Ariotti, Nicholas; Parton, Robert G; Hancock, John F

    2014-03-01

    Lipid-anchored Ras GTPases form transient, spatially segregated nanoclusters on the plasma membrane that are essential for high-fidelity signal transmission. The lipid composition of Ras nanoclusters, however, has not previously been investigated. High-resolution spatial mapping shows that different Ras nanoclusters have distinct lipid compositions, indicating that Ras proteins engage in isoform-selective lipid sorting and accounting for different signal outputs from different Ras isoforms. Phosphatidylserine is a common constituent of all Ras nanoclusters but is only an obligate structural component of K-Ras nanoclusters. Segregation of K-Ras and H-Ras into spatially and compositionally distinct lipid assemblies is exquisitely sensitive to plasma membrane phosphatidylserine levels. Phosphatidylserine spatial organization is also modified by Ras nanocluster formation. In consequence, Ras nanoclusters engage in remote lipid-mediated communication, whereby activated H-Ras disrupts the assembly and operation of spatially segregated K-Ras nanoclusters. Computational modeling and experimentation reveal that complex effects of caveolin and cortical actin on Ras nanoclustering are similarly mediated through regulation of phosphatidylserine spatiotemporal dynamics. We conclude that phosphatidylserine maintains the lateral segregation of diverse lipid-based assemblies on the plasma membrane and that lateral connectivity between spatially remote lipid assemblies offers important previously unexplored opportunities for signal integration and signal processing.

  18. Ras-mutant cancer cells display B-Raf binding to Ras that activates extracellular signal-regulated kinase and is inhibited by protein kinase A phosphorylation.

    PubMed

    Li, Yanping; Takahashi, Maho; Stork, Philip J S

    2013-09-20

    The small G protein Ras regulates proliferation through activation of the mitogen-activated protein (MAP) kinase (ERK) cascade. The first step of Ras-dependent activation of ERK signaling is Ras binding to members of the Raf family of MAP kinase kinase kinases, C-Raf and B-Raf. Recently, it has been reported that in melanoma cells harboring oncogenic Ras mutations, B-Raf does not bind to Ras and does not contribute to basal ERK activation. For other types of Ras-mutant tumors, the relative contributions of C-Raf and B-Raf are not known. We examined non-melanoma cancer cell lines containing oncogenic Ras mutations and express both C-Raf and B-Raf isoforms, including the lung cancer cell line H1299 cells. Both B-Raf and C-Raf were constitutively bound to oncogenic Ras and contributed to Ras-dependent ERK activation. Ras binding to B-Raf and C-Raf were both subject to inhibition by the cAMP-dependent protein kinase PKA. cAMP inhibited the growth of H1299 cells and Ras-dependent ERK activation via PKA. PKA inhibited the binding of Ras to both C-Raf and B-Raf through phosphorylations of C-Raf at Ser-259 and B-Raf at Ser-365, respectively. These studies demonstrate that in non-melanocytic Ras-mutant cancer cells, Ras signaling to B-Raf is a significant contributor to ERK activation and that the B-Raf pathway, like that of C-Raf, is a target for inhibition by PKA. We suggest that cAMP and hormones coupled to cAMP may prove useful in dampening the effects of oncogenic Ras in non-melanocytic cancer cells through PKA-dependent actions on B-Raf as well as C-Raf.

  19. Ras regulation of DNA-methylation and cancer

    SciTech Connect

    Patra, Samir Kumar

    2008-04-01

    Genome wide hypomethylation and regional hypermethylation of cancer cells and tissues remain a paradox, though it has received a convincing confirmation that epigenetic switching systems, including DNA-methylation represent a fundamental regulatory mechanism that has an impact on genome maintenance and gene transcription. Methylated cytosine residues of vertebrate DNA are transmitted by clonal inheritance through the strong preference of DNA methyltransferase, DNMT1, for hemimethylated-DNA. Maintenance of methylation patterns is necessary for normal development of mice, and aberrant methylation patterns are associated with many human tumours. DNMT1 interacts with many proteins during cell cycle progression, including PCNA, p53, EZH2 and HP1. Ras family of GTPases promotes cell proliferation by its oncogenic nature, which transmits signals by multiple pathways in both lipid raft dependent and independent fashion. DNA-methylation-mediated repression of DNA-repair protein O6-methylguanine DNA methyltransferase (MGMT) gene and increased rate of K-Ras mutation at codon for amino acids 12 and 13 have been correlated with a secondary role for Ras-effector homologues (RASSFs) in tumourigenesis. Lines of evidence suggest that DNA-methylation associated repression of tumour suppressors and apoptotic genes and ceaseless proliferation of tumour cells are regulated in part by Ras-signaling. Control of Ras GTPase signaling might reduce the aberrant methylation and accordingly may reduce the risk of cancer development.

  20. Oncogenic Ras influences the expression of multiple lncRNAs.

    PubMed

    Kotake, Yojiro; Naemura, Madoka; Kitagawa, Kyoko; Niida, Hiroyuki; Tsunoda, Toshiyuki; Shirasawa, Senji; Kitagawa, Masatoshi

    2016-08-01

    Recent ultrahigh-density tiling array and large-scale transcriptome analysis have revealed that large numbers of long non-coding RNAs (lncRNAs) are transcribed in mammals. Several lncRNAs have been implicated in transcriptional regulation, organization of nuclear structure, and post-transcriptional processing. However, the regulation of expression of lncRNAs is less well understood. Here, we show that the exogenous and endogenous expression of an oncogenic form of small GTPase Ras (called oncogenic Ras) decrease the expression of lncRNA ANRIL (antisense non-coding RNA in the INK4 locus), which is involved in the regulation of cellular senescence. We also show that forced expression of oncogenic Ras increases the expression of lncRNA PANDA (p21 associated ncRNA DNA damage activated), which is involved in the regulation of apoptosis. Microarray analysis demonstrated that expression of multiple lncRNAs fluctuated by forced expression of oncogenic Ras. These findings indicate that oncogenic Ras regulates the expression of a large number of lncRNAs including functional lncRNAs, such as ANRIL and PANDA.

  1. Identification of a farnesol analog as a Ras function inhibitor using both an in vivo Ras activation sensor and a phenotypic screening approach

    PubMed Central

    Srinivasan, Kamalakkannan; Subramanian, Thangaiah; Spielmann, H. Peter

    2013-01-01

    Mutations in Ras isoforms such as K-Ras, N-Ras, and H-Ras contribute to roughly 85, 15, and 1 % of human cancers, respectively. Proper membrane targeting of these Ras isoforms, a prerequisite for Ras activity, requires farnesylation or geranylgeranylation at the C-terminal CAAX box. We devised an in vivo screening strategy based on monitoring Ras activation and phenotypic physiological outputs for assaying synthetic Ras function inhibitors (RFI). Ras activity was visualized by the trans-location of RBDRaf1-GFP to activated Ras at the plasma membrane. By using this strategy, we screened one synthetic farnesyl substrate analog (AGOH) along with nine putative inhibitors and found that only m-CN-AGOH inhibited Ras activation. Phenotypic analysis of starving cells could be used to monitor polarization, motility, and the inability of these treated cells to aggregate properly during fruiting body formation. Incorporation of AGOH and m-CN-AGOH to cellular proteins was detected by western blot. These screening assays can be incorporated into a high throughput screening format using Dictyostelium discoideum and automated microscopy to determine effective RFIs. These RFI candidates can then be further tested in mammalian systems. PMID:24194124

  2. Muscarinic receptors transform NIH 3T3 cells through a Ras-dependent signalling pathway inhibited by the Ras-GTPase-activating protein SH3 domain.

    PubMed Central

    Mattingly, R R; Sorisky, A; Brann, M R; Macara, I G

    1994-01-01

    Expression of certain subtypes of human muscarinic receptors in NIH 3T3 cells provides an agonist-dependent model of cellular transformation by formation of foci in response to carbachol. Although focus formation correlates with the ability of the muscarinic receptors to activate phospholipase C, the actual mitogenic signal transduction pathway is unknown. Through cotransfection experiments and measurement of the activation state of native and epitope-tagged Ras proteins, the contributions of Ras and Ras GTPase-activating protein (Ras-GAP) to muscarinic receptor-dependent transformation were defined. Transforming muscarinic receptors were able to activate Ras, and such activation was required for transformation because focus formation was inhibited by coexpression of either Ras with a dominant-negative mutation or constructs of Ras-GAP that include the catalytic domain. Coexpression of the N-terminal region of GAP or of its isolated SH3 (Src homology 3) domain, but not its SH2 domain, was also sufficient to suppress muscarinic receptor-dependent focus formation. Point mutations at conserved residues in the Ras-GAP SH3 domain reversed its action, leading to an increase in carbachol-dependent transformation. The inhibitory effect of expression of the Ras-GAP SH3 domain occurs proximal to Ras activation and is selective for the mitogenic pathway activated by carbachol, as cellular transformation by either v-Ras or trkA/nerve growth factor is unaffected. Images PMID:7969134

  3. Imaging characteristics of ball lens

    NASA Astrophysics Data System (ADS)

    Li, Qinghui; Shao, Xiaopeng

    2014-05-01

    In most digital imaging applications, high-resolution imaging or videos are usually desired for later processing and analysis. The desire for high-resolution stems from two principal application areas: improvement of pictorial information for human interpretation, and helping representation for automatic machine preception. While the image sensors limit the spatial resolution of the image, the image details are also limited by the optical system, due to diffraction, and aberration1. Monocentric lens are an attractive option for gigapixel camera because the symmetrical design focuses light identically coming from any direction. Marks and Brady proposed a monocentric lens design imaging 40 gigapixels with an f-number of 2.5 and resolving 2 arcsec over a 120 degrees field of view2. Recently, Cossairt, Miau, and Nayer proposed a proof-of-concept gigapixel computational camera consisting of a large ball lens shared by several small planar sensors coupled with a deblurring step3. The design consists of a ball element resulting in a lens that is both inexpensive to produce and easy to align. Because the resolution of spherical lens is fundamentally limited by geometric aberrations, the imaging characteristics of the ball lens is expressed by the geometrical aberrations, in which the general equations for the primary aberration of the ball lens are given. The effect of shifting the stop position on the aberrations of a ball lens is discussed. The variation of the axial chromatic aberration with the Abbe V-number when the refraction index takes different values is analyzed. The variation of the third-order spherical aberration ,the fifth-order spherical aberration and the spherical aberration obtained directly from ray tracing with the f-number is discussed. The other imaging evaluation merits, such as the spot diagram, the modulation transfer function(MTF) and the encircled energy are also described. Most of the analysis of the ball lens is carried out using OSLO optics

  4. Pediatric genetic disorders of lens.

    PubMed

    Nihalani, Bharti R

    2014-12-01

    Pediatric genetic disorders of lens include various cataractous and non-cataractous anomalies. The purpose of this review is to help determine the genetic cause based on the lens appearance, ocular and systemic associations. Children with bilateral cataracts require a comprehensive history, ophthalmic and systemic examination to guide further genetic evaluation. With advancements in genetics, it is possible to determine the genetic mutations and assess phenotype genotype correlation in different lens disorders. The genetic diagnosis helps the families to better understand the disorder and develop realistic expectations as to the course of their child's disorder. PMID:27625879

  5. Pediatric genetic disorders of lens

    PubMed Central

    Nihalani, Bharti R.

    2014-01-01

    Pediatric genetic disorders of lens include various cataractous and non-cataractous anomalies. The purpose of this review is to help determine the genetic cause based on the lens appearance, ocular and systemic associations. Children with bilateral cataracts require a comprehensive history, ophthalmic and systemic examination to guide further genetic evaluation. With advancements in genetics, it is possible to determine the genetic mutations and assess phenotype genotype correlation in different lens disorders. The genetic diagnosis helps the families to better understand the disorder and develop realistic expectations as to the course of their child's disorder.

  6. A liquid crystal adaptive lens

    NASA Technical Reports Server (NTRS)

    Kowel, S. T.; Cleverly, D.

    1981-01-01

    Creation of an electronically controlled liquid crystal lens for use as a focusing mechanism in a multi-element lens system or as an adaptive optical element is analyzed. Varying the index of refraction is shown to be equivalent to the shaping of a solid refracting material. Basic characteristics of liquid crystals, essential for the creation of a lens, are reviewed. The required variation of index of refraction is provided by choosing appropriate electrode voltages. The configuration required for any incoming polarization is given and its theoretical performance in terms of modulation transfer function derived.

  7. Nanoceria have no genotoxic effect on human lens epithelial cells

    NASA Astrophysics Data System (ADS)

    Pierscionek, Barbara K.; Li, Yuebin; Yasseen, Akeel A.; Colhoun, Liza M.; Schachar, Ronald A.; Chen, Wei

    2010-01-01

    There are no treatments for reversing or halting cataract, a disease of the structural proteins in the eye lens, that has associations with other age-related degenerative conditions such as Alzheimer's disease. The incidence of cataract and associated conditions is increasing as the average age of the population rises. Protein folding diseases are difficult to assess in vivo as proteins and their age-related changes are assessed after extraction. Nanotechnology can be used to investigate protein changes in the intact lens as well as for a potential means of drug delivery. Nanoparticles, such as cerium oxide (CeO2) which have antioxidant properties, may even be used as a means of treating cataract directly. Prior to use in treatments, nanoparticle genotoxicity must be tested to assess the extent of any DNA or chromosomal damage. Sister chromatid exchanges were measured and DNA damage investigated using the alkaline COMET assay on cultured human lens epithelial cells, exposed to 5 and 10 µg ml-1 of CeO2 nanoparticles (nanoceria). Nanoceria at these dosages did not cause any DNA damage or significant increases in the number of sister chromatid exchanges. The absence of genotoxic effects on lens cells suggests that nanoceria, in the doses and exposures tested in this study, are not deleterious to the eye lens and have the potential for use in studying structural alterations, in developing non-surgical cataract treatments and in investigating other protein folding diseases.

  8. Lens system for a photo ion spectrometer

    DOEpatents

    Gruen, Dieter M.; Young, Charles E.; Pellin, Michael J.

    1990-01-01

    A lens system in a photo ion spectrometer for manipulating a primary ion beam and ionized atomic component. The atomic components are removed from a sample by a primary ion beam using the lens system, and the ions are extracted for analysis. The lens system further includes ionization resistant coatings for protecting the lens system.

  9. Lens system for a photo ion spectrometer

    DOEpatents

    Gruen, D.M.; Young, C.E.; Pellin, M.J.

    1990-11-27

    A lens system in a photo ion spectrometer for manipulating a primary ion beam and ionized atomic component is disclosed. The atomic components are removed from a sample by a primary ion beam using the lens system, and the ions are extracted for analysis. The lens system further includes ionization resistant coatings for protecting the lens system. 8 figs.

  10. Characterization and developmental expression of a Drosophila ras oncogene.

    PubMed Central

    Mozer, B; Marlor, R; Parkhurst, S; Corces, V

    1985-01-01

    We cloned a Drosophila melanogaster ras gene (Dmras64B) on the basis of its homology to the ras oncogen from Harvey murine sarcoma virus. This gene mapped at chromosomal position 64B on the left arm of the third chromosome. Sequencing of Dmras64B revealed extensive amino acid homology with the proteins encoded by the human and Saccharomyces cerevisiae ras genes. The coding region of the Drosophila gene is interrupted by two introns located in different positions with respect to its human counterpart. Dmras64B encodes three different RNAs (1.6, 2.1, and 2.6 kilobases long) that are constantly expressed throughout the development of the fly. Images PMID:3921827

  11. Hepatitis B virus HBx protein activates Ras-GTP complex formation and establishes a Ras, Raf, MAP kinase signaling cascade.

    PubMed Central

    Benn, J; Schneider, R J

    1994-01-01

    Hepatitis B virus produces a small (154-amino acid) transcriptional transactivating protein, HBx, which is required for viral infection and has been implicated in virus-mediated liver oncogenesis. However, the molecular mechanism for HBx activity and its possible influence on cell proliferation have remained obscure. A number of studies suggest that HBx may stimulate transcription by indirectly activating transcription factors, possibly by influencing cell signaling pathways. We now present biochemical evidence that HBx activates Ras and rapidly induces a cytoplasmic signaling cascade linking Ras, Raf, and mitogen-activated protein kinase (MAP kinase), leading to transcriptional transactivation. HBx strongly elevates levels of GTP-bound Ras, activated and phosphorylated Raf, and tyrosine-phosphorylated and activated MAP kinase. Transactivation of transcription factor AP-1 by HBx is blocked by inhibition of Ras or Raf activities but not by inhibition of Ca(2+)- and diacylglycerol-dependent protein kinase C. HBx was also found to stimulate DNA synthesis in serum-starved cells. The hepatitis B virus HBx protein therefore stimulates Ras-GTP complex formation and promotes downstream signaling through Raf and MAP kinases, and may influence cell proliferation. Images PMID:7937954

  12. Inactivation of RASA1 promotes melanoma tumorigenesis via R-Ras activation

    PubMed Central

    Sung, Hyeran; Kanchi, Krishna L.; Wang, Xue; Hill, Kristen S.; Messina, Jane L.; Lee, Ji-Hyun; Kim, Youngchul; Dees, Nathan D.; Ding, Li; Teer, Jamie K.; Yang, Shengyu; Sarnaik, Amod A.; Sondak, Vernon K.; Mulé, James J.; Wilson, Richard K.; Weber, Jeffrey S.; Kim, Minjung

    2016-01-01

    Inactivation of Ras GTPase activating proteins (RasGAPs) can activate Ras, increasing the risk for tumor development. Utilizing a melanoma whole genome sequencing (WGS) data from 13 patients, we identified two novel, clustered somatic missense mutations (Y472H and L481F) in RASA1 (RAS p21 protein activator 1, also called p120RasGAP). We have shown that wild type RASA1, but not identified mutants, suppresses soft agar colony formation and tumor growth of BRAF mutated melanoma cell lines via its RasGAP activity toward R-Ras (related RAS viral (r-ras) oncogene homolog) isoform. Moreover, R-Ras increased and RASA1 suppressed Ral-A activation among Ras downstream effectors. In addition to mutations, loss of RASA1 expression was frequently observed in metastatic melanoma samples on melanoma tissue microarray (TMA) and a low level of RASA1 mRNA expression was associated with decreased overall survival in melanoma patients with BRAF mutations. Thus, these data support that RASA1 is inactivated by mutation or by suppressed expression in melanoma and that RASA1 plays a tumor suppressive role by inhibiting R-Ras, a previously less appreciated member of the Ras small GTPases. PMID:26993606

  13. Galectin-1 dimers can scaffold Raf-effectors to increase H-ras nanoclustering

    PubMed Central

    Blaževitš, Olga; Mideksa, Yonatan G.; Šolman, Maja; Ligabue, Alessio; Ariotti, Nicholas; Nakhaeizadeh, Hossein; Fansa, Eyad K.; Papageorgiou, Anastassios C.; Wittinghofer, Alfred; Ahmadian, Mohammad R.; Abankwa, Daniel

    2016-01-01

    Galectin-1 (Gal-1) dimers crosslink carbohydrates on cell surface receptors. Carbohydrate-derived inhibitors have been developed for cancer treatment. Intracellularly, Gal-1 was suggested to interact with the farnesylated C-terminus of Ras thus specifically stabilizing GTP-H-ras nanoscale signalling hubs in the membrane, termed nanoclusters. The latter activity may present an alternative mechanism for how overexpressed Gal-1 stimulates tumourigenesis. Here we revise the current model for the interaction of Gal-1 with H-ras. We show that it indirectly forms a complex with GTP-H-ras via a high-affinity interaction with the Ras binding domain (RBD) of Ras effectors. A computationally generated model of the Gal-1/C-Raf-RBD complex is validated by mutational analysis. Both cellular FRET as well as proximity ligation assay experiments confirm interaction of Gal-1 with Raf proteins in mammalian cells. Consistently, interference with H-rasG12V-effector interactions basically abolishes H-ras nanoclustering. In addition, an intact dimer interface of Gal-1 is required for it to positively regulate H-rasG12V nanoclustering, but negatively K-rasG12V nanoclustering. Our findings suggest stacked dimers of H-ras, Raf and Gal-1 as building blocks of GTP-H-ras-nanocluster at high Gal-1 levels. Based on our results the Gal-1/effector interface represents a potential drug target site in diseases with aberrant Ras signalling. PMID:27087647

  14. Lens design based on lens form parameters using Gaussian brackets

    NASA Astrophysics Data System (ADS)

    Yuan, Xiangyu; Cheng, Xuemin

    2014-11-01

    The optical power distribution and the symmetry of the lens components are two important attributes that decide the ultimate lens performance and characteristics. Lens form parameters W and S are the key criteria describing the two attributes mentioned above. Lens components with smaller W and S will have a good nature of aberration balance and perform well in providing good image quality. Applying the Gaussian brackets, the two lens form parameters and the Seidel Aberration Coefficients are reconstructed. An initial lens structure can be analytically described by simultaneous equations of Seidel Aberration Coefficients and third-order aberration theory. Adding the constraints of parameters W and S in the solving process, a solution with a proper image quality and aberration distribution is achieved. The optical properties and image quality of the system based on the parameters W and S are also analyzed in this article. In the method, the aberration distribution can be controlled to some extent in the beginning of design, so that we can reduce some workload of optimization later.

  15. Survey of intraocular lens material and design.

    PubMed

    Doan, Kim T; Olson, Randall J; Mamalis, Nick

    2002-02-01

    Modern cataract surgery is constantly evolving and improving in terms of lens material and design. Researchers and physicians strive to obtain better refractive correction with smaller wound size and minimizing host cell response to limit the proliferation of lens epithelial cells leading to opacification of the lens capsule. Intraocular lens material varies in water content, refractive index, and tensile strength. Intraocular lens design has undergone revisions to prohibit lens epithelial cell migration and reflection of internal and external light. The evolution of intraocular lens and extracapsular cataract surgery has lead to faster postoperative recovery and better visual outcomes.

  16. Ras-related TC21 is activated by mutation in a breast cancer cell line, but infrequently in breast carcinomas in vivo.

    PubMed Central

    Barker, K. T.; Crompton, M. R.

    1998-01-01

    Activating ras mutations are found in many types of human tumour. Mutations in Harvey (H-), Kirsten (K-) and neuronal (N-) ras are, however, rarely found in breast carcinomas. TC21 is a ras family member that shares close homology to H-, K- and N-ras, and activating mutations have been found in ovarian carcinoma and leiomyosarcoma cell lines. We have examined panels of cDNAs from breast, ovarian and cervical cell lines, and primary and metastatic breast tumours for mutations in TC21 using a single-strand conformational polymorphism (SSCP)-based assay. One breast cancer cell line, CAL51, exhibited an altered SSCP pattern, compared with normal tissue, which was due to an A-T base change in codon 72, causing a predicted Gln-Leu activating mutation. Of nine primary and 15 metastatic breast tumour cDNAs analysed, none exhibited an altered pattern by SSCP. The apparently wild-type pattern by SSCP analysis was confirmed by sequence analysis of some of the cDNAs assayed. Thus, we conclude that mutations in TC21 are uncommon in breast carcinomas. Images Figure 1 Figure 2 Figure 3 PMID:9703274

  17. Detection of Ki-ras mutations in tissue and plasma samples of patients with pancreatic cancer using PNA-mediated PCR clamping and hybridisation probes

    PubMed Central

    Däbritz, J; Hänfler, J; Preston, R; Stieler, J; Oettle, H

    2005-01-01

    In the present study, we combined the PCR-clamping approach with melting curve analysis using mutant specific hybridisation probes and wild-type specific peptide nucleic acids (PNAs) to determine the genotypes of the most frequent point mutation in codon 12 of the proto-oncogene Ki-ras in tissue and plasma samples of patients with pancreatic cancer. The sensitivity of our assay was 1–5 × 10−5. The melting curve analysis of tissue samples of four patients revealed two valine mutations, one none-valine mutation and one wild-type sequence. Ki-ras alterations were found in 28% of DNAs (18 out of 64) of nonrelated plasma samples of 10 patients with ductal adenocarcinoma of the pancreas. The valine mutation was the predominantly detected gene alteration (83%). Out of ten patients investigated, four patients (40%) became positive during clinical observation with respect to Ki-ras mutation. All four patients exhibited progressive disease and high levels of tumour marker CA 19-9. In conclusion, the one-step procedure discribed may be a useful clinical tool for analysing Ki-ras point mutations in tissue and plasmas samples. In addition, this method can be adapted for simultanous detection of multiple mutations and quantitation. PMID:15655549

  18. Connexin23 deletion does not affect lens transparency.

    PubMed

    Berthoud, Viviana M; Minogue, Peter J; Snabb, Joseph I; Dzhashiashvili, Yulia; Novak, Layne A; Zoltoski, Rebecca K; Popko, Brian; Beyer, Eric C

    2016-05-01

    While connexin46 (Cx46) and connexin50 (Cx50) are crucial for maintaining lens transparency and growth, the contributions of a more recently identified lens fiber connexin, Cx23, are poorly understood. Therefore, we studied the consequences of absence of Cx23 in mouse lenses. Cx23-null mice were generated by homologous Cre recombination. Cx23 mRNA was abundantly expressed in wild type lenses, but not in Cx23-null lenses. The transparency and refractive properties of Cx23-null lenses were similar to wild type lenses when examined by darkfield microscopy. Neither the focusing ability nor the light scattering was altered in the Cx23-null lenses. While both Cx46 and Cx50 localized to appositional fiber cell membranes (as in wild type lenses), their levels were consistently (but not significantly) decreased in homozygous Cx23-null lenses. These results suggest that although Cx23 expression can influence the abundance of the co-expressed lens fiber connexins, heterozygous or homozygous expression of a Cx23-null allele does not alter lens transparency.

  19. The Tumor Suppressor DiRas3 Forms a Complex with H-Ras and C-RAF Proteins and Regulates Localization, Dimerization, and Kinase Activity of C-RAF*

    PubMed Central

    Baljuls, Angela; Beck, Matthias; Oenel, Ayla; Robubi, Armin; Kroschewski, Ruth; Hekman, Mirko; Rudel, Thomas; Rapp, Ulf R.

    2012-01-01

    The maternally imprinted Ras-related tumor suppressor gene DiRas3 is lost or down-regulated in more than 60% of ovarian and breast cancers. The anti-tumorigenic effect of DiRas3 is achieved through several mechanisms, including inhibition of cell proliferation, motility, and invasion, as well as induction of apoptosis and autophagy. Re-expression of DiRas3 in cancer cells interferes with the signaling through Ras/MAPK and PI3K. Despite intensive research, the mode of interference of DiRas3 with the Ras/RAF/MEK/ERK signal transduction is still a matter of speculation. In this study, we show that DiRas3 associates with the H-Ras oncogene and that activation of H-Ras enforces this interaction. Furthermore, while associated with DiRas3, H-Ras is able to bind to its effector protein C-RAF. The resulting multimeric complex consisting of DiRas3, C-RAF, and active H-Ras is more stable than the two protein complexes H-Ras·C-RAF or H-Ras·DiRas3, respectively. The consequence of this complex formation is a DiRas3-mediated recruitment and anchorage of C-RAF to components of the membrane skeleton, suppression of C-RAF/B-RAF heterodimerization, and inhibition of C-RAF kinase activity. PMID:22605333

  20. Contact lens management of keratoconus.

    PubMed

    Downie, Laura E; Lindsay, Richard G

    2015-07-01

    Contact lenses are the primary form of visual correction for patients with keratoconus. Contemporary advances in contact lens designs and materials have significantly expanded the available fitting options for patients with corneal ectasia. Furthermore, imaging technology, such as corneal topography and anterior segment optical coherence tomography, can be applied to both gain insight into corneal microstructural changes and to guide contact lens fitting. This paper provides a comprehensive review of the range of contact lens modalities, including soft lenses, hybrid designs, rigid lenses, piggyback configurations, corneo-scleral, mini-scleral and scleral lenses that are currently available for the optical management of keratoconus. The review also discusses the importance of monitoring for disease progression in patients with keratoconus, in particular children, who tend to undergo more rapid progressive changes, so as to facilitate appropriate modification to contact lens fitting and/or potential referral for corneal collagen cross-linking treatment, as appropriate. PMID:26104589

  1. Intraocular lens fixation with dacron.

    PubMed

    Peyman, G A; Koziol, J E

    1978-10-01

    To overcome the problem of postoperative lens dislocation, we evaluated a new means of lens fixation. Our experimental studies in rabbits and primates demonstrated that Dacron polyethylene terephtalate induced a cellular reaction from either the anterior or posterior iris surface when placed in contact with the iris, thereby establishing a bond between the Dacron fibers and the iris. Dacron mesh can be attached to the distal portion of either the anterior or posterior loops of a Binkhorst iris clip (4-loop) lens. In the rabbit eye, lens fixation occurred within five days; in the primate eye, 30 days. When combined with silk, Dacron produced tissue ingrowth in the primate eye within 14 days. No unwanted reaction occurred in any animal with the Dacron and silk combination. Being biodegradable, the silk induced faster cellular ingrowth than the Dacron. However, Dacron, which is not biodegradable, provided a permanent means of fixation. PMID:155053

  2. Genetic polymorphisms of the RAS-cytokine pathway and chronic kidney disease

    PubMed Central

    Kanetsky, Peter; Raj, Dominic

    2008-01-01

    Chronic kidney disease (CKD) in children is irreversible. It is associated with renal failure progression and atherosclerotic cardiovascular (CV) abnormalities. Nearly 60% of children with CKD are affected since birth with congenital or inherited kidney disorders. Preliminary evidence primarily from adult CKD studies indicates common genetic risk factors for CKD and atherosclerotic CV disease. Although multiple physiologic pathways share common genes for CKD and CV disease, substantial evidence supports our attention to the renin angiotensin system (RAS) and the interlinked inflammatory cascade because they modulate the progressions of renal and CV disease. Gene polymorphisms in the RAS-cytokine pathway, through altered gene expression of inflammatory cytokines, are potential factors that modulate the rate of CKD progression and CV abnormalities in patients with CKD. For studying such hypotheses, the cooperative efforts among scientific groups and the availability of robust and affordable technologies to genotype thousands of single nucleotide polymorphisms (SNPs) across the genome make genome-wide association studies an attractive paradigm for studying polygenic diseases such as CKD. Although attractive, such studies should be interpreted carefully, with a fundamental understanding of their potential weaknesses. Nevertheless, whole-genome association studies for diabetic nephropathy and future studies pertaining to other types of CKD will offer further insight for the development of targeted interventions to treat CKD and associated atherosclerotic CV abnormalities in the pediatric CKD population. PMID:18481112

  3. Single lens laser beam shaper

    DOEpatents

    Liu, Chuyu; Zhang, Shukui

    2011-10-04

    A single lens bullet-shaped laser beam shaper capable of redistributing an arbitrary beam profile into any desired output profile comprising a unitary lens comprising: a convex front input surface defining a focal point and a flat output portion at the focal point; and b) a cylindrical core portion having a flat input surface coincident with the flat output portion of the first input portion at the focal point and a convex rear output surface remote from the convex front input surface.

  4. Notch-1 expression levels in 3T3-L1 cells influence ras signaling and transformation by oncogenic ras.

    PubMed

    Ruiz-Hidalgo, M J; Garcés, C; Laborda, J

    1999-04-01

    Notch proteins participate in interactions between several cell types involved on the specification of numerous cell fates during development. We previously showed that enforced downregulation of Notch-1 expression prevented adipogenesis of 3T3-L1 cells. Since adipogenesis of 3T3-L1 cells can be induced by oncogenic ras, we studied whether this was also the case in 3T3-L1 cells with decreased levels of Notch-1 expression. We found that oncogenic ras induces transformation and not differentiation of 3T3-L1 cells with diminished levels of Notch-1. This result suggests that Notch-1 is implicated in the interpretation of signals leading to activation of p21 Ras.

  5. Lens specific RLIP76 transgenic mice show a phenotype similar to microphthalmia.

    PubMed

    Sahu, Mukesh; Sharma, Rajendra; Yadav, Sushma; Wakamiya, Maki; Chaudhary, Pankaj; Awasthi, Sanjay; Awasthi, Yogesh C

    2014-01-01

    RALBP1/RLIP76 is a ubiquitously expressed protein, involved in promotion and regulation of functions initiated by Ral and R-Ras small GTPases. Presence of multiple domains in its structure enables RLIP76 to be involved in a number of physiological processes such as endocytosis, exocytosis, mitochondrial fission, actin cytoskeleton remodeling, and transport of exogenous and endogenous toxicants. Previously, we have established that RLIP76 provides protection to ocular tissues against oxidative stress by transporting the glutathione-conjugates of the toxic, electrophilic products of lipid peroxidation generated during oxidative stress. Therefore, we developed lens specific RLIP76 transgenic mice (lensRLIP76 Tg) to elucidate the role of RLIP76 in protection against oxidative stress, but these transgenic mice showed impaired lens development and a phenotype with small eyes similar to that observed in microphthalmia. These findings prompted us to investigate the mechanisms via which RLIP76 affects lens and eye development. In the present study, we report engineering of lensRLIP76 Tg mice, characterization of the associated phenotype, and the possible molecular mechanisms that lead to the impaired development of eye and lens in these mice. The results of microarray array analysis indicate that the genes involved in pathways for G-Protein signaling, actin cytoskeleton reorganization, endocytosis, and apoptosis are affected in these transgenic mice. The expression of transcription factors, Pax6, Hsf1, and Hsf4b known to be involved in lens development is down regulated in the lens of these Tg mice. However, the expression of heat shock proteins (Hsps), the downstream targets of Hsfs, is differentially affected in the lens showing down regulation of Hsp27, Hsp40, up regulation of Hsp60, and no effect on Hsp70 and Hsp90 expression. The disruption in the organization of actin cytoskeleton of these Tg mice was associated with the inhibition of the activation of Cdc42 and

  6. Rasputin, the Drosophila homologue of the RasGAP SH3 binding protein, functions in ras- and Rho-mediated signaling.

    PubMed

    Pazman, C; Mayes, C A; Fanto, M; Haynes, S R; Mlodzik, M

    2000-04-01

    The small GTPase Ras plays an important role in many cellular signaling processes. Ras activity is negatively regulated by GTPase activating proteins (GAPs). It has been proposed that RasGAP may also function as an effector of Ras activity. We have identified and characterized the Drosophila homologue of the RasGAP-binding protein G3BP encoded by rasputin (rin). rin mutants are viable and display defects in photoreceptor recruitment and ommatidial polarity in the eye. Mutations in rin/G3BP genetically interact with components of the Ras signaling pathway that function at the level of Ras and above, but not with Raf/MAPK pathway components. These interactions suggest that Rin is required as an effector in Ras signaling during eye development, supporting an effector role for RasGAP. The ommatidial polarity phenotypes of rin are similar to those of RhoA and the polarity genes, e.g. fz and dsh. Although rin/G3BP interacts genetically with RhoA, affecting both photoreceptor differentiation and polarity, it does not interact with the gain-of-function genotypes of fz and dsh. These data suggest that Rin is not a general component of polarity generation, but serves a function specific to Ras and RhoA signaling pathways.

  7. Design, synthesis and biological evaluation of sugar-derived Ras inhibitors.

    PubMed

    Peri, Francesco; Airoldi, Cristina; Colombo, Sonia; Martegani, Enzo; van Neuren, Anske Stephanie; Stein, Matthias; Marinzi, Chiara; Nicotra, Francesco

    2005-10-01

    The design and synthesis of novel Ras inhibitors with a bicyclic scaffold derived from the natural sugar D-arabinose are presented. Molecular modelling showed that these ligands can bind Ras by accommodating the aromatic moieties and the phenylhydroxylamino group in a cavity near the Switch II region of the protein. All the synthetic compounds were active in inhibiting nucleotide exchange on p21 human Ras in vitro, and two of them selectively inhibited Ras-dependent cell growth in vivo.

  8. 1,25-Dihydroxyvitamin D inhibits glutamine metabolism in Harvey-ras transformed MCF10A human breast epithelial cell.

    PubMed

    Zhou, Xuanzhu; Zheng, Wei; Nagana Gowda, G A; Raftery, Daniel; Donkin, Shawn S; Bequette, Brian; Teegarden, Dorothy

    2016-10-01

    Breast cancer is the second most common cancer among women in the US. The active form of vitamin D, 1,25-dihydroxyvitamin D (1,25(OH)2D), is proposed to inhibit cellular processes and to prevent breast cancer. The current studies investigated the effect of 1,25(OH)2D on glutamine metabolism during cancer progression employing Harvey-ras oncogene transformed MCF10A human breast epithelial cells (MCF10A-ras). Treatment with 1,25(OH)2D significantly reduced intracellular glutamine and glutamate levels measured by nuclear magnetic resonance (NMR) by 23±2% each. Further, 1,25(OH)2D treatment reduced glutamine and glutamate flux, determined by [U-(13)C5] glutamine tracer kinetics, into the TCA cycle by 31±0.2% and 17±0.4%, respectively. The relative levels of mRNA and protein abundance of the major glutamine transporter, solute linked carrier family 1 member A5 (SLC1A5), was significantly decreased by 1,25(OH)2D treatment in both MCF10A-ras cells and MCF10A which overexpress ErbB2 (HER-2/neu). Consistent with these results, glutamine uptake was reduced by 1,25(OH)2D treatment and the impact was eliminated with the SLC1A5 inhibitor L-γ-Glutamyl-p-nitroanilide (GPNA). A consensus sequence to the vitamin D responsive element (VDRE) was identified in silico in the SLC1A5 gene promoter, and site-directed mutagenesis analyses with reporter gene studies demonstrate a functional negative VDRE in the promoter of the SLC1A5 gene. siRNA-SLC1A5 transfection in MCF10A-ras cells significantly reduced SLC1A5 mRNA expression as well as decreased viable cell number similar to 1,25(OH)2D treatment. SLC1A5 knockdown also induced an increase in apoptotic cells in MCF10A-ras cells. These results suggest 1,25(OH)2D alters glutamine metabolism in MCF10A-ras cells by inhibiting glutamine uptake and utilization, in part through down-regulation of SLC1A5 transcript abundance. Thus, 1,25(OH)2D down-regulation of the glutamine transporter, SLC1A5, may facilitate vitamin D prevention of breast

  9. Regulation of Raf-1 and Raf-1 mutants by Ras-dependent and Ras-independent mechanisms in vitro.

    PubMed

    Dent, P; Reardon, D B; Morrison, D K; Sturgill, T W

    1995-08-01

    The serine/threonine kinase Raf-1 functions downstream from Ras to activate mitogen-activated protein kinase kinase, but the mechanisms of Raf-1 activation are incompletely understood. To dissect these mechanisms, wild-type and mutant Raf-1 proteins were studied in an in vitro system with purified plasma membranes from v-Ras- and v-Src-transformed cells (transformed membranes). Wild-type (His)6- and FLAG-Raf-1 were activated in a Ras- and ATP-dependent manner by transformed membranes; however, Raf-1 proteins that are kinase defective (K375M), that lack an in vivo site(s) of regulatory tyrosine (YY340/341FF) or constitutive serine (S621A) phosphorylation, that do not bind Ras (R89L), or that lack an intact zinc finger (CC165/168SS) were not. Raf-1 proteins lacking putative regulatory sites for an unidentified kinase (S259A) or protein kinase C (S499A) were activated but with apparently reduced efficiency. The kinase(s) responsible for activation by Ras or Src may reside in the plasma membrane, since GTP loading of plasma membranes from quiescent NIH 3T3 cells (parental membranes) induced de novo capacity to activate Raf-1. Wild-type Raf-1, possessing only basal activity, was not activated by parental membranes in the absence of GTP loading. In contrast, Raf-1 Y340D, possessing significant activity, was, surprisingly, stimulated by parental membranes in a Ras-independent manner. The results suggest that activation of Raf-1 by phosphorylation may be permissive for further modulation by another membrane factor, such as a lipid. A factor(s) extracted with methanol-chloroform from transformed membranes or membranes from Sf9 cells coexpressing Ras and SrcY527F significantly enhanced the activity of Raf-1 Y340D or active Raf-1 but not that of inactive Raf-1. Our findings suggest a model for activation of Raf-1, wherein (i) Raf-1 associates with Ras-GTP, (ii) Raf-1 is activated by tyrosine and/or serine phosphorylation, and (iii) Raf-1 activity is further increased by a

  10. Attenuation of TGF-β signaling suppresses premature senescence in a p21-dependent manner and promotes oncogenic Ras-mediated metastatic transformation in human mammary epithelial cells

    PubMed Central

    Lin, Shu; Yang, Junhua; Elkahloun, Abdel G.; Bandyopadhyay, Abhik; Wang, Long; Cornell, John E.; Yeh, I-Tien; Agyin, Joseph; Tomlinson, Gail; Sun, Lu-Zhe

    2012-01-01

    The molecular mechanisms that drive triple-negative, basal-like breast cancer progression are elusive. Few molecular targets have been identified for the prevention or treatment of this disease. Here we developed a series of isogenic basal-like human mammary epithelial cells (HMECs) with altered transforming growth factor-β (TGF-β) sensitivity and different malignancy, resembling a full spectrum of basal-like breast carcinogenesis, and determined the molecular mechanisms that contribute to oncogene-induced transformation of basal-like HMECs when TGF-β signaling is attenuated. We found that expression of a dominant-negative type II receptor (DNRII) of TGF-β abrogated autocrine TGF-β signaling in telomerase-immortalized HMECs and suppressed H-Ras-V12–induced senescence-like growth arrest (SLGA). Furthermore, coexpression of DNRII and H-Ras-V12 rendered HMECs highly tumorigenic and metastatic in vivo in comparison with H-Ras-V12–transformed HMECs that spontaneously escaped H-Ras-V12–induced SLGA. Microarray analysis revealed that p21 was the major player mediating Ras-induced SLGA, and attenuated or loss of p21 expression contributed to the escape from SLGA when autocrine TGF-β signaling was blocked in HMECs. Furthermore, knockdown of p21 also suppressed H-Ras-V12–induced SLGA. Our results identify that autocrine TGF-β signaling is an integral part of the cellular anti-transformation network by suppressing the expression of a host of genes, including p21-regulated genes, that mediate oncogene-induced transformation in basal-like breast cancer. PMID:22357622

  11. Fibronectin-induced proliferation in thyroid cells is mediated by alphavbeta3 integrin through Ras/Raf-1/MEK/ERK and calcium/CaMKII signals.

