Science.gov

Sample records for pc mutant caused

  1. Polycystic Kidney Disease in the Medaka (Oryzias latipes) pc Mutant Caused by a Mutation in the Gli-Similar3 (glis3) Gene

    PubMed Central

    Hashimoto, Hisashi; Miyamoto, Rieko; Watanabe, Naoki; Shiba, Dai; Ozato, Kenjiro; Inoue, Chikako; Kubo, Yuko; Koga, Akihiko; Jindo, Tomoko; Narita, Takanori; Naruse, Kiyoshi; Ohishi, Kazuko; Nogata, Keiko; Shin-I, Tadasu; Asakawa, Shuichi; Shimizu, Nobuyoshi; Miyamoto, Tomotsune; Mochizuki, Toshio; Yokoyama, Takahiko; Hori, Hiroshi; Takeda, Hiroyuki; Kohara, Yuji; Wakamatsu, Yuko

    2009-01-01

    Polycystic kidney disease (PKD) is a common hereditary disease in humans. Recent studies have shown an increasing number of ciliary genes that are involved in the pathogenesis of PKD. In this study, the Gli-similar3 (glis3) gene was identified as the causal gene of the medaka pc mutant, a model of PKD. In the pc mutant, a transposon was found to be inserted into the fourth intron of the pc/glis3 gene, causing aberrant splicing of the pc/glis3 mRNA and thus a putatively truncated protein with a defective zinc finger domain. pc/glis3 mRNA is expressed in the epithelial cells of the renal tubules and ducts of the pronephros and mesonephros, and also in the pancreas. Antisense oligonucleotide-mediated knockdown of pc/glis3 resulted in cyst formation in the pronephric tubules of medaka fry. Although three other glis family members, glis1a, glis1b and glis2, were found in the medaka genome, none were expressed in the embryonic or larval kidney. In the pc mutant, the urine flow rate in the pronephros was significantly reduced, which was considered to be a direct cause of renal cyst formation. The cilia on the surface of the renal tubular epithelium were significantly shorter in the pc mutant than in wild-type, suggesting that shortened cilia resulted in a decrease in driving force and, in turn, a reduction in urine flow rate. Most importantly, EGFP-tagged pc/glis3 protein localized in primary cilia as well as in the nucleus when expressed in mouse renal epithelial cells, indicating a strong connection between pc/glis3 and ciliary function. Unlike human patients with GLIS3 mutations, the medaka pc mutant shows none of the symptoms of a pancreatic phenotype, such as impaired insulin expression and/or diabetes, suggesting that the pc mutant may be suitable for use as a kidney-specific model for human GLIS3 patients. PMID:19609364

  2. Mutants of PC12 cells with altered cyclic AMP responses

    SciTech Connect

    Block, T.; Kon, C.; Breckenridge, B.M.

    1984-10-01

    PCl2 cells, derived from a rat pheochromocytoma, were mutagenized and selected in media containing agents known to elevate intracellular concentrations of cyclic AMP (cAMP). More than 40 clones were isolated by selection with cholera toxin or 2-chloroadenosine or both. The variants that were deficient in accumulating cAMP were obtained by using a protocol in which 1 ..mu..m 8-bromo-cAMP was included in addition to the agonist. Certain of these variants were partially characterized with respect to the site of altered cAMP metabolism. The profiles of adenylate cyclase activity responsiveness of certain variants to guanosine-5'-(BETA,..gamma..-imido) triphosphate and to forskolin resembled those of UNC and cyc phenotypes of S49 lymphoma cells, which are functionally deficient in the GTP-sensitive coupling protein, N/sub s/. Other variants were characterized by increased cyclic nucleotide phosphodiesterase activity at low substrate concentration. Diverse morphological traits were observed among the variants, but it was not possible to assign them to a particular cAMP phenotype. Two revertants of a PCl2 mutant were isolated and observed to have regained a cellular cAMP response to 2-chloroadenosine and to forskolin. It is hoped that these PCl2 mutants will have utility for defining cAMP-mediated functions, including any links to the action of nerve growth factor, in cells derived from the neural crest.

  3. Neferine attenuates the protein level and toxicity of mutant huntingtin in PC-12 cells via induction of autophagy.

    PubMed

    Wong, Vincent Kam Wai; Wu, An Guo; Wang, Jing Rong; Liu, Liang; Law, Betty Yuen-Kwan

    2015-02-18

    Mutant huntingtin aggregation is highly associated with the pathogenesis of Huntington's disease, an adult-onset autosomal dominant disorder, which leads to a loss of motor control and decline in cognitive function. Recent literature has revealed the protective role of autophagy in neurodegenerative diseases through degradation of mutant toxic proteins, including huntingtin or a-synuclein. Through the GFP-LC3 autophagy detection platform, we have  identified  neferine,  isolated  from  the  lotus  seed  embryo  of Nelumbo nucifera, which is able to induce autophagy through an AMPK-mTOR-dependent pathway. Furthermore, by overexpressing huntingtin with 74 CAG repeats (EGFP-HTT 74) in PC-12 cells, neferine reduces both the protein level and toxicity of mutant huntingtin through an autophagy-related gene 7 (Atg7)-dependent mechanism. With the variety of novel active compounds present in medicinal herbs, our current study suggests the possible protective mechanism of an autophagy inducer isolated from Chinese herbal medicine, which is crucial for its further development into a potential therapeutic agent for neurodegenerative disorders in the future.

  4. Process optimisation for the biosynthesis of cellulase by Bacillus PC-BC6 and its mutant derivative Bacillus N3 using submerged fermentation.

    PubMed

    Abdullah, Roheena; Zafar, Wajeeha; Nadeem, Muhammad; Iqtedar, Mehwish; Naz, Shagufta; Syed, Quratulain; Kaleem, Afshan

    2015-01-01

    This study deals with optimisation of cultural conditions for enhanced production of cellulase by Bacillus PC-BC6 and its mutant derivative Bacillus N3. Influence of different variables including incubation time, temperature, inoculum size, pH, nitrogen sources and metal ions has been studied. The optimum conditions for cellulase production were incubation period of 72 h, inoculum size 4% incubation temperature 37°C, pH 7, 0.25% ammonium sulphate, 0.2% peptone as inorganic and organic nitrogen source in case of Bacillus PC-BC6. In case of mutant Bacillus N3, optimal conditions were incubation period of 48 h, incubation temperature 37°C, inoculum size 3%, pH 7, 0.2% ammonium chloride and 0.15% yeast extract. Presence of MnSO4 and CaCl2 enhances the enzyme production by Bacillus PC-BC6 and mutant Bacillus N3, respectively. This study was innovative and successful in producing cellulase economically by using cheap indigenous substrate Saccharum spontaneum.

  5. Pyrin gene and mutants thereof, which cause familial Mediterranean fever

    DOEpatents

    Kastner, Daniel L.; Aksentijevichh, Ivona; Centola, Michael; Deng, Zuoming; Sood, Ramen; Collins, Francis S.; Blake, Trevor; Liu, P. Paul; Fischel-Ghodsian, Nathan; Gumucio, Deborah L.; Richards, Robert I.; Ricke, Darrell O.; Doggett, Norman A.; Pras, Mordechai

    2003-09-30

    The invention provides the nucleic acid sequence encoding the protein associated with familial Mediterranean fever (FMF). The cDNA sequence is designated as MEFV. The invention is also directed towards fragments of the DNA sequence, as well as the corresponding sequence for the RNA transcript and fragments thereof. Another aspect of the invention provides the amino acid sequence for a protein (pyrin) associated with FMF. The invention is directed towards both the full length amino acid sequence, fusion proteins containing the amino acid sequence and fragments thereof. The invention is also directed towards mutants of the nucleic acid and amino acid sequences associated with FMF. In particular, the invention discloses three missense mutations, clustered in within about 40 to 50 amino acids, in the highly conserved rfp (B30.2) domain at the C-terminal of the protein. These mutants include M6801, M694V, K695R, and V726A. Additionally, the invention includes methods for diagnosing a patient at risk for having FMF and kits therefor.

  6. A clinical and molecular review of ubiquitous glucose-6-phosphatase deficiency caused by G6PC3 mutations.

    PubMed

    Banka, Siddharth; Newman, William G

    2013-06-13

    The G6PC3 gene encodes the ubiquitously expressed glucose-6-phosphatase enzyme (G-6-Pase β or G-6-Pase 3 or G6PC3). Bi-allelic G6PC3 mutations cause a multi-system autosomal recessive disorder of G6PC3 deficiency (also called severe congenital neutropenia type 4, MIM 612541). To date, at least 57 patients with G6PC3 deficiency have been described in the literature.G6PC3 deficiency is characterized by severe congenital neutropenia, recurrent bacterial infections, intermittent thrombocytopenia in many patients, a prominent superficial venous pattern and a high incidence of congenital cardiac defects and uro-genital anomalies. The phenotypic spectrum of the condition is wide and includes rare manifestations such as maturation arrest of the myeloid lineage, a normocellular bone marrow, myelokathexis, lymphopaenia, thymic hypoplasia, inflammatory bowel disease, primary pulmonary hypertension, endocrine abnormalities, growth retardation, minor facial dysmorphism, skeletal and integument anomalies amongst others. Dursun syndrome is part of this extended spectrum. G6PC3 deficiency can also result in isolated non-syndromic severe neutropenia. G6PC3 mutations in result in reduced enzyme activity, endoplasmic reticulum stress response, increased rates of apoptosis of affected cells and dysfunction of neutrophil activity.In this review we demonstrate that loss of function in missense G6PC3 mutations likely results from decreased enzyme stability. The condition can be diagnosed by sequencing the G6PC3 gene. A number of G6PC3 founder mutations are known in various populations and a possible genotype-phenotype relationship also exists. G6PC3 deficiency should be considered as part of the differential diagnoses in any patient with unexplained congenital neutropenia.Treatment with G-CSF leads to improvement in neutrophil numbers, prevents infections and improves quality of life. Mildly affected patients can be managed with prophylactic antibiotics. Untreated G6PC3 deficiency can

  7. Induction of cellular prion protein (PrPc) under hypoxia inhibits apoptosis caused by TRAIL treatment

    PubMed Central

    Lee, Ju-Hee; Moon, Ji-Hong; Kim, Sung-Wook; Lee, You-Jin; Park, Sang-Youel

    2015-01-01

    Hypoxia decreases cytotoxic responses to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) protein. Cellular prion protein (PrPc) is regulated by HIF-1α in neurons. We hypothesized that PrPc is involved in hypoxia-mediated resistance to TRAIL-induced apoptosis. We found that hypoxia induced PrPc protein and inhibited TRAIL-induced apoptosis. Thus silencing of PrPc increased TRAIL-induced apoptosis under hypoxia. Overexpression of PrPc protein using an adenoviral vector inhibited TRAIL-induced apoptosis. In xenograft model in vivo, shPrPc transfected cells were more sensitive to TRAIL-induced apoptosis than in shMock transfected cells. Molecular chemo-therapy approaches based on the regulation of PrPc expression need to address anti-tumor function of TRAIL under hypoxia. Molecular chemo-therapy approaches based on the regulation of PrPc expression need to address anti-tumor function of TRAIL under hypoxia. PMID:25742790

  8. Onjisaponin B Derived from Radix Polygalae Enhances Autophagy and Accelerates the Degradation of Mutant α-Synuclein and Huntingtin in PC-12 Cells

    PubMed Central

    Wu, An-Guo; Wong, Vincent Kam-Wai; Xu, Su-Wei; Chan, Wai-Kit; Ng, Choi-In; Liu, Liang; Law, Betty Yuen-Kwan

    2013-01-01

    Emerging evidence indicates important protective roles being played by autophagy in neurodegenerative disorders through clearance of aggregate-prone or mutant proteins. In the current study, we aimed to identify autophagy inducers from Chinese medicinal herbs as a potential neuroprotective agent that enhances the clearance of mutant huntingtin and α-synuclein in PC-12 cells. Through intensive screening using the green fluorescent protein-light chain 3 (GFP-LC3) autophagy detection platform, we found that the ethanol extracts of Radix Polygalae (Yuan Zhi) were capable of inducing autophagy. Further investigation showed that among three single components derived from Radix Polygalae—i.e., polygalacic acid, senegenin and onjisaponin B—onjisaponin B was able to induce autophagy and accelerate both the removal of mutant huntingtin and A53T α-synuclein, which are highly associated with Huntington disease and Parkinson disease, respectively. Our study further demonstrated that onjisaponin B induces autophagy via the AMPK-mTOR signaling pathway. Therefore, findings in the current study provide detailed insights into the protective mechanism of a novel autophagy inducer, which is valuable for further investigation as a new candidate agent for modulating neurodegenerative disorders through the reduction of toxicity and clearance of mutant proteins in the cellular level. PMID:24248062

  9. Nuclear protein import is reduced in cells expressing nuclear envelopathy-causing lamin A mutants

    SciTech Connect

    Busch, Albert; Kiel, Tilman; Heupel, Wolfgang-M.; Wehnert, Manfred; Huebner, Stefan

    2009-08-15

    Lamins, which form the nuclear lamina, not only constitute an important determinant of nuclear architecture, but additionally play essential roles in many nuclear functions. Mutations in A-type lamins cause a wide range of human genetic disorders (laminopathies). The importance of lamin A (LaA) in the spatial arrangement of nuclear pore complexes (NPCs) prompted us to study the role of LaA mutants in nuclear protein transport. Two mutants, causing prenatal skin disease restrictive dermopathy (RD) and the premature aging disease Hutchinson Gilford progeria syndrome, were used for expression in HeLa cells to investigate their impact on the subcellular localization of NPC-associated proteins and nuclear protein import. Furthermore, dynamics of the LaA mutants within the nuclear lamina were studied. We observed affected localization of NPC-associated proteins, diminished lamina dynamics for both LaA mutants and reduced nuclear import of representative cargo molecules. Intriguingly, both LaA mutants displayed similar effects on nuclear morphology and functions, despite their differences in disease severity. Reduced nuclear protein import was also seen in RD fibroblasts and impaired lamina dynamics for the nucleoporin Nup153. Our data thus represent the first study of a direct link between LaA mutant expression and reduced nuclear protein import.

  10. Differential interaction between iron and mutant alpha-synuclein causes distinctive Parkinsonian phenotypes in Drosophila.

    PubMed

    Zhu, Zhou-Jing; Wu, Ka-Chun; Yung, Wing-Ho; Qian, Zhong-Ming; Ke, Ya

    2016-04-01

    Alpha-synuclein aggregation is the central hallmark of both sporadic and familial Parkinson's disease (PD). Patients with different PD-causing genetic defects of alpha-synuclein usually show distinctive clinical features that are atypical to sporadic PD. Iron accumulation is invariably found in PD. Recent studies showed that mutant and wild-type alpha-synuclein may have differential interaction with iron and mutant alpha-synuclein toxicity could be preferentially exacerbated by iron. We hence hypothesized that iron overload could selectively influence mutant alpha-synuclein toxicity and disease phenotypes. To test the hypothesis, we investigated if Drosophila melanogaster over-expressing A53T, A30P, and wild-type (WT) alpha-synuclein have different responses to iron treatment. We showed that iron treatment induced similar reduction of survival rate in all flies but induced a more severe motor decline in A53T and A30P mutant alpha-synuclein expressing flies, suggesting interaction between mutant alpha-synuclein and iron. Although no significant difference in total head iron content was found among these flies, we demonstrated that iron treatment induced selective DA neuron loss in motor-related PPM3 cluster only in the flies that express A53T and A30P mutant alpha-synuclein. We provided the first in vivo evidence that iron overload could induce distinctive neuropathology and disease phenotypes in mutant but not WT alpha-synuclein expressing flies, providing insights to the cause of clinical features selectively exhibited by mutant alpha-synuclein carriers.

  11. Erwinia amylovora pyrC mutant causes fire blight despite pyrimidine auxotrophy.

    PubMed

    Ramos, L S; Sinn, J P; Lehman, B L; Pfeufer, E E; Peter, K A; McNellis, T W

    2015-06-01

    Erwinia amylovora bacteria cause fire blight disease, which affects apple and pear production worldwide. The Erw. amylovora pyrC gene encodes a predicted dihydroorotase enzyme involved in pyrimidine biosynthesis. Here, we discovered that the Erw. amylovora pyrC244::Tn5 mutant was a uracil auxotroph. Unexpectedly, the Erw. amylovora pyrC244::Tn5 mutant grew as well as the wild-type in detached immature apple and pear fruits. Fire blight symptoms caused by the pyrC244::Tn5 mutant in immature apple and pear fruits were attenuated compared to those caused by the wild-type. The pyrC244::Tn5 mutant also caused severe fire blight symptoms in apple tree shoots. A plasmid-borne copy of the wild-type pyrC gene restored prototrophy and symptom induction in apple and pear fruit to the pyrC244::Tn5 mutant. These results suggest that Erw. amylovora can obtain sufficient pyrimidine from the host to support bacterial growth and fire blight disease development, although de novo pyrimidine synthesis by Erw. amylovora is required for full symptom development in fruits. Significance and impact of the study: This study provides information about the fire blight host-pathogen interaction. Although the Erwinia amylovora pyrC mutant was strictly auxotrophic for pyrimidine, it grew as well as the wild-type in immature pear and apple fruits and caused severe fire blight disease in apple trees. This suggests that Erw. amylovora can obtain sufficient pyrimidines from host tissue to support growth and fire blight disease development. This situation contrasts with findings in some human bacterial pathogens, which require de novo pyrimidine synthesis for growth in host blood, for example.

  12. Phenotype Sequencing: Identifying the Genes That Cause a Phenotype Directly from Pooled Sequencing of Independent Mutants

    PubMed Central

    Harper, Marc A.; Chen, Zugen; Toy, Traci; Machado, Iara M. P.; Nelson, Stanley F.; Liao, James C.; Lee, Christopher J.

    2011-01-01

    Random mutagenesis and phenotype screening provide a powerful method for dissecting microbial functions, but their results can be laborious to analyze experimentally. Each mutant strain may contain 50–100 random mutations, necessitating extensive functional experiments to determine which one causes the selected phenotype. To solve this problem, we propose a “Phenotype Sequencing” approach in which genes causing the phenotype can be identified directly from sequencing of multiple independent mutants. We developed a new computational analysis method showing that 1. causal genes can be identified with high probability from even a modest number of mutant genomes; 2. costs can be cut many-fold compared with a conventional genome sequencing approach via an optimized strategy of library-pooling (multiple strains per library) and tag-pooling (multiple tagged libraries per sequencing lane). We have performed extensive validation experiments on a set of E. coli mutants with increased isobutanol biofuel tolerance. We generated a range of sequencing experiments varying from 3 to 32 mutant strains, with pooling on 1 to 3 sequencing lanes. Our statistical analysis of these data (4099 mutations from 32 mutant genomes) successfully identified 3 genes (acrB, marC, acrA) that have been independently validated as causing this experimental phenotype. It must be emphasized that our approach reduces mutant sequencing costs enormously. Whereas a conventional genome sequencing experiment would have cost $7,200 in reagents alone, our Phenotype Sequencing design yielded the same information value for only $1200. In fact, our smallest experiments reliably identified acrB and marC at a cost of only $110–$340. PMID:21364744

  13. Silicon phthalocyanine Pc 4 and red light causes apoptosis in HIV-infected cells.

    PubMed

    Ben-Hur, E; Oetjen, J; Horowitz, B

    1997-03-01

    The silicon phthalocyanine HOSiPcOSi(CH3)2(CH2)3 N(CH3)2 (Pc 4), is being studied as a photosensitizer for virus inactivation in red blood cell concentrates (RBCC). The RBCC spiked with cell-free human immunodeficiency virus (HIV) or with HIV actively replicating in the T-lymphocytic cell line CEM can be successfully inactivated (> or = 6 log10) when exposed to 2 microM Pc 4 and 90 J/cm2 red light (600-800 nm). Inactivation of > or = 6 log10 inducible HIV in the latently infected promonocytic cell line U1 occurred at 22.5 J/cm2 (H. Margolis-Nunno et al., Transfusion 36, 743-750, 1996). In order to understand the reason for the increased susceptibility of U1 to photosensitized inactivation we looked for induction of apoptosis by photodynamic treatment (PDT). Agarose gel electrophoresis was used to observe the appearance of a characteristic 180-200 base pair DNA ladder, which can indicate apoptosis. Using this assay it is shown that Pc 4 treatment induced apoptosis in U1 cells in a light dose-dependent manner, starting 30 min after light exposure. Using the ApopTag Plus kit (which attaches a fluorescent label to the 3'-OH ends of the degraded DNA) and flow cytometry, the percentage of cells undergoing apoptosis was quantitated. At 10.5 J/cm2, 3 h after light exposure, about 92.5% of the cells were apoptotic. Under these conditions 99% of the cells eventually die. The CEM cells similarly treated underwent apoptosis at slower kinetics and required higher light doses. Other cell lines latently infected with HIV (ACH-2 and OM 10.1) were as sensitive as U1 to HIV inactivation by Pc 4-PDT (H. Margolis-Nunno et al., Transfusion 36, 743-750, 1996) and underwent apoptosis at a similar kinetic. These results suggest that the enhanced inactivation of HIV in latently infected cells compared to CEM cells by Pc 4-PDT may be due, at least in part, to apoptosis in the former.

  14. Prp8 retinitis pigmentosa mutants cause defects in the transition between the catalytic steps of splicing.

    PubMed

    Mayerle, Megan; Guthrie, Christine

    2016-05-01

    Pre-mRNA splicing must occur with high fidelity and efficiency for proper gene expression. The spliceosome uses DExD/H box helicases to promote on-pathway interactions while simultaneously minimizing errors. Prp8 and Snu114, an EF2-like GTPase, regulate the activity of the Brr2 helicase, promoting RNA unwinding by Brr2 at appropriate points in the splicing cycle and repressing it at others. Mutations linked to retinitis pigmentosa (RP), a disease that causes blindness in humans, map to the Brr2 regulatory region of Prp8. Previous in vitro studies of homologous mutations in Saccharomyces cerevisiaes how that Prp8-RP mutants cause defects in spliceosome activation. Here we show that a subset of RP mutations in Prp8 also causes defects in the transition between the first and second catalytic steps of splicing. Though Prp8-RP mutants do not cause defects in splicing fidelity, they result in an overall decrease in splicing efficiency. Furthermore, genetic analyses link Snu114 GTP/GDP occupancy to Prp8-dependent regulation of Brr2. Our results implicate the transition between the first and second catalytic steps as a critical place in the splicing cycle where Prp8-RP mutants influence splicing efficiency. The location of the Prp8-RP mutants, at the "hinge" that links the Prp8 Jab1-MPN regulatory "tail" to the globular portion of the domain, suggests that these Prp8-RP mutants inhibit regulated movement of the Prp8 Jab1/MPN domain into the Brr2 RNA binding channel to transiently inhibit Brr2. Therefore, in Prp8-linked RP, disease likely results not only from defects in spliceosome assembly and activation, but also because of defects in splicing catalysis.

  15. Familial Parkinson mutant alpha-synuclein causes dopamine neuron dysfunction in transgenic Caenorhabditis elegans.

    PubMed

    Kuwahara, Tomoki; Koyama, Akihiko; Gengyo-Ando, Keiko; Masuda, Mayumi; Kowa, Hisatomo; Tsunoda, Makoto; Mitani, Shohei; Iwatsubo, Takeshi

    2006-01-06

    Mutations in alpha-synuclein gene cause familial form of Parkinson disease, and deposition of wild-type alpha-synuclein as Lewy bodies occurs as a hallmark lesion of sporadic Parkinson disease and dementia with Lewy bodies, implicating alpha-synuclein in the pathogenesis of Parkinson disease and related neurodegenerative diseases. Dopamine neurons in substantia nigra are the major site of neurodegeneration associated with alpha-synuclein deposition in Parkinson disease. Here we establish transgenic Caenorhabditis elegans (TG worms) that overexpresses wild-type or familial Parkinson mutant human alpha-synuclein in dopamine neurons. The TG worms exhibit accumulation of alpha-synuclein in the cell bodies and neurites of dopamine neurons, and EGFP labeling of dendrites is often diminished in TG worms expressing familial Parkinson disease-linked A30P or A53T mutant alpha-synuclein, without overt loss of neuronal cell bodies. Notably, TG worms expressing A30P or A53T mutant alpha-synuclein show failure in modulation of locomotory rate in response to food, which has been attributed to the function of dopamine neurons. This behavioral abnormality was accompanied by a reduction in neuronal dopamine content and was treatable by administration of dopamine. These phenotypes were not seen upon expression of beta-synuclein. The present TG worms exhibit dopamine neuron-specific dysfunction caused by accumulation of alpha-synuclein, which would be relevant to the genetic and compound screenings aiming at the elucidation of pathological cascade and therapeutic strategies for Parkinson disease.

  16. Controlling aggregation propensity in A53T mutant of alpha-synuclein causing Parkinson's disease

    SciTech Connect

    Kumar, Sonu; Sarkar, Anita; Sundar, Durai

    2009-09-18

    Understanding {alpha}-synuclein in terms of fibrillization, aggregation, solubility and stability is fundamental in Parkinson's disease (PD). The three familial mutations, namely, A30P, E46K and A53T cause PD because the hydrophobic regions in {alpha}-synuclein acquire {beta}-sheet configuration, and have a propensity to fibrillize and form amyloids that cause cytotoxicity and neurodegeneration. On simulating the native form and mutants (A30P, E46K and A53T) of {alpha}-synuclein in water solvent, clear deviations are observed in comparison to the all-helical 1XQ8 PDB structure. We have identified two crucial residues, {sup 40}Val and {sup 74}Val, which play key roles in {beta}-sheet aggregation in the hydrophobic regions 36-41 and 68-78, respectively, leading to fibrillization and amyloidosis in familial (A53T) PD. We have also identified V40D{sub V}74D, a double mutant of A53T (the most amyloidogenic mutant). The simultaneous introduction of these two mutations in A53T nearly ends its aggregation propensity, increases its solubility and positively enhances its thermodynamic stability.

  17. Short stature caused by a mutant growth hormone with an antagonistic effect.

    PubMed

    Takahashi, Y; Kaji, H; Okimura, Y; Goji, K; Abe, H; Chihara, K

    1996-10-01

    The molecular basis of biologically inactive GH remained unclear until recently. We have very recently reported a child with short stature and a mutant GH caused by a single missense mutation in the GH-1 gene, which itself cannot transduce the GH-signal to the cells but can blunt the action of wild-type GH by virtue of its greater affinity for the GH binding protein (GHBP)/GH receptor. Briefly the clinical features of the patient are: At the age of 4.9 years his height was 81.7 cm (-6.1 SD) and bone age was 2 years. The patient's serum insulin-like growth factor-1 (IGF-1) concentration was 34 ng/ml. The basal serum GH concentration ranged from 7.0 to 14.0 ng/ml and peak concentrations after insulin hypoglycemia, arginine and L-dopa were 38.0, 15.0 and 35.0 ng/ml, respectively. A heterozygous single base substitution was identified in the GH-1 gene of the proband, predicted to convert codon 77 from arginine to cysteine. Isoelectric focusing revealed the presence of an abnormal GH peak in addition to a normal GH peak. The affinity of expressed mutant GH to GHBP was approximately 6 times higher than that of wild-type GH. The mutant GH not only failed to stimulate tyrosine phosphorylation by itself, but it also inhibited the activity of wild-type GH when added simultaneously even in a one tenth dose of wild-type GH. The child whom we reported is therefore the first case of short stature caused by mutant GH with an antagonistic effect.

  18. Spinocerebellar ataxia type 13 mutant potassium channel alters neuronal excitability and causes locomotor deficits in zebrafish.

    PubMed

    Issa, Fadi A; Mazzochi, Christopher; Mock, Allan F; Papazian, Diane M

    2011-05-04

    Whether changes in neuronal excitability can cause neurodegenerative disease in the absence of other factors such as protein aggregation is unknown. Mutations in the Kv3.3 voltage-gated K(+) channel cause spinocerebellar ataxia type 13 (SCA13), a human autosomal-dominant disease characterized by locomotor impairment and the death of cerebellar neurons. Kv3.3 channels facilitate repetitive, high-frequency firing of action potentials, suggesting that pathogenesis in SCA13 is triggered by changes in electrical activity in neurons. To investigate whether SCA13 mutations alter excitability in vivo, we expressed the human dominant-negative R420H mutant subunit in zebrafish. The disease-causing mutation specifically suppressed the excitability of Kv3.3-expressing, fast-spiking motor neurons during evoked firing and fictive swimming and, in parallel, decreased the precision and amplitude of the startle response. The dominant-negative effect of the mutant subunit on K(+) current amplitude was directly responsible for the reduced excitability and locomotor phenotype. Our data provide strong evidence that changes in excitability initiate pathogenesis in SCA13 and establish zebrafish as an excellent model system for investigating how changes in neuronal activity impair locomotor control and cause cell death.

  19. IBMPFD Disease-Causing Mutant VCP/p97 Proteins Are Targets of Autophagic-Lysosomal Degradation

    PubMed Central

    Bayraktar, Oznur; Akkoc, Yunus; Eberhart, Karin; Kosar, Ali

    2016-01-01

    The ubiquitin-proteasome system (UPS) degrades soluble proteins and small aggregates, whereas macroautophagy (autophagy herein) eliminates larger protein aggregates, tangles and even whole organelles in a lysosome-dependent manner. VCP/p97 was implicated in both pathways. VCP/p97 mutations cause a rare multisystem disease called IBMPFD (Inclusion Body Myopathy with Paget’s Disease and Frontotemporal Dementia). Here, we studied the role IBMPFD-related mutants of VCP/p97 in autophagy. In contrast with the wild-type VCP/p97 protein or R155C or R191Q mutants, the P137L mutant was aggregate-prone. We showed that, unlike commonly studied R155C or R191Q mutants, the P137L mutant protein stimulated both autophagosome and autolysosome formation. Moreover, P137L mutant protein itself was a substrate of autophagy. Starvation- and mTOR inhibition-induced autophagy led to the degradation of the P137L mutant protein, while preserving the wild-type and functional VCP/p97. Strikingly, similar to the P137L mutant, other IBMPFD-related VCP/p97 mutants, namely R93C and G157R mutants induced autophagosome and autolysosome formation; and G157R mutant formed aggregates that could be cleared by autophagy. Therefore, cellular phenotypes caused by P137L mutant expression were not isolated observations, and some other IBMPFD disease-related VCP/p97 mutations could lead to similar outcomes. Our results indicate that cellular mechanisms leading to IBMPFD disease may be various, and underline the importance of studying different disease-associated mutations in order to better understand human pathologies and tailor mutation-specific treatment strategies. PMID:27768726

  20. Increased prevalence of mutant null alleles that cause hereditary fructose intolerance in the American population.

    PubMed

    Coffee, Erin M; Yerkes, Laura; Ewen, Elizabeth P; Zee, Tiffany; Tolan, Dean R

    2010-02-01

    Mutations in the aldolase B gene (ALDOB) impairing enzyme activity toward fructose-1-phosphate cleavage cause hereditary fructose intolerance (HFI). Diagnosis of the disease is possible by identifying known mutant ALDOB alleles in suspected patients; however, the frequencies of mutant alleles can differ by population. Here, 153 American HFI patients with 268 independent alleles were analyzed to identify the prevalence of seven known HFI-causing alleles (A149P, A174D, N334K, Delta4E4, R59Op, A337V, and L256P) in this population. Allele-specific oligonucleotide hybridization analysis was performed on polymerase chain reaction (PCR)-amplified genomic DNA from these patients. In the American population, the missense mutations A149P and A174D are the two most common alleles, with frequencies of 44% and 9%, respectively. In addition, the nonsense mutations Delta4E4 and R59Op are the next most common alleles, with each having a frequency of 4%. Together, the frequencies of all seven alleles make up 65% of HFI-causing alleles in this population. Worldwide, these same alleles make up 82% of HFI-causing mutations. This difference indicates that screening for common HFI alleles is more difficult in the American population. Nevertheless, a genetic screen for diagnosing HFI in America can be improved by including all seven alleles studied here. Lastly, identification of HFI patients presenting with classic symptoms and who have homozygous null genotypes indicates that aldolase B is not required for proper development or metabolic maintenance.

  1. Increased prevalence of mutant null alleles that cause hereditary fructose intolerance in the American population

    PubMed Central

    Coffee, Erin M.; Yerkes, Laura; Ewen, Elizabeth P.; Zee, Tiffany

    2010-01-01

    Mutations in the aldolase B gene (ALDOB) impairing enzyme activity toward fructose-1-phosphate cleavage cause hereditary fructose intolerance (HFI). Diagnosis of the disease is possible by identifying known mutant ALDOB alleles in suspected patients; however, the frequencies of mutant alleles can differ by population. Here, 153 American HFI patients with 268 independent alleles were analyzed to identify the prevalence of seven known HFI-causing alleles (A149P, A174D, N334K, Δ4E4, R59Op, A337V, and L256P) in this population. Allele-specific oligonucleotide hybridization analysis was performed on polymerase chain reaction (PCR)-amplified genomic DNA from these patients. In the American population, the missense mutations A149P and A174D are the two most common alleles, with frequencies of 44% and 9%, respectively. In addition, the nonsense mutations Δ4E4 and R59Op are the next most common alleles, with each having a frequency of 4%. Together, the frequencies of all seven alleles make up 65% of HFI-causing alleles in this population. Worldwide, these same alleles make up 82% of HFI-causing mutations. This difference indicates that screening for common HFI alleles is more difficult in the American population. Nevertheless, a genetic screen for diagnosing HFI in America can be improved by including all seven alleles studied here. Lastly, identification of HFI patients presenting with classic symptoms and who have homozygous null genotypes indicates that aldolase B is not required for proper development or metabolic maintenance. PMID:20033295

  2. What can long-lived mutants tell us about mechanisms causing aging and lifespan variation in natural environments?

    PubMed

    Briga, Michael; Verhulst, Simon

    2015-11-01

    Long-lived mutants of model organisms have brought remarkable progress in our understanding of aging mechanisms. However, long-lived mutants are usually maintained in optimal standardized laboratory environments (SLEs), and it is not obvious to what extent insights from long-lived mutants in SLEs can be generalized to more natural environments. To address this question, we reviewed experiments that compared the fitness and lifespan advantage of long-lived mutants relative to wild type controls in SLEs and more challenging environments in various model organisms such as yeast Saccharomyces cerevisiae, the nematode worm Caenorhabditis elegans, the fruitfly Drosophila melanogaster and the mouse Mus musculus. In competition experiments over multiple generations, the long-lived mutants had a lower fitness relative to wild type controls, and this disadvantage was the clearest when the environment included natural challenges such as limited food (N=6 studies). It is well known that most long-lived mutants have impaired reproduction, which provides one reason for the fitness disadvantage. However, based on 12 experiments, we found that the lifespan advantage of long-lived mutants is diminished in more challenging environments, often to the extent that the wild type controls outlive the long-lived mutants. Thus, it appears that information on aging mechanisms obtained from long-lived mutants in SLEs may be specific to such environments, because those same mechanisms do not extend lifespan in more natural environments. This suggests that different mechanisms cause variation in aging and lifespan in SLEs compared to natural populations.

  3. Oculomotor instabilities in zebrafish mutant belladonna: a behavioral model for congenital nystagmus caused by axonal misrouting.

    PubMed

    Huang, Ying-Yu; Rinner, Oliver; Hedinger, Patrik; Liu, Shih-Chii; Neuhauss, Stephan C F

    2006-09-27

    A large fraction of homozygous zebrafish mutant belladonna (bel) larvae display a reversed optokinetic response (OKR) that correlates with failure of the retinal ganglion cells to cross the midline and form the optic chiasm. Some of these achiasmatic mutants display strong spontaneous eye oscillations (SOs) in the absence of motion in the surround. The presentation of a stationary grating was necessary and sufficient to evoke SO. Both OKR reversal and SO depend on vision and are contrast sensitive. We built a quantitative model derived from bel fwd (forward) eye behaviors. To mimic the achiasmatic condition, we reversed the sign of the retinal slip velocity in the model, thereby successfully reproducing both reversed OKR and SO. On the basis of the OKR data, and with the support of the quantitative model, we hypothesize that the reversed OKR and the SO can be completely attributed to RGC misrouting. The strong resemblance between the SO and congenital nystagmus (CN) seen in humans with defective retinotectal projections implies that CN, of so far unknown etiology, may be directly caused by a projection defect.

  4. Reduction of germ cells in the Odysseus null mutant causes male fertility defect in Drosophila melanogaster.

    PubMed

    Cheng, Ya-Jen; Fang, Shu; Tsaur, Shun-Chern; Chen, Yi-Ling; Fu, Hua-Wen; Patel, Nipam H; Ting, Chau-Ti

    2012-01-01

    Odysseus (OdsH) has been identified as a hybrid male sterility gene between Drosophila mauritiana and D. simulans with accelerated evolutionary rate in both expression and DNA sequence. Loss of a testis-specific expression of OdsH causes male fertility defect in D. melanogaster. Yet, the underlying mechanisms at the cellular level are unknown. In an attempt to identify the possible mechanisms and functional roles of OdsH in spermatogenesis, the cell numbers at different developmental stages during spermatogenesis between the OdsH null mutant and wild-type flies were compared. The results showed that the early developing germ cells, including spermatogonia and spermatocytes, were reduced in the OdsH mutant males. In addition, the number of germline stem cells in aged males was also reduced, presumably due to the disruption of germline stem cell maintenance, which resulted in more severe fertility defect. These results suggest that the function of the enhancement of sperm production by OdsH acted across males of all ages.

  5. Molecular pathogenesis of Spondylocheirodysplastic Ehlers-Danlos syndrome caused by mutant ZIP13 proteins

    PubMed Central

    Bin, Bum-Ho; Hojyo, Shintaro; Hosaka, Toshiaki; Bhin, Jinhyuk; Kano, Hiroki; Miyai, Tomohiro; Ikeda, Mariko; Kimura-Someya, Tomomi; Shirouzu, Mikako; Cho, Eun-Gyung; Fukue, Kazuhisa; Kambe, Taiho; Ohashi, Wakana; Kim, Kyu-Han; Seo, Juyeon; Choi, Dong-Hwa; Nam, Yeon-Ju; Hwang, Daehee; Fukunaka, Ayako; Fujitani, Yoshio; Yokoyama, Shigeyuki; Superti-Furga, Andrea; Ikegawa, Shiro; Lee, Tae Ryong; Fukada, Toshiyuki

    2014-01-01

    The zinc transporter protein ZIP13 plays critical roles in bone, tooth, and connective tissue development, and its dysfunction is responsible for the spondylocheirodysplastic form of Ehlers-Danlos syndrome (SCD-EDS, OMIM 612350). Here, we report the molecular pathogenic mechanism of SCD-EDS caused by two different mutant ZIP13 proteins found in human patients: ZIP13G64D, in which Gly at amino acid position 64 is replaced by Asp, and ZIP13ΔFLA, which contains a deletion of Phe-Leu-Ala. We demonstrated that both the ZIP13G64D and ZIP13ΔFLA protein levels are decreased by degradation via the valosin-containing protein (VCP)-linked ubiquitin proteasome pathway. The inhibition of degradation pathways rescued the protein expression levels, resulting in improved intracellular Zn homeostasis. Our findings uncover the pathogenic mechanisms elicited by mutant ZIP13 proteins. Further elucidation of these degradation processes may lead to novel therapeutic targets for SCD-EDS. PMID:25007800

  6. A requirement for recombinational repair in Saccharomyces cerevisiae is caused by DNA replication defects of mec1 mutants.

    PubMed Central

    Merrill, B J; Holm, C

    1999-01-01

    To examine the role of the RAD52 recombinational repair pathway in compensating for DNA replication defects in Saccharomyces cerevisiae, we performed a genetic screen to identify mutants that require Rad52p for viability. We isolated 10 mec1 mutations that display synthetic lethality with rad52. These mutations (designated mec1-srf for synthetic lethality with rad-fifty-two) simultaneously cause two types of phenotypes: defects in the checkpoint function of Mec1p and defects in the essential function of Mec1p. Velocity sedimentation in alkaline sucrose gradients revealed that mec1-srf mutants accumulate small single-stranded DNA synthesis intermediates, suggesting that Mec1p is required for the normal progression of DNA synthesis. sml1 suppressor mutations suppress both the accumulation of DNA synthesis intermediates and the requirement for Rad52p in mec1-srf mutants, but they do not suppress the checkpoint defect in mec1-srf mutants. Thus, it appears to be the DNA replication defects in mec1-srf mutants that cause the requirement for Rad52p. By using hydroxyurea to introduce similar DNA replication defects, we found that single-stranded DNA breaks frequently lead to double-stranded DNA breaks that are not rapidly repaired in rad52 mutants. Taken together, these data suggest that the RAD52 recombinational repair pathway is required to prevent or repair double-stranded DNA breaks caused by defective DNA replication in mec1-srf mutants. PMID:10511542

  7. Common increase of GATA-3 level in PC-12 cells by three teratogens causing autism spectrum disorders.

    PubMed

    Rout, Ujjwal K; Clausen, Pete

    2009-06-01

    Autism spectrum disorder (ASD) is a disease of neuro-developmental origin of uncertain etiology. The current understanding is that both genetic and environmental factors contribute to the development of ASD. Exposure to valproate, thalidomide and alcohol during gestation are amongst the environmental triggers that are associated with the development of ASD. These teratogens may disturb the ontogeny of the brain by altering the expression pattern of genes that regulate the normal development of the brain. In this study, a neuron-like PC-12 cell model was used to examine the effects of these compounds on the binding potential of 50 different transcription factors to understand the molecular mechanism/s that may be involved in the teratogenesis caused by these agents. Cells in culture were treated with low or high concentrations of teratogens within a range that are reported in the blood of individuals. A pronounced increase in GATA transcription factor binding was observed for all three teratogens. Furthermore, Western blot analysis showed that GATA-3 level in the nuclear fractions was enhanced by each of the three teratogens. Results suggest that altered gene expression pattern due to heightened GATA-3 activities in the fetral brains following exposure to these teratogens may contribute to the development of ASD.

  8. Expression of a mutant form of cellulose synthase AtCesA7 causes dominant negative effect on cellulose biosynthesis.

    PubMed

    Zhong, Ruiqin; Morrison, W Herbert; Freshour, Glenn D; Hahn, Michael G; Ye, Zheng-Hua

    2003-06-01

    Cellulose synthase catalytic subunits (CesAs) have been implicated in catalyzing the biosynthesis of cellulose, the major component of plant cell walls. Interactions between CesA subunits are thought to be required for normal cellulose synthesis, which suggests that incorporation of defective CesA subunits into cellulose synthase complex could potentially cause a dominant effect on cellulose synthesis. However, all CesA mutants so far reported have been shown to be recessive in terms of cellulose synthesis. In the course of studying the molecular mechanisms regulating secondary wall formation in fibers, we have found that a mutant allele of AtCesA7 gene in the fra5 (fragile fiber 5) mutant causes a semidominant phenotype in the reduction of fiber cell wall thickness and cellulose content. The fra5 missense mutation occurred in a conserved amino acid located in the second cytoplasmic domain of AtCesA7. Overexpression of the fra5 mutant cDNA in wild-type plants not only reduced secondary wall thickness and cellulose content but also decreased primary wall thickness and cell elongation. In contrast, overexpression of the fra6 mutant form of AtCesA8 did not cause any reduction in cell wall thickness and cellulose content. These results suggest that the fra5 mutant protein may interfere with the function of endogenous wild-type CesA proteins, thus resulting in a dominant negative effect on cellulose biosynthesis.

  9. Sodium/Iodide Symporter Mutant V270E Causes Stunted Growth but No Cognitive Deficiency

    PubMed Central

    Nicola, Juan Pablo; Reyna-Neyra, Andrea; Saenger, Paul; Rodriguez-Buritica, David F.; Gamez Godoy, José David; Muzumdar, Radhika; Amzel, L. Mario

    2015-01-01

    Context: Iodide (I−), an essential constituent of the thyroid hormones, is actively accumulated in the thyroid by the Na+/I− symporter (NIS), a key plasma membrane protein encoded by the slc5a5 gene. Mutations in slc5a5 cause I− transport defects (ITDs), autosomal-recessive disorders in which I− accumulation is totally or partially impaired, leading to congenital hypothyroidism. The characterization of NIS mutants has yielded significant insights into the molecular mechanism of NIS. Objective: This study aimed to determine the basis of a patient's ITD clinical phenotype, by sequencing her slc5a5 gene. Design: Genomic DNA was purified and the slc5a5 gene sequence determined. Functional in vitro studies were performed to characterize the V270E NIS mutant. Patient: The index patient was diagnosed with hypothyroidism with minimal radioiodide uptake in a normally located, although enlarged, thyroid gland. Results: We identified a new NIS mutation: V270E. The patient had the compound heterozygous NIS mutation R124H/V270E. R124H NIS has been characterized previously. We show that V270E markedly reduces I− uptake via a pronounced (but not total) impairment of the protein's plasma membrane targeting. Remarkably, V270E is intrinsically active. Therefore, a negative charge at position 270 interferes with NIS cell surface trafficking. The patient's minimal I− uptake enabled sufficient thyroid hormone biosynthesis to prevent cognitive impairment. Conclusions: A nonpolar residue at position 270, which all members of the SLC5A family have, is required for NIS plasma membrane targeting. PMID:26204134

  10. Generation of Pc 1 waves by the ion temperature anisotropy associated with fast shocks caused by sudden impulses

    NASA Technical Reports Server (NTRS)

    Mandt, M. E.; Lee, L. C.

    1991-01-01

    The high correlation of Pc 1 events with magnetospheric compressions is known. A mechanism is proposed which leads to the generation of Pc 1 waves. The interaction of a dynamic pressure pulse with the earth's bow shock leads to the formation of a weak fast-mode shock propagating into the magnetoshealth. The shock wave can pass right through a tangential discontinuity (magnetopause) and into the magnetosphere, without disturbing either of the structures. In a quasiperpendicular geometry, the shock wave exhibits anisotropic heating. This anisotropy drives unstable ion-cyclotron waves which can contribute to the generation of the Pc 1 waves which are detected. The viability of the mechanism is demonstrated with simulations. This mechanism could explain the peak in the occurrence of observed Pc 1 waves in the postnoon sector where a field-aligned discontinuity in the solar wind would most often be parallel to the magnetopause surface due to the average Parker-spiral magnetic-field configuration.

  11. Mutant alpha-synuclein causes age-dependent neuropathology in monkey brain.

    PubMed

    Yang, Weili; Wang, Guohao; Wang, Chuan-En; Guo, Xiangyu; Yin, Peng; Gao, Jinquan; Tu, Zhuchi; Wang, Zhengbo; Wu, Jing; Hu, Xintian; Li, Shihua; Li, Xiao-Jiang

    2015-05-27

    Parkinson's disease (PD) is an age-dependent neurodegenerative disease that often occurs in those over age 60. Although rodents and small animals have been used widely to model PD and investigate its pathology, their short life span makes it difficult to assess the aging-related pathology that is likely to occur in PD patient brains. Here, we used brain tissues from rhesus monkeys at 2-3, 7-8, and >15 years of age to examine the expression of Parkin, PINK1, and α-synuclein, which are known to cause PD via loss- or gain-of-function mechanisms. We found that α-synuclein is increased in the older monkey brains, whereas Parkin and PINK1 are decreased or remain unchanged. Because of the gain of toxicity of α-synuclein, we performed stereotaxic injection of lentiviral vectors expressing mutant α-synuclein (A53T) into the substantia nigra of monkeys and found that aging also increases the accumulation of A53T in neurites and its associated neuropathology. A53T also causes more extensive reactive astrocytes and axonal degeneration in monkey brain than in mouse brain. Using monkey brain tissues, we found that A53T interacts with neurofascin, an adhesion molecule involved in axon subcellular targeting and neurite outgrowth. Aged monkey brain tissues show an increased interaction of neurofascin with A53T. Overexpression of A53T causes neuritic toxicity in cultured neuronal cells, which can be attenuated by transfected neurofascin. These findings from nonhuman primate brains reveal age-dependent pathological and molecular changes that could contribute to the age-dependent neuropathology in PD.

  12. [Effect of microRNA on proliferation caused by mutant HBx in human hepatocytes].

    PubMed

    Fu, Xiao-yu; Tan, De-ming; Hou, Zhou-hua; Hu, Zhi-liang; Liu, Guo-zhen; Ouyang, Yi; Liu, Fei

    2012-08-01

    To study the effect of micro (mi)RNA on cellular proliferation induced by hepatitis B x protein, HBx, in human liver cells and to investigate the underlying molecular mechanism of this cancer-related effect. The human L02 hepatocyte cell line was stably transfected with HBx (L02/HBx) or an HBx mutant (L02/HBx-d382) that induces higher levels of cellular proliferation. The differential miRNA expression profiles were determined by microarray analysis and confirmed by real-time PCR. Two miRNAs, miR-338-3p and miR-551b, that were found to be significantly down-regulated in the L02/HBx-d382 cells were selected for further study and transfected individually into cells using the lipofectamine procedure. The cell survival rate was analyzed by MTT assay, and cell cycles were assessed by flow cytometry. Expressions of cyclinD1, cyclinG1, and E2F1 were assessed by real-time PCR and Western blotting. Compared with the microarray miRNA profile of L02/pcDNA3.0 cells, six miRNAs were up-regulated and five miRNAs were down-regulated in the L02/HBx-d382 cells, while four miRNAs were up-regulated and 12 were down-regulated in the L02/HBx cells. The microarray results were consistent with real-time PCR results. Transfection of miR-338-3p and miR-551b significantly inhibited the cell survival rates (P less than 0.001) and induced G0/G1 phase cycle arrest. According to MTT results: for L02/HBx-d382 cells, compared with lipofectamine or non-transfected (NC) controls, the t value of miR-338-3p was 10.402, 9.133 and the t value of miR-551b was 8.763, 7.403; for L02/HBx cells, compared with lipofectamine or NC controls, the t value of miR-338-3p was 9.105, 8.074 and the t value of miR-551b was 7.673, 7.52. According to flow cytometry results: for L02/HBx-d382 cells, compared with lipofectamine or NC controls, the t value of miR-338-3p was 12.173, 11.107 and the t value of miR-551b was 15.364, 13.377; for L02/HBx cells, compared with lipofectamine or NC controls, the t value of miR-338-3p

  13. A DltA mutant of Haemophilus ducreyi Is partially attenuated in its ability to cause pustules in human volunteers.

    PubMed

    Janowicz, Diane; Leduc, Isabelle; Fortney, Kate R; Katz, Barry P; Elkins, Christopher; Spinola, Stanley M

    2006-02-01

    Haemophilus ducreyi produces two outer membrane proteins, called DltA (H. ducreyi lectin A) and DsrA (H. ducreyi serum resistance A), that contribute to the ability of the organism to evade complement-mediated serum killing. In contrast to their isogenic parent strain, 35000HP, the DsrA mutant FX517 exhibits 0% survival in 50% normal human serum and the DltA mutant FX533 exhibits 23% survival. Compared to 35000HP, FX517 does not cause pustule formation in human volunteers. To test whether DltA was required for virulence in humans, seven volunteers were experimentally infected with 35000HP and FX533. Four subjects were inoculated with fixed doses of 35000HP (101 CFU or 130 CFU) at three sites on one arm and escalating doses of FX533 (range, 46 CFU to 915 CFU) at three sites on the other arm. Pustules only developed at mutant-injected sites at doses nearly twofold higher than that of the parent, suggesting that FX533 was partially attenuated. Three subjects were inoculated with similar doses of the parent (67 CFU) and mutant (104 CFU) at three sites. Pustules formed at five of nine parent sites and one of nine mutant sites. Overall, the papule and pustule formation rates for 35000HP and FX533 were similar for the trial. However, for the five subjects who received similar doses of the parent and mutant, pustules developed at 7 of 15 sites (46.7%; 95% confidence interval [CI], 16.9% to 76.5%) inoculated with the parent and at 1 of 15 (6.7%; 95% CI, 0.1% to 18.4%) sites inoculated with the mutant (P = 0.043). We concluded that the DltA mutant was attenuated in its ability to cause disease at doses similar to that of the parent.

  14. Parkinson's disease-associated mutant VPS35 causes mitochondrial dysfunction by recycling DLP1 complexes.

    PubMed

    Wang, Wenzhang; Wang, Xinglong; Fujioka, Hisashi; Hoppel, Charles; Whone, Alan L; Caldwell, Maeve A; Cullen, Peter J; Liu, Jun; Zhu, Xiongwei

    2016-01-01

    Mitochondrial dysfunction represents a critical step during the pathogenesis of Parkinson's disease (PD), and increasing evidence suggests abnormal mitochondrial dynamics and quality control as important underlying mechanisms. The VPS35 gene, which encodes a key component of the membrane protein-recycling retromer complex, is the third autosomal-dominant gene associated with PD. However, how VPS35 mutations lead to neurodegeneration remains unclear. Here we demonstrate that PD-associated VPS35 mutations caused mitochondrial fragmentation and cell death in cultured neurons in vitro, in mouse substantia nigra neurons in vivo and in human fibroblasts from an individual with PD who has the VPS35(D620N) mutation. VPS35-induced mitochondrial deficits and neuronal dysfunction could be prevented by inhibition of mitochondrial fission. VPS35 mutants showed increased interaction with dynamin-like protein (DLP) 1, which enhanced turnover of the mitochondrial DLP1 complexes via the mitochondria-derived vesicle-dependent trafficking of the complexes to lysosomes for degradation. Notably, oxidative stress increased the VPS35-DLP1 interaction, which we also found to be increased in the brains of sporadic PD cases. These results revealed a novel cellular mechanism for the involvement of VPS35 in mitochondrial fission, dysregulation of which is probably involved in the pathogenesis of familial, and possibly sporadic, PD.

  15. Susceptibility of Maize to Stalk Rot Caused by Fusarium graminearum Deoxynivalenol and Zearalenone Mutants.

    PubMed

    Quesada-Ocampo, L M; Al-Haddad, J; Scruggs, A C; Buell, C R; Trail, F

    2016-08-01

    Fusarium graminearum is a destructive pathogen of cereals that can cause stalk rot in maize. Stalk rot results in yield losses due to impaired grain filling, premature senescence, and lodging, which limits production and harvesting of ears. In addition, mycotoxins can make infected tissues unfit for silage. Our objectives were to evaluate the natural variation in stalk rot resistance among maize inbreds, to establish whether deoxynivalenol (DON)- and zearalenone (ZEA)-deficient strains are pathogenic on a panel of diverse inbreds, and to quantify the accumulation of DON in infected stalk tissue. Wild-type F. graminearum and mycotoxin mutants (DON and ZEA) were used to separately inoculate stalks of 9-week-old plants of 20 inbreds in the greenhouse. Plants were evaluated for lesion area at the inoculation point at 0, 2, 14, and 28 days postinoculation and tissues around lesions were sampled to determine the DON content. Regardless of their ability to produce DON or ZEA, all tested F. graminearum strains caused stalk rot; however, significant differences in disease levels were detected. Among the tested inbreds, Mp717 was resistant to all three F. graminearum strains while Mp317 and HP301 were only partially resistant. Accumulation of DON was significantly lower in infected stalks of the resistant and partially resistant inbreds than the susceptible inbreds. Analysis of the 20 inbreds using data from 17 simple-sequence repeats revealed population structure among the individuals; however, there was no association between genetic clustering and stalk rot resistance. These findings are an additional step toward breeding maize inbreds suitable for planting in fields infested with F. graminearum.

  16. Functional Loss of Bmsei Causes Thermosensitive Epilepsy in Contractile Mutant Silkworm, Bombyx mori

    PubMed Central

    Nie, Hongyi; Cheng, Tingcai; Huang, Xiaofeng; Zhou, Mengting; Zhang, Yinxia; Dai, Fangyin; Mita, Kazuei; Xia, Qingyou; Liu, Chun

    2015-01-01

    The thermoprotective mechanisms of insects remain largely unknown. We reported the Bombyx mori contractile (cot) behavioral mutant with thermo-sensitive seizures phenotype. At elevated temperatures, the cot mutant exhibit seizures associated with strong contractions, rolling, vomiting, and a temporary lack of movement. We narrowed a region containing cot to ~268 kb by positional cloning and identified the mutant gene as Bmsei which encoded a potassium channel protein. Bmsei was present in both the cell membrane and cytoplasm in wild-type ganglia but faint in cot. Furthermore, Bmsei was markedly decreased upon high temperature treatment in cot mutant. With the RNAi method and injecting potassium channel blockers, the wild type silkworm was induced the cot phenotype. These results demonstrated that Bmsei was responsible for the cot mutant phenotype and played an important role in thermoprotection in silkworm. Meanwhile, comparative proteomic approach was used to investigate the proteomic differences. The results showed that the protein of Hsp-1 and Tn1 were significantly decreased and increased on protein level in cot mutant after thermo-stimulus, respectively. Our data provide insights into the mechanism of thermoprotection in insect. As cot phenotype closely resembles human epilepsy, cot might be a potential model for the mechanism of epilepsy in future. PMID:26198671

  17. Functional Loss of Bmsei Causes Thermosensitive Epilepsy in Contractile Mutant Silkworm, Bombyx mori

    NASA Astrophysics Data System (ADS)

    Nie, Hongyi; Cheng, Tingcai; Huang, Xiaofeng; Zhou, Mengting; Zhang, Yinxia; Dai, Fangyin; Mita, Kazuei; Xia, Qingyou; Liu, Chun

    2015-07-01

    The thermoprotective mechanisms of insects remain largely unknown. We reported the Bombyx mori contractile (cot) behavioral mutant with thermo-sensitive seizures phenotype. At elevated temperatures, the cot mutant exhibit seizures associated with strong contractions, rolling, vomiting, and a temporary lack of movement. We narrowed a region containing cot to ~268 kb by positional cloning and identified the mutant gene as Bmsei which encoded a potassium channel protein. Bmsei was present in both the cell membrane and cytoplasm in wild-type ganglia but faint in cot. Furthermore, Bmsei was markedly decreased upon high temperature treatment in cot mutant. With the RNAi method and injecting potassium channel blockers, the wild type silkworm was induced the cot phenotype. These results demonstrated that Bmsei was responsible for the cot mutant phenotype and played an important role in thermoprotection in silkworm. Meanwhile, comparative proteomic approach was used to investigate the proteomic differences. The results showed that the protein of Hsp-1 and Tn1 were significantly decreased and increased on protein level in cot mutant after thermo-stimulus, respectively. Our data provide insights into the mechanism of thermoprotection in insect. As cot phenotype closely resembles human epilepsy, cot might be a potential model for the mechanism of epilepsy in future.

  18. Human congenital myopathy actin mutants cause myopathy and alter Z-disc structure in Drosophila flight muscle.

    PubMed

    Sevdali, Maria; Kumar, Vikash; Peckham, Michelle; Sparrow, John

    2013-03-01

    Over 190 mutations in the human skeletal muscle α-actin gene, ACTA1 cause congenital actin myopathies. We transgenically expressed six different mutant actins, G15R, I136M, D154N, V163L, V163M and D292V in Drosophila indirect flight muscles and investigated their effects in flies that express one wild type and one mutant actin copy. All the flies were flightless, and the IFMs showed incomplete Z-discs, disorganised actin filaments and 'zebra bodies'. No differences in levels of sarcomeric protein expression were observed, but tropomodulin staining was somewhat disrupted in D164N, V163L, G15R and V163M heterozygotes. A single copy of D292V mutant actin rescued the hypercontractile phenotypes caused by TnI and TnT mutants, suggesting that the D292V mutation interferes with thin filament regulation. Our results show that expression of actin mutations homologous to those in humans in the indirect flight muscles of Drosophila disrupt sarcomere organisation, with somewhat similar phenotypes to those observed in humans. Using Drosophila to study actin mutations may help aid our understanding of congential myopathies caused by actin mutations.

  19. In vivo expression of mammalian BiP ATPase mutants causes disruption of the endoplasmic reticulum.

    PubMed Central

    Hendershot, L M; Wei, J Y; Gaut, J R; Lawson, B; Freiden, P J; Murti, K G

    1995-01-01

    BiP possesses ATP binding/hydrolysis activities that are thought to be essential for its ability to chaperone protein folding and assembly in the endoplasmic reticulum (ER). We have produced a series of point mutations in a hamster BiP clone that inhibit ATPase activity and have generated a species-specific anti-BiP antibody to monitor the effects of mutant hamster BiP expression in COS monkey cells. The enzymatic inactivation of BiP did not interfere with its ability to bind to Ig heavy chains in vivo but did inhibit ATP-mediated release of heavy chains in vitro. Immunofluorescence staining and electron microscopy revealed vesiculation of the ER membranes in COS cells expressing BiP ATPase mutants. ER disruption was not observed when a "44K" fragment of BiP that did not include the protein binding domain was similarly mutated but was observed when the protein binding region of BiP was expressed without an ATP binding domain. This suggests that BiP binding to target proteins as an inactive chaperone is responsible for the ER disruption. This is the first report on the in vivo expression of mammalian BiP mutants and is demonstration that in vitro-identified ATPase mutants behave as dominant negative mutants when expressed in vivo. Images PMID:7612964

  20. Chloroplast dysfunction causes multiple defects in cell cycle progression in the Arabidopsis crumpled leaf mutant.

    PubMed

    Hudik, Elodie; Yoshioka, Yasushi; Domenichini, Séverine; Bourge, Mickaël; Soubigout-Taconnat, Ludivine; Mazubert, Christelle; Yi, Dalong; Bujaldon, Sandrine; Hayashi, Hiroyuki; De Veylder, Lieven; Bergounioux, Catherine; Benhamed, Moussa; Raynaud, Cécile

    2014-09-01

    The majority of research on cell cycle regulation is focused on the nuclear events that govern the replication and segregation of the genome between the two daughter cells. However, eukaryotic cells contain several compartmentalized organelles with specialized functions, and coordination among these organelles is required for proper cell cycle progression, as evidenced by the isolation of several mutants in which both organelle function and overall plant development were affected. To investigate how chloroplast dysfunction affects the cell cycle, we analyzed the crumpled leaf (crl) mutant of Arabidopsis (Arabidopsis thaliana), which is deficient for a chloroplastic protein and displays particularly severe developmental defects. In the crl mutant, we reveal that cell cycle regulation is altered drastically and that meristematic cells prematurely enter differentiation, leading to reduced plant stature and early endoreduplication in the leaves. This response is due to the repression of several key cell cycle regulators as well as constitutive activation of stress-response genes, among them the cell cycle inhibitor SIAMESE-RELATED5. One unique feature of the crl mutant is that it produces aplastidic cells in several organs, including the root tip. By investigating the consequence of the absence of plastids on cell cycle progression, we showed that nuclear DNA replication occurs in aplastidic cells in the root tip, which opens future research prospects regarding the dialogue between plastids and the nucleus during cell cycle regulation in higher plants.

  1. Plastidic phosphoglucomutase and ADP-glucose pyrophosphorylase mutants impair starch synthesis in rice pollen grains and cause male sterility

    PubMed Central

    Lee, Sang-Kyu; Eom, Joon-Seob; Hwang, Seon-Kap; Shin, Dongjin; An, Gynheung; Okita, Thomas W.; Jeon, Jong-Seong

    2016-01-01

    To elucidate the starch synthesis pathway and the role of this reserve in rice pollen, we characterized mutations in the plastidic phosphoglucomutase, OspPGM, and the plastidic large subunit of ADP-glucose (ADP-Glc) pyrophosphorylase, OsAGPL4. Both genes were up-regulated in maturing pollen, a stage when starch begins to accumulate. Progeny analysis of self-pollinated heterozygous lines carrying the OspPGM mutant alleles, osppgm-1 and osppgm-2, or the OsAGPL4 mutant allele, osagpl4-1, as well as reciprocal crosses between the wild type (WT) and heterozygotes revealed that loss of OspPGM or OsAGPL4 caused male sterility, with the former condition rescued by the introduction of the WT OspPGM gene. While iodine staining and transmission electron microscopy analyses of pollen grains from homozygous osppgm-1 lines produced by anther culture confirmed the starch null phenotype, pollen from homozygous osagpl4 mutant lines, osagpl4-2 and osagpl4-3, generated by the CRISPR/Cas system, accumulated small amounts of starch which were sufficient to produce viable seed. Such osagpl4 mutant pollen, however, was unable to compete against WT pollen successfully, validating the important role of this reserve in fertilization. Our results demonstrate that starch is mainly polymerized from ADP-Glc synthesized from plastidic hexose phosphates in rice pollen and that starch is an essential requirement for successful fertilization in rice. PMID:27588462

  2. Plastidic phosphoglucomutase and ADP-glucose pyrophosphorylase mutants impair starch synthesis in rice pollen grains and cause male sterility.

    PubMed

    Lee, Sang-Kyu; Eom, Joon-Seob; Hwang, Seon-Kap; Shin, Dongjin; An, Gynheung; Okita, Thomas W; Jeon, Jong-Seong

    2016-10-01

    To elucidate the starch synthesis pathway and the role of this reserve in rice pollen, we characterized mutations in the plastidic phosphoglucomutase, OspPGM, and the plastidic large subunit of ADP-glucose (ADP-Glc) pyrophosphorylase, OsAGPL4 Both genes were up-regulated in maturing pollen, a stage when starch begins to accumulate. Progeny analysis of self-pollinated heterozygous lines carrying the OspPGM mutant alleles, osppgm-1 and osppgm-2, or the OsAGPL4 mutant allele, osagpl4-1, as well as reciprocal crosses between the wild type (WT) and heterozygotes revealed that loss of OspPGM or OsAGPL4 caused male sterility, with the former condition rescued by the introduction of the WT OspPGM gene. While iodine staining and transmission electron microscopy analyses of pollen grains from homozygous osppgm-1 lines produced by anther culture confirmed the starch null phenotype, pollen from homozygous osagpl4 mutant lines, osagpl4-2 and osagpl4-3, generated by the CRISPR/Cas system, accumulated small amounts of starch which were sufficient to produce viable seed. Such osagpl4 mutant pollen, however, was unable to compete against WT pollen successfully, validating the important role of this reserve in fertilization. Our results demonstrate that starch is mainly polymerized from ADP-Glc synthesized from plastidic hexose phosphates in rice pollen and that starch is an essential requirement for successful fertilization in rice.

  3. Cystic fibrosis-adapted Pseudomonas aeruginosa quorum sensing lasR mutants cause hyperinflammatory responses.

    PubMed

    LaFayette, Shantelle L; Houle, Daniel; Beaudoin, Trevor; Wojewodka, Gabriella; Radzioch, Danuta; Hoffman, Lucas R; Burns, Jane L; Dandekar, Ajai A; Smalley, Nicole E; Chandler, Josephine R; Zlosnik, James E; Speert, David P; Bernier, Joanie; Matouk, Elias; Brochiero, Emmanuelle; Rousseau, Simon; Nguyen, Dao

    2015-07-01

    Cystic fibrosis lung disease is characterized by chronic airway infections with the opportunistic pathogen Pseudomonas aeruginosa and severe neutrophilic pulmonary inflammation. P. aeruginosa undergoes extensive genetic adaptation to the cystic fibrosis (CF) lung environment, and adaptive mutations in the quorum sensing regulator gene lasR commonly arise. We sought to define how mutations in lasR alter host-pathogen relationships. We demonstrate that lasR mutants induce exaggerated host inflammatory responses in respiratory epithelial cells, with increased accumulation of proinflammatory cytokines and neutrophil recruitment due to the loss of bacterial protease- dependent cytokine degradation. In subacute pulmonary infections, lasR mutant-infected mice show greater neutrophilic inflammation and immunopathology compared with wild-type infections. Finally, we observed that CF patients infected with lasR mutants have increased plasma interleukin-8 (IL-8), a marker of inflammation. These findings suggest that bacterial adaptive changes may worsen pulmonary inflammation and directly contribute to the pathogenesis and progression of chronic lung disease in CF patients.

  4. Mutant SNAP25B causes myasthenia, cortical hyperexcitability, ataxia, and intellectual disability

    PubMed Central

    Selcen, Duygu; Brengman, Joan

    2014-01-01

    Objective: To identify and characterize the molecular basis of a syndrome associated with myasthenia, cortical hyperexcitability, cerebellar ataxia, and intellectual disability. Methods: We performed in vitro microelectrode studies of neuromuscular transmission, performed exome and Sanger sequencing, and analyzed functional consequences of the identified mutation in expression studies. Results: Neuromuscular transmission at patient endplates was compromised by reduced evoked quantal release. Exome sequencing identified a dominant de novo variant, p.Ile67Asn, in SNAP25B, a SNARE protein essential for exocytosis of synaptic vesicles from nerve terminals and of dense-core vesicles from endocrine cells. Ca2+-triggered exocytosis is initiated when synaptobrevin attached to synaptic vesicles (v-SNARE) assembles with SNAP25B and syntaxin anchored in the presynaptic membrane (t-SNAREs) into an α-helical coiled-coil held together by hydrophobic interactions. Pathogenicity of the Ile67Asn mutation was confirmed by 2 measures. First, the Ca2+ triggered fusion of liposomes incorporating v-SNARE with liposomes containing t-SNAREs was hindered when t-SNAREs harbored the mutant SNAP25B moiety. Second, depolarization of bovine chromaffin cells transfected with mutant SNAP25B or with mutant plus wild-type SNAP25B markedly reduced depolarization-evoked exocytosis compared with wild-type transfected cells. Conclusion: Ile67Asn variant in SNAP25B is pathogenic because it inhibits synaptic vesicle exocytosis. We attribute the deleterious effects of the mutation to disruption of the hydrophobic α-helical coiled-coil structure of the SNARE complex by replacement of a highly hydrophobic isoleucine by a strongly hydrophilic asparagine. PMID:25381298

  5. Mutant DnaK chaperones cause ribosome assembly defects in Escherichia coli.

    PubMed Central

    Alix, J H; Guérin, M F

    1993-01-01

    To determine whether the biogenesis of ribosomes in Escherichia coli is the result of the self-assembly of their different constituents or involves the participation of additional factors, we have studied the influence of a chaperone, the product of the gene dnaK, on ribosome assembly in vivo. Using three thermosensitive (ts) mutants carrying the mutations dnaK756-ts, dnaK25-ts, and dnaK103-ts, we have observed the accumulation at nonpermissive temperature (45 degrees C) of ribosomal particles with different sedimentation constants--namely, 45S, 35S, and 25S along with the normal 30S and 50S ribosomal subunits. This is the result of a defect not in thermostability but in ribosome assembly at the nonpermissive temperature. These abnormal ribosomal particles are rescued if the mutant cells are returned to 30 degrees C. Thus, the product of the dnaK gene is implicated in ribosome biogenesis at high temperature. PMID:8105482

  6. Reversions to respiratory competence of omnipotent sup45 suppressor mutants may be caused by secondary sup45 mutations.

    PubMed

    Mironova, L N; Samsonova, M G; Zhouravleva, G A; Kulikov, V N; Soom, M J

    1995-02-01

    The molecular nature of the sup45 respiratory deficient omnipotent suppressor, and of three reversions to respiratory competence which removed the suppressor effect of the initial mutation, was examined. All reversions were caused by secondary sup45 mutations which indicates a direct connection between sup45 "respiratory" and "translational" functions. Computer analysis showed the local changes of Sup45 protein characteristics in the suppressor strain and revertants in comparison to the wild-type protein. The distribution of mutant sites in relation to evolutionary conserved, and tentatively functional, regions in the Sup45 protein is discussed.

  7. Common and new acyclovir resistant herpes simplex virus-1 mutants causing bilateral recurrent herpetic keratitis in an immunocompetent patient.

    PubMed

    Pan, Dongli; Kaye, Stephen B; Hopkins, Mark; Kirwan, Ruaidhri; Hart, Ian J; Coen, Donald M

    2014-02-01

    We investigated thymidine kinase (tk) mutants isolated during multiple episodes of recurrent bilateral acyclovir resistant herpes simplex keratitis in an immunocompetent patient. From one eye, we found a single guanine insertion, previously shown to greatly reduce TK expression, and from the other, a previously unidentified substitution, which genetic experiments confirmed confers drug resistance. The substitution, although distant from substrate binding sites, reduced thymidine phosphorylation 10-20-fold, and acyclovir phosphorylation >100-fold. This phenotype should permit reactivation from latency to cause recurrent disease. The results may have implications for the prevalence and prevention of acyclovir resistance in patients with herpes simplex keratitis.

  8. More than one mutant allele causes infantile Tay-Sachs disease in French-Canadians

    PubMed Central

    Hechtman, Peter; Kaplan, Feige; Bayleran, Janet; Boulay, Bernard; Andermann, Eva; de Braekeleer, Marc; Melançon, Serge; Lambert, Marie; Potier, Michel; Gagné, Richard; Kolodny, Edwin; Clow, Carol; Capua, Aniceta; Prevost, Claude; Scriver, Charles

    1990-01-01

    Two Tay-Sachs disease (TSD) patients of French-Canadian origin were shown by Myerowitz and Hogikyan to be homozygous for a 7.6-kb deletion mutation at the 5' end of the hexosaminidase A α-subunit gene. In order to determine whether all French-Canadian TSD patients were homozygotes for the deletion allele and to assess the geographic origins of TSD in this population, we ascertained 12 TSD families of French-Canadian origin and screened for occurrence of mutations associated with infantile TSD. DNA samples were obtained from 12 French-Canadian TSD families. Samples were analyzed using polymerase-chain-reaction (PCR) amplification followed by hybridization to allele-specific oligonucleotides (ASO) or by restriction analysis of PCR products. In some cases Southern analysis of genomic DNA was performed. Eighteen of the 22 independently segregating mutant chromosomes in this sample carried the 7.6-kb deletion mutation at the 5' end of the gene. One chromosome carried the 4-nucleotide insertion in exon 11 (a “Jewish” mutation). In this population no individuals were detected who had the substitution at the splice junction of exon 12 previously identified in Ashkenazi Jews. One chromosome carried an undescribed B1 mutation; this allele came from a parent of non-French-Canadian origin. Patients in three families carried TSD alleles different from any of the above mutations. The 5' deletion mutation clusters in persons originating in southeastern Quebec (Gaspé) and adjacent counties of northern New Brunswick. ImagesFigure 3Figure 4Figure 5Figure 6 PMID:2220821

  9. The flexibility of two tropomyosin mutants, D175N and E180G, that cause hypertrophic cardiomyopathy

    SciTech Connect

    Li, Xiaochuan; Suphamungmee, Worawit; Janco, Miro; Geeves, Michael A.; Marston, Steven B.; Fischer, Stefan; Lehman, William

    2012-08-03

    Highlights: Black-Right-Pointing-Pointer Well-known tropomyosin mutants, D175N and E180G are linked to cardiomyopathies. Black-Right-Pointing-Pointer The structural mechanics of D175N and E180G tropomyosins have been investigated. Black-Right-Pointing-Pointer D175N and E180G mutations increase both local and global tropomyosin flexibility. Black-Right-Pointing-Pointer In muscle, this increased flexibility will enhance myosin interactions on actin. Black-Right-Pointing-Pointer Extra myosin interaction can alter cardiac Ca{sup 2+}-switching, leading to dysfunction. -- Abstract: Point mutations targeting muscle thin filament proteins are the cause of a number of cardiomyopathies. In many cases, biological effects of the mutations are well-documented, whereas their structural and mechanical impact on filament assembly and regulatory function is lacking. In order to elucidate molecular defects leading to cardiac dysfunction, we have examined the structural mechanics of two tropomyosin mutants, E180G and D175N, which are associated with hypertrophic cardiomyopathy (HCM). Tropomyosin is an {alpha}-helical coiled-coil dimer which polymerizes end-to-end to create an elongated superhelix that wraps around F-actin filaments of muscle and non-muscle cells, thus modulating the binding of other actin-binding proteins. Here, we study how flexibility changes in the E180G and D175N mutants might affect tropomyosin binding and regulatory motion on F-actin. Electron microscopy and Molecular Dynamics simulations show that E180G and D175N mutations cause an increase in bending flexibility of tropomyosin both locally and globally. This excess flexibility is likely to increase accessibility of the myosin-binding sites on F-actin, thus destabilizing the low-Ca{sup 2+} relaxed-state of cardiac muscle. The resulting imbalance in the on-off switching mechanism of the mutants will shift the regulatory equilibrium towards Ca{sup 2+}-activation of cardiac muscle, as is observed in affected

  10. A pure chloride channel mutant of CLC-5 causes Dent's disease via insufficient V-ATPase activation.

    PubMed

    Satoh, Nobuhiko; Yamada, Hideomi; Yamazaki, Osamu; Suzuki, Masashi; Nakamura, Motonobu; Suzuki, Atsushi; Ashida, Akira; Yamamoto, Daisuke; Kaku, Yoshitsugu; Sekine, Takashi; Seki, George; Horita, Shoko

    2016-07-01

    Dent's disease is characterized by defective endocytosis in renal proximal tubules (PTs) and caused by mutations in the 2Cl(-)/H(+) exchanger, CLC-5. However, the pathological role of endosomal acidification in endocytosis has recently come into question. To clarify the mechanism of pathogenesis for Dent's disease, we examined the effects of a novel gating glutamate mutation, E211Q, on CLC-5 functions and endosomal acidification. In Xenopus oocytes, wild-type (WT) CLC-5 showed outward-rectifying currents that were inhibited by extracellular acidosis, but E211Q and an artificial pure Cl(-) channel mutant, E211A, showed linear currents that were insensitive to extracellular acidosis. Moreover, depolarizing pulse trains induced a robust reduction in the surface pH of oocytes expressing WT CLC-5 but not E211Q or E211A, indicating that the E211Q mutant functions as a pure Cl(-) channel similar to E211A. In HEK293 cells, E211A and E211Q stimulated endosomal acidification and hypotonicity-inducible vacuolar-type H(+)-ATPase (V-ATPase) activation at the plasma membrane. However, the stimulatory effects of these mutants were reduced compared with WT CLC-5. Furthermore, gene silencing experiments confirmed the functional coupling between V-ATPase and CLC-5 at the plasma membrane of isolated mouse PTs. These results reveal for the first time that the conversion of CLC-5 from a 2Cl(-)/H(+) exchanger into a Cl(-) channel induces Dent's disease in humans. In addition, defective endosomal acidification as a result of insufficient V-ATPase activation may still be important in the pathogenesis of Dent's disease.

  11. Podocyte-Specific Overexpression of Wild Type or Mutant Trpc6 in Mice Is Sufficient to Cause Glomerular Disease

    PubMed Central

    Kairath, Pamela; Carmona-Mora, Paulina; Molina, Jessica; Carpio, J. Daniel; Ruiz, Phillip; Mezzano, Sergio A.; Li, Jing; Wei, Changli; Reiser, Jochen; Young, Juan I.; Walz, Katherina

    2010-01-01

    Mutations in the TRPC6 calcium channel (Transient receptor potential channel 6) gene have been associated with familiar forms of Focal and Segmental Glomerulosclerosis (FSGS) affecting children and adults. In addition, acquired glomerular diseases are associated with increased expression levels of TRPC6. However, the exact role of TRPC6 in the pathogenesis of FSGS remains to be elucidated. In this work we describe the generation and phenotypic characterization of three different transgenic mouse lines with podocyte-specific overexpression of the wild type or any of two mutant forms of Trpc6 (P111Q and E896K) previously related to FSGS. Consistent with the human phenotype a non-nephrotic range of albuminuria was detectable in almost all transgenic lines. The histological analysis demonstrated that the transgenic mice developed a kidney disease similar to human FSGS. Differences of 2–3 folds in the presence of glomerular lesions were found between the non transgenic and transgenic mice expressing Trpc6 in its wild type or mutant forms specifically in podocytes. Electron microscopy of glomerulus from transgenic mice showed extensive podocyte foot process effacement. We conclude that overexpression of Trpc6 (wild type or mutated) in podocytes is sufficient to cause a kidney disease consistent with FSGS. Our results contribute to reinforce the central role of podocytes in the etiology of FSGS. These mice constitute an important new model in which to study future therapies and outcomes of this complex disease. PMID:20877463

  12. Biochemical defects of mutant nudel alleles causing early developmental arrest or dorsalization of the Drosophila embryo.

    PubMed Central

    LeMosy, E K; Leclerc, C L; Hashimoto, C

    2000-01-01

    The nudel gene of Drosophila is maternally required both for structural integrity of the egg and for dorsoventral patterning of the embryo. It encodes a structurally modular protein that is secreted by ovarian follicle cells. Genetic and molecular studies have suggested that the Nudel protein is also functionally modular, with a serine protease domain that is specifically required for ventral development. Here we describe biochemical and immunolocalization studies that provide insight into the molecular basis for the distinct phenotypes produced by nudel mutations and for the interactions between these alleles. Mutations causing loss of embryonic dorsoventral polarity result in a failure to activate the protease domain of Nudel. Our analyses support previous findings that catalytic activity of the protease domain is required for dorsoventral patterning and that the Nudel protease is auto-activated and reveal an important role for a region adjacent to the protease domain in Nudel protease function. Mutations causing egg fragility and early embryonic arrest result in a significant decrease in extracellular Nudel protein, due to defects in post-translational processing, stability, or secretion. On the basis of these and other studies of serine proteases, we suggest potential mechanisms for the complementary and antagonistic interactions between the nudel alleles. PMID:10628985

  13. A Carboxyl Ester Lipase (CEL) Mutant Causes Chronic Pancreatitis by Forming Intracellular Aggregates That Activate Apoptosis.

    PubMed

    Xiao, Xunjun; Jones, Gabrielle; Sevilla, Wednesday A; Stolz, Donna B; Magee, Kelsey E; Haughney, Margaret; Mukherjee, Amitava; Wang, Yan; Lowe, Mark E

    2016-10-28

    Patients with chronic pancreatitis (CP) frequently have genetic risk factors for disease. Many of the identified genes have been connected to trypsinogen activation or trypsin inactivation. The description of CP in patients with mutations in the variable number of tandem repeat (VNTR) domain of carboxyl ester lipase (CEL) presents an opportunity to study the pathogenesis of CP independently of trypsin pathways. We tested the hypothesis that a deletion and frameshift mutation (C563fsX673) in the CEL VNTR causes CP through proteotoxic gain-of-function activation of maladaptive cell signaling pathways including cell death pathways. HEK293 or AR42J cells were transfected with constructs expressing CEL with 14 repeats in the VNTR (CEL14R) or C563fsX673 CEL (CEL maturity onset diabetes of youth with a deletion mutation in the VNTR (MODY)). In both cell types, CEL MODY formed intracellular aggregates. Secretion of CEL MODY was decreased compared with that of CEL14R. Expression of CEL MODY increased endoplasmic reticulum stress, activated the unfolded protein response, and caused cell death by apoptosis. Our results demonstrate that disorders of protein homeostasis can lead to CP and suggest that novel therapies to decrease the intracellular accumulation of misfolded protein may be successful in some patients with CP.

  14. A missense mutant myostatin causes hyperplasia without hypertrophy in the mouse muscle.

    PubMed

    Nishi, Masumi; Yasue, Akihiro; Nishimatu, Shinichirou; Nohno, Tsutomu; Yamaoka, Takashi; Itakura, Mitsuo; Moriyama, Keiji; Ohuchi, Hideyo; Noji, Sumihare

    2002-04-26

    Myostatin, which is a member of the TGF-beta superfamily, is a negative regulator of skeletal muscle formation. Double-muscled Piedmontese cattle have a C313Y mutation in myostatin and show increased skeletal muscle mass which resulted from an increase of myofiber number (hyperplasia) without that of myofiber size (hypertrophy). To examine whether this mutation in myostatin gene affects muscle development in a dominant negative manner, we generated transgenic mice overexpressing the mutated gene. The transgenic mice exhibited dramatic increases in the skeletal muscle mass resulting from hyperplasia without hypertrophy. In contrast, it has been reported that a myostatin mutated at its cleavage site produces hypertrophy without hyperplasia in the muscle. Thus, these results suggest that (1) the myostatin containing the missense mutation exhibits a dominant negative activity and that (2) there are two types in the dominant negative form of myostatin, causing either hypertrophy or hyperplasia.

  15. A thermolabile aldolase A mutant causes fever-induced recurrent rhabdomyolysis without hemolytic anemia.

    PubMed

    Mamoune, Asmaa; Bahuau, Michel; Hamel, Yamina; Serre, Valérie; Pelosi, Michele; Habarou, Florence; Nguyen Morel, Marie-Ange; Boisson, Bertrand; Vergnaud, Sabrina; Viou, Mai Thao; Nonnenmacher, Luc; Piraud, Monique; Nusbaum, Patrick; Vamecq, Joseph; Romero, Norma; Ottolenghi, Chris; Casanova, Jean-Laurent; de Lonlay, Pascale

    2014-11-01

    Aldolase A deficiency has been reported as a rare cause of hemolytic anemia occasionally associated with myopathy. We identified a deleterious homozygous mutation in the ALDOA gene in 3 siblings with episodic rhabdomyolysis without hemolytic anemia. Myoglobinuria was always triggered by febrile illnesses. We show that the underlying mechanism involves an exacerbation of aldolase A deficiency at high temperatures that affected myoblasts but not erythrocytes. The aldolase A deficiency was rescued by arginine supplementation in vitro but not by glycerol, betaine or benzylhydantoin, three other known chaperones, suggesting that arginine-mediated rescue operated by a mechanism other than protein chaperoning. Lipid droplets accumulated in patient myoblasts relative to control and this was increased by cytokines, and reduced by dexamethasone. Our results expand the clinical spectrum of aldolase A deficiency to isolated temperature-dependent rhabdomyolysis, and suggest that thermolability may be tissue specific. We also propose a treatment for this severe disease.

  16. A Thermolabile Aldolase A Mutant Causes Fever-Induced Recurrent Rhabdomyolysis without Hemolytic Anemia

    PubMed Central

    Mamoune, Asmaa; Bahuau, Michel; Hamel, Yamina; Serre, Valérie; Pelosi, Michele; Habarou, Florence; Nguyen Morel, Marie-Ange; Boisson, Bertrand; Vergnaud, Sabrina; Viou, Mai Thao; Nonnenmacher, Luc; Piraud, Monique; Nusbaum, Patrick; Vamecq, Joseph; Romero, Norma; Ottolenghi, Chris; Casanova, Jean-Laurent; de Lonlay, Pascale

    2014-01-01

    Aldolase A deficiency has been reported as a rare cause of hemolytic anemia occasionally associated with myopathy. We identified a deleterious homozygous mutation in the ALDOA gene in 3 siblings with episodic rhabdomyolysis without hemolytic anemia. Myoglobinuria was always triggered by febrile illnesses. We show that the underlying mechanism involves an exacerbation of aldolase A deficiency at high temperatures that affected myoblasts but not erythrocytes. The aldolase A deficiency was rescued by arginine supplementation in vitro but not by glycerol, betaine or benzylhydantoin, three other known chaperones, suggesting that arginine-mediated rescue operated by a mechanism other than protein chaperoning. Lipid droplets accumulated in patient myoblasts relative to control and this was increased by cytokines, and reduced by dexamethasone. Our results expand the clinical spectrum of aldolase A deficiency to isolated temperature-dependent rhabdomyolysis, and suggest that thermolability may be tissue specific. We also propose a treatment for this severe disease. PMID:25392908

  17. Temperature Sensitivity Caused by Mutant Release Factor 1 Is Suppressed by Mutations That Affect 16S rRNA Maturation

    PubMed Central

    Kaczanowska, Magdalena; Rydén-Aulin, Monica

    2004-01-01

    To study the effect of slow termination on the protein synthesizing machinery, we isolated suppressors to a temperature-sensitive release factor 1 (RF1). Of 26 independent clones, five complementation groups have been identified, two of which are presented here. The first mutation disrupts a base pair in the transcription terminator stem for the rplM-rpsI operon, which encodes ribosomal proteins L13 and S9. We have found that this leads to readthrough of the terminator and that lower levels of transcript (compared to the results seen with the wild type) are found in the cell. This probably leads to decreased expression of the two proteins. The second mutation is a small deletion of the yrdC open reading frame start site, and it is not likely that the protein is expressed. Both mutant strains show an increased accumulation of 17S rRNA (immature 16S rRNA). Maturation of 16S rRNA is dependent on proper assembly of the ribosomal proteins, a process that is disturbed when proteins are missing. The function of the YrdC protein is not known, but it is able to bind to double-stranded RNA; therefore, we suggest that it is an assembly factor important for 30S subunit biogenesis. On the basis of our findings, we propose that lesser amounts of S9 or a lack of YrdC causes the maturation defect. We have shown that as a consequence of the maturation defect, fewer 70S ribosomes and polysomes are formed. This and other results suggest that it is the lowered concentration of functional ribosomes that suppresses the temperature sensitivity caused by the mutant RF1. PMID:15126466

  18. Effect of trehalose on the properties of mutant {gamma}PKC, which causes spinocerebellar ataxia type 14, in neuronal cell lines and cultured Purkinje cells.

    PubMed

    Seki, Takahiro; Abe-Seki, Nana; Kikawada, Takahiro; Takahashi, Hideyuki; Yamamoto, Kazuhiro; Adachi, Naoko; Tanaka, Shigeru; Hide, Izumi; Saito, Naoaki; Sakai, Norio

    2010-10-22

    Several missense mutations in the protein kinase Cγ (γPKC) gene have been found to cause spinocerebellar ataxia type 14 (SCA14), an autosomal dominant neurodegenerative disease. We previously demonstrated that the mutant γPKC found in SCA14 is susceptible to aggregation, which induces apoptotic cell death. The disaccharide trehalose has been reported to inhibit aggregate formation and to alleviate symptoms in cellular and animal models of Huntington disease, Alzheimer disease, and prion disease. Here, we show that trehalose can be incorporated into SH-SY5Y cells and reduces the aggregation of mutant γPKC-GFP, thereby inhibiting apoptotic cell death in SH-SY5Y cells and primary cultured Purkinje cells (PCs). Trehalose acts by directly stabilizing the conformation of mutant γPKC without affecting protein turnover. Trehalose was also found to alleviate the improper development of dendrites in PCs expressing mutant γPKC-GFP without aggregates but not in PCs with aggregates. In PCs without aggregates, trehalose improves the mobility and translocation of mutant γPKC-GFP, probably by inhibiting oligomerization and thereby alleviating the improper development of dendrites. These results suggest that trehalose counteracts various cellular dysfunctions that are triggered by mutant γPKC in both neuronal cell lines and primary cultured PCs by inhibiting oligomerization and aggregation of mutant γPKC.

  19. Mutant HSPB1 overexpression in neurons is sufficient to cause age-related motor neuronopathy in mice

    PubMed Central

    Srivastava, Amit K.; Renusch, Samantha R.; Naiman, Nicole E.; Gu, Shuping; Sneh, Amita; Arnold, W. David; Sahenk, Zarife; Kolb, Stephen J.

    2012-01-01

    The small heat shock protein HSPB1 is a multifunctional, α-crystallin-based protein that has been shown to be neuroprotective in animal models of motor neuron disease and peripheral nerve injury. Missense mutations in HSPB1 result in axonal Charcot-Marie-Tooth disease with minimal sensory involvement (CMT2F) and distal hereditary motor neuropathy type 2 (dHMN-II). These disorders are characterized by a selective loss of motor axons in peripheral nerve resulting in distal muscle weakness and often severe disability. To investigate the pathogenic mechanisms of HSPB1 mutations in motor neurons in vivo, we have developed and characterized transgenic PrP-HSPB1 and PrP-HSPB1(R136W) mice. These mice express the human HSPB1 protein throughout the nervous system including in axons of peripheral nerve. Although both mouse strains lacked obvious motor deficits, the PrP-HSPB1(R136W) mice developed an age-dependent motor axonopathy. Mutant mice showed axonal pathology in spinal cord and peripheral nerve with evidence of impaired neurofilament cytoskeleton, associated with organelle accumulation. Accompanying these findings, increases in the number of Schmidt-Lanterman incisures, as evidence of impaired axon-Schwann cell interactions, were present. These observations suggest that overexpression of HSPB1(R136W) in neurons is sufficient to cause pathological and electrophysiological changes in mice that are seen in patients with hereditary motor neuropathy. PMID:22521462

  20. Transgenic expression of the N525S-tuberin variant in Tsc2 mutant (Eker) rats causes dominant embryonic lethality

    PubMed Central

    Shiono, Masatoshi; Kobayashi, Toshiyuki; Takahashi, Riichi; Ueda, Masatsugu; Ishioka, Chikashi; Hino, Okio

    2014-01-01

    The Tsc2 product, tuberin, negatively regulates the mTOR pathway. We have exploited the Eker (Tsc2-mutant) rat system to analyse various Tsc2 mutations. Here, we focus on the N525S-Tsc2 variant (NSM), which is known to cause distinct symptoms in patients even though normal suppression of mTOR is observed. Unexpectedly, we were repeatedly unable to generate viable rats carrying the NSM transgene. Genotypic analysis revealed that most of the embryos carrying the transgene died around embryonic day after 14.5—similar to the stage of lethality observed for Eker homozygotes. Thus, the NSM transgene appeared to have a dominant lethal effect in our rat model. Further, no significant differences were observed for various signal transduction molecules in transiently expressed NSM cells compared to WT. These results indicate that a non-mTOR pathway, critical for embryogenesis, is being regulated by tuberin, providing a link between tuberin expression and the severity of Tsc2 mutation-related pathogenesis. PMID:25088526

  1. Cell surface expression of the epithelial Na channel and a mutant causing Liddle syndrome: a quantitative approach.

    PubMed

    Firsov, D; Schild, L; Gautschi, I; Mérillat, A M; Schneeberger, E; Rossier, B C

    1996-12-24

    The epithelial amiloride-sensitive sodium channel (ENaC) controls transepithelial Na+ movement in Na(+)-transporting epithelia and is associated with Liddle syndrome, an autosomal dominant form of salt-sensitive hypertension. Detailed analysis of ENaC channel properties and the functional consequences of mutations causing Liddle syndrome has been, so far, limited by lack of a method allowing specific and quantitative detection of cell-surface-expressed ENaC. We have developed a quantitative assay based on the binding of 125I-labeled M2 anti-FLAG monoclonal antibody (M2Ab*) directed against a FLAG reporter epitope introduced in the extracellular loop of each of the alpha, beta, and gamma ENaC subunits. Insertion of the FLAG epitope into ENaC sequences did not change its functional and pharmacological properties. The binding specificity and affinity (Kd = 3 nM) allowed us to correlate in individual Xenopus oocytes the macroscopic amiloride-sensitive sodium current (INa) with the number of ENaC wild-type and mutant subunits expressed at the cell surface. These experiments demonstrate that: (i) only heteromultimeric channels made of alpha, beta, and gamma ENaC subunits are maximally and efficiently expressed at the cell surface; (ii) the overall ENaC open probability is one order of magnitude lower than previously observed in single-channel recordings; (iii) the mutation causing Liddle syndrome (beta R564stop) enhances channel activity by two mechanisms, i.e., by increasing ENaC cell surface expression and by changing channel open probability. This quantitative approach provides new insights on the molecular mechanisms underlying one form of salt-sensitive hypertension.

  2. Mutant GlialCAM Causes Megalencephalic Leukoencephalopathy with Subcortical Cysts, Benign Familial Macrocephaly, and Macrocephaly with Retardation and Autism

    PubMed Central

    López-Hernández, Tania; Ridder, Margreet C.; Montolio, Marisol; Capdevila-Nortes, Xavier; Polder, Emiel; Sirisi, Sònia; Duarri, Anna; Schulte, Uwe; Fakler, Bernd; Nunes, Virginia; Scheper, Gert C.; Martínez, Albert; Estévez, Raúl; van der Knaap, Marjo S.

    2011-01-01

    Megalencephalic leukoencephalopathy with subcortical cysts (MLC) is a leukodystrophy characterized by early-onset macrocephaly and delayed-onset neurological deterioration. Recessive MLC1 mutations are observed in 75% of patients with MLC. Genetic-linkage studies failed to identify another gene. We recently showed that some patients without MLC1 mutations display the classical phenotype; others improve or become normal but retain macrocephaly. To find another MLC-related gene, we used quantitative proteomic analysis of affinity-purified MLC1 as an alternative approach and found that GlialCAM, an IgG-like cell adhesion molecule that is also called HepaCAM and is encoded by HEPACAM, is a direct MLC1-binding partner. Analysis of 40 MLC patients without MLC1 mutations revealed multiple different HEPACAM mutations. Ten patients with the classical, deteriorating phenotype had two mutations, and 18 patients with the improving phenotype had one mutation. Most parents with a single mutation had macrocephaly, indicating dominant inheritance. In some families with dominant HEPACAM mutations, the clinical picture and magnetic resonance imaging normalized, indicating that HEPACAM mutations can cause benign familial macrocephaly. In other families with dominant HEPACAM mutations, patients had macrocephaly and mental retardation with or without autism. Further experiments demonstrated that GlialCAM and MLC1 both localize in axons and colocalize in junctions between astrocytes. GlialCAM is additionally located in myelin. Mutant GlialCAM disrupts the localization of MLC1-GlialCAM complexes in astrocytic junctions in a manner reflecting the mode of inheritance. In conclusion, GlialCAM is required for proper localization of MLC1. HEPACAM is the second gene found to be mutated in MLC. Dominant HEPACAM mutations can cause either macrocephaly and mental retardation with or without autism or benign familial macrocephaly. PMID:21419380

  3. JAK3 mutants transform hematopoietic cells through JAK1 activation, causing T-cell acute lymphoblastic leukemia in a mouse model.

    PubMed

    Degryse, Sandrine; de Bock, Charles E; Cox, Luk; Demeyer, Sofie; Gielen, Olga; Mentens, Nicole; Jacobs, Kris; Geerdens, Ellen; Gianfelici, Valentina; Hulselmans, Gert; Fiers, Mark; Aerts, Stein; Meijerink, Jules P; Tousseyn, Thomas; Cools, Jan

    2014-11-13

    JAK3 is a tyrosine kinase that associates with the common γ chain of cytokine receptors and is recurrently mutated in T-cell acute lymphoblastic leukemia (T-ALL). We tested the transforming properties of JAK3 pseudokinase and kinase domain mutants using in vitro and in vivo assays. Most, but not all, JAK3 mutants transformed cytokine-dependent Ba/F3 or MOHITO cell lines to cytokine-independent proliferation. JAK3 pseudokinase mutants were dependent on Jak1 kinase activity for cellular transformation, whereas the JAK3 kinase domain mutant could transform cells in a Jak1 kinase-independent manner. Reconstitution of the IL7 receptor signaling complex in 293T cells showed that JAK3 mutants required receptor binding to mediate downstream STAT5 phosphorylation. Mice transplanted with bone marrow progenitor cells expressing JAK3 mutants developed a long-latency transplantable T-ALL-like disease, characterized by an accumulation of immature CD8(+) T cells. In vivo treatment of leukemic mice with the JAK3 selective inhibitor tofacitinib reduced the white blood cell count and caused leukemic cell apoptosis. Our data show that JAK3 mutations are drivers of T-ALL and require the cytokine receptor complex for transformation. These results warrant further investigation of JAK1/JAK3 inhibitors for the treatment of T-ALL.

  4. Special AT-rich Binding Protein-2 (SATB2) Differentially Affects Disease-causing p63 Mutant Proteins*

    PubMed Central

    Chung, Jacky; Grant, R. Ian; Kaplan, David R.; Irwin, Meredith S.

    2011-01-01

    p63, a p53 family member, is critical for proper skin and limb development and directly regulates gene expression in the ectoderm. Mice lacking p63 exhibit skin and craniofacial defects including cleft palate. In humans p63 mutations are associated with several distinct developmental syndromes. p63 sterile-α-motif domain, AEC (ankyloblepharon-ectodermal dysplasia-clefting)-associated mutations are associated with a high prevalence of orofacial clefting disorders, which are less common in EEC (ectrodactyly-ectodermal dysplasia-clefting) patients with DNA binding domain p63 mutations. However, the mechanisms by which these mutations differentially influence p63 function remain unclear, and interactions with other proteins implicated in craniofacial development have not been identified. Here, we show that AEC p63 mutations affect the ability of the p63 protein to interact with special AT-rich binding protein-2 (SATB2), which has recently also been implicated in the development of cleft palate. p63 and SATB2 are co-expressed early in development in the ectoderm of the first and second branchial arches, two essential sites where signaling is required for craniofacial patterning. SATB2 attenuates p63-mediated gene expression of perp (p53 apoptosis effector related to PMP-22), a critical downstream target gene during development, and specifically decreases p63 perp promoter binding. Interestingly, AEC but not EEC p63 mutations affect the ability of p63 to interact with SATB2 and the inhibitory effects of SATB2 on p63 transactivation of perp are most pronounced for AEC-associated p63 mutations. Our findings reveal a novel gain-of-function property of AEC-causing p63 mutations and identify SATB2 as the first p63 binding partner that differentially influences AEC and EEC p63 mutant proteins. PMID:21965674

  5. Identification of p38 MAPK and JNK as new targets for correction of Wilson disease‐causing ATP7B mutants

    PubMed Central

    Chesi, Giancarlo; Hegde, Ramanath N.; Iacobacci, Simona; Concilli, Mafalda; Parashuraman, Seetharaman; Festa, Beatrice Paola; Polishchuk, Elena V.; Di Tullio, Giuseppe; Carissimo, Annamaria; Montefusco, Sandro; Canetti, Diana; Monti, Maria; Amoresano, Angela; Pucci, Piero; van de Sluis, Bart; Lutsenko, Svetlana

    2016-01-01

    Wilson disease (WD) is an autosomal recessive disorder that is caused by the toxic accumulation of copper (Cu) in the liver. The ATP7B gene, which is mutated in WD, encodes a multitransmembrane domain adenosine triphosphatase that traffics from the trans‐Golgi network to the canalicular area of hepatocytes, where it facilitates excretion of excess Cu into the bile. Several ATP7B mutations, including H1069Q and R778L that are two of the most frequent variants, result in protein products, which, although still functional, remain in the endoplasmic reticulum. Thus, they fail to reach Cu excretion sites, resulting in the toxic buildup of Cu in the liver of WD patients. Therefore, correcting the location of these mutants by leading them to the appropriate functional sites in the cell should restore Cu excretion and would be beneficial to help large cohorts of WD patients. However, molecular targets for correction of endoplasmic reticulum‐retained ATP7B mutants remain elusive. Here, we show that expression of the most frequent ATP7B mutant, H1069Q, activates p38 and c‐Jun N‐terminal kinase signaling pathways, which favor the rapid degradation of the mutant. Suppression of these pathways with RNA interference or specific chemical inhibitors results in the substantial rescue of ATP7BH1069Q (as well as that of several other WD‐causing mutants) from the endoplasmic reticulum to the trans‐Golgi network compartment, in recovery of its Cu‐dependent trafficking, and in reduction of intracellular Cu levels. Conclusion: Our findings indicate p38 and c‐Jun N‐terminal kinase as intriguing targets for correction of WD‐causing mutants and, hence, as potential candidates, which could be evaluated for the development of novel therapeutic strategies to combat WD. (Hepatology 2016;63:1842‐1859) PMID:26660341

  6. Unraveling of the E-helices and Disruption of 4-Fold Pores Are Associated with Iron Mishandling in a Mutant Ferritin Causing Neurodegeneration

    SciTech Connect

    Baraibar, Martin A.; Muhoberac, Barry B.; Garringer, Holly J.; Hurley, Thomas D.; Vidal, Ruben

    2010-03-12

    Mutations in the coding sequence of the ferritin light chain (FTL) gene cause a neurodegenerative disease known as neuroferritinopathy or hereditary ferritinopathy, which is characterized by the presence of intracellular inclusion bodies containing the mutant FTL polypeptide and by abnormal accumulation of iron in the brain. Here, we describe the x-ray crystallographic structure and report functional studies of ferritin homopolymers formed from the mutant FTL polypeptide p.Phe167SerfsX26, which has a C terminus that is altered in amino acid sequence and length. The structure was determined and refined to 2.85 {angstrom} resolution and was very similar to the wild type between residues Ile-5 and Arg-154. However, instead of the E-helices normally present in wild type ferritin, the C-terminal sequences of all 24 mutant subunits showed substantial amounts of disorder, leading to multiple C-terminal polypeptide conformations and a large disruption of the normally tiny 4-fold axis pores. Functional studies underscored the importance of the mutant C-terminal sequence in iron-induced precipitation and revealed iron mishandling by soluble mutant FTL homopolymers in that only wild type incorporated iron when in direct competition in solution with mutant ferritin. Even without competition, the amount of iron incorporation over the first few minutes differed severalfold. Our data suggest that disruption at the 4-fold pores may lead to direct iron mishandling through attenuated iron incorporation by the soluble form of mutant ferritin and that the disordered C-terminal polypeptides may play a major role in iron-induced precipitation and formation of ferritin inclusion bodies in hereditary ferritinopathy.

  7. Analysis of the rice mutant dwarf and gladius leaf 1. Aberrant katanin-mediated microtubule organization causes up-regulation of gibberellin biosynthetic genes independently of gibberellin signaling.

    PubMed

    Komorisono, Masahiko; Ueguchi-Tanaka, Miyako; Aichi, Ikuko; Hasegawa, Yasuko; Ashikari, Motoyuki; Kitano, Hidemi; Matsuoka, Makoto; Sazuka, Takashi

    2005-08-01

    Molecular genetic studies of plant dwarf mutants have indicated that gibberellin (GA) and brassinosteroid (BR) are two major factors that determine plant height; dwarf mutants that are caused by other defects are relatively rare, especially in monocot species. Here, we report a rice (Oryza sativa) dwarf mutant, dwarf and gladius leaf 1 (dgl1), which exhibits only minimal response to GA and BR. In addition to the dwarf phenotype, dgl1 produces leaves with abnormally rounded tip regions. Positional cloning of DGL1 revealed that it encodes a 60-kD microtubule-severing katanin-like protein. The protein was found to be important in cell elongation and division, based on the observed cell phenotypes. GA biosynthetic genes are up-regulated in dgl1, but the expression of BR biosynthetic genes is not enhanced. The enhanced expression of GA biosynthetic genes in dgl1 is not caused by inappropriate GA signaling because the expression of these genes was repressed by GA3 treatment, and degradation of the rice DELLA protein SLR1 was triggered by GA3 in this mutant. Instead, aberrant microtubule organization caused by the loss of the microtubule-severing function of DGL1 may result in enhanced expression of GA biosynthetic genes in that enhanced expression was also observed in a BR-deficient mutant with aberrant microtubule organization. These results suggest that the function of DGL1 is important for cell and organ elongation in rice, and aberrant DGL1-mediated microtubule organization causes up-regulation of gibberellin biosynthetic genes independently of gibberellin signaling.

  8. Recovery of Nonpathogenic Mutant Bacteria from Tumors Caused by Several Agrobacterium tumefaciens Strains: a Frequent Event?▿

    PubMed Central

    Llop, Pablo; Murillo, Jesús; Lastra, Beatriz; López, María M.