    PubMed

    Illario, Maddalena; Cavallo, Anna Lina; Monaco, Sara; Di Vito, Ennio; Mueller, Frank; Marzano, Luigi A; Troncone, Giancarlo; Fenzi, Gianfranco; Rossi, Guido; Vitale, Mario

    2005-05-01

    We recently demonstrated in an immortalized thyroid cell line that integrin stimulation by fibronectin (FN) simultaneously activates two signaling pathways: Ras/Raf/MAPK kinase (Mek)/Erk and calcium Ca2+/calcium calmodulin-dependent kinase II (CaMKII). Both signals are necessary to stimulate Erk phosphorylation because CaMKII modulates Ras-induced Raf-1 activity. In this study we present evidence that extends these findings to normal human thyroid cells in primary culture, demonstrating its biological significance in a more physiological cell model. In normal thyroid cells, immobilized FN-induced activation of p21Ras and Erk phosphorylation. This pathway was responsible for FN-induced cell proliferation. Concurrent increase of intracellular Ca2+ concentration and CaMKII activation was observed. Both induction of p21Ras activity and increase of intracellular Ca2+ concentration were mediated by FN binding to alphavbeta3 integrin. Inhibition of the Ca2+/CaMKII signal pathway by calmodulin or CaMKII inhibitors completely abolished the FN-induced Erk phosphorylation. Binding to FN induced Raf-1 and CaMKII to form a protein complex, indicating that intersection between Ras/Raf/Mek/Erk and Ca2+/CaMKII signaling pathways occurred at Raf-1 level. Interruption of the Ca2+/CaMKII signal pathway arrested cell proliferation induced by FN. We also analyzed thyroid tumor cell lines that displayed concomitant aberrant integrin expression and signal transduction. These data confirm that integrin activation by FN in normal thyroid cells generates Ras/Raf/Mek/Erk and Ca2+/CaMKII signaling pathways and that both are necessary to stimulate cell proliferation, whereas in thyroid tumors integrin signaling is altered.

  12. Domain contributions to signaling specificity differences between Ras-guanine nucleotide releasing factor (Ras-GRF) 1 and Ras-GRF2.

    PubMed

    Jin, Shan-Xue; Bartolome, Christopher; Arai, Junko A; Hoffman, Laurel; Uzturk, B Gizem; Kumar-Singh, Rajendra; Waxham, M Neal; Feig, Larry A

    2014-06-01

    Ras-GRF1 (GRF1) and Ras-GRF2 (GRF2) constitute a family of similar calcium sensors that regulate synaptic plasticity. They are both guanine exchange factors that contain a very similar set of functional domains, including N-terminal pleckstrin homology, coiled-coil, and calmodulin-binding IQ domains and C-terminal Dbl homology Rac-activating domains, Ras-exchange motifs, and CDC25 Ras-activating domains. Nevertheless, they regulate different forms of synaptic plasticity. Although both GRF proteins transduce calcium signals emanating from NMDA-type glutamate receptors in the CA1 region of the hippocampus, GRF1 promotes LTD, whereas GRF2 promotes θ-burst stimulation-induced LTP (TBS-LTP). GRF1 can also mediate high frequency stimulation-induced LTP (HFS-LTP) in mice over 2-months of age, which involves calcium-permeable AMPA-type glutamate receptors. To add to our understanding of how proteins with similar domains can have different functions, WT and various chimeras between GRF1 and GRF2 proteins were tested for their abilities to reconstitute defective LTP and/or LTD in the CA1 hippocampus of Grf1/Grf2 double knock-out mice. These studies revealed a critical role for the GRF2 CDC25 domain in the induction of TBS-LTP by GRF proteins. In contrast, the N-terminal pleckstrin homology and/or coiled-coil domains of GRF1 are key to the induction of HFS-LTP by GRF proteins. Finally, the IQ motif of GRF1 determines whether a GRF protein can induce LTD. Overall, these findings show that for the three forms of synaptic plasticity that are regulated by GRF proteins in the CA1 hippocampus, specificity is encoded in only one or two domains, and a different set of domains for each form of synaptic plasticity.

  13. Induction of p27Kip1 degradation and anchorage independence by Ras through the MAP kinase signaling pathway.

    PubMed

    Kawada, M; Yamagoe, S; Murakami, Y; Suzuki, K; Mizuno, S; Uehara, Y

    1997-08-01

    While most untransformed cells require substrate attachment for growth (anchorage dependence), the oncogenic transformed cells lack this requirement (anchorage independence) and are often tumorigenic. However, the mechanism of loss of anchorage dependence is not fully understood. When rat normal fibroblasts were cultured in suspension without substrate attachment, the cell cycle arrested in G1 phase and the cyclin-dependent kinase inhibitor p27Kip1 protein and its mRNA accumulated. Conditional expression of oncogenic Ras induced the G1-S transition of the cell cycle and significantly shortened the half-life of p27Kip1 protein without altering its mRNA level. Inhibition of the activation of mitogen-activated protein (MAP) kinase by cyclic AMP-elevating agents and a MEK inhibitor prevented the oncogenic Ras-induced degradation of p27Kip1. These results suggest that the loss of substrate attachment induces the cell cycle arrest through the up-regulation of p27Kip1 mRNA, but the oncogenic Ras confers anchorage independence by accelerating p27Kip1 degradation through the activation of the MAP kinase signaling pathway. Furthermore, we have found that p27Kip1 is phosphorylated by MAP kinase in vitro and the phosphorylated p27Kip1 cannot bind to and inhibit cdk2.

  14. Senescence-Associated Secretory Phenotypes Reveal Cell-Nonautonomous Functions of Oncogenic RAS and the p53 Tumor Suppressor

    SciTech Connect

    Coppé, Jean-Philippe; Patil, Christopher; Rodier, Francis; Sun, Yu; Munoz, Denise; Goldstein, Joshua; Nelson, Peter; Desprez, Pierre-Yves; Campisi, Judith

    2008-10-24

    Cellular senescence suppresses cancer by arresting cell proliferation, essentially permanently, in response to oncogenic stimuli, including genotoxic stress. We modified the use of antibody arrays to provide a quantitative assessment of factors secreted by senescent cells. We show that human cells induced to senesce by genotoxic stress secrete myriad factors associated with inflammation and malignancy. This senescence-associated secretory phenotype (SASP) developed slowly over several days and only after DNA damage of sufficient magnitude to induce senescence. Remarkably similar SASPs developed in normal fibroblasts, normal epithelial cells, and epithelial tumor cells after genotoxic stress in culture, and in epithelial tumor cells in vivo after treatment of prostate cancer patients with DNA-damaging chemotherapy. In cultured premalignant epithelial cells, SASPs induced an epithelial-mesenchyme transition and invasiveness, hallmarks of malignancy, by a paracrine mechanism that depended largely on the SASP factors interleukin (IL)-6 and IL-8. Strikingly, two manipulations markedly amplified, and accelerated development of, the SASPs: oncogenic RAS expression, which causes genotoxic stress and senescence in normal cells, and functional loss of the p53 tumor suppressor protein. Both loss of p53 and gain of oncogenic RAS also exacerbated the promalignant paracrine activities of the SASPs. Our findings define a central feature of genotoxic stress-induced senescence. Moreover, they suggest a cell-nonautonomous mechanism by which p53 can restrain, and oncogenic RAS can promote, the development of age-related cancer by altering the tissue microenvironment.

  15. Protective Effects of Acetylation on the Pathological Reactions of the Lens Crystallins with Homocysteine Thiolactone

    PubMed Central

    Moafian, Zeinab; Khoshaman, Kazem; Oryan, Ahmad; Kurganov, Boris I.; Yousefi, Reza

    2016-01-01

    Various post-translational lens crystallins modifications result in structural and functional insults, contributing to the development of lens opacity and cataract disorders. Lens crystallins are potential targets of homocysteinylation, particularly under hyperhomocysteinemia which has been indicated in various eye diseases. Since both homocysteinylation and acetylation primarily occur on protein free amino groups, we applied different spectroscopic methods and gel mobility shift analysis to examine the possible preventive role of acetylation against homocysteinylation. Lens crystallins were extensively acetylated in the presence of acetic anhydride and then subjected to homocysteinylation in the presence of homocysteine thiolactone (HCTL). Extensive acetylation of the lens crystallins results in partial structural alteration and enhancement of their stability, as well as improvement of α-crystallin chaperone-like activity. In addition, acetylation partially prevents HCTL-induced structural alteration and aggregation of lens crystallins. Also, acetylation protects against HCTL-induced loss of α-crystallin chaperone activity. Additionally, subsequent acetylation and homocysteinylation cause significant proteolytic degradation of crystallins. Therefore, further experimentation is required in order to judge effectively the preventative role of acetylation on the structural and functional insults induced by homocysteinylation of lens crystallins. PMID:27706231

  16. Nucleophosmin and Nucleolin Regulate K-Ras Plasma Membrane Interactions and MAPK Signal Transduction*

    PubMed Central

    Inder, Kerry L.; Lau, Chiyan; Loo, Dorothy; Chaudhary, Natasha; Goodall, Andrew; Martin, Sally; Jones, Alun; van der Hoeven, Dharini; Parton, Robert G.; Hill, Michelle M.; Hancock, John F.

    2009-01-01

    The spatial organization of Ras proteins into nanoclusters on the inner leaflet of the plasma membrane is essential for high fidelity signaling through the MAPK pathway. Here we identify two selective regulators of K-Ras nanoclustering from a proteomic screen for K-Ras interacting proteins. Nucleophosmin (NPM) and nucleolin are predominantly localized to the nucleolus but also have extranuclear functions. We show that a subset of NPM and nucleolin localizes to the inner leaflet of plasma membrane and forms specific complexes with K-Ras but not other Ras isoforms. Active GTP-loaded and inactive GDP-loaded K-Ras both interact with NPM, although NPM-K-Ras binding is increased by growth factor receptor activation. NPM and nucleolin both stabilize K-Ras levels on the plasma membrane, but NPM concurrently increases the clustered fraction of GTP-K-Ras. The increase in nanoclustered GTP-K-Ras in turn enhances signal gain in the MAPK pathway. In summary these results reveal novel extranucleolar functions for NPM and nucleolin as regulators of K-Ras nanocluster formation and activation of the MAPK pathway. The study also identifies a new class of K-Ras nanocluster regulator that operates independently of the structural scaffold galectin-3. PMID:19661056

  17. Targeting Bcl-2 stability to sensitize cells harboring oncogenic ras.

    PubMed

    Peng, Bo; Ganapathy, Suthakar; Shen, Ling; Huang, Junchi; Yi, Bo; Zhou, Xiaodong; Dai, Wei; Chen, Changyan

    2015-09-01

    The pro-survival factor Bcl-2 and its family members are critical determinants of the threshold of the susceptibility of cells to apoptosis. Studies are shown that cells harboring an oncogenic ras were extremely sensitive to the inhibition of protein kinase C (PKC) and Bcl-2 could antagonize this apoptotic process. However, it remains unrevealed how Bcl-2 is being regulated in this apoptotic process. In this study, we investigate the role of Bcl-2 stability in sensitizing the cells harboring oncogenic K-ras to apoptosis triggered by PKC inhibitor GO6976. We demonstrated that Bcl-2 in Swiss3T3 cells ectopically expressing or murine lung cancer LKR cells harboring K-ras rapidly underwent ubiquitin-dependent proteasome pathway after the treatment of GO6976, accompanied with induction of apoptosis. In this process, Bcl-2 formed the complex with Keap-1 and Cul3. The mutation of serine-17 and deletion of BH-2 or 4 was required for Bcl-2 ubiquitination and degradation, which elevate the signal threshold for the induction of apoptosis in the cells following PKC inhibition. Thus, Bcl-2 appears an attractive target for the induction of apoptosis by PKC inhibition in cancer cells expressing oncogenic K-ras. PMID:26041886

  18. Minireview: physiological and pathological actions of RAS in the ovary.

    PubMed

    Fan, Heng-Yu; Richards, Joanne S

    2010-02-01

    The small G proteins of the RAS superfamily act as molecular switches in the transduction of cellular signals critical for a wide range of normal developmental events as well as pathological processes. However, the functions of Ras genes in ovarian cells have only started to be unveiled. RAS, most likely KRAS that is highly expressed in granulosa cells of growing follicles, appears crucial for mediating the gonadotropin-induced events associated with the unique physiological process of ovulation. By contrast, conditional expression of a constitutively active Kras(G12D) mutant in granulosa cells results in ovulation defects due to the complete disruption of normal follicular growth, cessation of granulosa cell proliferation, and blockage of granulosa cell apoptosis and differentiation. When the tumor suppressor Pten is disrupted conditionally in the Kras(G12D)-expressing granulosa cells, granulosa cell tumors fail to develop. However, ovarian surface epithelial cells expressing the same Pten;Kras(G12D) mutations rapidly become ovarian surface epithelial serous cystadenocarcinomas. In this minireview, we summarize some of the physiological as well as pathological functions of RAS in the rodent ovary, discuss the implications of the Kras(G12D) mutant mouse models for understanding human diseases such as premature ovarian failure and ovarian cancers, and highlight new questions raised by the results of recent studies.

  19. RAS and BRAF in metastatic colorectal cancer management

    PubMed Central

    Gong, Jun; Cho, May

    2016-01-01

    The treatment of metastatic colorectal cancer (mCRC) has been further refined with the development of monoclonal antibodies, cetuximab and panitumumab, towards the epidermal growth factor receptor (EGFR). Anti-EGFR therapy has afforded improved survival in those with wild-type RAS mCRC but provides no benefit and even harm in those with RAS-mutant tumors. BRAF mutations have also been shown to predict lack of clinically meaningful benefit to anti-EGFR therapy in mCRC. Mechanisms of resistance to EGFR blockade in wild-type RAS or BRAF metastatic colorectal tumors appear to converge on the mitogen-activated protein kinase (MAPK) signaling pathway. Clinical trials involving combined BRAF, EGFR, and/or MAPK kinase (MEK) inhibition have shown promising activity in BRAF-mutant mCRC. Here, we review pivotal clinical trials that have redefined our treatment approach in mCRC with respect to anti-EGFR therapy based on RAS and BRAF mutation status. Future studies will likely focus on improving efficacy of anti-EGFR-based therapy in mCRC through sustained MAPK pathway inhibition. PMID:27747083

  20. NF2 loss promotes oncogenic RAS-induced thyroid cancers via YAP-dependent transactivation of RAS proteins and sensitizes them to MEK inhibition

    PubMed Central

    Garcia-Rendueles, Maria E.R.; Ricarte-Filho, Julio C.; Untch, Brian R.; Landa, Iňigo; Knauf, Jeffrey A.; Voza, Francesca; Smith, Vicki E.; Ganly, Ian; Taylor, Barry S.; Persaud, Yogindra; Oler, Gisele; Fang, Yuqiang; Jhanwar, Suresh C.; Viale, Agnes; Heguy, Adriana; Huberman, Kety H.; Giancotti, Filippo; Ghossein, Ronald; Fagin, James A.

    2015-01-01

    Ch22q LOH is preferentially associated with RAS mutations in papillary and in poorly differentiated thyroid cancer (PDTC). The 22q tumor suppressor NF2, encoding merlin, is implicated in this interaction because of its frequent loss of function in human thyroid cancer cell lines. Nf2 deletion or Hras mutation are insufficient for transformation, whereas their combined disruption leads to murine PDTC with increased MAPK signaling. Merlin loss induces RAS signaling in part through inactivation of Hippo, which activates a YAP-TEAD transcriptional program. We find that the three RAS genes are themselves YAP-TEAD1 transcriptional targets, providing a novel mechanism of promotion of RAS-induced tumorigenesis. Moreover, pharmacological disruption of YAP-TEAD with verteporfin blocks RAS transcription and signaling, and inhibits cell growth. The increased MAPK output generated by NF2 loss in RAS-mutant cancers may inform therapeutic strategies, as it generates greater dependency on the MAPK pathway for viability. PMID:26359368

  1. Meibomian gland dysfunction and contact lens intolerance.

    PubMed

    Korb, D R; Henriquez, A S

    1980-03-01

    A study of a syndrome characterized by deficient or inadequate Meibomian gland secretions, minimal or transient symptoms suggestive of ocular dryness, fluorescein staining of the cornea (often detected only after delayed observation or sequential instillation of stain), and contact lens intolerance is described. Clinical and cytologic studies indicate that the syndrome is due to obstruction of the Meibomian gland orifices by desquamated epithelial cells that tend to aggregate in keratotic clusters, which results in alteration of the Meibomian glands' contribution to the precorneal tear film. Further complication may result from bacterial proliferation in the desquamated keratotic cells and the release of the bacteria and their toxic products into the precorneal tear film from these reservoirs in the excretory pathways of the Meibomian glands.

  2. Ras-mediated deregulation of the circadian clock in cancer.

    PubMed

    Relógio, Angela; Thomas, Philippe; Medina-Pérez, Paula; Reischl, Silke; Bervoets, Sander; Gloc, Ewa; Riemer, Pamela; Mang-Fatehi, Shila; Maier, Bert; Schäfer, Reinhold; Leser, Ulf; Herzel, Hanspeter; Kramer, Achim; Sers, Christine

    2014-01-01

    Circadian rhythms are essential to the temporal regulation of molecular processes in living systems and as such to life itself. Deregulation of these rhythms leads to failures in biological processes and eventually to the manifestation of pathological phenotypes including cancer. To address the questions as to what are the elicitors of a disrupted clock in cancer, we applied a systems biology approach to correlate experimental, bioinformatics and modelling data from several cell line models for colorectal and skin cancer. We found strong and weak circadian oscillators within the same type of cancer and identified a set of genes, which allows the discrimination between the two oscillator-types. Among those genes are IFNGR2, PITX2, RFWD2, PPARγ, LOXL2, Rab6 and SPARC, all involved in cancer-related pathways. Using a bioinformatics approach, we extended the core-clock network and present its interconnection to the discriminative set of genes. Interestingly, such gene signatures link the clock to oncogenic pathways like the RAS/MAPK pathway. To investigate the potential impact of the RAS/MAPK pathway - a major driver of colorectal carcinogenesis - on the circadian clock, we used a computational model which predicted that perturbation of BMAL1-mediated transcription can generate the circadian phenotypes similar to those observed in metastatic cell lines. Using an inducible RAS expression system, we show that overexpression of RAS disrupts the circadian clock and leads to an increase of the circadian period while RAS inhibition causes a shortening of period length, as predicted by our mathematical simulations. Together, our data demonstrate that perturbations induced by a single oncogene are sufficient to deregulate the mammalian circadian clock. PMID:24875049

  3. Ras-Mediated Deregulation of the Circadian Clock in Cancer

    PubMed Central

    Relógio, Angela; Thomas, Philippe; Medina-Pérez, Paula; Reischl, Silke; Bervoets, Sander; Gloc, Ewa; Riemer, Pamela; Mang-Fatehi, Shila; Maier, Bert; Schäfer, Reinhold; Leser, Ulf; Herzel, Hanspeter; Kramer, Achim; Sers, Christine

    2014-01-01

    Circadian rhythms are essential to the temporal regulation of molecular processes in living systems and as such to life itself. Deregulation of these rhythms leads to failures in biological processes and eventually to the manifestation of pathological phenotypes including cancer. To address the questions as to what are the elicitors of a disrupted clock in cancer, we applied a systems biology approach to correlate experimental, bioinformatics and modelling data from several cell line models for colorectal and skin cancer. We found strong and weak circadian oscillators within the same type of cancer and identified a set of genes, which allows the discrimination between the two oscillator-types. Among those genes are IFNGR2, PITX2, RFWD2, PPARγ, LOXL2, Rab6 and SPARC, all involved in cancer-related pathways. Using a bioinformatics approach, we extended the core-clock network and present its interconnection to the discriminative set of genes. Interestingly, such gene signatures link the clock to oncogenic pathways like the RAS/MAPK pathway. To investigate the potential impact of the RAS/MAPK pathway - a major driver of colorectal carcinogenesis - on the circadian clock, we used a computational model which predicted that perturbation of BMAL1-mediated transcription can generate the circadian phenotypes similar to those observed in metastatic cell lines. Using an inducible RAS expression system, we show that overexpression of RAS disrupts the circadian clock and leads to an increase of the circadian period while RAS inhibition causes a shortening of period length, as predicted by our mathematical simulations. Together, our data demonstrate that perturbations induced by a single oncogene are sufficient to deregulate the mammalian circadian clock. PMID:24875049

  4. Phenotypic Screening Identifies Protein Synthesis Inhibitors as H-Ras-Nanocluster-Increasing Tumor Growth Inducers.

    PubMed

    Najumudeen, Arafath K; Posada, Itziar M D; Lectez, Benoit; Zhou, Yong; Landor, Sebastian K-J; Fallarero, Adyary; Vuorela, Pia; Hancock, John; Abankwa, Daniel

    2015-12-15

    Ras isoforms H-, N-, and K-ras are each mutated in specific cancer types at varying frequencies and have different activities in cell fate control. On the plasma membrane, Ras proteins are laterally segregated into isoform-specific nanoscale signaling hubs, termed nanoclusters. As Ras nanoclusters are required for Ras signaling, chemical modulators of nanoclusters represent ideal candidates for the specific modulation of Ras activity in cancer drug development. We therefore conducted a chemical screen with commercial and in-house natural product libraries using a cell-based H-ras-nanoclustering FRET assay. Next to established Ras inhibitors, such as a statin and farnesyl-transferase inhibitor, we surprisingly identified five protein synthesis inhibitors as positive regulators. Using commonly employed cycloheximide as a representative compound, we show that protein synthesis inhibition increased nanoclustering and effector recruitment specifically of active H-ras but not of K-ras. Consistent with these data, cycloheximide treatment activated both Erk and Akt kinases and specifically promoted H-rasG12V-induced, but not K-rasG12V-induced, PC12 cell differentiation. Intriguingly, cycloheximide increased the number of mammospheres, which are enriched for cancer stem cells. Depletion of H-ras in combination with cycloheximide significantly reduced mammosphere formation, suggesting an exquisite synthetic lethality. The potential of cycloheximide to promote tumor cell growth was also reflected in its ability to increase breast cancer cell tumors grown in ovo. These results illustrate the possibility of identifying Ras-isoform-specific modulators using nanocluster-directed screening. They also suggest an unexpected feedback from protein synthesis inhibition to Ras signaling, which might present a vulnerability in certain tumor cell types. PMID:26568031

  5. Inhibition of Acid Sphingomyelinase Depletes Cellular Phosphatidylserine and Mislocalizes K-Ras from the Plasma Membrane.

    PubMed

    Cho, Kwang-Jin; van der Hoeven, Dharini; Zhou, Yong; Maekawa, Masashi; Ma, Xiaoping; Chen, Wei; Fairn, Gregory D; Hancock, John F

    2015-01-01

    K-Ras must localize to the plasma membrane for biological activity; thus, preventing plasma membrane interaction blocks K-Ras signal output. Here we show that inhibition of acid sphingomyelinase (ASM) mislocalizes both the K-Ras isoforms K-Ras4A and K-Ras4B from the plasma membrane to the endomembrane and inhibits their nanoclustering. We found that fendiline, a potent ASM inhibitor, reduces the phosphatidylserine (PtdSer) and cholesterol content of the inner plasma membrane. These lipid changes are causative because supplementation of fendiline-treated cells with exogenous PtdSer rapidly restores K-Ras4A and K-Ras4B plasma membrane binding, nanoclustering, and signal output. Conversely, supplementation with exogenous cholesterol restores K-Ras4A but not K-Ras4B nanoclustering. These experiments reveal different operational pools of PtdSer on the plasma membrane. Inhibition of ASM elevates cellular sphingomyelin and reduces cellular ceramide levels. Concordantly, delivery of recombinant ASM or exogenous ceramide to fendiline-treated cells rapidly relocalizes K-Ras4B and PtdSer to the plasma membrane. K-Ras4B mislocalization is also recapitulated in ASM-deficient Neimann-Pick type A and B fibroblasts. This study identifies sphingomyelin metabolism as an indirect regulator of K-Ras4A and K-Ras4B signaling through the control of PtdSer plasma membrane content. It also demonstrates the critical and selective importance of PtdSer to K-Ras4A and K-Ras4B plasma membrane binding and nanoscale spatial organization. PMID:26572827

  6. Oncogenicity of human N-ras oncogene and proto-oncogene introduced into retroviral vectors

    SciTech Connect

    Souyri, M.; Vigon, I.; Charon, M.; Tambourin, P. )

    1989-09-01

    The N-ras gene is the only member of the ras family which has never been naturally transduced into a retrovirus. In order to study the in vitro and in vivo oncogenicity of N-ras and to compare its pathogenicity to that of H-ras, the authors have inserted an activated or a normal form of human N-ras cDNA into a slightly modified Harvey murine sarcoma virus-derived vector in which the H-ras p21 coding region had been deleted. The resulting constructions were transfected into NIH 3T3 cells. The activated N-ras-containing construct (HSN) induced 10{sup 4} foci per {mu}g of DNA and was found to be as transforming as H-ras was. After infection of the transfected cells by either the ecotropic Moloney murine leukemia virus or the amphotropic 4070A helper viruses, rescued transforming viruses were injected into newborn mice. Both pseudotypes of HSN virus containing activated N-ras induced the typical Harvey disease with similar latency. However, they found that the virus which contained normal N-ras p21 (HSn) was also pathogenic and induced splenomegaly, lymphadenopathies, and sarcoma in mice after a latency of 3 to 7 weeks. In addition, Moloney murine leukemia virus pseudotypes of N-ras caused neurological disorders in 30% of the infected animals. These results differed markedly from those of previous experiments in which the authors had inserted the activated form of N-ras in the pSV(X) vector: the resulting SVN-ras virus was transforming on NIH 3T3 cells but was poorly oncogenic in vivo. Altogether, these data demonstrated unequivocally that N-ras is potentially as oncogenic as H-ras and that such oncogenic effect could depend on the vector environment.

  7. Phenotypic Screening Identifies Protein Synthesis Inhibitors as H-Ras-Nanocluster-Increasing Tumor Growth Inducers.

    PubMed

    Najumudeen, Arafath K; Posada, Itziar M D; Lectez, Benoit; Zhou, Yong; Landor, Sebastian K-J; Fallarero, Adyary; Vuorela, Pia; Hancock, John; Abankwa, Daniel

    2015-12-15

    Ras isoforms H-, N-, and K-ras are each mutated in specific cancer types at varying frequencies and have different activities in cell fate control. On the plasma membrane, Ras proteins are laterally segregated into isoform-specific nanoscale signaling hubs, termed nanoclusters. As Ras nanoclusters are required for Ras signaling, chemical modulators of nanoclusters represent ideal candidates for the specific modulation of Ras activity in cancer drug development. We therefore conducted a chemical screen with commercial and in-house natural product libraries using a cell-based H-ras-nanoclustering FRET assay. Next to established Ras inhibitors, such as a statin and farnesyl-transferase inhibitor, we surprisingly identified five protein synthesis inhibitors as positive regulators. Using commonly employed cycloheximide as a representative compound, we show that protein synthesis inhibition increased nanoclustering and effector recruitment specifically of active H-ras but not of K-ras. Consistent with these data, cycloheximide treatment activated both Erk and Akt kinases and specifically promoted H-rasG12V-induced, but not K-rasG12V-induced, PC12 cell differentiation. Intriguingly, cycloheximide increased the number of mammospheres, which are enriched for cancer stem cells. Depletion of H-ras in combination with cycloheximide significantly reduced mammosphere formation, suggesting an exquisite synthetic lethality. The potential of cycloheximide to promote tumor cell growth was also reflected in its ability to increase breast cancer cell tumors grown in ovo. These results illustrate the possibility of identifying Ras-isoform-specific modulators using nanocluster-directed screening. They also suggest an unexpected feedback from protein synthesis inhibition to Ras signaling, which might present a vulnerability in certain tumor cell types.

  8. Targeting the K-Ras/PDEδ protein-protein interaction: the solution for Ras-driven cancers or just another therapeutic mirage?

    PubMed

    Frett, Brendan; Wang, Yuanxiang; Li, Hong-Yu

    2013-10-01

    The holy grail, finally? After years of unsuccessful attempts at drugging the Ras oncogene, a recent paper by Zimmerman et al. has revealed the possibility of inhibiting Ras signaling on a clinically relevant level by blocking the K-Ras/PDEδ protein-protein interaction. The results, reported in Nature, are highlighted herein with future implications and directions to evaluate the full clinical potential of this research. PMID:23939923

  9. Delayed accumulation of lens material behind the foldable intraocular lens.

    PubMed

    Bhattacharjee, Harsha; Bhattacharjee, Kasturi; Bhattacharjee, Pankaj

    2007-01-01

    Foldable acrylic intraocular lenses (IOLs) are known to reduce posterior capsule opacification by preventing migration of lens epithelial cells with its square edge design and its property of tackiness. Studies have reported a mean adhesiveness to posterior capsule more than three times higher for certain acrylic foldable IOLs than polymethyl methacrylate IOLs. The authors would like to report two cases where the force of tackiness was compensated, thereby presenting with delayed accumulation of lens material in the capsular bags behind the IOL with temporary loss of vision. PMID:17951912

  10. Delayed accumulation of lens material behind the foldable intraocular lens.

    PubMed

    Bhattacharjee, Harsha; Bhattacharjee, Kasturi; Bhattacharjee, Pankaj

    2007-01-01

    Foldable acrylic intraocular lenses (IOLs) are known to reduce posterior capsule opacification by preventing migration of lens epithelial cells with its square edge design and its property of tackiness. Studies have reported a mean adhesiveness to posterior capsule more than three times higher for certain acrylic foldable IOLs than polymethyl methacrylate IOLs. The authors would like to report two cases where the force of tackiness was compensated, thereby presenting with delayed accumulation of lens material in the capsular bags behind the IOL with temporary loss of vision.

  11. Delayed accumulation of lens material behind the foldable intraocular lens

    PubMed Central

    Bhattacharjee, Kasturi; Bhattacharjee, Pankaj

    2007-01-01

    Foldable acrylic intraocular lenses (IOLs) are known to reduce posterior capsule opacification by preventing migration of lens epithelial cells with its square edge design and its property of tackiness. Studies have reported a mean adhesiveness to posterior capsule more than three times higher for certain acrylic foldable IOLs than polymethyl methacrylate IOLs. The authors would like to report two cases where the force of tackiness was compensated, thereby presenting with delayed accumulation of lens material in the capsular bags behind the IOL with temporary loss of vision. PMID:17951912

  12. Targeting the RAS pathway by mitogen-activated protein kinase inhibitors.

    PubMed

    Kiessling, Michael K; Rogler, Gerhard

    2015-01-01

    Targeting of oncogenic driver mutations with small-molecule inhibitors resulted in powerful treatment options for cancer patients in recent years. The RAS (rat sarcoma) pathway is among the most frequently mutated pathways in human cancer. Whereas targeting mutant Kirsten RAS (KRAS) remains difficult, mutant B rapidly accelerated fibrosarcoma (BRAF) kinase is an established drug target in cancer. Now data show that neuroblastoma RAS (NRAS) and even Harvey RAS (HRAS) mutations could be predictive markers for treatment with mitogen-activated protein kinase (MEK) inhibitors. This review discusses recent preclinical and clinical studies of MEK inhibitors in BRAF and RAS mutant cancer. PMID:26691679

  13. K-Ras Promotes Tumorigenicity through Suppression of Non-canonical Wnt Signaling.

    PubMed

    Wang, Man-Tzu; Holderfield, Matthew; Galeas, Jacqueline; Delrosario, Reyno; To, Minh D; Balmain, Allan; McCormick, Frank

    2015-11-19

    K-Ras and H-Ras share identical effectors and have similar properties; however, the high degree of tumor-type specificity associated with K-Ras and H-Ras mutations suggests that they have unique roles in oncogenesis. Here, we report that oncogenic K-Ras, but not H-Ras, suppresses non-canonical Wnt/Ca(2+) signaling, an effect that contributes strongly to its tumorigenic properties. K-Ras does this by binding to calmodulin and so reducing CaMKii activity and expression of Fzd8. Restoring Fzd8 in K-Ras mutant pancreatic cells suppresses malignancy, whereas depletion of Fzd8 in H-Ras(V12)-transformed cells enhances their tumor initiating capacity. Interrupting K-Ras-calmodulin binding using genetic means or by treatment with an orally active protein kinase C (PKC)-activator, prostratin, represses tumorigenesis in K-Ras mutant pancreatic cancer cells. These findings provide an alternative way to selectively target this "undruggable" protein.

  14. N-terminally myristoylated Ras proteins require palmitoylation or a polybasic domain for plasma membrane localization.

    PubMed

    Cadwallader, K A; Paterson, H; Macdonald, S G; Hancock, J F

    1994-07-01

    Plasma membrane targeting of Ras requires CAAX motif modifications together with a second signal from an adjacent polybasic domain or nearby cysteine palmitoylation sites. N-terminal myristoylation is known to restore membrane binding to H-ras C186S (C-186 is changed to S), a mutant protein in which all CAAX processing is abolished. We show here that myristoylated H-ras C186S is a substrate for palmitoyltransferase, despite the absence of C-terminal farnesylation, and that palmitoylation is absolutely required for plasma membrane targeting of myristoylated H-ras. Similarly, the polybasic domain is required for specific plasma membrane targeting of myristoylated K-ras. In contrast, the combination of myristoylation plus farnesylation results in the mislocalization of Ras to numerous intracellular membranes. Ras that is only myristoylated does not bind with a high affinity to any membrane. The specific targeting of Ras to the plasma membrane is therefore critically dependent on signals that are contained in the hypervariable domain but can be supported by N-terminal myristoylation or C-terminal prenylation. Interestingly, oncogenic Ras G12V that is localized correctly to the plasma membrane leads to mitogen-activated protein kinase activation irrespective of the combination of targeting signals used for localization, whereas Ras G12V that is mislocalized to the cytosol or to other membranes activates mitogen-activated protein kinase only if the Ras protein is farnesylated.

  15. Approach for targeting Ras with small molecules that activate SOS-mediated nucleotide exchange.

    PubMed

    Burns, Michael C; Sun, Qi; Daniels, R Nathan; Camper, DeMarco; Kennedy, J Phillip; Phan, Jason; Olejniczak, Edward T; Lee, Taekyu; Waterson, Alex G; Rossanese, Olivia W; Fesik, Stephen W

    2014-03-01

    Aberrant activation of the small GTPase Ras by oncogenic mutation or constitutively active upstream receptor tyrosine kinases results in the deregulation of cellular signals governing growth and survival in ∼30% of all human cancers. However, the discovery of potent inhibitors of Ras has been difficult to achieve. Here, we report the identification of small molecules that bind to a unique pocket on the Ras:Son of Sevenless (SOS):Ras complex, increase the rate of SOS-catalyzed nucleotide exchange in vitro, and modulate Ras signaling pathways in cells. X-ray crystallography of Ras:SOS:Ras in complex with these molecules reveals that the compounds bind in a hydrophobic pocket in the CDC25 domain of SOS adjacent to the Switch II region of Ras. The structure-activity relationships exhibited by these compounds can be rationalized on the basis of multiple X-ray cocrystal structures. Mutational analyses confirmed the functional relevance of this binding site and showed it to be essential for compound activity. These molecules increase Ras-GTP levels and disrupt MAPK and PI3K signaling in cells at low micromolar concentrations. These small molecules represent tools to study the acute activation of Ras and highlight a pocket on SOS that may be exploited to modulate Ras signaling.

  16. RasGRP1 Transgenic Mice Develop Cutaneous Squamous Cell Carcinomas in Response to Skin Wounding

    PubMed Central

    Diez, Federico R.; Garrido, Ann A.; Sharma, Amrish; Luke, Courtney T.; Stone, James C.; Dower, Nancy A.; Cline, J. Mark; Lorenzo, Patricia S.

    2009-01-01

    Models of epidermal carcinogenesis have demonstrated that Ras is a critical molecule involved in tumor initiation and progression. Previously, we have shown that RasGRP1 increases the susceptibility of mice to skin tumorigenesis when overexpressed in the epidermis by a transgenic approach, related to its ability to activate Ras. Moreover, RasGRP1 transgenic mice develop spontaneous papillomas and cutaneous squamous cell carcinomas, some of which appear to originate in sites of injury, suggesting that RasGRP1 may be responding to signals generated during the wound-healing process. In this study, we examined the response of the RasGRP1 transgenic animals to full-thickness incision wounding of the skin, and demonstrated that they respond by developing tumors along the wounded site. The tumors did not present mutations in the H-ras gene, but Rasgrp1 transgene dosage correlated with tumor susceptibility and size. Analysis of serum cytokines showed increased levels of granulocyte colony-stimulating factor in transgenic animals after wounding. Furthermore, in vitro experiments with primary keratinocytes showed that granulocyte colony-stimulating factor stimulated Ras activation, although RasGRP1 was dispensable for this effect. Since granulocyte colony-stimulating factor has been recently associated with proliferation of skin cancer cells, our results may help in the elucidation of pathways that activate Ras in the epidermis during tumorigenesis in the absence of oncogenic ras mutations. PMID:19497993

  17. Absence of K-Ras Reduces Proliferation and Migration But Increases Extracellular Matrix Synthesis in Fibroblasts.

    PubMed

    Muñoz-Félix, José M; Fuentes-Calvo, Isabel; Cuesta, Cristina; Eleno, Nélida; Crespo, Piero; López-Novoa, José M; Martínez-Salgado, Carlos

    2016-10-01

    The involvement of Ras-GTPases in the development of renal fibrosis has been addressed in the last decade. We have previously shown that H- and N-Ras isoforms participate in the regulation of fibrosis. Herein, we assessed the role of K-Ras in cellular processes involved in the development of fibrosis: proliferation, migration, and extracellular matrix (ECM) proteins synthesis. K-Ras knockout (KO) mouse embryonic fibroblasts (K-ras(-/-) ) stimulated with transforming growth factor-β1 (TGF-β1) exhibited reduced proliferation and impaired mobility than wild-type fibroblasts. Moreover, an increase on ECM production was observed in K-Ras KO fibroblasts in basal conditions. The absence of K-Ras was accompanied by reduced Ras activation and ERK phosphorylation, and increased AKT phosphorylation, but no differences were observed in TGF-β1-induced Smad signaling. The MEK inhibitor U0126 decreased cell proliferation independently of the presence of K-ras but reduced migration and ECM proteins expression only in wild-type fibroblasts, while the PI3K-AKT inhibitor LY294002 decreased cell proliferation, migration, and ECM synthesis in both types of fibroblasts. Thus, our data unveil that K-Ras and its downstream effector pathways distinctively regulate key biological processes in the development of fibrosis. Moreover, we show that K-Ras may be a crucial mediator in TGF-β1-mediated effects in this cell type. J. Cell. Physiol. 231: 2224-2235, 2016. © 2016 Wiley Periodicals, Inc.