    2009-01-01

    We have evaluated the interaction that bacterial genotypes and plant hosts have with the loss of pathogenicity in tumors, using seven Agrobacterium tumefaciens strains inoculated on 12 herbaceous and woody hosts. We performed a screening of the agrobacteria present inside the tumors, looking for nonpathogenic strains, and found a high variability of those strains in this niche. To verify the origin of the putative nonpathogenic mutant bacteria, we applied an efficient, reproducible, and specific randomly amplified polymorphic DNA analysis method. In contrast with previous studies, we recovered a very small percentage (0.01%) of nonpathogenic strains that can be considered true mutants. Of 5,419 agrobacterial isolates examined, 662 were nonpathogenic in tomato, although only 7 (from pepper and tomato tumors induced by two A. tumefaciens strains) could be considered to derive from the inoculated strain. Six mutants were affected in the transferred DNA (T-DNA) region; one of them contained IS426 inserted into the iaaM gene, whereas the whole T-DNA region was apparently deleted in three other mutants, and the virulence of the remaining two mutants was fully restored with the T-DNA genes as well. The plasmid profile was altered in six of the mutants, with changes in the size of the Ti plasmid or other plasmids and/or the acquisition of new plasmids. Our results also suggest that the frequent occurrence of nonpathogenic clones in the tumors is probably due to the preferential growth of nonpathogenic agrobacteria, of either endophytic or environmental origin, but different from the bacterial strain inducing the tumor. PMID:19700547

  9. Mirror Movement-Like Defects in Startle Behavior of Zebrafish dcc Mutants Are Caused by Aberrant Midline Guidance of Identified Descending Hindbrain Neurons

    PubMed Central

    Jain, Roshan A.; Bell, Hannah; Lim, Amy; Chien, Chi-Bin

    2014-01-01

    Mirror movements are involuntary movements on one side of the body that occur simultaneously with intentional movements on the contralateral side. Humans with heterozygous mutations in the axon guidance receptor DCC display such mirror movements, where unilateral stimulation results in inappropriate bilateral motor output. Currently, it is unclear whether mirror movements are caused by incomplete midline crossing and reduced commissural connectivity of DCC-dependent descending pathways or by aberrant ectopic ipsilateral axonal projections of normally commissural neurons. Here, we show that in response to unilateral tactile stimuli, zebrafish dcc mutant larvae perform involuntary turns on the inappropriate body side. We show that these mirror movement-like deficits are associated with axonal guidance defects of two identified groups of commissural reticulospinal hindbrain neurons. Moreover, we demonstrate that in dcc mutants, axons of these identified neurons frequently fail to cross the midline and instead project ipsilaterally. Whereas laser ablation of these neurons in wild-type animals does not affect turning movements, their ablation in dcc mutants restores turning movements. Thus, our results demonstrate that in dcc mutants, turns on the inappropriate side of the body are caused by aberrant ipsilateral axonal projections, and suggest that aberrant ipsilateral connectivity of a very small number of descending axons is sufficient to induce incorrect movement patterns. PMID:24553931

  10. Chloroplast Dysfunction Causes Multiple Defects in Cell Cycle Progression in the Arabidopsis crumpled leaf Mutant1[C][W

    PubMed Central

    Hudik, Elodie; Yoshioka, Yasushi; Domenichini, Séverine; Bourge, Mickaël; Soubigout-Taconnat, Ludivine; Mazubert, Christelle; Yi, Dalong; Bujaldon, Sandrine; Hayashi, Hiroyuki; De Veylder, Lieven; Bergounioux, Catherine; Benhamed, Moussa; Raynaud, Cécile

    2014-01-01

    The majority of research on cell cycle regulation is focused on the nuclear events that govern the replication and segregation of the genome between the two daughter cells. However, eukaryotic cells contain several compartmentalized organelles with specialized functions, and coordination among these organelles is required for proper cell cycle progression, as evidenced by the isolation of several mutants in which both organelle function and overall plant development were affected. To investigate how chloroplast dysfunction affects the cell cycle, we analyzed the crumpled leaf (crl) mutant of Arabidopsis (Arabidopsis thaliana), which is deficient for a chloroplastic protein and displays particularly severe developmental defects. In the crl mutant, we reveal that cell cycle regulation is altered drastically and that meristematic cells prematurely enter differentiation, leading to reduced plant stature and early endoreduplication in the leaves. This response is due to the repression of several key cell cycle regulators as well as constitutive activation of stress-response genes, among them the cell cycle inhibitor SIAMESE-RELATED5. One unique feature of the crl mutant is that it produces aplastidic cells in several organs, including the root tip. By investigating the consequence of the absence of plastids on cell cycle progression, we showed that nuclear DNA replication occurs in aplastidic cells in the root tip, which opens future research prospects regarding the dialogue between plastids and the nucleus during cell cycle regulation in higher plants. PMID:25037213

  11. Wing Defects in Drosophila xenicid Mutant Clones Are Caused by C-Terminal Deletion of Additional Sex Combs (Asx)

    PubMed Central

    Bischoff, Kara; Ballew, Anna C.; Simon, Michael A.; O'Reilly, Alana M.

    2009-01-01

    Background The coordinated action of genes that control patterning, cell fate determination, cell size, and cell adhesion is required for proper wing formation in Drosophila. Defects in any of these basic processes can lead to wing aberrations, including blisters. The xenicid mutation was originally identified in a screen designed to uncover regulators of adhesion between wing surfaces [1]. Principal Findings Here, we demonstrate that expression of the βPS integrin or the patterning protein Engrailed are not affected in developing wing imaginal discs in xenicid mutants. Instead, expression of the homeotic protein Ultrabithorax (Ubx) is strongly increased in xenicid mutant cells. Conclusion Our results suggest that upregulation of Ubx transforms cells from a wing blade fate to a haltere fate, and that the presence of haltere cells within the wing blade is the primary defect leading to the adult wing phenotypes observed. PMID:19956620

  12. Structural Mutants of the Spindle Pole Body Cause Distinct Alteration of Cytoplasmic Microtubules and Nuclear Dynamics in Multinucleated Hyphae

    PubMed Central

    Lang, Claudia; Grava, Sandrine; Finlayson, Mark; Trimble, Rhonda; Philippsen, Peter

    2010-01-01

    In the multinucleate fungus Ashbya gossypii, cytoplasmic microtubules (cMTs) emerge from the spindle pole body outer plaque (OP) in perpendicular and tangential directions. To elucidate the role of cMTs in forward/backward movements (oscillations) and bypassing of nuclei, we constructed mutants potentially affecting cMT nucleation or stability. Hyphae lacking the OP components AgSpc72, AgNud1, AgCnm67, or the microtubule-stabilizing factor AgStu2 grew like wild- type but showed substantial alterations in the number, length, and/or nucleation sites of cMTs. These mutants differently influenced nuclear oscillation and bypassing. In Agspc72Δ, only long cMTs were observed, which emanate tangentially from reduced OPs; nuclei mainly moved with the cytoplasmic stream but some performed rapid bypassing. Agnud1Δ and Agcnm67Δ lack OPs; short and long cMTs emerged from the spindle pole body bridge/half-bridge structures, explaining nuclear oscillation and bypassing in these mutants. In Agstu2Δ only very short cMTs emanated from structurally intact OPs; all nuclei moved with the cytoplasmic stream. Therefore, long tangential cMTs promote nuclear bypassing and short cMTs are important for nuclear oscillation. Our electron microscopy ultrastructural analysis also indicated that assembly of the OP occurs in a stepwise manner, starting with AgCnm67, followed by AgNud1 and lastly AgSpc72. PMID:20053682

  13. Mutant superoxide dismutase 1 (SOD1), a cause of amyotrophic lateral sclerosis, disrupts the recruitment of SMN, the spinal muscular atrophy protein to nuclear Cajal bodies.

    PubMed

    Kariya, Shingo; Re, Diane B; Jacquier, Arnaud; Nelson, Katelyn; Przedborski, Serge; Monani, Umrao R

    2012-08-01

    Spinal muscular atrophy (SMA) and amyotrophic lateral sclerosis (ALS) are among the most common motor neuron diseases to afflict the human population. A deficiency of the survival of motor neuron (SMN) protein causes SMA and is also reported to be an exacerbating factor in the development of ALS. However, pathways linking the two diseases have yet to be defined and it is not clear precisely how the pathology of ALS is aggravated by reduced SMN or whether mutant proteins underlying familial forms of ALS interfere with SMN-related biochemical pathways to exacerbate the neurodegenerative process. In this study, we show that mutant superoxide dismutase-1 (SOD1), a cause of familial ALS, profoundly alters the sub-cellular localization of the SMN protein, preventing the formation of nuclear 'gems' by disrupting the recruitment of the protein to Cajal bodies. Overexpressing the SMN protein in mutant SOD1 mice, a model of familial ALS, alleviates this phenomenon, most likely in a cell-autonomous manner, and significantly mitigates the loss of motor neurons in the spinal cord and in culture dishes. In the mice, the onset of the neuromuscular phenotype is delayed and motor function enhanced, suggestive of a therapeutic benefit for ALS patients treated with agents that augment the SMN protein. Nevertheless, this finding is tempered by an inability to prolong survival, a limitation most likely imposed by the inexorable denervation that characterizes ALS and eventually disrupts the neuromuscular synapses even in the presence of increased SMN.

  14. High-throughput FACS-based mutant screen identifies a gain-of-function allele of the Fusarium graminearum adenylyl cyclase causing deoxynivalenol over-production.

    PubMed

    Blum, Ailisa; Benfield, Aurélie H; Stiller, Jiri; Kazan, Kemal; Batley, Jacqueline; Gardiner, Donald M

    2016-05-01

    Fusarium head blight and crown rot, caused by the fungal plant pathogen Fusarium graminearum, impose a major threat to global wheat production. During the infection, plants are contaminated with mycotoxins such as deoxynivalenol (DON), which can be toxic for humans and animals. In addition, DON is a major virulence factor during wheat infection. However, it is not fully understood how DON production is regulated in F. graminearum. In order to identify regulators of DON production, a high-throughput mutant screen using Fluorescence Activated Cell Sorting (FACS) of a mutagenised TRI5-GFP reporter strain was established and a mutant over-producing DON under repressive conditions identified. A gain-of-function mutation in the F. graminearum adenylyl cyclase (FAC1), which is a known positive regulator of DON production, was identified as the cause of this phenotype through genome sequencing and segregation analysis. Our results show that the high-throughput mutant screening procedure developed here can be applied for identification of fungal proteins involved in diverse processes.

  15. Analyzing and Quantifying the Gain-of-Function Enhancement of IP3 Receptor Gating by Familial Alzheimer’s Disease-Causing Mutants in Presenilins

    PubMed Central

    Mak, Don-On Daniel; Cheung, King-Ho; Toglia, Patrick; Foskett, J. Kevin; Ullah, Ghanim

    2015-01-01

    Familial Alzheimer’s disease (FAD)-causing mutant presenilins (PS) interact with inositol 1,4,5-trisphosphate (IP3) receptor (IP3R) Ca2+ release channels resulting in enhanced IP3R channel gating in an amyloid beta (Aβ) production-independent manner. This gain-of-function enhancement of IP3R activity is considered to be the main reason behind the upregulation of intracellular Ca2+ signaling in the presence of optimal and suboptimal stimuli and spontaneous Ca2+ signals observed in cells expressing mutant PS. In this paper, we employed computational modeling of single IP3R channel activity records obtained under optimal Ca2+ and multiple IP3 concentrations to gain deeper insights into the enhancement of IP3R function. We found that in addition to the high occupancy of the high-activity (H) mode and the low occupancy of the low-activity (L) mode, IP3R in FAD-causing mutant PS-expressing cells exhibits significantly longer mean life-time for the H mode and shorter life-time for the L mode, leading to shorter mean close-time and hence high open probability of the channel in comparison to IP3R in cells expressing wild-type PS. The model is then used to extrapolate the behavior of the channel to a wide range of IP3 and Ca2+ concentrations and quantify the sensitivity of IP3R to its two ligands. We show that the gain-of-function enhancement is sensitive to both IP3 and Ca2+ and that very small amount of IP3 is required to stimulate IP3R channels in the presence of FAD-causing mutant PS to the same level of activity as channels in control cells stimulated by significantly higher IP3 concentrations. We further demonstrate with simulations that the relatively longer time spent by IP3R in the H mode leads to the observed higher frequency of local Ca2+ signals, which can account for the more frequent global Ca2+ signals observed, while the enhanced activity of the channel at extremely low ligand concentrations will lead to spontaneous Ca2+ signals in cells expressing FAD-causing

  16. Discordant measures of androgen-binding kinetics in two mutant androgen receptors causing mild or partial androgen insensitivity, respectively.

    PubMed

    Shkolny, D L; Beitel, L K; Ginsberg, J; Pekeles, G; Arbour, L; Pinsky, L; Trifiro, M A

    1999-02-01

    We have characterized two different mutations of the human androgen receptor (hAR) found in two unrelated subjects with androgen insensitivity syndrome (AIS): in one, the external genitalia were ambiguous (partial, PAIS); in the other, they were male, but small (mild, MAIS). Single base substitutions have been found in both individuals: E772A in the PAIS subject, and R871G in the MAIS patient. In COS-1 cells transfected with the E772A and R871G hARs, the apparent equilibrium dissociation constants (Kd) for mibolerone (MB) and methyltrienolone are normal. Nonetheless, the mutant hAR from the PAIS subject (E772A) has elevated nonequilibrium dissociation rate constants (k(diss)) for both androgens. In contrast, the MAIS subject's hAR (R871G) has k(diss) values that are apparently normal for MB and methyltrienolone; in addition, the R871G hAR's ability to bind MB resists thermal stress better than the hAR from the PAIS subject. The E772A and R871G hARs, therefore, confer the same pattern of discordant androgen-binding parameters in transfected COS-1 cells as observed previously in the subjects' genital skin fibroblasts. This proves their pathogenicity and correlates with the relative severity of the clinical phenotype. In COS-1 cells transfected with an androgen-responsive reporter gene, trans-activation was 50% of normal in cells containing either mutant hAR. However, mutant hAR-MB binding is unstable during prolonged incubation with MB, whereas normal hAR-MB binding increases. Thus, normal equilibrium dissociation constants alone, as determined by Scatchard analysis, may not be indicative of normal hAR function. An increased k(diss) despite a normal Kd for a given androgen suggests that it not only has increased egress from a mutant ligand-binding pocket, but also increased access to it. This hypothesis has certain implications in terms of the three-dimensional model of the ligand-binding domain of the nuclear receptor superfamily.

  17. Reduced immunogenicity of Arabidopsis hgl1 mutant N-glycans caused by altered accessibility of xylose and core fucose epitopes.

    PubMed

    Kaulfürst-Soboll, Heidi; Rips, Stephan; Koiwa, Hisashi; Kajiura, Hiroyuki; Fujiyama, Kazuhito; von Schaewen, Antje

    2011-07-01

    Arabidopsis N-glycosylation mutants with enhanced salt sensitivity show reduced immunoreactivity of complex N-glycans. Among them, hybrid glycosylation 1 (hgl1) alleles lacking Golgi α-mannosidase II are unique, because their glycoprotein N-glycans are hardly labeled by anti-complex glycan antibodies, even though they carry β1,2-xylose and α1,3-fucose epitopes. To dissect the contribution of xylose and core fucose residues to plant stress responses and immunogenic potential, we prepared Arabidopsis hgl1 xylT double and hgl1 fucTa fucTb triple mutants by crossing previously established T-DNA insertion lines and verified them by mass spectrometry analyses. Root growth assays revealed that hgl1 fucTa fucTb but not hgl1 xylT plants are more salt-sensitive than hgl1, hinting at the importance of core fucose modification and masking of xylose residues. Detailed immunoblot analyses with anti-β1,2-xylose and anti-α1,3-fucose rabbit immunoglobulin G antibodies as well as cross-reactive carbohydrate determinant-specific human immunoglobulin E antibodies (present in sera of allergy patients) showed that xylose-specific reactivity of hgl1 N-glycans is indeed reduced. Based on three-dimensional modeling of plant N-glycans, we propose that xylose residues are tilted by 30° because of untrimmed mannoses in hgl1 mutants. Glycosidase treatments of protein extracts restored immunoreactivity of hgl1 N-glycans supporting these models. Furthermore, among allergy patient sera, untrimmed mannoses persisting on the α1,6-arm of hgl1 N-glycans were inhibitory to immunoreaction with core fucoses to various degrees. In summary, incompletely trimmed glycoprotein N-glycans conformationally prevent xylose and, to lesser extent, core fucose accessibility. Thus, in addition to N-acetylglucosaminyltransferase I, Golgi α-mannosidase II emerges as a so far unrecognized target for lowering the immunogenic potential of plant-derived glycoproteins.

  18. The einkorn wheat (Triticum monococcum) mutant, maintained vegetative phase, is caused by a deletion in the VRN1 gene.

    PubMed

    Shitsukawa, Naoki; Ikari, Chihiro; Shimada, Sanae; Kitagawa, Satoshi; Sakamoto, Koichi; Saito, Hiroyuki; Ryuto, Hiromichi; Fukunishi, Nobuhisa; Abe, Tomoko; Takumi, Shigeo; Nasuda, Shuhei; Murai, Koji

    2007-04-01

    The einkorn wheat (Triticum monococcum) mutant, maintained vegetative phase (mvp), was induced by nitrogen ion-beam treatment and was identified by its inability to transit from the vegetative to reproductive phase. In our previous study, we showed that WAP1 (wheat APETALA1) is a key gene in the regulatory pathway that controls phase transition from vegetative to reproductive growth in common wheat. WAP1 is an ortholog of the VRN1 gene that is responsible for vernalization insensitivity in einkorn wheat. The mvp mutation resulted from deletion of the VRN1 coding and promoter regions, demonstrating that WAP1/VRN1 is an indispensable gene for phase transition in wheat. Expression analysis of flowering-related genes in mvp plants indicated that wheat GIGANTIA (GI), CONSTANS (CO) and SUPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1) genes either act upstream of or in a different pathway to WAP1/VRN1.

  19. A Novel Lamin A Mutant Responsible for Congenital Muscular Dystrophy Causes Distinct Abnormalities of the Cell Nucleus

    PubMed Central

    Barateau, Alice; Vadrot, Nathalie; Vicart, Patrick; Ferreiro, Ana; Mayer, Michèle; Héron, Delphine; Vigouroux, Corinne

    2017-01-01

    A-type lamins, the intermediate filament proteins participating in nuclear structure and function, are encoded by LMNA. LMNA mutations can lead to laminopathies such as lipodystrophies, premature aging syndromes (progeria) and muscular dystrophies. Here, we identified a novel heterozygous LMNA p.R388P de novo mutation in a patient with a non-previously described severe phenotype comprising congenital muscular dystrophy (L-CMD) and lipodystrophy. In culture, the patient’s skin fibroblasts entered prematurely into senescence, and some nuclei showed a lamina honeycomb pattern. C2C12 myoblasts were transfected with a construct carrying the patient’s mutation; R388P-lamin A (LA) predominantly accumulated within the nucleoplasm and was depleted at the nuclear periphery, altering the anchorage of the inner nuclear membrane protein emerin and the nucleoplasmic protein LAP2-alpha. The mutant LA triggered a frequent and severe nuclear dysmorphy that occurred independently of prelamin A processing, as well as increased histone H3K9 acetylation. Nuclear dysmorphy was not significantly improved when transfected cells were treated with drugs disrupting microtubules or actin filaments or modifying the global histone acetylation pattern. Therefore, releasing any force exerted at the nuclear envelope by the cytoskeleton or chromatin did not rescue nuclear shape, in contrast to what was previously shown in Hutchinson-Gilford progeria due to other LMNA mutations. Our results point to the specific cytotoxic effect of the R388P-lamin A mutant, which is clinically related to a rare and severe multisystemic laminopathy phenotype. PMID:28125586

  20. Immunization with a highly attenuated replication-competent herpes simplex virus type 1 mutant, HF10, protects mice from genital disease caused by herpes simplex virus type 2.

    PubMed

    Luo, Chenhong; Goshima, Fumi; Kamakura, Maki; Mutoh, Yoshifumi; Iwata, Seiko; Kimura, Hiroshi; Nishiyama, Yukihiro

    2012-01-01

    Genital herpes is an intractable disease caused mainly by herpes simplex virus (HSV) type 2 (HSV-2), and is a major concern in public health. A previous infection with HSV type 1 (HSV-1) enhances protection against primary HSV-2 infection to some extent. In this study, we evaluated the ability of HF10, a naturally occurring replication-competent HSV-1 mutant, to protect against genital infection in mice caused by HSV-2. Subcutaneous inoculation of HF10-immunized mice against lethal infection by HSV-2, and attenuated the development of genital ulcer diseases. Immunization with HF10 inhibited HSV-2 replication in the mouse vagina, reduced local inflammation, controlled emergence of neurological dysfunctions of HSV-2 infection, and increased survival. In HF10-immunized mice, we observed rapid and increased production of interferon-γ in the vagina in response to HSV-2 infection, and numerous CD4(+) and a few CD8(+) T cells localized to the infective focus. CD4(+) T cells invaded the mucosal subepithelial lamina propria. Thus, the protective effect of HF10 was related to induction of cellular immunity, mediated primarily by Th1 CD4(+) cells. These data indicate that the live attenuated HSV-1 mutant strain HF10 is a promising candidate antigen for a vaccine against genital herpes caused by HSV-2.

  1. Iron Loading-Induced Aggregation and Reduction of Iron Incorporation in Heteropolymeric Ferritin Containing a Mutant Light Chain that Causes Neurodegeneration

    PubMed Central

    Muhoberac, Barry B.; Baraibar, Martin A.; Vidal, Ruben

    2010-01-01

    Hereditary Ferritinopathy (HF) is a neurodegenerative disease characterized by intracellular ferritin inclusion bodies (IBs) and iron accumulation throughout the central nervous system. Ferritin IBs are composed of mutant ferritin light chain as well as wild type light (Wt-FTL) and heavy chain (FTH1) polypeptides. In vitro studies have shown that the mutant light chain polypeptide p.Phe167SerfsX26 (Mt-FTL) forms soluble ferritin 24-mer homopolymers having a specific structural disruption that explains its functional problems of reduced ability to incorporate iron and aggregation during iron loading. However, because ferritins are usually 24-mer heteropolymers and all three polypeptides are found in IBs, we investigated the properties of Mt-FTL/FTH1 and Mt-FTL/Wt-FTL heteropolymeric ferritins. We show here the facile assembly of Mt-FTL and FTH1 subunits into soluble ferritin heteropolymers, but their ability to incorporate iron was significantly reduced relative to Wt-FTL/FTH1 heteropolymers. In addition, Mt-FTL/FTH1 heteropolymers formed aggregates during iron loading, contrasting Wt-FTL/FTH1 heteropolymers and similar to what was seen for Mt-FTL homopolymers. The resulting precipitate contained both Mt-FTL and FTH1 polypeptides as do ferritin IBs in patients with HF. The presence of Mt-FTL subunits in Mt-FTL/Wt-FTL heteropolymers also caused iron loading-induced aggregation relative to Wt-FTL homopolymers, with the precipitate containing Mt- and Wt-FTL polypeptides again paralleling HF. Our data demonstrate that co-assembly with wild type subunits does not circumvent the functional problems caused by mutant subunits. Furthermore, the functional problems characterized here in heteropolymers that contain mutant subunits parallel those problems previously reported in homopolymers composed exclusively of mutant subunits, which strongly suggests that the structural disruption characterized previously in Mt-FTL homopolymers occurs in a similar manner and to a

  2. Caffeic Acid Phenethyl Ester Causes p21Cip1 Induction, Akt Signaling Reduction, and Growth Inhibition in PC-3 Human Prostate Cancer Cells

    PubMed Central

    Lin, Hui-Ping; Jiang, Shih Sheng; Chuu, Chih-Pin

    2012-01-01

    Caffeic acid phenethyl ester (CAPE) treatment suppressed proliferation, colony formation, and cell cycle progression in PC-3 human prostate cancer cells. CAPE decreased protein expression of cyclin D1, cyclin E, SKP2, c-Myc, Akt1, Akt2, Akt3, total Akt, mTOR, Bcl-2, Rb, as well as phosphorylation of Rb, ERK1/2, Akt, mTOR, GSK3α, GSK3β, PDK1; but increased protein expression of KLF6 and p21Cip1. Microarray analysis indicated that pathways involved in cellular movement, cell death, proliferation, and cell cycle were affected by CAPE. Co-treatment of CAPE with chemotherapeutic drugs vinblastine, paclitaxol, and estramustine indicated synergistic suppression effect. CAPE administration may serve as a potential adjuvant therapy for prostate cancer. PMID:22347457

  3. The Cataract-linked Mutant Connexin50D47A Causes Endoplasmic Reticulum Stress in Mouse Lenses.

    PubMed

    Berthoud, Viviana M; Minogue, Peter J; Lambert, Paul A; Snabb, Joseph I; Beyer, Eric C

    2016-08-19

    Mice expressing connexin50D47A (Cx50D47A) exhibit nuclear cataracts and impaired differentiation. Cx50D47A does not traffic properly, and homozygous mutant lenses show increased levels of the stress-responsive αB-crystallins. Therefore, we assessed whether expression of Cx50D47A led to endoplasmic reticulum (ER) stress in the lens in vivo Although pharmacologic induction of ER stress can be transduced by three different pathways, we found no evidence for activation of the IRE1α or ATF6 pathways in Cx50D47A-expressing lenses. In contrast, heterozygous and homozygous Cx50D47A lenses showed an increase in phosphorylated PERK immunoreactivity and in the ratio of phosphorylated to total EIF2α (2.4- and 3.3-fold, respectively) compared with wild type. Levels of ATF4 were similar in wild type and heterozygous lenses but elevated in homozygotes (391%). In both heterozygotes and homozygotes, levels of calreticulin protein were increased (184 and 262%, respectively), as was Chop mRNA (1.9- and 12.4-fold, respectively). CHOP protein was increased in homozygotes (384%). TUNEL staining was increased in Cx50D47A lenses, especially in homozygous mice. Levels of two factors that may be pro-survival, Irs2 and Trib3, were greatly increased in homozygous lenses. These results suggest that expression of Cx50D47A induces ER stress, triggering activation of the PERK-ATF4 pathway, which potentially contributes to the lens pathology and leads to increased expression of anti-apoptotic factors, allowing cell survival.

  4. Biomimicry 1: PC.

    PubMed

    Cumberland, D C; Gunn, J; Malik, N; Holt, C M

    1998-01-01

    The surface properties of stents can be modified by coating them, for example with a polymer. Phosphorylcoline (PC) is the major component of the outer layer of the cell membrane. The haemo- and biocompatibility of a PC-containing polymer is thus based on biomimicry, and has been confirmed by several experiments showing much reduced thrombogenicity of PC-coated surfaces, and porcine coronary artery implants showing no sign of adverse effect. Clinical experience with the PC-coated BiodivYsio appears favourable. The PC coating can be tailored for take up and controlled elution of various drugs for stent-based local delivery, a property which is being actively explored.

  5. SI PC104 Performance Test Report

    SciTech Connect

    Montelongo, S

    2005-12-16

    The Spectral Instruments (SI) PC104 systems associated with the SI-1000 CCD camera exhibited intermittent power problems during setup, test and operations which called for further evaluation and testing. The SI PC104 System is the interface between the SI-1000 CCD camera and its associated Diagnostic Controller (DC). As such, the SI PC104 must be a reliable, robust system capable of providing consistent performance in various configurations and operating conditions. This SI PC104 system consists of a stackable set of modules designed to meet the PC104+ Industry Standard. The SI PC104 System consists of a CPU module, SI Camera card, Media converter card, Video card and a I/O module. The root cause of power problems was identified as failing solder joints at the LEMO power connector attached to the SI Camera Card. The recommended solution was to provide power to the PC104 system via a PC104+ power supply module configured into the PC104 stack instead of thru the LEMO power connector. Test plans (2) were developed to test SI PC104 performance and identify any outstanding issues noted during extended operations. Test Plan 1 included performance and image acquisition tests. Test Plan 2 verified performance after implementing recommendations. Test Plan 2 also included verifying integrity of system files and driver installation after bootup. Each test plan was implemented to fully test against each set of problems noted. Test Plan presentations and Test Plan results are attached as appendices. Anticipated test results will show successful operation and reliable performance of the SI PC104 system receiving its power via a PC104 power supply module. A SI PC104 Usage Recommendation Memo will be sent out to the SI PC104 User Community. Recommendation memo(s) are attached as appendices.

  6. Two types of albino mutants in desert and migratory locusts are caused by gene defects in the same signaling pathway.

    PubMed

    Sugahara, Ryohei; Tanaka, Seiji; Jouraku, Akiya; Shiotsuki, Takahiro

    2017-04-15

    Albinism is caused by mutations in the genes involved in melanin production. Albino nymphs of Locusta migratoria and Schistocerca gregaria reared under crowded conditions are uniformly creamy-white in color. However, nothing is known about the molecular mechanisms underlying this phenomenon in locusts. The albino strain of L. migratoria is known to lack the dark-color-inducing neuropeptide corazonin (Crz). In this study, we report that this albino strain has a 10-base-pair deletion in the gene LmCRZ, which encodes Crz. This mutation was found to cause a frame-shift, resulting in a null mutation in Crz. On the other hand, the albino strain of S. gregaria is known to have an intact Crz. This strain was found to possess a single-nucleotide substitution in the middle of the Crz receptor-encoding gene, SgCRZR, which caused a nonsense mutation, resulting in a truncated receptor. Silencing of SgCRZR in wild-type S. gregaria nymphs greatly reduced the area and intensity of their black patterning, suggesting that the functional defect of SgCRZR likely causes the albinism. The expression level of SgCRZR in the albino S. gregaria was comparable to that in the wild type. Unlike the wild type, the albino strain of this locust did not show a phase-dependent shift in a morphometric trait controlled by Crz. From these results, we conclude that the mutations in LmCRZ and SgCRZR are responsible for the albinism in L. migratoria and S. gregaria, respectively, indicating that the two types of albinism are caused by different genetic defects in the same Crz signaling pathway.

  7. Snyder-Robinson Syndrome: Rescuing the Disease-Causing Effect of G56S mutant by Small Molecule Binding

    NASA Astrophysics Data System (ADS)

    Zhang, Zhe; Martiny, Virginie; Lagorce, David; Alexov, Emil; Miteva, Maria; Clemson University Team; Université Paris Diderot Team

    2013-03-01

    Snyder-Robinson Syndrome (SRS) is an X-linked mental retardation disorder, which is caused by defects in a particular gene coding for the spermine synthase (SMS) protein. Among the missense mutations known to be disease-causing is the G56S, which is positioned at the interface of the SMS homo-dimer. Previous computational and experimental investigations have shown that G56S mutation destabilizes the homo-dimer and thus greatly reduces the SMS enzymatic activity. In this study, we explore the possibility of mitigating the effect of G56S mutation by binding small molecules to suitable pockets around the mutation site. It is done by combined efforts of molecular dynamics simulations and in silico screening. The binding of selected molecules was calculated to fully compensate the effect of the mutation and rescue the wild type dimer affinity. This work was supported by NIH, NLM grant. No. 1R03LM009748

  8. AID induces double-strand breaks at immunoglobulin switch regions and c-MYC causing chromosomal translocations in yeast THO mutants.

    PubMed

    Ruiz, José F; Gómez-González, Belén; Aguilera, Andrés

    2011-02-01

    Transcription of the switch (S) regions of immunoglobulin genes in B cells generates stable R-loops that are targeted by Activation Induced Cytidine Deaminase (AID), triggering class switch recombination (CSR), as well as translocations with c-MYC responsible for Burkitt's lymphomas. In Saccharomyces cerevisiae, stable R-loops are formed co-transcriptionally in mutants of THO, a conserved nuclear complex involved in mRNP biogenesis. Such R-loops trigger genome instability and facilitate deamination by human AID. To understand the mechanisms that generate genome instability mediated by mRNP biogenesis impairment and by AID, we devised a yeast chromosomal system based on different segments of mammalian S regions and c-MYC for the analysis of chromosomal rearrangements in both wild-type and THO mutants. We demonstrate that AID acts in yeast at heterologous S and c-MYC transcribed sequences leading to double-strand breaks (DSBs) which in turn cause chromosomal translocations via Non-Homologous End Joining (NHEJ). AID-induced translocations were strongly enhanced in yeast THO null mutants, consistent with the idea that AID-mediated DSBs depend on R-loop formation. Our study not only provides new clues to understand the role of mRNP biogenesis in preventing genome rearrangements and the mechanism of AID-mediated genome instability, but also shows that, once uracil residues are produced by AID-mediated deamination, these are processed into DSBs and chromosomal rearrangements by the general and conserved DNA repair functions present from yeast to human cells.

  9. Cold-aggravated pain in humans caused by a hyperactive NaV1.9 channel mutant

    PubMed Central

    Leipold, Enrico; Hanson-Kahn, Andrea; Frick, Miya; Gong, Ping; Bernstein, Jonathan A.; Voigt, Martin; Katona, Istvan; Oliver Goral, R.; Altmüller, Janine; Nürnberg, Peter; Weis, Joachim; Hübner, Christian A.; Heinemann, Stefan H.; Kurth, Ingo

    2015-01-01

    Gain-of-function mutations in the human SCN11A-encoded voltage-gated Na+ channel NaV1.9 cause severe pain disorders ranging from neuropathic pain to congenital pain insensitivity. However, the entire spectrum of the NaV1.9 diseases has yet to be defined. Applying whole-exome sequencing we here identify a missense change (p.V1184A) in NaV1.9, which leads to cold-aggravated peripheral pain in humans. Electrophysiological analysis reveals that p.V1184A shifts the voltage dependence of channel opening to hyperpolarized potentials thereby conferring gain-of-function characteristics to NaV1.9. Mutated channels diminish the resting membrane potential of mouse primary sensory neurons and cause cold-resistant hyperexcitability of nociceptors, suggesting a mechanistic basis for the temperature dependence of the pain phenotype. On the basis of direct comparison of the mutations linked to either cold-aggravated pain or pain insensitivity, we propose a model in which the physiological consequence of a mutation, that is, augmented versus absent pain, is critically dependent on the type of NaV1.9 hyperactivity. PMID:26645915

  10. PC-FACS.

    PubMed

    Zhukovsky, Donna S

    2016-10-01

    PC-FACS(FastArticleCriticalSummaries for Clinicians inPalliativeCare) provides hospice and palliative care clinicians with concise summaries of the most important findings from more than 100 medical and scientific journals. If you have colleagues who would benefit from receiving PC-FACS, please encourage them to join the AAHPM at aahpm.org. Comments from readers are welcomed at pc-facs@aahpm.org.

  11. PC-FACS.

    PubMed

    Zhukovsky, Donna S

    2016-12-01

    PC-FACS(FastArticleCriticalSummaries for Clinicians inPalliativeCare) provides hospice and palliative care clinicians with concise summaries of the most important findings from more than 100 medical and scientific journals. If you have colleagues who would benefit from receiving PC-FACS, please encourage them to join the AAHPM at aahpm.org. Comments from readers are welcomed at pc-facs@aahpm.org.

  12. PC-FACS.

    PubMed

    Zhukovsky, Donna S

    2016-11-01

    PC-FACS(FastArticleCriticalSummaries for Clinicians inPalliativeCare) provides hospice and palliative care clinicians with concise summaries of the most important findings from more than 100 medical and scientific journals. If you have colleagues who would benefit from receiving PC-FACS, please encourage them to join the AAHPM at aahpm.org. Comments from readers are welcomed at pc-facs@aahpm.org.

  13. PC-FACS.

    PubMed

    Zhukovsky, Donna S

    2017-03-01

    PC-FACS(FastArticleCriticalSummaries for Clinicians inPalliativeCare) provides hospice and palliative care clinicians with concise summaries of the most important findings from more than 100 medical and scientific journals. If you have colleagues who would benefit from receiving PC-FACS, please encourage them to join the AAHPM at aahpm.org. Comments from readers are welcomed at pc-facs@aahpm.org.

  14. PC-FACS.

    PubMed

    Zhukovsky, Donna S

    2016-07-01

    PC-FACS(FastArticleCriticalSummaries for Clinicians inPalliativeCare) provides hospice and palliative care clinicians with concise summaries of the most important findings from more than 100 medical and scientific journals. If you have colleagues who would benefit from receiving PC-FACS, please encourage them to join the AAHPM at aahpm.org. Comments from readers are welcomed at pc-facs@aahpm.org.

  15. PC-FACS.

    PubMed

    Zhukovsky, Donna S

    2016-06-01

    PC-FACS(FastArticleCriticalSummaries for Clinicians inPalliativeCare) provides hospice and palliative care clinicians with concise summaries of the most important findings from more than 100 medical and scientific journals. If you have colleagues who would benefit from receiving PC-FACS, please encourage them to join the AAHPM at aahpm.org. Comments from readers are welcomed at pc-facs@aahpm.org.

  16. PC-FACS.

    PubMed

    Zhukovsky, Donna S

    2017-02-01

    PC-FACS(FastArticleCriticalSummaries for Clinicians inPalliativeCare) provides hospice and palliative care clinicians with concise summaries of the most important findings from more than 100 medical and scientific journals. If you have colleagues who would benefit from receiving PC-FACS, please encourage them to join the AAHPM at aahpm.org. Comments from readers are welcomed at pc-facs@aahpm.org.

  17. Structures of the troponin core domain containing the cardiomyopathy-causing mutants studied by small-angle X-ray scattering

    PubMed Central

    Matsuo, Tatsuhito; Takeda, Soichi; Oda, Toshiro; Fujiwara, Satoru

    2015-01-01

    Troponin (Tn), consisting of three subunits, TnC, TnI, and TnT, is a protein in the thin filaments in muscle, and, together with another thin-filament protein tropomyosin (Tm), plays a major role in regulation of muscle contraction. Various mutations of Tn cause familial hypertrophic cardiomyopathy. These mutations are directly related to aberrations in this regulatory mechanism. Here we focus on the mutations E244D and K247R of TnT, which reside in the middle of the pathway of the Ca2+-binding signal from TnC to Tm. These mutations induce an increase in the maximum tension of cardiac muscle without changes in Ca2+-sensitivity. As a first step toward elucidating the molecular mechanism underlying this functional aberration, we carried out small-angle X-ray scattering experiments on the Tn core domain containing the wild type subunits and those containing the mutant TnT in the absence and presence of Ca2+. Changes in the overall shape induced by the mutations were detected for the first time by the changes in the radius of gyration and the maximum dimension between the wild type and the mutants. Analysis of the scattering curves by model calculations shows that TnC adopts a dumbbell structure regardless of the mutations, and that the mutations change the distributions of the conformational ensembles so that the flexible N- and C-terminal regions of TnT become close to the center of the whole moelcule. This suggests, since these regions are related to the Tn-Tm interactions, that alteration of the Tn-Tm interactions induced by the mutations causes the functional aberration. PMID:27493864

  18. Astrocyte pathology in a human neural stem cell model of frontotemporal dementia caused by mutant TAU protein

    PubMed Central

    Hallmann, Anna-Lena; Araúzo-Bravo, Marcos J.; Mavrommatis, Lampros; Ehrlich, Marc; Röpke, Albrecht; Brockhaus, Johannes; Missler, Markus; Sterneckert, Jared; Schöler, Hans R.; Kuhlmann, Tanja; Zaehres, Holm; Hargus, Gunnar

    2017-01-01

    Astroglial pathology is seen in various neurodegenerative diseases including frontotemporal dementia (FTD), which can be caused by mutations in the gene encoding the microtubule-associated protein TAU (MAPT). Here, we applied a stem cell model of FTD to examine if FTD astrocytes carry an intrinsic propensity to degeneration and to determine if they can induce non-cell-autonomous effects in neighboring neurons. We utilized CRISPR/Cas9 genome editing in human induced pluripotent stem (iPS) cell-derived neural progenitor cells (NPCs) to repair the FTD-associated N279K MAPT mutation. While astrocytic differentiation was not impaired in FTD NPCs derived from one patient carrying the N279K MAPT mutation, FTD astrocytes appeared larger, expressed increased levels of 4R-TAU isoforms, demonstrated increased vulnerability to oxidative stress and elevated protein ubiquitination and exhibited disease-associated changes in transcriptome profiles when compared to astrocytes derived from one control individual and to the isogenic control. Interestingly, co-culture experiments with FTD astrocytes revealed increased oxidative stress and robust changes in whole genome expression in previously healthy neurons. Our study highlights the utility of iPS cell-derived NPCs to elucidate the role of astrocytes in the pathogenesis of FTD. PMID:28256506

  19. Inappropriate translation inhibition and P-body formation cause cold-sensitivity in tryptophan-auxotroph yeast mutants.

    PubMed

    Ballester-Tomás, Lidia; Prieto, Jose A; Alepuz, Paula; González, Asier; Garre, Elena; Randez-Gil, Francisca

    2017-02-01

    In response to different adverse conditions, most eukaryotic organisms, including Saccharomyces cerevisiae, downregulate protein synthesis through the phosphorylation of eIF2α (eukaryotic initiation factor 2α) by Gcn2, a highly conserved protein kinase. Gcn2 also controls the translation of Gcn4, a transcription factor involved in the induction of amino acid biosynthesis enzymes. Here, we have studied the functional role of Gcn2 and Gcn2-regulating proteins, in controlling translation during temperature downshifts of TRP1 and trp1 yeast cells. Our results suggest that neither cold-instigated amino acid limitation nor Gcn2 are involved in the translation suppression at low temperature. However, loss of TRP1 causes increased eIF2α phosphorylation, Gcn2-dependent polysome disassembly and overactivity of Gcn4, which result in cold-sensitivity. Indeed, knock-out of GCN2 improves cold growth of trp1 cells. Likewise, mutation of several Gcn2-regulators and effectors results in cold-growth effects. Remarkably, we found that Hog1, the osmoresponsive MAPK, plays a role in the regulatory mechanism of Gcn2-eIF2α. Finally, we demonstrated that P-body formation responds to a downshift in temperature in a TRP1-dependent manner and is required for cold tolerance.

  20. Use of a high-throughput screen to identify Leptospira mutants unable to colonize the carrier host or cause disease in the acute model of infection.

    PubMed

    Marcsisin, Renee A; Bartpho, Thanatchaporn; Bulach, Dieter M; Srikram, Amporn; Sermswan, Rasana W; Adler, Ben; Murray, Gerald L

    2013-10-01

    The molecular basis for leptospirosis infection and colonization remains poorly understood, with no efficient methods available for screening libraries of mutants for attenuation. We analysed the attenuation of leptospiral transposon mutants in vivo using a high-throughput method by infecting animals with pooled sets of transposon mutants. A total of 95 mutants was analysed by this method in the hamster model of acute infection, and one mutant was identified as attenuated (M1233, lb058 mutant). All virulence factors identified in Leptospira to date have been characterized in the acute model of infection, neglecting the carrier host. To address this, a BALB/c mouse colonization model was established. The lb058 mutant and two mutants defective in LPS synthesis were colonization deficient in the mouse model. By applying the high-throughput screening method, a further five colonization-deficient mutants were identified for the mouse model; these included two mutants in genes encoding proteins with a predicted role in iron uptake (LB191/HbpA and LB194). Two attenuated mutants had transposon insertions in either la0589 or la2786 (encoding proteins of unknown function). The final attenuated mutant had an unexpected deletion of genes la0969-la0975 at the point of transposon insertion. This is the first description of defined, colonization-deficient mutants in a carrier host for Leptospira. These mutants were either not attenuated or only weakly attenuated in the hamster model of acute leptospirosis, thus illustrating that different factors that may be required in the carrier and acute models of leptospiral infection. High-throughput screening can reduce the number of animals used in virulence studies and increase the capacity to screen mutants for attenuation, thereby enhancing the likelihood of detecting unique virulence factors. A comparison of virulence factors required in the carrier and acute models of infection will help to unravel colonization and dissemination

  1. Human P301L-Mutant Tau Expression in Mouse Entorhinal-Hippocampal Network Causes Tau Aggregation and Presynaptic Pathology but No Cognitive Deficits

    PubMed Central

    Harris, Julie A.; Koyama, Akihiko; Maeda, Sumihiro; Ho, Kaitlyn; Devidze, Nino; Dubal, Dena B.; Yu, Gui-Qiu; Masliah, Eliezer; Mucke, Lennart

    2012-01-01

    Accumulation of hyperphosphorylated tau in the entorhinal cortex (EC) is one of the earliest pathological hallmarks in patients with Alzheimer’s disease (AD). It can occur before significant Aβ deposition and appears to “spread” into anatomically connected brain regions. To determine whether this early-stage pathology is sufficient to cause disease progression and cognitive decline in experimental models, we overexpressed mutant human tau (hTauP301L) predominantly in layer II/III neurons of the mouse EC. Cognitive functions remained normal in mice at 4, 8, 12 and 16 months of age, despite early and extensive tau accumulation in the EC. Perforant path (PP) axon terminals within the dentate gyrus (DG) contained abnormal conformations of tau even in young EC-hTau mice, and phosphorylated tau increased with age in both the EC and PP. In old mice, ultrastructural alterations in presynaptic terminals were observed at PP-to-granule cell synapses. Phosphorylated tau was more abundant in presynaptic than postsynaptic elements. Human and pathological tau was also detected within hippocampal neurons of this mouse model. Thus, hTauP301L accumulation predominantly in the EC and related presynaptic pathology in hippocampal circuits was not sufficient to cause robust cognitive deficits within the age range analyzed here. PMID:23029293

  2. Cuticular Defects in Oryza sativa ATP-binding Cassette Transporter G31 Mutant Plants Cause Dwarfism, Elevated Defense Responses and Pathogen Resistance.

    PubMed

    Garroum, Imène; Bidzinski, Przemyslaw; Daraspe, Jean; Mucciolo, Antonio; Humbel, Bruno M; Morel, Jean-Benoit; Nawrath, Christiane

    2016-06-01

    The cuticle covers the surface of the polysaccharide cell wall of leaf epidermal cells and forms an essential diffusion barrier between plant and environment. Homologs of the ATP-binding cassette (ABC) transporter AtABCG32/HvABCG31 clade are necessary for the formation of a functional cuticle in both monocots and dicots. Here we characterize the osabcg31 knockout mutant and hairpin RNA interference (RNAi)-down-regulated OsABCG31 plant lines having reduced plant growth and a permeable cuticle. The reduced content of cutin in leaves and structural alterations in the cuticle and at the cuticle-cell wall interface in plants compromised in OsABCG31 expression explain the cuticle permeability. Effects of modifications of the cuticle on plant-microbe interactions were evaluated. The cuticular alterations in OsABCG31-compromised plants did not cause deficiencies in germination of the spores or the formation of appressoria of Magnaporthe oryzae on the leaf surface, but a strong reduction of infection structures inside the plant. Genes involved in pathogen resistance were constitutively up-regulated in OsABCG31-compromised plants, thus being a possible cause of the resistance to M. oryzae and the dwarf growth phenotype. The findings show that in rice an abnormal cuticle formation may affect the signaling of plant growth and defense.

  3. Single-cell genetic expression of mutant GABAA receptors causing Human genetic epilepsy alters dendritic spine and GABAergic bouton formation in a mutation-specific manner.

    PubMed

    Lachance-Touchette, Pamela; Choudhury, Mayukh; Stoica, Ana; Di Cristo, Graziella; Cossette, Patrick

    2014-01-01

    Mutations in genes encoding for GABAA receptor subunits is a well-established cause of genetic generalized epilepsy. GABA neurotransmission is implicated in several developmental processes including neurite outgrowth and synapse formation. Alteration in excitatory/inhibitory synaptic activities plays a critical role in epilepsy, thus here we investigated whether mutations in α1 subunit of GABAA receptor may affect dendritic spine and GABAergic bouton formation. In particular, we examined the effects of three mutations of the GABRA1 gene (D219N, A322D and K353delins18X) that were found in a cohort of French Canadian families with genetic generalized epilepsy. We used a novel single-cell genetic approach, by preparing cortical organotypic cultures from GABRA1 (flox/flox) mice and simultaneously inactivating endogenous GABRA1 and transfecting mutant α1 subunits in single glutamatergic pyramidal cells and basket GABAergic interneurons by biolistic transfection. We found that GABRA1 (-/-) GABAergic cells showed reduced innervation field, which was rescued by co-expressing α1-A322D and α1-WT but not α1-D219N. We further found that the expression of the most severe GABRA1 missense mutation (α1-A322D) induced a striking increase of spine density in pyramidal cells along with an increase in the number of mushroom-like spines. In addition, α1-A322D expression in GABAergic cells slightly increased perisomatic bouton density, whereas other mutations did not alter bouton formation. All together, these results suggest that the effects of different GABAAR mutations on GABAergic bouton and dendritic spine formation are specific to the mutation and cannot be always explained by a simple loss-of-function gene model. The use of single cell genetic manipulation in organotypic cultures may provide a better understanding of the specific and distinct neural circuit alterations caused by different GABAA receptor subunit mutations and will help define the pathophysiology of genetic

  4. Single-cell genetic expression of mutant GABAA receptors causing Human genetic epilepsy alters dendritic spine and GABAergic bouton formation in a mutation-specific manner

    PubMed Central

    Lachance-Touchette, Pamela; Choudhury, Mayukh; Stoica, Ana; Di Cristo, Graziella; Cossette, Patrick

    2014-01-01

    Mutations in genes encoding for GABAA receptor subunits is a well-established cause of genetic generalized epilepsy. GABA neurotransmission is implicated in several developmental processes including neurite outgrowth and synapse formation. Alteration in excitatory/inhibitory synaptic activities plays a critical role in epilepsy, thus here we investigated whether mutations in α1 subunit of GABAA receptor may affect dendritic spine and GABAergic bouton formation. In particular, we examined the effects of three mutations of the GABRA1 gene (D219N, A322D and K353delins18X) that were found in a cohort of French Canadian families with genetic generalized epilepsy. We used a novel single-cell genetic approach, by preparing cortical organotypic cultures from GABRA1flox/flox mice and simultaneously inactivating endogenous GABRA1 and transfecting mutant α1 subunits in single glutamatergic pyramidal cells and basket GABAergic interneurons by biolistic transfection. We found that GABRA1−/− GABAergic cells showed reduced innervation field, which was rescued by co-expressing α1-A322D and α1-WT but not α1-D219N. We further found that the expression of the most severe GABRA1 missense mutation (α1-A322D) induced a striking increase of spine density in pyramidal cells along with an increase in the number of mushroom-like spines. In addition, α1-A322D expression in GABAergic cells slightly increased perisomatic bouton density, whereas other mutations did not alter bouton formation. All together, these results suggest that the effects of different GABAAR mutations on GABAergic bouton and dendritic spine formation are specific to the mutation and cannot be always explained by a simple loss-of-function gene model. The use of single cell genetic manipulation in organotypic cultures may provide a better understanding of the specific and distinct neural circuit alterations caused by different GABAA receptor subunit mutations and will help define the pathophysiology of genetic

  5. The proteinopathy of D169G and K263E mutants at the RNA Recognition Motif (RRM) domain of tar DNA binding protein (tdp43) causing neurological disorders: A computational study.

    PubMed

    Bhandare, Vishwambhar Vishnu; Ramaswamy, Amutha

    2017-03-22

    One of the multitasking proteins, transactive response DNA-binding protein 43 (tdp43) plays a key role in RNA regulation and the two pathogenic mutations such as D169G and K263E, located at the RNA Recognition Motif (RRM) of tdp43, are reported to cause neurological disorders such as Amyotrophic Lateral Sclerosis (ALS) and Fronto Temporal Lobar Degeneration (FTLD). As the exploration of the proteinopathy demands both structural and functional characterization of mutants, a comparative analysis on the wild type and mutant tdp43 (D169G and K263E) and their complexes with RNA have been performed using computational approaches. Molecular dynamics simulations revealed comparatively stable mutant structures compared to wild type tdp43. Both mutants show lesser binding affinity towards RNA molecule when compared to the wild type tdp43. Some of the observed features, including the increased solvent accessible surface area, conformational flexibility as well as unfolding of tdp43 and the altered RNA conformation in tp43-RNA complex, reveal the susceptibility of these mutants to induce conformational changes in tdp43 for a possible aggregation in the cytoplasm. Particularly, the enhanced aggregation propensity of both mutants also evidences the higher probability of cytoplasmic aggregation of tdp43 mutants. Hence, the present analysis highlighting the structural and functional aspects of wild and mutant tdp43 will form the basis to gain insight into the proteinopathy of tdp43 and the related structure based drug discovery. Thus, tdp43 can be used as target to develop novel therapeutic approaches or drug designing.

  6. Pale-Green Phenotype of atl31 atl6 Double Mutant Leaves Is Caused by Disruption of 5-Aminolevulinic Acid Biosynthesis in Arabidopsis thaliana

    PubMed Central

    Maekawa, Shugo; Takabayashi, Atsushi; Huarancca Reyes, Thais; Yamamoto, Hiroko; Tanaka, Ayumi; Sato, Takeo; Yamaguchi, Junji

    2015-01-01

    Arabidopsis ubiquitin ligases ATL31 and homologue ATL6 control the carbon/nitrogen nutrient and pathogen responses. A mutant with the loss-of-function of both atl31 and atl6 developed light intensity-dependent pale-green true leaves, whereas the single knockout mutants did not. Plastid ultrastructure and Blue Native-PAGE analyses revealed that pale-green leaves contain abnormal plastid structure with highly reduced levels of thylakoid proteins. In contrast, the pale-green leaves of the atl31/atl6 mutant showed normal Fv/Fm. In the pale-green leaves of the atl31/atl6, the expression of HEMA1, which encodes the key enzyme for 5-aminolevulinic acid synthesis, the rate-limiting step in chlorophyll biosynthesis, was markedly down-regulated. The expression of key transcription factor GLK1, which directly promotes HEMA1 transcription, was also significantly decreased in atl31/atl6 mutant. Finally, application of 5-aminolevulinic acid to the atl31/atl6 mutants resulted in recovery to a green phenotype. Taken together, these findings indicate that the 5-aminolevulinic acid biosynthesis step was inhibited through the down-regulation of chlorophyll biosynthesis-related genes in the pale-green leaves of atl31/atl6 mutant. PMID:25706562

  7. PC-FACS.

    PubMed

    Abernethy, Amy P

    2012-03-01

    PC-FACS (Fast Article Critical Summaries for Clinicians in Palliative Care), an electronic publication of the American Academy of Hospice and Palliative Medicine, provides palliative care clinicians with concise summaries of the most important findings from more than 50 medical and scientific journals. Each month, structured summaries and insightful commentaries on 6-10 articles help palliative care clinicians stay on top of the research that is critical to contemporary practice. PC-FACS is free to AAHPM members and members can earn up to 3 CME credits quarterly. Following are excerpts from recent issues, and comments from readers are welcomed at resources@aahpm.org.