  18. The SNAP Strong Lens Survey

    SciTech Connect

    Marshall, P.

    2005-01-03

    Basic considerations of lens detection and identification indicate that a wide field survey of the types planned for weak lensing and Type Ia SNe with SNAP are close to optimal for the optical detection of strong lenses. Such a ''piggy-back'' survey might be expected even pessimistically to provide a catalogue of a few thousand new strong lenses, with the numbers dominated by systems of faint blue galaxies lensed by foreground ellipticals. After sketching out our strategy for detecting and measuring these galaxy lenses using the SNAP images, we discuss some of the scientific applications of such a large sample of gravitational lenses: in particular we comment on the partition of information between lens structure, the source population properties and cosmology. Understanding this partitioning is key to assessing strong lens cosmography's value as a cosmological probe.

  19. [Contact lens-related keratitis].

    PubMed

    Steiber, Zita; Berta, András; Módis, László

    2013-11-10

    Nowadays, keratitis, corneal infection due to wearing contact lens means an increasingly serious problem. Neglected cases may lead to corneal damage that can cause blindness in cases of otherwise healthy eyes. Early diagnosis based on the clinical picture and the typical patient history is an important way of prevention. Prophylaxis is substantial to avoid bacterial and viral infection that is highly essential in this group of diseases. Teaching contact lens wearers the proper contact lens care, storage, sterility, and hygiene regulations is of great importance. In case of corneal inflammation early accurate diagnosis supported by microbiological culture from contact lenses, storage boxes or cornea is very useful. Thereafter, targeted drug therapy or in therapy-resistant cases surgical treatment may even be necessary in order to sustain suitable visual acuity.

  20. A Prototype Antifungal Contact Lens

    PubMed Central

    Ciolino, Joseph B.; Hudson, Sarah P.; Mobbs, Ashley N.; Hoare, Todd R.; Iwata, Naomi G.; Fink, Gerald R.

    2011-01-01

    Purpose. To design a contact lens to treat and prevent fungal ocular infections. Methods. Curved contact lenses were created by encapsulating econazole-impregnated poly(lactic-co-glycolic) acid (PLGA) films in poly(hydroxyethyl methacrylate) (pHEMA) by ultraviolet photopolymerization. Release studies were conducted in phosphate-buffered saline at 37°C with continuous shaking. The contact lenses and their release media were tested in an antifungal assay against Candida albicans. Cross sections of the pre- and postrelease contact lenses were characterized by scanning electron microscopy and by Raman spectroscopy. Results. Econazole-eluting contact lenses provided extended antifungal activity against Candida albicans fungi. Fungicidal activity varied in duration and effectiveness depending on the mass of the econazole-PLGA film encapsulated in the contact lens. Conclusions. An econazole-eluting contact lens could be used as a treatment for fungal ocular infections. PMID:21527380

  1. Automated Fresnel lens tester system

    SciTech Connect

    Phipps, G.S.

    1981-07-01

    An automated data collection system controlled by a desktop computer has been developed for testing Fresnel concentrators (lenses) intended for solar energy applications. The system maps the two-dimensional irradiance pattern (image) formed in a plane parallel to the lens, whereas the lens and detector assembly track the sun. A point detector silicon diode (0.5-mm-dia active area) measures the irradiance at each point of an operator-defined rectilinear grid of data positions. Comparison with a second detector measuring solar insolation levels results in solar concentration ratios over the image plane. Summation of image plane energies allows calculation of lens efficiencies for various solar cell sizes. Various graphical plots of concentration ratio data help to visualize energy distribution patterns.

  2. Oncogenic K-Ras promotes proliferation in quiescent intestinal stem cells.

    PubMed

    Gierut, Jessica J; Lyons, Jesse; Shah, Manasvi S; Genetti, Casie; Breault, David T; Haigis, Kevin M

    2015-07-01

    K-Ras is a monomeric GTPase that controls cellular and tissue homeostasis. Prior studies demonstrated that mutationally activated K-Ras (K-Ras(G12D)) signals through MEK to promote expansion and hyperproliferation of the highly mitotically active transit-amplifying cells (TACs) in the intestinal crypt. Its effect on normally quiescent stem cells was unknown, however. Here, we have used an H2B-Egfp transgenic system to demonstrate that K-Ras(G12D) accelerates the proliferative kinetics of quiescent intestinal stem cells. As in the TAC compartment, the effect of mutant K-Ras on the quiescent stem cell is dependent upon activation of MEK. Mutant K-Ras is also able to increase self-renewal potential of intestinal stem cells following damage. These results demonstrate that mutant K-Ras can influence intestinal homeostasis on multiple levels.

  3. A Ras subfamily GTPase shows cell cycle-dependent nuclear localization

    PubMed Central

    Sutherland, Brent W.; Spiegelman, George B.; Weeks, Gerald

    2001-01-01

    Previously characterized Ras subfamily proteins have been found to be predominantly associated with the plasma membrane where they function in signal transduction pathways to convey extracellular signals to intracellular targets. Here, we provide evidence that the Dictyostelium Ras subfamily protein RasB has a novel subcellular localization and function. The protein is predominantly localized in the nucleus during most of the cell cycle. Furthermore, during mitosis and cytokinesis RasB assumes a diffuse cellular localization despite the fact that the nuclear membrane stays intact. The linkage between the position of RasB in the cell and division suggests that it may have a role in nuclear division. Consistent with this idea, rasB– cells exhibit severe growth defects and cells overexpressing an activated version of RasB are multinucleate. PMID:11606416

  4. Impaired Binding of 14-3-3 to C-RAF in Noonan Syndrome Suggests New Approaches in Diseases with Increased Ras Signaling▿

    PubMed Central

    Molzan, Manuela; Schumacher, Benjamin; Ottmann, Corinna; Baljuls, Angela; Polzien, Lisa; Weyand, Michael; Thiel, Philipp; Rose, Rolf; Rose, Micheline; Kuhenne, Philipp; Kaiser, Markus; Rapp, Ulf R.; Kuhlmann, Jürgen; Ottmann, Christian

    2010-01-01

    The Ras-RAF-mitogen-activated protein kinase (Ras-RAF-MAPK) pathway is overactive in many cancers and in some developmental disorders. In one of those disorders, namely, Noonan syndrome, nine activating C-RAF mutations cluster around Ser259, a regulatory site for inhibition by 14-3-3 proteins. We show that these mutations impair binding of 14-3-3 proteins to C-RAF and alter its subcellular localization by promoting Ras-mediated plasma membrane recruitment of C-RAF. By presenting biophysical binding data, the 14-3-3/C-RAFpS259 crystal structure, and cellular analyses, we indicate a mechanistic link between a well-described human developmental disorder and the impairment of a 14-3-3/target protein interaction. As a broader implication of these findings, modulating the C-RAFSer259/14-3-3 protein-protein interaction with a stabilizing small molecule may yield a novel potential approach for treatment of diseases resulting from an overactive Ras-RAF-MAPK pathway. PMID:20679480

  5. Computer-aided lens assembly.

    PubMed

    Tomlinson, Richard; Alcock, Rob; Petzing, Jon; Coupland, Jeremy

    2004-01-20

    We propose a computer-aided method of lens manufacture that allows assembly, adjustment, and test phases to be run concurrently until an acceptable level of optical performance is reached. Misalignment of elements within a compound lens is determined by a comparison of the results of physical ray tracing by use of an array of Gaussian laser beams with numerically obtained geometric ray traces. An estimate of misalignment errors is made, and individual elements are adjusted in an iterative manner until performance criteria are achieved. The method is illustrated for the alignment of an air-spaced doublet. PMID:14765916

  6. Liquid crystal Fresnel lens display

    NASA Astrophysics Data System (ADS)

    Wang, Xiao-Qian; Abhishek Kumar, Srivastava; Alwin Tam, Ming-Wai; Zheng, Zhi-Gang; Shen, Dong; Vladimir, Chigrinov G.; Kwok, Hoi-Sing

    2016-09-01

    A novel see-through display with a liquid crystal lens array was proposed. A liquid crystal Fresnel lens display (LCFLD) with a holographic screen was demonstrated. The proposed display system has high efficiency, simple fabrication, and low manufacturing cost due to the absence of a polarizer and color filter. Project supported by Partner State Key Laboratory on Advanced Displays and Optoelectronics Technologies HKUST, China, the National Natural Science Foundation of China (Grant Nos. 61435008 and 61575063), and the Fundamental Research Funds for the Central Universities, China (Grant No. WM1514036).

  7. Liquid crystal Fresnel lens display

    NASA Astrophysics Data System (ADS)

    Wang, Xiao-Qian; Abhishek Kumar, Srivastava; Alwin Tam, Ming-Wai; Zheng, Zhi-Gang; Shen, Dong; Vladimir, Chigrinov G.; Kwok, Hoi-Sing

    2016-09-01

    A novel see-through display with a liquid crystal lens array was proposed. A liquid crystal Fresnel lens display (LCFLD) with a holographic screen was demonstrated. The proposed display system has high efficiency, simple fabrication, and low manufacturing cost due to the absence of a polarizer and color filter. Project supported by Partner State Key Laboratory on Advanced Displays and Optoelectronics Technologies HKUST, China, the National Natural Science Foundation of China (Grant Nos. 61435008 and 61575063), and the Fundamental Research Funds for the Central Universities, China (Grant No. WM1514036).

  8. Liquid-filled varifocal lens on a chip

    NASA Astrophysics Data System (ADS)

    Choi, Seung Tae; Lee, Jeong Yub; Kwon, Jong Oh; Lee, Seungwan; Kim, Woonbae

    2009-02-01

    In this study we developed a liquid-filled varifocal lens operated by electroactive polymer actuators. A silicon wafer was structured with micromachining processes to have four microfluidic chambers and a circular hole working as an aperture. The structured silicon wafer (opaque frame) was bonded to a glass wafer (transparent frame), and thus microfluidic channels were formed between them. Top surface of the main frame was covered with a transparent elastomer membrane, and the internal volume confined by the membrane and the two frames was filled with optical fluid. In order to operate this varifocal lens system, multilayered P(VDF-TrFE-CFE) [poly(vinylidene fluoride-trifluoroethylene-clorofluoroethylene)] polymer actuators were also developed, which show relaxor ferroelectric behavior, and thus produce large electrostrictive strain. When an electric field is applied, the multilayered P(VDF-TrFE-CFE) polymer actuators push the optical fluid so that the elastomer membrane together with the internal fluid changes their shape, which alters the light path of the varifocal lens. The original shape of the elastomer membrane is restored by the elastic recovery of the P(VDF-TrFE-CFE) actuators when an applied electric field is removed. We observed that with the applied voltage of 40 V the varifocal lens changes the optical power of more than 30 diopters within 20 ms. Optical analysis showed that the deformation shape of the optical membrane can be successfully used to design phone camera modules with auto-focus function.

  9. Changes in the Eye Microbiota Associated with Contact Lens Wearing

    PubMed Central

    Price, Kenneth; Albert, Luong; Dodick, Jack; Park, Lisa

    2016-01-01

    ABSTRACT Wearing contact lenses has been identified as a risk factor for the development of eye conditions such as giant papillary conjunctivitis and keratitis. We hypothesized that wearing contact lenses is associated with changes in the ocular microbiota. We compared the bacterial communities of the conjunctiva and skin under the eye from 58 subjects and analyzed samples from 20 subjects (9 lens wearers and 11 non-lens wearers) taken at 3 time points using a 16S rRNA gene-based sequencing technique (V4 region; Illumina MiSeq). We found that using anesthetic eye drops before sampling decreases the detected ocular microbiota diversity. Compared to those from non-lens wearers, dry conjunctival swabs from lens wearers had more variable and skin-like bacterial community structures (UniFrac; P value = <0.001), with higher abundances of Methylobacterium, Lactobacillus, Acinetobacter, and Pseudomonas and lower abundances of Haemophilus, Streptococcus, Staphylococcus, and Corynebacterium (linear discriminant analysis [LDA] score = >3.0). The results indicate that wearing contact lenses alters the microbial structure of the ocular conjunctiva, making it more similar to that of the skin microbiota. Further research is needed to determine whether the microbiome structure provides less protection from ocular infections. PMID:27006462

  10. Contact lens wear at altitude: subcontact lens bubble formation.

    PubMed

    Flynn, W J; Miller, R E; Tredici, T J; Block, M G; Kirby, E E; Provines, W F

    1987-11-01

    A concern in the past regarding contact lens wear in aviation has been the fear of subcontact lens bubble formation. Previous reports have documented the occurrence of bubbles with hard (PMMA) lenses. Reported here are the results of contact lens bubble studies with soft hydrophilic and rigid gas-permeable lenses. Testing was accomplished in hypobaric chambers and onboard USAF transport aircraft. Hypobaric chamber flights were of three types: high-altitude flights up to 7,620 m (25,000 ft); explosive rapid decompressions from 2,438.4 m (8,000 ft) to 7,620 m (25,000 ft); and 4-h flights at 3,048 m (10,000 ft). Flights aboard transport aircraft typically had cabin pressures equivalent to 1,524-2,438.4 m (5,000-8,000 ft), and ranged in duration from 3 to 10 h. For subjects wearing rigid gas-permeable lenses, central bubbles were detected in 2 of 10 eyes and occurred at altitudes greater than 6,096 m (20,000 ft). With soft contact lenses, bubble formation was detected in approximately 24% (22 of 92 eyes) of the eyes tested, sometimes occurring at altitudes as low as 1,828.8 m (6,000 ft). Soft lens bubbles were always located at the limbus and were without sequela to vision or corneal epithelial integrity. Bubbles under the rigid lenses were primarily central, with potential adverse effects on vision and the corneal epithelium.

  11. Ensuring Safe Use of Contact Lens Solution

    MedlinePlus

    ... For Consumers Consumer Updates Ensuring Safe Use of Contact Lens Solution Share Tweet Linkedin Pin it More ... back to top Dos and Don'ts for Contact Lens Wearers DO: Always wash your hands before ...

  12. Up-regulation of IGF-1R by mutant RAS in leukemia and potentiation of RAS signaling inhibitors by small molecule inhibition of IGF-1R

    PubMed Central

    Weisberg, Ellen; Nonami, Atsushi; Chen, Zhao; Nelson, Erik; Chen, Yongfei; Liu, Feiyang; Cho, Haeyeon; Zhang, Jianming; Sattler, Martin; Mitsiades, Constantine; Wong, Kwok-Kin; Liu, Qingsong; Gray, Nathanael; Griffin, James D.

    2014-01-01

    Purpose Activating mutations in the RAS oncogene occur frequently in human leukemias. Direct targeting of RAS has proven to be challenging, although targeting of downstream RAS mediators, such as MEK, is currently being tested clinically. Given the complexity of RAS signaling, it is likely that combinations of targeted agents will be more effective than single agents. Experimental Design A chemical screen using RAS-dependent leukemia cells was developed to identify compounds with unanticipated activity in the presence of a MEK inhibitor, and led to identification of inhibitors of IGF-1R. Results were validated using cell-based proliferation assays and apoptosis, cell cycle, and gene knockdown assays, immunoprecipitation and immunoblotting, and a non-invasive in vivo bioluminescence model of acute myeloid leukemia (AML). Results Mechanistically, IGF-1R protein expression/activity was substantially increased in mutant RAS-expressing cells, and suppression of RAS led to decreases in IGF-1R. Synergy between MEK and IGF-1R inhibitors correlated with induction of apoptosis, inhibition of cell cycle progression, and decreased phospho-S6 and phospho-4E-BP1. In vivo, NSG mice tail vein-injected with OCI-AML3-luc+ cells showed significantly lower tumor burden following one week of daily oral administration of 50 mg/kg NVP-AEW541 (IGF-1R inhibitor) combined with 25 mg/kg AZD6244 (MEK inhibitor), as compared to mice treated with either agent alone. Drug combination effects observed in cell-based assays were generalized to additional mutant RAS-positive neoplasms. Conclusions The finding that downstream inhibitors of RAS signaling and IGF-1R inhibitors have synergistic activity warrants further clinical investigation of IGF-1R and RAS signaling inhibition as a potential treatment strategy for RAS-driven malignancies. PMID:25186968

  13. SimpLens: Interactive gravitational lensing simulator

    NASA Astrophysics Data System (ADS)

    Saha, Prasenjit; Williams, Liliya L. R.

    2016-06-01

    SimpLens illustrates some of the theoretical ideas important in gravitational lensing in an interactive way. After setting parameters for elliptical mass distribution and external mass, SimpLens displays the mass profile and source position, the lens potential and image locations, and indicate the image magnifications and contours of virtual light-travel time. A lens profile can be made shallower or steeper with little change in the image positions and with only total magnification affected.

  14. A broadband transformation-optics metasurface lens

    SciTech Connect

    Wan, Xiang; Xiang Jiang, Wei; Feng Ma, Hui; Jun Cui, Tie

    2014-04-14

    We present a transformational metasurface Luneburg lens based on the quasi-conformal mapping method, which has weakly anisotropic constitutive parameters. We design the metasurface lens using inhomogeneous artificial structures to realize the required surface refractive indexes. The transformational metasurface Luneburg lens is fabricated and the measurement results demonstrate very good performance in controlling the radiated surface waves.

  15. 21 CFR 886.3600 - Intraocular lens.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Intraocular lens. 886.3600 Section 886.3600 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) MEDICAL DEVICES OPHTHALMIC DEVICES Prosthetic Devices § 886.3600 Intraocular lens. (a) Identification. An intraocular lens is a device made of materials...

  16. Contact Lens-Related Eye Infections

    MedlinePlus

    ... Stories Español Eye Health / Eye Health A-Z Contact Lens-Related Eye Infections Sections Contact Lens-Related ... About Contact Lenses Proper Care of Contact Lenses Contact Lens-Related Eye Infections Written by: Kierstan Boyd ...

  17. New merit values for lens performance.

    NASA Astrophysics Data System (ADS)

    Nakagawa, J.

    1995-08-01

    New merit values for lens performance m=Σ(Qh)2 and m¯=Σ(Qh¯)2 were introduced, where Q is Abbe's invariant, h is paraxial ray height and the bar indicates principal ray. The studies showed that m and m¯ are useful for the evaluation of potentialities of lens systems and for the optimization of lens designs.

  18. In vivo human crystalline lens topography

    PubMed Central

    Ortiz, Sergio; Pérez-Merino, Pablo; Gambra, Enrique; de Castro, Alberto; Marcos, Susana

    2012-01-01

    Custom high-resolution high-speed anterior segment spectral domain optical coherence tomography (OCT) was used to characterize three-dimensionally (3-D) the human crystalline lens in vivo. The system was provided with custom algorithms for denoising and segmentation of the images, as well as for fan (scanning) and optical (refraction) distortion correction, to provide fully quantitative images of the anterior and posterior crystalline lens surfaces. The method was tested on an artificial eye with known surfaces geometry and on a human lens in vitro, and demonstrated on three human lenses in vivo. Not correcting for distortion overestimated the anterior lens radius by 25% and the posterior lens radius by more than 65%. In vivo lens surfaces were fitted by biconicoids and Zernike polynomials after distortion correction. The anterior lens radii of curvature ranged from 10.27 to 14.14 mm, and the posterior lens radii of curvature ranged from 6.12 to 7.54 mm. Surface asphericities ranged from −0.04 to −1.96. The lens surfaces were well fitted by quadrics (with variation smaller than 2%, for 5-mm pupils), with low amounts of high order terms. Surface lens astigmatism was significant, with the anterior lens typically showing horizontal astigmatism (Z22 ranging from −11 to −1 µm) and the posterior lens showing vertical astigmatism (Z22 ranging from 6 to 10 µm). PMID:23082289

  19. Tevatron electron lens magnetic system

    SciTech Connect

    Vladimir Shiltsev et al.

    2001-07-12

    In the framework of collaboration between IHEP and FNAL, a magnetic system of the Tevatron Electron Lens (TEL) has been designed and built. The TEL is currently installed in the superconducting ring of the Tevatron proton-antiproton collider and used for experimental studies of beam-beam compensation [1].

  20. The Fyodorov Sputnik intraocular lens.

    PubMed

    Kwitko, M L

    1979-04-01

    The author has implanted 197 Fyodorov intraocular lenses. With careful selection of patients, good surgical judgment, and meticulous surgery, a degree of success can be obtained with this lens, which will equal that of conventional cataract surgery. The surgical technique of implantation will be described. PMID:537770

  1. A Lens to the Enterprise.

    ERIC Educational Resources Information Center

    Zemsky, Robert, Ed.

    1999-01-01

    This essay is based on a series of roundtables convened through the Knight Collaborative National Medical Education Roundtable. It reports that the challenges and transformations experienced in recent years by community-based medical schools and clinical campuses offer a lens to the whole higher education enterprise, and asks the fundamental…

  2. Gamma Band Activity in the RAS-intracellular mechanisms

    PubMed Central

    Garcia-Rill, E.; Kezunovic, N.; D’Onofrio, S.; Luster, B.; Hyde, J.; Bisagno, V.; Urbano, F.J.

    2014-01-01

    Gamma band activity participates in sensory perception, problem solving, and memory. This review considers recent evidence showing that cells in the reticular activating system (RAS) exhibit gamma band activity, and describes the intrinsic membrane properties behind such manifestation. Specifically, we discuss how cells in the mesopontine pedunculopontine nucleus (PPN), intralaminar parafascicular nucleus (Pf), and pontine Subcoeruleus nucleus dorsalis (SubCD) all fire in the gamma band range when maximally activated, but no higher. The mechanisms involve high threshold, voltage-dependent P/Q-type calcium channels or sodium-dependent subthreshold oscillations. Rather than participating in the temporal binding of sensory events as in the cortex, gamma band activity in the RAS may participate in the processes of preconscious awareness, and provide the essential stream of information for the formulation of many of our actions. We address three necessary next steps resulting from these discoveries, an intracellular mechanism responsible for maintaining gamma band activity based on persistent G-protein activation, separate intracellular pathways that differentiate between gamma band activity during waking vs during REM sleep, and an intracellular mechanism responsible for the dysregulation in gamma band activity in schizophrenia. These findings open several promising research avenues that have not been thoroughly explored. What are the effects of sleep or REM sleep deprivation on these RAS mechanisms? Are these mechanisms involved in memory processing during waking and/or during REM sleep? Does gamma band processing differ during waking vs REM sleep after sleep or REM sleep deprivation? PMID:24309750

  3. Plant farnesyltransferase can restore yeast Ras signaling and mating

    SciTech Connect

    Yalovsky, S.; Callan, K.L.; Narita, J.O.

    1997-04-01

    Farnesyltransferase (FTase) is a heterodimeric enzyme that modifies a group of proteins, including Ras, in mammals and yeasts. Plant FTase {alpha} and {beta} subunits were cloned from tomato and expressed in the yeast Saccharomyces cerevisiae to assess their functional conservation in farnesylating Ras and a-factor proteins, which are important for cell growth and mating. The tomato FTase {beta} subunit (LeFTB) alone was unable to complement the growth defect of ram1{del} mutant yeast strains in which the chromosomal FTase {beta} subunit gene was deleted, but coexpression of LeFTB with the plant {alpha} subunit gene (LeFTA) restored normal growth, Ras membrane association, and mating. LeFTB contains a novel 66-amino-acid sequence domain whose deletion reduces the efficiency of tomato FTase to restore normal growth to yeast ram1{del} strains. Coexpression of LeFTA and LeFTB in either yeast or insect cells yielded a functional enzyme that correctly farnesylated CaaX-motif-containing peptides. Despite their low degree of sequence homology, yeast and plant FTases shared similar in vivo and in vitro substrate specificities, demonstrating that this enzymatic modification of proteins with intermediates from the isoprenoid biosynthesis pathway is conserved in evolutionarily divergent eukaryotes. 56 refs., 7 figs., 1 tab.

  4. Identification of Proteins that Modify Cataract of the Eye Lens

    PubMed Central

    Hoehenwarter, Wolfgang; Tang, Yajun; Ackermann, Renate; Pleissner, Klaus-Peter; Schmid, Monika; Stein, Robert; Zimny-Arndt, Ursula; Kumar, Nalin M.; Jungblut, Peter R.

    2010-01-01

    The occurrence of a nuclear cataract in the eye lens due to disruption of theα3Cx46 connexin gene, Gja3, is dependent on strain background in a mouse model, implicating factors that modify the pathology. The differences upon cataractogenesis in the urea soluble proteins of the lens of two mouse strains, C57BL/6J and 129/SvJ, were analyzed by a comparative proteomics approach. Determination of the complete proteome of an organ offers the opportunity to characterize at a molecular level, differences in gene expression and post-translational modifications occurring during pathology and between individuals. The abundance of 63 protein species was altered between the strains. A unique aspect of this study is the identification of chaperonin subunit 6A, mortalin, ERp29 and syntaxin binding protein 6 in the eye lens. DNA polymorphisms resulting in non-conservative amino acid changes that led to altered physicochemical properties of the proteins were detected for mortalin, chaperonin subunit 6A, annexin A1 and possibly gamma N crystallin. The results show HSP27/25 and/or ERp29 are the likely major modifying factors for cataractogenesis. Extension of the results suggests that small heat shock proteins have a major role for influencing cataract formation in humans. PMID:19003866

  5. Regulation of autophagy and chloroquine sensitivity by oncogenic RAS in vitro is context-dependent.

    PubMed

    Morgan, Michael J; Gamez, Graciela; Menke, Christina; Hernandez, Ariel; Thorburn, Jacqueline; Gidan, Freddi; Staskiewicz, Leah; Morgan, Shellie; Cummings, Christopher; Maycotte, Paola; Thorburn, Andrew

    2014-10-01

    Chloroquine (CQ) is an antimalarial drug and late-stage inhibitor of autophagy currently FDA-approved for use in the treatment of rheumatoid arthritis and other autoimmune diseases. Based primarily on its ability to inhibit autophagy, CQ and its derivative, hydroxychloroquine, are currently being investigated as primary or adjuvant therapy in multiple clinical trials for cancer treatment. Oncogenic RAS has previously been shown to regulate autophagic flux, and cancers with high incidence of RAS mutations, such as pancreatic cancer, have been described in the literature as being particularly susceptible to CQ treatment, leading to the hypothesis that oncogenic RAS makes cancer cells dependent on autophagy. This autophagy "addiction" suggests that the mutation status of RAS in tumors could identify patients who would be more likely to benefit from CQ therapy. Here we show that RAS mutation status itself is unlikely to be beneficial in such a patient selection because oncogenic RAS does not always promote autophagy addiction. Moreover, oncogenic RAS can have opposite effects on both autophagic flux and CQ sensitivity in different cells. Finally, for any given cell type, the positive or negative effect of oncogenic RAS on autophagy does not necessarily predict whether RAS will promote or inhibit CQ-mediated toxicity. Thus, although our results confirm that different tumor cell lines display marked differences in how they respond to autophagy inhibition, these differences can occur irrespective of RAS mutation status and, in different contexts, can either promote or reduce chloroquine sensitivity of tumor cells.

  6. Ca2+/calmodulin binds and dissociates K-RasB from membrane.

    PubMed

    Sidhu, Ranjinder S; Clough, Richard R; Bhullar, Rajinder P

    2003-05-16

    We have investigated the interaction of calmodulin (CaM) with Ras-p21 and the significance of this association. All Ras-p21 isoforms tested (H-, K-, and N-Ras) were detected in the particulate fraction of human platelets and MCF-7 cells, a human breast cancer cell line. In MCF-7 cells, H- and N-Ras were also detected in the cytosolic fraction. K-RasB from platelet and MCF-7 cell lysates was found to bind CaM in a Ca2+ -dependent but GTPgammaS-independent manner. The yeast two-hybrid analysis demonstrated that K-RasB binds to CaM in vivo. Incubation of isolated membranes from platelet and MCF-7 cells with CaM caused dissociation of only K-RasB from membranes in a Ca2+ -dependent manner. CaM antagonist, W7, inhibited dissociation of K-RasB. Addition of platelet or MCF-7 cytosol alone to isolated platelet membranes did not cause dissociation of K-RasB and only addition of exogenous CaM caused dissociation. The results suggest a potential role for Ca2+/CaM in the regulation of K-RasB function.

  7. B- and C-RAF display essential differences in their binding to Ras: the isotype-specific N terminus of B-RAF facilitates Ras binding.

    PubMed

    Fischer, Andreas; Hekman, Mirko; Kuhlmann, Jürgen; Rubio, Ignacio; Wiese, Stefan; Rapp, Ulf R

    2007-09-01

    Recruitment of RAF kinases to the plasma membrane was initially proposed to be mediated by Ras proteins via interaction with the RAF Ras binding domain (RBD). Data reporting that RAF kinases possess high affinities for particular membrane lipids support a new model in which Ras-RAF interactions may be spatially restricted to the plane of the membrane. Although the coupling features of Ras binding to the isolated RAF RBD were investigated in great detail, little is known about the interactions of the processed Ras with the functional and full-length RAF kinases. Here we present a quantitative analysis of the binding properties of farnesylated and nonfarnesylated H-Ras to both full-length B- and C-RAF in the presence and absence of lipid environment. Although isolated RBD fragments associate with high affinity to both farnesylated and nonfarnesylated H-Ras, the full-length RAF kinases revealed fundamental differences with respect to Ras binding. In contrast to C-RAF that requires farnesylated H-Ras, cytosolic B-RAF associates effectively and with significantly higher affinity with both farnesylated and nonfarnesylated H-Ras. To investigate the potential farnesyl binding site(s) we prepared several N-terminal fragments of C-RAF and found that in the presence of cysteine-rich domain only the farnesylated form of H-Ras binds with high association rates. The extreme N terminus of B-RAF turned out to be responsible for the facilitation of lipid independent Ras binding to B-RAF, since truncation of this region resulted in a protein that changed its kinase properties and resembles C-RAF. In vivo studies using PC12 and COS7 cells support in vitro results. Co-localization measurements using labeled Ras and RAF documented essential differences between B- and C-RAF with respect to association with Ras. Taken together, these data suggest that the activation of B-RAF, in contrast to C-RAF, may take place both at the plasma membrane and in the cytosolic environment.

  8. Presence of Ras guanyl nucleotide-releasing protein in striosomes of the mature and developing rat.

    PubMed

    Pierret, P; Mechawar, N; Vallée, A; Patel, J; Priestley, J V; Dunn, R J; Dower, N A; Stone, J C; Richardson, P M

    2002-01-01

    Ras signal transduction pathways have been implicated as key regulators in neuroplasticity and synaptic transmission in the brain. These pathways can be modulated by Ras guanyl nucleotide exchange factors, (GEF) which activate Ras proteins by catalysing the exchange of GDP for GTP. Ras guanyl nucleotide-releasing protein (RasGRP), a recently discovered Ras GEF, that links diacylglycerol and probably calcium to Ras signaling pathways, is expressed in brain as well as in T-cells. Here, we have used a highly selective monoclonal antibody against RasGRP to localize this protein within the striatum and related forebrain structures of developing and adult rats. RasGRP immunolabeling was found to be widespread in the mature and developing rat forebrain. Most notably, it presented a prominent patchy distribution throughout the striatum at birth and at all postnatal ages examined. These patches were found to correspond with the striosomal compartment of the striatum, as identified by micro-opioid receptor labeling in the adult. RasGRP-immunoreactivity was also observed in the matrix-like compartment surrounding these patches/striosomes but appeared later in development and was always weaker than in the patches. In both striatal compartments, RasGRP was exclusively expressed by medium-sized spiny neurons and showed no preference for neurons that project either directly or indirectly to the substantia nigra. At the ultrastructural level, immunogold labeling of RasGRP was confined to the cell bodies and dendritic shafts of these output neurons. We conclude that the prominent expression of RasGRP in striosomes may be of significance for diacylglycerol signaling in the striatum, and could be of importance for the processing of limbic-related activity within the basal ganglia.

  9. Geomorphic changes in Ras Al-Subiyah area, Kuwait

    NASA Astrophysics Data System (ADS)

    Al Hurban, A.; El-Gamily, H.; El-Sammak, A.

    2008-06-01

    The Ras Al-Subiyah area is considered one of the most promising areas in Kuwait for future development. This development will include a new town called Subiyah and its associated infrastructure. This area is also being considered as the location for connection between Boubyan Island, which is now undergoing major development and the Kuwait mainland. The present study investigates the geomorphology of the Ras Al-Sabiyah area in the northern sector of Kuwait. The study area is generally flat, and it is located west of the Jal Az-Zor escarpment. It is bordered on the east by the Khor Al-Sabiyah tidal channel and on the south by Kuwait Bay. The area receives sediments from several sources; currently the most important are aeolian sediments and the deposition of mud delivered through the Khor Al-Sabiyah from the Iraqi marshes. The study area has been subjected to severe environmental changes due to the Gulf wars and the drainage of Iraqi marshes and the associated artificial changes in fluvial system. Twenty-two surface sediments were collected from the Ras Al-Subiyah area. Samples were collected to include the main geomorphologic characteristic features of the study area. Field observations and remote sensing images from 1990 and 2001 were used to produce an updated geomorphologic map for the Ras Al-Subiyah and a map showing geomorphic changes between 1990 and 2001. Grain size of the surface sediment ranges from gravel to medium sand. In general, grain size statistical analysis indicates that most of the areas are composed of two or more classes of sands transported and deposited from different sources including aeolian, sabkhas, river and the bays. The variability in the grain size statistical parameters may be attributed to the complexity of surface morphology as well as the diversity in the type of depositional environment in the Ras Al-Subiyah area. The total area subjected to change during the 12-year period (1990 2001) is about 32 km2 as calculated using GIS

  10. The role of lens epithelium in sugar cataract formation.

    PubMed

    Robison, W G; Houlder, N; Kinoshita, J H

    1990-06-01

    Previous evidence has shown clearly that sugar cataract formation results from unusually high intracellular levels of polyol. Documentation of polyol-related histological changes in the cortical fiber cells of the equatorial zone has been extensive. However, little attention has been given to the early changes in the lens epithelial cells, in spite of the fact that the highest level of aldose reductase is found in this layer of the lens. Also, cultured lens epithelial cells exposed to high sugar levels exhibit rapid accumulation of polyol and show ultrastructural alterations. Therefore, a study was designed to evaluate the role of the lens epithelium in sugar cataract formation. Specifically, an attempt was made to localize the earliest fine structural lesions in intact lenses of galactose-fed rats and to test their relation to aldose reductase. Rats were fed either a normal diet or a 50% galactose diet with or without sorbinil, an aldose reductase inhibitor. Rats were killed at varying periods of time ranging from 6 to 96 hr, and the eyes were processed for light and electron microscopy. The first detectable abnormalities occurred after 36 hr of galactose feeding, and were limited to the central lens epithelium. Cell edema, apparent dilution of cytoplasm, rounding of nuclei, aberrant intracellular vacuoles, and loss of normal tortuosity of cell boundaries were the salient lesions. No changes were detectable in the equatorial zone until 48 hr, and no deviation from the control structure was found in any of the rats treated with an aldose reductase inhibitor.(ABSTRACT TRUNCATED AT 250 WORDS)

  11. A stochastic model of eye lens growth.

    PubMed

    Šikić, Hrvoje; Shi, Yanrong; Lubura, Snježana; Bassnett, Steven

    2015-07-01

    The size and shape of the ocular lens must be controlled with precision if light is to be focused sharply on the retina. The lifelong growth of the lens depends on the production of cells in the anterior epithelium. At the lens equator, epithelial cells differentiate into fiber cells, which are added to the surface of the existing fiber cell mass, increasing its volume and area. We developed a stochastic model relating the rates of cell proliferation and death in various regions of the lens epithelium to deposition of fiber cells and radial lens growth. Epithelial population dynamics were modeled as a branching process with emigration and immigration between proliferative zones. Numerical simulations were in agreement with empirical measurements and demonstrated that, operating within the strict confines of lens geometry, a stochastic growth engine can produce the smooth and precise growth necessary for lens function. PMID:25816743

  12. Formation of Micro Lens by Laser Polymerization

    NASA Astrophysics Data System (ADS)

    Mori, Akira; Horiuchi, Takashi; Mizumachi, Manabu; Seino, Satoshi; Nakagawa, Takuya; Suzuki, Kaoru

    Recently, a micro lens has been demanded in uniting a laser device and an optical fiber. We have fabricated a new type of plastic micro lens by laser polymerization. The amount of the resin polymerized by exposing laser light, namely light-curing, depends on the laser power and exposing time. The shape of the lens can be controlled by changing the condition of laser irradiation. In this paper, the characteristic of the lens formed by this method was examined. Moreover, the relation between the lens shape and the condition of laser irradiation was investigated, and the condition to reducing a transverse spherical aberration was examined. As the result, the lens of 390μm in diameter was formed. The area which can be used for light coupling from a laser diode to a multimode fiber will be 81 % in the total lens area.

  13. Repression of CD24 surface protein expression by oncogenic Ras is relieved by inhibition of Raf but not MEK or PI3K

    PubMed Central

    Pallegar, Nikitha K.; Ayre, D. Craig; Christian, Sherri L.

    2015-01-01

    CD24 is a dynamically regulated cell surface protein. High expression of CD24 leads to progression of lung, prostrate, colon, and pancreatic cancers, among others. In contrast, low expression of CD24 leads to cell proliferation and metastasis of breast cancer stem cells (BCSCs). Activating mutations in Ras are found in 30% of all human cancers. Oncogenic Ras constitutively stimulates the Raf, PI3K, and Ral GDS signaling pathways, leading to cellular transformation. Previous studies have shown that expression of oncogenic Ras in breast cancer cells generates CD24− cells from CD24+ cells. However, the molecular mechanisms involved in the generation of CD24− cells were not determined. Here, we demonstrate that oncogenic Ras (RasV12) expression suppresses CD24 mRNA, protein, and promoter levels when expressed in NIH/3T3 cells. Furthermore, activation of only the Raf pathway was sufficient to downregulate CD24 mRNA and protein expression to levels similar to those seen in with RasV12 expression. In contrast, activation of the PI3K pathway downregulated mRNA expression with a partial effect on protein expression whereas activation of the RalGDS pathway only partially affected protein expression. Surprisingly, inhibition of MEK with U0126 only partially restored CD24 mRNA expression but not surface protein expression. In contrast, inhibition of Raf with sorafenib did not restore CD24 mRNA expression but significantly increased the proportion of RasV12 cells expressing CD24. Therefore, the Raf pathway is the major repressor of CD24 mRNA and protein expression, with PI3K also able to substantially inhibit CD24 expression. Moreover, these data indicate that the levels of CD24 mRNA and surface protein are independently regulated. Although inhibition of Raf by sorafenib only partially restored CD24 expression, sorafenib should still be considered as a potential therapeutic strategy to alter CD24 expression in CD24− cells, such as BCSCs. PMID:26301220

  14. Transformation with Oncogenic Ras and the Simian Virus 40 T Antigens Induces Caspase-Dependent Sensitivity to Fatty Acid Biosynthetic Inhibition

    PubMed Central

    Xu, Shihao; Spencer, Cody M.

    2015-01-01

    drive the transformation of normal cells to the cancerous state. These oncogenic alterations induce metabolic changes and dependencies that can be targeted to kill cancerous cells. Here, we find that the cellular transformation resulting from combined expression of the SV40 early region with an oncogenic Ras allele is sufficient to induce cellular susceptibility to fatty acid biosynthetic inhibition. Inhibition of fatty acid biosynthesis in these cells resulted in programmed cell death, which could be rescued by supplementing the medium with nonsaturated fatty acids. Similar results were observed with the expression of oncogenic Ras in nontransformed breast epithelial cells. Combined, our results suggest that specific oncogenic alleles induce metabolic dependencies that can be exploited to selectively kill cancerous cells. PMID:25855740

  15. Characterization of a novel oncogenic K-ras mutation in colon cancer

    SciTech Connect

    Akagi, Kiwamu . E-mail: akagi@cancer-c.pref.saitama.jp; Uchibori, Ryosuke; Yamaguchi, Kensei; Kurosawa, Keiko; Tanaka, Yoichiro; Kozu, Tomoko

    2007-01-19

    Activating mutations of RAS are frequently observed in subsets of human cancers, indicating that RAS activation is involved in tumorigenesis. Here, we identified and characterized a novel G to T transversion mutation of the K-ras gene at the third position of codon 19 (TTG) which substituted phenylalanine for leucine in 3 primary colon carcinomas. Biological and biochemical activity was examined using transformed NIH3T3 cells expressing mutant or wild-type K-ras. Transformants harboring the K-ras mutation at codon 19 showed proliferative capacity under serum-starved conditions, less contact inhibition, anchorage-independent growth, tumorigenicity in nude mice and elevation of active Ras-GTP levels. These results indicated that this novel mutation possesses high oncogenic activity.

  16. RasGRF1 regulates proliferation and metastatic behavior of human alveolar rhabdomyosarcomas.

    PubMed

    Tarnowski, Maciej; Schneider, Gabriela; Amann, Gabriele; Clark, Geoffrey; Houghton, Peter; Barr, Frederic G; Kenner, Lukas; Ratajczak, Mariusz Z; Kucia, Magda

    2012-09-01

    The involvement of the Ras superfamily of GTPases in the pathogenesis of rhabdomysarcoma (RMS) is not well understood. While mutant H-Ras leads to embryonal RMS (ERMS) formation in experimental animals and in Costello syndrome patients, no data exists on the potential role of Ras GTPases in the pathogenesis of alveolar RMS (ARMS). To address this issue better, we focused on the role of the GTP exchange factor RasGRF1 in this process. We observed that, in comparison to normal skeletal muscle cells, RasGRF1 mRNA is upregulated in the majority of human ARMS cell lines and subsequently confirmed its high expression in patient samples. By employing confocal microscopy analysis, we observed RasGRF1 accumulation in cell filopodia, which suggests its involvement in ARMS cell migration. Furthermore, we observed that RasGRF1 becomes phosphorylated in ARMS after stimulation by several pro-metastatic factors, such as SDF-1 and HGF/SF, as well as after exposure to growth-promoting Igf-2 and insulin. More importantly, activation of RasGRF1 expression correlated with activation of p42/44 MAPK and AKT. When the expression of RasGRF1 was down-regulated in ARMS cells by an shRNA strategy, these RasGRF1-kd RMS cells did not respond to stimulation by SDF-1, HGF/SF, Igf-2 or insulin by phosphorylation of p42/44 MAPK and AKT and lost their chemotactic responsiveness; however, their adhesion was not affected. We also observed that RasGRF1-kd ARMS cells proliferated at a very low rate in vitro, and, more importantly, after inoculation into immunodeficient SCID/beige inbred mice they formed significantly smaller tumors. We conclude that RasGRF1 plays an important role in ARMS pathogenesis and is a new potential therapeutic target to inhibit ARMS growth.

  17. A novel role for flotillin-1 in H-Ras-regulated breast cancer aggressiveness.

    PubMed

    Koh, Minsoo; Yong, Hae-Young; Kim, Eun-Sook; Son, Hwajin; Jeon, You Rim; Hwang, Jin-Sun; Kim, Myeong-Ok; Cha, Yujin; Choi, Wahn Soo; Noh, Dong-Young; Lee, Kyung-Min; Kim, Ki-Bum; Lee, Jae-Seon; Kim, Hyung Joon; Kim, Haemin; Kim, Hong-Hee; Kim, Eun Joo; Park, So Yeon; Kim, Hoe Suk; Moon, Woo Kyung; Choi Kim, Hyeong-Reh; Moon, Aree

    2016-03-01

    Elevated expression and aberrant activation of Ras have been implicated in breast cancer aggressiveness. H-Ras, but not N-Ras, induces breast cell invasion. A crucial link between lipid rafts and H-Ras function has been suggested. This study sought to identify the lipid raft protein(s) responsible for H-Ras-induced tumorigenicity and invasiveness of breast cancer. We conducted a comparative proteomic analysis of lipid raft proteins from invasive MCF10A human breast epithelial cells engineered to express active H-Ras and non-invasive cells expressing active N-Ras. Here, we identified a lipid raft protein flotillin-1 as an important regulator of H-Ras activation and breast cell invasion. Flotillin-1 was required for epidermal growth factor-induced activation of H-Ras, but not that of N-Ras, in MDA-MB-231 triple-negative breast cancer (TNBC) cells. Flotillin-1 knockdown inhibited the invasiveness of MDA-MB-231 and Hs578T TNBC cells in vitro and in vivo. In xenograft mouse tumor models of these TNBC cell lines, we showed that flotillin-1 played a critical role in tumor growth. Using human breast cancer samples, we provided clinical evidence for the metastatic potential of flotillin-1. Membrane staining of flotillin-1 was positively correlated with metastatic spread (p = 0.013) and inversely correlated with patient disease-free survival rates (p = 0.005). Expression of flotillin-1 was associated with H-Ras in breast cancer, especially in TNBC (p < 0.001). Our findings provide insight into the molecular basis of Ras isoform-specific interplay with flotillin-1, leading to tumorigenicity and aggressiveness of breast cancer.

  18. Chimeric proteins define variable and essential regions of Ha-ras-encoded protein

    SciTech Connect

    Lowe, D.G.; Ricketts, M.; Levinson, A.D.; Goeddel, D.V.

    1988-02-01

    The biological role of amino acid differences between the human 21-kDa Ha-ras protein (p21) and the human 23-kDa R-ras protein (p23) was investigated by engineering mutant Ha-ras p21 molecules containing divergent amino acid sequences from R-ras p23. Variant amino acids from R-ras p23 regions 1-30, 52-57, 67-78, 1-30 and 67-78 together, and 112-124 were substituted for the corresponding Ha-ras p21 amino acid regions 1-4, 26-31, 41-52, 1-4 and 41-52 together, and 86-98, respectively. Rat fibroblasts transfected with genes encoding these position-12 valine-substituted chimeric Ha-ras proteins displayed the same properties of morphological transformation and anchorage-independent growth as Ha-ras T24 oncogene-transformed fibroblasts. However, substitution of variant amino acids from the 80 C-terminal residues (amino acids 138-218) of R-ras p23 for the corresponding p21 amino acids (residues 112-189) inactivated the transforming activity of position-12 valine-substituted p21. The converse substitution of Ha-ras p21 C-terminal residues into R-ras p23 did not result in transformation by position-38 valine-substituted p232. These data are discussed in terms of the structure of ras proteins and the nature of interactions determining the specificity of effector function.

  19. The hypervariable region of K-Ras4B is responsible for its specific interactions with Calmodulin

    PubMed Central

    Abraham, Sherwin J.; Nolet, Ryan P.; Calvert, Richard J.; Anderson, Lucy M.; Gaponenko, Vadim

    2009-01-01

    K-Ras4B belongs to the family of p21 Ras GTPases, which play an important role in cell proliferation, survival and motility. The p21 Ras proteins such as K-Ras4B, K-Ras4A, H-Ras, and N-Ras, share 85% sequence homology and activate very similar signaling pathways. Only the C-terminal hypervariable regions differ significantly. A growing body of literature demonstrates that each Ras isoform possesses unique functions in normal physiological processes as well as in pathogenesis. One of the central questions in the field of Ras biology is how these very similar proteins achieve such remarkable specificity in protein-protein interactions that regulate signal transduction pathways. Here we explore specific binding of K-Ras4B to calmodulin. Using NMR techniques and isothermal titration calorimetry we demonstrate that the hypervariable region of K-Ras contributes in a major way to the interaction with calmodulin while the catalytic domain of K-Ras4B provides a way to control the interaction by nucleotide binding. The hypervariable region of K-Ras4B binds specifically to the C-terminal domain of Ca2+-loaded calmodulin with micromolar affinity, while the GTP-γ-S loaded catalytic domain of K-Ras4B may interact with the N-terminal domain of calmodulin. PMID:19583261

  20. The hypervariable region of K-Ras4B is responsible for its specific interactions with calmodulin.

    PubMed

    Abraham, Sherwin J; Nolet, Ryan P; Calvert, Richard J; Anderson, Lucy M; Gaponenko, Vadim

    2009-08-18

    K-Ras4B belongs to the family of p21 Ras GTPases, which play an important role in cell proliferation, survival, and motility. The p21 Ras proteins, such as K-Ras4B, K-Ras4A, H-Ras, and N-Ras, share 85% sequence homology and activate very similar signaling pathways. Only the C-terminal hypervariable regions differ significantly. A growing body of literature demonstrates that each Ras isoform possesses unique functions in normal physiological processes as well as in pathogenesis. One of the central questions in the field of Ras biology is how these very similar proteins achieve such remarkable specificity in protein-protein interactions that regulate signal transduction pathways. Here we explore specific binding of K-Ras4B to calmodulin. Using NMR techniques and isothermal titration calorimetry, we demonstrate that the hypervariable region of K-Ras4B contributes in a major way to the interaction with calmodulin, while the catalytic domain of K-Ras4B provides a way to control the interaction by nucleotide binding. The hypervariable region of K-Ras4B binds specifically to the C-terminal domain of Ca(2+)-loaded calmodulin with micromolar affinity, while the GTP-gamma-S-loaded catalytic domain of K-Ras4B may interact with the N-terminal domain of calmodulin.

  1. Lens Coupled Quantum Cascade Laser

    NASA Technical Reports Server (NTRS)

    Hu, Qing (Inventor); Lee, Alan Wei Min (Inventor)

    2013-01-01

    Terahertz quantum cascade (QC) devices are disclosed that can operate, e.g., in a range of about 1 THz to about 10 THz. In some embodiments, QC lasers are disclosed in which an optical element (e.g., a lens) is coupled to an output facet of the laser's active region to enhance coupling of the lasing radiation from the active region to an external environment. In other embodiments, terahertz amplifier and tunable terahertz QC lasers are disclosed.

  2. Influence of aging and caloric restriction on activation of Ras/MAPK, calcineurin, and CaMK-IV activities in rat T cells.

    PubMed

    Pahlavani, M A; Vargas, D M

    2000-02-01

    The signaling cascade mediated by Ras (p21ras) and MAPK (mitogen-activated protein kinase) and calcium/calmodulin regulating enzymes, calcineurin (CaN) and CaMK-IV, are considered to be essential for T-cell growth and function. In the present study, the effect of aging and caloric restriction (CR) on the induction of Ras and MAPK activation by concanavalin A (ConA) was studied. Splenic T cells were isolated from young (4-6 months) and old (22-24 months) rats that had free access to food (control group), and from caloric restricted old (22-24 months) rats that beginning at 6 weeks of age were fed 60%(40% caloric restriction) of the diet consumed by the control rats. We found that the induction of Ras activity in T cells isolated from control old rats was lower (P<0.001) than that in control young rats. However, the levels of Ras activity in T cells isolated from CR old rats were similar to the levels in the age-matched control rats. The induction of MAPK activity in T cells isolated from control old rats and CR old rats was significantly less than in T cells isolated from control young rats, and caloric restriction significantly (P<0.05) reduced the age-related decline in MAPK activation. We also measured the induction of CaN and CaMK-IV activities by ConA in T cells from control young and old and CR old rats. The induction of both CaN and CaMK-IV activity decreased with age. Caloric restriction significantly (P<0.05) reduced the age-related decline in CaN activity, but had no significant effect on CaMK-IV activity. The changes in Ras/MAPK activation and in CaN and CaMK-IV activity with age or with CR were not associated with alterations in their corresponding protein levels. Thus, caloric restriction has a differential effect on the activation of the upstream signaling molecules that are altered with age.

  3. Direct Attack on RAS: Intramolecular Communication and Mutation-Specific Effects.

    PubMed

    Marcus, Kendra; Mattos, Carla

    2015-04-15

    The crystal structure of RAS was first solved 25 years ago. In spite of tremendous and sustained efforts, there are still no drugs in the clinic that directly target this major driver of human cancers. Recent success in the discovery of compounds that bind RAS and inhibit signaling has fueled renewed enthusiasm, and in-depth understanding of the structure and function of RAS has opened new avenues for direct targeting. To succeed, we must focus on the molecular details of the RAS structure and understand at a high-resolution level how the oncogenic mutants impair function. Structural networks of intramolecular communication between the RAS active site and membrane-interacting regions on the G-domain are disrupted in oncogenic mutants. Although conserved across the isoforms, these networks are near hot spots of protein-ligand interactions with amino acid composition that varies among RAS proteins. These differences could have an effect on stabilization of conformational states of interest in attenuating signaling through RAS. The development of strategies to target these novel sites will add a fresh direction in the quest to conquer RAS-driven cancers. Clin Cancer Res; 21(8); 1810-8. ©2015 AACR. See all articles in this CCR Focus section, "Targeting RAS-Driven Cancers." PMID:25878362

  4. GILZ mediates the antiproliferative activity of glucocorticoids by negative regulation of Ras signaling

    PubMed Central

    Ayroldi, Emira; Zollo, Ornella; Bastianelli, Alessandra; Marchetti, Cristina; Agostini, Massimiliano; Di Virgilio, Rosa; Riccardi, Carlo

    2007-01-01

    Tsc22d3 coding for glucocorticoid-induced leucine zipper (GILZ) was initially identified as a dexamethasone-responsive gene involved in the control of T lymphocyte activation and apoptosis. However, the physiological role of this molecule and its function in the biological activity of glucocorticoids (GCs) has not been clarified. Here, we demonstrate that GILZ interacts directly with Ras in vitro and in vivo as shown by GILZ and Ras coimmunoprecipitation and colocalization upon PMA activation in primary mouse spleen T lymphocytes and thymus cells. The analysis of GILZ mutants showed that they bound Ras through the tuberous sclerosis complex box (TSC) and, depending on the Ras activation level, formed a trimeric complex with Ras and Raf, which we previously identified as a GILZ binder. As a consequence of these interactions, GILZ diminished the activation of Ras and Raf downstream targets including ERK1/2, AKT/PKB serine/threonine kinase, and retinoblastoma (Rb) phosphorylation and cyclin D1 expression, leading to inhibition of Ras- and Raf-dependent cell proliferation and Ras-induced NIH-3T3 transformation. GILZ silencing resulted in an increase in concanavalin A–induced T cell proliferation and, most notably, inhibition of dexamethasone antiproliferative effects. Together, these findings indicate that GILZ serves as a negative regulator of Ras- and Raf-induced proliferation and is an important mediator of the antiproliferative effect of GCs. PMID:17492054

  5. Ras-GTP dimers activate the mitogen-activated protein kinase (MAPK) pathway

    SciTech Connect

    Nan, Xiaolin; Tamgüney, Tanja M.; Collisson, Eric A.; Lin, Li -Jung; Pitt, Cameron; Galeas, Jacqueline; Lewis, Sophia; Gray, Joe W.; McCormick, Frank; Chu, Steven

    2015-06-16

    Rat sarcoma (Ras) GTPases regulate cell proliferation and survival through effector pathways including Raf-MAPK, and are the most frequently mutated genes in human cancer. Although it is well established that Ras activity requires binding to both GTP and the membrane, details of how Ras operates on the cell membrane to activate its effectors remain elusive. Efforts to target mutant Ras in human cancers to therapeutic benefit have also been largely unsuccessful. Here we show that Ras-GTP forms dimers to activate MAPK. We used quantitative photoactivated localization microscopy (PALM) to analyze the nanoscale spatial organization of PAmCherry1-tagged KRas 4B (hereafter referred to KRas) on the cell membrane under various signaling conditions. We found that at endogenous expression levels KRas forms dimers, and KRasG12D, a mutant that constitutively binds GTP, activates MAPK. Overexpression of KRas leads to formation of higher order Ras nanoclusters. Conversely, at lower expression levels, KRasG12D is monomeric and activates MAPK only when artificially dimerized. Moreover, dimerization and signaling of KRas are both dependent on an intact CAAX (C, cysteine; A, aliphatic; X, any amino acid) motif that is also known to mediate membrane localization. These results reveal a new, dimerization-dependent signaling mechanism of Ras, and suggest Ras dimers as a potential therapeutic target in mutant Ras-driven tumors.

  6. Nitrative and oxidative DNA damage caused by K-ras mutation in mice

    SciTech Connect

    Ohnishi, Shiho; Saito, Hiromitsu; Suzuki, Noboru; Ma, Ning; Hiraku, Yusuke; Murata, Mariko; Kawanishi, Shosuke

    2011-09-23

    Highlights: {yields} Mutated K-ras in transgenic mice caused nitrative DNA damage, 8-nitroguanine. {yields} The mutagenic 8-nitroguanine seemed to be generated by iNOS via Ras-MAPK signal. {yields} Mutated K-ras produces additional mutagenic lesions, as a new oncogenic role. -- Abstract: Ras mutation is important for carcinogenesis. Carcinogenesis consists of multi-step process with mutations in several genes. We investigated the role of DNA damage in carcinogenesis initiated by K-ras mutation, using conditional transgenic mice. Immunohistochemical analysis revealed that mutagenic 8-nitroguanine and 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) were apparently formed in adenocarcinoma caused by mutated K-ras. 8-Nitroguanine was co-localized with iNOS, eNOS, NF-{kappa}B, IKK, MAPK, MEK, and mutated K-ras, suggesting that oncogenic K-ras causes additional DNA damage via signaling pathway involving these molecules. It is noteworthy that K-ras mutation mediates not only cell over-proliferation but also the accumulation of mutagenic DNA lesions, leading to carcinogenesis.

  7. Mutant K-RAS Promotes Invasion and Metastasis in Pancreatic Cancer Through GTPase Signaling Pathways

    PubMed Central

    Padavano, Julianna; Henkhaus, Rebecca S; Chen, Hwudaurw; Skovan, Bethany A; Cui, Haiyan; Ignatenko, Natalia A

    2015-01-01

    Pancreatic ductal adenocarcinoma is one of the most aggressive malignancies, characterized by the local invasion into surrounding tissues and early metastasis to distant organs. Oncogenic mutations of the K-RAS gene occur in more than 90% of human pancreatic cancers. The goal of this study was to investigate the functional significance and downstream effectors of mutant K-RAS oncogene in the pancreatic cancer invasion and metastasis. We applied the homologous recombination technique to stably disrupt K-RAS oncogene in the human pancreatic cell line MiaPaCa-2, which carries the mutant K-RASG12C oncogene in both alleles. Using in vitro assays, we found that clones with disrupted mutant K-RAS gene exhibited low RAS activity, reduced growth rates, increased sensitivity to the apoptosis inducing agents, and suppressed motility and invasiveness. In vivo assays showed that clones with decreased RAS activity had reduced tumor formation ability in mouse xenograft model and increased survival rates in the mouse orthotopic pancreatic cancer model. We further examined molecular pathways downstream of mutant K-RAS and identified RhoA GTP activating protein 5, caveolin-1, and RAS-like small GTPase A (RalA) as key effector molecules, which control mutant K-RAS-dependent migration and invasion in MiaPaCa-2 cells. Our study provides rational for targeting RhoA and RalA GTPase signaling pathways for inhibition of pancreatic cancer metastasis. PMID:26512205

  8. Mechanisms of Membrane Binding of Small GTPase K-Ras4B Farnesylated Hypervariable Region*

    PubMed Central

    Jang, Hyunbum; Abraham, Sherwin J.; Chavan, Tanmay S.; Hitchinson, Ben; Khavrutskii, Lyuba; Tarasova, Nadya I.; Nussinov, Ruth; Gaponenko, Vadim

    2015-01-01

    K-Ras4B belongs to a family of small GTPases that regulates cell growth, differentiation and survival. K-ras is frequently mutated in cancer. K-Ras4B association with the plasma membrane through its farnesylated and positively charged C-terminal hypervariable region (HVR) is critical to its oncogenic function. However, the structural mechanisms of membrane association are not fully understood. Here, using confocal microscopy, surface plasmon resonance, and molecular dynamics simulations, we observed that K-Ras4B can be distributed in rigid and loosely packed membrane domains. Its membrane binding domain interaction with phospholipids is driven by membrane fluidity. The farnesyl group spontaneously inserts into the disordered lipid microdomains, whereas the rigid microdomains restrict the farnesyl group penetration. We speculate that the resulting farnesyl protrusion toward the cell interior allows oligomerization of the K-Ras4B membrane binding domain in rigid microdomains. Unlike other Ras isoforms, K-Ras4B HVR contains a single farnesyl modification and positively charged polylysine sequence. The high positive charge not only modulates specific HVR binding to anionic phospholipids but farnesyl membrane orientation. Phosphorylation of Ser-181 prohibits spontaneous farnesyl membrane insertion. The mechanism illuminates the roles of HVR modifications in K-Ras4B targeting microdomains of the plasma membrane and suggests an additional function for HVR in regulation of Ras signaling. PMID:25713064

  9. Multiple cellular proteins modulate the dynamics of K-ras association with the plasma membrane.

    PubMed

    Bhagatji, Pinkesh; Leventis, Rania; Rich, Rebecca; Lin, Chen-ju; Silvius, John R

    2010-11-17

    Although specific proteins have been identified that regulate the membrane association and facilitate intracellular transport of prenylated Rho- and Rab-family proteins, it is not known whether cellular proteins fulfill similar roles for other prenylated species, such as Ras-family proteins. We used a previously described method to evaluate how several cellular proteins, previously identified as potential binding partners (but not effectors) of K-ras4B, influence the dynamics of K-ras association with the plasma membrane. Overexpression of either PDEδ or PRA1 enhances, whereas knockdown of either protein reduces, the rate of dissociation of K-ras from the plasma membrane. Inhibition of calmodulin likewise reduces the rate of K-ras dissociation from the plasma membrane, in this case in a manner specific for the activated form of K-ras. By contrast, galectin-3 specifically reduces the rate of plasma membrane dissociation of activated K-ras, an effect that is blocked by the K-ras antagonist farnesylthiosalicylic acid (salirasib). Multiple cellular proteins thus control the dynamics of membrane association and intercompartmental movement of K-ras to an important degree even under basal cellular conditions.

  10. Ras-GTP dimers activate the Mitogen-Activated Protein Kinase (MAPK) pathway

    PubMed Central

    Nan, Xiaolin; Tamgüney, Tanja M.; Collisson, Eric A.; Lin, Li-Jung; Pitt, Cameron; Galeas, Jacqueline; Lewis, Sophia; Gray, Joe W.; McCormick, Frank; Chu, Steven

    2015-01-01

    Rat sarcoma (Ras) GTPases regulate cell proliferation and survival through effector pathways including Raf-MAPK, and are the most frequently mutated genes in human cancer. Although it is well established that Ras activity requires binding to both GTP and the membrane, details of how Ras operates on the cell membrane to activate its effectors remain elusive. Efforts to target mutant Ras in human cancers to therapeutic benefit have also been largely unsuccessful. Here we show that Ras-GTP forms dimers to activate MAPK. We used quantitative photoactivated localization microscopy (PALM) to analyze the nanoscale spatial organization of PAmCherry1-tagged KRas 4B (hereafter referred to KRas) on the cell membrane under various signaling conditions. We found that at endogenous expression levels KRas forms dimers, and KRasG12D, a mutant that constitutively binds GTP, activates MAPK. Overexpression of KRas leads to formation of higher order Ras nanoclusters. Conversely, at lower expression levels, KRasG12D is monomeric and activates MAPK only when artificially dimerized. Moreover, dimerization and signaling of KRas are both dependent on an intact CAAX (C, cysteine; A, aliphatic; X, any amino acid) motif that is also known to mediate membrane localization. These results reveal a new, dimerization-dependent signaling mechanism of Ras, and suggest Ras dimers as a potential therapeutic target in mutant Ras-driven tumors. PMID:26080442

  11. Formation of the Ras dimer is essential for Raf-1 activation.

    PubMed

    Inouye, K; Mizutani, S; Koide, H; Kaziro, Y

    2000-02-11

    Although it is well established that Ras requires membrane localization for activation of its target molecule, Raf-1, the reason for this requirement is not fully understood. In this study, we found that modified Ras, which is purified from Sf9 cells, could activate Raf-1 in a cell-free system, when incorporated into liposome. Using a bifunctional cross-linker and a protein-fragmentation complementation assay, we detected dimer formation of Ras in the liposome and in the intact cells, respectively. These results suggest that dimerization of Ras in the lipid membrane is essential for activation of Raf-1. To support this, we found that, when fused to glutathione S-transferase (GST), unprocessed Ras expressed in Escherichia coli could bypass the requirement for liposome. A Ras-dependent Raf-1 activator, which we previously reported (Mizutani, S., Koide, H., and Kaziro, Y. (1998) Oncogene 16, 2781-2786), was still required for Raf-1 activation by GST-Ras. Furthermore, an enforced dimerization of unmodified oncogenic Ras mutant in human embryonic kidney (HEK) 293 cells, using a portion of gyrase B or estrogen receptor, also resulted in activation of Raf-1. From these results, we conclude that membrane localization allows Ras to form a dimer, which is essential, although not sufficient, for Raf-1 activation.

  12. A primary cardiac leiomyosarcoma with mutation at H-ras codon 12.

    PubMed

    Parissis, J; Arvanitis, D; Sourvinos, G; Spandidos, D

    1997-01-01

    The presence of activating ras mutations in a cardiac leiomyosarcoma which occurred in the right atrium of the heart of a female patient was examined. The tumor had the appearance of leiomyosarcoma in rutine histopathological examination and the definite diagnosis was confirmed by a positive immunohistochemical reaction to smooth muscle actin. Molecular analysis by polymerase chain reaction (PCR) restriction fragment length polymorphism (RFLP) technique showed a point mutation of H-ras gene at codon 12. To the best of our knowledge, this is the first report describing ras gene mutation in a cardiac leiomyosarcoma implying a role for the ras oncogenes in the development of this tumor.

  13. Ras-GTP dimers activate the mitogen-activated protein kinase (MAPK) pathway

    DOE PAGESBeta

    Nan, Xiaolin; Tamgüney, Tanja M.; Collisson, Eric A.; Lin, Li -Jung; Pitt, Cameron; Galeas, Jacqueline; Lewis, Sophia; Gray, Joe W.; McCormick, Frank; Chu, Steven

    2015-06-16

    Rat sarcoma (Ras) GTPases regulate cell proliferation and survival through effector pathways including Raf-MAPK, and are the most frequently mutated genes in human cancer. Although it is well established that Ras activity requires binding to both GTP and the membrane, details of how Ras operates on the cell membrane to activate its effectors remain elusive. Efforts to target mutant Ras in human cancers to therapeutic benefit have also been largely unsuccessful. Here we show that Ras-GTP forms dimers to activate MAPK. We used quantitative photoactivated localization microscopy (PALM) to analyze the nanoscale spatial organization of PAmCherry1-tagged KRas 4B (hereafter referredmore » to KRas) on the cell membrane under various signaling conditions. We found that at endogenous expression levels KRas forms dimers, and KRasG12D, a mutant that constitutively binds GTP, activates MAPK. Overexpression of KRas leads to formation of higher order Ras nanoclusters. Conversely, at lower expression levels, KRasG12D is monomeric and activates MAPK only when artificially dimerized. Moreover, dimerization and signaling of KRas are both dependent on an intact CAAX (C, cysteine; A, aliphatic; X, any amino acid) motif that is also known to mediate membrane localization. These results reveal a new, dimerization-dependent signaling mechanism of Ras, and suggest Ras dimers as a potential therapeutic target in mutant Ras-driven tumors.« less

  14. CARD9 mediates Dectin-1–induced ERK activation by linking Ras-GRF1 to H-Ras for antifungal immunity

    PubMed Central

    Tang, Bing; Zhu, Le-Le; Liu, Yan-Hui; Zhao, Xue-Qiang; Gorjestani, Sara; Hsu, Yen-Michael S.; Yang, Long; Guan, Jian-Hong; Xu, Guo-Tong

    2014-01-01

    Dectin-1 functions as a pattern recognition receptor for sensing fungal infection. It has been well-established that Dectin-1 induces innate immune responses through caspase recruitment domain-containing protein 9 (CARD9)–mediated NF-κB activation. In this study, we find that CARD9 is dispensable for NF-κB activation induced by Dectin-1 ligands, such as curdlan or Candida albicans yeast. In contrast, we find that CARD9 regulates H-Ras activation by linking Ras-GRF1 to H-Ras, which mediates Dectin-1–induced extracellular signal-regulated protein kinase (ERK) activation and proinflammatory responses when stimulated by their ligands. Mechanistically, Dectin-1 engagement initiates spleen tyrosine kinase (Syk)–dependent Ras-GRF1 phosphorylation, and the phosphorylated Ras-GRF1 recruits and activates H-Ras through forming a complex with CARD9, which leads to activation of ERK downstream. Finally, we show that inhibiting ERK activation significantly accelerates the death of C. albicans–infected mice, and this inhibitory effect is dependent on CARD9. Together, our studies reveal a molecular mechanism by which Dectin-1 induces H-Ras activation that leads to ERK activation for host innate immune responses against fungal infection. PMID:25267792

  15. New Scleral Lens For Electroretinography

    NASA Astrophysics Data System (ADS)

    Charlier, J. R.; Grall, Y.; Legargasson, J. F.

    1986-05-01

    This paper presents a scleral lens specifically designed for the implementation of new electroretinography (E.R.G.) procedures including ganzeld ERG, pattern ERG and optic fiber ERG. Ganzfeld ERG requires a direct, uniform illumination of the retina and is usually obtained within a ball stimulation which provides precisely controlled stimulation conditions. Pattern ERG is related to the electrical activity of ganglion cells and is produced by a structured visual stimulus, for instance a cherkerboard reversal. Fiber optics stimulation is a promising new technique involving an optic fiber which connects the scleral lens to a remote light stimulator. The new scleral lens is made out of silicon. Silicon is an ideal material for ERG applications. It is highly permeable to oxygen. It eliminates the risk of corneal abrasion and it provides good comfort to the patient. Two layers of conductive silicon are used for recording the bioelectric potential difference of the cornea with respect to the eye lids. An optically transparent window allows for the projection of structured images on the retina. A wide angle conic aperture is provided for ganzfeld stimulation. The same cone is used as a blepharostat and as a plug for connecting the optic fiber, once the transparent window is centered with respect to the pupil entrance.

  16. Aquaporin 0 plays a pivotal role in refractive index gradient development in mammalian eye lens to prevent spherical aberration

    SciTech Connect

    Kumari, S. Sindhu; Varadaraj, Kulandaiappan

    2014-10-03

    . Transmission and scanning electron micrographs of lenses of both mouse models showed increased extracellular space between fiber cells. Water content determination study showed increase in water in the lenses of these mouse models. In summary, lens transparency, CTCA and compact packing of fiber cells were affected due to the loss of 50% AQP0 leading to larger extracellular space, more water content and SA, possibly due to alteration in RING. To our knowledge, this is the first report identifying the role of AQP0 in RING development to ward off lens SA during focusing.

  17. Sp1 binding plays a critical role in Erb-B2- and v-ras-mediated downregulation of alpha2-integrin expression in human mammary epithelial cells.

    PubMed Central

    Ye, J; Xu, R H; Taylor-Papadimitriou, J; Pitha, P M

    1996-01-01

    The human alpha2-integrin gene is transcriptionally downregulated in a nontumorigenic human mammary epithelial cell line, MTSV1-7, and its clonal variant HB2, overexpressing the Erb-B2 oncogene. In this study, we have used deletion mutations within the alpha2-integrin promoter inserted 5' of the chloramphenicol acetyltransferase or luciferase reporter genes to identify the element that is responsible for the Erb-B2-mediated downregulation. The results of the transient-transfection assay showed that the Sp1 binding element located in the core region (positions --64 to +1) of the alpha2-integrin promoter plays an essential role in the alpha2-integrin promoter activity and its downregulation by Erb-B2. By gel shift assay, we have demonstrated that this element binds with a high degree of affinity not only to Sp1, but also to Sp3. The downregulation of the alpha2-integrin promoter activity could also be achieved by overexpression of v-Hras (v-ras), suggesting that the signals generated by Erb-B2, which lead to downregulation of the alpha2-integrin gene expression, may proceed through the ras pathway. Both the Erb-B2- and the v-ras-overexpressing cells exhibited a Sp1 DNA binding activity lower than that of the parental line, while the relative levels of Sp1 protein in these cells were not altered. The Erb-B2- and v-ras-mediated downregulation could be reversed by the overexpression of Sp1 and by a dominant negative variant of ras (rasN17), confirming the importance of Sp1 and the ras pathway. The inhibitory effects of Erb-B2 on transcriptional activity of the alpha2-integrin promoter were observed in transient-cotransfection assays using alpha2-integrin reporter plasmids and plasmids expressing the Erb-B2 or v-ras oncogene. The same effects were seen when an alpha2-integrin reporter gene construct was transfected into MTSV1-7 or HB2 cells permanently overexpressing Erb-B2 or v-ras. The effects of Erb-B2 or v-ras on the transcriptional activity of the alpha2-integrin

  18. Rethinking contact lens associated keratitis.

    PubMed

    Efron, Nathan; Morgan, Philip B

    2006-09-01

    This review presents a critical analysis of the literature relating to the use of binomial and polynomial classification schemes for categorising corneal infiltrative events (CIEs) associated with contact lens wear and the epidemiology of such events. The results of the Manchester Keratitis Study-a 12-month, prospective, hospital-based epidemiological study of contact lens wearer suffering from CIEs-are used as a tool to challenge and test traditional thinking in relation to contact lens associated keratitis. An innovative aspect of this study is the use of a novel clinical severity matrix to systematically score the severity of CIEs based on 10 key signs and symptoms. The ambiguities inherent in using binomial classification schemes (such as, microbial versus sterile, ulcerative versus non-ulcerative etcetera) are highlighted. The failure of a polynomial scheme-due to extensive classification overlap between proposed sub-types of CIEs-is demonstrated using a Venn diagram. A cartographic analysis reveals that infiltrates tend to occur in the superior cornea of patients wearing extended wear silicone hydrogel lenses, in the central cornea of patients wearing daily wear hydrogel daily disposable lenses and in the peripheral cornea of patients wearing daily wear hydrogel (excluding daily disposable) lenses. Infiltrates that occur more towards the limbus are less severe. The incidence of CIEs is higher when contact lenses are worn overnight. Logistic analysis reveals that the risk of developing a severe CIE is five times greater with conventional hydrogel extended wear versus silicone hydrogel extended wear. The male gender, smoking, a healthy eye and body, and the late Winter months are associated with an increased risk of developing CIEs. The rate of significant visual loss as a result of developing a CIE is low. Two key conclusions are drawn from this work, which represent a radical rethinking of this potentially sight-threatening condition. CIEs should be

  19. H-Ras Increases Urokinase Expression and Cell Invasion in Genetically Modified Human Astrocytes Through Ras/Raf/MEK Signaling Pathway

    PubMed Central

    ZHAO, YUNGE; XIAO, AIZHEN; DIPIERRO, CHARLES G.; ABDEL-FATTAH, RANA; AMOS, SAMSON; REDPATH, GERARD T.; CARPENTER, JOAN E.; PIEPER, RUSSELL O.; HUSSAINI, ISA M.

    2008-01-01

    Previous study reported that the activation of Ras pathway cooperated with E6/E7-mediated inactivation of p53/pRb to transform immortalized normal human astrocytes (NHA/hTERT) into intracranial tumors strongly resembling human astrocytomas. The mechanism of how H-Ras contributes to astrocytoma formation is unclear. Using genetically modified NHA cells (E6/E7/hTERT and E6/E7/hTERT/Ras cells) as models, we investigated the mechanism of Ras-induced tumorigenesis. The overexpression of constitutively active H-RasV12 in E6/E7/hTERT cells robustly increased the levels of urokinase plasminogen activator (uPA) mRNA, protein, activity and invasive capacity of the E6/E7/hTERT/Ras cells. However, the expressions of MMP-9 and MMP-2 did not significantly change in the E6/E7/hTERT and E6/E7/hTERT/Ras cells. Furthermore, E6/E7/hTERT/Ras cells also displayed higher level of uPA activity and were more invasive than E6/E7/hTERT cells in 3D culture, and formed an intracranial tumor mass in a NOD-SCID mouse model. uPA specific inhibitor (B428) and uPA neutralizing antibody decreased uPA activity and invasion in E6/E7/hTERT/Ras cells. uPA-deficient U-1242 glioblastoma cells were less invasive in vitro and exhibited reduced tumor growth and infiltration into normal brain in xenograft mouse model. Inhibitors of Ras (FTA), Raf (Bay 54−9085) and MEK (UO126), but not of phosphatidylinositol 3-kinase (PI3K) (LY294002) and of protein kinase C (BIM) pathways, inhibited uPA activity and cell invasion. Our results suggest that H-Ras increased uPA expression and activity via the Ras/Raf/MEK signaling pathway leading to enhanced cell invasion and this may contribute to increased invasive growth properties of astrocytomas. PMID:18383343

  20. An Active RFID Accountability System (RAS) for Constrained Wireless Environments

    SciTech Connect

    Barker, Alan M; Hanson, Gregory R; Sexton, Angela Kay; Jones Jr, J P; Freer, Eva B; Sjoreen, Andrea L

    2011-01-01

    A team from Oak Ridge National Laboratory (ORNL) has developed an RFID Accountability System (RAS) that allows items with active RFID tags to be tracked in environments where tags may not be able to transmit their location continuously. The system uses activators that transmit a short range signal. Active RFID tags are in a sleep state until they encounter an activator. Then they transmit a signal that is picked up by the antennas installed throughout the building. This paper presents the theory of operation, application areas, lessons learned, and key features developed over the course of seven years of development and use.

  1. Ras protein expression as a marker for breast cancer

    PubMed Central

    CALAF, GLORIA M.; ABARCA-QUINONES, JORGE

    2016-01-01

    Breast cancer, the most common neoplasm in women of all ages, is the leading cause of cancer-related mortality in women worldwide. Markers to help to predict the risk of progression and ultimately provide non-surgical treatment options would be of great benefit. At present, there are no available molecular markers to predict the risk of carcinoma in situ progression to invasive cancer; therefore, all women diagnosed with this type of malignancy must undergo surgery. Breast cancer is a heterogeneous complex disease, and different patients respond differently to different treatments. In breast cancer, analysis using immunohistochemical markers remains an essential component of routine pathological examinations, and plays an import role in the management of the disease by providing diagnostic and prognostic strategies. The aim of the present study was to identify a marker that can be used as a prognostic tool for breast cancer. For this purpose, we firstly used an established breast cancer model. MCF-10F, a spontaneously immortalized breast epithelial cell line was transformed by exposure to estrogen and radiation. MCF-10F cells were exposed to low doses of high linear energy transfer (LET) α particles (150 keV/μm) of radiation, and subsequently cultured in the presence of 17β-estradiol. Three cell lines were used: i) MCF-10F cells as a control; ii) Alpha5 cells, a malignant and tumorigenic cell line; and iii) Tumor2 cells derived from Alpha5 cells injected into nude mice. Secondly, we also used normal, benign and malignant breast specimens obtained from biopsies. The results revealed that the MCF-10F cells were negative for c-Ha-Ras protein expression; however, the Alpha5 and Tumor2 cell lines were positive for c-Ha-Ras protein expression. The malignant breast samples were also strongly positive for c-Ha-Ras expression. The findings of our study indicate that c-Ha-Ras protein expression may be used as a marker to predict the progression of breast cancer; this

  2. Regulation of H-Ras-driven MAPK signaling, transformation and tumorigenesis, but not PI3K signaling and tumor progression, by plasma membrane microdomains.