  8. PC-FACS.

    PubMed

    Zhukovsky, Donna S

    2012-04-01

    PC-FACS (Fast Article Critical Summaries for Clinicians in Palliative Care), an electronic publication of the American Academy of Hospice and Palliative Medicine, provides palliative care clinicians with concise summaries of the most important findings from more than 50 medical and scientific journals. Each month, structured summaries and insightful commentaries on 6-10 articles help palliative care clinicians stay on top of the research that is critical to contemporary practice. PC-FACS is free to AAHPM members. Following are excerpts from recent issues, and comments from readers are welcomed at resources@aahpm.org.

  9. Peak Pc Prediction in Conjunction Analysis: Conjunction Assessment Risk Analysis. Pc Behavior Prediction Models

    NASA Technical Reports Server (NTRS)

    Vallejo, J.J.; Hejduk, M.D.; Stamey, J. D.

    2015-01-01

    Satellite conjunction risk typically evaluated through the probability of collision (Pc). Considers both conjunction geometry and uncertainties in both state estimates. Conjunction events initially discovered through Joint Space Operations Center (JSpOC) screenings, usually seven days before Time of Closest Approach (TCA). However, JSpOC continues to track objects and issue conjunction updates. Changes in state estimate and reduced propagation time cause Pc to change as event develops. These changes a combination of potentially predictable development and unpredictable changes in state estimate covariance. Operationally useful datum: the peak Pc. If it can reasonably be inferred that the peak Pc value has passed, then risk assessment can be conducted against this peak value. If this value is below remediation level, then event intensity can be relaxed. Can the peak Pc location be reasonably predicted?

  10. dBRWD3 Regulates Tissue Overgrowth and Ectopic Gene Expression Caused by Polycomb Group Mutations

    PubMed Central

    Shih, Hsueh-Tzu; Chen, Wei-Yu; Liu, Kwei-Yan; Shih, Zong-Siou; Chen, Yi-Jyun; Hsieh, Paul-Chen; Kuo, Kuan-Lin; Huang, Kuo-How; Hsu, Pang-Hung; Liu, Ya-Wen; Tsai, Yu-Chen; Wu, June-Tai

    2016-01-01

    To maintain a particular cell fate, a unique set of genes should be expressed while another set is repressed. One way to repress gene expression is through Polycomb group (PcG) proteins that compact chromatin into a silent configuration. In addition to cell fate maintenance, PcG proteins also maintain normal cell physiology, for example cell cycle. In the absence of PcG, ectopic activation of the PcG-repressed genes leads to developmental defects and malignant tumors. Little is known about the molecular nature of ectopic gene expression; especially what differentiates expression of a given gene in the orthotopic tissue (orthotopic expression) and the ectopic expression of the same gene due to PcG mutations. Here we present that ectopic gene expression in PcG mutant cells specifically requires dBRWD3, a negative regulator of HIRA/Yemanuclein (YEM)-mediated histone variant H3.3 deposition. dBRWD3 mutations suppress both the ectopic gene expression and aberrant tissue overgrowth in PcG mutants through a YEM-dependent mechanism. Our findings identified dBRWD3 as a critical regulator that is uniquely required for ectopic gene expression and aberrant tissue overgrowth caused by PcG mutations. PMID:27588417

  11. Melanoma loss-of-function mutants in Xiphophorus caused by Xmrk-oncogene deletion and gene disruption by a transposable element.

    PubMed Central

    Schartl, M; Hornung, U; Gutbrod, H; Volff, J N; Wittbrodt, J

    1999-01-01

    The overexpression of the Xmrk oncogene (ONC-Xmrk) in pigment cells of certain Xiphophorus hybrids has been found to be the primary change that results in the formation of malignant melanoma. Spontaneous mutant stocks have been isolated that have lost the ability to induce tumor formation when crossed with Xiphophorus helleri. Two of these loss-of-function mutants were analyzed for genetic defects in ONC-Xmrk's. In the lof-1 mutant a novel transposable element, TX-1, has jumped into ONC-Xmrk, leading to a disruption of the gene and a truncated protein product lacking the carboxyterminal domain of the receptor tyrosine kinase. TX-1 is obviously an active LTR-containing retrotransposon in Xiphophorus that was not found in other fish species outside the family Poeciliidae. Surprisingly, it does not encode any protein, suggesting the existence of a helper function for this retroelement. In the lof-2 mutant the entire ONC-Xmrk gene was found to be deleted. These data show that ONC-Xmrk is indeed the tumor-inducing gene of Xiphophorus and thus the critical constituent of the tumor (Tu) locus. PMID:10545466

  12. The tomato homolog of the gene encoding UV-damaged DNA binding protein 1 (DDB1) underlined as the gene that causes the high pigment-1 mutant phenotype.

    PubMed

    Lieberman, Michal; Segev, Orit; Gilboa, Nehama; Lalazar, Avraham; Levin, Ilan

    2004-05-01

    A tomato EST sequence, highly homologous to the human and Arabidopsis thaliana UV-damaged DNA binding protein 1 (DDB1), was mapped to the centromeric region of the tomato chromosome 2. This region was previously shown to harbor the HP-1 gene, encoding the high pigment-1 ( hp-1) and the high pigment-1(w) ( hp-1(w)) mutant phenotypes. Recent results also show that the A. thaliana DDB1 protein interacts both genetically and biochemically with the protein encoded by DEETIOLATED1, a gene carrying three tomato mutations that are in many respects isophenotypic to hp-1: high pigment-2 ( hp-2), high pigment-2(j) ( hp-2(j)) and dark green ( dg). The entire coding region of the DDB1 gene was sequenced in an hp-1 mutant and its near-isogenic normal plant in the cv. Ailsa Craig background, and also in an hp-1(w) mutant and its isogenic normal plant in the GT breeding line background. Sequence analysis revealed a single A(931)-to-T(931) base transversion in the coding sequence of the DDB1 gene in the hp-1 mutant plants. This transversion results in the substitution of the conserved asparagine at position 311 to a tyrosine residue. In the hp-1(w) mutant, on the other hand, a single G(2392)-to-A(2392) transition was observed, resulting in the substitution of the conserved glutamic acid at position 798 to a lysine residue. The single nucleotide polymorphism that differentiates hp-1 mutant and normal plants in the cv. Ailsa Craig background was used to design a pyrosequencing genotyping system. Analysis of a resource F(2) population segregating for the hp-1 mutation revealed a very strong linkage association between the DDB1 locus and the photomorphogenic response of the seedlings, measured as hypocotyl length (25mutant phenotypes.

  13. Easy PC Astronomy

    NASA Astrophysics Data System (ADS)

    Duffett-Smith, Peter

    1996-11-01

    Easy PC Astronomy is the perfect book for everyone who wants to make easy and accurate astronomical calculations. The author supplies a simple but powerful script language called AstroScript on a disk, ready to use on any IBM PC-type computer. Equipped with this software, readers can compute complex but interesting astronomical results within minutes: from the time of moonrise or moonset anywhere in the world on any date, to the display of a lunar or solar eclipse on the computer screen--all within a few minutes of opening the book! The Sky Graphics feature of the software displays a detailed image of the sky as seen from any point on earth--at any time in the future or past--showing the constellations, planets, and a host of other features. Readers need no expert knowledge of astronomy, math or programming; the author provides full details of the calculations and formulas, which the reader can absorb or ignore as desired, and a comprehensive glossary of astronomical terms. Easy PC Astronomy is of immediate practical use to beginning and advanced amateur astronomers, students at all levels, science teachers, and research astronomers. Peter Duffett-Smith is at the Cavendish Laboratory of the University of Cambridge and is the author of Astronomy with Your Personal Computer (Cambridge University Press, 1990) and Practical Astronomy with Your Calculator (Cambridge University Press, 1989).

  14. The zebrafish early arrest mutants.

    PubMed

    Kane, D A; Maischein, H M; Brand, M; van Eeden, F J; Furutani-Seiki, M; Granato, M; Haffter, P; Hammerschmidt, M; Heisenberg, C P; Jiang, Y J; Kelsh, R N; Mullins, M C; Odenthal, J; Warga, R M; Nüsslein-Volhard, C

    1996-12-01

    This report describes mutants of the zebrafish having phenotypes causing a general arrest in early morphogenesis. These mutants identify a group of loci making up about 20% of the loci identified by mutants with visible morphological phenotypes within the first day of development. There are 12 Class I mutants, which fall into 5 complementation groups and have cells that lyse before morphological defects are observed. Mutants at three loci, speed bump, ogre and zombie, display abnormal nuclei. The 8 Class II mutants, which fall into 6 complementation groups, arrest development before cell lysis is observed. These mutants seemingly stop development in the late segmentation stages, and maintain a body shape similar to a 20 hour embryo. Mutations in speed bump, ogre, zombie, specter, poltergeist and troll were tested for cell lethality by transplanting mutant cells into wild-type hosts. With poltergeist, transplanted mutant cells all survive. The remainder of the mutants tested were autonomously but conditionally lethal: mutant cells, most of which lyse, sometimes survive to become notochord, muscles, or, in rare cases, large neurons, all cell types which become postmitotic in the gastrula. Some of the genes of the early arrest group may be necessary for progression though the cell cycle; if so, the survival of early differentiating cells may be based on having their terminal mitosis before the zygotic requirement for these genes.

  15. slender rice, a constitutive gibberellin response mutant, is caused by a null mutation of the SLR1 gene, an ortholog of the height-regulating gene GAI/RGA/RHT/D8.

    PubMed

    Ikeda, A; Ueguchi-Tanaka, M; Sonoda, Y; Kitano, H; Koshioka, M; Futsuhara, Y; Matsuoka, M; Yamaguchi, J

    2001-05-01

    The rice slender mutant (slr1-1) is caused by a single recessive mutation and results in a constitutive gibberellin (GA) response phenotype. The mutant elongates as if saturated with GAs. In this mutant, (1) elongation was unaffected by an inhibitor of GA biosynthesis, (2) GA-inducible alpha-amylase was produced by the aleurone layers without gibberellic acid application, and (3) endogenous GA content was lower than in the wild-type plant. These results indicate that the product of the SLR1 gene is an intermediate of the GA signal transduction pathway. SLR1 maps to OsGAI in rice and has significant homology with height-regulating genes, such as RHT-1Da in wheat, D8 in maize, and GAI and RGA in Arabidopsis. The GAI gene family is likely to encode transcriptional factors belonging to the GRAS gene superfamily. DNA sequence analysis revealed that the slr1-1 mutation is a single basepair deletion of the nuclear localization signal domain, resulting in a frameshift mutation that abolishes protein production. Furthermore, introduction of a 6-kb genomic DNA fragment containing the wild-type SLR1 gene into the slr1-1 mutant restored GA sensitivity to normal. These results indicate that the slr1-1 mutant is caused by a loss-of-function mutation of the SLR1 gene, which is an ortholog of GAI, RGA, RHT, and D8. We also succeeded in producing GA-insensitive dwarf rice by transforming wild-type rice with a modified SLR1 gene construct that has a 17-amino acid deletion affecting the DELLA region. Thus, we demonstrate opposite GA response phenotypes depending on the type of mutations in SLR1.

  16. The rice semi-dwarf mutant sd37, caused by a mutation in CYP96B4, plays an important role in the fine-tuning of plant growth.

    PubMed

    Zhang, Jie; Liu, Xiaoqiang; Li, Shuyu; Cheng, Zhukuan; Li, Chuanyou

    2014-01-01

    Plant cytochrome P450 has diverse roles in developmental processes and in the response to environmental cues. Here, we characterized the rice (Oryza sativa L ssp. indica cultivar 3037) semi-dwarf mutant sd37, in which the gene CYP96B4 (Cytochrome P450 96B subfamily) was identified and confirmed as the target by map-based cloning and a complementation test. A point mutation in the SRS2 domain of CYP96B4 resulted in a threonine to lysine substitution in the sd37 mutant. Examination of the subcellular localization of the protein revealed that SD37 was ER-localized protein. And SD37 was predominantly expressed in the shoot apical meristem and developing leaf and root maturation zone but not in the root apical meristem. The sd37 leaves, panicles, and seeds were smaller than those of the wild type. Histological analysis further revealed that a decrease in cell number in the mutant, specifically in the shoots, was the main cause of the dwarf phenotype. Microarray analysis demonstrated that the expression of several cell division-related genes was disturbed in the sd37 mutant. In addition, mutation or strongly overexpression of SD37 results in dwarf plants but moderate overexpression increases plant height. These data suggest that CYP96B4 may be an important regulator of plant growth that affects plant height in rice.

  17. Computational diagnosis of protein conformational diseases: short molecular dynamics simulations reveal a fast unfolding of r-LDL mutants that cause familial hypercholesterolemia.

    PubMed

    Cuesta-López, S; Falo, F; Sancho, J

    2007-01-01

    The molecular basis of conformational diseases frequently resides in mutant proteins constituting a subset of the vast mutational space. While the subtleties of protein structure point to molecular dynamics (MD) techniques as promising tools for an efficient exploration of such a space, the average size of proteins and the time scale of unfolding events make this goal difficult with present computational capabilities. We show here, nevertheless, that an efficient approach is already feasible for modular proteins. Familial hypercholesterolemia (FH) is a conformational disease linked to mutations in the gene encoding the low density lipoprotein receptor. A high percentage of these mutations has been found in the seven small modular binding repeats of the receptor. Taking advantage of its small size, we have performed an in depth MD study of the fifth binding repeat. Fast unfolding dynamics have been observed in the absence of a structural bound calcium ion, which agrees with its reported essential role in the stability of the module. In addition, several mutations detected in FH patients have been analyzed, starting from the native conformation. Our results indicate that in contrast with the wild type protein and an innocuous control mutant, disease-related mutants experience, in short simulation times (2-8 ns), gross departures from the native state that lead to unfolded conformations and, in some cases, to binding site desorganization deriving in calcium release. Computational diagnosis of mutations leading to conformational diseases seems thus feasible, at least for small or modular pathogenic proteins.

  18. PDGFRA-mutant syndrome.

    PubMed

    Ricci, Riccardo; Martini, Maurizio; Cenci, Tonia; Carbone, Arnaldo; Lanza, Paola; Biondi, Alberto; Rindi, Guido; Cassano, Alessandra; Larghi, Alberto; Persiani, Roberto; Larocca, Luigi M

    2015-07-01

    Germline PDGFRA mutations cause multiple heterogeneous gastrointestinal mesenchymal tumors. In its familial form this disease, which was formerly termed intestinal neurofibromatosis/neurofibromatosis 3b (INF/NF3b), has been included among familial gastrointestinal stromal tumors (GISTs) because of its genotype, described when GIST was the only known PDGFRA-mutant gastrointestinal tumor. Shortly afterwards, however, inflammatory fibroid polyps also revealed PDGFRA mutations. Subsequently, gastrointestinal CD34+ 'fibrous tumors' of uncertain classification were described in a germline PDGFRA-mutant context. Our aim was to characterize the syndrome produced by germline PDGFRA mutations and establish diagnostic criteria and management strategies for this hitherto puzzling disease. We studied a kindred displaying multiple gastrointestinal mesenchymal tumors, comparing it with published families/individuals with possible analogous conditions. We identified a novel inherited PDGFRA mutation (P653L), constituting the third reported example of familial PDGFRA mutation. In adult mutants we detected inflammatory fibroid polyps, gastric GISTs and gastrointestinal fibrous tumors of uncertain nosology. We demonstrate that the syndrome formerly defined as INF/NF3b (exemplified by the family reported herein) is simplistically considered a form of familial GIST, because inflammatory fibroid polyps often prevail. Fibrous tumors appear variants of inflammatory fibroid polyps. 'INF/NF3b' and 'familial GIST' are misleading terms which we propose changing to 'PDGFRA-mutant syndrome'. In this condition, unlike KIT-dependent familial GIST syndromes, if present, GISTs are stomach-restricted and diffuse Cajal cell hyperplasia is not observed. This restriction of GISTs to the stomach in PDGFRA-mutant syndrome: (i) focuses oncological concern on gastric masses, as inflammatory fibroid polyps are benign; (ii) supports a selective role of gastric environment for PDGFRA mutations to elicit GISTs

  19. Genome sequencing of the Trichoderma reesei QM9136 mutant identifies a truncation of the transcriptional regulator XYR1 as the cause for its cellulase-negative phenotype

    DOE PAGES

    Lichius, Alexander; Bidard, Frédérique; Buchholz, Franziska; ...

    2015-04-20

    Trichoderma reesei is the main industrial source of cellulases and hemicellulases required for the hydrolysis of biomass to simple sugars, which can then be used in the production of biofuels and biorefineries. The highly productive strains in use today were generated by classical mutagenesis. As byproducts of this procedure, mutants were generated that turned out to be unable to produce cellulases. In order to identify the mutations responsible for this inability, we sequenced the genome of one of these strains, QM9136, and compared it to that of its progenitor T. reesei QM6a.

  20. Different Mutant/Wild-Type p53 Combinations Cause a Spectrum of Increased Invasive Potential in Nonmalignant Immortalized Human Mammary Epithelial Cells1

    PubMed Central

    Junk, Damian J; Vrba, Lukas; Watts, George S; Oshiro, Marc M; Martinez, Jesse D; Futscher, Bernard W

    2008-01-01

    Aberrations of p53 occur in most, if not all, human cancers. In breast cancer, p53 mutation is the most common genetic defect related to a single gene. Immortalized human mammary epithelial cells resemble the earliest forms of aberrant breast tissue growth but do not express many malignancy-associated phenotypes. We created a model of human mammary epithelial tumorigenesis by infecting hTERT-HME1 immortalized human mammary epithelial cells expressing wild-type p53 with four different mutant p53 constructs to determine the role of p53 mutation on the evolution of tumor phenotypes. We demonstrate that different mutant/wild-type p53 heterozygous models generate loss of function, dominant negative activity, and a spectrum of gain of function activities that induce varying degrees of invasive potential. We suggest that this model can be used to elucidate changes that occur in early stages of human mammary epithelial tumorigenesis. These changes may constitute novel biomarkers or reveal novel treatment modalities that could inhibit progression from primary to metastatic breast disease. PMID:18472962

  1. Intracellular Transport, Assembly, and Degradation of Wild-Type and Disease-linked Mutant Gap Junction Proteins

    PubMed Central

    VanSlyke, Judy K.; Deschenes, Suzanne M.; Musil, Linda S.

    2000-01-01

    More than 130 different mutations in the gap junction integral plasma membrane protein connexin32 (Cx32) have been linked to the human peripheral neuropathy X-linked Charcot–Marie–Tooth disease (CMTX). How these various mutants are processed by the cell and the mechanism(s) by which they cause CMTX are unknown. To address these issues, we have studied the intracellular transport, assembly, and degradation of three CMTX-linked Cx32 mutants stably expressed in PC12 cells. Each mutant had a distinct fate: E208K Cx32 appeared to be retained in the endoplasmic reticulum (ER), whereas both the E186K and R142W mutants were transported to perinuclear compartments from which they trafficked either to lysosomes (R142W Cx32) or back to the ER (E186K Cx32). Despite these differences, each mutant was soluble in nonionic detergent but unable to assemble into homomeric connexons. Degradation of both mutant and wild-type connexins was rapid (t1/2 < 3 h) and took place at least in part in the ER by a process sensitive to proteasome inhibitors. The mutants studied are therefore unlikely to cause disease by accumulating in degradation-resistant aggregates but instead are efficiently cleared from the cell by quality control processes that prevent abnormal connexin molecules from traversing the secretory pathway. PMID:10848620

  2. Chemical chaperone treatment reduces intracellular accumulation of mutant collagen IV and ameliorates the cellular phenotype of a COL4A2 mutation that causes haemorrhagic stroke.

    PubMed

    Murray, Lydia S; Lu, Yinhui; Taggart, Aislynn; Van Regemorter, Nicole; Vilain, Catheline; Abramowicz, Marc; Kadler, Karl E; Van Agtmael, Tom

    2014-01-15

    Haemorrhagic stroke accounts for ∼20% of stroke cases and porencephaly is a clinical consequence of perinatal cerebral haemorrhaging. Here, we report the identification of a novel dominant G702D mutation in the collagen domain of COL4A2 (collagen IV alpha chain 2) in a family displaying porencephaly with reduced penetrance. COL4A2 is the obligatory protein partner of COL4A1 but in contrast to most COL4A1 mutations, the COL4A2 mutation does not lead to eye or kidney disease. Analysis of dermal biopsies from a patient and his unaffected father, who also carries the mutation, revealed that both display basement membrane (BM) defects. Intriguingly, defective collagen IV incorporation into the dermal BM was observed in the patient only and was associated with endoplasmic reticulum (ER) retention of COL4A2 in primary dermal fibroblasts. This intracellular accumulation led to ER stress, unfolded protein response activation, reduced cell proliferation and increased apoptosis. Interestingly, the absence of ER retention of COL4A2 and ER stress in cells from the unaffected father indicate that accumulation and/or clearance of mutant COL4A2 from the ER may be a critical modifier for disease development. Our analysis also revealed that mutant collagen IV is degraded via the proteasome. Importantly, treatment of patient cells with a chemical chaperone decreased intracellular COL4A2 levels, ER stress and apoptosis, demonstrating that reducing intracellular collagen accumulation can ameliorate the cellular phenotype of COL4A2 mutations. Importantly, these data highlight that manipulation of chaperone levels, intracellular collagen accumulation and ER stress are potential therapeutic options for collagen IV diseases including haemorrhagic stroke.

  3. Conditional Expression of Parkinson disease-related Mutant α-synuclein in the Midbrain Dopaminergic Neurons causes Progressive Neurodegeneration and Degradation of Transcription Factor Nuclear Receptor Related 1

    PubMed Central

    Lin, Xian; Parisiadou, Loukia; Sgobio, Carmelo; Liu, Guoxiang; Yu, Jia; Sun, Lixin; Shim, Hoon; Gu, Xing-Long; Luo, Jing; Long, Cai-Xia; Ding, Jinhui; Mateo, Yolanda; Sullivan, Patricia H.; Wu, Ling-Gang; Goldstein, David S.; Lovinger, David; Cai, Huaibin

    2012-01-01

    α-synuclein(α-syn) plays a prominent role in the degeneration of midbrain dopaminergic (mDA) neurons in Parkinson disease (PD). However, only a few studies on α-syn have been carried out in the mDA neurons in vivo, which may be attributed to a lack of α-syn transgenic mice that develop PD-like severe degeneration of mDA neurons. To gain mechanistic insights into the α-syn-induced mDA neurodegeneration, we generated a new line of tetracycline-regulated inducible transgenic mice that overexpressed the PD-related α-syn A53T missense mutation in the mDA neurons. Here we show that the mutant mice developed profound motor disabilities and robust mDA neurodegeneration, resembling some key motor and pathological phenotypes of PD. We further systematically examined the subcellular abnormalities appeared in the mDA neurons of mutant mice, and observed a profound decrease of dopamine release, the fragmentation of Golgi apparatus, and impairments of autophagy/lysosome degradation pathways in these neurons. To further understand the specific molecular events leading to the α-syn-dependent degeneration of mDA neurons, we found that over-expression of α-syn promoted a proteasome-dependent degradation of nuclear receptor related 1 protein (Nurr1); while inhibition of Nurr1 degradation ameliorated the α-syn-induced loss of mDA neurons. Given that Nurr1 plays an essential role in maintaining the normal function and survival of mDA neurons, our studies suggest that the α-syn-mediated suppression of Nurr1 protein expression may contribute to the preferential vulnerability of mDA neurons in the pathogenesis of PD. PMID:22764233

  4. PC index and magnetic substorms

    NASA Astrophysics Data System (ADS)

    Troshichev, Oleg; Janzhura, Alexander; Sormakov, Dmitry; Podorozhkina, Nataly

    PC index is regarded as a proxy of the solar wind energy that entered into the magnetosphere as distinct from the AL and Dst indices, which are regarded as characteristics of the energy that realize in the magnetosphere in form of substorm and magnetic storms. This conclusion is based on results of analysis of relationships between the polar cap magnetic activity (PC-index) and parameters of the solar wind, on the one hand, relationships between changes of PC and development of magnetospheric substorms (AL-index) and magnetic storms (Dst-index), on the other hand. This paper describes in detail the following main results which demonstrate a strong connection between the behavior of PC and development of magnetic disturbances in the auroral zone: (1) magnetic substorms are preceded by the РС index growth (isolated and extended substorms) or long period of stationary PC (postponed substorms), (2) the substorm sudden onsets are definitely related to such PC signatures as leap and reverse, which are indicative of sharp increase of the PC growth rate, (3) substorms generally start to develop when the PC index exceeds the threshold level ~ 1.5±0.5 mV/m, irrespective of the substorm growth phase duration and type of substorm, (4) linear dependency of AL values on PC is typical of all substorm events irrespective of type and intensity of substorm.

  5. Rough Endoplasmic Reticulum (rER) Trafficking Errors by Different Classes of Mutant DSPP Cause the Dominant Negative Effects in both Dentinogenesis Imperfecta and Dentin Dysplasia by Entrapping Normal DSPP

    PubMed Central

    von Marschall, Zofia; Mok, Seeun; Phillips, Matthew D.; McKnight, Dianalee A.; Fisher, Larry W.

    2012-01-01

    Families with nonsyndromic dentinogenesis imperfecta (DGI) and the milder, dentin dysplasia (DD), have mutations in one allele of the dentin sialophosphoprotein (DSPP) gene. Because loss of a single Dspp allele in mice (and likely, humans) causes no dental phenotype, the mechanism(s) underling the dominant-negative effects were investigated. DSPP mutations occur in three classes. (The first class, the mid-leader missense mutation, Y6D, was not investigated in this report.) All other 5' mutations of DSPP result in changes/loss in the first three amino acids (IPV) of mature DSPP or, for A15V, some retention of the hydrophobic leader sequence. All of this second class of mutations caused mutant DSPP to be retained in the rER of transfected HEK293 cells. Trafficking out of the rER by co-expressed normal DSPP was reduced in a dose-responsive manner, probably due to formation of Ca2+-dependent complexes with the retained mutant DSPP. IPV-like sequences begin many secreted Ca2+-binding proteins, and changing the third amino acid to the charged aspartate (D) in three other acidic proteins also caused increased rER accumulation. Both the leader-retaining A15V and the long string of hydrophobic amino acids resulting from all known frameshift mutations within the 3'-encoded Ca2+-binding repeat domain (third class of mutations) caused retention by association of the mutant proteins with rER membranes. More 5' frameshift mutations result in longer mutant hydrophobic domains but the milder phenotype, DD, probably due to lower effectiveness of the remaining, shorter Ca2+-binding domain in capturing normal DSPP protein within the rER. This study presents evidence of a shared underlying mechanism of capturing of normal DSPP by two different classes of DSPP mutations and offers an explanation for the mild (DD-II) versus severe (DGI-II & III) nonsyndromic dentin phenotypes. Evidence is also presented that many acidic, Ca2+-binding proteins may use the same IPV-like receptor

  6. Multifunctional human transcriptional coactivator protein PC4 is a substrate of Aurora kinases and activates the Aurora enzymes.

    PubMed

    Dhanasekaran, Karthigeyan; Kumari, Sujata; Boopathi, Ramachandran; Shima, Hiroki; Swaminathan, Amrutha; Bachu, Mahesh; Ranga, Udaykumar; Igarashi, Kazuhiko; Kundu, Tapas K

    2016-03-01

    Positive coactivator 4 (PC4), a human transcriptional coactivator, is involved in diverse processes like chromatin organization and transcription regulation. It is hyperphosphorylated during mitosis, with unknown significance. For the first time, we demonstrate the function of PC4 outside the nucleus upon nuclear envelope breakdown. A fraction of PC4 associates with Aurora A and Aurora B and undergoes phosphorylation, following which PC4 activates both Aurora A and B to sustain optimal kinase activity to maintain the phosphorylation gradient for the proper functioning of the mitotic machinery. This mitotic role is evident in PC4 knockdown cells where the defects are rescued only by the catalytically active Aurora kinases, but not the kinase-dead mutants. Similarly, the PC4 phosphodeficient mutant failed to rescue such defects. Hence, our observations establish a novel mitotic function of PC4 that might be dependent on Aurora kinase-mediated phosphorylation.

  7. Thymoquinone, a bioactive component of black caraway seeds, causes G1 phase cell cycle arrest and apoptosis in triple-negative breast cancer cells with mutant p53.

    PubMed

    Sutton, Kimberly M; Greenshields, Anna L; Hoskin, David W

    2014-01-01

    Thymoquinone (TQ) from black caraway seeds has several anticancer activities; however, its effect on triple-negative breast cancer (TNBC) cells that lack functional tumor suppressor p53 is not known. Here, we explored the growth inhibitory effect of TQ on 2 TNBC cell lines with mutant p53. Cell metabolism assays showed that TQ inhibited TNBC cell growth without affecting normal cell growth. Flow cytometric analyses of TQ-treated TNBC cells showed G1 phase cell cycle arrest and apoptosis characterized by the loss of mitochondrial membrane integrity. Western blots of lysates from TQ-treated TNBC cells showed cytochrome c and apoptosis-inducing factor in the cytoplasm, as well as caspase-9 activation consistent with the mitochondrial pathway of apoptosis. Caspase-8 was also activated in TQ-treated TNBC cells, although the mechanism of activation is not clear at this time. Importantly, TQ-induced apoptosis was only partially inhibited by zVAD-fmk, indicating a role for caspase-independent effector molecules. Poly(ADP-ribose) polymerase cleavage and increased γH2AX, as well as reduced Akt phosphorylation and decreased expression of X-linked inhibitor of apoptosis, were evident in TQ-treated cells. Finally, TQ enhanced cisplatin- and docetaxel-induced cytotoxicity. These findings suggest that TQ could be useful in the management of TNBC, even when functional p53 is absent.

  8. Wanted: A Solid, Reliable PC

    ERIC Educational Resources Information Center

    Goldsborough, Reid

    2004-01-01

    This article discusses PC reliability, one of the most pressing issues regarding computers. Nearly a quarter century after the introduction of the first IBM PC and the outset of the personal computer revolution, PCs have largely become commodities, with little differentiating one brand from another in terms of capability and performance. Most of…

  9. NASA PC software evaluation project

    NASA Technical Reports Server (NTRS)

    Dominick, Wayne D. (Editor); Kuan, Julie C.

    1986-01-01

    The USL NASA PC software evaluation project is intended to provide a structured framework for facilitating the development of quality NASA PC software products. The project will assist NASA PC development staff to understand the characteristics and functions of NASA PC software products. Based on the results of the project teams' evaluations and recommendations, users can judge the reliability, usability, acceptability, maintainability and customizability of all the PC software products. The objective here is to provide initial, high-level specifications and guidelines for NASA PC software evaluation. The primary tasks to be addressed in this project are as follows: to gain a strong understanding of what software evaluation entails and how to organize a structured software evaluation process; to define a structured methodology for conducting the software evaluation process; to develop a set of PC software evaluation criteria and evaluation rating scales; and to conduct PC software evaluations in accordance with the identified methodology. Communication Packages, Network System Software, Graphics Support Software, Environment Management Software, General Utilities. This report represents one of the 72 attachment reports to the University of Southwestern Louisiana's Final Report on NASA Grant NGT-19-010-900. Accordingly, appropriate care should be taken in using this report out of context of the full Final Report.

  10. Current insights into the role of PKA phosphorylation in CFTR channel activity and the pharmacological rescue of cystic fibrosis disease-causing mutants.

    PubMed

    Chin, Stephanie; Hung, Maurita; Bear, Christine E

    2017-01-01

    Cystic fibrosis transmembrane conductance regulator (CFTR) channel gating is predominantly regulated by protein kinase A (PKA)-dependent phosphorylation. In addition to regulating CFTR channel activity, PKA phosphorylation is also involved in enhancing CFTR trafficking and mediating conformational changes at the interdomain interfaces of the protein. The major cystic fibrosis (CF)-causing mutation is the deletion of phenylalanine at position 508 (F508del); it causes many defects that affect CFTR trafficking, stability, and gating at the cell surface. Due to the multiple roles of PKA phosphorylation, there is growing interest in targeting PKA-dependent signaling for rescuing the trafficking and functional defects of F508del-CFTR. This review will discuss the effects of PKA phosphorylation on wild-type CFTR, the consequences of CF mutations on PKA phosphorylation, and the development of therapies that target PKA-mediated signaling.

  11. Altered neuronal mitochondrial coenzyme A synthesis in neurodegeneration with brain iron accumulation caused by abnormal processing, stability, and catalytic activity of mutant pantothenate kinase 2.

    PubMed

    Kotzbauer, Paul T; Truax, Adam C; Trojanowski, John Q; Lee, Virginia M-Y

    2005-01-19

    Mutations in the pantothenate kinase 2 (PANK2) gene have been identified in patients with neurodegeneration with brain iron accumulation (NBIA; formerly Hallervorden-Spatz disease). However, the mechanisms by which these mutations cause neurodegeneration are unclear, especially given the existence of multiple pantothenate kinase genes in humans and multiple PanK2 transcripts with potentially different subcellular localizations. We demonstrate that PanK2 protein is localized to mitochondria of neurons in human brain, distinguishing it from other pantothenate kinases that do not possess mitochondrial-targeting sequences. PanK2 protein translated from the most 5' start site is sequentially cleaved at two sites by the mitochondrial processing peptidase, generating a long-lived 48 kDa mature protein identical to that found in human brain extracts. The mature protein catalyzes the initial step in coenzyme A (CoA) synthesis but displays feedback inhibition in response to species of acyl CoA rather than CoA itself. Some, but not all disease-associated point mutations result in significantly reduced catalytic activity. The most common mutation, G521R, results in marked instability of the intermediate PanK2 isoform and reduced production of the mature isoform. These results suggest that NBIA is caused by altered neuronal mitochondrial lipid metabolism caused by mutations disrupting PanK2 protein levels and catalytic activity.

  12. 75 FR 62002 - Airworthiness Directives; Pilatus Aircraft Ltd. Models PC-6, PC-6-H1, PC-6-H2, PC-6/350, PC-6/350...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2010-10-07

    ... Limitations Section (ALS). For PC-6 models other than B2-H2 and B2- H4, no ALS at all is included in the AMM... and B2-H4 models. For PC-6 models other than B2-H2 and B2-H4, a new ALS document has been implemented... Limitations Section (ALS). For PC-6 models other than B2-H2 and B2- H4, no ALS at all is included in the...

  13. 76 FR 5467 - Airworthiness Directives; Pilatus Aircraft Ltd. Models PC-6, PC-6-H1, PC-6-H2, PC-6/350, PC-6/350...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-02-01

    ... Limitations Section (ALS). For PC-6 models other than B2-H2 and B2- H4, no ALS at all is included in the AMM... and B2-H4 models. For PC-6 models other than B2-H2 and B2-H4, a new ALS document has been implemented... Limitations Section (ALS). For PC-6 models other than B2-H2 and B2- H4, no ALS at all is included in the...

  14. Muscle dystrophy-causing ΔK32 lamin A/C mutant does not impair functions of nucleoplasmic LAP2α - lamin A/C complexes in mice

    PubMed Central

    Pilat, Ursula; Dechat, Thomas; Bertrand, Anne T; Woisetschläger, Nikola; Gotic, Ivana; Spilka, Rita; Biadasiewicz, Katarzyna; Bonne, Gisèle; Foisner, Roland

    2015-01-01

    Summary A-type lamins are components of the nuclear lamina, a filamentous network of the nuclear envelope in metazoans that supports nuclear architecture. In addition, lamin A/C can also be found in the nuclear interior. This nucleoplasmic lamin pool is soluble in physiological buffer, depends on the presence of the lamin-binding protein, Lamina-associated polypeptide 2α (LAP2α) and regulates cell cycle progression in tissue progenitor cells. ΔK32 mutations in A-type lamins cause severe congenital muscle disease in humans and a muscle maturation defect in LmnaΔK32/ΔK32 knock-in mice. At molecular level, mutant ΔK32 lamin A/C protein levels were reduced and all mutant lamin A/C was soluble and mislocalized to the nucleoplasm. To test the role of LAP2α in nucleoplasmic ΔK32 lamin A/C regulation and functions, we deleted LAP2α in LmnaΔK32/ΔK32 knock-in mice. In double mutant mice the LmnaΔK32/ΔK32- linked muscle defect was unaffected. LAP2α interacted with mutant lamin A/C, but unlike wild-type lamin A/C, the intranuclear localization of ΔK32 lamin A/C was not affected by loss of LAP2α. In contrast, loss of LAP2α in LmnaΔK32/ΔK32 mice impaired the regulation of tissue progenitor cells like in lamin A/C wild type animals. These data indicate that a LAP2α-independent assembly defect of ΔK32 lamin A/C is predominant for the mouse pathology, while the LAP2α-linked functions of nucleoplasmic lamin A/C in the regulation of tissue progenitor cells are not affected in LmnaΔK32/ΔK32 mice. PMID:23444379

  15. Anxiety, memory impairment, and locomotor dysfunction caused by a mutant thyroid hormone receptor α1 can be ameliorated by T3 treatment

    PubMed Central

    Venero, César; Guadaño-Ferraz, Ana; Herrero, Ana Isabel; Nordström, Kristina; Manzano, Jimena; de Escobar, Gabriella Moreale; Bernal, Juan; Vennström, Björn

    2005-01-01

    The transcriptional properties of unliganded thyroid hormone receptors are thought to cause the misdevelopment during hypothyroidism of several functions essential for adult life. To specifically determine the role of unliganded thyroid hormone receptor α1 (TRα1) in neuronal tissues, we introduced a mutation into the mouse TRα1 gene that lowers affinity to thyroid hormone (TH) 10-fold. The resulting heterozygous mice exhibit several distinct neurological abnormalities: extreme anxiety, reduced recognition memory, and locomotor dysfunction. The anxiety and memory deficiencies were relieved by treatment with high levels of TH in adulthood, an effect that correlated with a normalization of GABAergic inhibitory interneurons in the hippocampal CA1 region. In contrast, a post-natal TH treatment was necessary and sufficient for ameliorating the adult locomotor dysfunction. Here, the hormone treatment normalized the otherwise delayed cerebellar development. The data thus identify two novel and distinct functions of an unliganded TRα1 during development and adulthood, respectively. PMID:16131613

  16. Intravitreal injection of ciliary neurotrophic factor (CNTF) causes peripheral remodeling and does not prevent photoreceptor loss in canine RPGR mutant retina.

    PubMed

    Beltran, William A; Wen, Rong; Acland, Gregory M; Aguirre, Gustavo D

    2007-04-01

    Ciliary neurotrophic factor (CNTF) rescues photoreceptors in several animal models of retinal degeneration and is currently being evaluated as a potential treatment for retinitis pigmentosa in humans. This study was conducted to test whether CNTF prevents photoreceptor cell loss in XLPRA2, an early onset canine model of X-linked retinitis pigmentosa caused by a frameshift mutation in RPGR exon ORF15. Four different treatment regimens of CNTF were tested in XLPRA2 dogs. Under anesthesia, the animals received at different ages an intravitreal injection of 12 microg of CNTF in the left eye. The right eye served as a control and was injected with a similar volume of phosphate buffered saline (PBS). Ocular examinations were performed regularly during the treatment periods. At termination, the dogs were euthanatized, eyes collected and the retinas were processed for embedding in optimal cutting temperature (OCT) medium. The outer nuclear layer (ONL) thickness was evaluated on H&E sections and values in both CNTF- and PBS-treated eyes were compared. Morphologic alterations in the peripheral retina were characterized by immunohistochemistry using cell-specific markers. Cell proliferation in the retinas was examined on semi-thin plastic sections, and by BrdU pulse-labeling and Ki67 immunohistochemistry on cryosections. All CNTF-treated eyes showed early clinical signs of corneal epitheliopathy, subcapsular cataracts and uveitis. No statistically significant difference in ONL thickness was seen between the CNTF- and PBS-injected eyes. Prominent retinal remodeling that consisted in an abnormal increase in the number of rods, and in misplacement of some rods, cones, bipolar and Müller cells, was observed in the peripheral retina of CNTF-treated eyes. This was only seen when CNTF was in injected before the age at which the canine retina reaches full maturation. In XLPRA2 dogs, intravitreal injections of CNTF failed to prevent photoreceptors from undergoing cell death in the

  17. Intravitreal injection of ciliary neurotrophic factor (CNTF) causes peripheral remodeling and does not prevent photoreceptor loss in canine RPGR mutant retina

    PubMed Central

    Beltran, William A.; Wen, Rong; Acland, Gregory M.; Aguirre, Gustavo D.

    2009-01-01

    Ciliary neurotrophic factor (CNTF) rescues photoreceptors in several animal models of retinal degeneration and is currently being evaluated as a potential treatment for retinitis pigmentosa in humans. This study was conducted to test whether CNTF prevents photoreceptor cell loss in XLPRA2, an early onset canine model of X-linked retinitis pigmentosa caused by a frameshift mutation in RPGR exon ORF15. Four different treatment regimens of CNTF were tested in XLPRA2 dogs. Under anesthesia, the animals received at different ages an intravitreal injection of 12 μg of CNTF in the left eye. The right eye served as a control and was injected with a similar volume of phosphate buffered saline (PBS). Ocular examinations were performed regularly during the treatment periods. At termination, the dogs were euthanatized, eyes collected and the retinas were processed for embedding in optimal cutting temperature (OCT) medium. The outer nuclear layer (ONL) thickness was evaluated on H&E sections and values in both CNTF- and PBS-treated eyes were compared. Morphologic alterations in the peripheral retina were characterized by immunohistochemistry using cell-specific markers. Cell proliferation in the retinas was examined on semi-thin plastic sections, and by BrdU pulse-labeling and Ki67 immunohistochemistry on cryosections. All CNTF-treated eyes showed early clinical signs of corneal epitheliopathy, subcapsular cataracts and uveitis. No statistically significant difference in ONL thickness was seen between the CNTF- and PBS-injected eyes. Prominent retinal remodeling that consisted in an abnormal increase in the number of rods, and in misplacement of some rods, cones, bipolar and Müller cells, was observed in the peripheral retina of CNTF-treated eyes. This was only seen when CNTF was in injected before the age at which the canine retina reaches full maturation. In XLPRA2 dogs, intravitreal injections of CNTF failed to prevent photoreceptors from undergoing cell death in the

  18. Overexpression of metallothionein-I, a copper-regulating protein, attenuates intracellular copper dyshomeostasis and extends lifespan in a mouse model of amyotrophic lateral sclerosis caused by mutant superoxide dismutase-1.

    PubMed

    Tokuda, Eiichi; Okawa, Eriko; Watanabe, Shunsuke; Ono, Shin-Ichi

    2014-03-01

    Over 170 mutations in superoxide dismutase-1 (SOD1) cause familial amyotrophic lateral sclerosis (ALS), a lethal motor neuron disease. Although the molecular properties of SOD1 mutants differ considerably, we have recently shown that intracellular copper dyshomeostasis is a common pathogenic feature of different SOD1 mutants. Thus, the potentiation of endogenous copper regulation could be a therapeutic strategy. In this study, we investigated the effects of the overexpression of metallothionein-I (MT-I), a major copper-regulating protein, on the disease course of a mouse model of ALS (SOD1(G93A)). Using double transgenic techniques, we found that the overexpression of MT-I in SOD1(G93A) mice significantly extended the lifespan and slowed disease progression, but the effects on disease onset were modest. Genetically induced MT-I normalized copper dyshomeostasis in the spinal cord without influencing SOD1 enzymatic activity. The overexpression of MT-I in SOD1(G93A) mice markedly attenuated the pathological features of the mice, including the death of motor neurons, the degeneration of ventral root axons, the atrophy of skeletal muscles, and the activation of glial cells. Double transgenic mice also showed a decreased level of SOD1 aggregates within the glial cells of the spinal cord. Furthermore, the overexpression of MT-I in SOD1(G93A) mice reduced the number of spheroid-shaped astrocytes cleaved by active caspase-3. We concluded that therapeutic strategies aimed at the potentiation of copper regulation by MT-I could be of benefit in cases of ALS caused by SOD1 mutations.

  19. Cyclin B2 undergoes cell cycle-dependent nuclear translocation and, when expressed as a non-destructible mutant, causes mitotic arrest in HeLa cells

    PubMed Central

    1992-01-01

    Cyclin proteins form complexes with members of the p34cdc2 kinase family and they are essential components of the cell cycle regulatory machinery. They are thought to determine the timing of activation, the subcellular distribution, and/or the substrate specificity of cdc2- related kinases, but their precise mode of action remains to be elucidated. Here we report the cloning and sequencing of avian cyclin B2. Based on the use of monospecific antibodies raised against bacterially expressed protein, we also describe the subcellular distribution of cyclin B2 in chick embryo fibroblasts and in DU249 hepatoma cells. By indirect immunofluorescence microscopy we show that cyclin B2 is cytoplasmic during interphase of the cell cycle, but undergoes an abrupt translocation to the cell nucleus at the onset of mitotic prophase. Finally, we have examined the phenotypic consequences of expressing wild-type and mutated versions of avian cyclin B2 in HeLa cells. We found that expression of cyclin B2 carrying a mutation at arginine 32 (to serine) caused HeLa cells to arrest in a pseudomitotic state. Many of the arrested cells displayed multiple mitotic spindles, suggesting that the centrosome cycle had continued in spite of the cell cycle arrest. PMID:1532584

  20. The Caenorhabditis elegans PcG-like gene sop-2 regulates the temporal and sexual specificities of cell fates.

    PubMed

    Cai, Qingchun; Sun, Yinyan; Huang, Xinxin; Guo, Cong; Zhang, Yuxia; Zhu, Zuoyan; Zhang, Hong

    2008-03-01

    How spatial, temporal, and sexual specific cues are integrated to specify distinct cell fates during multicellular organism development is largely unknown. Here we demonstrate that the Caenorhabditis elegans PcG-like gene sop-2 determines the temporal and sexual specificities of a row of hypodermal seam cells, in addition to specifying their positional identities. Loss-of-function of sop-2 causes premature expression of adult fates at larval stages. sop-2 acts upstream of lin-29 in the heterochronic pathway and genetically interacts with other heterochronic genes in specifying the temporal fates of seam cells at different larval stages. We show that the number of ALG-1-containing P bodies is increased in seam cells in sop-2 mutants. Furthermore, the microRNA-mediated repression of a heterochronic gene reporter is enhanced in sop-2 mutants. Mutations in sop-2 also cause partial hermaphrodite-to-male sexual transformations. The homeotic transformations, heterochronic defects, and sexual transformations can occur concomitantly in sop-2 mutants. In summary, our studies reveal that sop-2 integrates spatial, temporal, and sexual cues during C. elegans development.

  1. CtBP Levels Control Intergenic Transcripts, PHO/YY1 DNA Binding, and PcG Recruitment to DNA

    PubMed Central

    Basu, Arindam; Atchison, Michael L.

    2013-01-01

    Carboxy-terminal binding protein (CtBP) is a well-known corepressor of several DNA binding transcription factors in Drosophila as well as in mammals. CtBP is implicated in Polycomb Group (PcG) complex-mediated transcriptional repression because it can bind to some PcG proteins, and mutation of the ctbp gene in flies results in lost PcG protein recruitment to Polycomb Response Elements (PREs) and lost PcG repression. However, the mechanism of reduced PcG DNA binding in CtBP mutant backgrounds is unknown. We show here that in a Drosophila CtBP mutant background, intergenic transcripts are induced across several PRE sequences and this corresponds to reduced DNA binding by PcG proteins Pleiohomeotic (PHO) and Polycomb (Pc), and reduced trimethylation of histone H3 on lysine 27, a hallmark of PcG repression. Restoration of CtBP levels by expression of a CtBP transgene results in repression of intergenic transcripts, restored PcG binding, and elevated trimethylation of H3 on lysine 27. Our results support a model in which CtBP regulates expression of intergenic transcripts that controls DNA binding by PcG proteins and subsequent histone modifications and transcriptional activity. PMID:20082324

  2. D2PC sensitivity analysis

    SciTech Connect

    Lombardi, D.P.

    1992-08-01

    The Chemical Hazard Prediction Model (D2PC) developed by the US Army will play a critical role in the Chemical Stockpile Emergency Preparedness Program by predicting chemical agent transport and dispersion through the atmosphere after an accidental release. To aid in the analysis of the output calculated by D2PC, this sensitivity analysis was conducted to provide information on model response to a variety of input parameters. The sensitivity analysis focused on six accidental release scenarios involving chemical agents VX, GB, and HD (sulfur mustard). Two categories, corresponding to conservative most likely and worst case meteorological conditions, provided the reference for standard input values. D2PC displayed a wide variety of sensitivity to the various input parameters. The model displayed the greatest overall sensitivity to wind speed, mixing height, and breathing rate. For other input parameters, sensitivity was mixed but generally lower. Sensitivity varied not only with parameter, but also over the range of values input for a single parameter. This information on model response can provide useful data for interpreting D2PC output.

  3. Cooperation between SMYD3 and PC4 drives a distinct transcriptional program in cancer cells

    PubMed Central

    Kim, Jin-Man; Kim, Kyunghwan; Schmidt, Thomas; Punj, Vasu; Tucker, Haley; Rice, Judd C.; Ulmer, Tobias S.; An, Woojin

    2015-01-01

    SET and MYND domain containing protein 3 (SMYD3) is a histone methyltransferase, which has been implicated in cell growth and cancer pathogenesis. Increasing evidence suggests that SMYD3 can influence distinct oncogenic processes by acting as a gene-specific transcriptional regulator. However, the mechanistic aspects of SMYD3 transactivation and whether SMYD3 acts in concert with other transcription modulators remain unclear. Here, we show that SMYD3 interacts with the human positive coactivator 4 (PC4) and that such interaction potentiates a group of genes whose expression is linked to cell proliferation and invasion. SMYD3 cooperates functionally with PC4, because PC4 depletion results in the loss of SMYD3-mediated H3K4me3 and target gene expression. Individual depletion of SMYD3 and PC4 diminishes the recruitment of both SMYD3 and PC4, indicating that SMYD3 and PC4 localize at target genes in a mutually dependent manner. Artificial tethering of a SMYD3 mutant incapable of binding to its cognate elements and interacting with PC4 to target genes is sufficient for achieving an active transcriptional state in SMYD3-deficient cells. These observations suggest that PC4 contributes to SMYD3-mediated transactivation primarily by stabilizing SMYD3 occupancy at target genes. Together, these studies define expanded roles for SMYD3 and PC4 in gene regulation and provide an unprecedented documentation of their cooperative functions in stimulating oncogenic transcription. PMID:26350217

  4. A novel homozygous no-stop mutation in G6PC gene from a Chinese patient with glycogen storage disease type Ia.

    PubMed

    Gu, Lei-Lei; Li, Xin-Hua; Han, Yue; Zhang, Dong-Hua; Gong, Qi-Ming; Zhang, Xin-Xin

    2014-02-25

    Glycogen storage disease type Ia (GSD-Ia) is an autosomal recessive genetic disorder resulting in hypoglycemia, hepatomegaly and growth retardation. It is caused by mutations in the G6PC gene encoding Glucose-6-phosphatase. To date, over 80 mutations have been identified in the G6PC gene. Here we reported a novel mutation found in a Chinese patient with abnormal transaminases, hypoglycemia, hepatomegaly and short stature. Direct sequencing of the coding region and splicing-sites in the G6PC gene revealed a novel no-stop mutation, p.*358Yext*43, leading to a 43 amino-acid extension of G6Pase. The expression level of mutant G6Pase transcripts was only 7.8% relative to wild-type transcripts. This mutation was not found in 120 chromosomes from 60 unrelated healthy control subjects using direct sequencing, and was further confirmed by digestion with Rsa I restriction endonuclease. In conclusion, we revealed a novel no-stop mutation in this study which expands the spectrum of mutations in the G6PC gene. The molecular genetic analysis was indispensable to the diagnosis of GSD-Ia for the patient.

  5. Remote estimation of phycocyanin (PC) for inland waters coupled with YSI PC fluorescence probe.

    PubMed

    Song, Kaishan; Li, Lin; Tedesco, Lenore; Clercin, Nicole; Hall, Bob; Li, Shuai; Shi, Kun; Liu, Dawei; Sun, Ying

    2013-08-01

    Nuisance cyanobacterial blooms degrade water resources through accelerated eutrophication, odor generation, and production of toxins that cause adverse effects on human health. Quick and effective methods for detecting cyanobacterial abundance in drinking water supplies are urgently needed to compliment conventional laboratory methods, which are costly and time consuming. Hyperspectral remote sensing can be an effective approach for rapid assessment of cyanobacterial blooms. Samples (n=250) were collected from five drinking water sources in central Indiana (CIN), USA, and South Australia (SA), which experience nuisance cyanobacterial blooms. In situ hyperspectral data were used to develop models by relating spectral signal with handheld fluorescence probe (YSI 6600 XLM-SV) measured phycocyanin (PC in cell/ml), a proxy pigment unique for indicating the presence of cyanobacteria. Three-band model (TBM), which is effective for chlorophyll-a estimates, was tuned to quantify cyanobacteria coupled with the PC probe measured cyanobacteria. As a comparison, two band model proposed by Simis et al. (Limnol Oceanogr, 50(11): 237-245, 2005; denoted as SM05) was paralleled to evaluate TBM model performance. Our observation revealed a high correlation between measured and estimated PC for SA dataset (R (2) =0.96; range: 534-20,200 cell/ml) and CIN dataset (R (2) =0.88; range: 1,300-44,500 cell/ml). The potential of this modeling approach for imagery data were assessed by simulated ESA/Centinel3/OLCI spectra, which also resulted in satisfactory performance with the TBM for both SA dataset (RMSE % =26.12) and CIN dataset (RMSE % =34.49). Close relationship between probe-measured PC and laboratory measured cyanobacteria biovolume was observed (R (2) =0.93, p<0.0001) for the CIN dataset, indicating a stable performance for PC probe. Based on our observation, field spectroscopic measurement coupled with PC probe measurements can provide quantitative cyanobacterial bloom

  6. The single-strand DNA binding activity of human PC4 preventsmutagenesis and killing by oxidative DNA damage

    SciTech Connect

    Wang, Jen-Yeu; Sarker, Altaf Hossain; Cooper, Priscilla K.; Volkert, Michael R.