    PubMed

    Michael, J V; Wurtzel, J G T; Goldfinger, L E

    2016-05-30

    In this study, we assessed the contributions of plasma membrane (PM) microdomain targeting to the functions of H-Ras and R-Ras. These paralogs have identical effector-binding regions, but variant C-terminal targeting domains (tDs) which are responsible for lateral microdomain distribution: activated H-Ras targets to lipid ordered/disordered (Lo/Ld) domain borders, and R-Ras to Lo domains (rafts). We hypothesized that PM distribution regulates Ras-effector interactions and downstream signaling. We used tD swap mutants, and assessed effects on signal transduction, cell proliferation, transformation and tumorigenesis. R-Ras harboring the H-Ras tD (R-Ras-tH) interacted with Raf, and induced Raf and ERK phosphorylation similar to H-Ras. R-Ras-tH stimulated proliferation and transformation in vitro, and these effects were blocked by both MEK and PI3K inhibition. Conversely, the R-Ras tD suppressed H-Ras-mediated Raf activation and ERK phosphorylation, proliferation and transformation. Thus, Ras access to Raf at the PM is sufficient for MAPK activation and is a principal component of Ras mitogenesis and transformation. Fusion of the R-Ras extended N-terminal domain to H-Ras had no effect on proliferation, but inhibited transformation and tumor progression, indicating that the R-Ras N-terminus also contributes negative regulation to these Ras functions. PI3K activation was tD independent; however, H-Ras was a stronger activator of PI3K than R-Ras, with either tD. PI3K inhibition nearly ablated transformation by R-Ras-tH, H-Ras and H-Ras-tR, whereas MEK inhibition had a modest effect on Ras-tH-driven transformation but no effect on H-Ras-tR transformation. R-Ras-tH supported tumor initiation, but not tumor progression. While H-Ras-tR-induced transformation was reduced relative to H-Ras, tumor progression was robust and similar to H-Ras. H-Ras tumor growth was moderately suppressed by MEK inhibition, which had no effect on H-Ras-tR tumor growth. In contrast, PI3K inhibition

  3. Graphene plasmonic lens for manipulating energy flow

    NASA Astrophysics Data System (ADS)

    Wang, Guoxi; Liu, Xueming; Lu, Hua; Zeng, Chao

    2014-02-01

    Manipulating the energy flow of light is at the heart of modern information and communication technologies. Because photons are uncharged, it is still difficult to effectively control them by electrical means. Here, we propose a graphene plasmonic (GP) lens to efficiently manipulate energy flow by elaborately designing the thickness of the dielectric spacer beneath the graphene sheet. Different from traditional metal-based lenses, the proposed graphene plasmonic lens possesses the advantages of tunability and excellent confinement of surface plasmons. It is found that the proposed lens can be utilized to focus and collimate the GP waves propagating along the graphene sheet. Particularly, the lens is dispersionless over a wide frequency range and the performance of lens can be flexibly tuned by adjusting the bias voltage. As an application of such a lens, the image transfer of two point sources with a separation of λ0/30 is demonstrated.

  4. Graphene plasmonic lens for manipulating energy flow

    PubMed Central

    Wang, Guoxi; Liu, Xueming; Lu, Hua; Zeng, Chao

    2014-01-01

    Manipulating the energy flow of light is at the heart of modern information and communication technologies. Because photons are uncharged, it is still difficult to effectively control them by electrical means. Here, we propose a graphene plasmonic (GP) lens to efficiently manipulate energy flow by elaborately designing the thickness of the dielectric spacer beneath the graphene sheet. Different from traditional metal-based lenses, the proposed graphene plasmonic lens possesses the advantages of tunability and excellent confinement of surface plasmons. It is found that the proposed lens can be utilized to focus and collimate the GP waves propagating along the graphene sheet. Particularly, the lens is dispersionless over a wide frequency range and the performance of lens can be flexibly tuned by adjusting the bias voltage. As an application of such a lens, the image transfer of two point sources with a separation of λ0/30 is demonstrated. PMID:24517981

  5. Optmization design of zoom lens systems

    NASA Astrophysics Data System (ADS)

    Li, Xiaotong; Cen, Zhaofeng

    2002-09-01

    A zoom lens system is usually composed of several components. Some of the components can be moved to change the focal length or magnification. Zoom lens system design is more complicated than fixed-focus lens design due to the moving of some components. In this paper, an optimization method that is used to design zoom lens systems is presented. Using this method, the Gaussian parameters of zoom lens systems are optimized at first, and then the initial structure parameters in each component are generated and optimized. At last the aberration balance is made using multi-configuration. In this paper the flowchart of optimization design for such complex optical systems is showed and the algorithms are described. As a conclusion, the relationship between power distribution, initial structure and the aberrations is considered at the beginning, the evaluation criteria are reliable and efficiency for designing zoom lens systems.

  6. Nitric oxide mediates N-methyl-D-aspartate receptor-induced activation of p21ras.

    PubMed

    Yun, H Y; Gonzalez-Zulueta, M; Dawson, V L; Dawson, T M

    1998-05-12

    N-methyl-D-aspartate (NMDA) glutamate receptor-mediated increases in intracellular calcium are thought to play a critical role in synaptic plasticity. The mechanisms by which changes in cytoplasmic calcium transmit the glutamate signal to the nucleus, which is ultimately important for long-lasting neuronal responses, are poorly understood. We show that NMDA receptor stimulation leads to activation of p21(ras) (Ras) through generation of nitric oxide (NO) via neuronal NO synthase. The competitive NO synthase inhibitor, L-nitroarginine methyl ester, prevents Ras activation elicited by NMDA and this effect is competitively reversed by the NO synthase substrate, L-arginine. NMDA receptor stimulation fails to activate Ras in neuronal cultures from mice lacking neuronal NO synthase. NMDA-induced Ras activation occurs through a cGMP-independent pathway as 1H-[1,2,4]oxadiazolo[4,3-alpha]quinoxalin-1-one (ODQ), a potent and selective inhibitor of guanylyl cyclase, has no effect on NMDA receptor-induced activation of Ras, and the cell-permeable cGMP analog, 8Br-cGMP, does not activate Ras. Furthermore, NO directly activates immunoprecipitated Ras from neurons. NMDA also elicits tyrosine phosphorylation of extracellular signal-regulated kinases, a downstream effector pathway of Ras, through a NO/non-cGMP dependent mechanism, thus supporting the physiologic relevance of endogenous NO regulation of Ras. These results suggest that Ras is a physiologic target of endogenously produced NO and indicates a signaling pathway for NMDA receptor activation that may be important for long-lasting neuronal responses.

  7. PEA-15 potentiates H-Ras mediated epithelial cell transformation through Phospholipase D

    PubMed Central

    Sulzmaier, Florian J.; Valmiki, Mohana K. Gudur; Nelson, Deirdre A.; Caliva, Maisel J.; Geerts, Dirk; Matter, Michelle L.; White, Eileen P.; Ramos, Joe W.

    2011-01-01

    The small GTPase H-Ras is a proto-oncogene that activates a variety of different pathways including the extracellular-signal-regulated kinase mitogen-activated protein kinase (ERK/MAPK) pathway. H-Ras is mutated in many human malignancies and these mutations cause the protein to be constitutively active. PEA-15 blocks ERK-dependent gene transcription and inhibits proliferation by sequestering ERK in the cytoplasm. We therefore investigated whether PEA-15 influences H-Ras mediated transformation. We found that PEA-15 does not block H-Ras activated proliferation when H-Ras is constitutively active. We show instead that in H-Ras transformed mouse kidney epithelial cells, co-expression of PEA-15 resulted in enhanced soft agar colony growth and increased tumor growth in vivo. Overexpression of both H-Ras and PEA-15 resulted in accelerated G1/S cell cycle transition and increased activation of the ERK signaling pathway. PEA-15 mediated these effects through activation of its binding partner phospholipase D1 (PLD1). Inhibition of PLD1 or interference with PEA-15/PLD1 binding blocked PEA-15’s ability to increase ERK activation. Our findings reveal a novel mechanism by which PEA-15 positively regulates Ras/ERK signaling and increases the proliferation of H-Ras transformed epithelial cells through enhanced PLD1 expression and activation. Thus, our work provides a surprising mechanism by which PEA-15 augments H-Ras driven transformation. These data reveal that PEA-15 not only suppresses ERK signaling and tumorigenesis but can alternatively enhance tumorigenesis in the context of active Ras. PMID:22105357

  8. Lateral shear interferometry with holo shear lens

    NASA Astrophysics Data System (ADS)

    Joenathan, C.; Mohanty, R. K.; Sirohi, R. S.

    1984-12-01

    A simple method for obtaining lateral shear using holo shear lenses (HSL) has been discussed. This simple device which produces lateral shears in the orthogonal directions has been used for lens testing. The holo shear lens is placed at or near the focus of the lens to be tested. It has also been shown that HSL can be used in speckle shear interferometry as it performs both the functions of shearing and imaging.

  9. No consistent pattern of mutations in p53 and ras genes in liver tumors of rat treated with the drinking water mutagen 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX).

    PubMed

    Komulainen, H; Hakulinen, P; Servomaa, K; Makkonen, K; Vasara, R; Mäki-Paakkanen, J; Kosma, V M

    2000-01-01

    The frequency of point mutations in p53 (exons 4-7) and in Ki-ras, Ha-ras, and N-ras (exons 1 and 2) and the expression of p53 protein were evaluated in the liver tumors of Wistar rats of a 104-week carcinogenicity study on 3-chloro-4-(dichloromethyl)-5-hydroxy-2(5H)-furanone (MX), a chlorine disinfection by-product in drinking water. Mutations were analyzed in 16 hepatocellular adenomas, 7 hepatocellular carcinomas, 23 cholangiomas, and 2 cholangiocarcinomas of the MX-treated animals and one hepatocellular carcinoma and cholangiocarcinoma in control animals using PCR-SSCP (polymerase chain reaction-single-strand conformation polymorphism) or PCR-TGGE (temperature gradient gel electrophoresis) and direct sequencing. The expression of the p53 protein (wild-type and mutated protein) was detected by immunohistochemistry (CM5 antibody). The expression of p53 and that of the proliferating cell nuclear antigen (PCNA, 19 A2) were also evaluated in livers of female animals exposed to MX for 1 week, 3 weeks, or 18 weeks. Altogether, four mutations were found in p53 in three tumors, in two hepatocellular adenomas, and one cholangiocarcinoma, all in females receiving the highest MX dose (6. 6 mg/kg/day) of the study. Three of the mutations were G:C --> A:T transitions and one was an A:T --> T:A transversion. The mutations were scattered at different codons and positions of the codon. One hepatocellular adenoma contained two p53 mutations. All cholangiomas and cholangiocarcinomas, but no hepatocellular adenomas and carcinomas, overexpressed the p53 protein. MX treatment did not induce p53 expression at any age in the liver or alter the expression of the PCNA in the liver of younger animals. The p53 protein was overexpressed in hyperplastic bile ducts in aged rats but not in bile ducts of younger rats (up to 24 weeks). No mutations were observed in either Ki-ras, Ha-ras, or N-ras of the liver tumors. These data suggest that point mutations in p53, Ki-ras, Ha-ras, and N-ras are

  10. Role of the lens capsule on the mechanical accommodative response in a lens stretcher

    PubMed Central

    Ziebarth, Noël M.; Borja, David; Arrieta, Esdras; Aly, Mohamed; Manns, Fabrice; Dortonne, Isabelle; Nankivil, Derek; Jain, Rakhi; Parel, Jean-Marie

    2009-01-01

    Purpose To determine if changes in elastic properties of the lens capsule ex vivo with age contribute to the forces required to accommodate. Methods Post-mortem human (n=22; age average: 41±17years, range: 6–71 years) and cynomolgus monkey (n=19; age average: 7.7±1.8 years, range: 4.2–10 years) tissues including the lens, capsule, zonules, ciliary body, and sclera were mounted in an optomechanical lens stretching system. Starting at zero load, the sclera was symmetrically stretched to 2mm in 0.25 steps at a speed of 0.1mm.s−1. The load and lens diameter were measured at each step. The lens contents were removed through a mini-capsulorhexis. The stretching cycles were repeated on the empty capsular bag. The forces required to stretch the natural lens and empty bag were quantified as a function of age and compared. Results The force required to stretch the empty lens capsule was independent of age (human=2.6–34.9g/mm [25.2–342.7mN/mm], monkey=8.2–21.3g/mm [80.3–208.6mN/mm]). The ratio of the force required to stretch the empty lens capsule to the force required to stretch the natural lens decreases with age in human and monkey lenses (p=0.003, p=0.72, respectively). Conclusions The mechanical properties of the empty lens capsule assessed ex vivo in a lens stretcher remain constant with age, suggesting that the changes in elasticity of the lens capsule do not play a significant role in presbyopia. In young eyes, the lens capsule determines the force required to stretch the whole lens. The age-related increase in force required to stretch the lens is due to changes in the lens contents. PMID:18515568

  11. Elasticity of the eye's crystalline lens: A Brillouin light scattering study.

    NASA Astrophysics Data System (ADS)

    Bailey, S.; Gump, J.; Sooryakumar, R.; Jayaprakash, C.; Venkiteshwar, M. S.; Bullimore, M.; Twa, M.

    2009-03-01

    Focusing the eye on a near object results in an increase in its optical power brought about by contraction of the ciliary muscles and an increase in the lens surface curvature. Distant vision occurs when the muscular force flattens the lens. Central to the ability of the lens to alter shape are its mechanical properties. Thus, given that hardening of the lens would impede deformation and reduce its ability to undergo the changes required for accommodation, a noninvasive approach to measure the elastic properties of the lens is valuable. We present results of Brillouin scattering from bovine and human lenses (from the organ donor program at The Ohio State University) that measure their high frequency acoustic response. These measurements are conducted with a few milli-watts of laser power and, in the case of bovine lenses, from entire intact eye globes, allow the stiffness of the lens to be mapped across its cross-section. The results will be compared to values of the shear- and bulk-moduli determined from other techniques and the implications of differences in these moduli discussed.

  12. Optical density of the crystalline lens

    SciTech Connect

    Hemenger, R.P.

    1982-01-01

    The optical density for the noncataractous crystalline lens is written as a sum of two terms, each with a specific dependence on wavelength. The first term, proportional to 1/lambda 2, represents all light-scattering processes in the lens. The second term, assumed significant only for lambda less than or equal to 500 nm, accounts for absorption by lens pigments. By analyzing transmittance data on lenses of subjects aged 21 to 63 years, a spectrum for light absorption by lens pigment is derived and it is shown to be essentially the same for all of the lenses.

  13. Stretchable Binary Fresnel Lens for Focus Tuning

    PubMed Central

    Li, Xueming; Wei, Lei; Poelma, René H.; Vollebregt, Sten; Wei, Jia; Urbach, Hendrik Paul; Sarro, Pasqualina M.; Zhang, Guo Qi

    2016-01-01

    This paper presents a tuneable binary amplitude Fresnel lens produced by wafer-level microfabrication. The Fresnel lens is fabricated by encapsulating lithographically defined vertically aligned carbon nanotube (CNT) bundles inside a polydimethyl-siloxane (PDMS) layer. The composite lens material combines the excellent optical absorption properties of the CNT with the transparency and stretchability of the PDMS. By stretching the elastomeric composite in radial direction, the lens focal length is tuned. Good focusing response is demonstrated and a large focus change (≥24%) was achieved by stretching lenses up to 11.4%. PMID:27139747

  14. A Magnification Lens for Interactive Volume Visualization

    SciTech Connect

    LaMar, E; Hamann, B; Joy, K I

    2001-07-19

    Volume visualization of large data sets suffers from the same problem that many other visualization modalities suffer from: either one can visualize the entire data set and lose small details or visualize a small region and lose the context. In this paper, they present a magnification lens technique for volume visualization. While the notion of a magnification-lens is not new, and other techniques attempt to simulate the physical properties of a magnifying lens, their contribution is in developing a magnification lens that is fast, can be implemented using a fairly small software overhead, and has a natural, intuitive appearance. The issue with magnification lens is the border, or transition, region. The lens center and exterior have a constant zoom factor, and are simple to render. It is the border region that blends between the external and interior magnification, and has a non-constant magnification. They use the perspective-correct textures capability, available in most current graphics systems, to produce a lens with a tessellated border region that approximates linear compression with respect to the radius of the magnification lens. They discuss how a cubic border can mitigate the discontinuities resulting from the use of a linear function, without significant performance loss. They discuss various issues concerning development of a three-dimensional magnification lens.

  15. Stretchable Binary Fresnel Lens for Focus Tuning

    NASA Astrophysics Data System (ADS)

    Li, Xueming; Wei, Lei; Poelma, René H.; Vollebregt, Sten; Wei, Jia; Urbach, Hendrik Paul; Sarro, Pasqualina M.; Zhang, Guo Qi

    2016-05-01

    This paper presents a tuneable binary amplitude Fresnel lens produced by wafer-level microfabrication. The Fresnel lens is fabricated by encapsulating lithographically defined vertically aligned carbon nanotube (CNT) bundles inside a polydimethyl-siloxane (PDMS) layer. The composite lens material combines the excellent optical absorption properties of the CNT with the transparency and stretchability of the PDMS. By stretching the elastomeric composite in radial direction, the lens focal length is tuned. Good focusing response is demonstrated and a large focus change (≥24%) was achieved by stretching lenses up to 11.4%.

  16. Endoscopic inspection using a panoramic annular lens

    NASA Technical Reports Server (NTRS)

    Gilbert, John A.; Matthys, Donald R.

    1991-01-01

    The objective of this one year study was to design, build, and demonstrate a prototype system for cavity inspection. A cylindrical view of the cavity interior was captured in real time through a compound lens system consisting of a unique panoramic annular lens and a collector lens. Images, acquired with a digitizing camera and stored in a desktop computer, were manipulated using image processing software to aid in visual inspection and qualitative analysis. A detailed description of the lens and its applications is given.

  17. Algorithm design of liquid lens inspection system

    NASA Astrophysics Data System (ADS)

    Hsieh, Lu-Lin; Wang, Chun-Chieh

    2008-08-01

    In mobile lens domain, the glass lens is often to be applied in high-resolution requirement situation; but the glass zoom lens needs to be collocated with movable machinery and voice-coil motor, which usually arises some space limits in minimum design. In high level molding component technology development, the appearance of liquid lens has become the focus of mobile phone and digital camera companies. The liquid lens sets with solid optical lens and driving circuit has replaced the original components. As a result, the volume requirement is decreased to merely 50% of the original design. Besides, with the high focus adjusting speed, low energy requirement, high durability, and low-cost manufacturing process, the liquid lens shows advantages in the competitive market. In the past, authors only need to inspect the scrape defect made by external force for the glass lens. As to the liquid lens, authors need to inspect the state of four different structural layers due to the different design and structure. In this paper, authors apply machine vision and digital image processing technology to administer inspections in the particular layer according to the needs of users. According to our experiment results, the algorithm proposed can automatically delete non-focus background, extract the region of interest, find out and analyze the defects efficiently in the particular layer. In the future, authors will combine the algorithm of the system with automatic-focus technology to implement the inside inspection based on the product inspective demands.

  18. Coherence and frequency in the reticular activating system (RAS).

    PubMed

    Garcia-Rill, Edgar; Kezunovic, Nebojsa; Hyde, James; Simon, Christen; Beck, Paige; Urbano, Francisco J

    2013-06-01

    This review considers recent evidence showing that cells in the reticular activating system (RAS) exhibit (1) electrical coupling mainly in GABAergic cells, and (2) gamma band activity in virtually all of the cells. Specifically, cells in the mesopontine pedunculopontine nucleus (PPN), intralaminar parafascicular nucleus (Pf), and pontine dorsal subcoeruleus nucleus dorsalis (SubCD) (1) show electrical coupling, and (2) all fire in the beta/gamma band range when maximally activated, but no higher. The mechanism behind electrical coupling is important because the stimulant modafinil was shown to increase electrical coupling. We also provide recent findings demonstrating that all cells in the PPN and Pf have high threshold, voltage-dependent P/Q-type calcium channels that are essential to gamma band activity. On the other hand, all SubCD, and some PPN, cells manifested sodium-dependent subthreshold oscillations. A novel mechanism for sleep-wake control based on transmitter interactions, electrical coupling, and gamma band activity is described. We speculate that continuous sensory input will modulate coupling and induce gamma band activity in the RAS that could participate in the processes of preconscious awareness, and provide the essential stream of information for the formulation of many of our actions.

  19. 4-Hydroxytamoxifen induces autophagic death through K-Ras degradation.

    PubMed

    Kohli, Latika; Kaza, Niroop; Coric, Tatjana; Byer, Stephanie J; Brossier, Nicole M; Klocke, Barbara J; Bjornsti, Mary-Ann; Carroll, Steven L; Roth, Kevin A

    2013-07-15

    Tamoxifen is widely used to treat estrogen receptor-positive breast cancer. Recent findings that tamoxifen and its derivative 4-hydroxytamoxifen (OHT) can exert estrogen receptor-independent cytotoxic effects have prompted the initiation of clinical trials to evaluate its use in estrogen receptor-negative malignancies. For example, tamoxifen and OHT exert cytotoxic effects in malignant peripheral nerve sheath tumors (MPNST) where estrogen is not involved. In this study, we gained insights into the estrogen receptor-independent cytotoxic effects of OHT by studying how it kills MPNST cells. Although caspases were activated following OHT treatment, caspase inhibition provided no protection from OHT-induced death. Rather, OHT-induced death in MPNST cells was associated with autophagic induction and attenuated by genetic inhibition of autophagic vacuole formation. Mechanistic investigations revealed that OHT stimulated autophagic degradation of K-Ras, which is critical for survival of MPNST cells. Similarly, we found that OHT induced K-Ras degradation in breast, colon, glioma, and pancreatic cancer cells. Our findings describe a novel mechanism of autophagic death triggered by OHT in tumor cells that may be more broadly useful clinically in cancer treatment.

  20. The PDZ Protein Canoe/AF-6 Links Ras-MAPK, Notch and Wingless/Wnt Signaling Pathways by Directly Interacting with Ras, Notch and Dishevelled

    PubMed Central

    Carmena, Ana; Speicher, Stephan; Baylies, Mary

    2006-01-01

    Over the past few years, it has become increasingly apparent that signal transduction pathways are not merely linear cascades; they are organized into complex signaling networks that require high levels of regulation to generate precise and unique cell responses. However, the underlying regulatory mechanisms by which signaling pathways cross-communicate remain poorly understood. Here we show that the Ras-binding protein Canoe (Cno)/AF-6, a PDZ protein normally associated with cellular junctions, is a key modulator of Wingless (Wg)/Wnt, Ras-Mitogen Activated Protein Kinase (MAPK) and Notch (N) signaling pathways cross-communication. Our data show a repressive effect of Cno/AF-6 on these three signaling pathways through physical interactions with Ras, N and the cytoplasmic protein Dishevelled (Dsh), a key Wg effector. We propose a model in which Cno, through those interactions, actively coordinates, at the membrane level, Ras-MAPK, N and Wg signaling pathways during progenitor specification. PMID:17183697

  1. Expression of truncated PITX3 in the developing lens leads to microphthalmia and aphakia in mice.

    PubMed

    Wada, Kenta; Matsushima, Yoshibumi; Tada, Tomoki; Hasegawa, Sayaka; Obara, Yo; Yoshizawa, Yasuhiro; Takahashi, Gou; Hiai, Hiroshi; Shimanuki, Midori; Suzuki, Sari; Saitou, Junichi; Yamamoto, Naoki; Ichikawa, Masumi; Watanabe, Kei; Kikkawa, Yoshiaki

    2014-01-01

    Microphthalmia is a severe ocular disorder, and this condition is typically caused by mutations in transcription factors that are involved in eye development. Mice carrying mutations in these transcription factors would be useful tools for defining the mechanisms underlying developmental eye disorders. We discovered a new spontaneous recessive microphthalmos mouse mutant in the Japanese wild-derived inbred strain KOR1/Stm. The homozygous mutant mice were histologically characterized as microphthalmic by the absence of crystallin in the lens, a condition referred to as aphakia. By positional cloning, we identified the nonsense mutation c.444C>A outside the genomic region that encodes the homeodomain of the paired-like homeodomain transcription factor 3 gene (Pitx3) as the mutation responsible for the microphthalmia and aphakia. We examined Pitx3 mRNA expression of mutant mice during embryonic stages using RT-PCR and found that the expression levels are higher than in wild-type mice. Pitx3 over-expression in the lens during developmental stages was also confirmed at the protein level in the microphthalmos mutants via immunohistochemical analyses. Although lens fiber differentiation was not observed in the mutants, strong PITX3 protein signals were observed in the lens vesicles of the mutant lens. Thus, we speculated that abnormal PITX3, which lacks the C-terminus (including the OAR domain) as a result of the nonsense mutation, is expressed in mutant lenses. We showed that the expression of the downstream genes Foxe3, Prox1, and Mip was altered because of the Pitx3 mutation, with large reductions in the lens vesicles in the mutants. Similar profiles were observed by immunohistochemical analysis of these proteins. The expression profiles of crystallins were also altered in the mutants. Therefore, we speculated that the microphthalmos/aphakia in this mutant is caused by the expression of truncated PITX3, resulting in the abnormal expression of downstream targets and

  2. Expression of Truncated PITX3 in the Developing Lens Leads to Microphthalmia and Aphakia in Mice

    PubMed Central

    Wada, Kenta; Matsushima, Yoshibumi; Tada, Tomoki; Hasegawa, Sayaka; Obara, Yo; Yoshizawa, Yasuhiro; Takahashi, Gou; Hiai, Hiroshi; Shimanuki, Midori; Suzuki, Sari; Saitou, Junichi; Yamamoto, Naoki; Ichikawa, Masumi; Watanabe, Kei; Kikkawa, Yoshiaki

    2014-01-01

    Microphthalmia is a severe ocular disorder, and this condition is typically caused by mutations in transcription factors that are involved in eye development. Mice carrying mutations in these transcription factors would be useful tools for defining the mechanisms underlying developmental eye disorders. We discovered a new spontaneous recessive microphthalmos mouse mutant in the Japanese wild-derived inbred strain KOR1/Stm. The homozygous mutant mice were histologically characterized as microphthalmic by the absence of crystallin in the lens, a condition referred to as aphakia. By positional cloning, we identified the nonsense mutation c.444C>A outside the genomic region that encodes the homeodomain of the paired-like homeodomain transcription factor 3 gene (Pitx3) as the mutation responsible for the microphthalmia and aphakia. We examined Pitx3 mRNA expression of mutant mice during embryonic stages using RT-PCR and found that the expression levels are higher than in wild-type mice. Pitx3 over-expression in the lens during developmental stages was also confirmed at the protein level in the microphthalmos mutants via immunohistochemical analyses. Although lens fiber differentiation was not observed in the mutants, strong PITX3 protein signals were observed in the lens vesicles of the mutant lens. Thus, we speculated that abnormal PITX3, which lacks the C-terminus (including the OAR domain) as a result of the nonsense mutation, is expressed in mutant lenses. We showed that the expression of the downstream genes Foxe3, Prox1, and Mip was altered because of the Pitx3 mutation, with large reductions in the lens vesicles in the mutants. Similar profiles were observed by immunohistochemical analysis of these proteins. The expression profiles of crystallins were also altered in the mutants. Therefore, we speculated that the microphthalmos/aphakia in this mutant is caused by the expression of truncated PITX3, resulting in the abnormal expression of downstream targets and

  3. Profiling of transcripts and proteins modulated by K-ras oncogene in the lung tissues of K-ras transgenic mice by omics approaches.

    PubMed

    Lee, Sojung; Kang, Jungwoo; Cho, Minchul; Seo, Eunhee; Choi, Heesook; Kim, Eunjin; Kim, Junghee; Kim, Heejong; Kang, Gum Yong; Kim, Kwang Pyo; Park, Young-Ho; Yu, Dae-Yeul; Yum, Young Na; Park, Sue-Nie; Yoon, Do-Young

    2009-01-01

    The mutated K-ras gene is involved in approximately 30% of human cancers. In order to search for K-ras oncogene-induced modulators in lung tissues of K-ras transgenic mice, we performed microarray and proteomics (LC/ESI-MS/MS) analysis. Genes (RAB27b RAS family, IL-1RA, IL-33, chemokine ligand 6, epiregulin, EGF-like domain and cathepsin) related to cancer development (Wnt signaling pathway) and inflammation (chemokine/cytokine signaling pathway, Toll receptor signaling) were up-regulated while genes (troponin, tropomodulin 2, endothelial lipase, FGFR4, integrin alpha8 and adenylate cyclase 8) related to the tumor suppression such as p53 pathway, TGF-beta signaling pathway and cadherin signaling pathway were down-regulated by K-ras oncogene. Proteomics approach revealed that up-regulated proteins in lung adenomas of K-ras mice were classified as follows: proteins related to the metabolism/catabolism (increased from 7 to 22% by K-ras gene), proteins related to translation/transcription and nucleotide (from 4 to 6%), proteins related to signal transduction (from 3 to 5%), proteins related to phosphorylation (from 1 to 2%). ATP synthase, Ras oncogene family, cytochrome c oxidase, flavoprotein, TEF 1, adipoprotein A-1 BP, glutathione oxidase, fatty acid BP 4, diaphorase 1, MAPK4 and transgelin were up-regulated by K-ras oncogene. However, integrin alpha1, Ras-interacting protein (Rain), endothelin-converting enzyme-1d and splicing factor 3b were down-regulated. These studies suggest that genes related to cancer development and inflammation were up-regulated while genes related to the tumor suppression were down-regulated by K-ras, resulting in the tumor growth. Putative biomarkers such as cell cycle related genes (Cdc37), cancer cell adhesion (Glycam 1, integrin alpha8, integrin alphaX and Clec4n), signal transduction (Tlr2, IL-33, and Ccbp2), migration (Ccr1, Ccl6, and diaphorase 1 (Cyb5r3) and cancer development (epiregulin) can be useful for diagnosis and as

  4. Profiling of transcripts and proteins modulated by K-ras oncogene in the lung tissues of K-ras transgenic mice by omics approaches.

    PubMed

    Lee, Sojung; Kang, Jungwoo; Cho, Minchul; Seo, Eunhee; Choi, Heesook; Kim, Eunjin; Kim, Junghee; Kim, Heejong; Kang, Gum Yong; Kim, Kwang Pyo; Park, Young-Ho; Yu, Dae-Yeul; Yum, Young Na; Park, Sue-Nie; Yoon, Do-Young

    2009-01-01

    The mutated K-ras gene is involved in approximately 30% of human cancers. In order to search for K-ras oncogene-induced modulators in lung tissues of K-ras transgenic mice, we performed microarray and proteomics (LC/ESI-MS/MS) analysis. Genes (RAB27b RAS family, IL-1RA, IL-33, chemokine ligand 6, epiregulin, EGF-like domain and cathepsin) related to cancer development (Wnt signaling pathway) and inflammation (chemokine/cytokine signaling pathway, Toll receptor signaling) were up-regulated while genes (troponin, tropomodulin 2, endothelial lipase, FGFR4, integrin alpha8 and adenylate cyclase 8) related to the tumor suppression such as p53 pathway, TGF-beta signaling pathway and cadherin signaling pathway were down-regulated by K-ras oncogene. Proteomics approach revealed that up-regulated proteins in lung adenomas of K-ras mice were classified as follows: proteins related to the metabolism/catabolism (increased from 7 to 22% by K-ras gene), proteins related to translation/transcription and nucleotide (from 4 to 6%), proteins related to signal transduction (from 3 to 5%), proteins related to phosphorylation (from 1 to 2%). ATP synthase, Ras oncogene family, cytochrome c oxidase, flavoprotein, TEF 1, adipoprotein A-1 BP, glutathione oxidase, fatty acid BP 4, diaphorase 1, MAPK4 and transgelin were up-regulated by K-ras oncogene. However, integrin alpha1, Ras-interacting protein (Rain), endothelin-converting enzyme-1d and splicing factor 3b were down-regulated. These studies suggest that genes related to cancer development and inflammation were up-regulated while genes related to the tumor suppression were down-regulated by K-ras, resulting in the tumor growth. Putative biomarkers such as cell cycle related genes (Cdc37), cancer cell adhesion (Glycam 1, integrin alpha8, integrin alphaX and Clec4n), signal transduction (Tlr2, IL-33, and Ccbp2), migration (Ccr1, Ccl6, and diaphorase 1 (Cyb5r3) and cancer development (epiregulin) can be useful for diagnosis and as

  5. Paediatric intraocular lens implants: accuracy of lens power calculations.

    PubMed

    O'Gallagher, M K; Lagan, M A; Mulholland, C P; Parker, M; McGinnity, G; McLoone, E M

    2016-09-01

    PurposeThis study aims to evaluate the accuracy of lens prediction formulae on a paediatric population.MethodsA retrospective case-note review was undertaken of patients under 8 years old who underwent cataract surgery with primary lens implantation in a regional referral centre for paediatric ophthalmology, excluding those whose procedure was secondary to trauma. Biometric and refractive data were analysed for 43 eyes, including prediction errors (PE). Statistical measures used included mean absolute error (MAE), median absolute error (MedAE), Student's t-test and Lin's correlation coefficient.ResultsThe mean PE using the SRK-II formula was +0.96 D (range -2.47D to +2.41 D, SD 1.33 D, MAE 1.38 D, MedAE 1.55, n=15). The mean PE was smaller using SRK/T (-0.18 D, range -3.25 D to +3.95 D, SD 1.70 D, MAE 1.30 D, MedAE 1.24, n=27). We performed an analysis of the biometry data using four different formula (Hoffer Q, Holladay 1, SRK-II and SRK/T). Hoffer Q showed a smaller MedAE than other formulae but also a myopic bias.ConclusionOur clinical data suggest SRK/T was more accurate in predicting post-operative refraction in this cohort of paediatric patients undergoing cataract surgery. Hoffer Q may have improved accuracy further.

  6. Society News: PhD theses could win prizes; Last chance for IYA2009 grants; New Fellows; RAS Fellows win prizes; Need a job? Need staff? RAS Library Saturdays

    NASA Astrophysics Data System (ADS)

    2009-08-01

    Fellows who are PhD student supervisors should be on the lookout for exceptionally good work from research students submitting their theses this year, for nomination for the RAS Michael Penston Astronomy Prize and the RAS Keith Runcorn Prize. The RAS is offering one last chance to apply for grants towards International Year of Astronomy activities, but you'll have to apply soon. The Society sends congratulations to Fellows of the RAS who have recently received prestigious awards for their work.

  7. Study Illuminates K-Ras4B Activation, Which May Help Predict Drug Resistance | Poster

    Cancer.gov

    Until recently, researchers studying RAS, a family of proteins involved in transmitting signals within cells, believed that the exchange of guanosine 5’-diphosphate (GDP) by guanosine triphosphate (GTP) was sufficient to activate the protein. Once activated, RAS can cause unintended and overactive signaling in cells, which can lead to cell division and, ultimately, cancer.

  8. Genome-wide gene expression analysis identifies K-ras as a regulator of alcohol intake.

    PubMed

    Repunte-Canonigo, Vez; van der Stap, Lena D; Chen, Jihuan; Sabino, Valentina; Wagner, Ulrich; Zorrilla, Eric P; Schumann, Gunter; Roberts, Amanda J; Sanna, Pietro Paolo

    2010-06-21

    Adaptations in the anterior cingulate cortex (ACC) have been implicated in alcohol and drug addiction. To identify genes that may contribute to excessive drinking, here we performed microarray analyses in laser microdissected rat ACC after a single or repeated administration of an intoxicating dose of alcohol (3 g/kg). Expression of the small G protein K-ras was differentially regulated following both single and repeated alcohol administration. We also observed that voluntary alcohol intake in K-ras heterozygous null mice (K-ras(+/-)) did not increase after withdrawal from repeated cycles of intermittent ethanol vapor exposure, unlike in their wild-type littermates. To identify K-ras regulated pathways, we then profiled gene expression in the ACC of K-ras(+/-), heterozygous null mice for the K-ras negative regulator Nf1 (Nf1(+/-)) and wild-type mice following repeated administration of an intoxicating dose of alcohol. Pathway analysis showed that alcohol differentially affected various pathways in a K-ras dependent manner - some of which previously shown to be regulated by alcohol - including the insulin/PI3K pathway, the NF-kappaB, the phosphodiesterases (PDEs) pathway, the Jak/Stat and the adipokine signaling pathways. Altogether, the data implicate K-ras-regulated pathways in the regulation of excessive alcohol drinking after a history of dependence.

  9. K-Ras(G12C) inhibitors allosterically control GTP affinity and effector interactions

    NASA Astrophysics Data System (ADS)

    Ostrem, Jonathan M.; Peters, Ulf; Sos, Martin L.; Wells, James A.; Shokat, Kevan M.

    2013-11-01

    Somatic mutations in the small GTPase K-Ras are the most common activating lesions found in human cancer, and are generally associated with poor response to standard therapies. Efforts to target this oncogene directly have faced difficulties owing to its picomolar affinity for GTP/GDP and the absence of known allosteric regulatory sites. Oncogenic mutations result in functional activation of Ras family proteins by impairing GTP hydrolysis. With diminished regulation by GTPase activity, the nucleotide state of Ras becomes more dependent on relative nucleotide affinity and concentration. This gives GTP an advantage over GDP and increases the proportion of active GTP-bound Ras. Here we report the development of small molecules that irreversibly bind to a common oncogenic mutant, K-Ras(G12C). These compounds rely on the mutant cysteine for binding and therefore do not affect the wild-type protein. Crystallographic studies reveal the formation of a new pocket that is not apparent in previous structures of Ras, beneath the effector binding switch-II region. Binding of these inhibitors to K-Ras(G12C) disrupts both switch-I and switch-II, subverting the native nucleotide preference to favour GDP over GTP and impairing binding to Raf. Our data provide structure-based validation of a new allosteric regulatory site on Ras that is targetable in a mutant-specific manner.

  10. A RAS renaissance: emerging targeted therapies for KRAS-mutated non-small cell lung cancer.

    PubMed

    Vasan, Neil; Boyer, Julie L; Herbst, Roy S

    2014-08-01

    Of the numerous oncogenes implicated in human cancer, the most common and perhaps the most elusive to target pharmacologically is RAS. Since the discovery of RAS in the 1960s, numerous studies have elucidated the mechanism of activity, regulation, and intracellular trafficking of the RAS gene products, and of its regulatory pathways. These pathways yielded druggable targets, such as farnesyltransferase, during the 1980s to 1990s. Unfortunately, early clinical trials investigating farnesyltransferase inhibitors yielded disappointing results, and subsequent interest by pharmaceutical companies in targeting RAS waned. However, recent advances including the identification of novel regulatory enzymes (e.g., Rce1, Icmt, Pdeδ), siRNA-based synthetic lethality screens, and fragment-based small-molecule screens, have resulted in a "Ras renaissance," signified by new Ras and Ras pathway-targeted therapies that have led to new clinical trials of patients with Ras-driven cancers. This review gives an overview of KRas signaling pathways with an emphasis on novel targets and targeted therapies, using non-small cell lung cancer as a case example.

  11. College Students vs. RAs: A Programmatic Comparison of Ideas about Alcohol and Drugs.

    ERIC Educational Resources Information Center

    Coll, Kenneth M.; Darby, Merlin D.

    This study compared differences between resident assistants (RAs) and conduct code violators in alcohol and drug knowledge, reported use, and perceptions of reasons for abuse and role of the institution. Twenty conduct code violators participated in an alcohol and drug education program and 35 RAs participated in an abbreviated program as part of…

  12. Inhibition of malignant thyroid carcinoma cell proliferation by Ras and galectin-3 inhibitors

    PubMed Central

    Menachem, A; Bodner, O; Pastor, J; Raz, A; Kloog, Y

    2015-01-01

    Anaplastic Thyroid carcinoma is an extremely aggressive solid tumor that resists most treatments and is almost always fatal. Galectin-3 (Gal-3) is an important marker for thyroid carcinomas and a scaffold of the K-Ras protein. S-trans, transfarnesylthiosalicylic acid (FTS; Salirasib) is a Ras inhibitor that inhibits the active forms of Ras proteins. Modified citrus pectin (MCP) is a water-soluble citrus-fruit-derived polysaccharide fiber that specifically inhibits Gal-3. The aim of this study was to develop a novel drug combination designed to treat aggressive anaplastic thyroid carcinoma. Combined treatment with FTS and MCP inhibited anaplastic thyroid cells proliferation in vitro by inducing cell cycle arrest and increasing apoptosis rate. Immunoblot analysis revealed a significant decrease in Pan-Ras, K-Ras, Ras-GTP, p-ERK, p53, and Gal-3 expression levels and significant increase in p21 expression levels. In nude mice, treatment with FTS and MCP inhibited tumor growth. Levels of Gal-3, K-Ras-GTP, and p-ERK were significantly decreased. To conclude, our results suggest K-Ras and Gal-3 as potential targets in anaplastic thyroid tumors and herald a novel treatment for highly aggressive anaplastic thyroid carcinoma. PMID:27551476

  13. Rational design of small molecule inhibitors targeting the Ras GEF, SOS1

    PubMed Central

    Evelyn, Chris R.; Duan, Xin; Biesiada, Jacek; Seibel, William L.; Meller, Jaroslaw; Zheng, Yi

    2014-01-01

    Summary Ras GTPases regulate intracellular signaling involved in cell proliferation. Elevated Ras signaling activity has been associated with human cancers. Ras activation is catalyzed by guanine-nucleotide exchange factors (GEFs), of which SOS1 is a major member that transduces receptor tyrosine kinase signaling to Ras. We have developed a rational approach coupling virtual screening with experimental screening in identifying small-molecule inhibitors targeting the catalytic site of SOS1 and SOS1-regulated Ras activity. A lead inhibitor, NSC-658497, is found to bind to SOS1, competitively suppresses SOS1-Ras interaction, and dose-dependently inhibits SOS1 GEF activity. Mutagenesis and structure-activity relationship studies map the NSC-658497 site of action to the SOS1 catalytic site, and define the chemical moieties in the inhibitor essential for the activity. NSC-658497 showed dose-dependent efficacy in inhibiting Ras, downstream signaling activities, and associated cell proliferation. These studies establish a proof of principle for rational design of small-molecule inhibitors targeting Ras GEF enzymatic activity. PMID:25455859

  14. Rational design of small molecule inhibitors targeting the Ras GEF, SOS1.

    PubMed

    Evelyn, Chris R; Duan, Xin; Biesiada, Jacek; Seibel, William L; Meller, Jaroslaw; Zheng, Yi

    2014-12-18

    Ras GTPases regulate intracellular signaling involved in cell proliferation. Elevated Ras signaling activity has been associated with human cancers. Ras activation is catalyzed by guanine nucleotide exchange factors (GEFs), of which SOS1 is a major member that transduces receptor tyrosine kinase signaling to Ras. We have developed a rational approach coupling virtual screening with experimental screening in identifying small-molecule inhibitors targeting the catalytic site of SOS1 and SOS1-regulated Ras activity. A lead inhibitor, NSC-658497, was found to bind to SOS1, competitively suppress SOS1-Ras interaction, and dose-dependently inhibit SOS1 GEF activity. Mutagenesis and structure-activity relationship studies map the NSC-658497 site of action to the SOS1 catalytic site, and define the chemical moieties in the inhibitor essential for the activity. NSC-658497 showed dose-dependent efficacy in inhibiting Ras, downstream signaling activities, and associated cell proliferation. These studies establish a proof of principle for rational design of small-molecule inhibitors targeting Ras GEF enzymatic activity.