    2004-02-01

    Human positive cofactor 4 (PC4) is a transcriptional coactivator with a highly conserved single-strand DNA (ssDNA) binding domain of unknown function. We identified PC4 as a suppressor of the oxidative mutator phenotype of the Escherichia coli fpg mutY mutant and demonstrate that this suppression requires its ssDNA binding activity. Yeast mutants lacking their PC4 ortholog Sub1 are sensitive to hydrogen peroxide and exhibit spontaneous and peroxide induced hypermutability. PC4 expression suppresses the peroxide sensitivity of the yeast sub l{Delta} mutant, suggesting that the human protein has a similar function. A role for yeast and human proteins in DNA repair is suggested by the demonstration that Sub1 acts in a peroxide-resistance pathway involving Rad2 and by the physical interaction of PC4 with the human Rad2 homolog XPG. We show XPG recruits PC4 to a bubble-containing DNA substrate with resulting displacement of XPG and formation of a PC4-DNA complex. We discuss the possible requirement for PC4 in either global or transcription-coupled repair of oxidative DNA damage to mediate the release of XPG bound to its substrate.

  7. Photodynamic therapy with Pc 4 induces apoptosis of Candida albicans.

    PubMed

    Lam, Minh; Jou, Paul C; Lattif, Ali A; Lee, Yoojin; Malbasa, Christi L; Mukherjee, Pranab K; Oleinick, Nancy L; Ghannoum, Mahmoud A; Cooper, Kevin D; Baron, Elma D

    2011-01-01

    The high prevalence of drug resistance necessitates the development of novel antifungal agents against infections caused by opportunistic fungal pathogens, such as Candida albicans. Elucidation of apoptosis in yeast-like fungi may provide a basis for future therapies. In mammalian cells, photodynamic therapy (PDT) has been demonstrated to generate reactive oxygen species, leading to immediate oxidative modifications of biological molecules and resulting in apoptotic cell death. In this report, we assess the in vitro cytotoxicity and mechanism of PDT, using the photosensitizer Pc 4, in planktonic C. albicans. Confocal image analysis confirmed that Pc 4 localizes to cytosolic organelles, including mitochondria. A colony formation assay showed that 1.0 μM Pc 4 followed by light at 2.0 J cm(-2) reduced cell survival by 4 logs. XTT (2,3-bis[2-methoxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxyanilide) assay revealed that Pc 4-PDT impaired fungal metabolic activity, which was confirmed using the FUN-1 (2-chloro-4-[2,3-dihydro-3-methyl-(benzo-1,3-thiazol-2-yl)-methylidene]-1-phenylquinolinium iodide) fluorescence probe. Furthermore, we observed changes in nuclear morphology characteristic of apoptosis, which were substantiated by increased externalization of phosphatidylserine and DNA fragmentation following Pc 4-PDT. These data indicate that Pc 4-PDT can induce apoptosis in C. albicans. Therefore, a better understanding of the process will be helpful, as PDT may become a useful treatment option for candidiasis.

  8. Poster session ELIPGRID-PC

    SciTech Connect

    Davidson, J.R.

    1995-02-01

    ELIPGRID-PC, a new personal computer program, has been developed to provide easy access to Singer`s ELIPGRID algorithm for hot-spot detection probabilities. Three features of the program are the ability to determine: (1) the grid size required for specified conditions, (2) the smallest hot spot that can be sampled with a given probability, and (3) the approximate grid size resulting from specified conditions and sampling cost. ELIPGRID-PC also provides probability of detection versus cost data for graphing with spreadsheets or graphics software. The program has been successfully tested using Singer`s published ELIPGRID results. An apparent error in the published ELIPGRID code has been uncovered and an appropriate modification incorporated into the new program.

  9. Increased vulnerability of hippocampal neurons from presenilin-1 mutant knock-in mice to amyloid beta-peptide toxicity: central roles of superoxide production and caspase activation.

    PubMed

    Guo, Q; Sebastian, L; Sopher, B L; Miller, M W; Ware, C B; Martin, G M; Mattson, M P

    1999-03-01

    Many cases of early-onset inherited Alzheimer's disease (AD) are caused by mutations in the presenilin-1 (PS1) gene. Overexpression of PS1 mutations in cultured PC12 cells increases their vulnerability to apoptosis-induced trophic factor withdrawal and oxidative insults. We now report that primary hippocampal neurons from PS1 mutant knock-in mice, which express the human PS1M146V mutation at normal levels, exhibit increased vulnerability to amyloid beta-peptide toxicity. The endangering action of mutant PS1 was associated with increased superoxide production, mitochondrial membrane depolarization, and caspase activation. The peroxynitrite-scavenging antioxidant uric acid and the caspase inhibitor benzyloxycarbonyl-Val-Ala-Asp-fluoromethyl ketone protected hippocampal neurons expressing mutant PS1 against cell death induced by amyloid beta-peptide. Increased oxidative stress may contribute to the pathogenic action of PS1 mutations, and antioxidants may counteract the adverse property of such AD-linked mutations.

  10. The isoforms of proprotein convertase PC5 are sorted to different subcellular compartments

    PubMed Central

    1996-01-01

    The proprotein convertase PC5 is encoded by multiple mRNAs, two of which give rise to the COOH-terminal variant isoforms PC5-A (915 amino acids [aa]) and PC5-B (1877 aa). To investigate the differences in biosynthesis and sorting between these two proteins, we generated stably transfected AtT-20 cell lines expressing each enzyme individually and examined their respective processing pattern and subcellular localization. Biosynthetic analyses coupled to immunofluorescence studies demonstrated that the shorter and soluble PC5-A is sorted to regulated secretory granules. In contrast, the COOH- terminally extended and membrane-bound PC5-B is located in the Golgi. The presence of a sorting signal in the COOH-terminal 38 amino acids unique to PC5-A was demonstrated by the inefficient entry into the regulated secretory pathway of a mutant lacking this segment. EM of pancreatic cells established the presence of immunoreactive PC5 in glucagon-containing granules, demonstrating the sorting of this protein to dense core secretory granules in endocrine cells. Thus, a single PC5 gene generates COOH-terminally modified isoforms with different sorting signals directing these proteins to distinct subcellular localization, thereby allowing them to process their appropriate substrates. PMID:8947550

  11. PC25{trademark} product and manufacturing experience

    SciTech Connect

    Hall, E.W.; Riley, W.C.; Sandelli, G.J.

    1996-12-31

    Product and manufacturing experience accumulated since the beginning of PC25. A production in 1991 provides a strong base of demonstration and experience for establishing future improvements to the PC25 power plant.

  12. The Pseudophosphatase MK-STYX Induces Neurite-Like Outgrowths in PC12 Cells

    PubMed Central

    Flowers, Brittany M.; Rusnak, Lauren E.; Wong, Kristen E.; Banks, Dallas A.; Munyikwa, Michelle R.; McFarland, Alexander G.; Hinton, Shantá D.

    2014-01-01

    The rat pheochromocytoma PC12 cell line is a widely used system to study neuronal differentiation for which sustained activation of the extracellular signaling related kinase (ERK) pathway is required. Here, we investigate the function of MK-STYX [MAPK (mitogen-activated protein kinase) phosphoserine/threonine/tyrosine-binding protein] in neuronal differentiation. MK-STYX is a member of the MAPK phosphatase (MKP) family, which is generally responsible for dephosphorylating the ERKs. However, MK-STYX lacks catalytic activity due to the absence of the nucleophilic cysteine in the active site signature motif HC(X5)R that is essential for phosphatase activity. Despite being catalytically inactive, MK-STYX has been shown to play a role in important cellular pathways, including stress responses. Here we show that PC12 cells endogenously express MK-STYX. In addition, MK-STYX, but not its catalytically active mutant, induced neurite-like outgrowths in PC12 cells. Furthermore, MK-STYX dramatically increased the number of cells with neurite extensions in response to nerve growth factor (NGF), whereas the catalytically active mutant did not. MK-STYX continued to induce neurites in the presence of a MEK (MAP kinase kinase) inhibitor suggesting that MK-STYX does not act through the Ras-ERK/MAPK pathway but is involved in another pathway whose inactivation leads to neuronal differentiation. RhoA activity assays indicated that MK-STYX induced extensions through the Rho signaling pathway. MK-STYX decreased RhoA activation, whereas RhoA activation increased when MK-STYX was down-regulated. Furthermore, MK-STYX affected downstream players of RhoA such as the actin binding protein cofilin. The presence of MK-STYX decreased the phosphorylation of cofilin in non NGF stimulated cells, but increased its phosphorylation in NGF stimulated cells, whereas knocking down MK-STYX caused an opposite effect. Taken together our data suggest that MK-STYX may be a regulator of RhoA signaling, and

  13. Acyl-chain remodeling of dioctanoyl-phosphatidylcholine in Saccharomyces cerevisiae mutant defective in de novo and salvage phosphatidylcholine synthesis

    SciTech Connect

    Kishino, Hideyuki; Eguchi, Hiroki; Takagi, Keiko; Horiuchi, Hiroyuki; Fukuda, Ryouichi; Ohta, Akinori

    2014-03-07

    Highlights: • Dioctanoyl-PC (diC8PC) supported growth of a yeast mutant defective in PC synthesis. • diC8PC was converted to PC species containing longer acyl residues in the mutant. • Both acyl residues of diC8PC were replaced by longer fatty acids in vitro. • This system will contribute to the elucidation of the acyl chain remodeling of PC. - Abstract: A yeast strain, in which endogenous phosphatidylcholine (PC) synthesis is controllable, was constructed by the replacement of the promoter of PCT1, encoding CTP:phosphocholine cytidylyltransferase, with GAL1 promoter in a double deletion mutant of PEM1 and PEM2, encoding phosphatidylethanolamine methyltransferase and phospholipid methyltransferase, respectively. This mutant did not grow in the glucose-containing medium, but the addition of dioctanoyl-phosphatidylcholine (diC8PC) supported its growth. Analyses of the metabolism of {sup 13}C-labeled diC8PC ((methyl-{sup 13}C){sub 3}-diC8PC) in this strain using electrospray ionization tandem mass spectrometry revealed that it was converted to PC species containing acyl residues of 16 or 18 carbons at both sn-1 and sn-2 positions. In addition, both acyl residues of (methyl-{sup 13}C){sub 3}-diC8PC were replaced with 16:1 acyl chains in the in vitro reaction using the yeast cell extract in the presence of palmitoleoyl-CoA. These results indicate that PC containing short acyl residues was remodeled to those with acyl chains of physiological length in yeast.

  14. Spontaneous Pneumocystis carinii pneumonia in immunodeficient mutant scid mice. Natural history and pathobiology.

    PubMed Central

    Roths, J. B.; Marshall, J. D.; Allen, R. D.; Carlson, G. A.; Sidman, C. L.

    1990-01-01

    The opportunistic pathogen Pneumocystis carinii (Pc) poses a major clinical health problem in individuals with immune deficiency, including those patients with human immunodeficiency (HIV)-associated acquired immune deficiency disease (AIDS). Heretofore, in vivo investigations of the biology of Pc and pathogenesis of pneumocystosis have generally employed steroid-induced immune suppression with antibiotic prophylaxis and protein deprivation. This approach has many drawbacks, chief among them being the widespread, multiple interacting effects caused by the inducing agents. Athymic (nude) mice and rats have been used, but are less than ideal, as the immune defect primarily affects T lymphocytes. This article describes the natural history, pathobiology, and environmental effects on Pc pneumonitis in nonaxenically housed mice homozygous for the autosomal recessive mutation 'severe combined immunodeficiency' (scid), which almost totally lack both cell-mediated and antibody-mediated immune functions. The predictability, unequivocal expression, high morbidity, and well-defined genetic basis make scid/scid mutant mice the model of choice for in vivo studies of spontaneous pneumocystosis. Images Figure 3 Figure 6 PMID:2349968

  15. Spontaneous Pneumocystis carinii pneumonia in immunodeficient mutant scid mice. Natural history and pathobiology.

    PubMed

    Roths, J B; Marshall, J D; Allen, R D; Carlson, G A; Sidman, C L

    1990-05-01

    The opportunistic pathogen Pneumocystis carinii (Pc) poses a major clinical health problem in individuals with immune deficiency, including those patients with human immunodeficiency (HIV)-associated acquired immune deficiency disease (AIDS). Heretofore, in vivo investigations of the biology of Pc and pathogenesis of pneumocystosis have generally employed steroid-induced immune suppression with antibiotic prophylaxis and protein deprivation. This approach has many drawbacks, chief among them being the widespread, multiple interacting effects caused by the inducing agents. Athymic (nude) mice and rats have been used, but are less than ideal, as the immune defect primarily affects T lymphocytes. This article describes the natural history, pathobiology, and environmental effects on Pc pneumonitis in nonaxenically housed mice homozygous for the autosomal recessive mutation 'severe combined immunodeficiency' (scid), which almost totally lack both cell-mediated and antibody-mediated immune functions. The predictability, unequivocal expression, high morbidity, and well-defined genetic basis make scid/scid mutant mice the model of choice for in vivo studies of spontaneous pneumocystosis.

  16. Identifying representative drug resistant mutants of HIV

    PubMed Central

    2015-01-01

    Background Drug resistance is one of the most important causes for failure of anti-AIDS treatment. During therapy, multiple mutations accumulate in the HIV genome, eventually rendering the drugs ineffective in blocking replication of the mutant virus. The huge number of possible mutants precludes experimental analysis to explore the molecular mechanisms of resistance and develop improved antiviral drugs. Results In order to solve this problem, we have developed a new algorithm to reveal the most representative mutants from the whole drug resistant mutant database based on our newly proposed unified protein sequence and 3D structure encoding method. Mean shift clustering and multiple regression analysis were applied on genotype-resistance data for mutants of HIV protease and reverse transcriptase. This approach successfully chooses less than 100 mutants with the highest resistance to each drug out of about 10K in the whole database. When considering high level resistance to multiple drugs, the numbers reduce to one or two representative mutants. Conclusion This approach for predicting the most representative mutants for each drug has major importance for experimental verification since the results provide a small number of representative sequences, which will be amenable for in vitro testing and characterization of the expressed mutant proteins. PMID:26678327

  17. An equine herpesvirus 1 mutant with a lacZ insertion between open reading frames 62 and 63 is replication competent and causes disease in the murine respiratory model.

    PubMed

    Csellner, H; Walker, C; Love, D N; Whalley, J M

    1998-01-01

    An equine herpesvirus 1 (EHV-1) mutant was constructed by inserting a lacZ expression cassette into the intergenic region upstream of gene 62 (glycoprotein L; gL) and downstream of gene 63 (a homologue of the herpes simplex virus transcriptional activator ICP0). The recombinant lacZ62/63-EHV-1 had similar growth kinetics in cell culture to those of the parental wild type (wt) virus, with indistinguishable cytopathic effects and plaque morphology. Reverse transcriptase PCR confirmed that the lacZ insertion did not interfere with transcription of gL and immunoblot analysis indicated there was no modification to late gene expression as monitored by synthesis of EHV-1 glycoproteins C and D. The parental EHV-1 isolate HVS25A used here had almost identical nucleotide sequence to that published for isolate Ab4, in a 1200 bp region surrounding the insert, but lacked a HindIII site corresponding to Ab4 position 109,048. The lacZ62/63-EHV-1 caused respiratory disease in BALB/c mice with clinical signs, histopathology and virus titres in lungs throughout days 1-5 post infection similar to those induced by wt EHV-1. X-gal staining for beta-galactosidase expression in murine lungs clearly demonstrated EHV-1 infection in cells of the bronchiolar epithelium and pulmonary parenchyma, with a peak of infection evident at day 2 post infection, when up to 50% of bronchioles demonstrated blue-staining and thus virus-infected epithelial cells. The construction of this replication competent virus carrying a reporter gene identifies a site for insertion of foreign genes and will facilitate studies on the pathogenesis of EHV-1.

  18. Cadmium-sensitive, cad1 mutants of Arabidopsis thaliana are phytochelatin deficient.

    PubMed Central

    Howden, R; Goldsbrough, P B; Andersen, C R; Cobbett, C S

    1995-01-01

    An allelic series of cad1, cadmium-sensitive mutants of Arabidopsis thaliana, was isolated. These mutants were sensitive to cadmium to different extents and were deficient in their ability to form cadmium-peptide complexes as detected by gel-filtration chromatography. Each mutant was deficient in its ability to accumulate phytochelatins (PCs) as detected by high-performance liquid chromatography and the amount of PCs accumulated by each mutant correlated with its degree of sensitivity to cadmium. The mutants had wild-type levels of glutathione, the substrate for PC biosynthesis, and in vitro assays demonstrated that each of the mutants was deficient in PC synthase activity. These results demonstrate conclusively the importance of PCs for cadmium tolerance in plants. PMID:7770517

  19. Mutation in cpsf6/CFIm68 (Cleavage and Polyadenylation Specificity Factor Subunit 6) causes short 3'UTRs and disturbs gene expression in developing embryos, as revealed by an analysis of primordial germ cell migration using the medaka mutant naruto.

    PubMed

    Sasado, Takao; Kondoh, Hisato; Furutani-Seiki, Makoto; Naruse, Kiyoshi

    2017-01-01

    Our previous studies analyzing medaka mutants defective in primordial germ cell (PGC) migration identified cxcr4b and cxcr7, which are both receptors of the chemokine sdf1/cxcl12, as key regulators of PGC migration. Among PGC migration mutants, naruto (nar) is unique in that the mutant phenotype includes gross morphological abnormalities of embryos, suggesting that the mutation affects a broader range of processes. A fine genetic linkage mapping and genome sequencing showed the nar gene encodes Cleavage and Polyadenylation Specificity Factor subunit 6 (CPSF6/CFIm68). CPSF6 is a component of the Cleavage Factor Im complex (CFIm) which plays a key role in pre-mRNA 3'-cleavage and polyadenylation. 3'RACE of sdf1a/b and cxcr7 transcripts in the mutant embryos indicated shorter 3'UTRs with poly A additions occurring at more upstream positions than wild-type embryos, suggesting CPSF6 functions to prevent premature 3'UTR cleavage. In addition, expression of the coding region sequences of sdf1a/b in nar mutants was more anteriorly extended in somites than wild-type embryos, accounting for the abnormally extended distribution of PGCs in nar mutants. An expected consequence of shortening 3'UTR is the escape from the degradation mechanism mediated by microRNAs interacting with distal 3'UTR sequence. The abnormal expression pattern of sdf1a coding sequence may be at least partially accounted for by this mechanism. Given the pleiotropic effects of nar mutation, further analysis using the nar mutant will reveal processes in which CPSF6 plays essential regulatory roles in poly A site selection and involvement of 3'UTRs in posttranscriptional gene regulation in various genes in vivo.

  20. Mutation in cpsf6/CFIm68 (Cleavage and Polyadenylation Specificity Factor Subunit 6) causes short 3'UTRs and disturbs gene expression in developing embryos, as revealed by an analysis of primordial germ cell migration using the medaka mutant naruto

    PubMed Central

    Kondoh, Hisato; Furutani-Seiki, Makoto; Naruse, Kiyoshi

    2017-01-01

    Our previous studies analyzing medaka mutants defective in primordial germ cell (PGC) migration identified cxcr4b and cxcr7, which are both receptors of the chemokine sdf1/cxcl12, as key regulators of PGC migration. Among PGC migration mutants, naruto (nar) is unique in that the mutant phenotype includes gross morphological abnormalities of embryos, suggesting that the mutation affects a broader range of processes. A fine genetic linkage mapping and genome sequencing showed the nar gene encodes Cleavage and Polyadenylation Specificity Factor subunit 6 (CPSF6/CFIm68). CPSF6 is a component of the Cleavage Factor Im complex (CFIm) which plays a key role in pre-mRNA 3'-cleavage and polyadenylation. 3'RACE of sdf1a/b and cxcr7 transcripts in the mutant embryos indicated shorter 3’UTRs with poly A additions occurring at more upstream positions than wild-type embryos, suggesting CPSF6 functions to prevent premature 3’UTR cleavage. In addition, expression of the coding region sequences of sdf1a/b in nar mutants was more anteriorly extended in somites than wild-type embryos, accounting for the abnormally extended distribution of PGCs in nar mutants. An expected consequence of shortening 3'UTR is the escape from the degradation mechanism mediated by microRNAs interacting with distal 3’UTR sequence. The abnormal expression pattern of sdf1a coding sequence may be at least partially accounted for by this mechanism. Given the pleiotropic effects of nar mutation, further analysis using the nar mutant will reveal processes in which CPSF6 plays essential regulatory roles in poly A site selection and involvement of 3'UTRs in posttranscriptional gene regulation in various genes in vivo. PMID:28253363

  1. Loss of the Ciliary Kinase Nek8 Causes Left-Right Asymmetry Defects

    PubMed Central

    Manning, Danielle K.; Sergeev, Mikhail; van Heesbeen, Roy G.; Wong, Michael D.; Oh, Jin-Hee; Liu, Yan; Henkelman, R. Mark; Drummond, Iain; Shah, Jagesh V.

    2012-01-01

    A missense mutation in mouse Nek8, which encodes a ciliary kinase, produces the juvenile cystic kidneys (jck) model of polycystic kidney disease, but the functions of Nek8 are incompletely understood. Here, we generated a Nek8-null allele and found that homozygous mutant mice die at birth and exhibit randomization of left-right asymmetry, cardiac anomalies, and glomerular kidney cysts. The requirement for Nek8 in left-right patterning is conserved, as knockdown of the zebrafish ortholog caused randomized heart looping. Ciliogenesis was intact in Nek8-deficient embryos and cells, but we observed misexpression of left-sided marker genes early in development, suggesting that nodal ciliary signaling was perturbed. We also generated jck/Nek8 compound heterozygotes; these mutants developed less severe cystic disease than jck homozygotes and provided genetic evidence that the jck allele may encode a gain-of-function protein. Notably, NEK8 and polycystin-2 (PC2) proteins interact, and we found that Nek8−/− and Pkd2−/− embryonic phenotypes are strikingly similar. Nek8-deficient embryos and cells did express PC2 normally, which localized properly to the cilia. However, similar to cells lacking PC2, NEK8-depleted inner medullary collecting duct cells exhibited a defective response to fluid shear, suggesting that NEK8 may play a role in mediating PC2-dependent signaling. PMID:23274954

  2. Functional Analysis of Mouse G6pc1 Mutations Using a Novel In Situ Assay for Glucose-6-Phosphatase Activity and the Effect of Mutations in Conserved Human G6PC1/G6PC2 Amino Acids on G6PC2 Protein Expression

    PubMed Central

    Boortz, Kayla A.; Syring, Kristen E.; Pound, Lynley D.; Wang, Yingda; Oeser, James K.; O’Brien, Richard M.

    2016-01-01

    Elevated fasting blood glucose (FBG) has been associated with increased risk for development of type 2 diabetes. Single nucleotide polymorphisms (SNPs) in G6PC2 are the most important common determinants of variations in FBG in humans. Studies using G6pc2 knockout mice suggest that G6pc2 regulates the glucose sensitivity of insulin secretion. G6PC2 and the related G6PC1 and G6PC3 genes encode glucose-6-phosphatase catalytic subunits. This study describes a functional analysis of 22 non-synonymous G6PC2 SNPs, that alter amino acids that are conserved in human G6PC1, mouse G6pc1 and mouse G6pc2, with the goal of identifying variants that potentially affect G6PC2 activity/expression. Published data suggest strong conservation of catalytically important amino acids between all four proteins and the related G6PC3 isoform. Because human G6PC2 has very low glucose-6-phosphatase activity we used an indirect approach, examining the effect of these SNPs on mouse G6pc1 activity. Using a novel in situ functional assay for glucose-6-phosphatase activity we demonstrate that the amino acid changes associated with the human G6PC2 rs144254880 (Arg79Gln), rs149663725 (Gly114Arg) and rs2232326 (Ser324Pro) SNPs reduce mouse G6pc1 enzyme activity without affecting protein expression. The Arg79Gln variant alters an amino acid mutation of which, in G6PC1, has previously been shown to cause glycogen storage disease type 1a. We also demonstrate that the rs368382511 (Gly8Glu), rs138726309 (His177Tyr), rs2232323 (Tyr207Ser) rs374055555 (Arg293Trp), rs2232326 (Ser324Pro), rs137857125 (Pro313Leu) and rs2232327 (Pro340Leu) SNPs confer decreased G6PC2 protein expression. In summary, these studies identify multiple G6PC2 variants that have the potential to be associated with altered FBG in humans. PMID:27611587

  3. Functional Analysis of Mouse G6pc1 Mutations Using a Novel In Situ Assay for Glucose-6-Phosphatase Activity and the Effect of Mutations in Conserved Human G6PC1/G6PC2 Amino Acids on G6PC2 Protein Expression.

    PubMed

    Boortz, Kayla A; Syring, Kristen E; Pound, Lynley D; Wang, Yingda; Oeser, James K; O'Brien, Richard M

    2016-01-01

    Elevated fasting blood glucose (FBG) has been associated with increased risk for development of type 2 diabetes. Single nucleotide polymorphisms (SNPs) in G6PC2 are the most important common determinants of variations in FBG in humans. Studies using G6pc2 knockout mice suggest that G6pc2 regulates the glucose sensitivity of insulin secretion. G6PC2 and the related G6PC1 and G6PC3 genes encode glucose-6-phosphatase catalytic subunits. This study describes a functional analysis of 22 non-synonymous G6PC2 SNPs, that alter amino acids that are conserved in human G6PC1, mouse G6pc1 and mouse G6pc2, with the goal of identifying variants that potentially affect G6PC2 activity/expression. Published data suggest strong conservation of catalytically important amino acids between all four proteins and the related G6PC3 isoform. Because human G6PC2 has very low glucose-6-phosphatase activity we used an indirect approach, examining the effect of these SNPs on mouse G6pc1 activity. Using a novel in situ functional assay for glucose-6-phosphatase activity we demonstrate that the amino acid changes associated with the human G6PC2 rs144254880 (Arg79Gln), rs149663725 (Gly114Arg) and rs2232326 (Ser324Pro) SNPs reduce mouse G6pc1 enzyme activity without affecting protein expression. The Arg79Gln variant alters an amino acid mutation of which, in G6PC1, has previously been shown to cause glycogen storage disease type 1a. We also demonstrate that the rs368382511 (Gly8Glu), rs138726309 (His177Tyr), rs2232323 (Tyr207Ser) rs374055555 (Arg293Trp), rs2232326 (Ser324Pro), rs137857125 (Pro313Leu) and rs2232327 (Pro340Leu) SNPs confer decreased G6PC2 protein expression. In summary, these studies identify multiple G6PC2 variants that have the potential to be associated with altered FBG in humans.

  4. PC/104 Embedded IOCs at Jefferson Lab

    SciTech Connect

    Jianxun Yan, Trent Allison, Sue Witherspoon, Anthony Cuffe

    2009-10-01

    Jefferson Lab has developed embedded IOCs based on PC/104 single board computers (SBC) for low level control systems. The PC/104 IOCs run EPICS on top of the RTEMS operating system. Two types of control system configurations are used in different applications, PC/104 SBC with commercial PC/104 I/O cards and PC/104 SBC with custom designed FPGA-based boards. RTEMS was built with CEXP shell to run on the PC/104 SBC. CEXP shell provides the function of dynamic object loading, which is similar to the widely used VxWorks operating system. Standard software configurations were setup for PC/104 IOC application development to provide a familiar format for new projects as well as ease the conversion of applications from VME based IOCs to PC/104 IOCs. Many new projects at Jefferson Lab are going to employ PC/104 SBCs as IOCs and some applications have already been running them for accelerator operations. The PC/104 - RTEMS IOC provides a free open source Real-Time Operating System (RTOS), low cost/maintenance, easily installed/ configured, flexible, and reliable solution for accelerator control and 12GeV Upgrade projects.

  5. HBV genotypes prevalence, precore and basal core mutants in Morocco.

    PubMed

    Baha, Warda; Ennaji, My Mustapha; Lazar, Fatiha; Melloul, Marouane; El Fahime, Elmostafa; El Malki, Abdelouahad; Bennani, Abdelouaheb

    2012-08-01

    The study of hepatitis B virus (HBV) genomic heterogeneity has become a major issue in investigations aimed at understanding the relationship between HBV mutants and the wide spectrum of clinical and pathological conditions associated with HBV infection. The objective of the current study was to find out the pattern of HBV genotypes circulating in Morocco and to investigate the precore (PC) and basal core promoter (BCP) mutants' status in Moroccan chronic hepatitis B patients. Viral genotypes were determined in 221 chronic carriers using INNO-LiPA HBV assay and hemi-nested PCR. Phylogenetic analysis was performed in 70 samples, and multiplex PCR method was used to confirm some genotyping results. PC and CP mutants were determined using Inno-Lipa. All isolates were successfully genotyped. The genotype distribution was D in 90.45% of cases, A (5.9%), E (1 case), and mixed genotypes (5 A/D and 2 D/F) in 3.17% patients. HBV carried in the HBV/D samples could be assigned to D7 (63.3%), D1 (32.7%) and 2% of strains to each D4 and D5, all HBV/A belonged to A2 subgenotype and HBV/E strain could not be sub-genotyped. In 70 studied strains, HBV mutants were detected in 88.6% of cases; PC mutants were detected in (40%) of patients and 21.5% present a mixture of wild type and G1896A mutation. BCP mutants were observed in 65.7% of cases, 22.9% were found to have the T1762/1764A double mutation, 18.6% had A1762/1764T mutation and 22.9% of patients showed the A1762T/G1764A double mutation with either A1762T/G1764T mutation. Co-infection by PC and BCP mutants was detected in 52.9% of cases. Movement from place to place most likely shapes the observed genotype distribution and consequent prevalence of genotypes other than A2 or D7 in this population. High circulation of PC and BCP mutants is common in chronic hepatitis B infection in Morocco.

  6. Saccharomyces cerevisiae aldolase mutants.

    PubMed Central

    Lobo, Z

    1984-01-01

    Six mutants lacking the glycolytic enzyme fructose 1,6-bisphosphate aldolase have been isolated in the yeast Saccharomyces cerevisiae by inositol starvation. The mutants grown on gluconeogenic substrates, such as glycerol or alcohol, and show growth inhibition by glucose and related sugars. The mutations are recessive, segregate as one gene in crosses, and fall in a single complementation group. All of the mutants synthesize an antigen cross-reacting to the antibody raised against yeast aldolase. The aldolase activity in various mutant alleles measured as fructose 1,6-bisphosphate cleavage is between 1 to 2% and as condensation of triose phosphates to fructose 1,6-bisphosphate is 2 to 5% that of the wild-type. The mutants accumulate fructose 1,6-bisphosphate from glucose during glycolysis and dihydroxyacetone phosphate during gluconeogenesis. This suggests that the aldolase activity is absent in vivo. PMID:6384192

  7. The deubiquitinating enzyme UBPy/USP8 interacts with TrkA and inhibits neuronal differentiation in PC12 cells.

    PubMed

    Ceriani, Michela; Amigoni, Loredana; D'Aloia, Alessia; Berruti, Giovanna; Martegani, Enzo

    2015-04-10

    The tropomyosin-related kinase (Trk) family of receptor tyrosine kinases controls synaptic function, plasticity and sustains differentiation, morphology, and neuronal cell survival. Understanding Trk receptors down-regulation and recycling is a crucial step to point out sympathetic and sensory neuron function and survival. PC12 cells derived from pheochromocytoma of the rat adrenal medulla have been widely used as a model system for studies of neuronal differentiation as they respond to nerve growth factor (NGF) with a dramatic change in phenotype and acquire a number of properties characteristic of sympathetic neurons. In this study we demonstrated that in PC12 cells the TrkA receptor interacts with the deubiquitinating enzyme USP8/UBPy in a NGF-dependent manner and that it is deubiquitinated in vivo and in vitro by USP8. USP8 overexpression blocked NGF-induced neurites outgrowth while the overexpression of the catalytically inactive mutant USP8/UBPy(C748A) caused a marked increase of cell differentiation. Localization and biochemical experiments have point out that USP8 and TrkA partially co-localize in endosomes after NGF stimulation. Finally we have studied the role played by USP8 on TrkA turnover; using specific siRNA for USP8 we found that USP8 knockdown increases TrkA half-life, suggesting that the deubiquitinating activity of USP8 promotes TrkA degradation.

  8. AMPS/PC - AUTOMATIC MANUFACTURING PROGRAMMING SYSTEM

    NASA Technical Reports Server (NTRS)

    Schroer, B. J.

    1994-01-01

    The AMPS/PC system is a simulation tool designed to aid the user in defining the specifications of a manufacturing environment and then automatically writing code for the target simulation language, GPSS/PC. The domain of problems that AMPS/PC can simulate are manufacturing assembly lines with subassembly lines and manufacturing cells. The user defines the problem domain by responding to the questions from the interface program. Based on the responses, the interface program creates an internal problem specification file. This file includes the manufacturing process network flow and the attributes for all stations, cells, and stock points. AMPS then uses the problem specification file as input for the automatic code generator program to produce a simulation program in the target language GPSS. The output of the generator program is the source code of the corresponding GPSS/PC simulation program. The system runs entirely on an IBM PC running PC DOS Version 2.0 or higher and is written in Turbo Pascal Version 4 requiring 640K memory and one 360K disk drive. To execute the GPSS program, the PC must have resident the GPSS/PC System Version 2.0 from Minuteman Software. The AMPS/PC program was developed in 1988.

  9. Different Phenotypes of the Two Chinese Probands with the Same c.889G>A (p.C162Y) Mutation in COCH Gene Verify Different Mechanisms Underlying Autosomal Dominant Nonsyndromic Deafness 9

    PubMed Central

    Wang, Qi; Fei, Peipei; Gu, Hongbo; Zhang, Yanmei; Ke, Xiaomei; Liu, Yuhe

    2017-01-01

    Objectives By analyzing the different phenotypes of two Chinese DFNA9 families with the same mutation located in the intervening region between the LCCL and vWFA domains of cochlin and testing the functional changes in the mutant cochlin, we investigated the different pathogeneses for mutations in LCCL and vWFA domains. Methods Targeted next-generation sequencing for deafness-related genes was used to identify the mutation in the proband in family #208. The probands of family #208 and family #32 with the same p.C162Y mutation were followed for more than 3 years to evaluate the progression of hearing loss and vestibular dysfunction using pure-tone audiometry, caloric testing, electrocochleogram, vestibular-evoked myogenic potential, and video head-impulse test. The disruption of normal cleavage to produce secreted LCCL domain fragments and the tendency to form aggregations of mutant cochlins were tested by in vitro cell experiments. Results The two families showed different clinical symptoms. Family #32 was identified as having early-onset, progressive sensorineural hearing loss, similar to the symptoms in DFNA9 patients with cochlin mutations in the vWFA domain. The proband of family #208 endured late-onset recurrent paroxysmal vertigo attacks and progressively deteriorating hearing, similar to symptoms in those with cochlin mutations in the LCCL domain. We therefore suggest that the disrupted cleavage of the LCCL domain fragment is likely to cause vestibular dysfunction, and aggregation of mutant cochlin caused by mutations in the vWFA domain is responsible for early-onset hearing loss. The p.C162Y mutation causes either disruption of LCCL domain fragment cleavage or aggregation of mutant cochlin, resulting in the different phenotypes in the two families. Conclusion This study demonstrates that DFNA9 families with the same genotype may have significantly different phenotypes. The mutation site in cochlin is related to the pathological mechanism underlying the

  10. Understanding the muscular dystrophy caused by deletion of choline kinase beta in mice.

    PubMed

    Wu, Gengshu; Sher, Roger B; Cox, Gregory A; Vance, Dennis E

    2009-05-01

    Choline kinase in mice is encoded by two genes, Chka and Chkb. Disruption of murine Chka leads to embryonic lethality, whereas a spontaneously occurring genomic deletion in murine Chkb results in neonatal bone deformity and hindlimb muscular dystrophy. We have investigated the mechanism by which a lack of choline kinase beta, encoded by Chkb, causes hindlimb muscular dystrophy. The biosynthesis of phosphatidylcholine (PC) is impaired in the hindlimbs of Chkb -/- mice, with an accumulation of choline and decreased amount of phosphocholine. The activity of CTP: phosphocholine cytidylyltransferase is also decreased in the hindlimb muscle of mutant mice. Concomitantly, the activities of PC phospholipase C and phospholipase A2 are increased. The mitochondria in Chkb -/- mice are abnormally large and exhibit decreased inner membrane potential. Despite the muscular dystrophy in Chkb -/- mice, we observed increased expression of insulin like growth factor 1 and proliferating cell nuclear antigen. However, regeneration of hindlimb muscles of Chkb -/- mice was impaired when challenged with cardiotoxin. Injection of CDP-choline increased PC content of hindlimb muscle and decreased creatine kinase activity in plasma of Chkb -/- mice. We conclude that the hindlimb muscular dystrophy in Chkb -/- mice is due to attenuated PC biosynthesis and enhanced catabolism of PC.

  11. PC-based fault finder

    SciTech Connect

    Bengiamin, N.N. ); Jensen, C.A. . Electrical Engineering Dept. Otter Tail Power Co., Fergus Falls, MN . System Protection Group); McMahon, H. )

    1993-07-01

    Electric utilities are continually pressed to stay competitive while meeting the increasing demand of today's sophisticated customer. Advances in electron equipment and the improved array of electric driven devices are setting new standards for improved reliability and quality of service. Besides the specifications on voltage and frequency regulation and the permitted harmonic content, to name a few, the number and duration of service interruptions have a dramatic direct effect on the customer. Accurate fault locating reduces transmission line patrolling and is of particular significance in repairing long lines in rough terrain. Shortened outage times, reduced equipment degrading and stress on the system, fast restored service, and improved revenue are immediate outcomes of fast fault locating which insure minimum loss of system security. This article focuses on a PC-based (DOS) computer program that has unique features for identifying the type of fault and its location on overhead transmission/distribution lines. Balanced and unbalanced faults are identified and located accurately while accounting for changes in conductor sizes and network configuration. The presented concepts and methodologies have been spurred by Otter Tail Power's need for an accurate fault locating scheme to accommodate multiple feeders with mixed lone configurations. A case study based on a section of the Otter Tail network is presented to illustrate the features and capabilities of the developed software.

  12. Effect of K doping on CuPc: C60 heterojunctions

    NASA Astrophysics Data System (ADS)

    Cheng, Chiu-Ping; Chen, Wen-Yen; Wei, Ching-Hsuan; Pi, Tun-Wen

    2011-12-01

    Here, the electronic properties of K-doped copper phthalocyanine (CuPc): C60 heterojunctions are studied via synchrotron-radiation photoemission. The K-doped heterointerfaces were obtained by means of C60 on K1.5CuPc and CuPc on K3C60. The photoelectron spectra show that the potassium prefers to combine with C60. At the C60/K1.5CuPc interface, the K diffuses and transfers negative charge into the C60 overlayer, while no strong chemical reaction could be found at the CuPc/K3C60 interface. A significant shift of the vacuum level was observed in both cases, which was caused by the charge transfer for the C60/K1.5CuPc and by the induced density of interface states (IDIS) dipole for the CuPc/K3C60. The energy level diagrams show that using C60 adsorption on a K-doped CuPc film is good for the improvement of photovoltaic devices. However, the inverse process, that of CuPc on a K-doped C60, is unfavorable for the photovoltaic effect.

  13. Endonuclease IV (nfo) mutant of Escherichia coli.

    PubMed Central

    Cunningham, R P; Saporito, S M; Spitzer, S G; Weiss, B

    1986-01-01

    A cloned gene, designated nfo, caused overproduction of an EDTA-resistant endonuclease specific for apurinic-apyrimidinic sites in DNA. The sedimentation coefficient of the enzyme was similar to that of endonuclease IV. An insertion mutation was constructed in vitro and transferred from a plasmid to the Escherichia coli chromosome. nfo mutants had an increased sensitivity to the alkylating agents methyl methanesulfonate and mitomycin C and to the oxidants tert-butyl hydroperoxide and bleomycin. The nfo mutation enhanced the killing of xth (exonuclease III) mutants by methyl methanesulfonate, H2O2, tert-butyl hydroperoxide, and gamma rays, and it enhanced their mutability by methyl methanesulfonate. It also increased the temperature sensitivity of an xth dut (dUTPase) mutant that is defective in the repair of uracil-containing DNA. These results are consistent with earlier findings that endonuclease IV and exonuclease III both cleave DNA 5' to an apurinic-apyrimidinic site and that exonuclease III is more active. However, nfo mutants were more sensitive to tert-butyl hydroperoxide and to bleomycin than were xth mutants, suggesting that endonuclease IV might recognize some lesions that exonuclease III does not. The mutants displayed no marked increase in sensitivity to 254-nm UV radiation, and the addition of an nth (endonuclease III) mutation to nfo or nfo xth mutants did not significantly increase their sensitivity to any of the agents tested. Images PMID:2430946

  14. The mutant phenotype associated with P-element alleles of the vestigial locus in Drosophila melanogaster may be caused by a readthrough transcript initiated at the P-element promoter.

    PubMed

    Hodgetts, R B; O'Keefe, S L

    2001-04-01

    We report here the isolation of a new P-element-induced allele of the vestigial locus vg(2a33), the molecular characterization of which allows us to propose a unifying explanation of the phenotypes of the large number of vestigial P-element alleles that now exists. The first P-element allele of vestigial to be isolated was vg(21), which results in a very weak mutant wing phenotype that is suppressed in the P cytotype. By destabilizing vg(2a33) in a dysgenic cross, we isolated the vg(2a33) allele, which exhibits a moderate mutant wing phenotype and is not suppressed by the P cytotype. The new allele is characterized by a 46-bp deletion that removes the 3'-proximal copy of the 11-bp internal repeat from the P element of vg(21). To understand how this subtle difference between the two alleles leads to a rather pronounced difference in their phenotypes, we mapped both the vg and P-element transcription units present in wild type and mutants. Using both 5'-RACE and S1 protection, we found that P-element transcription is initiated 19 bp farther upstream than previously thought. Using primer extension, the start of vg transcription was determined to lie 435 bp upstream of the longest cDNA recovered to date and upstream of the P-element insertion site. Our discovery that the P element is situated within the first vg exon has prompted a reassessment of the large body of genetic data on a series of alleles derived from vg(21). Our current hypothesis to explain the degree of variation in the mutant phenotypes and their response to the P repressor invokes a critical RNA secondary structure in the vg transcript, the formation of which is hindered by a readthrough transcript initiated at the P-element promoter.

  15. Propagation mechanism of daytime Pc 3-4 pulsations observed at synchronous orbit and multiple ground-based stations

    NASA Technical Reports Server (NTRS)

    Yumoto, K.; Saito, T.; Akasofu, S.-I.; Tsurutani, B. T.; Smith, E. J.

    1985-01-01

    Observational data obtained during the last two decades show that the amplitude of daytime Pc 3-4 magnetic pulsations is controlled by the solar wind conditions. The high degree of correlation between the solar wind parameters and Pc 3-4 pulsations in the dayside magnetosphere suggests that the ultimate cause of the daytime Pc 3-4 pulsations must be the interaction of the solar wind with the earth's magnetosphere. The present paper is concerned with details regarding the control of the properties of the Pc 3-4 pulsations by the solar wind parameters, taking into account observations made at multiple ground-based stations. It is attempted to establish the relation between the daytime Pc 3-4 pulsations at the ground stations and the compressional Pc 3-4 waves in the magnetosphere. Attention is given to the most probable propagation mechanism of the daytime Pc 3-4 pulsations in the magnetosphere.

  16. Drosophila melanogaster dHCF interacts with both PcG and TrxG epigenetic regulators.

    PubMed

    Rodriguez-Jato, Sara; Busturia, Ana; Herr, Winship

    2011-01-01

    Repression and activation of gene transcription involves multiprotein complexes that modify chromatin structure. The integration of these complexes at regulatory sites can be assisted by co-factors that link them to DNA-bound transcriptional regulators. In humans, one such co-factor is the herpes simplex virus host-cell factor 1 (HCF-1), which is implicated in both activation and repression of transcription. We show here that disruption of the gene encoding the Drosophila melanogaster homolog of HCF-1, dHCF, leads to a pleiotropic phenotype involving lethality, sterility, small size, apoptosis, and morphological defects. In Drosophila, repressed and activated transcriptional states of cell fate-determining genes are maintained throughout development by Polycomb Group (PcG) and Trithorax Group (TrxG) genes, respectively. dHCF mutant flies display morphological phenotypes typical of TrxG mutants and dHCF interacts genetically with both PcG and TrxG genes. Thus, dHCF inactivation enhances the mutant phenotypes of the Pc PcG as well as brm and mor TrxG genes, suggesting that dHCF possesses Enhancer of TrxG and PcG (ETP) properties. Additionally, dHCF interacts with the previously established ETP gene skd. These pleiotropic phenotypes are consistent with broad roles for dHCF in both activation and repression of transcription during fly development.

  17. Genome sequencing of the Trichoderma reesei QM9136 mutant identifies a truncation of the transcriptional regulator XYR1 as the cause for its cellulase-negative phenotype

    SciTech Connect

    Lichius, Alexander; Bidard, Frédérique; Buchholz, Franziska; Le Crom, Stéphane; Martin, Joel; Schackwitz, Wendy; Austerlitz, Tina; Grigoriev, Igor V; Baker, Scott E; Margeot, Antoine; Seiboth, Bernhard; Kubicek, Christian P

    2015-04-20

    Trichoderma reesei is the main industrial source of cellulases and hemicellulases required for the hydrolysis of biomass to simple sugars, which can then be used in the production of biofuels and biorefineries. The highly productive strains in use today were generated by classical mutagenesis. As byproducts of this procedure, mutants were generated that turned out to be unable to produce cellulases. In order to identify the mutations responsible for this inability, we sequenced the genome of one of these strains, QM9136, and compared it to that of its progenitor T. reesei QM6a.

  18. Scale-PC shielding analysis sequences

    SciTech Connect

    Bowman, S.M.

    1996-05-01

    The SCALE computational system is a modular code system for analyses of nuclear fuel facility and package designs. With the release of SCALE-PC Version 4.3, the radiation shielding analysis community now has the capability to execute the SCALE shielding analysis sequences contained in the control modules SAS1, SAS2, SAS3, and SAS4 on a MS- DOS personal computer (PC). In addition, SCALE-PC includes two new sequences, QADS and ORIGEN-ARP. The capabilities of each sequence are presented, along with example applications.

  19. Autism-associated R451C mutation in neuroligin3 leads to activation of the unfolded protein response in a PC12 Tet-On inducible system

    PubMed Central

    Ulbrich, Lisa; Favaloro, Flores Lietta; Trobiani, Laura; Marchetti, Valentina; Patel, Vruti; Pascucci, Tiziana; Comoletti, Davide; Marciniak, Stefan J.; De Jaco, Antonella

    2015-01-01

    Several forms of monogenic heritable autism spectrum disorders are associated with mutations in the neuroligin genes. The autism-linked substitution R451C in neuroligin3 induces local misfolding of its extracellular domain, causing partial retention in the ER (endoplasmic reticulum) of expressing cells. We have generated a PC12 Tet-On cell model system with inducible expression of wild-type or R451C neuroligin3 to investigate whether there is activation of the UPR (unfolded protein response) as a result of misfolded protein retention. As a positive control for protein misfolding, we also expressed the mutant G221R neuroligin3, which is known to be completely retained within the ER. Our data show that overexpression of either R451C or G221R mutant proteins leads to the activation of all three signalling branches of the UPR downstream of the stress sensors ATF6 (activating transcription factor 6), IRE1 (inositol-requiring enzyme 1) and PERK [PKR (dsRNA-dependent protein kinase)-like endoplasmic reticulum kinase]. Each branch displayed different activation profiles that partially correlated with the degree of misfolding caused by each mutation. We also show that up-regulation of BiP (immunoglobulin heavy-chain-binding protein) and CHOP [C/EBP (CCAAT/enhancer-binding protein)-homologous protein] was induced by both mutant proteins but not by wild-type neuroligin3, both in proliferative cells and cells differentiated to a neuron-like phenotype. Collectively, our data show that mutant R451C neuroligin3 activates the UPR in a novel cell model system, suggesting that this cellular response may have a role in monogenic forms of autism characterized by misfolding mutations. PMID:26621873

  20. Autism-associated R451C mutation in neuroligin3 leads to activation of the unfolded protein response in a PC12 Tet-On inducible system.

    PubMed

    Ulbrich, Lisa; Favaloro, Flores Lietta; Trobiani, Laura; Marchetti, Valentina; Patel, Vruti; Pascucci, Tiziana; Comoletti, Davide; Marciniak, Stefan J; De Jaco, Antonella

    2016-02-15

    Several forms of monogenic heritable autism spectrum disorders are associated with mutations in the neuroligin genes. The autism-linked substitution R451C in neuroligin3 induces local misfolding of its extracellular domain, causing partial retention in the ER (endoplasmic reticulum) of expressing cells. We have generated a PC12 Tet-On cell model system with inducible expression of wild-type or R451C neuroligin3 to investigate whether there is activation of the UPR (unfolded protein response) as a result of misfolded protein retention. As a positive control for protein misfolding, we also expressed the mutant G221R neuroligin3, which is known to be completely retained within the ER. Our data show that overexpression of either R451C or G221R mutant proteins leads to the activation of all three signalling branches of the UPR downstream of the stress sensors ATF6 (activating transcription factor 6), IRE1 (inositol-requiring enzyme 1) and PERK [PKR (dsRNA-dependent protein kinase)-like endoplasmic reticulum kinase]. Each branch displayed different activation profiles that partially correlated with the degree of misfolding caused by each mutation. We also show that up-regulation of BiP (immunoglobulin heavy-chain-binding protein) and CHOP [C/EBP (CCAAT/enhancer-binding protein)-homologous protein] was induced by both mutant proteins but not by wild-type neuroligin3, both in proliferative cells and cells differentiated to a neuron-like phenotype. Collectively, our data show that mutant R451C neuroligin3 activates the UPR in a novel cell model system, suggesting that this cellular response may have a role in monogenic forms of autism characterized by misfolding mutations.

  1. Lamin A/C sustains PcG protein architecture, maintaining transcriptional repression at target genes

    PubMed Central

    Cesarini, Elisa; Mozzetta, Chiara; Marullo, Fabrizia; Gregoretti, Francesco; Gargiulo, Annagiusi; Columbaro, Marta; Cortesi, Alice; Antonelli, Laura; Di Pelino, Simona; Squarzoni, Stefano; Palacios, Daniela; Zippo, Alessio; Bodega, Beatrice; Oliva, Gennaro

    2015-01-01

    Beyond its role in providing structure to the nuclear envelope, lamin A/C is involved in transcriptional regulation. However, its cross talk with epigenetic factors—and how this cross talk influences physiological processes—is still unexplored. Key epigenetic regulators of development and differentiation are the Polycomb group (PcG) of proteins, organized in the nucleus as microscopically visible foci. Here, we show that lamin A/C is evolutionarily required for correct PcG protein nuclear compartmentalization. Confocal microscopy supported by new algorithms for image analysis reveals that lamin A/C knock-down leads to PcG protein foci disassembly and PcG protein dispersion. This causes detachment from chromatin and defects in PcG protein–mediated higher-order structures, thereby leading to impaired PcG protein repressive functions. Using myogenic differentiation as a model, we found that reduced levels of lamin A/C at the onset of differentiation led to an anticipation of the myogenic program because of an alteration of PcG protein–mediated transcriptional repression. Collectively, our results indicate that lamin A/C can modulate transcription through the regulation of PcG protein epigenetic factors. PMID:26553927

  2. Functional roles of PC-PLC and Cdc20 in the cell cycle, proliferation, and apoptosis.

    PubMed

    Chen, Zhiwei; Yu, Yongfeng; Fu, Da; Li, Ziming; Niu, Xiaoming; Liao, Meilin; Lu, Shun

    2010-06-01

    Phosphatidylcholine-specific phospholipase C (PC-PLC) is the major enzyme in the Phosphatidylcholine (PC) cycle and is involved in many long-term cellular responses such as activation, proliferation, and differentiation events. Cell division cycle 20 homolog (Cdc20) is an essential cell-cycle regulator required for the completion of mitosis. Our previous studies identified the interaction between PC-PLC and Cdc20. Through the interaction, Cdc20 could mediate the degradation of PC-PLC by Cdc20-mediated ubiquitin proteasome pathway (UPP). In this study, we found that PC-PLC might not be involved in cancer metastasis. Inhibition of PC-PLC by D609 could cause cell proliferation inhibition and apoptosis inhibition in CBRH-7919 cells. Inhibition of PC-PLC could also influence the cell cycle by arresting the cells in G1 phase, and Cdc20 might be involved in these processes. Taken together, in this report, we provided new evidence for the functional roles of PC-PLC and Cdc20 in the cell cycle, proliferation, and apoptosis in CBRH-7919 cells.

  3. Lamin A/C sustains PcG protein architecture, maintaining transcriptional repression at target genes.

    PubMed

    Cesarini, Elisa; Mozzetta, Chiara; Marullo, Fabrizia; Gregoretti, Francesco; Gargiulo, Annagiusi; Columbaro, Marta; Cortesi, Alice; Antonelli, Laura; Di Pelino, Simona; Squarzoni, Stefano; Palacios, Daniela; Zippo, Alessio; Bodega, Beatrice; Oliva, Gennaro; Lanzuolo, Chiara

    2015-11-09

    Beyond its role in providing structure to the nuclear envelope, lamin A/C is involved in transcriptional regulation. However, its cross talk with epigenetic factors--and how this cross talk influences physiological processes--is still unexplored. Key epigenetic regulators of development and differentiation are the Polycomb group (PcG) of proteins, organized in the nucleus as microscopically visible foci. Here, we show that lamin A/C is evolutionarily required for correct PcG protein nuclear compartmentalization. Confocal microscopy supported by new algorithms for image analysis reveals that lamin A/C knock-down leads to PcG protein foci disassembly and PcG protein dispersion. This causes detachment from chromatin and defects in PcG protein-mediated higher-order structures, thereby leading to impaired PcG protein repressive functions. Using myogenic differentiation as a model, we found that reduced levels of lamin A/C at the onset of differentiation led to an anticipation of the myogenic program because of an alteration of PcG protein-mediated transcriptional repression. Collectively, our results indicate that lamin A/C can modulate transcription through the regulation of PcG protein epigenetic factors.