  15. Inhibition of malignant thyroid carcinoma cell proliferation by Ras and galectin-3 inhibitors.

    PubMed

    Menachem, A; Bodner, O; Pastor, J; Raz, A; Kloog, Y

    2015-01-01

    Anaplastic Thyroid carcinoma is an extremely aggressive solid tumor that resists most treatments and is almost always fatal. Galectin-3 (Gal-3) is an important marker for thyroid carcinomas and a scaffold of the K-Ras protein. S-trans, transfarnesylthiosalicylic acid (FTS; Salirasib) is a Ras inhibitor that inhibits the active forms of Ras proteins. Modified citrus pectin (MCP) is a water-soluble citrus-fruit-derived polysaccharide fiber that specifically inhibits Gal-3. The aim of this study was to develop a novel drug combination designed to treat aggressive anaplastic thyroid carcinoma. Combined treatment with FTS and MCP inhibited anaplastic thyroid cells proliferation in vitro by inducing cell cycle arrest and increasing apoptosis rate. Immunoblot analysis revealed a significant decrease in Pan-Ras, K-Ras, Ras-GTP, p-ERK, p53, and Gal-3 expression levels and significant increase in p21 expression levels. In nude mice, treatment with FTS and MCP inhibited tumor growth. Levels of Gal-3, K-Ras-GTP, and p-ERK were significantly decreased. To conclude, our results suggest K-Ras and Gal-3 as potential targets in anaplastic thyroid tumors and herald a novel treatment for highly aggressive anaplastic thyroid carcinoma. PMID:27551476

  16. Multilayer Laue Lens Sequence Compiler

    2005-10-01

    For the growth of a new kind of x-ray focusing optic called a multilayer Laue lens, a device is constructed in which each layer of alernating high-z and low-z is placed in the appropriate place according to the Fresnel zone plate law. This requires that each layer have a different layer thickness. Because each layer is grown using DC magnetron sputter deposition, these layer thicknesses are not only dictated by the zone plate law, butmore » are adjusted to account for various drifting in the growth chamber due to target erosion, etc.« less

  17. Multilayer Laue Lens Sequence Compiler

    SciTech Connect

    Conley, Roy; Liu, Chian

    2005-10-01

    For the growth of a new kind of x-ray focusing optic called a multilayer Laue lens, a device is constructed in which each layer of alernating high-z and low-z is placed in the appropriate place according to the Fresnel zone plate law. This requires that each layer have a different layer thickness. Because each layer is grown using DC magnetron sputter deposition, these layer thicknesses are not only dictated by the zone plate law, but are adjusted to account for various drifting in the growth chamber due to target erosion, etc.

  18. Plasmonic lens for ultraviolet wavelength

    NASA Astrophysics Data System (ADS)

    Takeda, Minoru; Tanimoto, Takuya; Inoue, Tsutomu; Aizawa, Kento

    2016-09-01

    A plasmonic lens (PL) is one of the promising photonic devices utilizing the surface plasmon wave. In this study, we have newly developed a PL with a 3.5 µm diameter for a wavelength of 375 nm (ultraviolet region). It is composed of multiple circular slit apertures milled in aluminum (Al) thin film. We have simulated the electric field distribution of the PL, and confirmed that a tightly focused beam spot of subwavelength size in the far-field region was attained. We have also measured the focusing characteristics of the PL using a near-field scanning optical microscope (NSOM) and compared them with the calculated results.

  19. Health Through the Urban Lens

    PubMed Central

    2008-01-01

    Cities are now the major sites of human habitation worldwide, a trend that will continue for the foreseeable future, not only in the developed world but in developing countries. Urban residence impacts health and health prospects both positively and negatively through a complex mix of exposures and mechanisms. In addition, cities concentrate population subsets of various demographic, economic, and social characteristics, some with particular health risks and vulnerabilities. Looking at health through the urban lens allows increased understanding of disparate risks and emphasizes the essentiality of collaborative efforts in protecting and enhancing the health of populations, especially those living in cities. PMID:18668367

  20. K-ras gene mutation in gall bladder carcinomas and dysplasia.

    PubMed Central

    Ajiki, T; Fujimori, T; Onoyama, H; Yamamoto, M; Kitazawa, S; Maeda, S; Saitoh, Y

    1996-01-01

    Epithelial dysplasia of gall bladder is an important precancerous lesion of gall bladder carcinogenesis. To investigate the frequency of K-ras gene mutation in gall bladder carcinoma and dysplasia, K-ras codon 12 mutations were investigated by the polymerase chain reaction/restriction enzyme based method following direct sequencing. Mutation was detected in 59% (30 of 51) of gall bladder carcinomas, in 73% (8 of 11) of gall bladder dysplasia in gall stone cases, and in 0% of the normal gall bladder epithelium. There was, however, no correlation between K-ras mutation and clinicopathological factors of gall bladder carcinoma. K-ras gene mutation occurs even in gall bladder dysplasia at an incidence similar to that in carcinomas, suggesting that testing for K-ras gene mutation may prove useful as an adjunct to bile cytological or biopsy analysis. Images Figure 1 Figure 2 Figure 3 PMID:8675098

  1. A new member of the ras gene superfamily identified in a rat liver cell line.

    PubMed Central

    Bucci, C; Frunzio, R; Chiariotti, L; Brown, A L; Rechler, M M; Bruni, C B

    1988-01-01

    A new member of the ras genes superfamily was isolated from a cDNA library derived from a rat liver cell line (BRL-3A). The predicted 201 amino acids ras-like protein shows 30-35% homology with other members of the ras and ras-related gene products so far described. Conserved features include the GTP-binding and hydrolysis domains and the carboxyl terminal cysteine residues. A protein of the expected size (Mr 23,000) was synthesized in an in vitro transcription-translation system. The BRL-ras gene is present in single copy in the rat genome and is ubiquitously expressed at high levels in all tissues and cell lines examined. Images PMID:3057452

  2. Fas-induced programmed cell death is mediated by a Ras-regulated O2- synthesis.

    PubMed Central

    Gulbins, E; Brenner, B; Schlottmann, K; Welsch, J; Heinle, H; Koppenhoefer, U; Linderkamp, O; Coggeshall, K M; Lang, F

    1996-01-01

    Fas induces apoptosis in lymphocytes via a poorly defined intracellular signalling cascade. Previously, we have demonstrated the involvement and significance of a signalling cascade from the Fas receptor via sphingomyelinases and ceramide to Ras in Fas-induced apoptosis. Here we demonstrate rapid and transient synthesis of reactive oxygen intermediates (ROI) via activation of Ras after Fas. Genetic inhibition of Ras by transfection of transdominant inhibitory N17Ras blocked Fas-mediated ROI synthesis and programmed cell death. Likewise, the antioxidants N-acetyl-cysteine and N-t-butyl-phenylnitrone abolished Fas-induced cell death, pointing to an important role for Ras-triggered ROI synthesis in Fas-mediated programmed cell death. Images Figure 1 Figure 3 PMID:8943716

  3. Ras gene activation in gastric adenocarcinoma of Chinese patients in Taiwan

    SciTech Connect

    Tzeng, C.C.; Lee, W.Y.; Jin, Y.T.

    1994-09-01

    In order to assess the implication of mutational activation of members of the ras family of cellular proto-oncogenes in the development of gastric cancers in Chinese patients, a series of 55 cases of gastric adenocarcinoma in Taiwan was studied. Genomic deoxyribonucleic acid obtained from formalin-fixed paraffin-embedded archival tumor tissue was amplified by polymerase chain reaction and then analyzed by dot blot hybridation assay with allele-specific oligonucleotide probes to detect mutations at codons 12, 13, and 61 of c-Ki-ras, c-Ha-ras, and c-N-ras. Twelve (12.8%) of the 55 carcinomas examined harbored a point mutation. Of the 12 mutations, 8 (66.6%) were detected in Ha-ras codon 12. Our result is consistent with reports from mainland China and Korea, but different from those of Japan and the United States. This difference is probably attributable to different eating and drinking habits.

  4. cpRAS: a novel circularly permuted RAS-like GTPase domain with a highly scattered phylogenetic distribution.

    PubMed

    Elias, Marek; Novotny, Marian

    2008-01-01

    A recent systematic survey suggested that the YRG (or YawG/YlqF) family with the G4-G5-G1-G2-G3 order of the conserved GTPase motifs represents the only possible circularly permuted variation of the canonical GTPase structure. Here we show that a different circularly permuted GTPase domain actually does exist, conforming to the pattern G3-G4-G5-G1-G2. The domain, dubbed cpRAS, is a variant of RAS family GTPases and occurs in two types of larger proteins, either inserted into a region homologous to a bacterial group of proteins classified as COG2373 and potentially related to the alpha-2-macroglobulin family (so far a single protein in Dictyostelium) or in combination with a von Willebrand factor type A (VWA) domain. For the latter protein type, which was found in a few metazoans and several distantly related protists, existence in the common ancestor of opisthokonts, Amoebozoa and excavates followed by at least eight independent losses may be inferred. Our findings thus bring further evidence for the importance of parallel reduction of ancestral complexity in the eukaryotic evolution.

  5. Propiconazole-enhanced hepatic cell proliferation is associated with dysregulation of the cholesterol biosynthesis pathway leading to activation of Erk1/2 through Ras farnesylation.

    PubMed

    Murphy, Lynea A; Moore, Tanya; Nesnow, Stephen

    2012-04-15

    Propiconazole is a mouse hepatotumorigenic fungicide designed to inhibit CYP51, a key enzyme in the biosynthesis of ergosterol in fungi and is widely used in agriculture to prevent fungal growth. Metabolomic studies in mice revealed that propiconazole increased levels of hepatic cholesterol metabolites and bile acids, and transcriptomic studies revealed that genes within the cholesterol biosynthesis, cholesterol metabolism and bile acid biosyntheses pathways were up-regulated. Hepatic cell proliferation was also increased by propiconazole. AML12 immortalized hepatocytes were used to study propiconazole's effects on cell proliferation focusing on the dysregulation of cholesterol biosynthesis and resulting effects on Ras farnesylation and Erk1/2 activation as a primary pathway. Mevalonate, a key intermediate in the cholesterol biosynthesis pathway, increases cell proliferation in several cancer cell lines and tumors in vivo and serves as the precursor for isoprenoids (e.g. farnesyl pyrophosphate) which are crucial in the farnesylation of the Ras protein by farnesyl transferase. Farnesylation targets Ras to the cell membrane where it is involved in signal transduction, including the mitogen-activated protein kinase (MAPK) pathway. In our studies, mevalonic acid lactone (MVAL), a source of mevalonic acid, increased cell proliferation in AML12 cells which was reduced by farnesyl transferase inhibitors (L-744,832 or manumycin) or simvastatin, an HMG-CoA reductase inhibitor, indicating that this cell system responded to alterations in the cholesterol biosynthesis pathway. Cell proliferation in AML12 cells was increased by propiconazole which was reversed by co-incubation with L-744,832 or simvastatin. Increasing concentrations of exogenous cholesterol muted the proliferative effects of propiconazole and the inhibitory effects of L-733,832, results ascribed to reduced stimulation of the endogenous cholesterol biosynthesis pathway. Western blot analysis of subcellular

  6. Hyperglycemia Promotes K-Ras-Induced Lung Tumorigenesis through BASCs Amplification

    PubMed Central

    Micucci, Carla; Orciari, Silvia; Catalano, Alfonso

    2014-01-01

    Oncogenic K-Ras represents the most common molecular change in human lung adenocarcinomas, the major histologic subtype of non–small cell lung cancer (NSCLC). The presence of K-Ras mutation is associated with a poor prognosis, but no effective treatment strategies are available for K-Ras -mutant NSCLC. Epidemiological studies report higher lung cancer mortality rates in patients with type 2 diabetes. Here, we use a mouse model of K-Ras-mediated lung cancer on a background of chronic hyperglycemia to determine whether elevated circulating glycemic levels could influence oncogenic K-Ras-mediated tumor development. Inducible oncogenic K-Ras mouse model was treated with subtoxic doses of streptozotocin (STZ) to induce chronic hyperglycemia. We observed increased tumor mass and higher grade of malignancy in STZ treated diabetic mice analyzed at 4, 12 and 24 weeks, suggesting that oncogenic K-Ras increased lung tumorigenesis in hyperglycemic condition. This promoting effect is achieved by expansion of tumor-initiating lung bronchio-alveolar stem cells (BASCs) in bronchio-alveolar duct junction, indicating a role of hyperglycemia in the activity of K-Ras-transformed putative lung stem cells. Notably, after oncogene K-Ras activation, BASCs show upregulation of the glucose transporter (Glut1/Slc2a1), considered as an important player of the active control of tumor cell metabolism by oncogenic K-Ras. Our novel findings suggest that anti-hyperglycemic drugs, such as metformin, may act as therapeutic agent to restrict lung neoplasia promotion and progression. PMID:25144301

  7. Insights into K-Ras 4B regulation by post-translational lysine acetylation.

    PubMed

    Knyphausen, Philipp; Lang, Franziska; Baldus, Linda; Extra, Antje; Lammers, Michael

    2016-10-01

    Ras is a molecular switch cycling between an active, GTP-bound and an inactive, GDP-bound state. Mutations in Ras, mostly affecting the off-switch, are found in many human tumours. Recently, it has been shown that K-Ras 4B is targeted by lysine acetylation at K104. Based on results obtained for an acetylation mimetic Ras mutant (K104Q), it was hypothesised that K104-acetylation might interfere with its oncogenicity by impairing SOS-catalysed guanine-nucleotide exchange. We prepared site-specifically K104-acetylated K-Ras 4B and the corresponding oncogenic mutant protein G12V using the genetic-code expansion concept. We found that SOS-catalysed nucleotide exchange, also of allosterically activated SOS, was neither affected by acetylation of K104 in wildtype K-Ras 4B nor in the G12V mutant, suggesting that glutamine is a poor mimetic for acetylation at this site. In vitro, the lysine-acetyltransferases CBP and p300 were able to acetylate both, wildtype and G12V K-Ras 4B. In addition to K104 we identified further acetylation sites in K-Ras 4B, including K147, within the important G5/SAK-motif. However, the intrinsic and the SOS-catalysed nucleotide exchange was not affected by K147-acetylation of K-Ras 4B. Finally, we show that Sirt2 and HDAC6 do neither deacetylate K-Ras 4B if acetylated at K104 nor if acetylated at K147 in vitro.

  8. Lead identification for the K-Ras protein: virtual screening and combinatorial fragment-based approaches

    PubMed Central

    Pathan, Akbar Ali Khan; Panthi, Bhavana; Khan, Zahid; Koppula, Purushotham Reddy; Alanazi, Mohammed Saud; Sachchidanand; Parine, Narasimha Reddy; Chourasia, Mukesh

    2016-01-01

    Objective Kirsten rat sarcoma (K-Ras) protein is a member of Ras family belonging to the small guanosine triphosphatases superfamily. The members of this family share a conserved structure and biochemical properties, acting as binary molecular switches. The guanosine triphosphate-bound active K-Ras interacts with a range of effectors, resulting in the stimulation of downstream signaling pathways regulating cell proliferation, differentiation, and apoptosis. Efforts to target K-Ras have been unsuccessful until now, placing it among high-value molecules against which developing a therapy would have an enormous impact. K-Ras transduces signals when it binds to guanosine triphosphate by directly binding to downstream effector proteins, but in case of guanosine diphosphate-bound conformation, these interactions get disrupted. Methods In the present study, we targeted the nucleotide-binding site in the “on” and “off” state conformations of the K-Ras protein to find out suitable lead compounds. A structure-based virtual screening approach has been used to screen compounds from different databases, followed by a combinatorial fragment-based approach to design the apposite lead for the K-Ras protein. Results Interestingly, the designed compounds exhibit a binding preference for the “off” state over “on” state conformation of K-Ras protein. Moreover, the designed compounds’ interactions are similar to guanosine diphosphate and, thus, could presumably act as a potential lead for K-Ras. The predicted drug-likeness properties of these compounds suggest that these compounds follow the Lipinski’s rule of five and have tolerable absorption, distribution, metabolism, excretion and toxicity values. Conclusion Thus, through the current study, we propose targeting only “off” state conformations as a promising strategy for the design of reversible inhibitors to pharmacologically inhibit distinct conformations of K-Ras protein. PMID:27217775

  9. Across the universe of K-RAS mutations in non-small-cell-lung cancer.

    PubMed

    Piva, Sheila; Ganzinelli, Monica; Garassino, Marina Chiara; Caiola, Elisa; Farina, Gabriella; Broggini, Massimo; Marabese, Mirko

    2014-01-01

    RAS family proteins are important signaling molecules that regulate cell growth, survival and differentiation by coupling receptor activation to downstream effector pathways. Three distinct genes encode for the three different proteins H-, K-, and N- RAS. These proteins share high sequence homology, particularly at the N-Terminal domain. Among them, K-RAS is one of the most frequently mutated in human cancer. The majority of the mutations present in K-RAS are at codon 12 (from 80 to 100%) followed by codon 13 and 61. In all cases, aminoacid change leads to a constitutively activated protein. K-RAS mutations have a role in tumor development as well as in tumor progression and resistance. Despite the various studies which have been published, the prognostic and predictive role of K-RAS mutations is still under debate. Keeping in mind that the glycine present at position 12 can be substituted by valine, aspartic acid or cysteine, it could be well understood that each different substitution plays a different role in K-RAS-dependent processes. The present article focuses on the molecular and biological characteristics of K-RAS protein, its role in NSCLC tumor development and progression. We also present an overview of the preclinical models both in vitro and in vivo available to determine the role of K-RAS in tumor progression and response to treatment and on the recent results obtained in this field. Finally, we have considered the impact of KRAS mutations in clinical practice, analyzing the different recent trials that have taken into consideration K-RAS.

  10. Lack of R-Ras Leads to Increased Vascular Permeability in Ischemic Retinopathy

    PubMed Central

    Vähätupa, Maria; Prince, Stuart; Vataja, Suvi; Mertimo, Teija; Kataja, Marko; Kinnunen, Kati; Marjomäki, Varpu; Uusitalo, Hannu; Komatsu, Masanobu; Järvinen, Tero A.H.; Uusitalo–Järvinen, Hannele

    2016-01-01

    Purpose The role of R-Ras in retinal angiogenesis and vascular permeability was evaluated in an oxygen-induced retinopathy (OIR) model using R-Ras knockout (KO) mice and in human diabetic neovascular membranes. Methods Mice deficient for R-Ras and their wild-type (WT) littermates were subjected to 75% oxygen from postnatal day 7 (P7) to P12 and then returned to room air. At P17 retinal vascularization was examined from whole mounts, and retinal vascular permeability was studied using Miles assay. Real-time RT-PCR, Western blotting, and immunohistochemistry were used to assess the expression of R-Ras in retina during development or in the OIR model. The degree of pericyte coverage and vascular endothelial (VE)-cadherin expression on WT and R-Ras KO retinal blood vessels was quantified using confocal microscopy. The correlation of R-Ras with vascular endothelial growth factor receptor 2 (VEGFR2) and human serum albumin on human proliferative diabetic retinopathy membranes was assessed using immunohistochemistry. Results In retina, R-Ras expression was mostly restricted to the vasculature. Retinal vessels in the R-Ras KO mice were significantly more permeable than WT controls in the OIR model. A significant reduction in the direct physical contact between pericytes and blood vessel endothelium as well as reduced VE-cadherin immunostaining was found in R-Ras–deficient mice. In human proliferative diabetic retinopathy neovascular membranes, R-Ras expression negatively correlated with increased vascular leakage and expression of VEGFR2, a marker of blood vessel immaturity. Conclusions Our results suggest that R-Ras has a role in controlling retinal vessel maturation and stabilization in ischemic retinopathy and provides a potential target for pharmacologic manipulation to treat diabetic retinopathy. PMID:27654416

  11. Change of optical design thought about focusing of zoom lens

    NASA Astrophysics Data System (ADS)

    Hagimori, Hitoshi

    2015-09-01

    Zoom lens has been developed around lens applications of consumer still camera and TV broadcast cameras from about 1960s. Among, zoom lens as an interchangeable lens of a single-lens camera has made the most significant evolution in technically. In this paper, I describe the change of optical design concept about focusing function in zoom lens including introduction of some topic specific lenses.

  12. Mechanically tunable photonic crystal lens

    NASA Astrophysics Data System (ADS)

    Cui, Y.; Tamma, V. A.; Lee, J.-B.; Park, W.

    2010-08-01

    We designed, fabricated and characterized MEMS-enabled mechanically-tunable photonic crystal lens comprised of 2D photonic crystal and symmetrical electro-thermal actuators. The 2D photonic crystal was made of a honeycomb-lattice of 340 nm thick, 260 nm diameter high-index silicon rods embedded in low-index 10 μm thick SU-8 cladding. Silicon input waveguide and deflection block were also fabricated for light in-coupling and monitoring of focused spot size, respectively. When actuated, the electro-thermal actuators induced mechanical strain which changed the lattice constant of the photonic crystal and consequently modified the photonic band structure. This in turn modified the focal-length of the photonic crystal lens. The fabricated device was characterized using a tunable laser (1400~1602 nm) and an infrared camera during actuation. At the wavelength of 1450 nm, the lateral light spot size observed at the deflection block gradually decreased 40%, as applied current increased from 0 to 0.7 A, indicating changes in focal length in response to the mechanical stretching.

  13. Fabricating customized hydrogel contact lens

    PubMed Central

    Childs, Andre; Li, Hao; Lewittes, Daniella M.; Dong, Biqin; Liu, Wenzhong; Shu, Xiao; Sun, Cheng; Zhang, Hao F.

    2016-01-01

    Contact lenses are increasingly used in laboratories for in vivo animal retinal imaging and pre-clinical studies. The lens shapes often need modification to optimally fit corneas of individual test subjects. However, the choices from commercially available contact lenses are rather limited. Here, we report a flexible method to fabricate customized hydrogel contact lenses. We showed that the fabricated hydrogel is highly transparent, with refractive indices ranging from 1.42 to 1.45 in the spectra range from 400 nm to 800 nm. The Young’s modulus (1.47 MPa) and hydrophobicity (with a sessile drop contact angle of 40.5°) have also been characterized experimentally. Retinal imaging using optical coherence tomography in rats wearing our customized contact lenses has the quality comparable to the control case without the contact lens. Our method could significantly reduce the cost and the lead time for fabricating soft contact lenses with customized shapes, and benefit the laboratorial-used contact lenses in pre-clinical studies. PMID:27748361

  14. Fabricating customized hydrogel contact lens

    NASA Astrophysics Data System (ADS)

    Childs, Andre; Li, Hao; Lewittes, Daniella M.; Dong, Biqin; Liu, Wenzhong; Shu, Xiao; Sun, Cheng; Zhang, Hao F.

    2016-10-01

    Contact lenses are increasingly used in laboratories for in vivo animal retinal imaging and pre-clinical studies. The lens shapes often need modification to optimally fit corneas of individual test subjects. However, the choices from commercially available contact lenses are rather limited. Here, we report a flexible method to fabricate customized hydrogel contact lenses. We showed that the fabricated hydrogel is highly transparent, with refractive indices ranging from 1.42 to 1.45 in the spectra range from 400 nm to 800 nm. The Young’s modulus (1.47 MPa) and hydrophobicity (with a sessile drop contact angle of 40.5°) have also been characterized experimentally. Retinal imaging using optical coherence tomography in rats wearing our customized contact lenses has the quality comparable to the control case without the contact lens. Our method could significantly reduce the cost and the lead time for fabricating soft contact lenses with customized shapes, and benefit the laboratorial-used contact lenses in pre-clinical studies.

  15. Impedance of the amphibian lens.

    PubMed

    Duncan, G; Patmore, L; Pynsent, P B

    1981-03-01

    1. The electrical resistance of the perfused frog lens was measured using separate internal current passing and voltage measuring electrodes. 2. The resistance values obtained using voltage clamp and direct and alternating current techniques were in good agreement. 3. The voltage transients induced in response to current steps were multi-exponential in form. Increasing the external K concentration reduced both the amplitude of the voltage response and the rise time. 4. The impedance characteristics were investigated in more detail using alternating current analysis techniques. 5. In an equivalent-circuit modelling study it was assumed that there were two major pathways for current flow in the lens. The first through the surface membranes and the second through the inner fibre membranes via the narrow extracellular spaces. 6. The experimental impedance loci could not be adequately fitted by a simple two time constant model and a third time constant was introduced which may represent diffusion polarization effects in the extracellular spaces. 7. The three time constant model gave good and consistent fits to impedance data from a number of preparations. 8. The form of the impedance loci was also dependent on the external K concentration, but the only fitted parameter which changed consistently with external K was the surface membrane resistance (Rs).

  16. Impedance of the amphibian lens.

    PubMed Central

    Duncan, G; Patmore, L; Pynsent, P B

    1981-01-01

    1. The electrical resistance of the perfused frog lens was measured using separate internal current passing and voltage measuring electrodes. 2. The resistance values obtained using voltage clamp and direct and alternating current techniques were in good agreement. 3. The voltage transients induced in response to current steps were multi-exponential in form. Increasing the external K concentration reduced both the amplitude of the voltage response and the rise time. 4. The impedance characteristics were investigated in more detail using alternating current analysis techniques. 5. In an equivalent-circuit modelling study it was assumed that there were two major pathways for current flow in the lens. The first through the surface membranes and the second through the inner fibre membranes via the narrow extracellular spaces. 6. The experimental impedance loci could not be adequately fitted by a simple two time constant model and a third time constant was introduced which may represent diffusion polarization effects in the extracellular spaces. 7. The three time constant model gave good and consistent fits to impedance data from a number of preparations. 8. The form of the impedance loci was also dependent on the external K concentration, but the only fitted parameter which changed consistently with external K was the surface membrane resistance (Rs). PMID:6973626

  17. Visual evoked potential latency and contrast sensitivity in patients with posterior chamber intraocular lens implants.

    PubMed Central

    Howe, J. W.; Mitchell, K. W.; Mahabaleswara, M.; Abdel-Khalek, M. N.

    1986-01-01

    An electrophysiological investigation of visual evoked potential (VEP) latency and contrast sensitivity was performed in a group of 13 patients who had undergone extracapsular cataract surgery with posterior chamber lens implantation. In spite of good postoperative visual acuity, abnormalities were detected in nine of the group (69%). This study suggests that, even with successfully implanted lenses, there may be a reduction in visual function which could be the result of altered transmission through the plastic lenticulus or fibrosis of the posterior lens capsule, and/or subtle changes in retinal architecture, not observed ophthalmoscopically. PMID:3801366

  18. Electrically switchable liquid crystal Fresnel lens using UV-modified alignment film.

    PubMed

    Jeng, Shie-Chang; Hwang, Shug-June; Horng, Jing-Shyang; Lin, Kuo-Ren

    2010-12-01

    A simple method to make a switchable liquid crystal (LC) Fresnel lens with high diffraction efficiency and a low driving voltage was proposed based on the photo-induced surface modification of the vertical alignment layer. UV illumination alters the pretilt angle of alignment layers, a Fresnel zone-distribution hybrid alignment in the homeotropic LC cell can be straightforwardly achieved through UV exposure, yielding a concentric structure of the Fresnel phase LC lens. A remarkable diffraction efficiency of ~31.4%, close to the measured diffraction efficiency of the used Fresnel-zone-plate mask of 32%, was detected using a linearly polarized incident beam. PMID:21164982

  19. Variational data assimilation system "INM RAS - Black Sea"

    NASA Astrophysics Data System (ADS)

    Parmuzin, Eugene; Agoshkov, Valery; Assovskiy, Maksim; Giniatulin, Sergey; Zakharova, Natalia; Kuimov, Grigory; Fomin, Vladimir

    2013-04-01

    Development of Informational-Computational Systems (ICS) for Data Assimilation Procedures is one of multidisciplinary problems. To study and solve these problems one needs to apply modern results from different disciplines and recent developments in: mathematical modeling; theory of adjoint equations and optimal control; inverse problems; numerical methods theory; numerical algebra and scientific computing. The problems discussed above are studied in the Institute of Numerical Mathematics of the Russian Academy of Science (INM RAS) in ICS for Personal Computers (PC). Special problems and questions arise while effective ICS versions for PC are being developed. These problems and questions can be solved with applying modern methods of numerical mathematics and by solving "parallelism problem" using OpenMP technology and special linear algebra packages. In this work the results on the ICS development for PC-ICS "INM RAS - Black Sea" are presented. In the work the following problems and questions are discussed: practical problems that can be studied by ICS; parallelism problems and their solutions with applying of OpenMP technology and the linear algebra packages used in ICS "INM - Black Sea"; Interface of ICS. The results of ICS "INM RAS - Black Sea" testing are presented. Efficiency of technologies and methods applied are discussed. The work was supported by RFBR, grants No. 13-01-00753, 13-05-00715 and by The Ministry of education and science of Russian Federation, project 8291, project 11.519.11.1005 References: [1] V.I. Agoshkov, M.V. Assovskii, S.A. Lebedev, Numerical simulation of Black Sea hydrothermodynamics taking into account tide-forming forces. Russ. J. Numer. Anal. Math. Modelling (2012) 27, No.1, 5-31 [2] E.I. Parmuzin, V.I. Agoshkov, Numerical solution of the variational assimilation problem for sea surface temperature in the model of the Black Sea dynamics. Russ. J. Numer. Anal. Math. Modelling (2012) 27, No.1, 69-94 [3] V.B. Zalesny, N.A. Diansky, V

  20. Contact lens sensors in ocular diagnostics.

    PubMed

    Farandos, Nicholas M; Yetisen, Ali K; Monteiro, Michael J; Lowe, Christopher R; Yun, Seok Hyun

    2015-04-22

    Contact lenses as a minimally invasive platform for diagnostics and drug delivery have emerged in recent years. Contact lens sensors have been developed for analyzing the glucose composition of tears as a surrogate for blood glucose monitoring and for the diagnosis of glaucoma by measuring intraocular pressure. However, the eye offers a wider diagnostic potential as a sensing site and therefore contact lens sensors have the potential to improve the diagnosis and treatment of many diseases and conditions. With advances in polymer synthesis, electronics and micro/nanofabrication, contact lens sensors can be produced to quantify the concentrations of many biomolecules in ocular fluids. Non- or minimally invasive contact lens sensors can be used directly in a clinical or point-of-care setting to monitor a disease state continuously. This article reviews the state-of-the-art in contact lens sensor fabrication, their detection, wireless powering, and readout mechanisms, and integration with mobile devices and smartphones. High-volume manufacturing considerations of contact lenses are also covered and a case study of an intraocular pressure contact lens sensor is provided as an example of a successful product. This Review further analyzes the contact lens market and the FDA regulatory requirements for commercialization of contact lens sensors.

  1. Analysis of a Thin Optical Lens Model

    ERIC Educational Resources Information Center

    Ivchenko, Vladimir V.

    2011-01-01

    In this article a thin optical lens model is considered. It is shown that the limits of its applicability are determined not only by the ratio between the thickness of the lens and the modules of the radii of curvature, but above all its geometric type. We have derived the analytical criteria for the applicability of the model for different types…

  2. Mathematical Lens: How Much Can You Bench?

    ERIC Educational Resources Information Center

    Bolognese, Chris A.

    2013-01-01

    "How Much Can You Bench?" appears in the "Mathematical Lens" section of "Mathematics Teacher." "Mathematical Lens" uses photographs as a springboard for mathematical inquiry and appears in every issue of "Mathematics Teacher." This month the mathematics behind the photograph includes finding areas…

  3. A Fisheye Lens for Many Point PDV

    SciTech Connect

    Frogget, B.

    2011-11-01

    The features of the fisheye lens are illustrated, including a design with reflector prisms. The fisheye fiber map and the beam footprint are shown. Fisheye rough-angle metrology was done and results presented. Next steps are given, including a smaller top fisheye lens element, longer reflector prisms with better mounting, and different fiber arrangements.

  4. Mathematical Lens: Iron Gate, Metulla, Israel

    ERIC Educational Resources Information Center

    Mathematics Teacher, 2005

    2005-01-01

    The "Mathematical Lens" feature of "Mathematics Teacher," uses photographs as a springboard for mathematical inquiry. The goal of this feature is to encourage readers to see patterns and relationships that they can think about and extend in a mathematically playful way. In this edition of "Mathematical Lens," students analyze the shapes in an iron…

  5. Propiconazole-enhanced hepatic cell proliferation is associated with dysregulation of the cholesterol biosynthesis pathway leading to activation of Erk1/2 through Ras farnesylation

    SciTech Connect

    Murphy, Lynea A.; Moore, Tanya; Nesnow, Stephen

    2012-04-15

    Propiconazole is a mouse hepatotumorigenic fungicide designed to inhibit CYP51, a key enzyme in the biosynthesis of ergosterol in fungi and is widely used in agriculture to prevent fungal growth. Metabolomic studies in mice revealed that propiconazole increased levels of hepatic cholesterol metabolites and bile acids, and transcriptomic studies revealed that genes within the cholesterol biosynthesis, cholesterol metabolism and bile acid biosyntheses pathways were up-regulated. Hepatic cell proliferation was also increased by propiconazole. AML12 immortalized hepatocytes were used to study propiconazole's effects on cell proliferation focusing on the dysregulation of cholesterol biosynthesis and resulting effects on Ras farnesylation and Erk1/2 activation as a primary pathway. Mevalonate, a key intermediate in the cholesterol biosynthesis pathway, increases cell proliferation in several cancer cell lines and tumors in vivo and serves as the precursor for isoprenoids (e.g. farnesyl pyrophosphate) which are crucial in the farnesylation of the Ras protein by farnesyl transferase. Farnesylation targets Ras to the cell membrane where it is involved in signal transduction, including the mitogen-activated protein kinase (MAPK) pathway. In our studies, mevalonic acid lactone (MVAL), a source of mevalonic acid, increased cell proliferation in AML12 cells which was reduced by farnesyl transferase inhibitors (L-744,832 or manumycin) or simvastatin, an HMG-CoA reductase inhibitor, indicating that this cell system responded to alterations in the cholesterol biosynthesis pathway. Cell proliferation in AML12 cells was increased by propiconazole which was reversed by co-incubation with L-744,832 or simvastatin. Increasing concentrations of exogenous cholesterol muted the proliferative effects of propiconazole and the inhibitory effects of L-733,832, results ascribed to reduced stimulation of the endogenous cholesterol biosynthesis pathway. Western blot analysis of subcellular

  6. Revisiting G3BP1 as a RasGAP Binding Protein: Sensitization of Tumor Cells to Chemotherapy by the RasGAP 317–326 Sequence Does Not Involve G3BP1

    PubMed Central

    Annibaldi, Alessandro; Dousse, Aline; Martin, Sophie; Tazi, Jamal; Widmann, Christian

    2011-01-01

    RasGAP is a multifunctional protein that controls Ras activity and that is found in chromosomal passenger complexes. It also negatively or positively regulates apoptosis depending on the extent of its cleavage by caspase-3. RasGAP has been reported to bind to G3BP1 (RasGAP SH3-domain-binding protein 1), a protein regulating mRNA stability and stress granule formation. The region of RasGAP (amino acids 317–326) thought to bind to G3BP1 corresponds exactly to the sequence within fragment N2, a caspase-3-generated fragment of RasGAP, that mediates sensitization of tumor cells to genotoxins. While assessing the contribution of G3BP1 in the anti-cancer function of a cell-permeable peptide containing the 317–326 sequence of RasGAP (TAT-RasGAP317–326), we found that, in conditions where G3BP1 and RasGAP bind to known partners, no interaction between G3BP1 and RasGAP could be detected. TAT-RasGAP317–326 did not modulate binding of G3BP1 to USP10, stress granule formation or c-myc mRNA levels. Finally, TAT-RasGAP317–326 was able to sensitize G3BP1 knock-out cells to cisplatin-induced apoptosis. Collectively these results indicate that G3BP1 and its putative RasGAP binding region have no functional influence on each other. Importantly, our data provide arguments against G3BP1 being a genuine RasGAP-binding partner. Hence, G3BP1-mediated signaling may not involve RasGAP. PMID:22205990

  7. Revisiting G3BP1 as a RasGAP binding protein: sensitization of tumor cells to chemotherapy by the RasGAP 317-326 sequence does not involve G3BP1.

    PubMed

    Annibaldi, Alessandro; Dousse, Aline; Martin, Sophie; Tazi, Jamal; Widmann, Christian

    2011-01-01

    RasGAP is a multifunctional protein that controls Ras activity and that is found in chromosomal passenger complexes. It also negatively or positively regulates apoptosis depending on the extent of its cleavage by caspase-3. RasGAP has been reported to bind to G3BP1 (RasGAP SH3-domain-binding protein 1), a protein regulating mRNA stability and stress granule formation. The region of RasGAP (amino acids 317-326) thought to bind to G3BP1 corresponds exactly to the sequence within fragment N2, a caspase-3-generated fragment of RasGAP, that mediates sensitization of tumor cells to genotoxins. While assessing the contribution of G3BP1 in the anti-cancer function of a cell-permeable peptide containing the 317-326 sequence of RasGAP (TAT-RasGAP₃₁₇₋₃₂₆), we found that, in conditions where G3BP1 and RasGAP bind to known partners, no interaction between G3BP1 and RasGAP could be detected. TAT-RasGAP₃₁₇₋₃₂₆ did not modulate binding of G3BP1 to USP10, stress granule formation or c-myc mRNA levels. Finally, TAT-RasGAP₃₁₇₋₃₂₆ was able to sensitize G3BP1 knock-out cells to cisplatin-induced apoptosis. Collectively these results indicate that G3BP1 and its putative RasGAP binding region have no functional influence on each other. Importantly, our data provide arguments against G3BP1 being a genuine RasGAP-binding partner. Hence, G3BP1-mediated signaling may not involve RasGAP.

  8. Hepatitis C virus core protein cooperates with ras and transforms primary rat embryo fibroblasts to tumorigenic phenotype.