  4. Regulation of phospholipid synthesis in phosphatidylserine synthase-deficient (chol) mutants of Saccharomyces cerevisiae.

    PubMed Central

    Letts, V A; Henry, S A

    1985-01-01

    chol mutants of Saccharomyces cerevisiae are deficient in the synthesis of the phospholipid phosphatidylserine owing to lowered activity of the membrane-associated enzyme phosphatidylserine synthase. chol mutants are auxotrophic for ethanolamine or choline and, in the absence of these supplements, cannot synthesize phosphatidylethanolamine or phosphatidylcholine (PC). We exploited these characteristics of the chol mutants to examine the regulation of phospholipid metabolism in S. cerevisiae. Macromolecular synthesis and phospholipid metabolism were examined in chol cells starved for ethanolamine. As expected, when chol mutants were starved for ethanolamine, the rates of synthesis of the phospholipids phosphatidylethanolamine and PC declined rapidly. Surprisingly, however, coupled to the decline in PC biosynthesis was a simultaneous decrease in the overall rate of phospholipid synthesis. In particular, the rate of synthesis of phosphatidylinositol decreased in parallel with the decline in PC biosynthesis. The results obtained suggest that the slowing of PC biosynthesis in ethanolamine-starved chol cells leads to a coordinated decrease in the synthesis of all phospholipids. However, under conditions of ethanolamine deprivation in chol cells, the cytoplasmic enzyme inositol-1-phosphate synthase could not be repressed by exogenous inositol, and the endogenous synthesis of the phospholipid precursor inositol appeared to be elevated. The implications of these findings with respect to the coordinated regulation of phospholipid synthesis are discussed. Images PMID:2991194

  5. Isolation and characterization of unusual gin mutants.

    PubMed Central

    Klippel, A; Cloppenborg, K; Kahmann, R

    1988-01-01

    Site-specific inversion of the G segment in phage Mu DNA is promoted by two proteins, the DNA invertase Gin and the host factor FIS. Recombination occurs if the recombination sites (IR) are arranged as inverted repeats and a recombinational enhancer sequence is present in cis. Intermolecular reactions as well as deletions between direct repeats of the IRs rarely occur. Making use of a fis- mutant of Escherichia coli we have devised a scheme to isolate gin mutants that have a FIS independent phenotype. This mutant phenotype is caused by single amino acid changes at five different positions of gin. The mutant proteins display a whole set of new properties in vivo: they promote inversions, deletions and intermolecular recombination in an enhancer- and FIS-independent manner. The mutants differ in recombination activity. The most active mutant protein was analysed in vitro. The loss of site orientation specificity was accompanied with the ability to recombine even linear substrates. We discuss these results in connection with the role of the enhancer and FIS protein in the wild-type situation. Images PMID:2974801

  6. Magnetospheric filter effect for Pc 3 Alfven mode waves

    NASA Technical Reports Server (NTRS)

    Zhang, X.; Comfort, R. H.; Gallagher, D. L.; Green, J. L.; Musielak, Z. E.; Moore, T. E.

    1995-01-01

    We present a ray-tracing study of the propagation of Pc 3 Alfven mode waves originating at the dayside magnetopause. This study reveals interesting features of magnetospheric filter effect for these waves. Pc 3 Alfven mode waves cannot penetrate to low Earth altitudes unless the wave frequency is below approximately 30 mHz. Configurations of the dispersion curves and the refractive index show that the gyroresonance and pseudo-cutoff introduced by the heavy ion O(+) block the waves. When the O(+) concentration is removed from the plasma composition, the barriers caused by the O(+) no longer exist, and waves with much higher frequencies than 30 mHz can penetrate to low altitudes. The result that the 30 mHz or lower frequency Alfven waves can be guided to low altitudes agrees with ground-based power spectrum observation at high altitudes.

  7. Magnetospheric filter effect for Pc 3 Alfven mode waves

    NASA Technical Reports Server (NTRS)

    Zhang, X.; Comfort, R. H.; Gallagher, D. L.; Green, J. L.; Musielak, Z. E.; Moore, T. E.

    1994-01-01

    We present a ray-tracing study of the propagation of Pc 3 Alfven mode waves originating at the dayside magnetopause. This study reveals interesting features of a magnetospheric filter effect for these waves. Pc 3 Alfven mode waves cannot penetrate to low Earth altitudes unless the wave frequency is below approximately 30 mHz. Configurations of the dispersion curves and the refractive index show that the gyroresonance and pseudo-cutoff introduced by the heavy ion O(+) block the waves. When the O(+) concentration is removed from the plasma composition, the barriers caused by the O(+) no longer exist, and waves with much higher frequencies than 30 mHz can penetrate to low altitudes. The result that the 30 mHz or lower frequency Alfven waves can be guided to low altitudes agrees with ground-based power spectrum observations at high latitudes.

  8. DIGLIB. PC-DOS Graphics Subroutine Library

    SciTech Connect

    Burleson, R.R.

    1989-02-01

    DIGLIB is a collection of general graphics subroutines. It was designed to be small, reasonably fast, device-independent, and compatible with DEC-supplied operating systems for VAXes, PDP-11s, and LSI-11s, and the DOS operating system for IBM PCs and IBM-compatible machines. DIGLIB/PC runs on IBM PCs under PC-DOS or MS-DOS. The software is readily usable by casual programmers for two-dimensional plotting.

  9. Mutant fatty acid desaturase

    DOEpatents

    Shanklin, John; Cahoon, Edgar B.

    2004-02-03

    The present invention relates to a method for producing mutants of a fatty acid desaturase having a substantially increased activity towards fatty acid substrates with chains containing fewer than 18 carbons relative to an unmutagenized precursor desaturase having an 18 carbon atom chain length substrate specificity. The method involves inducing one or more mutations in the nucleic acid sequence encoding the precursor desaturase, transforming the mutated sequence into an unsaturated fatty acid auxotroph cell such as MH13 E. coli, culturing the cells in the absence of supplemental unsaturated fatty acids, thereby selecting for recipient cells which have received and which express a mutant fatty acid desaturase with an elevated specificity for fatty acid substrates having chain lengths of less than 18 carbon atoms. A variety of mutants having 16 or fewer carbon atom chain length substrate specificities are produced by this method. Mutant desaturases produced by this method can be introduced via expression vectors into prokaryotic and eukaryotic cells and can also be used in the production of transgenic plants which may be used to produce specific fatty acid products.

  10. The catalytic domain CysPc of the DEK1 calpain is functionally conserved in land plants.

    PubMed

    Liang, Zhe; Demko, Viktor; Wilson, Robert C; Johnson, Kenneth A; Ahmad, Rafi; Perroud, Pierre-François; Quatrano, Ralph; Zhao, Sen; Shalchian-Tabrizi, Kamran; Otegui, Marisa S; Olsen, Odd-Arne; Johansen, Wenche

    2013-09-01

    DEK1, the single calpain of land plants, is a member of the ancient membrane bound TML-CysPc-C2L calpain family that dates back 1.5 billion years. Here we show that the CysPc-C2L domains of land plant calpains form a separate sub-clade in the DEK1 clade of the phylogenetic tree of plants. The charophycean alga Mesostigma viride DEK1-like gene is clearly divergent from those in land plants, suggesting that a major evolutionary shift in DEK1 occurred during the transition to land plants. Based on genetic complementation of the Arabidopsis thaliana dek1-3 mutant using CysPc-C2L domains of various origins, we show that these two domains have been functionally conserved within land plants for at least 450 million years. This conclusion is based on the observation that the CysPc-C2L domains of DEK1 from the moss Physcomitrella patens complements the A. thaliana dek1-3 mutant phenotype. In contrast, neither the CysPc-C2L domains from M. viride nor chimeric animal-plant calpains complement this mutant. Co-evolution analysis identified differences in the interactions between the CysPc-C2L residues of DEK1 and classical calpains, supporting the view that the two enzymes are regulated by fundamentally different mechanisms. Using the A. thaliana dek1-3 complementation assay, we show that four conserved amino acid residues of two Ca²⁺-binding sites in the CysPc domain of classical calpains are conserved in land plants and functionally essential in A. thaliana DEK1.

  11. Use of Topical PC-NSAIDs to Treat Burn Injury and Pain

    DTIC Science & Technology

    2015-08-01

    AWARD NUMBER: W81XWH-14-2-0016 TITLE: Use of Topical PC-NSAIDs to Treat Burn Injury and Pain PRINCIPAL INVESTIGATOR: Lenard Lichtenberger...TITLE AND SUBTITLE Use of Topical PC-NSAIDs to Treat Burn Injury and Pain 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-14-2-0016 5c. PROGRAM...for continued development of Indo-PC for treatment of acute pain caused by 2nd degree burn wounds. 15. SUBJECT TERMS Burn, phospholipid, NSAID

  12. Accumulation of Pol Mutations Selected by HLA-B*52:01-C*12:02 Protective Haplotype-Restricted Cytotoxic T Lymphocytes Causes Low Plasma Viral Load Due to Low Viral Fitness of Mutant Viruses.

    PubMed

    Murakoshi, Hayato; Koyanagi, Madoka; Chikata, Takayuki; Rahman, Mohammad Arif; Kuse, Nozomi; Sakai, Keiko; Gatanaga, Hiroyuki; Oka, Shinichi; Takiguchi, Masafumi

    2017-02-15

    HLA-B*52:01-C*12:02, which is the most abundant haplotype in Japan, has a protective effect on disease progression in HIV-1-infected Japanese individuals, whereas HLA-B*57 and -B*27 protective alleles are very rare in Japan. A previous study on HLA-associated polymorphisms demonstrated that the number of HLA-B*52:01-associated mutations at four Pol positions was inversely correlated with plasma viral load (pVL) in HLA-B*52:01-negative individuals, suggesting that the transmission of HIV-1 with these mutations could modulate the pVL in the population. However, it remains unknown whether these mutations were selected by HLA-B*52:01-restricted CTLs and also reduced viral fitness. In this study, we identified two HLA-B*52:01-restricted and one HLA-C*12:02-restricted novel cytotoxic T-lymphocyte (CTL) epitopes in Pol. Analysis using CTLs specific for these three epitopes demonstrated that these CTLs failed to recognize mutant epitopes or more weakly recognized cells infected with mutant viruses than wild-type virus, supporting the idea that these mutations were selected by the HLA-B*52:01- or HLA-C*12:02-restricted T cells. We further showed that these mutations reduced viral fitness, although the effect of each mutation was weak. The present study demonstrated that the accumulation of these Pol mutations selected by HLA-B*52:01- or HLA-C*12:02-restricted CTLs impaired viral replication capacity and thus reduced the pVL. The fitness cost imposed by the mutations partially accounted for the effect of the HLA-B*52:01-C*12:02 haplotype on clinical outcome, together with the effect of HLA-B*52:01-restricted CTLs on viral replication, which had been previously demonstrated.

  13. Cytotoxic Effects of the Ethanol Bane Skin Extract in Human Prostate Cancer Pc3 Cells

    PubMed Central

    Amiri, Maryam; Kazerouni, Faranak; Namaki, Saeed; Darbandi Tamijani, Hassan; Rahimipour, Hooman; Boroumand, Nasrin; Barghi, Siyamak; Ebrahimi, Nazanin; Gheibi Hayat, Seyed Mohammad

    2016-01-01

    Background: It is extensively supposed that vegetarian diet could affect cancer progress and increase the influence of formal chemotherapy. Objectives: The present study was designed to determine the effect of the ethanol Bane skin extract against chemo resistant prostate cancer PC3 cells. Materials and Methods: PC3 and L929 cells were cultivated and then incubated in the ethanol Bane skin extract with various concentrations of 0.78, 1.5, 3.13, 6.25, 12.5 mg/mL in 3 times 24, 48, 72 hours. Cytotoxic effect of the ethanol Bane skin extract on PC3 and L929 cells was examined by MTT assay after 24, 48, and 72 hours. Morphology of PC3 cells was evaluated by Gimsa staining. Results: The ethanol Bane skin extract inhibited proliferation and caused cell death with IC50 values of 2.8 mg/mL on PC3 cells and the IC50 was 6.1 mg/mL on l929 cells. Morphological changes and apoptotic bodies were observed in PC3 cells faced with the ethanol Bane skin extract by staining with Gimsa. Conclusions: The ethanol Bane skin extract could repress the growth of PC3 cell line. This inhibitory effect of the Bane extract depended on the dose and the time on PC3. The result of this study shows that the ethanol Bane skin extract includes photochemical and inhibitory function against proliferation and inducer of apoptosis in human prostate cancer PC3 cells and also has less cytotoxic effect on l929 than PC3 cells. The ethanol Bane skin extract might be a good candidate for the new herbal anticancer drug. PMID:27482333

  14. CARES/PC - CERAMICS ANALYSIS AND RELIABILITY EVALUATION OF STRUCTURES

    NASA Technical Reports Server (NTRS)

    Szatmary, S. A.

    1994-01-01

    The beneficial properties of structural ceramics include their high-temperature strength, light weight, hardness, and corrosion and oxidation resistance. For advanced heat engines, ceramics have demonstrated functional abilities at temperatures well beyond the operational limits of metals. This is offset by the fact that ceramic materials tend to be brittle. When a load is applied, their lack of significant plastic deformation causes the material to crack at microscopic flaws, destroying the component. CARES/PC performs statistical analysis of data obtained from the fracture of simple, uniaxial tensile or flexural specimens and estimates the Weibull and Batdorf material parameters from this data. CARES/PC is a subset of the program CARES (COSMIC program number LEW-15168) which calculates the fast-fracture reliability or failure probability of ceramic components utilizing the Batdorf and Weibull models to describe the effects of multi-axial stress states on material strength. CARES additionally requires that the ceramic structure be modeled by a finite element program such as MSC/NASTRAN or ANSYS. The more limited CARES/PC does not perform fast-fracture reliability estimation of components. CARES/PC estimates ceramic material properties from uniaxial tensile or from three- and four-point bend bar data. In general, the parameters are obtained from the fracture stresses of many specimens (30 or more are recommended) whose geometry and loading configurations are held constant. Parameter estimation can be performed for single or multiple failure modes by using the least-squares analysis or the maximum likelihood method. Kolmogorov-Smirnov and Anderson-Darling goodness-of-fit tests measure the accuracy of the hypothesis that the fracture data comes from a population with a distribution specified by the estimated Weibull parameters. Ninety-percent confidence intervals on the Weibull parameters and the unbiased value of the shape parameter for complete samples are provided

  15. Adapting PC104Plus for Space

    NASA Technical Reports Server (NTRS)

    Abbott, Larry; Cox, Gary; Nguyen, Hai

    2000-01-01

    This article addresses the issues associated with adapting the commercial PC104Plus standard and its associated architecture to the requirements of space applications. In general, space applications exhibit extreme constraints on power, weight, and volume. EMI and EMC are also issues of significant concern. Additionally, space applications have to survive high radiation environment. Finally, NASA is always concerned about achieving cost effective solutions that are compatible with safety and launch constraints. Weight and volume constraints are directly related to high launch cost. Power on the other hand is not only related to the high launch costs, but are related to the problem of dissipating the resulting heat once in space. The article addresses why PC104Plus is an appropriate solution for the weight and volume issues. The article also addresses what NASA did electrically to reduce power consumption and mechanically dissipate the associated heat in a microgravity and vacuum environment, and how these solutions allow NASA to integrate various sizes of ruggedized custom PC104 boards with COTS, PC104 complaint boards for space applications. In addition to the mechanical changes to deal with thermal dissipation NASA also made changes to minimize EMI. Finally, radiation issues are addressed as well as the architectural and testing solutions and the implications for use of COTS PC104Plus boards.

  16. APC-PC Combined Scheme in Gilbert Two State Model: Proposal and Study

    NASA Astrophysics Data System (ADS)

    Bulo, Yaka; Saring, Yang; Bhunia, Chandan Tilak

    2016-08-01

    In an automatic repeat request (ARQ) scheme, a packet is retransmitted if it gets corrupted due to transmission errors caused by the channel. However, an erroneous packet may contain both erroneous bits and correct bits and hence it may still contain useful information. The receiver may be able to combine this information from multiple erroneous copies to recover the correct packet. Packet combining (PC) is a simple and elegant scheme of error correction in transmitted packet, in which two received copies are XORed to obtain the bit location of erroneous bits. Thereafter, the packet is corrected by bit inversion of bit located as erroneous. Aggressive packet combining (APC) is a logic extension of PC primarily designed for wireless communication with objective of correcting error with low latency. PC offers higher throughput than APC, but PC does not correct double bit errors if occur in same bit location of erroneous copies of the packet. A hybrid technique is proposed to utilize the advantages of both APC and PC while attempting to remove the limitation of both. In the proposed technique, applications of APC-PC on Gilbert two state model has been studied. The simulation results show that the proposed technique offers better throughput than the conventional APC and lesser packet error rate than PC scheme.

  17. Broad spectral sensitivity and improved efficiency in CuPc/Sub-Pc organic photovoltaic devices

    NASA Astrophysics Data System (ADS)

    Kumar, Hemant; Kumar, Pankaj; Bhardwaj, Ramil; Sharma, G. D.; Chand, Suresh; Jain, S. C.; Kumar, Vikram

    2009-01-01

    We demonstrate organic photovoltaic devices incorporating two donors, namely, copper phthalocyanine (CuPc) and boron sub-phthalocyanine chloride (Sub-Pc) in association with single acceptor fullerene (C60) with sensitivity extending across the visible solar spectrum. It has been found that the absorption in different spectral regions in CuPc and Sub-Pc results in efficient harvesting of incident light photons which leads to enhanced power conversion efficiency (η). An enhancement in η from 0.64%, in the device architecture indium-tin-oxide (ITO)/CuPc(20 nm)/C60(40 nm)/bathophenanthroline (BPhen) (8 nm)/Al(150 nm), to ~1.3% in the optimized device having a 2 nm layer of Sub-Pc in the geometry ITO/CuPc(18 nm)/Sub-Pc(2 nm)/C60 (40 nm)/BPhen (8 nm)/Al(150 nm) has been observed. This enhancement in η is dominantly attributed to the increment in short circuit current density (Jsc) due to efficient photon harvesting by incorporation of dual donors.

  18. The IBM PC at NASA Ames

    NASA Technical Reports Server (NTRS)

    Peredo, James P.

    1988-01-01

    Like many large companies, Ames relies very much on its computing power to get work done. And, like many other large companies, finding the IBM PC a reliable tool, Ames uses it for many of the same types of functions as other companies. Presentation and clarification needs demand much of graphics packages. Programming and text editing needs require simpler, more-powerful packages. The storage space needed by NASA's scientists and users for the monumental amounts of data that Ames needs to keep demand the best database packages that are large and easy to use. Availability to the Micom Switching Network combines the powers of the IBM PC with the capabilities of other computers and mainframes and allows users to communicate electronically. These four primary capabilities of the PC are vital to the needs of NASA's users and help to continue and support the vast amounts of work done by the NASA employees.

  19. PcchiB1, encoding a class V chitinase, is affected by PcVelA and PcLaeA, and is responsible for cell wall integrity in Penicillium chrysogenum.

    PubMed

    Kamerewerd, Jens; Zadra, Ivo; Kürnsteiner, Hubert; Kück, Ulrich

    2011-11-01

    Penicillin production in Penicillium chrysogenum is controlled by PcVelA and PcLaeA, two components of the regulatory velvet-like complex. Comparative microarray analysis with mutants lacking PcVelA or PcLaeA revealed a set of 62 common genes affected by the loss of both components. A downregulated gene in both knockout strains is PcchiB1, potentially encoding a class V chitinase. Under nutrient-depleted conditions, transcript levels of PcchiB1 are strongly upregulated, and the gene product contributes to more than 50 % of extracellular chitinase activity. Functional characterization by generating PcchiB1-disruption strains revealed that PcChiB1 is responsible for cell wall integrity and pellet formation in P. chrysogenum. Further, fluorescence microscopy with a DsRed-labelled chitinase suggests a cell wall association of the protein. An unexpected phenotype occurred when knockout strains were grown on media containing N-acetylglucosamine as the sole C and N source, where, in contrast to the recipient, a penicillin producer strain, the mutants and an ancestral strain show distinct mycelial growth. We discuss the relevance of this class V chitinase for morphology in an industrially important fungus.

  20. Misfolded opsin mutants display elevated β-sheet structure.

    PubMed

    Miller, Lisa M; Gragg, Megan; Kim, Tae Gyun; Park, Paul S-H

    2015-10-07

    Mutations in rhodopsin can cause misfolding and aggregation of the receptor, which leads to retinitis pigmentosa, a progressive retinal degenerative disease. The structure adopted by misfolded opsin mutants and the associated cell toxicity is poorly understood. Förster resonance energy transfer (FRET) and Fourier transform infrared (FTIR) microspectroscopy were utilized to probe within cells the structures formed by G188R and P23H opsins, which are misfolding mutants that cause autosomal dominant retinitis pigmentosa. Both mutants formed aggregates in the endoplasmic reticulum and exhibited altered secondary structure with elevated β-sheet and reduced α-helical content. The newly formed β-sheet structure may facilitate the aggregation of misfolded opsin mutants. The effects observed for the mutants were unrelated to retention of opsin molecules in the endoplasmic reticulum itself.

  1. Misfolded opsin mutants display elevated β -sheet structure

    DOE PAGES

    Miller, Lisa M.; Gragg, Megan; Kim, Tae Gyun; ...

    2015-09-07

    Mutations in rhodopsin can cause misfolding and aggregation of the receptor, which leads to retinitis pigmentosa, a progressive retinal degenerative disease. The structure adopted by misfolded opsin mutants and the associated cell toxicity is poorly understood. Förster resonance energy transfer (FRET) and Fourier transform infrared (FTIR) microspectroscopy were utilized to probe within cells the structures formed by G188R and P23H opsins, which are misfolding mutants that cause autosomal dominant retinitis pigmentosa. Also, both mutants formed aggregates in the endoplasmic reticulum and exhibited altered secondary structure with elevated β-sheet and reduced α-helical content. The newly formed β-sheet structure may facilitate themore » aggregation of misfolded opsin mutants. In conclusion, the effects observed for the mutants were unrelated to retention of opsin molecules in the endoplasmic reticulum itself.« less

  2. Neurosecretory Habituation in PC12 Cells: Modulation During Parallel Habituation

    NASA Astrophysics Data System (ADS)

    Martin, Paul T.; Koshland, Daniel E., Jr.

    1995-05-01

    PC12 cells habituate during repetitive stimulation with acetylcholine, bradykinin, or high potassium. Interspersing these stimulants did not affect the rate of habituation of the others, but it could modulate the amplitude of the norepinephrine secretion each could achieve. Stimulation with acetylcholine inhibited norepinephrine secretion caused by high potassium and bradykinin stimulation, while high potassium had no effect on acetylcholine or bradykinin, and bradykinin increased secretion caused by acetylcholine. Changes in norepinephrine secretion resulting from any of these stimulants correlated with changes in internal calcium levels. Cyclic AMP-, protein kinase C-, and calmodulin-dependent second messenger pathways all modulated norepinephrine secretion caused by acetylcholine and high potassium and showed a distinct hierarchy in their effectiveness. These data demonstrate that different receptor pathways can change the norepinephrine response of one another while not changing the levels of the molecules responsible for habituation.

  3. Protective effects of ginsenoside Rg1 against colistin sulfate-induced neurotoxicity in PC12 cells.

    PubMed

    Jiang, Guo-Zheng; Li, Ji-Chang

    2014-03-01

    The present study aimed to examine the protective effect of ginsenoside Rg1 against colistin-induced neurotoxicity in cultured rat pheochromocytoma (PC12) cells. Ginsenoside Rg1 was shown to elevate cell viability, decrease levels of malondialdehyde and intracellular reactive oxygen species, enhance activity of superoxide dismutase and glutathione, and decrease the release of cytochrome-c, formation of DNA fragmentation in colistin-treated PC12 cells. Ginsenoside Rg1 also reversed the increased caspase-9 and -3 mRNA levels caused by colistin in PC12 cells. These results suggest that ginsenoside Rg1 exerts a neuroprotective effect on colistin-induced neurotoxicity in PC12 cells, at least in part, via the inhibition of oxidative stress, prevention of apoptosis mediated via mitochondria pathway. Co-administration of ginsenoside Rg1 highlights the potential to increase the therapeutic index of colistin.

  4. Mutations in GANAB, Encoding the Glucosidase IIα Subunit, Cause Autosomal-Dominant Polycystic Kidney and Liver Disease.

    PubMed

    Porath, Binu; Gainullin, Vladimir G; Cornec-Le Gall, Emilie; Dillinger, Elizabeth K; Heyer, Christina M; Hopp, Katharina; Edwards, Marie E; Madsen, Charles D; Mauritz, Sarah R; Banks, Carly J; Baheti, Saurabh; Reddy, Bharathi; Herrero, José Ignacio; Bañales, Jesús M; Hogan, Marie C; Tasic, Velibor; Watnick, Terry J; Chapman, Arlene B; Vigneau, Cécile; Lavainne, Frédéric; Audrézet, Marie-Pierre; Ferec, Claude; Le Meur, Yannick; Torres, Vicente E; Harris, Peter C

    2016-06-02

    Autosomal-dominant polycystic kidney disease (ADPKD) is a common, progressive, adult-onset disease that is an important cause of end-stage renal disease (ESRD), which requires transplantation or dialysis. Mutations in PKD1 or PKD2 (∼85% and ∼15% of resolved cases, respectively) are the known causes of ADPKD. Extrarenal manifestations include an increased level of intracranial aneurysms and polycystic liver disease (PLD), which can be severe and associated with significant morbidity. Autosomal-dominant PLD (ADPLD) with no or very few renal cysts is a separate disorder caused by PRKCSH, SEC63, or LRP5 mutations. After screening, 7%-10% of ADPKD-affected and ∼50% of ADPLD-affected families were genetically unresolved (GUR), suggesting further genetic heterogeneity of both disorders. Whole-exome sequencing of six GUR ADPKD-affected families identified one with a missense mutation in GANAB, encoding glucosidase II subunit α (GIIα). Because PRKCSH encodes GIIβ, GANAB is a strong ADPKD and ADPLD candidate gene. Sanger screening of 321 additional GUR families identified eight further likely mutations (six truncating), and a total of 20 affected individuals were identified in seven ADPKD- and two ADPLD-affected families. The phenotype was mild PKD and variable, including severe, PLD. Analysis of GANAB-null cells showed an absolute requirement of GIIα for maturation and surface and ciliary localization of the ADPKD proteins (PC1 and PC2), and reduced mature PC1 was seen in GANAB(+/-) cells. PC1 surface localization in GANAB(-/-) cells was rescued by wild-type, but not mutant, GIIα. Overall, we show that GANAB mutations cause ADPKD and ADPLD and that the cystogenesis is most likely driven by defects in PC1 maturation.

  5. Characterization of shrunken endosperm mutants in barley.

    PubMed

    Ma, Jian; Jiang, Qian-Tao; Wei, Long; Wang, Ji-Rui; Chen, Guo-Yue; Liu, Ya-Xi; Li, Wei; Wei, Yu-Ming; Liu, Chunji; Zheng, You-Liang

    2014-04-10

    Despite numerous studies on shrunken endosperm mutants caused by either maternal tissues (seg) or kernel per se (sex) in barley, the molecular mechanism for all of the eight seg mutants (seg1-seg8) and some sex mutants is yet to be uncovered. In this study, we determined the amylose content, characterized granule-binding proteins, analyzed the expression of key genes involved in starch synthesis, and examined starch granule structure of both normal (Bowman and Morex) and shrunken endosperm (seg1, seg3, seg4a, seg4b, seg5, seg6, seg7, and sex1) barley accessions. Our results showed that amylose contents of shrunken endosperm mutants ranged from 8.9% (seg4a) to 25.8% (seg1). SDS-PAGE analysis revealed that 87 kDa proteins corresponding to the starch branching enzyme II (SBEII) and starch synthase II (SSII) were not present in seg1, seg3, seg6, and seg7 mutants. Real-time quantitative PCR (RT-qPCR) analysis indicated that waxy expression levels of seg1, seg3, seg6, and seg7 mutants decreased in varying degrees to lower levels until 27 days after anthesis (DAA) after reaching the peak at 15-21 DAA, which differed from the pattern of normal barley accessions. Further characterization of waxy alleles revealed 7 non-synonymous single nucleotide polymorphisms (SNPs) in the coding sequences and 16 SNPs and 8 indels in the promoter sequences of the mutants. Results from starch granule by scanning electron microscopy (SEM) indicated that, in comparison with normal barley accessions, seg4a, seg4b, and sex1 had fewer starch granules per grain; seg3 and seg6 had less small B-type granules; some large A-type granules in seg7 had a hollow surface. These results improve our understanding about effects of seg and sex mutants on starch biosynthesis and granule structure during endosperm development and provide information for identification of key genes responsible for these shrunken endosperm mutants.

  6. Mathematics Instruction and the Tablet PC

    ERIC Educational Resources Information Center

    Fister, K. Renee; McCarthy, Maeve L.

    2008-01-01

    The use of tablet PCs in teaching is a relatively new phenomenon. A cross between a notebook computer and a personal digital assistant (PDA), the tablet PC has all of the features of a notebook with the additional capability that the screen can also be used for input. Tablet PCs are usually equipped with a stylus that allows the user to write on…

  7. Jargon that Computes: Today's PC Terminology.

    ERIC Educational Resources Information Center

    Crawford, Walt

    1997-01-01

    Discusses PC (personal computer) and telecommunications terminology in context: Integrated Services Digital Network (ISDN); Asymmetric Digital Subscriber Line (ADSL); cable modems; satellite downloads; T1 and T3 lines; magnitudes ("giga-,""nano-"); Central Processing Unit (CPU); Random Access Memory (RAM); Universal Serial Bus…

  8. Multitasking Operating Systems for the IBM PC.

    ERIC Educational Resources Information Center

    Owen, G. Scott

    1985-01-01

    The ability of a microcomputer to execute several programs at the same time is called "multitasking." The nature and use of one multitasking operating system Concurrent PC-DOS from Digital Research (the developers of the CP/M operating system) are discussed. (JN)

  9. Combating adverse selection in secondary PC markets.

    PubMed

    Hickey, Stewart W; Fitzpatrick, Colin

    2008-04-15

    Adverse selection is a significant contributor to market failure in secondary personal computer (PC) markets. Signaling can act as a potential solution to adverse selection and facilitate superior remarketing of second-hand PCs. Signaling is a means whereby usage information can be utilized to enhance consumer perception of both value and utility of used PCs and, therefore, promote lifetime extension for these systems. This can help mitigate a large portion of the environmental impact associated with PC system manufacture. In this paper, the computer buying and selling behavior of consumers is characterized via a survey of 270 Irish residential users. Results confirm the existence of adverse selection in the Irish market with 76% of potential buyers being unwilling to purchase and 45% of potential vendors being unwilling to sell a used PC. The so-called "closet affect" is also apparent with 78% of users storing their PC after use has ceased. Results also indicate that consumers place a higher emphasis on specifications when considering a second-hand purchase. This contradicts their application needs which are predominantly Internet and word-processing/spreadsheet/presentation applications, 88% and 60% respectively. Finally, a market solution utilizing self monitoring and reporting technology (SMART) sensors for the purpose of real time usage monitoring is proposed, that can change consumer attitudes with regard to second-hand computer equipment.

  10. PC Kiosk Trends in Rural India

    ERIC Educational Resources Information Center

    Toyama, Kentaro; Kiri, Karishma; Menon, Deepak; Sethi, Suneet; Pal, Joyojeet; Srinivasan, Janaki

    2006-01-01

    This article presents a series of preliminary, quantitative results on rural PC kiosks in India. An analysis of the data confirms many expected trends and correlations and shows that kiosks still face the challenge of sustainability as a business. This study is based on questionnaires presented to kiosk operators and customers of kiosks operated…

  11. Experience in setting up a PC cluster

    NASA Astrophysics Data System (ADS)

    Lin, Ganghua; Zhang, Mei

    2004-09-01

    In this paper we summary and present our thinking and experience in setting up a PC cluster, with a consideration that the described thinking and experience may be relevant to or useful for those who intend to buy a similar cluster in the near future.

  12. Stretch Your PC Dollars--Buy Clones.

    ERIC Educational Resources Information Center

    True, John

    1986-01-01

    Relates how the story of how San Francisco State University evaluated IBM PC look-alikes, considered some of the risks involved, and decided to purchase over 100 of them. Questions of compatibility, vendor longevity, support, and other risk management issues are discussed. (Author/MLW)

  13. Wf/pc Cycle 1 Cal: Non-Sv PC Flats 1

    NASA Astrophysics Data System (ADS)

    MacKenty, John

    1990-12-01

    This program takes "uniform illuminated" pictures of the earth to obtain the "instrument signature" for flat field corrections. This proposal contains observations for all PC filters used by GOs and GTOs during Cycle 1 which were not included in the SV program. The camera - filter combinations done in this proposal are: PC: F368M,F588N,F631N,F648M,F875M,F1042M These are the PC flats which are not expected to saturate on the bright earth.

  14. ECB deacylase mutants

    DOEpatents

    Arnold, Frances H.; Shao, Zhixin; Zhao, Huimin; Giver, Lorraine J.

    2002-01-01

    A method for in vitro mutagenesis and recombination of polynucleotide sequences based on polymerase-catalyzed extension of primer oligonucleotides is disclosed. The method involves priming template polynucleotide(s) with random-sequences or defined-sequence primers to generate a pool of short DNA fragments with a low level of point mutations. The DNA fragments are subjected to denaturization followed by annealing and further enzyme-catalyzed DNA polymerization. This procedure is repeated a sufficient number of times to produce full-length genes which comprise mutants of the original template polynucleotides. These genes can be further amplified by the polymerase chain reaction and cloned into a vector for expression of the encoded proteins.

  15. Mutant presenilin2 promotes apoptosis through the p53/miR-34a axis in neuronal cells.

    PubMed

    Li, Liu-Hong; Tu, Qiu-Yun; Deng, Xiao-Hua; Xia, Jian; Hou, De-Ren; Guo, Ke; Zi, Xiao-Hong

    2017-05-01

    Neurodegenerative disorders have attracted attention in last decades due to their high incidence in the world. The p53/miR-34a axis triggers apoptosis and suppresses viability in multiple types of cells, but little is known about its role in neurodegenerative diseases. In this study, we showed that presenilin (PS)-2, a major gene associated with familial Alzheimer's disease (AD) could trigger the apoptosis through the p53/miR-34a axis in PC12 cells. First we found that PC12 cell viability was downregulated by PS-2 and mutant PS-2 overexpression, especially by mutant PS-2 overexpression. Then, we established a mutant PS-2-overexpressing PC12 cell line and confirmed that mutant PS-2 induced not only p53 but also miR-34a expression. The transfection of miR-34a inhibitor reversed PS-2-induced effects on cellular viability and apoptosis. Mutant PS-2 overexpression promoted caspase-3 expression, reduced Sirt1 and Bcl-2 expression, all of which were miR-34a downstream genes related with cell apoptosis. Moreover, mutant PS-2 also activated the p53/miR-34a axis and induced apoptosis in AD transgenic mice brain. These results implied that mutant PS-2 might promote the apoptosis of neuronal cells through triggering the p53/miR-34a axis. Altogether our results provide a novel insight into neurodegenerative disease and deepen our understandings of AD pathogenic processes.

  16. PC1/3, PC2 and PC5/6A are targeted to dense core secretory granules by a common mechanism.

    PubMed

    Dikeakos, Jimmy D; Mercure, Chantal; Lacombe, Marie-Josée; Seidah, Nabil G; Reudelhuber, Timothy L

    2007-08-01

    There are seven members of the proprotein convertase (PC) family of secreted serine proteases that cleave their substrates at basic amino acids, thereby activating a variety of hormones, growth factors, and viruses. PC1/3, PC2 and PC5/6A are the only members of the PC family that are targeted to dense core secretory granules, where they carry out the processing of proteins that are secreted from the cell in a regulated manner. Previous studies have identified alpha-helices in the C-termini of the PC1/3 and PC2 proteases that are required for this subcellular targeting. In the current study, we demonstrate that a predicted alpha-helix in the C-terminus of PC5/6A is also critical for the ability of this domain to target a heterologous protein to the regulated secretory pathway of mouse endocrine AtT-20 cells. Analysis of the subcellular distribution of fusion proteins containing the C-terminal domains of PC1/3, PC2 and PC5/6A confirmed that all three domains have the capacity to redirect a constitutively secreted protein to the granule-containing cytoplasmic extensions. Analysis of the predicted structures formed by these three granule-sorting helices shows a correlation between their granule-sorting efficiency and the clustering of hydrophobic amino acids in their granule-targeting helices.

  17. Binding of bovine T194A PrPC by PrPSc-specific antibodies

    PubMed Central

    Madampage, Claudia A; Määttänen, Pekka; Marciniuk, Kristen; Brownlie, Robert; Andrievskaia, Olga; Potter, Andrew; Cashman, Neil R; Lee, Jeremy S; Napper, Scott

    2013-01-01

    Transmissible spongiform encephalopathies (TSEs) are fatal neurodegenerative diseases that are based on the misfolding of a cellular prion protein (PrPC) into an infectious, pathological conformation (PrPSc). There is proof-of-principle evidence that a prion vaccine is possible but this is tempered with concerns of the potential dangers associated with induction of immune responses to a widely-expressed self-protein. By targeting epitopes that are specifically exposed upon protein misfolding, our group developed a vaccine that induces PrPSc-specific antibody responses. Here we consider the ability of this polyclonal antibody (SN6b) to bind to a mutant of PrPC associated with spontaneous prion disease. Polyclonal antibodies were selected to mimic the vaccination outcome and also explore all possible protein conformations of the recombinant bovine prion protein with mutation T194A [bPrP(T194A)]. This mutant is a homolog of the human T183A mutation of PrPC that is associated with early onset of familial dementia. With nanopore analysis, under non-denaturing conditions, we observed binding of the SN6b antibody to bPrP(T194A). This interaction was confirmed through ELISAs as well as immunoprecipitation of the recombinant and cellularly expressed forms of bPrP(T194A). This interaction did not promote formation of a protease resistant conformation of PrP in vitro. Collectively, these findings support the disease-specific approach for immunotherapy of prion diseases but also suggest that the concept of conformation-specific immunotherapy may be complicated in individuals who are genetically predisposed to PrPC misfolding. PMID:23787697

  18. Nerve growth factor and epidermal growth factor stimulate clusterin gene expression in PC12 cells.

    PubMed Central

    Gutacker, C; Klock, G; Diel, P; Koch-Brandt, C

    1999-01-01

    Clusterin (apolipoprotein J) is an extracellular glycoprotein that might exert functions in development, cell death and lipid transport. Clusterin gene expression is elevated at sites of tissue remodelling, such as differentiation and apoptosis; however, the signals responsible for this regulation have not been identified. We use here the clusterin gene as a model system to examine expression in PC12 cells under the control of differentiation and proliferation signals produced by nerve growth factor (NGF) and by epidermal growth factor (EGF) respectively. NGF induced clusterin mRNA, which preceded neurite outgrowth typical of neuronal differentiation. EGF also activated the clusterin mRNA, demonstrating that both proliferation and differentiation signals regulate the gene. To localize NGF- and EGF-responsive elements we isolated the clusterin promoter and tested it in PC12 cell transfections. A 2.5 kb promoter fragment and two 1.5 and 0.3 kb deletion mutants were inducible by NGF and EGF. The contribution to this response of a conserved activator protein 1 (AP-1) motif located in the 0.3 kb fragment was analysed by mutagenesis. The mutant promoter was not inducible by NGF or EGF, which identifies the AP-1 motif as an element responding to both factors. Binding studies with PC12 nuclear extracts showed that AP-1 binds to this sequence in the clusterin promoter. These findings suggest that NGF and EGF, which give differential gene regulation in PC12 cells, resulting in neuronal differentiation and proliferation respectively, use the common Ras/extracellular signal-regulated kinase/AP-1 signalling pathway to activate clusterin expression. PMID:10215617

  19. PC12 differentiation on biopolymer nanostructures

    NASA Astrophysics Data System (ADS)

    Cecchini, Marco; Bumma, Giorgia; Serresi, Michela; Beltram, Fabio

    2007-12-01

    The study of nervous system regeneration and axonal outgrowth control are relevant in several research areas, like neurophysiology or biomedical engineering. Among the elements that control neuron dynamics, the host substrate topography is a key parameter in determining cell differentiation. We present time-lapse experiments to analyze the differentiation dynamics of PC12 cells on nanopatterned biocompatible substrates. 200 nm depth gratings were fabricated on tissue-culture polystyrene substrates by nanoimprint lithography; different linewidths and pitches were compared down to 500 nm and 1000 nm, respectively. PC12 cells were cultured on these substrates and, following NGF administration to the medium, body morphology, cell movement and neuritogenesis were monitored at different time periods. In addition to demonstrating guided differentiation, our studies show complex time variations in body morphology and axon length, and guided cell movement. We show unstable synaptic connections and cell-body polarization, and the competition between topographical guidance and cell-cell interactions.

  20. IBM PC enhances the world's future

    NASA Technical Reports Server (NTRS)

    Cox, Jozelle

    1988-01-01

    Although the purpose of this research is to illustrate the importance of computers to the public, particularly the IBM PC, present examinations will include computers developed before the IBM PC was brought into use. IBM, as well as other computing facilities, began serving the public years ago, and is continuing to find ways to enhance the existence of man. With new developments in supercomputers like the Cray-2, and the recent advances in artificial intelligence programming, the human race is gaining knowledge at a rapid pace. All have benefited from the development of computers in the world; not only have they brought new assets to life, but have made life more and more of a challenge everyday.

  1. A PC based fault diagnosis expert system

    NASA Technical Reports Server (NTRS)

    Marsh, Christopher A.

    1990-01-01

    The Integrated Status Assessment (ISA) prototype expert system performs system level fault diagnosis using rules and models created by the user. The ISA evolved from concepts to a stand-alone demonstration prototype using OPS5 on a LISP Machine. The LISP based prototype was rewritten in C and the C Language Integrated Production System (CLIPS) to run on a Personal Computer (PC) and a graphics workstation. The ISA prototype has been used to demonstrate fault diagnosis functions of Space Station Freedom's Operation Management System (OMS). This paper describes the development of the ISA prototype from early concepts to the current PC/workstation version used today and describes future areas of development for the prototype.

  2. Virtual Reality at the PC Level

    NASA Technical Reports Server (NTRS)

    Dean, John

    1998-01-01

    The main objective of my research has been to incorporate virtual reality at the desktop level; i.e., create virtual reality software that can be run fairly inexpensively on standard PC's. The standard language used for virtual reality on PC's is VRML (Virtual Reality Modeling Language). It is a new language so it is still undergoing a lot of changes. VRML 1.0 came out only a couple years ago and VRML 2.0 came out around last September. VRML is an interpreted language that is run by a web browser plug-in. It is fairly flexible in terms of allowing you to create different shapes and animations. Before this summer, I knew very little about virtual reality and I did not know VRML at all. I learned the VRML language by reading two books and experimenting on a PC. The following topics are presented: CAD to VRML, VRML 1.0 to VRML 2.0, VRML authoring tools, VRML browsers, finding virtual reality applications, the AXAF project, the VRML generator program, web communities and future plans.

  3. Spent fuel pin temperature PC code

    SciTech Connect

    Fischer, L.E.

    1985-03-01

    During an annual outage, a Pressurized Water Reactor (PWR) may discharge 60 or more spent fuel bundles into its storage pool. Most early PWRs were built to store 3 to 5 years of spent fuel in their pools and are beginning to exceed their capacities. One method currently being developed and licensed for expanding spent fuel storage capabilities is the dry storage of spent fuel in large casks. To reduce the probability of gross failures of fuel cladding during dry storage in casks, the fuel pin temperatures must be shown to remain within acceptable limits. LLNL has developed, for the Nuclear Regulatory Commission, a personal computer (PC) code for calculating fuel pin temperatures on the IBM PC. The code uses the Wooton-Epstein Correlation to calculate the pin temperatures and has been benchmarked against test data. An iterative type of solution is used to calculate the fuel pin temperatures for specified heat fluxes and pin configurations. The PC code is useful in performing confirmatory analyses and comparing the results with those submitted by applicants applying for storage licenses. 5 references, 2 tables.

  4. Huntington's disease cerebrospinal fluid seeds aggregation of mutant huntingtin

    PubMed Central

    Tan, Z; Dai, W; van Erp, T G M; Overman, J; Demuro, A; Digman, M A; Hatami, A; Albay, R; Sontag, E M; Potkin, K T; Ling, S; Macciardi, F; Bunney, W E; Long, J D; Paulsen, J S; Ringman, J M; Parker, I; Glabe, C; Thompson, L M; Chiu, W; Potkin, S G

    2015-01-01

    Huntington's disease (HD), a progressive neurodegenerative disease, is caused by an expanded CAG triplet repeat producing a mutant huntingtin protein (mHTT) with a polyglutamine-repeat expansion. Onset of symptoms in mutant huntingtin gene-carrying individuals remains unpredictable. We report that synthetic polyglutamine oligomers and cerebrospinal fluid (CSF) from BACHD transgenic rats and from human HD subjects can seed mutant huntingtin aggregation in a cell model and its cell lysate. Our studies demonstrate that seeding requires the mutant huntingtin template and may reflect an underlying prion-like protein propagation mechanism. Light and cryo-electron microscopy show that synthetic seeds nucleate and enhance mutant huntingtin aggregation. This seeding assay distinguishes HD subjects from healthy and non-HD dementia controls without overlap (blinded samples). Ultimately, this seeding property in HD patient CSF may form the basis of a molecular biomarker assay to monitor HD and evaluate therapies that target mHTT. PMID:26100538

  5. Poliovirus Mutants Resistant to Neutralization with Soluble Cell Receptors

    NASA Astrophysics Data System (ADS)

    Kaplan, Gerardo; Peters, David; Racaniello, Vincent R.

    1990-12-01

    Poliovirus mutants resistant to neutralization with soluble cellular receptor were isolated. Replication of soluble receptor-resistant (srr) mutants was blocked by a monoclonal antibody directed against the HeLa cell receptor for poliovirus, indicating that the mutants use this receptor to enter cells. The srr mutants showed reduced binding to HeLa cells and cell membranes. However, the reduced binding phenotype did not have a major impact on viral replication, as judged by plaque size and one-step growth curves. These results suggest that the use of soluble receptors as antiviral agents could lead to the selection of neutralization-resistant mutants that are able to bind cell surface receptors, replicate, and cause disease.

  6. Cell-cycle-dependent PC-PLC regulation by APC/C(Cdc20)-mediated ubiquitin-proteasome pathway.

    PubMed

    Fu, Da; Ma, Yushui; Wu, Wei; Zhu, Xuchao; Jia, Chengyou; Zhao, Qianlei; Zhang, Chunyi; Wu, Xing Zhong

    2009-07-01

    Phosphatidylcholine-specific phospholipase C (PC-PLC) is involved in the cell signal transduction, cell proliferation, and apoptosis. The mechanism of its action, however, has not been fully understood, particularly, the role of PC-PLC in the cell cycle. In the present study, we found that cell division cycle 20 homolog (Cdc20) and PC-PLC were co-immunoprecipitated reciprocally by either antibody in rat hepatoma cells CBRH-7919 as well as in rat liver tissue. Using confocal microscopy, we found that PC-PLC and Cdc20 were co-localized in the perinuclear endoplasmic reticulum region (the "juxtanuclear quality control" compartment, JUNQ). The expression level and activities of PC-PLC changed in a cell-cycle-dependent manner and were inversely correlated with the expression of Cdc20. Intriguingly, Cdc20 overexpression altered the subcellular localization and distribution of PC-PLC, and caused PC-PLC degradation by the ubiquitin proteasome pathway (UPP). Taken together, our data indicate that PC-PLC regulation in cell cycles is controlled by APC/C(Cdc20)-mediated UPP.

  7. Impaired Terminal Differentiation of Hippocampal Granule Neurons and Defective Contextual Memory in PC3/Tis21 Knockout Mice

    PubMed Central

    Costanzi, Marco; Leonardi, Luca; Cinà, Irene; Micheli, Laura; Nutini, Michele; Longone, Patrizia; Oh, S. Paul; Cestari, Vincenzo; Tirone, Felice

    2009-01-01

    Neurogenesis in the dentate gyrus of the adult hippocampus has been implicated in neural plasticity and memory, but the molecular mechanisms controlling the proliferation and differentiation of newborn neurons and their integration into the synaptic circuitry are still largely unknown. To investigate this issue, we have analyzed the adult hippocampal neurogenesis in a PC3/Tis21-null mouse model. PC3/Tis21 is a transcriptional co-factor endowed with antiproliferative and prodifferentiative properties; indeed, its upregulation in neural progenitors has been shown to induce exit from cell cycle and differentiation. We demonstrate here that the deletion of PC3/Tis21 causes an increased proliferation of progenitor cells in the adult dentate gyrus and an arrest of their terminal differentiation. In fact, in the PC3/Tis21-null hippocampus postmitotic undifferentiated neurons accumulated, while the number of terminally differentiated neurons decreased of 40%. As a result, PC3/Tis21-null mice displayed a deficit of contextual memory. Notably, we observed that PC3/Tis21 can associate to the promoter of Id3, an inhibitor of proneural gene activity, and negatively regulates its expression, indicating that PC3/Tis21 acts upstream of Id3. Our results identify PC3/Tis21 as a gene required in the control of proliferation and terminal differentiation of newborn neurons during adult hippocampal neurogenesis and suggest its involvement in the formation of contextual memories. PMID:20020054

  8. PC, a Novel Oral Insecticidal Toxin from Bacillus bombysepticus Involved in Host Lethality via APN and BtR-175

    PubMed Central

    Lin, Ping; Cheng, Tingcai; Jin, Shengkai; Wu, Yuqian; Fu, Bohua; Long, Renwen; Zhao, Ping; Xia, Qingyou

    2015-01-01

    Insect pests have developed resistance to chemical insecticides, insecticidal toxins as bioinsecticides or genetic protection built into crops. Consequently, novel, orally active insecticidal toxins would be valuable biological alternatives for pest control. Here, we identified a novel insecticidal toxin, parasporal crystal toxin (PC), from Bacillus bombysepticus (Bb). PC shows oral pathogenic activity and lethality towards silkworms and Cry1Ac-resistant Helicoverpa armigera strains. In vitro assays, PC after activated by trypsin binds to BmAPN4 and BtR-175 by interacting with CR7 and CR12 fragments. Additionally, trypsin-activated PC demonstrates cytotoxicity against Sf9 cells expressing BmAPN4, revealing that BmAPN4 serves as a functional receptor that participates in Bb and PC pathogenicity. In vivo assay, knocking out BtR-175 increased the resistance of silkworms to PC. These data suggest that PC is the first protein with insecticidal activity identified in Bb that is capable of causing silkworm death via receptor interactions, representing an important advance in our understanding of the toxicity of Bb and the contributions of interactions between microbial pathogens and insects to disease pathology. Furthermore, the potency of PC as an insecticidal protein makes it a good candidate for inclusion in integrated agricultural pest management systems. PMID:26057951

  9. Arabidopsis MET1 cytosine methyltransferase mutants.

    PubMed Central

    Kankel, Mark W; Ramsey, Douglas E; Stokes, Trevor L; Flowers, Susan K; Haag, Jeremy R; Jeddeloh, Jeffrey A; Riddle, Nicole C; Verbsky, Michelle L; Richards, Eric J

    2003-01-01

    We describe the isolation and characterization of two missense mutations in the cytosine-DNA-methyltransferase gene, MET1, from the flowering plant Arabidopsis thaliana. Both missense mutations, which affect the catalytic domain of the protein, led to a global reduction of cytosine methylation throughout the genome. Surprisingly, the met1-2 allele, with the weaker DNA hypomethylation phenotype, alters a well-conserved residue in methyltransferase signature motif I. The stronger met1-1 allele caused late flowering and a heterochronic delay in the juvenile-to-adult rosette leaf transition. The distribution of late-flowering phenotypes in a mapping population segregating met1-1 indicates that the flowering-time phenotype is caused by the accumulation of inherited defects at loci unlinked to the met1 mutation. The delay in flowering time is due in part to the formation and inheritance of hypomethylated fwa epialleles, but inherited defects at other loci are likely to contribute as well. Centromeric repeat arrays hypomethylated in met1-1 mutants are partially remethylated when introduced into a wild-type background, in contrast to genomic sequences hypomethylated in ddm1 mutants. ddm1 met1 double mutants were constructed to further our understanding of the mechanism of DDM1 action and the interaction between two major genetic loci affecting global cytosine methylation levels in Arabidopsis. PMID:12663548

  10. Methods of producing protoporphyrin IX and bacterial mutants therefor

    SciTech Connect

    Zhou, Jizhong; Qiu, Dongru; He, Zhili; Xie, Ming

    2016-03-01

    The presently disclosed inventive concepts are directed in certain embodiments to a method of producing protoporphyrin IX by (1) cultivating a strain of Shewanella bacteria in a culture medium under conditions suitable for growth thereof, and (2) recovering the protoporphyrin IX from the culture medium. The strain of Shewanella bacteria comprises at least one mutant hemH gene which is incapable of normal expression, thereby causing an accumulation of protoporphyrin IX. In certain embodiments of the method, the strain of Shewanella bacteria is a strain of S. loihica, and more specifically may be S. loihica PV-4. In certain embodiments, the mutant hemH gene of the strain of Shewanella bacteria may be a mutant of shew_2229 and/or of shew_1140. In other embodiments, the presently disclosed inventive concepts are directed to mutant strains of Shewanella bacteria having at least one mutant hemH gene which is incapable of normal expression, thereby causing an accumulation of protoporphyrin IX during cultivation of the bacteria. In certain embodiments the strain of Shewanella bacteria is a strain of S. loihica, and more specifically may be S. loihica PV-4. In certain embodiments, the mutant hemH gene of the strain of Shewanella bacteria may be a mutant of shew_2229 and/or shew_1140.

  11. 39 CFR 501.16 - PC postage payment methodology.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 39 Postal Service 1 2010-07-01 2010-07-01 false PC postage payment methodology. 501.16 Section 501.16 Postal Service UNITED STATES POSTAL SERVICE POSTAGE PROGRAMS AUTHORIZATION TO MANUFACTURE AND DISTRIBUTE POSTAGE EVIDENCING SYSTEMS § 501.16 PC postage payment methodology. (a) The PC Postage customer...

  12. The USL NASA PC R and D development environment standards

    NASA Technical Reports Server (NTRS)

    Dominick, Wayne D. (Editor); Moreau, Dennis R.

    1984-01-01

    The development environment standards which have been established in order to control usage of the IBM PC/XT development systems and to prevent interference between projects being currently developed on the PC's are discussed. The standards address the following areas: scheduling PC resources; login/logout procedures; training; file naming conventions; hard disk organization; diskette care; backup procedures; and copying policies.

  13. Isolation of a Defective Prion Mutant from Natural Scrapie

    PubMed Central

    Migliore, Sergio; Cosseddu, Gian Mario; Pirisinu, Laura; Riccardi, Geraldina; Nonno, Romolo

    2016-01-01

    It is widely known that prion strains can mutate in response to modification of the replication environment and we have recently reported that prion mutations can occur in vitro during amplification of vole-adapted prions by Protein Misfolding Cyclic Amplification on bank vole substrate (bvPMCA). Here we exploited the high efficiency of prion replication by bvPMCA to study the in vitro propagation of natural scrapie isolates. Although in vitro vole-adapted PrPSc conformers were usually similar to the sheep counterpart, we repeatedly isolated a PrPSc mutant exclusively when starting from extremely diluted seeds of a single sheep isolate. The mutant and faithful PrPSc conformers showed to be efficiently autocatalytic in vitro and were characterized by different PrP protease resistant cores, spanning aa ∼155–231 and ∼80–231 respectively, and by different conformational stabilities. The two conformers could thus be seen as different bona fide PrPSc types, putatively accounting for prion populations with different biological properties. Indeed, once inoculated in bank vole the faithful conformer was competent for in vivo replication while the mutant was unable to infect voles, de facto behaving like a defective prion mutant. Overall, our findings confirm that prions can adapt and evolve in the new replication environments and that the starting population size can affect their evolutionary landscape, at least in vitro. Furthermore, we report the first example of “authentic” defective prion mutant, composed of brain-derived PrPC and originating from a natural scrapie isolate. Our results clearly indicate that the defective mutant lacks of some structural characteristics, that presumably involve the central region ∼90–155, critical for infectivity but not for in vitro replication. Finally, we propose a molecular mechanism able to account for the discordant in vitro and in vivo behavior, suggesting possible new paths for investigating the molecular bases of

  14. Wf/pc Cycle 1 Calib: Non-Sv PC Flats 2

    NASA Astrophysics Data System (ADS)

    MacKenty, John

    1990-12-01

    This program takes "uniform illuminated" pictures of the earth to obtain the "instrument signature" for flat field corrections. This proposal series contains observations for all PC filters used by GOs and GTOs during Cycle 1 which were not included in the SV program. The camera - filter combinations done in this proposal are: PC: F569W, F606W, F675W, F725LP, F791W, F814W, F850LP (these are done in combination with F122M) These are the PC flats which are expected to saturate on the bright earth without a neutral density filter. The following observations are also included to permit correction for the F122M filter: F517N+F122M, F517N, F664N+F122M, F664N, F718M+F122M, F718M, F875M+F122M, F875M

  15. IBM PC/IX operating system evaluation plan

    NASA Technical Reports Server (NTRS)

    Dominick, Wayne D. (Editor); Granier, Martin; Hall, Philip P.; Triantafyllopoulos, Spiros

    1984-01-01

    An evaluation plan for the IBM PC/IX Operating System designed for IBM PC/XT computers is discussed. The evaluation plan covers the areas of performance measurement and evaluation, software facilities available, man-machine interface considerations, networking, and the suitability of PC/IX as a development environment within the University of Southwestern Louisiana NASA PC Research and Development project. In order to compare and evaluate the PC/IX system, comparisons with other available UNIX-based systems are also included.

  16. Human Liver Cell Trafficking Mutants: Characterization and Whole Exome Sequencing

    PubMed Central

    Yuan, Fei; Snapp, Erik L.; Novikoff, Phyllis M.; Suadicani, Sylvia O.; Spray, David C.; Potvin, Barry; Wolkoff, Allan W.; Stanley, Pamela

    2014-01-01

    The HuH7 liver cell mutant Trf1 is defective in membrane trafficking and is complemented by the casein kinase 2α subunit CK2α’’. Here we identify characteristic morphologies, trafficking and mutational changes in six additional HuH7 mutants Trf2-Trf7. Trf1 cells were previously shown to be severely defective in gap junction functions. Using a Lucifer yellow transfer assay, remarkable attenuation of gap junction communication was revealed in each of the mutants Trf2-Trf7. Electron microscopy and light microscopy of thiamine pyrophosphatase showed that several mutants exhibited fragmented Golgi apparatus cisternae compared to parental HuH7 cells. Intracellular trafficking was investigated using assays of transferrin endocytosis and recycling and VSV G secretion. Surface binding of transferrin was reduced in all six Trf2-Trf7 mutants, which generally correlated with the degree of reduced expression of the transferrin receptor at the cell surface. The mutants displayed the same transferrin influx rates as HuH7, and for efflux rate, only Trf6 differed, having a slower transferrin efflux rate than HuH7. The kinetics of VSV G transport along the exocytic pathway were altered in Trf2 and Trf5 mutants. Genetic changes unique to particular Trf mutants were identified by exome sequencing, and one was investigated in depth. The novel mutation Ile34Phe in the GTPase RAB22A was identified in Trf4. RNA interference knockdown of RAB22A or overexpression of RAB22AI34F in HuH7 cells caused phenotypic changes characteristic of the Trf4 mutant. In addition, the Ile34Phe mutation reduced both guanine nucleotide binding and hydrolysis activities of RAB22A. Thus, the RAB22A Ile34Phe mutation appears to contribute to the Trf4 mutant phenotype. PMID:24466322

  17. Human liver cell trafficking mutants: characterization and whole exome sequencing.

    PubMed

    Yuan, Fei; Snapp, Erik L; Novikoff, Phyllis M; Suadicani, Sylvia O; Spray, David C; Potvin, Barry; Wolkoff, Allan W; Stanley, Pamela

    2014-01-01

    The HuH7 liver cell mutant Trf1 is defective in membrane trafficking and is complemented by the casein kinase 2α subunit CK2α''. Here we identify characteristic morphologies, trafficking and mutational changes in six additional HuH7 mutants Trf2-Trf7. Trf1 cells were previously shown to be severely defective in gap junction functions. Using a Lucifer yellow transfer assay, remarkable attenuation of gap junction communication was revealed in each of the mutants Trf2-Trf7. Electron microscopy and light microscopy of thiamine pyrophosphatase showed that several mutants exhibited fragmented Golgi apparatus cisternae compared to parental HuH7 cells. Intracellular trafficking was investigated using assays of transferrin endocytosis and recycling and VSV G secretion. Surface binding of transferrin was reduced in all six Trf2-Trf7 mutants, which generally correlated with the degree of reduced expression of the transferrin receptor at the cell surface. The mutants displayed the same transferrin influx rates as HuH7, and for efflux rate, only Trf6 differed, having a slower transferrin efflux rate than HuH7. The kinetics of VSV G transport along the exocytic pathway were altered in Trf2 and Trf5 mutants. Genetic changes unique to particular Trf mutants were identified by exome sequencing, and one was investigated in depth. The novel mutation Ile34Phe in the GTPase RAB22A was identified in Trf4. RNA interference knockdown of RAB22A or overexpression of RAB22AI34F in HuH7 cells caused phenotypic changes characteristic of the Trf4 mutant. In addition, the Ile34Phe mutation reduced both guanine nucleotide binding and hydrolysis activities of RAB22A. Thus, the RAB22A Ile34Phe mutation appears to contribute to the Trf4 mutant phenotype.

  18. The Local Population of White Dwarfs within 25 pc

    NASA Astrophysics Data System (ADS)

    Holberg, Jay B.; Oswalt, Terry D.; Sion, Edward M.

    2015-01-01

    We have extended the detailed survey of the local white dwarf population from 20 pc to 25 pc, effectively doubling the sample volume to now include 231 stars. The present 25 pc has an estimated completeness of 70% (the corresponding 20 pc sample is now 85% complete). The space density of white dwarfs remains at 4.8 ± 0.5 x 10-3 pc-3. There exists a curious excess of single stars in the sample 70% vs 30% in systems with one or more companions. A pronounced apparent deficiency remains between the eleven known Sirius-like systems present in the 20 pc sample and only a single such system presently known in the extended 25 pc sample. Also demonstrated, using explicit individual white dwarf cooling ages, is the feasibility of estimating the white dwarf birth rates over the last ~ 5 Gyr.This work is supported by NSF grant AST-1413537

  19. Epilepsy-Related Slack Channel Mutants Lead to Channel Over-Activity by Two Different Mechanisms.

    PubMed

    Tang, Qiong-Yao; Zhang, Fei-Fei; Xu, Jie; Wang, Ran; Chen, Jian; Logothetis, Diomedes E; Zhang, Zhe

    2016-01-05

    Twelve sodium-activated potassium channel (KCNT1, Slack) genetic mutants have been identified from severe early-onset epilepsy patients. The changes in biophysical properties of these mutants and the underlying mechanisms causing disease remain elusive. Here, we report that seven of the 12 mutations increase, whereas one mutation decreases, the channel's sodium sensitivity. Two of the mutants exhibit channel over-activity only when the intracellular Na(+) ([Na(+)]i) concentration is ∼80 mM. In contrast, single-channel data reveal that all 12 mutants increase the maximal open probability (Po). We conclude that these mutant channels lead to channel over-activity predominantly by increasing the ability of sodium binding to activate the channel, which is indicated by its maximal Po. The sodium sensitivity of these epilepsy causing mutants probably determines the [Na(+)]i concentration at which these mutants exert their pathological effects.

  20. Silymarin protects against acrylamide-induced neurotoxicity via Nrf2 signalling in PC12 cells.

    PubMed

    Li, Liang; Sun, Hong-Yang; Liu, Wei; Zhao, Hong-Yu; Shao, Mei-Li

    2017-04-01

    Silymarin (SM) is a well-known antioxidant, anti-inflammatory and anti-cancer compound extracted from the milk thistle. Here, we investigated the protective effect of SM against acrylamide (AA)-induced neurotoxicity, mainly caused by oxidative stress, via activation of the nuclear transcription factor E2-related factor 2 (Nrf2) signalling pathway in PC12 cells. The MTT reduction assay was used to measure cell viability in various drug-treated groups and demonstrated that SM could increase cell viability in AA-treated PC12 cells. We then measured the reactive oxygen species (ROS) levels by the peroxide-sensitive fluorescent probe DCFH-DA and intracellular glutathione (GSH) and malondialdehyde (MDA) levels by absorption spectrophotometry. Our data revealed that SM could reduce ROS and MDA levels and increase GSH levels in AA-induced PC12 cells. To identify a potential mechanism for SM-induced protection, we measured the mRNA and protein expression levels of Nrf2 and its downstream target antioxidants glutathione peroxidase (Gpx), glutamate cysteine ligase catalytic subunit (GCLC) and glutamate cysteine ligase modifier subunit (GCLM) by quantitative real-time PCR and Western blot, respectively. The results suggested that SM could activate Nrf2 signalling and increase the expression of Nrf2, Gpx, GCLC and GCLM in AA-treated PC12 cells. In conclusion, SM can effectively alleviate AA-induced neurotoxicity in PC12 cells.

  1. Gab1 mediates neurite outgrowth, DNA synthesis, and survival in PC12 cells.

    PubMed

    Korhonen, J M; Saïd, F A; Wong, A J; Kaplan, D R

    1999-12-24

    The Gab1-docking protein has been shown to regulate phosphatidylinositol 3-kinase PI3K activity and potentiate nerve growth factor (NGF)-induced survival in PC12 cells. Here, we investigated the potential of Gab1 to induce neurite outgrowth and DNA synthesis, two other important aspects of NGF-induced neuronal differentiation of PC12 cells and NGF-independent survival. We generated a recombinant adenovirus encoding hemagglutinin (HA)-epitope-tagged Gab1 and expressed this protein in PC12 cells. HA-Gab1 was constitutively tyrosine-phosphorylated in PC12 cells and induced the phosphorylation of Akt/protein kinase B and p44/42 mitogen-activated protein kinase. HA-Gab1-stimulated a 10-fold increase in neurite outgrowth in the absence of NGF and a 5-fold increase in NGF-induced neurite outgrowth. HA-Gab1 also stimulated DNA synthesis and caused NGF-independent survival in PC12 cells. Finally, we found that HA-Gab1-induced neuritogenesis was completely suppressed by pharmacological inhibition of mitogen-activated protein kinase kinase (MEK) activity and 50% suppressed by inhibition of PI3K activity. In contrast, HA-Gab1-stimulated cell survival was efficiently suppressed only by inhibition of both PI3K and MEK activities. These results indicate that Gab1 is capable of mediating differentiation, DNA synthesis, and cell survival and uses both PI3K and MEK signaling pathways to achieve its effects.

  2. PcFKH1, a novel regulatory factor from the forkhead family, controls the biosynthesis of penicillin in Penicillium chrysogenum.

    PubMed

    Domínguez-Santos, Rebeca; García-Estrada, Carlos; Kosalková, Katarina; Prieto, Carlos; Santamarta, Irene; Martín, Juan-Francisco

    2015-08-01

    Penicillin biosynthesis in Penicillium chrysogenum (re-identified as Penicillium rubens) is a good example of a biological process subjected to complex global regulatory networks and serves as a model to study fungal secondary metabolism. The winged-helix family of transcription factors recently described, which includes the forkhead type of proteins, is a key type of regulatory proteins involved in this process. In yeasts and humans, forkhead transcription factors are involved in different processes (cell cycle regulation, cell death control, pre-mRNA processing and morphogenesis); one member of this family of proteins has been identified in the P. chrysogenum genome (Pc18g00430). In this work, we have characterized this novel transcription factor (named PcFKH1) by generating knock-down mutants and overexpression strains. Results clearly indicate that PcFKH1 positively controls antibiotic biosynthesis through the specific interaction with the promoter region of the penDE gene, thus regulating penDE mRNA levels. PcFKH1 also binds to the pcbC promoter, but with low affinity. In addition, it also controls other ancillary genes of the penicillin biosynthetic process, such as phlA (encoding phenylacetyl CoA ligase) and ppt (encoding phosphopantetheinyl transferase). PcFKH1 also plays a role in conidiation and spore pigmentation, but it does not seem to be involved in hyphal morphology or cell division in the improved laboratory reference strain Wisconsin 54-1255. A genome-wide analysis of processes putatively coregulated by PcFKH1 and PcRFX1 (another winged-helix transcription factor) in P. chrysogenum provided evidence of the global effect of these transcription factors in P. chrysogenum metabolism.

  3. PC-based Multiple Information System Interface (PC/MISI) detailed design and implementation plan

    NASA Technical Reports Server (NTRS)

    Dominick, Wayne D. (Editor); Hall, Philip P.

    1985-01-01

    The design plan for the personal computer multiple information system interface (PC/MISI) project is discussed. The document is intended to be used as a blueprint for the implementation of the system. Each component is described in the detail necessary to allow programmers to implement the system. A description of the system data flow and system file structures is given.

  4. THE ORIGIN OF OB CLUSTERS: FROM 10 pc TO 0.1 pc

    SciTech Connect

    Liu Hauyu Baobab; Wang Ke; Ho, Paul T. P.; Zhang Qizhou; Quintana-Lacaci, Guillermo; Li Zhiyun; Zhang Zhiyu E-mail: kwang@cfa.harvard.edu E-mail: quintana@iram.es E-mail: zl4h@virginia.edu

    2012-01-20

    We observe the 1.2 mm continuum emission around the OB cluster-forming region G10.6-0.4, using the MAMBO-2 bolometer array of the IRAM 30 m telescope and the Submillimeter Array (SMA). Comparison of the Spitzer 24 {mu}m and 8 {mu}m images with our 1.2 mm continuum maps reveal an ionization front of an H II region, the photon-dominated layer, and several 5 pc scale filaments that follow the outer edge of the photon-dominated layer. The filaments, which are resolved in the MAMBO-2 observations, show regularly spaced parsec-scale molecular clumps, embedded with a cluster of dense molecular cores as shown in the SMA 0.87 mm observations. Toward the center of the G10.6-0.4 region, the combined SMA+IRAM 30 m continuum image reveals several parsec-scale protrusions. They may continue down to within 0.1 pc of the geometric center of a dense 3 pc scale structure, where a 200 M{sub Sun} OB cluster resides. The observed filaments may facilitate mass accretion onto the central cluster-forming region in the presence of strong radiative and mechanical stellar feedback. Their filamentary geometry may also facilitate fragmentation. We did not detect any significant polarized emission at 0.87 mm in the inner 1 pc region with SMA.

  5. Glycosaminoglycan Sulphation Affects the Seeded Misfolding of a Mutant Prion Protein

    PubMed Central

    Lawson, Victoria A.; Lumicisi, Brooke; Welton, Jeremy; Machalek, Dorothy; Gouramanis, Katrina; Klemm, Helen M.; Stewart, James D.; Masters, Colin L.; Hoke, David E.; Collins, Steven J.; Hill, Andrew F.

    2010-01-01

    Background The accumulation of protease resistant conformers of the prion protein (PrPres) is a key pathological feature of prion diseases. Polyanions, including RNA and glycosaminoglycans have been identified as factors that contribute to the propagation, transmission and pathogenesis of prion disease. Recent studies have suggested that the contribution of these cofactors to prion propagation may be species specific. Methodology/Principal Finding In this study a cell-free assay was used to investigate the molecular basis of polyanion stimulated PrPres formation using brain tissue or cell line derived murine PrP. Enzymatic depletion of endogenous nucleic acids or heparan sulphate (HS) from the PrPC substrate was found to specifically prevent PrPres formation seeded by mouse derived PrPSc. Modification of the negative charge afforded by the sulphation of glycosaminoglycans increased the ability of a familial PrP mutant to act as a substrate for PrPres formation, while having no effect on PrPres formed by wildtype PrP. This difference may be due to the observed differences in the binding of wild type and mutant PrP for glycosaminoglycans. Conclusions/Significance Cofactor requirements for PrPres formation are host species and prion strain specific and affected by disease associated mutations of the prion protein. This may explain both species and strain dependent propagation characteristics and provide insights into the underlying mechanisms of familial prion disease. It further highlights the challenge of designing effective therapeutics against a disease which effects a range of mammalian species, caused by range of aetiologies and prion strains. PMID:20808809

  6. (TFPP)Eu[Pc(OPh)8]Eu[Pc(OPh)8]/CuPc two-component bilayer heterojunction-based organic transistors with high ambipolar performance.

    PubMed

    Gao, Dameng; Zhang, Xia; Kong, Xia; Chen, Yanli; Jiang, Jianzhuang

    2015-02-04

    Organic thin film transistor (OTFT) devices fabricated by the solution-based QLS technique from a mixed (phthalocyaninato)(porphyrinato) europium complex (TFPP)Eu[Pc(OPh)8]Eu[Pc(OPh)8] exhibit air-stable ambipolar performance with mobilities of 6.0 × 10(-5) cm(2) V(-1) s(-)1 for holes and 1.4 × 10(-4) cm(2) V(-1) s(-1) for electrons, respectively. In good contrast, the two-component bilayer heterojunction thin film devices constructed by directly growing (TFPP)Eu[Pc(OPh)8]Eu[Pc(OPh)8] on vacuum deposited (VCD) CuPc film using solution based QLS method were revealed to show unprecedented ambipolar performance with carrier mobilities of 0.16 cm(2) V(-1) s(-1) for holes and 0.30 cm(2) V(-1) s(-1) for electrons. In addition to the intrinsic role of p-type organic semiconductor, the VCD CuPc film on the substrate also acts as a good template that induces significant improvement over the molecular ordering of triple-decker compound in the film. In particular, it results in the change in the aggregation mode of (TFPP)Eu[Pc(OPh)8]Eu[Pc(OPh)8] from J-type in the single-layer film to H-type in the bilayer film according to the UV-vis, XRD, and AFM observations.

  7. METCAN-PC - METAL MATRIX COMPOSITE ANALYZER

    NASA Technical Reports Server (NTRS)

    Murthy, P. L.

    1994-01-01

    High temperature metal matrix composites offer great potential for use in advanced aerospace structural applications. The realization of this potential however, requires concurrent developments in (1) a technology base for fabricating high temperature metal matrix composite structural components, (2) experimental techniques for measuring their thermal and mechanical characteristics, and (3) computational methods to predict their behavior. METCAN (METal matrix Composite ANalyzer) is a computer program developed to predict this behavior. METCAN can be used to computationally simulate the non-linear behavior of high temperature metal matrix composites (HT-MMC), thus allowing the potential payoff for the specific application to be assessed. It provides a comprehensive analysis of composite thermal and mechanical performance. METCAN treats material nonlinearity at the constituent (fiber, matrix, and interphase) level, where the behavior of each constituent is modeled accounting for time-temperature-stress dependence. The composite properties are synthesized from the constituent instantaneous properties by making use of composite micromechanics and macromechanics. Factors which affect the behavior of the composite properties include the fabrication process variables, the fiber and matrix properties, the bonding between the fiber and matrix and/or the properties of the interphase between the fiber and matrix. The METCAN simulation is performed as point-wise analysis and produces composite properties which are readily incorporated into a finite element code to perform a global structural analysis. After the global structural analysis is performed, METCAN decomposes the composite properties back into the localized response at the various levels of the simulation. At this point the constituent properties are updated and the next iteration in the analysis is initiated. This cyclic procedure is referred to as the integrated approach to metal matrix composite analysis. METCAN-PC

  8. Photodynamic therapy with the phthalocyanine photosensitizer Pc 4 of SW480 human colon cancer xenografts in athymic mice.

    PubMed

    Whitacre, C M; Feyes, D K; Satoh, T; Grossmann, J; Mulvihill, J W; Mukhtar, H; Oleinick, N L

    2000-05-01

    Photodynamic therapy (PDT) using the silicon phthalocyanine photosensitizer Pc 4 [HOSiPcOSi(CH3)2(CH2)3N-(CH3)2] is an oxidative stress associated with induction of apoptosis in various cell types. We assessed the effectiveness of Pc 4-PDT on SW480 colon cancer xenografts grown in athymic nude mice. Animals bearing xenografts were treated with 1 mg/kg body weight Pc 4 and 48 h later were irradiated with 150 J/cm2 672-nm light from a diode laser delivered at 150 mW/cm2. Biochemical studies were performed in xenografts resected at various time points up to 26 h after Pc 4-PDT treatment, whereas tumor size was evaluated over a 4-week period in parallel experiments. In the tumors resected for biochemical studies, apoptosis was visualized by activation of caspase-9 and caspase-3 and a gradual increase in the cleavage of the nuclear enzyme poly(ADP-ribose) polymerase (PARP) to a maximum of approximately 60% of the total PARP present at approximately 26 h. At that time all Pc 4-PDT-treated tumors had regressed significantly. Two signaling responses that have previously been shown to be associated with Pc 4-PDT-induced apoptosis in cultured cells, p38 mitogen-activated protein kinase and p21/WAF1/Cip1, were examined. A marked increase in phosphorylation of p38 was observed within 1 h after Pc 4-PDT without changes in levels of the p38 protein. Levels of p21 were not altered in the xenografts in correspondence with the presence of mutant p53 in SW480 cells. Evaluation of tumor size showed that tumor growth resumed after a delay of 9-15 days. Our results suggest that: (a) Pc 4-PDT is effective in the treatment of SW480 human colon cancer xenografts independent of p53 status; (b) PARP cleavage may be mediated by caspase-9 and caspase-3 activation in the Pc 4-PDT-treated tumors; and (c) p38 phosphorylation may be a trigger of apoptosis in response to PDT in vivo in this tumor model.

  9. [Report on the VDÄPC Fellowship].

    PubMed

    Sarantopoulos, E

    2016-04-01

    "Orandum est ut sit mens sana in corpore sano" ("You should pray for a healthy mind in a healthy body"). This phrase is a shortened citation from the satirical work of the Roman poet Juvenal. It highlights the significant role of physical as well as mental health. Aesthetic ideals have existed since the Archaic age, but they are not necessarily the same in different continents. Being familiar with aesthetic ideals in different cultures might help to accommodate patients' needs and wishes in today's globalised world. Therefore, fellowship programs such as the program organised by the Association of German Aesthetic Plastic Surgeons ("VDÄPC") are very important for young plastic surgeons who are interested in improving their surgical skills and experience. After all, aesthetic surgery is a dynamic specialty, which requires aesthetic plastic surgeons to undergo continued medical education.

  10. Genetic analysis of sex chromosomal meiotic mutants in Drosophilia melanogaster.

    PubMed

    Baker, B S; Carpenter, A T

    1972-06-01

    A total of 209 ethyl methanesulfonate-treated X chromosomes were screened for meiotic mutants that either (1) increased sex or fourth chromosome nondisjunction at either meiotic division in males; (2) allowed recombination in such males; (3) increased nondisjunction of the X chromosome at either meiotic division in females; or (4) caused such females, when mated to males heterozygous for Segregation-Distorter (SD) and a sensitive homolog to alter the strength of meiotic drive in males.-Twenty male-specific meiotic mutants were found. Though the rates of nondisjunction differed, all twenty mutants were qualitatively similar in that (1) they alter the disjunction of the X chromosome from the Y chromosome; (2) among the recovered sex-chromosome exceptional progeny, there is a large excess of those derived from nullo-XY as compared to XY gametes; (3) there is a negative correlation between the frequency of sex-chromosome exceptional progeny and the frequency of males among the regular progeny. In their effects on meiosis these mutants are similar to In(1)sc(4L)sc(8R), which is deleted for the basal heterochromatin. These mutants, however, have normal phenotypes and viabilities when examined as X/0 males, and furthermore, a mapping of two of the mutants places them in the euchromatin of the X chromosome. It is suggested that these mutants are in genes whose products are involved in insuring the proper functioning of the basal pairing sites which are deleted in In(1)sc(4L)sc(8R), and in addition that there is a close connection, perhaps causal, between the disruption of normal X-Y pairing (and, therefore, disjunction) and the occurrence of meiotic drive in the male.-Eleven mutants were found which increased nondisjunction in females. These mutants were characterized as to (1) the division at which they acted; (2) their effect on recombination; (3) their dominance; (4) their effects on disjunction of all four chromosome pairs. Five female mutants caused a nonuniform

  11. Pathway Controlled Penetration (PcP)

    SciTech Connect

    Knight, Earl E.; Rougier, Esteban; Zubelewicz, Aleksander

    2012-08-29

    The technical approach employs advanced computational simulation tools to demonstrate how current assets can destroy RWK-RFI-12-0001's HDBT, a tunnel complex with two portals built into the base of a granite mountain. The granite over layer is assumed to be 60 meters thick over both portals and 80 meters over the facility's mission space. Key S&T is the completed development of a highly innovative viscoplastic fracture material model, 3D parallel gas-fracture capabilities into FDEM, and a stochastic handling of the material properties. Phase I - Develop and validate code simulation tools: (1) develop, incorporate and validate AZ-Frac material model for granite; and (2) Develop and incorporate gas-driven-fracture modeling into LANL's FDEM MUNROU code; (3) Develop and incorporate stochastic features into FDEM modeling. Phase II - Conduct PcP analysis on above HDBT: (1) Acquire HDBT design data, develop simulation model; and (2) Evaluate and select most promising defeat alternative. Phase III - Deliver code, train Service target analysts, and conduct simulations against real world HDBTs. PcP uses advanced computer simulations to enhance HDBT functional defeat efforts. Newly developed material models that account for fractural energy coupled with the finite discrete element methodology (FDEM) will provide targeting packages that will create penetration avenues for current or future lethality options. This novel computational approach requires full 3D geologic and structure characterization as well as significant high performance computing capabilities. The goal is to distinctively alter the targeting paradigm by leveraging critical DoD assets along with insitu geologic strata. In other words, assets will utilize underground rock structure to their benefit by creating rubbilization zones that will allow pathway controlled penetration.

  12. Ibrutinib selectively and irreversibly targets EGFR (L858R, Del19) mutant but is moderately resistant to EGFR (T790M) mutant NSCLC Cells

    PubMed Central

    Wang, Wenchao; Hu, Chen; Ye, Zi; Zhao, Zheng; Wang, Li; Li, Xixiang; Yu, Kailin; Liu, Juan; Wu, Jiaxin; Yan, Xiao-E; Zhao, Peng; Wang, Jinhua; Wang, Chu; Weisberg, Ellen L.; Gray, Nathanael S.; Yun, Cai-Hong; Liu, Jing; Chen, Liang; Liu, Qingsong

    2015-01-01

    Through comprehensive comparison study, we found that ibrutinib, a clinically approved covalent BTK kinase inhibitor, was highly active against EGFR (L858R, del19) mutant driven NSCLC cells, but moderately active to the T790M ‘gatekeeper’ mutant cells and not active to wild-type EGFR NSCLC cells. Ibrutinib strongly affected EGFR mediated signaling pathways and induced apoptosis and cell cycle arrest (G0/G1) in mutant EGFR but not wt EGFR cells. However, ibrutinib only slowed down tumor progression in PC-9 and H1975 xenograft models. MEK kinase inhibitor, GSK1120212, could potentiate ibrutinib's effect against the EGFR (L858R/T790M) mutation in vitro but not in vivo. These results suggest that special drug administration might be required to achieve best clinical response in the ongoing phase I/II clinical trial with ibrutinib for NSCLC. PMID:26375053

  13. Rhizobium phaseoli symbiotic mutants with transposon Tn5 insertions.

    PubMed Central

    Noel, K D; Sanchez, A; Fernandez, L; Leemans, J; Cevallos, M A

    1984-01-01

    Rhizobium phaseoli CFN42 DNA was mutated by random insertion of Tn5 from suicide plasmid pJB4JI to obtain independently arising strains that were defective in symbiosis with Phaseolus vulgaris but grew normally outside the plant. When these mutants were incubated with the plant, one did not initiate visible nodule tissue (Nod-), seven led to slow nodule development (Ndv), and two led to superficially normal early nodule development but lacked symbiotic nitrogenase activity (Sna-). The Nod- mutant lacked the large transmissible indigenous plasmid pCFN42d that has homology to Klebsiella pneumoniae nitrogenase (nif) genes. The other mutants had normal plasmid content. In the two Sna- mutants and one Ndv mutant, Tn5 had inserted into plasmid pCFN42d outside the region of nif homology. The insertions of the other Ndv mutants were apparently in the chromosome. They were not in plasmids detected on agarose gels, and, in contrast to insertions on indigenous plasmids, they were transmitted in crosses to wild-type strain CFN42 at the same frequency as auxotrophic markers and with the same enhancement of transmission by conjugation plasmid R68.45. In these Ndv mutants the Tn5 insertions were the same as or very closely linked to mutations causing the Ndv phenotype. However, in two mutants with Tn5 insertions on plasmid pCFN42d, an additional mutation on the same plasmid, rather than Tn5, was responsible for the Sna- or Ndv phenotype. When plasmid pJB4JI was transferred to two other R. phaseoli strains, analysis of symbiotic mutants was complicated by Tn5-containing deleted forms of pJB4JI that were stably maintained. Images PMID:6325385

  14. Effect of a dominant inhibitory Ha-ras mutation on neuronal differentiation of PC12 cells.

    PubMed Central

    Szeberényi, J; Cai, H; Cooper, G M

    1990-01-01

    A dominant inhibitory mutation of Ha-ras which changes Ser-17 to Asn-17 in the gene product p21 [p21 (Asn-17)Ha-ras] has been used to investigate the role of ras in neuronal differentiation of PC12 cells. The growth of PC12 cells, in contrast to NIH 3T3 cells, was not inhibited by p21(Asn-17)Ha-ras expression. However, PC12 cells expressing the mutant Ha-ras protein showed a marked inhibition of morphological differentiation induced by nerve growth factor (NGF) or fibroblast growth factor (FGF). These cells, however, were still able to respond with neurite outgrowth to dibutyryl cyclic AMP and 12-O-tetradecanoylphorbol-13-acetate (TPA). Induction of early-response genes (fos, jun, and zif268) by NGF and FGF but not by TPA was also inhibited by high levels of p21(Asn-17)Ha-ras. However, lower levels of p21(Asn-17) expression were sufficient to block neuronal differentiation without inhibiting induction of these early-response genes. Induction of the secondary-response genes SCG10 and transin by NGF, like morphological differentiation, was inhibited by low levels of p21(Asn-17) whether or not induction of early-response genes was blocked. Therefore, although inhibition of ras function can inhibit early-response gene induction, this is not required to block morphological differentiation or secondary-response gene expression. These results suggest that ras proteins are involved in at least two different pathways of signal transduction from the NGF receptor, which can be distinguished by differential sensitivity to p21(Asn-17)Ha-ras. In addition, ras and protein kinase C can apparently induce early-response gene expression by independent pathways in PC12 cells. Images PMID:2118994

  15. Microcomputer Applications with PC LAN (Local Area Network) in Battleships.

    DTIC Science & Technology

    1988-12-01

    9 G. INTEL 80386 MICROPROCESSOR ARCHITECTURE 9 1. Cache Strategy 10 2. Memory Management 10 3. Multitasking 11 4. Software...shrinks, it becomes more clear that the traditional modes of battle planning and management are no longer practical. Many of these problems, like...Card in it. Each PC must run the PC Network program to manage its participation in the network. Each PC must have a cable to connect it to the

  16. ELIPGRID-PC: A PC program for calculating hot spot probabilities

    SciTech Connect

    Davidson, J.R.

    1994-10-01

    ELIPGRID-PC, a new personal computer program has been developed to provide easy access to Singer`s 1972 ELIPGRID algorithm for hot-spot detection probabilities. Three features of the program are the ability to determine: (1) the grid size required for specified conditions, (2) the smallest hot spot that can be sampled with a given probability, and (3) the approximate grid size resulting from specified conditions and sampling cost. ELIPGRID-PC also provides probability of hit versus cost data for graphing with spread-sheets or graphics software. The program has been successfully tested using Singer`s published ELIPGRID results. An apparent error in the original ELIPGRID code has been uncovered and an appropriate modification incorporated into the new program.

  17. Features of Pc5 pulsations in the geomagnetic field, auroral luminosity, and Riometer absorption

    NASA Astrophysics Data System (ADS)

    Belakhovsky, V. B.; Pilipenko, V. A.; Samsonov, S. N.; Lorentsen, D.

    2016-01-01

    Simultaneous morning Pc5 pulsations ( f ~ 3-5 mHz) in the geomagnetic field, aurora intensities (in the 557.7 and 630.0 nm oxygen emissions and the 471.0 nm nitrogen emission), and riometer absorption, were studied based on the CARISMA, CANMOS, and NORSTAR network data for the event of January 1, 2000. According to the GOES-8 satellite observations, these Pc5 geomagnetic pulsations are observed as incompressible Alfvén waves with toroidal polarization in the magnetosphere. Although the Pc5 pulsation frequencies in auroras, the geomagnetic field, and riometer absorption are close to one another, stable phase relationships are not observed between them. Far from all trains of geomagnetic Pc5 pulsations are accompanied by corresponding auroral pulsations; consequently, geomagnetic pulsations are primary with respect to auroral pulsations. Both geomagnetic and auroral pulsations propagate poleward, and the frequency decreases with increasing geomagnetic latitude. When auroral Pc5 pulsations appear, the ratio of the 557.7/630.0 nm emission intensity sharply increases, which indicates that auroral pulsations result from not simply modulated particle precipitation but also an additional periodic acceleration of auroral electrons by the wave field. A high correlation is not observed between Pc5 pulsations in auroras and the riometer absorption, which indicates that these pulsations have a common source but different generation mechanisms. Auroral luminosity modulation is supposedly related to the interaction between Alfvén waves and the region with the field-aligned potential drop above the auroral ionosphere, and riometer absorption modulation is caused by the scattering of energetic electrons by VLF noise pulsations.

  18. Electronic properties of CuPc and H2Pc: an experimental and theoretical study.

    PubMed

    Nardi, Marco Vittorio; Detto, Francesca; Aversa, Lucrezia; Verucchi, Roberto; Salviati, Giancarlo; Iannotta, Salvatore; Casarin, Maurizio

    2013-08-21

    Phthalocyanine (H2Pc) and its open-shell copper complex (CuPc) deposited on amorphous gold films have been studied by combining the outcomes of several synchrotron based spectroscopic tools (X-ray photoelectron spectroscopy, UV photoelectron spectroscopy and near-edge X-ray absorption fine structure, NEXAFS, spectroscopy) with those of density functional theory (DFT) calculations. The assignment of experimental evidence has been guided by the results of DFT numerical experiments carried out on isolated molecules. With specific reference to CuPc NEXAFS data collected at the N K-edge, they have been assigned by using the open-shell time-dependent DFT (TDDFT) in the framework of the zeroth order regular approximation (ZORA) scalar relativistic approach. The agreement between theory and experiment has been found to be satisfactory, thus indicating that the open-shell TDDFT (F. Wang and T. Ziegler, Mol. Phys., 2004, 102, 2585) may be used with some confidence to look into the X-ray absorption spectroscopy results pertinent to transition metal complexes. As far as the metal-ligand interaction is concerned, the combined use of NEXAFS spectroscopy and DFT outcomes ultimately testified the significant ionic contribution characterizing the bonding between the metal centre and the nitrogen atoms of the phthalocyanine coordinative pocket.

  19. Cerebellar Expression of the Neurotrophin Receptor p75 in Naked-Ataxia Mutant Mouse.

    PubMed

    Rahimi Balaei, Maryam; Jiao, Xiaodan; Ashtari, Niloufar; Afsharinezhad, Pegah; Ghavami, Saeid; Marzban, Hassan

    2016-01-15

    Spontaneous mutation in the lysosomal acid phosphatase 2 (Acp2) mouse (nax--naked-ataxia mutant mouse) correlates with severe cerebellar defects including ataxia, reduced size and abnormal lobulation as well as Purkinje cell (Pc) degeneration. Loss of Pcs in the nax cerebellum is compartmentalized and harmonized to the classic pattern of gene expression of the cerebellum in the wild type mouse. Usually, degeneration starts in the anterior and posterior zones and continues to the central and nodular zones of cerebellum. Studies have suggested that the p75 neurotrophin receptor (NTR) plays a role in Pc degeneration; thus, in this study, we investigated the p75NTR pattern and protein expression in the cerebellum of the nax mutant mouse. Despite massive Pc degeneration that was observed in the nax mouse cerebellum, p75NTR pattern expression was similar to the HSP25 pattern in nax mice and comparable with wild type sibling cerebellum. In addition, immunoblot analysis of p75NTR protein expression did not show any significant difference between nax and wild type sibling (p > 0.5). In comparison with wild type counterparts, p75NTR pattern expression is aligned with the fundamental cytoarchitecture organization of the cerebellum and is unchanged in the nax mouse cerebellum despite the severe neurodevelopmental disorder accompanied with Pc degeneration.

  20. [Isolation and primary identification of methylotrophic yeast Hansenula polymorpha mutants for peroxisome biogenesis].

    PubMed

    Kurbatova, E M; Dutova, T A; Serkova, N N; Rabinovich, Ia M; Trotsenko, Iu A

    2004-05-01

    After exposure of cells of the methylotrophic yeast Hansenula polymorpha HF246 leu1-1 to N-nitro-N-nitrosoguanidine, a collection of 227 mutants unable to grow on methanol at elevated temperature (45 degrees C) was obtained. Ninety four ts mutants (35% of the total number of mutants), which were unable to grow on methanol only at 45 degrees C but could grow at optimal temperature (37 degrees C), were isolated. Complementation analysis of mutants using 12 deletion mutants for genes of peroxisome biogenesis (PEX) (available in this yeast species by the beginning of our work) allowed to assign 51 mutants (including 16 ts) to the separate group of mutants unable to complement deletion mutants with defects in eight PEX genes. These mutants were classified into three groups: group 1 contained 10 pex10 mutants (4 ts mutants among them); group 2 included 19 mutants that failed to complement other pex testers: 1 pex1; 2 pex4 (1 ts); 6 pex5 (5 ts); 3 pex8; 6 (3ts)- pex19; group 3 contained 22 "multiple" mutants. In mutants of group 3, hybrids with several testers do not grow on methanol. All mutants (51) carried recessive mutations, except for mutant 108, in which the mutation was dominant only at 30 degrees C, which suggests that it is ts-dominant. Recombination analysis of mutants belonging to group 2 revealed that only five mutants (two pex5 and three pex8) carried mutations for the corresponding PEX genes. The remaining 14 mutants yielded methanol-utilizing segregants in an arbitrarily chosen sample of hybrids with the pex tester, which indicates mutation location in other genes. In 19 mutants, random analysis of ascospores from hybrids obtained upon crossing mutants of group 3 with a strain lacking peroxisomal disorders (ade11) revealed a single mutation causing the appearance of a multiple phenotype. A more detailed study of two mutants from this group allowed the localization of this mutation in the only PEX gene (PEX or PEX2). The revealed disorder of complementation

  1. Regulation of phospholipid synthesis in phosphatidylserine synthase-deficient (chol) mutants of Saccharomyces cerevisiae

    SciTech Connect

    Letts, V.A.; Henry, S.A.

    1985-08-01

    Saccharomyces cerevisiae mutants, chol, are deficient in the synthesis of the phospholipid phosphatidylserine owing to lowered activity of the membrane-associated enzyme phosphatidylserine synthase. These mutants are auxotrophic for ethanolamine or choline and, in the absence of these supplements, cannot synthesize phosphatidylethanolamine or phosphatidylcholine (PC). The authors exploited these characteristics of the chol mutants to examine the regulation of phospholipid metabolism in S. cerevisiae. Macromolecular synthesis and phospholipid metabolism were examined in chol cells starved for ethanolamine. Coupled to the decline in PC biosynthesis was a simultaneous decrease in the overall rate of phospholipid synthesis. In particular, the rate of synthesis of phosphatidylinositol decreased in parallel with the decline in PC biosynthesis. However, under conditions of ethanolamine deprivation in chol cells, the cytoplasmic enzyme inositol-1-phosphate synthase could not be repressed by exogenous inositol, and the endogenous synthesis of the phospholipid precursor inositol appeared to be elevated. The implications of these findings with respect to the coordinated regulation of phospholipid synthesis are discussed.

  2. Inhibition of Prohormone Convertases PC1/3 and PC2 by 2,5-Dideoxystreptamine Derivatives

    PubMed Central

    Vivoli, Mirella; Caulfield, Thomas R.; Martínez-Mayorga, Karina; Johnson, Alan T.; Jiao, Guan-Sheng

    2012-01-01

    The prohormone convertases PC1/3 and PC2 are eukaryotic serine proteases involved in the proteolytic maturation of peptide hormone precursors and are implicated in a variety of pathological conditions, including obesity, diabetes, and neurodegenerative diseases. In this work, we screened 45 compounds obtained by derivatization of a 2,5-dideoxystreptamine scaffold with guanidinyl and aryl substitutions for convertase inhibition. We identified four promising PC1/3 competitive inhibitors and three PC2 inhibitors that exhibited various inhibition mechanisms (competitive, noncompetitive, and mixed), with sub- and low micromolar inhibitory potency against a fluorogenic substrate. Low micromolar concentrations of certain compounds blocked the processing of the physiological substrate proglucagon. The best PC2 inhibitor effectively inhibited glucagon synthesis, a known PC2-mediated process, in a pancreatic cell line; no cytotoxicity was observed. We also identified compounds that were able to stimulate both 87 kDa PC1/3 and PC2 activity, behavior related to the presence of aryl groups on the dideoxystreptamine scaffold. By contrast, inhibitory activity was associated with the presence of guanidinyl groups. Molecular modeling revealed interactions of the PC1/3 inhibitors with the active site that suggest structural modifications to further enhance potency. In support of kinetic data suggesting that PC2 inhibition probably occurs via an allosteric mechanism, we identified several possible allosteric binding sites using computational searches. It is noteworthy that one compound was found to both inhibit PC2 and stimulate PC1/3. Because glucagon acts in functional opposition to insulin in blood glucose homeostasis, blocking glucagon formation and enhancing proinsulin cleavage with a single compound could represent an attractive therapeutic approach in diabetes. PMID:22169851

  3. Quantification of PrPC in bovine peripheral tissues: Analysis in wild-type and PrPC-deficient cattle.

    PubMed

    Kobayashi, Shin-Ichi; Ano, Yasuhisa; Sakudo, Akikazu; Yukawa, Masayoshi; Sigiura, Katsuaki; Manabe, Noboru; Nakayama, Hiroyuki; Onodera, Takashi

    2009-01-01

    Cellular PrP (PrPC) is necessary for bovine spongiform encephalopathy (BSE) infection. The purpose of the present experiment was the quantification of PrPC in peripheral tissues to assess the risk of BSE infection from these tissues. The tissue distribution of PrPC was examined by a sandwich enzyme-linked immunosorbent assay (sELISA) and histochemical analysis. PrPC-deficient cows were used as a negative control. The sELISA revealed that the brain contained the highest PrPC content (10.7 µg/g tissue), while other organs/tissues harbored lower amounts, in decreasing order as follows: longissimus capitis muscle, iliocostalis thoracis muscle, splenius muscle, biceps femoris muscle, triceps brachii muscle, longissimus thoracis muscle, ileum, jejunum, duodenum, colon, cecum, apex linguae, omotransversarius muscle, posterior part of the corpus linguae, anterior part of the corpus linguae and radix linguae (5.2- to 31-fold less PrPC than the brain). In the tissue/organs of PrP-deficient cows, PrPC levels were under the limit of detection. Histochemical analysis showed that PrPC was expressed in nerve cells in intestinal tissues. The presence of PrPC in the bovine tongue, skeletal muscles and intestines raises the possibility of PrPSc accumulation in these tissues, indicating that these organs/tissues may serve as potential sources of BSE infection.

  4. Physical Properties of PC-PMMA Multilayers

    NASA Astrophysics Data System (ADS)

    Rahman, Arifur; Baer, Eric; Chipara, Alin Cristian; Vajtai, Robert; Ajayan, Pullickel M.; Hinthorne, James; Elamin, Ibrahim; Chipara, Mircea; Eric Baer Collaboration; Pullickel Ajayan Collaboration; Mircea Chipara Collaboration

    2015-03-01

    Multilayers of polycarbonate (PC) and polymethylmethacrylate (PMMA) have been obtained by the layer multiplying coextrusion method. Each sample (1024 layers, of equal thickness, with individual thickness between 10 and 200 nm) has been investigated at room temperature by Wide Angle X-Ray Scattering (WAXS) using a Bruker Discovery 8 spectrometer (Cu K α radiation), Raman spectroscopy (Bruker Senterra confocal Raman spectrometer operating at 785 nm), FTIR spectroscopy (Tensor 27 Bruker), and UV-Vis spectroscopy. Further details about the glass transition temperature in these samples have been obtained by Dynamical Mechanical Analysis, DMA, (TA Instruments Q800) at various frequencies in the range 1 to 100 Hz. Isothermal Differential Scanning Calorimetry, DSC, (TA Instruments Q200) was used to investigate the effect of the thickness of the polymeric film on the crystallization processes. Non-isothermal DSC measurements aimed at the identification and location of the main phase transitions (glass, crystallization, and melting) occurring in these multilayers. The effects of confinement on the phase transitions occurring in these multilayers are discussed in detail.

  5. Pc-Based Floating Point Imaging Workstation

    NASA Astrophysics Data System (ADS)

    Guzak, Chris J.; Pier, Richard M.; Chinn, Patty; Kim, Yongmin

    1989-07-01

    The medical, military, scientific and industrial communities have come to rely on imaging and computer graphics for solutions to many types of problems. Systems based on imaging technology are used to acquire and process images, and analyze and extract data from images that would otherwise be of little use. Images can be transformed and enhanced to reveal detail and meaning that would go undetected without imaging techniques. The success of imaging has increased the demand for faster and less expensive imaging systems and as these systems become available, more and more applications are discovered and more demands are made. From the designer's perspective the challenge to meet these demands forces him to attack the problem of imaging from a different perspective. The computing demands of imaging algorithms must be balanced against the desire for affordability and flexibility. Systems must be flexible and easy to use, ready for current applications but at the same time anticipating new, unthought of uses. Here at the University of Washington Image Processing Systems Lab (IPSL) we are focusing our attention on imaging and graphics systems that implement imaging algorithms for use in an interactive environment. We have developed a PC-based imaging workstation with the goal to provide powerful and flexible, floating point processing capabilities, along with graphics functions in an affordable package suitable for diverse environments and many applications.

  6. Attention selection, distractor suppression and N2pc.

    PubMed

    Mazza, Veronica; Turatto, Massimo; Caramazza, Alfonso

    2009-01-01

    N2pc is generally interpreted as the electrocortical correlate of the distractor-suppression mechanisms through which attention selection takes place in humans. Here, we present data that challenge this common N2pc interpretation. In Experiment 1, multiple distractors induced greater N2pc amplitudes even when they facilitated target identification, despite the suppression account of the N2pc predicted the contrary; in Experiment 2, spatial proximity between target and distractors did not affect the N2pc amplitude, despite resulting in more interference in response times; in Experiment 3, heterogeneous distractors delayed response times but did not elicit a greater N2pc relative to homogeneous distractors again in contrast with what would have predicted the suppression hypothesis. These results do not support the notion that the N2pc unequivocally mirrors distractor-suppression processes. We propose that the N2pc indexes mechanisms involved in identifying and localizing relevant stimuli in the scene through enhancement of their features and not suppression of distractors.

  7. PC2 Ovotransferrin: Characterization and Alternative Immunotherapeutic Activity

    PubMed Central

    Chiurciu, Constantin; Chiurciu, Viorica; Oporanu, Mariana; Pătrașcu, Ionel Victor; Mihai, Iuliana; Tablică, Mădălina

    2017-01-01

    Characterization and evaluation of immunotherapeutic potential of ovotransferrin PC2 (OTf PC2) were performed in this study. The ovoprotein was obtained from egg white from hens immunized with bacterial antigens, pathogenic for humans. For the negative control samples, OTf was extracted from eggs collected from Specific Pathogen-Free (SPF) hens and purified by affinity chromatography on Protein G-agarose column with two eluting peaks: I, representing ovalbumin, and II, ovotransferrin. The final apo-OTf form was reached by successive precipitation with ammonium sulfate and citric acid and the holo-OTf form by saturating the apo-form with FeCl3. Multiple OTf PC2 samples were analyzed through Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE) and, based on the molecular marker migration model, the ovotransferrin (76.5 kDa) and ovalbumin (45 kDa) were detected. The agglutination reaction exhibited statistically significant high specificity of the multiple OTf PC2, by reacting with the antigens used for hens' immunization. Following ELISA, it was established that OTf PC2 from hyperimmune eggs has specificity for all antigens; the antibody titer was high, indicating that OTf PC2 possesses immunological properties similar to immunoglobulin Y (Ig Y). This study suggests that OTf PC2 immunological activity may play a crucial role in the prevention and treatment of infections resistant to antibiotics and OTf PC2 can also act as a valuable nutraceutical.

  8. Chromosomal Distribution of PcG Proteins during Drosophila Development

    PubMed Central

    Nègre, Nicolas; Hennetin, Jérôme; Sun, Ling V; Lavrov, Sergey; Bellis, Michel; White, Kevin P

    2006-01-01

    Polycomb group (PcG) proteins are able to maintain the memory of silent transcriptional states of homeotic genes throughout development. In Drosophila, they form multimeric complexes that bind to specific DNA regulatory elements named PcG response elements (PREs). To date, few PREs have been identified and the chromosomal distribution of PcG proteins during development is unknown. We used chromatin immunoprecipitation (ChIP) with genomic tiling path microarrays to analyze the binding profile of the PcG proteins Polycomb (PC) and Polyhomeotic (PH) across 10 Mb of euchromatin. We also analyzed the distribution of GAGA factor (GAF), a sequence-specific DNA binding protein that is found at most previously identified PREs. Our data show that PC and PH often bind to clustered regions within large loci that encode transcription factors which play multiple roles in developmental patterning and in the regulation of cell proliferation. GAF co-localizes with PC and PH to a limited extent, suggesting that GAF is not a necessary component of chromatin at PREs. Finally, the chromosome-association profile of PC and PH changes during development, suggesting that the function of these proteins in the regulation of some of their target genes might be more dynamic than previously anticipated. PMID:16613483

  9. Hydrogen bonds in PC{sub 61}BM solids

    SciTech Connect

    Sheng, Chun-Qi; Li, Wen-Jie; Du, Ying-Ying; Chen, Guang-Hua; Chen, Zheng; Li, Hai-Yang; Li, Hong-Nian

    2015-09-15

    We have studied the hydrogen bonds in PC{sub 61}BM solids. Inter-molecular interaction is analyzed theoretically for the well-defined monoclinic (P2{sub 1}/n) structure. The results indicate that PC{sub 61}BM combines into C–H⋯O{sub d} bonded molecular chains, where O{sub d} denotes the doubly-bonded O atom of PC{sub 61}BM. The molecular chains are linked together by C–H⋯O{sub s} bonds, where O{sub s} denotes the singly-bonded O atom of PC{sub 61}BM. To reveal the consequences of hydrogen bond formation on the structural properties of PC{sub 61}BM solids (not limited to the monoclinic structure), we design and perform some experiments for annealed samples with the monoclinic (P2{sub 1}/n) PC{sub 61}BM as starting material. The experiments include differential scanning calorimetry, X-ray diffraction and infrared absorption measurements. Structural phase transitions are observed below the melting point. The C–H⋯O{sub d} bonds seem persisting in the altered structures. The inter-molecular hydrogen bonds can help to understand the phase separation in polymer/PC{sub 61}BM blends and may be responsible for the existence of liquid PC{sub 61}BM.