    PubMed Central

    Ray, R B; Lagging, L M; Meyer, K; Ray, R

    1996-01-01

    We have previously demonstrated that hepatitis C virus (HCV) core protein regulates cellular protooncogenes at the transcriptional level; this observation implicates core protein in the alteration of normal hepatocyte growth. In the present study, the transforming potential of the HCV core gene was investigated by using primary rat embryo fibroblast (REF) cells which were transfected with or without cooperative oncogenes. Integration of the HCV core gene resulted in expression of the viral protein in REF stable transformants. REF cells cotransfected with HCV core and H-ras genes became transformed and exhibited rapid proliferation, anchor-independent growth, and tumor formation in athymic nude mice. Results from these studies suggest that the core protein plays an important role in the regulation of HCV-infected cell growth and in the transformation to tumorigenic phenotype. These observations suggest a possible mechanism for this viral protein in the pathogenesis of hepatocellular carcinoma in HCV-infected humans. PMID:8676467

  9. The impact of the genetic background in the Noonan syndrome phenotype induced by K-RasV14I

    PubMed Central

    Hernández-Porras, Isabel; Jiménez-Catalán, Beatriz; Schuhmacher, Alberto J; Guerra, Carmen

    2015-01-01

    Noonan syndrome (NS) is an autosomal dominant genetic disorder characterized by short stature, craniofacial dysmorphism, and congenital heart defects. A significant fraction of NS-patients also develop myeloproliferative disorders. The penetrance of these defects varies considerably among patients. In this study, we have examined the effect of 2 genetic backgrounds (C57BL/6J.OlaHsd and 129S2/SvPasCrl) on the phenotypes displayed by a mouse model of NS induced by germline expression of the mutated K-RasV14I allele, one of the most frequent NS-KRAS mutations. Our results suggest the presence of genetic modifiers associated to the genetic background that are essential for heart development and function at early stages of postnatal life as well as in the severity of the haematopoietic alterations. PMID:26458870

  10. Oncogenic Ras activation of Raf/mitogen-activated protein kinase-independent pathways is sufficient to cause tumorigenic transformation.

    PubMed Central

    Khosravi-Far, R; White, M A; Westwick, J K; Solski, P A; Chrzanowska-Wodnicka, M; Van Aelst, L; Wigler, M H; Der, C J

    1996-01-01

    Substantial evidence supports a critical role for the activation of the Raf-1/MEK/mitogen-activated protein kinase pathway in oncogenic Ras-mediated transformation. For example, dominant negative mutants of Raf-1, MEK, and mitogen-activated protein kinase all inhibit Ras transformation. Furthermore, the observation that plasma membrane-localized Raf-1 exhibits the same transforming potency as oncogenic Ras suggests that Raf-1 activation alone is sufficient to mediate full Ras transforming activity. However, the recent identification of other candidate Ras effectors (e.g., RalGDS and phosphatidylinositol-3 kinase) suggests that activation of other downstream effector-mediated signaling pathways may also mediate Ras transforming activity. In support of this, two H-Ras effector domain mutants, H-Ras(12V, 37G) and H-Ras(12V, 40C), which are defective for Raf binding and activation, induced potent tumorigenic transformation of some strains of NIH 3T3 fibroblasts. These Raf-binding defective mutants of H-Ras induced a transformed morphology that was indistinguishable from that induced by activated members of Rho family proteins. Furthermore, the transforming activities of both of these mutants were synergistically enhanced by activated Raf-1 and inhibited by the dominant negative RhoA(19N) mutant, indicating that Ras may cause transformation that occurs via coordinate activation of Raf-dependent and -independent pathways that involves Rho family proteins. Finally, cotransfection of H-Ras(12V, 37G) and H-Ras(12V, 40C) resulted in synergistic cooperation of their focus-forming activities, indicating that Ras activates at least two Raf-independent, Ras effector-mediated signaling events. PMID:8668210

  11. Regulation of Ras Localization and Cell Transformation by Evolutionarily Conserved Palmitoyltransferases

    PubMed Central

    Young, Evelin; Zheng, Ze-Yi; Wilkins, Angela D.; Jeong, Hee-Tae; Li, Min; Lichtarge, Olivier

    2014-01-01

    Ras can act on the plasma membrane (PM) to mediate extracellular signaling and tumorigenesis. To identify key components controlling Ras PM localization, we performed an unbiased screen to seek Schizosaccharomyces pombe mutants with reduced PM Ras. Five mutants were found with mutations affecting the same gene, S. pombe erf2 (sp-erf2), encoding sp-Erf2, a palmitoyltransferase, with various activities. sp-Erf2 localizes to the trans-Golgi compartment, a process which is mediated by its third transmembrane domain and the Erf4 cofactor. In fission yeast, the human ortholog zDHHC9 rescues the phenotypes of sp-erf2 null cells. In contrast, expressing zDHHC14, another sp-Erf2-like human protein, did not rescue Ras1 mislocalization in these cells. Importantly, ZDHHC9 is widely overexpressed in cancers. Overexpressing ZDHHC9 promotes, while repressing it diminishes, Ras PM localization and transformation of mammalian cells. These data strongly demonstrate that sp-Erf2/zDHHC9 palmitoylates Ras proteins in a highly selective manner in the trans-Golgi compartment to facilitate PM targeting via the trans-Golgi network, a role that is most certainly critical for Ras-driven tumorigenesis. PMID:24248599

  12. The Thyroid Hormone Receptor Is a Suppressor of ras-Mediated Transcription, Proliferation, and Transformation

    PubMed Central

    García-Silva, Susana; Aranda, Ana

    2004-01-01

    The thyroid hormone triiodothyronine (T3) has a profound effect on growth, differentiation, and metabolism in higher organisms. Here we demonstrate that T3 inhibits ras-induced proliferation in neuroblastoma cells and blocks induction of cyclin D1 expression by the oncogene. The hormone, at physiological concentrations, strongly antagonizes the transcriptional response mediated by the Ras/mitogen-activated protein kinase/ribosomal-S6 subunit kinase (Rsk) signaling pathway in cells expressing thyroid hormone receptors (TRs). T3 blocks the response to the oncogenic forms of the three ras isoforms (H-, K-, and N-ras) and both TRα and TRβ can mediate this action. The main target for induction of cyclin D1 transcription by oncogenic ras in neuroblastoma cells is a cyclic AMP response element (CRE) located in proximal promoter sequences, and T3 represses the transcriptional activity of b-Zip transcription factors such as CREB (CRE-binding protein) or ATF-2 (activation transcription factor 2) that are direct targets of Rsk2 and bind to this sequence. The hormone also blocks fibroblast transformation by oncogenic ras when TR is expressed. Furthermore, TRs act as suppressors of tumor formation by the oncogene in vivo in nude mice. The TRβ isoform has stronger antitransforming properties than the α isoform and can inhibit tumorigenesis even in hypothyroid mice. These results show the existence of a previously unrecognized transcriptional cross talk between the TRs and the ras oncogene which influences relevant processes such as cell proliferation, transformation, or tumorigenesis. PMID:15314161

  13. Transgenic activation of Ras in neurons increases synapse formation in mouse neocortex.

    PubMed

    Seeger, G; Gärtner, U; Arendt, Th

    2005-06-01

    The small G protein Ras, which is a molecular switch in neurotrophic signal transduction, is implicated in synaptic plasticity and synapse development during ontogeny and in the adult nervous system. To characterise the involvement of Ras-dependent signaling in synaptogenesis, the cortical synapse-to-neuron ratio was investigated in synRas mice overexpressing Val12-Ha-Ras in postmitotic neurons (introduced by Heumann, 2000). The number of synapses per neuron was analysed in cortical layers II/III of the somatosensory cortex at different stages of postnatal development by stereological methods. The synapse-to-neuron ratio was still identical in wild-type and synRas mice at postnatal day 4 before the onset of transgene expression. At P12, P47 and in the adult, analyses revealed a significant increase in the synapse-to-neuron ratio in synRas mice which correlated with the strength of transgene expression. The data presented here provide evidence that Ras activity might be profoundly involved in synaptogenesis by reinforcing the formation or maintenance of synapses during the development and in the adult.

  14. Measuring Ras-family GTP levels in vivo--running hot and cold.

    PubMed

    Castro, Ariel F; Rebhun, John F; Quilliam, Lawrence A

    2005-10-01

    The detection of Ras-family GTPase activity is important in the determination of cell signaling events elicited by numerous ligands and cellular processes. This has been made much easier in recent years by the use of glutathione S-transferase (GST)-fused Ras binding domains. These domains from downstream effectors such as Raf and RalGDS preferentially bind the GTP-bound Ras proteins enabling their extraction and subsequent quantification by immunoblotting. Despite this advance, effectors that efficiently discriminate between GTP- and GDP-bound states are not available for many Ras-family members. While this hampers the ability to detect activity in tissue specimens, it is still possible to metabolically label cells with (32)Pi to load the GTP/GDP pool with labeled nucleotides, immunoprecipitate the Ras protein and detect the bound label following thin layer chromatographic separation and exposure to film or a phosphorimager. Using a transfection system and antibodies that recognize epitope tags one can test the ability of a protein to work as a GEF or GAP for a certain GTPase. Alternatively, if an immunoprecipitating antibody is available to the target GTPase, then analysis of endogenous GTP/GDP ratio is possible. Here we describe the detection of M-Ras and Rap1 activity by GST-RBD pull-down as well as that of Rheb and epitope-tagged R-Ras by classical metabolic labeling and immunoprecipitation.

  15. The Ras-Erk-ETS-Signaling Pathway Is a Drug Target for Longevity

    PubMed Central

    Slack, Cathy; Alic, Nazif; Foley, Andrea; Cabecinha, Melissa; Hoddinott, Matthew P.; Partridge, Linda

    2015-01-01

    Summary Identifying the molecular mechanisms that underlie aging and their pharmacological manipulation are key aims for improving lifelong human health. Here, we identify a critical role for Ras-Erk-ETS signaling in aging in Drosophila. We show that inhibition of Ras is sufficient for lifespan extension downstream of reduced insulin/IGF-1 (IIS) signaling. Moreover, direct reduction of Ras or Erk activity leads to increased lifespan. We identify the E-twenty six (ETS) transcriptional repressor, Anterior open (Aop), as central to lifespan extension caused by reduced IIS or Ras attenuation. Importantly, we demonstrate that adult-onset administration of the drug trametinib, a highly specific inhibitor of Ras-Erk-ETS signaling, can extend lifespan. This discovery of the Ras-Erk-ETS pathway as a pharmacological target for animal aging, together with the high degree of evolutionary conservation of the pathway, suggests that inhibition of Ras-Erk-ETS signaling may provide an effective target for anti-aging interventions in mammals. Video Abstract PMID:26119340

  16. Extended RAS analysis for anti-epidermal growth factor therapy in patients with metastatic colorectal cancer.

    PubMed

    Hecht, J Randolph; Douillard, Jean-Yves; Schwartzberg, Lee; Grothey, Axel; Kopetz, Scott; Rong, Alan; Oliner, Kelly S; Sidhu, Roger

    2015-09-01

    RAS family proteins (including KRAS and NRAS) play important roles in the epidermal growth factor receptor (EGFR) signaling pathway. Mutations in RAS genes (occurring at loci in exons 2, 3, and 4) often result in constitutive activation of RAS proteins and persistent downstream signaling. Mutations in KRAS exon 2 (codon 12/13) are an established predictor of lack of response to the anti-EGFR monoclonal antibodies cetuximab and panitumumab in patients with metastatic colorectal cancer (mCRC), and have been used routinely in clinical practice to identify patients unlikely to derive benefit from these therapies. However, a meaningful proportion of patients with mCRC have tumors bearing other mutations in RAS genes. Recent studies have demonstrated that evaluation of an extended panel of RAS mutations—including mutations in KRAS exon 2, 3, and 4 and NRAS exons 2, 3, and 4—can better define the patient population that is unlikely to benefit from anti-EGFR therapy, with concomitant improvements in outcomes in the more highly selected RAS wild-type group. This discovery has changed the practice of oncology and has the potential to spare patients from exposure to ineffective therapy. In the near future, it is important for the oncology community to validate extended RAS analysis assays and make certain that patients who are candidates for anti-EGFR therapy undergo appropriate testing and treatment.

  17. Lin28-let7 Modulates Radiosensitivity of Human Cancer Cells With Activation of K-Ras

    SciTech Connect

    Oh, Jee-Sun.; Kim, Jae-Jin; Byun, Ju-Yeon; Kim, In-Ah

    2010-01-15

    Purpose: To evaluate the potential of targeting Lin28-let7 microRNA regulatory network for overcoming the radioresistance of cancer cells having activated K-Ras signaling. Methods and Materials: A549 lung carcinoma cells and ASPC1 pancreatic cancer cells possessing K-RAS mutation were transfected with pre-let7a microRNA or Lin28 siRNA, respectively. Clonogenic assay, quantitative reverse transcription polymerase chain reaction, and Western analysis were performed. The effects of Lin28 on SQ20B cells having wild-type K-RAS, and a normal fibroblast were also assessed. Results: The overexpression of let-7a decreased expression of K-Ras and radiosensitized A549 cells. Inhibition of Lin28, a repressor of let-7, attenuated K-Ras expression and radiosensitized A549 and ASPC1 cells. Neither SQ20B cells expressing wild-type K-RAS nor HDF, the normal human fibroblasts, were radiosensitized by this approach. Conclusions: The Lin28-let7 regulatory network may be a potentially useful therapeutic target for overcoming the radioresistance of human cancers having activated K-Ras signaling.

  18. Assessment of the chemosensitizing activity of TAT-RasGAP317-326 in childhood cancers.

    PubMed

    Chevalier, Nadja; Gross, Nicole; Widmann, Christian

    2015-01-01

    Although current anti-cancer protocols are reasonably effective, treatment-associated long-term side effects, induced by lack of specificity of the anti-cancer procedures, remain a challenging problem in pediatric oncology. TAT-RasGAP317-326 is a RasGAP-derived cell-permeable peptide that acts as a sensitizer to various anti-cancer treatments in adult tumor cells. In the present study, we assessed the effect of TAT-RasGAP317-326 in several childhood cancer cell lines. The RasGAP-derived peptide-induced cell death was analyzed in several neuroblastoma, Ewing sarcoma and leukemia cell lines (as well as in normal lymphocytes). Cell death was evaluated using flow cytometry methods in the absence or in the presence of the peptide in combination with various genotoxins used in the clinics (4-hydroperoxycyclophosphamide, etoposide, vincristine and doxorubicin). All tested pediatric tumors, in response to at least one genotoxin, were sensitized by TAT-RasGAP317-326. The RasGAP-derived peptide did not increase cell death of normal lymphocytes, alone or in combination with the majority of the tested chemotherapies. Consequently, TAT-RasGAP317-326 may benefit children with tumors by increasing the efficacy of anti-cancer therapies notably by allowing reductions in anti-cancer drug dosage and the associated drug-induced side effects.

  19. The Ras-Erk-ETS-Signaling Pathway Is a Drug Target for Longevity.

    PubMed

    Slack, Cathy; Alic, Nazif; Foley, Andrea; Cabecinha, Melissa; Hoddinott, Matthew P; Partridge, Linda

    2015-07-01

    Identifying the molecular mechanisms that underlie aging and their pharmacological manipulation are key aims for improving lifelong human health. Here, we identify a critical role for Ras-Erk-ETS signaling in aging in Drosophila. We show that inhibition of Ras is sufficient for lifespan extension downstream of reduced insulin/IGF-1 (IIS) signaling. Moreover, direct reduction of Ras or Erk activity leads to increased lifespan. We identify the E-twenty six (ETS) transcriptional repressor, Anterior open (Aop), as central to lifespan extension caused by reduced IIS or Ras attenuation. Importantly, we demonstrate that adult-onset administration of the drug trametinib, a highly specific inhibitor of Ras-Erk-ETS signaling, can extend lifespan. This discovery of the Ras-Erk-ETS pathway as a pharmacological target for animal aging, together with the high degree of evolutionary conservation of the pathway, suggests that inhibition of Ras-Erk-ETS signaling may provide an effective target for anti-aging interventions in mammals.

  20. RAS testing in metastatic colorectal cancer: excellent reproducibility amongst 17 Dutch pathology centers

    PubMed Central

    Boleij, Annemarie; Tops, Bastiaan B.J.; Rombout, Paul D.M.; Dequeker, Elizabeth M.; Ligtenberg, Marjolijn J.L.; van Krieken, J. Han

    2015-01-01

    In 2013 the European Medicine Agency (EMA) restricted the indication for anti-EGFR targeted therapy to metastatic colorectal cancer (mCRC) with a wild-type RAS gene, increasing the need for reliable RAS mutation testing. We evaluated the completeness and reproducibility of RAS-testing in the Netherlands. From 17 laboratories, tumor DNA of the first 10 CRC cases tested in 2014 in routine clinical practice was re-tested by a reference laboratory using a custom next generation sequencing panel. In total, 171 CRC cases were re-evaluated for hotspot mutations in KRAS, NRAS and BRAF. Most laboratories had introduced complete RAS-testing (65%) and BRAF-testing (71%) by January 2014. The most employed method for all hotspot regions was Sanger sequencing (range 35.7 – 49.2%). The reference laboratory detected all mutations that had been found in the participating laboratories (n = 92), plus 10 additional mutations. This concerned three RAS and seven BRAF mutations that were missed due to incomplete testing of the participating laboratory. Overall, the concordance of tests performed by both the reference and participating laboratory was 100% (163/163; κ-static 1.0) for RAS and 100% (144/144; κ-static 1.0) for BRAF. Our study shows that RAS and BRAF mutations can be reproducibly assessed using a variety of testing methods. PMID:25944693

  1. RAS testing in metastatic colorectal cancer: excellent reproducibility amongst 17 Dutch pathology centers.

    PubMed

    Boleij, Annemarie; Tops, Bastiaan B J; Rombout, Paul D M; Dequeker, Elizabeth M; Ligtenberg, Marjolijn J L; van Krieken, J Han

    2015-06-20

    In 2013 the European Medicine Agency (EMA) restricted the indication for anti-EGFR targeted therapy to metastatic colorectal cancer (mCRC) with a wild-type RAS gene, increasing the need for reliable RAS mutation testing. We evaluated the completeness and reproducibility of RAS-testing in the Netherlands. From 17 laboratories, tumor DNA of the first 10 CRC cases tested in 2014 in routine clinical practice was re-tested by a reference laboratory using a custom next generation sequencing panel. In total, 171 CRC cases were re-evaluated for hotspot mutations in KRAS, NRAS and BRAF. Most laboratories had introduced complete RAS-testing (65%) and BRAF-testing (71%) by January 2014. The most employed method for all hotspot regions was Sanger sequencing (range 35.7 - 49.2%). The reference laboratory detected all mutations that had been found in the participating laboratories (n = 92), plus 10 additional mutations. This concerned three RAS and seven BRAF mutations that were missed due to incomplete testing of the participating laboratory. Overall, the concordance of tests performed by both the reference and participating laboratory was 100% (163/163; κ-static 1.0) for RAS and 100% (144/144; κ-static 1.0) for BRAF. Our study shows that RAS and BRAF mutations can be reproducibly assessed using a variety of testing methods. PMID:25944693

  2. Cross-talk between Ras and Rho signalling pathways in transformation favours proliferation and increased motility.

    PubMed

    Sahai, E; Olson, M F; Marshall, C J

    2001-02-15

    Transformation by oncogenic Ras requires the function of the Rho family GTPases. We find that Ras-transformed cells have elevated levels of RhoA-GTP, which functions to inhibit the expression of the cell cycle inhibitor p21/Waf1. These high levels of Rho-GTP are not a direct consequence of Ras signalling but are selected for in response to sustained ERK-MAP kinase signalling. While the elevated levels of Rho-GTP control the level of p21/Waf, they no longer regulate the formation of actin stress fibres in transformed cells. We show that the sustained ERK-MAP kinase signalling resulting from transformation by oncogenic Ras down-regulates ROCK1 and Rho-kinase, two Rho effectors required for actin stress fibre formation. The repression of Rho- dependent stress fibre formation by ERK-MAP kinase signalling contributes to the increased motility of Ras-transformed fibroblasts. Overexpression of the ROCK target LIM kinase restores actin stress fibres and inhibits the motility of Ras-transformed fibroblasts. We propose a model in which Ras and Rho signalling pathways cross-talk to promote signalling pathways favouring transformation.

  3. Explosive plane-wave lens

    DOEpatents

    Marsh, S.P.

    1988-03-08

    An explosive plane-wave air lens which enables a spherical wave form to be converted to a planar wave without the need to specially machine or shape explosive materials is described. A disc-shaped impactor having a greater thickness at its center than around its periphery is used to convert the spherical wave into a plane wave. When the wave reaches the impactor, the center of the impactor moves first because the spherical wave reaches the center of the impactor first. The wave strikes the impactor later in time as one moves radially along the impactor. Because the impactor is thinner as one moves radially outward, the velocity of the impactor is greater at the periphery than at the center. An acceptor explosive is positioned so that the impactor strikes the acceptor simultaneously. Consequently, a plane detonation wave is propagated through the acceptor explosive. 4 figs.

  4. Explosive plane-wave lens

    DOEpatents

    Marsh, Stanley P.

    1988-01-01

    An explosive plane-wave air lens which enables a spherical wave form to be converted to a planar wave without the need to specially machine or shape explosive materials is described. A disc-shaped impactor having a greater thickness at its center than around its periphery is used to convert the spherical wave into a plane wave. When the wave reaches the impactor, the center of the impactor moves first because the spherical wave reaches the center of the impactor first. The wave strikes the impactor later in time as one moves radially along the impactor. Because the impactor is thinner as one moves radially outward, the velocity of the impactor is greater at the periphery than at the center. An acceptor explosive is positioned so that the impactor strikes the acceptor simultaneously. Consequently, a plane detonation wave is propagated through the acceptor explosive.

  5. Explosive plane-wave lens

    DOEpatents

    Marsh, S.P.

    1987-03-12

    An explosive plane-wave air lens which enables a spherical wave form to be converted to a planar wave without the need to specially machine or shape explosive materials is described. A disc-shaped impactor having a greater thickness at its center than around its periphery is used to convert the spherical wave into a plane wave. When the wave reaches the impactor, the center of the impactor moves first because the spherical wave reaches the center of the impactor first. The wave strikes the impactor later in time as one moves radially along the impactor. Because the impactor is thinner as one moves radially outward, the velocity of the impactor is greater at the periphery than at the center. An acceptor explosive is positioned so that the impactor strikes the acceptor simultaneously. Consequently, a plane detonation wave is propagated through the acceptor explosive. 3 figs., 3 tabs.

  6. Studies on contact lens materials.

    PubMed

    Alyanak, H; Aksoy, S; Hasirci, N

    1991-02-01

    The development of plastics with the optical properties of glass led promptly to their use as contact lenses and intra-ocular lenses to rectify certain visual defects. Research to improve these polymeric materials is continuous but there is not much in the literature since most of the findings are patented. In this work, polymethyl methacrylate, the most commonly used lens material was chosen as the base material and its co and terpolymers were prepared using 2-hydroxyethyl methacrylate, N-vinyl-2-pyrrolidinone, hexamethyl disiloxane, and polypropylene glycol. The transparency, refractive index, contact angle, density, equilibrium water content, and percent hydration properties were examined. Theoretical values were calculated for linear expansion and oxygen permeation from the density and hydration values.

  7. Antisense treatment directed against mutated Ki-ras in human colorectal adenocarcinoma

    PubMed Central

    Andreyev, H; Ross, P; Cunningham, D; Clarke, P

    2001-01-01

    BACKGROUND—Kirsten ras (Ki-ras) mutations are common in gastrointestinal cancer and one codon 12 mutation, glycine to valine, is particularly aggressive in colorectal cancer.
AIMS—To investigate if this valine point mutation could be targeted with antisense oligonucleotides and to determine the efficacy of any antisense/mRNA interaction.
METHODS—Twenty nine antisense oligonucleotides were screened against target and control Ki-ras RNA in a cell free system and against target and control cell lines in culture.
RESULTS—The activity and specificity of the oligonucleotides varied. Results for the individual oligonucleotides were consistent in a cell free model and in cell culture using two different uptake promoters. Only one oligonucleotide was specific in its cleavage of target Ki-ras mRNA in the cell free system and appeared specific in cell culture, although changes in Ki-ras mRNA and protein expression following a single treatment could not be detected. Experiments in the cell free system showed that the point mutation is relatively inaccessible to oligonucleotides. Other sites on the Ki-ras RNA molecule, away from the point mutation, can be targeted more effectively.
CONCLUSIONS—Successful targeting of the clinically relevant Ki-ras point mutation with antisense oligonucleotides is difficult because of RNA structure at the mutated site and is inefficient compared with other sites on the Ki-ras mRNA.


Keywords: Ki-ras mutation; antisense treatment; colorectal carcinoma PMID:11156646

  8. Dominant inhibitory Ras delays Sindbis virus-induced apoptosis in neuronal cells.

    PubMed Central

    Joe, A K; Ferrari, G; Jiang, H H; Liang, X H; Levine, B

    1996-01-01

    Mature neurons are more resistant than dividing cells or differentiating neurons to Sindbis virus-induced apoptotic death. Therefore, we hypothesized that mitogenic signal transduction pathways may influence susceptibility to Sindbis virus-induced apoptosis. Since Ras, a 21-kDa GTP-binding protein, plays an important role in cellular proliferation and neuronal differentiation, we investigated the effect of an inducible dominant inhibitory Ras on Sindbis virus-induced death of a rat pheochromocytoma cell line, PC12 cells. Dexamethasone induction of dominant inhibitory Ras (Ha Ras(Asn17)) expression in transfected PC12 cell lines (MMTV-M17-21 and GSrasDN6 cells) resulted in a marked delay in Sindbis virus-induced apoptosis, compared with infected, uninduced cells. The delay in death after Sindbis virus infection in induced versus uninduced PC12 cells was not associated with differences in viral titers or viral infectivity. No delay in Sindbis virus-induced apoptosis was observed in Ha Ras(Asn17)-transfected PC12 cells if dexamethasone induction was initiated less than 12 h before Sindbis virus infection or in wild-type PC12 cells infected with a chimeric Sindbis virus construct that expresses Ha Ras(Asn17). The delay in Sindbis virus-induced apoptosis in induced Ha Ras(Asn17)-transfected PC12 cells was associated with a decrease in cellular DNA synthesis as measured by 5'-bromo-2'-deoxyuridine incorporation. Thus, in PC12 cells, inducible dominant inhibitory Ras inhibits cellular proliferation and delays Sindbis virus-induced apoptosis. These findings suggest that a Ras-dependent signaling pathway is a determinant of neuronal susceptibility to Sindbis virus-induced apoptosis. PMID:8892895

  9. Post-translational processing of purified human K-ras in Xenopus oocytes.

    PubMed

    Kaplan, J B; Sass, P M

    1991-01-01

    Membrane localization of ras p21 involves a complex series of post-translational processing events, including S-farnesylation of Cys-186, removal of three carboxyl-terminal amino acid residues, and methylation of the carboxyl-terminal farnesylcysteine residue. Palmitoylation of cysteine residues within the hypervariable region (amino acids 165-185) is also required for membrane localization of mammalian H-, N-, and K-ras(A). For K-ras(B), which contains no cysteine residues within the hypervariable region, a polybasic domain substitutes for palmitoylation as a second signal for plasma membrane targeting. In order to investigate the localization of K-ras(B) to the plasma membrane, we purified wild-type and mutant human K-ras(B) proteins from strains of E. coli harboring bacterial expression plasmids and injected them into Xenopus laevis oocytes. Our results show that wild-type and activated K-ras(B) proteins can be post-translationally modified and can induce meiotic maturation in Xenopus oocytes. A mutation at Cys-186 (Cys to Gly) abolished the ability of activated K-ras(B) to induce meiosis. Deprivation of isoprenyl precursors by the addition of lovastatin, a drug that blocks the synthesis of mevalonate, also abolished the ability of activated K-ras(B) to induce meiosis, although this inhibition could be overcome by the addition of exogenous mevalonate. Lovastatin did not block meiotic maturation induced by microinjection of purified mos protein, a component of the cytostatic factor that arrests Xenopus oocytes at the first meiotic prophase. These results indicate that post-translational isoprenylation of K-ras(B) is essential for plasma membrane targeting and induction of meiotic maturation in Xenopus oocytes and that further isoprenyl modification of proteins downstream from mos signal transduction is not essential for this process. PMID:16296004

  10. Variable focus crystal diffraction lens

    SciTech Connect

    Smither, R.K.

    1988-11-01

    A new method has been developed to control the shape of the surface of a diffracting crystal that will allow it to function as a variable focus crystal diffraction lens, for focusing photon beams from a synchrotron source. The new method uses thermal gradients in the crystal to control the shape of the surface of the crystal in two dimensions and allows one to generate both spherical and ellipsoidal surface shapes. In this work the thermal gradient was generated by core drilling two sets of cooling channels in a silicon crystal so that cooling or heating fluids could be circulated through the crystal at two different levels. The first set of channels is close to the surface of the crystal where the photon beam strikes it. The second set of channels is equal distant from the back surface. If a concave surface is desired, the fluid in the channels just below the surface exposed to the beam is cooler than the fluid circulating through the channels near the back surface. If a convex surface is desired, then the cooling fluid in the upper channels near the surface exposed to the incident photon beam, is warmer than the fluid in the lower channels. The focal length of the crystal lens is varied by varying the thermal gradient in the crystal. This approach can also be applied to the first crystal in a high power synchrotron beam line to eliminate the bowing and other thermal distortions of the crystal caused by the high heat load. 6 refs., 8 figs., 3 tabs.

  11. Microbial contamination of contact lens storage cases and domestic tap water of contact lens wearers.

    PubMed

    Üstüntürk, Miray; Zeybek, Zuhal

    2012-11-01

    Contact lenses have been widely used as an alternative to spectacles both in developed and developing countries. However, under certain circumstances, adverse responses can occur during contact lens wear and several microorganisms--including bacteria, fungi, and free living amoebae--can cause several eye infections in wearers. Extended wear of contact lenses is the major risk factor of eye infections such as microbial keratitis, besides contaminated contact lens storage case, contaminated lens care solutions, and inaccurate contact lens handling. In this study, we collected contact lens storage case and domestic tap water samples from 50 asymptomatic contact lens wearers. We determined that total aerobic mesophilic bacteria were isolated in 45 (90 %), Gram negative rod bacteria were isolated in 20 (40 %), Pseudomonas spp. were isolated in 2 (4 %) and fungi were isolated in 18 (36 %) out of 50 contact lens storage cases. Free living amoebae were not detected in investigated contact lens storage cases. At the same time, out of 50, total aerobic mesophilic bacteria were isolated in 34 (68 %), fungi were isolated in 15 (30 %) and free living amoebae were isolated in 15 (30 %) domestic tap water samples. No Gram-negative rod bacteria and Pseudomonas spp. were detected in investigated water samples. Two contact lens case samples and two tap water samples were excluded from the analysis for Pseudomonas spp. for technical reasons. According to our findings, inadequate contact lens maintenance during lens wear may result in the contamination of contact lens storage cases. This situation can lead to severe eye infections in contact lens wearers over time.

  12. Singlet mega-pixel resolution lens

    NASA Astrophysics Data System (ADS)

    Lin, Chen-Hung; Lin, Hoang Yan; Chang, Horng

    2008-03-01

    There always exist some new challenges for lens designers to keep their old-line technology update. To minimize lens volume is one of the most notified examples. In this paper we designed a single thick lens, constructed by using one oblique (reflective) surface, apart from two conventional refractive surfaces, to bend the optical path of the optical system to achieve this goal. Detail design procedure, including system layout and lens performance diagrams, will be presented. Following the first order layout, we applied aspherical form to the two refractive surfaces in order to correct the spherical aberration up to an acceptable condition. Then, the reduced aberrations such as coma, astigmatism, field curvature and distortion can easily be corrected with some calculations related to spherical aberration as shown in the publication of H. H. Hopkins (1950). Plastic material is used in the design, because the aspherical surfaces can then be manufactured in a more cost effective way. The final specification of the design is: EFL is 4.6 mm, the F number is 2.8, the over all thickness of lens is 3.6 mm, its MTF is 0.3 at 227 lp/mm in center field and chief ray angle is less than 15 degrees. Lens data as well as optical performance curves are also presented in the paper. In conclusion we have successfully finished a mega-pixel resolution lens design and its overall thickness is compatible with the state of the art.

  13. Microindentation of the young porcine ocular lens.

    PubMed

    Reilly, Matthew; Ravi, Nathan

    2009-04-01

    Debate regarding the mechanisms of how the eye changes focus (accommodation) and why this ability is lost with age (presbyopia) has recently been rejoined due to the advent of surgical procedures for the correction of presbyopia. Due to inherent confounding factors in both in vivo and in vitro measurement techniques, mechanical modeling of the behavior of the ocular lens in accommodation has been attempted to settle the debate. However, a paucity of reliable mechanical property measurements has proven problematic in the development of a successful mechanical model of accommodation. Instrumented microindentation was utilized to directly measure the local elastic modulus and dynamic response at various locations in the lens. The young porcine lens exhibits a large modulus gradient with the highest modulus appearing at the center of the nucleus and exponentially decreasing with distance. The loss tangent was significantly higher in the decapsulated lens and the force waveform amplitude decreased significantly upon removal of the lens capsule. The findings indicate that localized measurements of the lens' mechanical properties are necessary to achieve accurate quantitative parameters suitable for mechanical modeling efforts. The results also indicate that the lens behaves as a crosslinked gel rather than as a collection of individual arched fiber cells.

  14. Synergism between K-rasVal12 and mutant Apc accelerates murine large intestinal tumourigenesis.

    PubMed

    Luo, Feijun; Poulogiannis, George; Ye, Hongtao; Hamoudi, Rifat; Arends, Mark J

    2011-07-01

    K-ras (KRAS) is mutated in 40-50% of human colorectal adenomas and carcinomas and plays key roles in cell proliferation, apoptosis, motility and differentiation, but its functional contribution to intestinal tumourigenesis in vivo remains incompletely understood. We have previously crossed K-rasVal12 transgenic mice with Ah-Cre mice to produce K-rasVal12/Cre offspring that inducibly express K-rasVal12 4A and 4B in the intestines, but this alone showed no significant effect on intestinal adenoma formation. Here, we crossed these mice with Min mice to evaluate the effect of K-rasVal12 and Apc mutation on intestinal tumourigenesis in vivo. The double mutant K-rasVal12/Cre/ApcMin/+ mice showed a moderate (1.86-fold) increase in adenomas in the small intestines, but a striking acceleration (6-fold increase) of large intestinal adenoma formation (P<0.01) and significantly reduced survival (by ~5 weeks) compared with control ApcMin/+ mice (P<0.01). There was recombination of the mutant K-rasVal12 transgene in 80% of large intestinal adenomas with expression of both K-rasVal12 4A and 4B isoform transcripts and expression of K-RasVal12 protein. The large intestinal adenomas showed immunohistochemical evidence of activation of MapK, Akt and Wnt signaling pathways and this was confirmed by quantitative RT-PCR analysis of relative transcript expression levels of target genes using a panel of 23 selected genes evaluated in both adenomas and non-tumour-bearing intestines. Several genes including Tiam1, Gastrin, CD44, uPA, Igfbp4, VEGF and Cox-2 that are known to be transcriptionally regulated by activation of the Wnt signaling pathway were found to be expressed at higher levels in the large intestinal adenomas from K-rasVal12/Cre/ApcMin/+ mice compared with those from controls, although other Wnt signaling pathway target genes remained unchanged. These data show that intestinal expression of K-rasVal12 accelerates Apc-initiated intestinal adenomagenesis in vivo with

  15. Isolation and characterization of temperature-sensitive mutations in the RAS2 and CYR1 genes of Saccharomyces cerevisiae

    SciTech Connect

    Mitsuzawa, Hiroshi; Uno, Isao; Ishikawa, Tatsuo ); Oshima, Takehiro )

    1989-12-01

    The yeast Saccharomyces cerevisiae contains two ras homologues, RAS1 and RAS2, whose products have been shown to modulate the activity of adenylate cyclase encoded by the CYR1 gene. To isolate temperature-sensitive mutations in the RAS2 gene, the authors constructed a plasmid carrying a RAS2 gene whose expression is under the control of the galactose-inducible GAL1 promoter. A ras1 strain transformed with this plasmid was subjected to ethyl methanesfulfonate mutagenesis and nystatin enrichment. Screening of approximately 13,000 mutagenized colonies for galactose-dependent growth at a high temperature (37{degree}) yielded six temperature-sensitive ras2(ras2{sup ts}) mutations and one temperature-sensitive cry1 (cyr1{sup ts}) mutation than can be suppressed by overexpression or increased dosage of RAS2. Some ras2{sup ts} mutations were shown to be suppressed by an extra copy of CYR1. Therefore increased dosage of either RAS2 or CYR1 can suppress the temperature sensitivity caused by a mutation in the other.

  16. Tear analysis in contact lens wearers.

    PubMed Central

    Farris, R L

    1985-01-01

    Tear analysis in contact lens wearers was compared with tear analysis in aphakics without contact lens wear and normal phakic patients. Subjects were divided into five groups: group 1, aphakic without contact lens; group 2, phakic with daily-wear hard contact lens; group 3, phakic with daily-wear soft contact lens; group 4, phakic with extended-wear soft contact lens; and group 5, aphakic with extended-wear soft contact lens. The experimental groups were compared with age- and sex-matched control groups for statistical analysis of tear variables by means of the Student's t-test. The variables measured were tear osmolarity, tear albumin, and lysozyme and lactoferrin concentrations in basal and reflex tears. Highly significant elevations of tear osmolarity were found in aphakic subjects without contact lenses. Less significant differences in tear osmolarity were found in phakic subjects with hard daily-wear lenses or with extended-wear soft lenses. Tear albumin, lysozyme, and lactoferrin in basal and reflex tears were not significantly different in the different groups of contact lens wearers or in the group of aphakic subjects without contact lenses compared with their control groups. Individual variations in tear albumin, lysozyme, and lactoferrin appeared to be responsible for the inability to demonstrate significant differences in tear composition in association with the wearing of different types of contact lenses. Older and aphakic patients demonstrated a tendency to have increased concentrations of proteins in the tears compared with younger, phakic contact lens wearers and normal controls without contact lenses. PMID:3914131

  17. Proposal of RAS-diuretic vs. RAS-calcium antagonist strategies in high-risk hypertension: insight from the 24-hour ambulatory blood pressure profile and central pressure.

    PubMed

    Kario, Kazuomi

    2010-01-01

    I here propose an individualized renin angiotensin system (RAS) inhibitor-based combination therapy with calcium-channel blockers (CCBs) or with diuretics, based on the 24-hr ambulatory blood pressure (BP) profiles and central pressure in relation to the target organ damage in high-risk hypertensive patients. For high-risk patients with increased circulating volume, such as that caused by chronic kidney disease (CKD) or congestive heart failure (CHF), who are likely to exhibit a non-dipper/riser pattern of nocturnal BP fall, diuretics are recommended in combination with a RAS inhibitor to reduce nocturnal BP preferentially. For high-risk patients with arterial diseases such as cardiovascular disease and increased arterial stiffness, who are likely to exhibit exaggerated BP variability, such as morning BP surge and day-to-day BP variability, a CCB is recommended for use in combination with a RAS inhibitor to reduce BP variability and central BP. In particular, bedtime dosing of a RAS inhibitor targeting sleep-early morning activation of RAS may be particularly effective for cardiorenal protection. PMID:20728424

  18. Adjustable Focus Optical Correction Lens (AFOCL)

    NASA Technical Reports Server (NTRS)

    Peters, Bruce R.

    2001-01-01

    This report describes a metrology plan that was developed for the characterization of PLZT-based devices, such as the Adjustable Focus Optical Correction Lens (AFOCL) in support of and as part of the deliverables for NASA contract NAS8-00118. The areas to be investigated include intensiometric effects (those that limit or alter the intensity of the light transmitted through the optic); interferometric effects (the phase change induced through the optic); and polarimetric effects (evaluating the differential lag between two polarization states propagating through the optic). These distinct phenomena are often coupled together in real applications consequently, there is a need to develop different standardized testing apparatus to: (1) isolate one effect from another; (2) gather information for understanding the physical effects; (3) anchor wavefront corrector modeling efforts; (4) develop the ability to decouple different effects; (5) demonstrate the suitability of PLZT technology to perform wavefront correction. The Center for Applied Optics (CAO) at the University of Alabama in Huntsville (UAH) is skilled in the characterization of transmission wavefront shaping devices using traditional interferometers available within the CAO Optical Metrology Laboratory and their Advanced Polarization Test Facility. Besides the imaging and interferometers available, the polarimetry facility has at its disposal, a Mueller Matrix Imaging Polarimeter (MMIP) which is well suited to the characterization of SLMs, polarizers, and thin film coatings within the visible and near-IR spectrums. In addition, the phase-shifting interferometry facilities at NASA-MSFC and the unique interferometers they processes are some of the most advanced available and may be of value especially for performing real-time optical performance evaluation of AFOCL test components.

  19. The elastic constants of the human lens.

    PubMed

    Fisher, R F

    1971-01-01

    1. When the lens is spun around its antero-posterior polar axis in an apparatus designed for the purpose, high speed photography can be used to record its changing profile. By this method a variable radial centrifugal force can be applied to the lens which mimics the pull of the zonule.2. If the lens is not stressed at its centre beyond 100 Nm(-2) it behaves as a truly elastic body. When stressed beyond this limit visco-elastic strain is produced at its poles.3. The human lens has isotropic elastic properties at the extremes of life, but at the other times Young's Modulus of Elasticity varies with the direction in which it is measured.4. Young's Modulus of Elasticity of the lens varies with age, polar elasticity and equatorial elasticity, at birth being 0.75 x 10(3) and 0.85 x 10(3) Nm(-2) respectively, while at 63 years of age both are equal to 3 x 10(3) Nm(-2).5. A comparison of Young's Modulus of the young human lens with that of the rabbit and cat shows that the polar elasticity of the lenses of these animals was 5 times greater in the young rabbit, and 21 times greater in the adult cat. Equatorial elasticities of the rabbit and human lens were equal, while in the cat the equatorial elasticity was four times greater.6. A mathematical model showing the lens substance possessing a nucleus of lower isotropic elasticity than that of the isotropic elastic cortex surrounding it, accounts for the difference between polar and equatorial elasticity of the intact adult lens.7. The implications of these findings are discussed in relation to:(i) accommodation and the rheological properties of the lens;(ii) possible differences in the physical state of the lenticular proteins in the cortex and nucleus which may account for the senile variations in Young's Modulus of Elasticity in these regions of the lens;(iii) the loss of accommodation due solely to an increase in Young's Modulus of Elasticity of the lens between the ages of 15 and 60. This would amount to 44% of the total

  20. Negative refraction makes a perfect lens

    PubMed

    Pendry

    2000-10-30

    With a conventional lens sharpness of the image is always limited by the wavelength of light. An unconventional alternative to a lens, a slab of negative refractive index material, has the power to focus all Fourier components of a 2D image, even those that do not propagate in a radiative manner. Such "superlenses" can be realized in the microwave band with current technology. Our simulations show that a version of the lens operating at the frequency of visible light can be realized in the form of a thin slab of silver. This optical version resolves objects only a few nanometers across.

  1. Negative Refraction Makes a Perfect Lens

    NASA Astrophysics Data System (ADS)

    Pendry, J. B.

    2000-10-01

    With a conventional lens sharpness of the image is always limited by the wavelength of light. An unconventional alternative to a lens, a slab of negative refractive index material, has the power to focus all Fourier components of a 2D image, even those that do not propagate in a radiative manner. Such ``superlenses'' can be realized in the microwave band with current technology. Our simulations show that a version of the lens operating at the frequency of visible light can be realized in the form of a thin slab of silver. This optical version resolves objects only a few nanometers across.

  2. Gravitational lens time delays and gravitational waves

    SciTech Connect

    Frieman, J.A. Department of Astronomy Astrophysics, University of Chicago, Chicago, Illinois 60637 ); Harari, D.D.; Surpi, G.C. )

    1994-10-15

    Using Fermat's principle, we analyze the effects of very long wavelength gravitational waves upon the images of a gravitationally lensed quasar. We show that the lens equation in the presence of gravity waves is equivalent to that of a lens with a different alignment between source, deflector, and observer in the absence of gravity waves. Contrary to a recent claim, we conclude that measurements of time delays in gravitational lenses cannot serve as a method to detect or constrain a stochastic background of gravitational waves of cosmological wavelengths, because the wave-induced time delay is observationally indistinguishable from an intrinsic time delay due to the lens geometry.

  3. Lens-Free Imaging for Biological Applications

    PubMed Central

    Kim, Sang Bok; Bae, Hojae; Koo, Kyo-in; Dokmeci, Mehmet R.; Ozcan, Aydogan; Khademhosseini, Ali

    2013-01-01

    Lens-free (or lensless) imaging is emerging as a cost-effective, compact, and lightweight detection method that can serve numerous biological applications. Lens-free imaging can generate high-resolution images within a field-portable platform, which is ideal for affordable point-of-care devices aiming at resource-limited settings. In this mini-review, we first describe different modes of operation for lens-free imaging and then highlight several recent biological applications of this emerging platform technology. PMID:22357607

  4. The elastic constants of the human lens

    PubMed Central

    Fisher, R. F.

    1971-01-01

    1. When the lens is spun around its antero-posterior polar axis in an apparatus designed for the purpose, high speed photography can be used to record its changing profile. By this method a variable radial centrifugal force can be applied to the lens which mimics the pull of the zonule. 2. If the lens is not stressed at its centre beyond 100 Nm-2 it behaves as a truly elastic body. When stressed beyond this limit visco-elastic strain is produced at its poles. 3. The human lens has isotropic elastic properties at the extremes of life, but at the other times Young's Modulus of Elasticity varies with the direction in which it is measured. 4. Young's Modulus of Elasticity of the lens varies with age, polar elasticity and equatorial elasticity, at birth being 0·75 × 103 and 0·85 × 103 Nm-2 respectively, while at 63 years of age both are equal to 3 × 103 Nm-2. 5. A comparison of Young's Modulus of the young human lens with that of the rabbit and cat shows that the polar elasticity of the lenses of these animals was 5 times greater in the young rabbit, and 21 times greater in the adult cat. Equatorial elasticities of the rabbit and human lens were equal, while in the cat the equatorial elasticity was four times greater. 6. A mathematical model showing the lens substance possessing a nucleus of lower isotropic elasticity than that of the isotropic elastic cortex surrounding it, accounts for the difference between polar and equatorial elasticity of the intact adult lens. 7. The implications of these findings are discussed in relation to: (i) accommodation and the rheological properties of the lens; (ii) possible differences in the physical state of the lenticular proteins in the cortex and nucleus which may account for the senile variations in Young's Modulus of Elasticity in these regions of the lens; (iii) the loss of accommodation due solely to an increase in Young's Modulus of Elasticity of the lens between the ages of 15 and 60. This would amount to 44% of the

  5. Electrowetting based infrared lens using ionic liquids

    NASA Astrophysics Data System (ADS)

    Hu, Xiaodong; Zhang, Shiguo; Liu, Yu; Qu, Chao; Lu, Liujin; Ma, Xiangyuan; Zhang, Xiaoping; Deng, Youquan

    2011-11-01

    We demonstrated an infrared variable focus ionic liquids lens using electrowetting, which could overcome the problems caused by use of water, e.g., evaporation and poor thermostability, while keeping good optical transparency in visible light and near-infrared region. Besides, the type of lens (convex or concave) could be tuned by applied voltage or refractive index of ILs used, and the transmittance was measured to exceed 90% over the spectrum of visible light and near-infrared. We believe this infrared variable focus ionic liquids lens has a great application prospect in both visible light and infrared image systems.

  6. Development of a tantalum pentoxide Luneberg lens

    NASA Technical Reports Server (NTRS)

    Bryan, D. A.; Chubb, C. R.; Powers, J. K.; Reed, W. R.; Dalke, E. A.; Tomaschke, H. E.

    1982-01-01

    A process has been developed for the fabrication of a tantalum pentoxide waveguide Luneburg lens as the input collimator for an optical signal processing circuit on a silicon substrate, such as an integrated wavelength demultiplexer. The development of such a lens involved improvement of the deposition mask profile, reduction of surface scattering by underlaying the lens, reduction of edge scattering by using shims under the mask, and prediction and measurement of the TE-TM polarization aberration. It is shown that polarization aberration significantly affects the design of a demultiplexer system.

  7. Evaluation of the toric implantable collamer lens for simultaneous treatment of myopia and astigmatism.

    PubMed

    Price, Marianne O; Price, Francis W

    2015-01-01

    Myopic astigmatism is a prevalent condition that can be treated with spectacles, contact lenses, or laser refractive surgery. However, these treatment options have functional limitations at higher levels of refractive error. The toric implantable collamer lens is designed to treat a broad range of refractive error, generally up to -18 diopters with +1 to +6 diopters of astigmatism. Approval for a more limited treatment range of up to 15 diopters of myopia with +1 to +4 diopters of astigmatism is being sought in the US, where this device has not yet received marketing approval. Surgical correction of high-myopic astigmatism can be life-altering and allow people to participate in activities that were not previously feasible because of visual limitations. The toric implantable collamer lens is implanted behind the iris and in front of the natural crystalline lens. With earlier lens designs, it was necessary to create an iridectomy or iridotomy to prevent pupillary block. The newest toric implantable collamer lens model has a small central hole that is not visually noticeable. This eliminates the need to create a hole in the iris, thereby enhancing the safety of the procedure.

  8. A gene expression signature associated with “K-Ras addiction” reveals regulators of EMT and tumor cell survival

    PubMed Central

    Singh, Anurag; Greninger, Patricia; Rhodes, Daniel; Koopman, Louise; Violette, Sheila; Bardeesy, Nabeel; Settleman, Jeff

    2009-01-01

    SUMMARY K-Ras mutations occur frequently in epithelial cancers. Using shRNAs to deplete K-Ras in lung and pancreatic cancer cell lines harboring K-Ras mutations, two classes were identified—lines that do or do not require K-Ras to maintain viability. Comparing these two classes of cancer cells revealed a gene expression signature in K-Ras-dependent cells, associated with a well-differentiated epithelial phenotype, which was also seen in primary tumors. Several of these genes encode pharmacologically tractable proteins, such as Syk and Ron kinases and integrin beta6, depletion of which induces epithelial-mesenchymal transformation (EMT) and apoptosis specifically in K-Ras-dependent cells. These findings indicate that epithelial differentiation and tumor cell viability are associated, and that EMT regulators in “K-Ras-addicted” cancers represent candidate therapeutic targets. SIGNIFICANCE K-Ras is the most frequently mutated oncogene in solid tumors and when aberrantly activated, is a potent tumor initiator. However, the identification of the critical effectors of K-Ras-mediated tumorigenesis and the development of clinically effective therapeutic strategies in this setting remain challenging. We have found that cancer cell lines harboring K-Ras mutations can be broadly classified into K-Ras-dependent and K-Ras-independent groups. By establishing a gene expression signature that can distinguish these two groups, we identified genes that are specifically up-regulated in K-Ras-dependent cells and are required for their viability. Therefore, the K-Ras dependency signature has revealed several potential therapeutic targets in a subset of otherwise pharmacologically intractable human cancers. PMID:19477428

  9. Farnesyl transferase inhibitor FTI-277 inhibits breast cell invasion and migration by blocking H-Ras activation

    PubMed Central

    Lee, Kyung Hun; Koh, Minsoo; Moon, Aree

    2016-01-01

    Hyperactive Ras promotes proliferation and malignant phenotypic conversion of cells in cancer. Ras protein must be associated with cellular membranes for its oncogenic activities through post-translational modifications, including farnesylation. Farnesyltransferase (FTase) is essential for H-Ras membrane targeting, and H-Ras, but not N-Ras, has been demonstrated to cause an invasive phenotype in MCF10A breast epithelial cells. In the present study, it was observed that an FTase inhibitor (FTI), FTI-277, blocked epidermal growth factor (EGF)-induced H-Ras activation, but not N-Ras activation in MDA-MB-231 cells, which express wild-type H-Ras and N-Ras. FTI-277 exerted a more potent inhibitory effect on the proliferation of H-Ras-MCF10A cells and Hs578T breast cancer cells expressing an active mutant of H-Ras than that of MDA-MB-231 cells. The invasive/migratory phenotypes of the H-Ras-MCF10A and Hs578T cells were effectively inhibited by FTI-277 treatment. By contrast, FTI-277 did not affect the invasive/migratory phenotypes of MDA-MB-231 cells. However, the EGF-induced invasion of MDA-MB-231 cells was decreased by FTI-277, implicating that FTI-277 inhibits breast cell invasion and migration by blocking H-Ras activation. Taken together, the results of the present study suggest that FTase inhibition by FTI-277 may be an effective strategy for targeting H-Ras-mediated proliferation, migration and invasion of breast cells. PMID:27602167

  10. Expression of a dominant-negative Ras mutant does not affect stimulation of glucose uptake and glycogen synthesis by insulin.

    PubMed

    Dorrestijn, J; Ouwens, D M; Van den Berghe, N; Bos, J L; Maassen, J A

    1996-05-01

    It has previously been shown that insulin-induced stimulation of glucose uptake and glycogen synthesis requires activation of phosphatidylinositol-3-kinase (PI3kinase). Insulin also induces formation of RasGTP in cells and various studies have yielded inconsistent data with respect to the contribution of signalling pathways activated by RasGTP, to insulin-stimulated glucose uptake and glycogen synthesis. We have examined the requirement of RasGTP-mediated signalling for these insulin responses by expression of a dominant negative mutant of Ras (RasN17) in cells by vaccinia virus mediated gene transfer. This Ras-mutant abrogates the signalling pathways mediated by endogenous RasGTP. Subsequently, the ability of insulin to stimulate 2-deoxyglucose uptake and glycogen was examined. We observed that expression of RasN17 in 3T3L1 adipocytes did not affect the stimulation of hexose uptake by insulin. Similarly, expression of RasN17 in A14 cells, an NIH 3T3-derived cell line with high expression of insulin receptors, did not affect insulin-induced stimulation of glycogen synthesis. In both cell lines, insulin-induced phosphorylation of Mapkinase (Erk1,2) was abrogated after expression of RasN17, demonstrating the functional interference by RasN17 with signalling mediated by endogenous RasGTP. Wortmannin, an inhibitor of PI3kinase, abolished dose-dependently the insulin-induced stimulation of hexose uptake and glycogen synthesis without an effect on RasGTP levels in both cell types. We conclude that stimulation of glucose transport and glycogen synthesis by insulin occurs independently of RasGTP-mediated signalling.

  11. Requirement of the NF-κB Subunit p65/RelA for K-Ras-Induced Lung Tumorigenesis

    PubMed Central

    Basseres, Daniela S.; Ebbs, Aaron; Levantini, Elena; Baldwin, Albert S.

    2010-01-01

    K-Ras-induced lung cancer is a very common disease, for which there are currently no effective therapies. Because therapy directly targeting the activity of oncogenic Ras has been unsuccessful, a different approach for novel therapy design is to identify critical Ras downstream oncogenic targets. Given that oncogenic Ras proteins activate the transcription factor NF-κB, and the importance of NF-κB in oncogenesis, we hypothesized that NF-κB would be an important K-Ras target in lung cancer. To address this hypothesis, we generated an NF-κB-EGFP reporter mouse model of K-Ras-induced lung cancer and determined that K-Ras activates NF-κB in lung tumors in situ. Furthermore, a mouse model was generated where activation of oncogenic K-Ras in lung cells was coupled with inactivation of the NF-κB subunit p65/RelA. In this model, deletion of p65/RelA reduces the number of K-Ras-induced lung tumors both in the presence and absence of the tumor suppressor p53. Lung tumors with loss of p65/RelA have higher numbers of apoptotic cells, reduced spread and lower grade. Using lung cell lines expressing oncogenic K-Ras, we show that NF-κB is activated in these cells in a K-Ras-dependent manner and that NF-κB activation by K-Ras requires IKKβ kinase activity. Taken together, these results demonstrate the importance of the NF-κB subunit p65/RelA in K-Ras induced lung transformation and identify IKKβ as a potential therapeutic target for K-Ras-induced lung cancer. PMID:20406971

  12. Farnesyl transferase inhibitor FTI-277 inhibits breast cell invasion and migration by blocking H-Ras activation

    PubMed Central

    Lee, Kyung Hun; Koh, Minsoo; Moon, Aree

    2016-01-01

    Hyperactive Ras promotes proliferation and malignant phenotypic conversion of cells in cancer. Ras protein must be associated with cellular membranes for its oncogenic activities through post-translational modifications, including farnesylation. Farnesyltransferase (FTase) is essential for H-Ras membrane targeting, and H-Ras, but not N-Ras, has been demonstrated to cause an invasive phenotype in MCF10A breast epithelial cells. In the present study, it was observed that an FTase inhibitor (FTI), FTI-277, blocked epidermal growth factor (EGF)-induced H-Ras activation, but not N-Ras activation in MDA-MB-231 cells, which express wild-type H-Ras and N-Ras. FTI-277 exerted a more potent inhibitory effect on the proliferation of H-Ras-MCF10A cells and Hs578T breast cancer cells expressing an active mutant of H-Ras than that of MDA-MB-231 cells. The invasive/migratory phenotypes of the H-Ras-MCF10A and Hs578T cells were effectively inhibited by FTI-277 treatment. By contrast, FTI-277 did not affect the invasive/migratory phenotypes of MDA-MB-231 cells. However, the EGF-induced invasion of MDA-MB-231 cells was decreased by FTI-277, implicating that FTI-277 inhibits breast cell invasion and migration by blocking H-Ras activation. Taken together, the results of the present study suggest that FTase inhibition by FTI-277 may be an effective strategy for targeting H-Ras-mediated proliferation, migration and invasion of breast cells.

  13. NMR characterization of full-length farnesylated and non-farnesylated H-Ras and its implications for Raf activation.

    PubMed

    Thapar, Roopa; Williams, Jason G; Campbell, Sharon L

    2004-11-01

    The C terminus, also known as the hypervariable region (residues 166-189), of H-, N-, and K-Ras proteins has sequence determinants necessary for full activation of downstream effectors such as Raf kinase and PI-3 kinase as well as for the correct targeting of Ras proteins to lipid rafts and non-raft membranes. There is considerable interest in understanding how residues in the extreme C terminus of the different Ras proteins and farnesylation of the CaaX box cysteine affect Ras membrane localization and allosteric activation of Raf kinase. To provide insights into the structural and dynamic changes that occur in Ras upon farnesylation, we have used NMR spectroscopy to compare the properties of truncated H-Ras (1-166), to non-processed full-length H-Ras (residues 1-185) and full-length (1-189) farnesylated H-Ras. We report that the C-terminal helix alpha-5 extends to residue N172, and the remaining 17 amino acid residues in the C terminus are conformationally averaged in solution. Removal of either 23 or 18 amino acid residues from the C terminus of full length H-Ras generates truncated H-Ras (1-166) and H-Ras (1-171) proteins, respectively, that have been structurally characterized and are biochemical active. Here we report that C-terminal truncation of H-Ras results in minor structural and dynamic perturbations that are propagated throughout the H-Ras protein including increased flexibility of the central beta-sheet and the C-terminal helix alpha-5. Ordering of residues in loop-2, which is involved in Raf CRD binding is also observed. Farnesylation of full-length H-Ras at C186 does not result in detectable conformational changes in H-Ras. Chemical shift mapping studies of farnesylated and non-farnesylated forms of H-Ras with the Raf-CRD show that the farnesyl moiety, the extreme H-Ras C terminus and residues 23-30, contribute to H-Ras:Raf-CRD interactions, thereby increasing the affinity of H-Ras for the Raf-CRD.

  14. Design, Evaluation and GCM-Performance of a New Parameterization for Microphysics of Clouds with Relaxed Arakawa-Schubert Scheme (McRas)

    NASA Technical Reports Server (NTRS)

    Sud, Y. C.; Walker, G. K.

    1998-01-01

    altered by the choice of time constant and incloud critical cloud water amount regulators for auto-conversion. The CRF and its feedbacks also have a profound effect on the ITCZ. Even though somewhat weaker than observed, the McRAS-GCM simulation produces robust 30-60 day oscillations in the 200 hPa velocity potential. Two ensembles of 4-summer (July, August, September) simulations, one each for 1987 and 1988 show that the McRAS-GCM simulates realistic and statistically significant precipitation differences over India, Central America, and tropical Africa. Several seasonal simulations were performed with McRAS-GEOS II GCM for the summer (June-July- August) and winter (December-January-February) periods to determine how the simulated clouds and CRFs would be affected by: i) advection of clouds; ii) cloud top entrainment instability, iii) cloud water inhomogeneity correction, and (iv) cloud production and dissipation in different cloud-processes. The results show that each of these processes contributes to the simulated cloud-fraction and CRF.

  15. 21 CFR 886.1390 - Flexible diagnostic Fresnel lens.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Flexible diagnostic Fresnel lens. 886.1390 Section... (CONTINUED) MEDICAL DEVICES OPHTHALMIC DEVICES Diagnostic Devices § 886.1390 Flexible diagnostic Fresnel lens. (a) Identification. A flexible diagnostic Fresnel lens is a device that is a very thin lens which...

  16. 21 CFR 886.1390 - Flexible diagnostic Fresnel lens.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Flexible diagnostic Fresnel lens. 886.1390 Section... (CONTINUED) MEDICAL DEVICES OPHTHALMIC DEVICES Diagnostic Devices § 886.1390 Flexible diagnostic Fresnel lens. (a) Identification. A flexible diagnostic Fresnel lens is a device that is a very thin lens which...

  17. Coculture with intraocular lens material-activated macrophages induces an inflammatory phenotype in lens epithelial cells.

    PubMed

    Pintwala, Robert; Postnikoff, Cameron; Molladavoodi, Sara; Gorbet, Maud

    2015-03-01

    Cataracts are the leading cause of blindness worldwide, requiring surgical implantation of an intraocular lens. Despite evidence of leukocyte ingress into the postoperative lens, few studies have investigated the leukocyte response to intraocular lens materials. A novel coculture model was developed to examine macrophage activation by hydrophilic acrylic (poly(2-hydroxyethyl methacrylate)) and hydrophobic acrylic (polymethylmethacrylate) commercial intraocular lens. The human monocytic cell line THP-1 was differentiated into macrophages and cocultured with human lens epithelial cell line (HLE-B3) with or without an intraocular lens for one, two, four, or six days. Using flow cytometry and confocal microscopy, expression of the macrophage activation marker CD54 (intercellular adhesion molecule-1) and production of reactive oxygen species via the fluorogenic probe 2',7'-dichlorodihydrofluorescein diacetate were examined in macrophages. α-Smooth muscle actin, a transdifferentiation marker, was characterized in lens epithelial cells. The poly(2-hydroxyethyl methacrylate) intraocular lens prevented adhesion but induced significant macrophage activation (p < 0.03) versus control (no intraocular lens), while the polymethylmethacrylate intraocular lens enabled adhesion and multinucleated fusion, but induced no significant activation. Coculture with either intraocular lens increased reactive oxygen species production in macrophages after one day (p < 0.03) and increased expression of α-smooth muscle actin in HLE B-3 after six days, although only poly(2-hydroxyethyl methacrylate) induced a significant difference versus control (p < 0.01). Our results imply that-contrary to prior uveal biocompatibility understanding-macrophage adherence is not necessary for a strong inflammatory response to an intraocular lens, with hydrophilic surfaces inducing higher activation than hydrophobic surfaces. These findings provide a new method of inquiry into uveal

  18. Coculture with intraocular lens material-activated macrophages induces an inflammatory phenotype in lens epithelial cells.

    PubMed

    Pintwala, Robert; Postnikoff, Cameron; Molladavoodi, Sara; Gorbet, Maud

    2015-03-01

    Cataracts are the leading cause of blindness worldwide, requiring surgical implantation of an intraocular lens. Despite evidence of leukocyte ingress into the postoperative lens, few studies have investigated the leukocyte response to intraocular lens materials. A novel coculture model was developed to examine macrophage activation by hydrophilic acrylic (poly(2-hydroxyethyl methacrylate)) and hydrophobic acrylic (polymethylmethacrylate) commercial intraocular lens. The human monocytic cell line THP-1 was differentiated into macrophages and cocultured with human lens epithelial cell line (HLE-B3) with or without an intraocular lens for one, two, four, or six days. Using flow cytometry and confocal microscopy, expression of the macrophage activation marker CD54 (intercellular adhesion molecule-1) and production of reactive oxygen species via the fluorogenic probe 2',7'-dichlorodihydrofluorescein diacetate were examined in macrophages. α-Smooth muscle actin, a transdifferentiation marker, was characterized in lens epithelial cells. The poly(2-hydroxyethyl methacrylate) intraocular lens prevented adhesion but induced significant macrophage activation (p < 0.03) versus control (no intraocular lens), while the polymethylmethacrylate intraocular lens enabled adhesion and multinucleated fusion, but induced no significant activation. Coculture with either intraocular lens increased reactive oxygen species production in macrophages after one day (p < 0.03) and increased expression of α-smooth muscle actin in HLE B-3 after six days, although only poly(2-hydroxyethyl methacrylate) induced a significant difference versus control (p < 0.01). Our results imply that-contrary to prior uveal biocompatibility understanding-macrophage adherence is not necessary for a strong inflammatory response to an intraocular lens, with hydrophilic surfaces inducing higher activation than hydrophobic surfaces. These findings provide a new method of inquiry into uveal

  19. Luneburg and flat lens based on graded photonic crystal

    NASA Astrophysics Data System (ADS)

    Liu, Wei; Sun, Xiaohong; Gao, Minglei; Wang, Shuai

    2016-04-01

    Square-lattice graded photonic crystals employed for designing Luneburg and Flat Lens is presented. Comparable simulation of the Luneburg lens with TE and TM polarizations predicts that TM lens possesses of enlarged transmission bandwidth and strengthened focusing ability, in comparison with TE lens. As a typical simplified counterpart, the evolution of focusing intensity and numerical aperture of the flat lens is achieved. What is more, those Luneburg and Flat Lens can withstand imperfect gradients in structure design. This will provide a guidance to produce a high quality focusing lens with small size, short focal length and large numerical aperture applied in the integrated photonic devices.

  20. Alteration of EGFR Spatiotemporal Dynamics Suppresses Signal Transduction

    PubMed Central

    Turk, Harmony F.; Barhoumi, Rola; Chapkin, Robert S.

    2012-01-01

    The epidermal growth factor receptor (EGFR), which regulates cell growth and survival, is integral to colon tumorigenesis. Lipid rafts play a role in regulating EGFR signaling, and docosahexaenoic acid (DHA) is known to perturb membrane domain organization through changes in lipid rafts. Therefore, we investigated the mechanistic link between EGFR function and DHA. Membrane incorporation of DHA into immortalized colonocytes altered the lateral organization of EGFR. DHA additionally increased EGFR phosphorylation but paradoxically suppressed downstream signaling. Assessment of the EGFR-Ras-ERK1/2 signaling cascade identified Ras GTP binding as the locus of the DHA-induced disruption of signal transduction. DHA also antagonized EGFR signaling capacity by increasing receptor internalization and degradation. DHA suppressed cell proliferation in an EGFR-dependent manner, but cell proliferation could be partially rescued by expression of constitutively active Ras. Feeding chronically-inflamed, carcinogen-injected C57BL/6 mice a fish oil containing diet enriched in DHA recapitulated the effects on the EGFR signaling axis observed in cell culture and additionally suppressed tumor formation. We conclude that DHA-induced alteration in both the lateral and subcellular localization of EGFR culminates in the suppression of EGFR downstream signal transduction, which has implications for the molecular basis of colon cancer prevention by DHA. PMID:22761867

  1. Bilateral lens luxation and intracapsular lens extractions in a Matshchie's tree kangaroo.

    PubMed

    McLean, Nancy Johnstone; Zimmerman, Ralph

    2015-01-01

    An adult, female, captive, Matshchie's tree kangaroo was diagnosed with an anterior lens luxation in the right eye and a lens subluxation in the left eye. Both eyes were treated surgically with intracapsular lens extractions. A 360° rhegmatogenous retinal detachment was diagnosed 6 months postoperatively in the left eye. Aphakic vision was maintained in the right eye 9 months postoperatively. Based on family history and the lack of antecedent ocular disease, the lens luxations were presumed to be inherited and veterinarians should be aware of this condition within the captive tree kangaroo population.

  2. Bilateral lens luxation and intracapsular lens extractions in a Matshchie's tree kangaroo.

    PubMed

    McLean, Nancy Johnstone; Zimmerman, Ralph

    2015-01-01

    An adult, female, captive, Matshchie's tree kangaroo was diagnosed with an anterior lens luxation in the right eye and a lens subluxation in the left eye. Both eyes were treated surgically with intracapsular lens extractions. A 360° rhegmatogenous retinal detachment was diagnosed 6 months postoperatively in the left eye. Aphakic vision was maintained in the right eye 9 months postoperatively. Based on family history and the lack of antecedent ocular disease, the lens luxations were presumed to be inherited and veterinarians should be aware of this condition within the captive tree kangaroo population. PMID:25135107

  3. RISK FACTORS FOR CONTACT LENS INDUCED PAPILLARY CONJUNCTIVITIS ASSOCIATED WITH SILICONE HYDROGEL CONTACT LENS WEAR

    PubMed Central

    Tagliaferri, Angela; Love, Thomas E.; Szczotka-Flynn, Loretta

    2014-01-01

    BACKGROUND Contact lens induced papillary conjunctivitis (CLPC) continues to be a major cause of dropout during contact lens extended wear. This retrospective study explores risk factors for the development of CLPC during silicone hydrogel lens extended wear. METHODS Data from 205 subjects enrolled in the Longitudinal Analysis of Silicone Hydrogel Contact Lens (LASH) study wearing lotrafilcon A silicone hydrogel lenses for up to 30 days of continuous wear were used to determine risk factors for CLPC in this secondary analysis of the main cohort. The main covariates of interest included substantial lens-associated bacterial bioburden, and topographically determined lens base curve-to-cornea fitting relationships. Additional covariates of interest included history of prior adverse events, time of year, race, education level, gender and other subject demographics. Statistical analyses included univariate logistic regression to assess the impact of potential risk factors on the binary CLPC outcome, and Cox proportional hazards regression to describe the impact of those factors on time-to-CLPC diagnosis. RESULTS Across 12 months of follow-up, 52 subjects (25%) experienced CLPC. No associations were found between CLPC development and the presence of bacterial bioburden, lens-to-cornea fitting relationships, history of prior adverse events, gender or race. CLPC development followed the same seasonal trends as the local peaks in environmental allergans. CONCLUSIONS Lens fit and biodeposits, in the form of lens associated bacterial bioburden, were not associated with the development of CLPC during extended wear with lotrafilcon A silicone hydrogel lenses. PMID:24681609

  4. Restoring lens capsule integrity enhances lens regeneration in New Zealand albino rabbits and cats.

    PubMed

    Gwon, A; Gruber, L J; Mantras, C

    1993-11-01

    In studies conducted by numerous investigators for 150 years, lenses regenerated following endocapsular lens extraction in New Zealand albino rabbits have been irregular in shape, appearing primarily doughnut-shaped as a result of lack of lens growth at the site of the anterior capsulotomy and its adhesion to the posterior capsule. In the present study, we restored the lens capsule integrity by inserting a collagen patch at the time of surgery to seal the anterior capsulotomy and to improve the shape and structure of the regenerated lenses. We then filled the capsule bag with air to prevent adhesions between the anterior and posterior capsule and maintain capsule tautness and shape. Lens regeneration was first noted as early as one to two weeks. Regenerated lens filled approximately 50% of the capsule bag at two weeks and 100% by five weeks. Subsequent growth was in the anterior-posterior direction and measured by A-scan biometry. Lens thickness increased by 0.3 mm per month. The regenerated lenses were spherical with normal cortical structure and a nuclear opacity. In conclusion, restoration of lens capsular integrity with a collagen patch following endocapsular lens extraction enhanced the shape, structure, and growth rate of the regenerated lenses. In addition, lens regeneration was shown to occur in two cats.

  5. Membrane potential modulates plasma membrane phospholipid dynamics and K-Ras signaling

    PubMed Central

    Zhou, Yong; Wong, Ching-On; Cho, Kwang-jin; van der Hoeven, Dharini; Liang, Hong; Thakur, Dhananiay P.; Luo, Jialie; Babic, Milos; Zinsmaier, Konrad E.; Zhu, Michael X.; Hu, Hongzhen; Venkatachalam, Kartik; Hancock, John F.

    2015-01-01

    Plasma membrane depolarization can trigger cell proliferation, but how membrane potential influences mitogenic signaling is uncertain. Here, we show that plasma membrane depolarization induces nanoscale reorganization of phosphatidylserine and phosphatidylinositol 4,5-bisphosphate but not other anionic phospholipids. K-Ras, which is targeted to the plasma membrane by electrostatic interactions with phosphatidylserine, in turn undergoes enhanced nanoclustering. Depolarization-induced changes in phosphatidylserine and K-Ras plasma membrane organization occur in fibroblasts, excitable neuroblastoma cells, and Drosophila neurons in vivo and robustly amplify K-Ras–dependent mitogen-activated protein kinase (MAPK) signaling. Conversely, plasma membrane repolarization disrupts K-Ras nanoclustering and inhibits MAPK signaling. By responding to voltage-induced changes in phosphatidylserine spatiotemporal dynamics, K-Ras nanoclusters set up the plasma membrane as a biological field-effect transistor, allowing membrane potential to control the gain in mitogenic signaling circuits. PMID:26293964

  6. A Plasma Lens for Magnetron Sputtering

    SciTech Connect

    Anders, Andre; Brown, Jeff

    2010-11-30

    A plasma lens, consisting of a solenoid and potential-defining ring electrodes, has been placed between a magnetron and substrates to be coated. Photography reveals qualitative information on excitation, ionization, and the transport of plasma to the substrate.

  7. Perfect lens makes a perfect trap.

    PubMed

    Lu, Zhaolin; Murakowski, Janusz; Schuetz, Christopher A; Shi, Shouyuan; Schneider, Garrett J; Samluk, Jesse P; Prather, Dennis W

    2006-03-20

    In this work, we present for the first time a new and realistic application of the "perfect lens", namely, electromagnetic traps (or tweezers). We combined two recently developed techniques, 3D negative refraction flat lenses (3DNRFLs) and optical tweezers, and experimentally demonstrated the very unique advantages of using 3DNRFLs for electromagnetic traps. Super-resolution and short focal distance of the flat lens result in a highly focused and strongly convergent beam, which is a key requirement for a stable and accurate electromagnetic trap. The translation symmetry of 3DNRFL provides translation-invariance for imaging, which allows an electromagnetic trap to be translated without moving the lens, and permits a trap array by using multiple sources with a single lens. Electromagnetic trapping was demonstrated using polystyrene particles in suspension, and subsequent to being trapped to a single point, they were then accurately manipulated over a large distance by simple movement of a 3DNRFL-imaged microwave monopole source.

  8. Methane Detector With Plastic Fresnel Lens

    NASA Technical Reports Server (NTRS)

    Grant, W. B.

    1986-01-01

    Laser detector for natural gas leaks modified by substitution of molded plastic lens for spherical mirror. By measuring relative attenuation at two wavelengths, detector used to check for methane escaping from pipelines above or below ground and from landfill.

  9. Optical switch based on electrowetting liquid lens

    NASA Astrophysics Data System (ADS)

    Li, Lei; Liu, Chao; Peng, Hua-Rong; Wang, Qiong-Hua

    2012-05-01

    In this paper, we propose an optical switch based on an electrowetting liquid lens. The device consists of an electrowetting liquid lens and a non-transparent cap with a pin hole. When the lens is actuated to be positive, the incident light can be converged on the pin hole and pass through the hole with less attenuation. When the lens is deformed to be negative, the incident light is diverged and most of light is blocked by the cap. Our results show that the system can provide high contrast ratio (˜800:1) and reasonable response time (˜88 ms). The proposed optical switch has potential application in light shutters, variable optical attenuators, and adaptive irises.

  10. Avoiding liability in contact lens practice.

    PubMed

    Classé, J G

    1994-01-01

    Contact lens practice is the leading source of liability claims involving optometrists. Claims most often allege negligence, although informed consent or product liability may also be causes of legal claims for damages. For daily wear patients, the most likely causes of claims are nonapproved use of lenses or solutions, monovision contact lenses, negligence by a contact lens technician, failure to verify lenses before dispensing, mismanagement of contact lens-related corneal abrasions, and inadequate monitoring of ocular health. For extended-wear patients, the most common sources of liability claims are inappropriate patient selection, inadequate training of patients, improper wearing schedules, improper management of contact lens-related corneal complications, and inadequate monitoring of ocular health.

  11. 23-gauge vitrectomy for retained lens material.

    PubMed

    Bhandari, Ramanath; Ernst, Benjamin J; Stafeeva, Ksenia; Mandava, Naresh; Quiroz-Mercado, Hugo

    2012-07-01

    A technique for removal of retained lens material is described with a three-port 23-gauge vitrectomy system. Removal of the core vitreous is first performed, followed by removal of the cortical vitreous. All vitreous adhesions to the lens are cleared. The cut rate is then decreased to 1,500 cuts per minute, and vacuum increased to 600 mm Hg. The cortical lens material is cleared first, and then the nuclear material is taken with the same vitrectomy probe using the light pipe to assist in crushing the nuclear fragments. With this technique, even large dense nuclear and cortical retained lens material can be removed from the vitreous chamber without the need for a fragmatome. PMID:22692723

  12. Luneburg lens and optical matrix algebra research

    NASA Technical Reports Server (NTRS)

    Wood, V. E.; Busch, J. R.; Verber, C. M.; Caulfield, H. J.

    1984-01-01

    Planar, as opposed to channelized, integrated optical circuits (IOCs) were stressed as the basis for computational devices. Both fully-parallel and systolic architectures are considered and the tradeoffs between the two device types are discussed. The Kalman filter approach is a most important computational method for many NASA problems. This approach to deriving a best-fit estimate for the state vector describing a large system leads to matrix sizes which are beyond the predicted capacities of planar IOCs. This problem is overcome by matrix partitioning, and several architectures for accomplishing this are described. The Luneburg lens work has involved development of lens design techniques, design of mask arrangements for producing lenses of desired shape, investigation of optical and chemical properties of arsenic trisulfide films, deposition of lenses both by thermal evaporation and by RF sputtering, optical testing of these lenses, modification of lens properties through ultraviolet irradiation, and comparison of measured lens properties with those expected from ray trace analyses.

  13. Gravitational Lens B0218+357

    NASA Video Gallery

    This movie illustrates the components of a gravitational lens system (a kind of natural telescope formed when a rare cosmic alignment allows the gravity of a massive object to bend and amplify ligh...

  14. Ras-induced epigenetic inactivation of the RRAD (Ras-related associated with diabetes) gene promotes glucose uptake in a human ovarian cancer model.

    PubMed

    Wang, Yan; Li, Guiling; Mao, Fengbiao; Li, Xianfeng; Liu, Qi; Chen, Lin; Lv, Lu; Wang, Xin; Wu, Jinyu; Dai, Wei; Wang, Guan; Zhao, Enfeng; Tang, Kai-Fu; Sun, Zhong Sheng

    2014-05-16

    RRAD (Ras-related associated with diabetes) is a small Ras-related GTPase that is frequently inactivated by DNA methylation of the CpG island in its promoter region in cancer tissues. However, the role of the methylation-induced RRAD inactivation in tumorigenesis remains unclear. In this study, the Ras-regulated transcriptome and epigenome were profiled by comparing T29H (a Ras(V12)-transformed human ovarian epithelial cell line) with T29 (an immortalized but non-transformed cell line) through reduced representation bisulfite sequencing and digital gene expression. We found that Ras(V12)-mediated oncogenic transformation was accompanied by RRAD promoter hypermethylation and a concomitant loss of RRAD expression. In addition, we found that the RRAD promoter was hypermethylated, and its transcription was reduced in ovarian cancer versus normal ovarian tissues. Treatment with the DNA methyltransferase inhibitor 5-aza-2'-deoxycytidine resulted in demethylation in the RRAD promoter and restored RRAD expression in T29H cells. Additionally, treatment with farnesyltransferase inhibitor FTI277 resulted in restored RRAD expression and inhibited DNA methytransferase expression and activity in T29H cells. By employing knockdown and overexpression techniques in T29 and T29H, respectively, we found that RRAD inhibited glucose uptake and lactate production by repressing the expression of glucose transporters. Finally, RRAD overexpression in T29H cells inhibited tumor formation in nude mice, suggesting that RRAD is a tumor suppressor gene. Our results indicate that Ras(V12)-mediated oncogenic transformation induces RRAD epigenetic inactivation, which in turn promotes glucose uptake and may contribute to ovarian cancer tumorigenesis.

  15. Ras-induced epigenetic inactivation of the RRAD (Ras-related associ