  10. PrPC from stem cells to cancer

    PubMed Central

    Martin-Lannerée, Séverine; Hirsch, Théo Z.; Hernandez-Rapp, Julia; Halliez, Sophie; Vilotte, Jean-Luc; Launay, Jean-Marie; Mouillet-Richard, Sophie

    2014-01-01

    The cellular prion protein PrPC was initially discovered as the normal counterpart of the pathological scrapie prion protein PrPSc, the main component of the infectious agent of Transmissible Spongiform Encephalopathies. While clues as to the physiological function of this ubiquitous protein were greatly anticipated from the development of knockout animals, PrP-null mice turned out to be viable and to develop without major phenotypic abnormalities. Notwithstanding, the discovery that hematopoietic stem cells from PrP-null mice have impaired long-term repopulating potential has set the stage for investigating into the role of PrPC in stem cell biology. A wealth of data have now exemplified that PrPC is expressed in distinct types of stem cells and regulates their self-renewal as well as their differentiation potential. A role for PrPC in the fate restriction of embryonic stem cells has further been proposed. Paralleling these observations, an overexpression of PrPC has been documented in various types of tumors. In line with the contribution of PrPC to stemness and to the proliferation of cancer cells, PrPC was recently found to be enriched in subpopulations of tumor-initiating cells. In the present review, we summarize the current knowledge of the role played by PrPC in stem cell biology and discuss how the subversion of its function may contribute to cancer progression. PMID:25364760

  11. Transforming PC Power Supplies into Smart Car Battery Conditioners

    ERIC Educational Resources Information Center

    Rodriguez-Ascariz, J. M.; Boquete-Vazquez, L.

    2011-01-01

    This paper describes a laboratory project consisting of a PC power supply modification into an intelligent car-battery conditioner with both wireless and wired networking capabilities. Adding a microcontroller to an average PC power supply transforms it into a flexible, intelligent device that can be configured and that is suitable to keep car…

  12. Anticancer effects of ethanolic neem leaf extract on prostate cancer cell line (PC-3).

    PubMed

    Kumar, Suresh; Suresh, P K; Vijayababu, M R; Arunkumar, A; Arunakaran, J

    2006-04-21

    Prostate cancer (PC) is the most prevalent cancer and the leading cause of male cancer death. Azadirachta indica (neem tree) has been used successfully centuries to reduce tumors by herbalists throughout Southeast Asia. Here the present study indicated that an ethanolic extract of neem has been shown to cause cell death of prostate cancer cells (PC-3) by inducing apoptosis as evidenced by a dose-dependent increase in DNA fragmentation and a decrease in cell viability. Western blot studies indicated that treatment with neem extract showed decreased level of Bcl-2, which is anti-apoptotic protein and increased the level of Bax protein. So the neem extract could be potentially effective against prostate cancer treatment.

  13. Plasma mutant α-galactosidase A protein and globotriaosylsphingosine level in Fabry disease.

    PubMed

    Tsukimura, Takahiro; Nakano, Sachie; Togawa, Tadayasu; Tanaka, Toshie; Saito, Seiji; Ohno, Kazuki; Shibasaki, Futoshi; Sakuraba, Hitoshi

    2014-01-01

    Fabry disease is an X-linked genetic disorder characterized by deficient activity of α-galactosidase A (GLA) and accumulation of glycolipids, and various GLA gene mutations lead to a wide range of clinical phenotypes from the classic form to the later-onset one. To investigate the biochemical heterogeneity and elucidate the basis of the disease using available clinical samples, we measured GLA activity, GLA protein and accumulated globotriaosylsphingosine (Lyso-Gb3), a biomarker of this disease, in plasma samples from Fabry patients. The analysis revealed that both the enzyme activity and the protein level were apparently decreased, and the enzyme activity was well correlated with the protein level in many Fabry patients. In these cases, a defect of biosynthesis or excessive degradation of mutant GLAs should be involved in the pathogenesis, and the residual protein level would determine the accumulation of Lyso-Gb3 and the severity of the disease. However, there are some exceptional cases, i.e., ones harboring p.C142Y, p.R112H and p.M296I, who exhibit a considerable amount of GLA protein. Especially, a subset of Fabry patients with p.R112H or p.M296I has been attracted interest because the patients exhibit almost normal plasma Lyso-Gb3 concentration. Structural analysis revealed that C142Y causes a structural change at the entrance of the active site. It will lead to a complete enzyme activity deficiency, resulting in a high level of plasma Lyso-Gb3 and the classic Fabry disease. On the other hand, it is thought that R112H causes a relatively large structural change on the molecular surface, and M296I a small one in a restricted region from the core to the surface, both the structural changes being far from the active site. These changes will cause not only partial degradation but also degeneration of the mutant GLA proteins, and the degenerated enzymes exhibiting small and residual activity remain and probably facilitate degradation of Lyso-Gb3 in plasma, leading

  14. Monte Carlo tests of the ELIPGRID-PC algorithm

    SciTech Connect

    Davidson, J.R.

    1995-04-01

    The standard tool for calculating the probability of detecting pockets of contamination called hot spots has been the ELIPGRID computer code of Singer and Wickman. The ELIPGRID-PC program has recently made this algorithm available for an IBM{reg_sign} PC. However, no known independent validation of the ELIPGRID algorithm exists. This document describes a Monte Carlo simulation-based validation of a modified version of the ELIPGRID-PC code. The modified ELIPGRID-PC code is shown to match Monte Carlo-calculated hot-spot detection probabilities to within {plus_minus}0.5% for 319 out of 320 test cases. The one exception, a very thin elliptical hot spot located within a rectangular sampling grid, differed from the Monte Carlo-calculated probability by about 1%. These results provide confidence in the ability of the modified ELIPGRID-PC code to accurately predict hot-spot detection probabilities within an acceptable range of error.

  15. Habituation in the Single Cell: Diminished Secretion of Norepinephrine with Repetitive Depolarization of PC12 Cells

    NASA Astrophysics Data System (ADS)

    McFadden, Philip N.; Koshland, Daniel E., Jr.

    1990-03-01

    Neuronally differentiated PC12 cells secrete decreasing amounts of [^3H]norepinephrine when repetitively stimulated by depolarizing concentrations of potassium ion. The decreasing response shows attributes that have been classically ascribed to response habituation, a behavior commonly observed in nervous systems but found here in a homogeneous cell type. Alteration of the habituation pattern was caused by activators of the protein kinase C pathway and of voltage-gated calcium channels.

  16. Deficiency in prohormone convertase PC1 impairs prohormone processing in Prader-Willi syndrome

    PubMed Central

    Burnett, Lisa C.; LeDuc, Charles A.; Sulsona, Carlos R.; Paull, Daniel; Rausch, Richard; Eddiry, Sanaa; Carli, Jayne F. Martin; Morabito, Michael V.; Skowronski, Alicja A.; Hubner, Gabriela; Zimmer, Matthew; Wang, Liheng; Day, Robert; Levy, Brynn; Dubern, Beatrice; Poitou, Christine; Clement, Karine; Rosenbaum, Michael; Salles, Jean Pierre; Tauber, Maithe; Egli, Dieter

    2016-01-01

    Prader-Willi syndrome (PWS) is caused by a loss of paternally expressed genes in an imprinted region of chromosome 15q. Among the canonical PWS phenotypes are hyperphagic obesity, central hypogonadism, and low growth hormone (GH). Rare microdeletions in PWS patients define a 91-kb minimum critical deletion region encompassing 3 genes, including the noncoding RNA gene SNORD116. Here, we found that protein and transcript levels of nescient helix loop helix 2 (NHLH2) and the prohormone convertase PC1 (encoded by PCSK1) were reduced in PWS patient induced pluripotent stem cell–derived (iPSC-derived) neurons. Moreover, Nhlh2 and Pcsk1 expression were reduced in hypothalami of fasted Snord116 paternal knockout (Snord116p–/m+) mice. Hypothalamic Agrp and Npy remained elevated following refeeding in association with relative hyperphagia in Snord116p–/m+ mice. Nhlh2-deficient mice display growth deficiencies as adolescents and hypogonadism, hyperphagia, and obesity as adults. Nhlh2 has also been shown to promote Pcsk1 expression. Humans and mice deficient in PC1 display hyperphagic obesity, hypogonadism, decreased GH, and hypoinsulinemic diabetes due to impaired prohormone processing. Here, we found that Snord116p–/m+ mice displayed in vivo functional defects in prohormone processing of proinsulin, pro-GH–releasing hormone, and proghrelin in association with reductions in islet, hypothalamic, and stomach PC1 content. Our findings suggest that the major neuroendocrine features of PWS are due to PC1 deficiency. PMID:27941249

  17. Color Shift Investigations for LED Secondary Optical Designs: Comparison between BPA-PC and PMMA

    NASA Astrophysics Data System (ADS)

    Lu, Guangjun; Yazdan Mehr, M.; van Driel, W. D.; Fan, Xuejun; Fan, Jiajie; Jansen, K. M. B.; Zhang, G. Q.

    2015-07-01

    Recently, color shift of LED-based lighting products has attracted much attention due to its increasing impact on the field application. However, significant research investigations on the color shift mechanisms are not publically available especially for important transmission materials used for secondary optical design. In this paper, broadly used such commercial materials (BPA-PC and PMMA) are experimentally investigated on the color shift effects during aging. Besides this, color shift mechanisms of degradation of transmittance are also studied. Results revealed: (1) Inconsistent degradation of wavelength-dependent transmittance induces the decrease of the blue/yellow light intensity ratio and thus gives rise to the color shift toward the yellow field, which is the color shift mechanism of BPA-PC; (2) Even for the non-aged BPA-PC, the transmittance varies with wavelength in the visible light field due to the chemistry of the material, which caused the change of intensify ratio of blue light to yellow light in the SPD, leading to color change in perception; (3) Oxidation plays a key role in the degradation of transmittance at around the peak wavelength of the blue light field, which is in correlation with the discoloration of thermally-aged BPA-PC materials. By contrast, for the PMMA specimen aged up to 3000 h, oxidation was neither occurred at 85 °C, nor with additional exposure to blue light, nor even with additional humidity of 85%RH.

  18. MtDNA depleted PC3 cells exhibit Warburg effect and cancer stem cell features

    PubMed Central

    Li, Xiaoran; Zhong, Yali; Lu, Jie; Axcrona, Karol; Eide, Lars; Syljuåsen, Randi G.; Peng, Qian; Wang, Junbai; Zhang, Hongquan; Goscinski, Mariusz Adam; Kvalheim, Gunnar; Nesland, Jahn M.; Suo, Zhenhe

    2016-01-01

    Reducing mtDNA content was considered as a critical step in the metabolism restructuring for cell stemness restoration and further neoplastic development. However, the connections between mtDNA depletion and metabolism reprograming-based cancer cell stemness in prostate cancers are still lack of studies. Here, we demonstrated that human CRPC cell line PC3 tolerated high concentration of the mtDNA replication inhibitor ethidium bromide (EtBr) and the mtDNA depletion triggered a universal metabolic remodeling process. Failure in completing that process caused lethal consequences. The mtDNA depleted (MtDP) PC3 cells could be steadily maintained in the special medium in slow cycling status. The MtDP PC3 cells contained immature mitochondria and exhibited Warburg effect. Furthermore, the MtDP PC3 cells were resistant to therapeutic treatments and contained greater cancer stem cell-like subpopulations: CD44+, ABCG2+, side-population and ALDHbright. In conclusion, these results highlight the association of mtDNA content, mitochondrial function and cancer cell stemness features. PMID:27248169

  19. Structure of the polycystic kidney disease TRP channel Polycystin-2 (PC2).

    PubMed

    Grieben, Mariana; Pike, Ashley C W; Shintre, Chitra A; Venturi, Elisa; El-Ajouz, Sam; Tessitore, Annamaria; Shrestha, Leela; Mukhopadhyay, Shubhashish; Mahajan, Pravin; Chalk, Rod; Burgess-Brown, Nicola A; Sitsapesan, Rebecca; Huiskonen, Juha T; Carpenter, Elisabeth P

    2017-02-01

    Mutations in either polycystin-1 (PC1 or PKD1) or polycystin-2 (PC2, PKD2 or TRPP1) cause autosomal-dominant polycystic kidney disease (ADPKD) through unknown mechanisms. Here we present the structure of human PC2 in a closed conformation, solved by electron cryomicroscopy at 4.2-Å resolution. The structure reveals a novel polycystin-specific 'tetragonal opening for polycystins' (TOP) domain tightly bound to the top of a classic transient receptor potential (TRP) channel structure. The TOP domain is formed from two extensions to the voltage-sensor-like domain (VSLD); it covers the channel's endoplasmic reticulum lumen or extracellular surface and encloses an upper vestibule, above the pore filter, without blocking the ion-conduction pathway. The TOP-domain fold is conserved among the polycystins, including the homologous channel-like region of PC1, and is the site of a cluster of ADPKD-associated missense variants. Extensive contacts among the TOP-domain subunits, the pore and the VSLD provide ample scope for regulation through physical and chemical stimuli.

  20. Cytotoxic, Genotoxic, and Neurotoxic Effects of Mg, Pb, and Fe on Pheochromocytoma (PC-12) Cells

    PubMed Central

    Sanders, Talia; Liu, Yi-Ming; Tchounwou, Paul B.

    2014-01-01

    Metals such as lead (Pb), magnesium (Mg), and iron (Fe) are ubiquitous in the environment as a result of natural occurrence and anthropogenic activities. Although Mg, Fe and others are considered essential elements, high level of exposure has been associated with severe adverse health effects including cardiovascular, hematological, nephrotoxic, hepatotoxic, and neurologic abnormalities in humans. In the present study we hypothesized that Mg, Pb, and Fe are cytotoxic, genotoxic and neurotoxic, and their toxicity is mediated through oxidative stress and alteration in protein expression. To test the hypothesis, we used the pheochromocytoma (PC-12) cell line as a neuro cell model and performed the LDH assay for cell viability, Comet assay for DNA damage, Western blot for oxidative stress, and HPLC-MS to assess the concentration levels of neurological biomarkers such as glutamate, dopamine (DA), and 3-methoxytyramine (3-MT). The results of this study clearly show that Mg, Pb, and Fe, respectively in the form of MgSO4, Pb(NO3)2, FeCl2, and FeCl3 induce cytotoxicity, oxidative stress, and genotoxicity in PC-12 cells. In addition, exposure to these metallic compounds caused significant changes in the concentration levels of glutamate, dopamine, and 3-MT in PC-12 cells. Taken together the findings suggest that MgSO4, Pb(NO3)2, FeCl2, and FeCl3 have the potential to induce substantial toxicity to PC-12 cells. PMID:24942330

  1. PC7 and the related proteases Furin and Pace4 regulate E-cadherin function during blastocyst formation.

    PubMed

    Bessonnard, Sylvain; Mesnard, Daniel; Constam, Daniel B

    2015-09-28

    The first cell differentiation in mammalian embryos segregates polarized trophectoderm cells from an apolar inner cell mass (ICM). This lineage decision is specified in compacted morulae by cell polarization and adhesion acting on the Yes-associated protein in the Hippo signaling pathway, but the regulatory mechanisms are unclear. We show that morula compaction and ICM formation depend on PC7 and the related proprotein convertases (PCs) Furin and Pace4 and that these proteases jointly regulate cell-cell adhesion mediated by E-cadherin processing. We also mapped the spatiotemporal activity profiles of these proteases by live imaging of a transgenic reporter substrate in wild-type and PC mutant embryos. Differential inhibition by a common inhibitor revealed that all three PCs are active in inner and outer cells, but in partially nonoverlapping compartments. E-cadherin processing by multiple PCs emerges as a novel mechanism to modulate cell-cell adhesion and fate allocation.

  2. PC7 and the related proteases Furin and Pace4 regulate E-cadherin function during blastocyst formation

    PubMed Central

    Mesnard, Daniel

    2015-01-01

    The first cell differentiation in mammalian embryos segregates polarized trophectoderm cells from an apolar inner cell mass (ICM). This lineage decision is specified in compacted morulae by cell polarization and adhesion acting on the Yes-associated protein in the Hippo signaling pathway, but the regulatory mechanisms are unclear. We show that morula compaction and ICM formation depend on PC7 and the related proprotein convertases (PCs) Furin and Pace4 and that these proteases jointly regulate cell–cell adhesion mediated by E-cadherin processing. We also mapped the spatiotemporal activity profiles of these proteases by live imaging of a transgenic reporter substrate in wild-type and PC mutant embryos. Differential inhibition by a common inhibitor revealed that all three PCs are active in inner and outer cells, but in partially nonoverlapping compartments. E-cadherin processing by multiple PCs emerges as a novel mechanism to modulate cell–cell adhesion and fate allocation. PMID:26416966

  3. PC viruses: How do they do that

    SciTech Connect

    Pichnarczyk, K.

    1992-07-01

    The topic of PC Viruses has been an issue for a number of years now. They've been reported in every major newspaper, tabloids, television and radio. People from all fields get viruses: government, private sector businesses, home computers, schools, computer software suppliers. A definition is proposed to introduce the virus phenomenon. Virus authors come from a variety of communities. Motives and ideologies of authors are discussed, and examples of viruses are offered. Also mentioned is the growing number of viruses developed, isolated, and never distributed to the public at large, but kept within the antivirus research community. Virus examples are offered as well. Viruses are distributed not only through bulletin boards and shareware, but also from areas previously assumed to be safe, including the threat of receiving a virus through a standard in-house function, such as an in-house hardware maintenance shop. Three categories of viruses are presented: File Infecter viruses, Boot Sector Infecters, and the new category of Directory Entry Infecter virus. Also discussed are crossover viruses, that is, viruses which utilize a variety of techniques to ensure survival. An explanation of what is occurring within every stage of various viruses is given. Replication strategies common to all three types is noted, mainly the two different replication strategies of memory resident infecters and active selection infecters. A detailed definition, description and application of a stealth virus is presented. Detection strategies are discussed as each topic in this section is completed; a high level schemata of the operation of various virus detection programs ispresented. Since most eradication today is done using virus detection/eradication software, this paper attempts to reveal the techniques used by these packages.Included in the paper is the topic of manual eradication.

  4. PC viruses: How do they do that?

    SciTech Connect

    Pichnarczyk, K.

    1992-07-01

    The topic of PC Viruses has been an issue for a number of years now. They`ve been reported in every major newspaper, tabloids, television and radio. People from all fields get viruses: government, private sector businesses, home computers, schools, computer software suppliers. A definition is proposed to introduce the virus phenomenon. Virus authors come from a variety of communities. Motives and ideologies of authors are discussed, and examples of viruses are offered. Also mentioned is the growing number of viruses developed, isolated, and never distributed to the public at large, but kept within the antivirus research community. Virus examples are offered as well. Viruses are distributed not only through bulletin boards and shareware, but also from areas previously assumed to be safe, including the threat of receiving a virus through a standard in-house function, such as an in-house hardware maintenance shop. Three categories of viruses are presented: File Infecter viruses, Boot Sector Infecters, and the new category of Directory Entry Infecter virus. Also discussed are crossover viruses, that is, viruses which utilize a variety of techniques to ensure survival. An explanation of what is occurring within every stage of various viruses is given. Replication strategies common to all three types is noted, mainly the two different replication strategies of memory resident infecters and active selection infecters. A detailed definition, description and application of a stealth virus is presented. Detection strategies are discussed as each topic in this section is completed; a high level schemata of the operation of various virus detection programs ispresented. Since most eradication today is done using virus detection/eradication software, this paper attempts to reveal the techniques used by these packages.Included in the paper is the topic of manual eradication.

  5. Run-09 pC polarimeter analysis

    SciTech Connect

    Alekseev, I.; Aschenauer, E.; Atoyan, G.; Bazilevsky, A.; Gill, R.; Huang, H.; Lee, S.; Li, X.; Makdisi, Y.; Morozov, B.; Nakagawa, I.; Svirida, D.; Zelenski, A.

    2010-08-01

    Analysis of PC polarimeter data at {radical}s = 200 and 500 GeV from Run9 is presented. Final polarization results, fill-by-fill, for blue and yellow beams, as to be used by RHIC experiments (in collisions) are released and collected in http://www4.rcf.bnl.gov/cnipol/pubdocs/Run09Offline/. Global relative systematic uncertainties {delta}P/P (to be considered as correlated from fill to fill) are 4.7% for 100 GeV beams, and 8.3% (12.1%) for blue (yellow) 250 GeV beams. For a product of two beam polarizations P{sub B} {center_dot} P{sub Y} (used in double spin asymmetry measurements) the relative uncertainty {delta}(P{sub B} {center_dot} P{sub Y})/(P{sub B} {center_dot} P{sub Y}) 8.8% for 100 GeV beams and 18.5% for 250 GeV beams. For the average between two beam polarization (P{sub B} + P{sub Y})/2 (used in single spin asymmetry measurements, when data from two polarized beams are combined) the relative uncertainty is 4.4% for 100 GeV beams and 9.2% for 250 GeV beams. Larger uncertainties for 250 GeV beams relate to significant rate related systematic effects experienced in the first part of Run9 (due to thicker targets used and smaller trans. beam size at higher beam energy).

  6. HOTSPOT Health Physics codes for the PC

    SciTech Connect

    Homann, S.G.

    1994-03-01

    The HOTSPOT Health Physics codes were created to provide Health Physics personnel with a fast, field-portable calculation tool for evaluating accidents involving radioactive materials. HOTSPOT codes are a first-order approximation of the radiation effects associated with the atmospheric release of radioactive materials. HOTSPOT programs are reasonably accurate for a timely initial assessment. More importantly, HOTSPOT codes produce a consistent output for the same input assumptions and minimize the probability of errors associated with reading a graph incorrectly or scaling a universal nomogram during an emergency. The HOTSPOT codes are designed for short-term (less than 24 hours) release durations. Users requiring radiological release consequences for release scenarios over a longer time period, e.g., annual windrose data, are directed to such long-term models as CAPP88-PC (Parks, 1992). Users requiring more sophisticated modeling capabilities, e.g., complex terrain; multi-location real-time wind field data; etc., are directed to such capabilities as the Department of Energy`s ARAC computer codes (Sullivan, 1993). Four general programs -- Plume, Explosion, Fire, and Resuspension -- calculate a downwind assessment following the release of radioactive material resulting from a continuous or puff release, explosive release, fuel fire, or an area contamination event. Other programs deal with the release of plutonium, uranium, and tritium to expedite an initial assessment of accidents involving nuclear weapons. Additional programs estimate the dose commitment from the inhalation of any one of the radionuclides listed in the database of radionuclides; calibrate a radiation survey instrument for ground-survey measurements; and screen plutonium uptake in the lung (see FIDLER Calibration and LUNG Screening sections).

  7. Mutant power: using mutant allele collections for yeast functional genomics.

    PubMed

    Norman, Kaitlyn L; Kumar, Anuj

    2016-03-01

    The budding yeast has long served as a model eukaryote for the functional genomic analysis of highly conserved signaling pathways, cellular processes and mechanisms underlying human disease. The collection of reagents available for genomics in yeast is extensive, encompassing a growing diversity of mutant collections beyond gene deletion sets in the standard wild-type S288C genetic background. We review here three main types of mutant allele collections: transposon mutagen collections, essential gene collections and overexpression libraries. Each collection provides unique and identifiable alleles that can be utilized in genome-wide, high-throughput studies. These genomic reagents are particularly informative in identifying synthetic phenotypes and functions associated with essential genes, including those modeled most effectively in complex genetic backgrounds. Several examples of genomic studies in filamentous/pseudohyphal backgrounds are provided here to illustrate this point. Additionally, the limitations of each approach are examined. Collectively, these mutant allele collections in Saccharomyces cerevisiae and the related pathogenic yeast Candida albicans promise insights toward an advanced understanding of eukaryotic molecular and cellular biology.

  8. Analysis of Mycobacterium avium subsp. paratuberculosis mutant libraries reveals loci-dependent transcription biases and strategies to novel mutant discovery

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Mycobacterium avium subsp. paratuberculosis (MAP) is the etiologic agent of Johne’s disease in ruminants and it has been implicated as a cause of Crohn’s disease in humans. The generation of comprehensive random mutant banks by transposon mutagenesis is a fundamental wide genomic technology utilized...

  9. Zebrafish Genomic Instability Mutants and Cancer Susceptibility

    PubMed Central

    Moore, Jessica L.; Rush, Lindsay M.; Breneman, Carol; Mohideen, Manzoor-Ali P. K.; Cheng, Keith C.

    2006-01-01

    Somatic loss of tumor suppressor gene function comprising the second hit of Knudson's two-hit hypothesis is important in human cancer. A genetic screen was performed in zebrafish (Danio rerio) to find mutations that cause genomic instability (gin), as scored by Streisinger's mosaic-eye assay that models this second hit. The assay, based on a visible test for loss of wild-type gene function at a single locus, golden, is representative of genomewide events. Twelve ENU-induced genomic instability (gin) mutations were isolated. Most mutations showed weak dominance in heterozygotes and all showed a stronger phenotype in homozygotes. Trans-heterozygosity for 7 of these mutations showed greatly enhanced instability. A variety of spontaneous tumors were found in heterozygous adults from all gin lines, consistent with the expectation that genomic instability (mutator) mutations can accelerate carcinogenesis. The incidence of spontaneous cancer at 30–34 months was increased 9.6-fold in heterozygotes for the mutant with the strongest phenotype, gin-10. Tumors were seen in skin, colon, kidney, liver, pancreas, ovary, testis, and neuronal tissues, with multiple tumors in some fish. The study of these mutants will add to our understanding of the mechanisms of somatic loss of gene function and how those mechanisms contribute to cancer susceptibility. PMID:16888336

  10. Identical substitutions in magnesium chelatase paralogs result in chlorophyll deficient soybean mutants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The soybean (Glycine max (L.) Merr.) chlorophyll deficient line MinnGold is a spontaneous mutant characterized by yellow foliage. Map-based cloning and transgenic complementation revealed that the mutant phenotype is caused by a non-synonymous nucleotide substitution in the third exon of a Mg-chelat...

  11. ISTUM PC: industrial sector technology use model for the IBM-PC

    SciTech Connect

    Roop, J.M.; Kaplan, D.T.

    1984-09-01

    A project to improve and enhance the Industrial Sector Technology Use Model (ISTUM) was originated in the summer of 1983. The project had dix identifiable objectives: update the data base; improve run-time efficiency; revise the reference base case; conduct case studies; provide technical and promotional seminars; and organize a service bureau. This interim report describes which of these objectives have been met and which tasks remain to be completed. The most dramatic achievement has been in the area of run-time efficiency. From a model that required a large proportion of the total resources of a mainframe computer and a great deal of effort to operate, the current version of the model (ISTUM-PC) runs on an IBM Personal Computer. The reorganization required for the model to run on a PC has additional advantages: the modular programs are somewhat easier to understand and the data base is more accessible and easier to use. A simple description of the logic of the model is given in this report. To generate the necessary funds for completion of the model, a multiclient project is proposed. This project will extend the industry coverage to all the industrial sectors, including the construction of process flow models for chemicals and petroleum refining. The project will also calibrate this model to historical data and construct a base case and alternative scenarios. The model will be delivered to clients and training provided. 2 references, 4 figures, 3 tables.

  12. Electrical Phenotypes of Calcium Transport Mutant Strains of a Filamentous Fungus, Neurospora crassa

    PubMed Central

    Hamam, Ahmed

    2012-01-01

    We characterized the electrical phenotypes of mutants with mutations in genes encoding calcium transporters—a mechanosensitive channel homolog (MscS), a Ca2+/H+ exchange protein (cax), and Ca2+-ATPases (nca-1, nca-2, nca-3)—as well as those of double mutants (the nca-2 cax, nca-2 nca-3, and nca-3 cax mutants). The electrical characterization used dual impalements to obtain cable-corrected current-voltage measurements. Only two types of mutants (the MscS mutant; the nca-2 mutant and nca-2-containing double mutants) exhibited lower resting potentials. For the nca-2 mutant, on the basis of unchanged conductance and cyanide-induced depolarization of the potential, the cause is attenuated H+-ATPase activity. The growth of the nca-2 mutant-containing strains was inhibited by elevated extracellular Ca2+ levels, indicative of lesions in Ca2+ homeostasis. However, the net Ca2+ effluxes of the nca-2 mutant, measured noninvasively with a self-referencing Ca2+-selective microelectrode, were similar to those of the wild type. All of the mutants exhibited osmosensitivity similar to that of the wild type (the turgor of the nca-2 mutant was also similar to that of the wild type), suggesting that Ca2+ signaling does not play a role in osmoregulation. The hyphal tip morphology and tip-localized mitochondria of the nca-2 mutant were similar to those of the wild type, even when the external [Ca2+] was elevated. Thus, although Ca2+ homeostasis is perturbed in the nca-2 mutant (B. J. Bowman et al., Eukaryot. Cell 10:654–661, 2011), the phenotype does not extend to tip growth or to osmoregulation but is revealed by lower H+-ATPase activity. PMID:22408225

  13. Ball-milled CuPc/TiO{sub 2} hybrid nanocomposite and its photocatalytic degradation of aqueous Rhodamine B

    SciTech Connect

    Mekprasart, W.; Vittayakorn, N.; Pecharapa, W.

    2012-11-15

    B dye solution using as-synthesized composites was investigated under ultraviolet irradiation. The optimized condition for RhB photodegradation was obtained for 1 wt.% CuPc/TiO{sub 2} nanocomposite with efficiency of 87.5% and reaction rate of 0.15 min{sup −1}. The significant improvement in photocatalytic activity of the hybrid composite may be associated to the enhancement of optical absorption and the inhibition of electron–hole recombination caused by the presence of CuPc in TiO{sub 2} matrix.

  14. Docosahexaenoic acid phospholipid differentially modulates the conformation of G90V and N55K rhodopsin mutants associated with retinitis pigmentosa.

    PubMed

    Dong, Xiaoyun; Herrera-Hernández, María Guadalupe; Ramon, Eva; Garriga, Pere

    2017-05-01

    Rhodopsin is the visual photoreceptor of the retinal rod cells that mediates dim light vision and a prototypical member of the G protein-coupled receptor superfamily. The structural stability and functional performance of rhodopsin are modulated by membrane lipids. Docosahexaenoic acid has been shown to interact with native rhodopsin but no direct evidence has been established on the effect of such lipid on the stability and regeneration of rhodopsin mutants associated with retinal diseases. The stability and regeneration of two thermosensitive mutants G90V and N55K, associated with the retinal degenerative disease retinitis pigmentosa, have been analyzed in docosohexaenoic phospholipid (1,2-didocosa-hexaenoyl-sn-glycero-3-phosphocholine; DDHA-PC) liposomes. G90V mutant reconstituted in DDHA-PC liposomes significantly increased its thermal stability, but N55K mutant showed similar thermal sensitivity both in dodecyl maltoside detergent solution and in DDHA-PC liposomes. The retinal release process, measured by fluorescence spectroscopy, became faster in the lipid system for the two mutants. The opsin conformation was stabilized for the G90V mutant allowing improved retinal uptake whereas no chromophore binding could be detected for N55K opsin after photoactivation. The results emphasize the distinct role of DHA on different phenotypic rhodopsin mutations associated with classical (G90V) and sector (N55K) retinitis pigmentosa.

  15. Possible generation mechanisms for Pc1 pearl structures in the ionosphere based on 6 years of ground observations in Canada, Russia, and Japan

    NASA Astrophysics Data System (ADS)

    Jun, Chae-Woo; Shiokawa, Kazuo; Connors, Martin; Schofield, Ian; Poddelsky, Igor; Shevtsov, Boris

    2016-05-01

    We investigate pearl structures (amplitude modulations) of Pc1 pulsations simultaneously observed at Athabasca (ATH, 54.7°N, 246.7°E, L = 4.3) in Canada, Magadan (MGD, 60.1°N, 150.7°E, L = 2.6) in Russia, and Moshiri (MOS, 44.4°N, 142.3°E, L = 1.5) in Japan. From 6 years of ground observations, from 2008 to 2013, we selected 84 Pc1 events observed simultaneously at the longitudinally separated stations (ATH and MGD) and 370 events observed at the latitudinally separated stations (MGD and MOS), all with high coherence (>0.7) of Pc1 waveforms. We calculated the cross-correlation coefficient (similarity: r) for the Pc1 pearl structures and found that more than half of the events in both pairs had low similarity (r < 0.7), indicating that most Pc1 waves exhibit different pearl structures at different stations. We found that high-similarity Pc1 pearl structures (r > 0.7) at the longitudinally separated stations are concentrated from 6 to 15 UT when both stations are in the nighttime. The similarity of Pc1 pearl structures tends to show a negative correlation with the standard deviation of the polarization angle in both pairs. The observed repetition period of Pc1 pearl structures has a clear positive correlation with the repetition period estimated from Pc1 bandwidth by assuming beating of different frequencies. From these results, we suggest that ionospheric beating effect could be a dominant process for the generation of Pc1 pearl structures. Beating processes in the ionosphere with a spatially distributed ionospheric source can cause the different shapes of Pc1 pearl structures at different observation points during ionospheric duct propagation.

  16. User`s guide and documentation manual for ``PC-Gel`` simulator

    SciTech Connect

    Chang, Ming-Ming; Gao, Hong W.

    1993-10-01

    PC-GEL is a three-dimensional, three-phase (oil, water, and gas) permeability modification simulator developed by incorporating an in-situ gelation model into a black oil simulator (BOAST) for personal computer application. The features included in the simulator are: transport of each chemical species of the polymer/crosslinker system in porous media, gelation reaction kinetics of the polymer with crosslinking agents, rheology of the polymer and gel, inaccessible pore volume to macromolecules, adsorption of chemical species on rock surfaces, retention of gel on the rock matrix, and permeability reduction caused by the adsorption of polymer and gel. The in-situ gelation model and simulator were validated against data reported in the literature. The simulator PC-GEL is useful for simulating and optimizing any combination of primary production, waterflooding, polymer flooding, and permeability modification treatments. A general background of permeability modification using crosslinked polymer gels is given in Section I and the governing equations, mechanisms, and numerical solutions of PC-GEL are given in Section II. Steps for preparing an input data file with reservoir and gel-chemical transport data, and recurrent data are described in Sections III and IV, respectively. Example data inputs are enclosed after explanations of each input line to help the user prepare data files. Major items of the output files are reviewed in Section V. Finally, three sample problems for running PC-GEL are described in Section VI, and input files and part of the output files of these problems are listed in the appendices. For the user`s reference a copy of the source code of PC-GEL computer program is attached in Appendix A.

  17. Latitude-independent Pc5 Geomagnetic Pulsations Associated With Field Line Resonance

    NASA Astrophysics Data System (ADS)

    Sung, S.; Kim, K.; Lee, D.; Cattell, C. A.; Andre, M.; Khotyaintsev, Y. V.

    2004-12-01

    The latitude-independent Pc5 pulsations with a spectral peak at ˜2.8 mHz were observed with IMAGE and SAMNET magnetometer array in the morning sector (0700-1000 local time) on April 29 (Day 119), 2001. The spectral amplitude had a local peak at ˜67° geomagnetic latitude, where a sudden phase change of ˜180° appeared. A vortical equivalent ionospheric current structure centered at latitude between 67° and 71° was observed during the Pc5 pulsations and the rotational sense of the current vortex was reversed for one cycle of the pulsation. During the interval of the enhancement of the Pc5 pulsations, the POLAR spacecraft in the morning side crossed near the magnetic shell (L ˜ 8) corresponding to the latitude where the spectral amplitude was maximum, and observed ˜2.8 mHz pulsations in the radial electric field and compressional magnetic field components. Since the toroidal mode Alfvén waves in the magnetosphere are characterized by an electric field perturbation in the radial direction, the simultaneous presence of the pulsations in both components indicates that a field line resonance (FLR) was driven by compressional Pc5 pulsations. Using solar wind data, we conformed that the compressional Pc5 pulsations at POLAR occurred during an interval of enhanced solar wind dynamic pressure. From the analysis of the ground magnetometer data and POLAR data, we suggest that latitude independent ground magnetic perturbations are caused by the vortical equivalent current generated by FLR-associated field-aligned currents.

  18. ADP ribosylation factor 1 is required for synaptic vesicle budding in PC12 cells.

    PubMed

    Faúndez, V; Horng, J T; Kelly, R B

    1997-08-11

    Carrier vesicle generation from donor membranes typically progresses through a GTP-dependent recruitment of coats to membranes. Here we explore the role of ADP ribosylation factor (ARF) 1, one of the GTP-binding proteins that recruit coats, in the production of neuroendocrine synaptic vesicles (SVs) from PC12 cell membranes. Brefeldin A (BFA) strongly and reversibly inhibited SV formation in vivo in three different PC12 cell lines expressing vesicle-associated membrane protein-T Antigen derivatives. Other membrane traffic events remained unaffected by the drug, and the BFA effects were not mimicked by drugs known to interfere with formation of other classes of vesicles. The involvement of ARF proteins in the budding of SVs was addressed in a cell-free reconstitution system (Desnos, C., L. Clift-O'Grady, and R.B. Kelly. 1995. J. Cell Biol. 130:1041-1049). A peptide spanning the effector domain of human ARF1 (2-17) and recombinant ARF1 mutated in its GTPase activity, both inhibited the formation of SVs of the correct size. During in vitro incubation in the presence of the mutant ARFs, the labeled precursor membranes acquired different densities, suggesting that the two ARF mutations block at different biosynthetic steps. Cell-free SV formation in the presence of a high molecular weight, ARF-depleted fraction from brain cytosol was significantly enhanced by the addition of recombinant myristoylated native ARF1. Thus, the generation of SVs from PC12 cell membranes requires ARF and uses its GTPase activity, probably to regulate coating phenomena.

  19. Changes in temperature preferences and energy homeostasis in dystroglycan mutants.

    PubMed

    Takeuchi, Ken-Ichi; Nakano, Yoshiro; Kato, Utako; Kaneda, Mizuho; Aizu, Masako; Awano, Wakae; Yonemura, Shigenobu; Kiyonaka, Shigeki; Mori, Yasuo; Yamamoto, Daisuke; Umeda, Masato

    2009-03-27

    Temperature affects the physiology, behavior, and evolution of organisms. We conducted mutagenesis and screens for mutants with altered temperature preference in Drosophila melanogaster and identified a cryophilic (cold-seeking) mutant, named atsugari (atu). Reduced expression of the Drosophila ortholog of dystroglycan (DmDG) induced tolerance to cold as well as preference for the low temperature. A sustained increase in mitochondrial oxidative metabolism caused by the reduced expression of DmDG accounted for the cryophilic phenotype of the atu mutant. Although most ectothermic animals do not use metabolically produced heat to regulate body temperature, our results indicate that their thermoregulatory behavior is closely linked to rates of mitochondrial oxidative metabolism and that a mutation in a single gene can induce a sustained change in energy homeostasis and the thermal responses.

  20. Defective Glycinergic Synaptic Transmission in Zebrafish Motility Mutants

    PubMed Central

    Hirata, Hiromi; Carta, Eloisa; Yamanaka, Iori; Harvey, Robert J.; Kuwada, John Y.

    2009-01-01

    Glycine is a major inhibitory neurotransmitter in the spinal cord and brainstem. Recently, in vivo analysis of glycinergic synaptic transmission has been pursued in zebrafish using molecular genetics. An ENU mutagenesis screen identified two behavioral mutants that are defective in glycinergic synaptic transmission. Zebrafish bandoneon (beo) mutants have a defect in glrbb, one of the duplicated glycine receptor (GlyR) β subunit genes. These mutants exhibit a loss of glycinergic synaptic transmission due to a lack of synaptic aggregation of GlyRs. Due to the consequent loss of reciprocal inhibition of motor circuits between the two sides of the spinal cord, motor neurons activate simultaneously on both sides resulting in bilateral contraction of axial muscles of beo mutants, eliciting the so-called ‘accordion’ phenotype. Similar defects in GlyR subunit genes have been observed in several mammals and are the basis for human hyperekplexia/startle disease. By contrast, zebrafish shocked (sho) mutants have a defect in slc6a9, encoding GlyT1, a glycine transporter that is expressed by astroglial cells surrounding the glycinergic synapse in the hindbrain and spinal cord. GlyT1 mediates rapid uptake of glycine from the synaptic cleft, terminating synaptic transmission. In zebrafish sho mutants, there appears to be elevated extracellular glycine resulting in persistent inhibition of postsynaptic neurons and subsequent reduced motility, causing the ‘twitch-once’ phenotype. We review current knowledge regarding zebrafish ‘accordion’ and ‘twitch-once’ mutants, including beo and sho, and report the identification of a new α2 subunit that revises the phylogeny of zebrafish GlyRs. PMID:20161699

  1. In vivo fitness and virulence of a drug-resistant herpes simplex virus 1 mutant.

    PubMed

    Pesola, Jean M; Coen, Donald M

    2007-05-01

    Two important issues regarding a virus mutant that is resistant to an antiviral drug are its ability to replicate in animal hosts (in vivo fitness) relative to other genetic variants, including wild type, and its ability to cause disease. These issues have been investigated for a herpes simplex virus 1 mutant that is resistant to thiourea compounds, which inhibit encapsidation of viral DNA. Following corneal inoculation of mice, the mutant virus replicated very similarly to its wild-type parent in the eye, trigeminal ganglion and brain. The mutant virus was as lethal to mice as its wild-type parent following this route of inoculation. Indeed, it exhibited increased virulence. Thus, unlike most drug-resistant virus mutants, this mutant retained in vivo fitness and virulence.

  2. A double-mutant collection targeting MAP kinase related genes in Arabidopsis for studying genetic interactions.

    PubMed

    Su, Shih-Heng; Krysan, Patrick J

    2016-12-01

    Mitogen-activated protein kinase cascades are conserved in all eukaryotes. In Arabidopsis thaliana there are approximately 80 genes encoding MAP kinase kinase kinases (MAP3K), 10 genes encoding MAP kinase kinases (MAP2K), and 20 genes encoding MAP kinases (MAPK). Reverse genetic analysis has failed to reveal abnormal phenotypes for a majority of these genes. One strategy for uncovering gene function when single-mutant lines do not produce an informative phenotype is to perform a systematic genetic interaction screen whereby double-mutants are created from a large library of single-mutant lines. Here we describe a new collection of 275 double-mutant lines derived from a library of single-mutants targeting genes related to MAP kinase signaling. To facilitate this study, we developed a high-throughput double-mutant generating pipeline using a system for growing Arabidopsis seedlings in 96-well plates. A quantitative root growth assay was used to screen for evidence of genetic interactions in this double-mutant collection. Our screen revealed four genetic interactions, all of which caused synthetic enhancement of the root growth defects observed in a MAP kinase 4 (MPK4) single-mutant line. Seeds for this double-mutant collection are publicly available through the Arabidopsis Biological Resource Center. Scientists interested in diverse biological processes can now screen this double-mutant collection under a wide range of growth conditions in order to search for additional genetic interactions that may provide new insights into MAP kinase signaling.

  3. Problem-Solving Test: Tryptophan Operon Mutants

    ERIC Educational Resources Information Center

    Szeberenyi, Jozsef

    2010-01-01

    This paper presents a problem-solving test that deals with the regulation of the "trp" operon of "Escherichia coli." Two mutants of this operon are described: in mutant A, the operator region of the operon carries a point mutation so that it is unable to carry out its function; mutant B expresses a "trp" repressor protein unable to bind…

  4. Nanoformulated cell-penetrating survivin mutant and its dual actions

    PubMed Central

    Sriramoju, Bhasker; Kanwar, Rupinder K; Kanwar, Jagat R

    2014-01-01

    In this study, we investigated the differential actions of a dominant-negative survivin mutant (SurR9-C84A) against cancerous SK-N-SH neuroblastoma cell lines and differentiated SK-N-SH neurons. In both the cases, the mutant protein displayed dual actions, where its effects were cytotoxic toward cancerous cells and proliferative toward the differentiated neurons. This can be explained by the fact that tumorous (undifferentiated SK-N-SH) cells have a high endogenous survivin pool and upon treatment with mutant SuR9-C84A causes forceful survivin expression. These events significantly lowered the microtubule dynamics and stability, eventually leading to apoptosis. In the case of differentiated SK-N-SH neurons that express negligible levels of wild-type survivin, the mutant indistinguishably behaved in a wild-type fashion. It also favored cell-cycle progression, forming the chromosome-passenger complex, and stabilized the microtubule-organizing center. Therefore, mutant SurR9-C84A represents a novel therapeutic with its dual actions (cytotoxic toward tumor cells and protective and proliferative toward neuronal cells), and hence finds potential applications against a variety of neurological disorders. In this study, we also developed a novel poly(lactic-co-glycolic acid) nanoparticulate formulation to surmount the hurdles associated with the delivery of SurR9-C84A, thus enhancing its effective therapeutic outcome. PMID:25045261

  5. PC-SPES (PDQ®)—Health Professional Version

    Cancer.gov

    Expert-reviewed information summary about the use of PC-SPES as a treatment for prostate cancer. Note: The information in this summary is no longer being updated and is provided for reference purposes only.

  6. OCEAN-PC and a distributed network for ocean data

    NASA Technical Reports Server (NTRS)

    Mclain, Douglas R.

    1992-01-01

    The Intergovernmental Oceanographic Commission (IOC) wishes to develop an integrated software package for oceanographic data entry and access in developing countries. The software, called 'OCEAN-PC', would run on low cost PC microcomputers and would encourage and standardize: (1) entry of local ocean observations; (2) quality control of the local data; (3) merging local data with historical data; (4) improved display and analysis of the merged data; and (5) international data exchange. OCEAN-PC will link existing MS-DOS oceanographic programs and data sets with table-driven format conversions. Since many ocean data sets are now being distributed on optical discs (Compact Discs - Read Only Memory, CD-ROM, Mass et al. 1987), OCEAN-PC will emphasize access to CD-ROMs.

  7. PC-SPES (PDQ®)—Patient Version

    Cancer.gov

    Expert-reviewed information summary about the use of PC-SPES as a treatment for prostate cancer. Note: The information in this summary is no longer being updated and is provided for reference purposes only.

  8. Software Reviews. PC Software for Artificial Intelligence Applications.

    ERIC Educational Resources Information Center

    Epp, Helmut; And Others

    1988-01-01

    Contrasts artificial intelligence and conventional programming languages. Reviews Personal Consultant Plus, Smalltalk/V, and Nexpert Object, which are PC-based products inspired by problem-solving paradigms. Provides information on background and operation of each. (RT)

  9. Single-event upset in advanced PowerPC microprocessors

    NASA Technical Reports Server (NTRS)

    Irom, F.; Swift, G. M.; Farmanesh, F.; Millward, D. G.

    2002-01-01

    Proton and heavy-ion single-event upset susceptibility has been measured for the MotorolaPowerPC7400. The results show that this advanced device has low upset susceptibility, despite the scaling and design advances.

  10. The toc132toc120 heterozygote mutant of Arabidopsis thaliana accumulates reduced levels of hexadecatrienoic acid.

    PubMed

    Afitlhile, Meshack; Duffield-Duncan, Kayla; Fry, Morgan; Workman, Samantha; Hum-Musser, Sue; Hildebrand, David

    2015-11-01

    A null and heterozygous mutant for the Arabidopsis thaliana TOC132 and TOC120 genes accumulates increased levels of 16:0 and decreased 16:3, suggesting altered homeostasis in fatty acid synthesis. The FAD5 gene encodes a plastid desaturase that catalyzes the first step in the synthesis of 16:3 in monogalactosyldiacylglycerol (MGDG). In non-acclimated toc132toc120+/- mutant plants, the FAD5 gene was repressed and this correlated with decreased levels of 16:3. In cold-acclimated mutant however, the FAD5 gene was upregulated and there was a small increase in 16:3 levels relative to the non-acclimated mutant plants. The MGD1 gene was expressed at control levels and the mutant accumulated levels of MGDG that were similar to the wild type. In the mutant however, MGDG had decreased 16:3 levels, suggesting that the activity of FAD5 desaturase was compromised. In the mutant, the FAD2 and FAD3 genes were downregulated but levels of 18:3-PC were increased, suggesting posttranscriptional regulation for the ER-localized fatty acid desaturases. The Toc120 or Toc159 receptor is likely to compensate for a defective Toc132 receptor. In the cold-acclimated mutant, the TOC159 gene was repressed ca. 300-fold, whereas the TOC120 gene was repressed 7-fold relative to the non-acclimated wild type. Thus, the TOC159 gene is more sensitive to cold-stress and might not compensate for defect in the TOC132 gene under these conditions. Overall, these data show that a mutation in the TOC132 gene results in decreased 16:3 levels, indicating the need for an intact Toc132/Toc120 receptor, presumably to facilitate the import of the FAD5 preprotein into chloroplasts.

  11. Neuroprotective effects of constituents of Eragrostis ferruginea against Aβ-induced toxicity in PC12 cells.

    PubMed

    Na, Chae Sun; Hong, Seong Su; Choi, Yun-Hyeok; Lee, Yong Ho; Hong, Sun Hee; Lim, Ji-Youn; Kang, Byeong Hoa; Park, So-Young; Lee, Dongho

    2010-07-01

    A new flavonoid, 7-demethylageconyflavone A (1), and five known compounds, tricin (2), ageconyflavone A (3), corylin (4), nectandrin B (5), and 4-ketopinoresinol (6) were isolated from the aerial parts of Eragrostis ferruginea. Their structures were determined using spectroscopic techniques, including 1D- and 2D-NMR. All compounds were tested for the neuroprotective effects against amyloid beta peptide (Abeta) using PC12 cells, a major cause of the pathology of Alzheimer's disease. Tricin (2) was found to have a neuroprotective effect with an ED(50) value of 20.3 microM against Abeta-induced toxicity in PC12 cells. Ageconyflavone A (3), nectandrin B (5) and 4-ketopinoresinol (6) demonstrated moderate neuroprotective effects with ED(50) values of 58.7, 44.1, and 54.8 microM, respectively.

  12. Mutant Huntingtin Disrupts the Nuclear Pore Complex.

    PubMed

    Grima, Jonathan C; Daigle, J Gavin; Arbez, Nicolas; Cunningham, Kathleen C; Zhang, Ke; Ochaba, Joseph; Geater, Charlene; Morozko, Eva; Stocksdale, Jennifer; Glatzer, Jenna C; Pham, Jacqueline T; Ahmed, Ishrat; Peng, Qi; Wadhwa, Harsh; Pletnikova, Olga; Troncoso, Juan C; Duan, Wenzhen; Snyder, Solomon H; Ranum, Laura P W; Thompson, Leslie M; Lloyd, Thomas E; Ross, Christopher A; Rothstein, Jeffrey D

    2017-04-05

    Huntington's disease (HD) is caused by an expanded CAG repeat in the Huntingtin (HTT) gene. The mechanism(s) by which mutant HTT (mHTT) causes disease is unclear. Nucleocytoplasmic transport, the trafficking of macromolecules between the nucleus and cytoplasm, is tightly regulated by nuclear pore complexes (NPCs) made up of nucleoporins (NUPs). Previous studies offered clues that mHTT may disrupt nucleocytoplasmic transport and a mutation of an NUP can cause HD-like pathology. Therefore, we evaluated the NPC and nucleocytoplasmic transport in multiple models of HD, including mouse and fly models, neurons transfected with mHTT, HD iPSC-derived neurons, and human HD brain regions. These studies revealed severe mislocalization and aggregation of NUPs and defective nucleocytoplasmic transport. HD repeat-associated non-ATG (RAN) translation proteins also disrupted nucleocytoplasmic transport. Additionally, overexpression of NUPs and treatment with drugs that prevent aberrant NUP biology also mitigated this transport defect and neurotoxicity, providing future novel therapy targets.

  13. A PC-Windows-Based program for geostatistical modeling application

    SciTech Connect

    Wu, G.G.; Yang, A.P.

    1994-12-31

    This paper describes a technically advanced, user-friendly, PC-Windows{sup TM} based reservoir simulation tool (SIMTOOLS) that allows construction of realistic reservoir models using a geostatistical approach. This PC-Windows based product has three application tools: Digitizing, mapping, and geostatistics. It has been designed primarily to enable reservoir engineers to apply the geostatistical gridding technique in mapping and reservoir simulation practices.

  14. Wf/pc Cycle 3 Calibration: Rapid Internal Monitor

    NASA Astrophysics Data System (ADS)

    MacKenty, John

    1992-06-01

    This test is to take repeated internal flats to test for contamination buildup on the optical surfaces or the reappearance of QEH. Part 1: INTFLATS in F555W are obtained every 4 days in both WFC and PC to check for measles or daisies and to monitor scattered light. Part 2: Sequential INTFLATS in F439W with PC are obtained every 7 days to check for QEH.

  15. Wf/pc Cycle 2 Calibration: Rapid Internal Monitor

    NASA Astrophysics Data System (ADS)

    MacKenty, John

    1991-07-01

    This test is to take repeated internal flats to test for contamination buildup on the optical surfaces or the reappearance of QEH. Part 1: INTFLATS in F555W are obtained every 4 days in both WFC and PC to check for measles or daisies and to monitor scattered light. Part 2: Sequential INTFLATS in F439W with PC are obtained every 7 days to check for QEH.

  16. Wf/pc Cycle 1 Calibration: Rapid Internal Monitor

    NASA Astrophysics Data System (ADS)

    MacKenty, John

    1990-12-01

    This test is to take repeated internal flats to test for contamination buildup on the optical surfaces or the reappearance of QEH. Part 1: INTFLATS in F555W are obtained every 4 days in both WFC and PC to check for measles or daisies and to monitor scattered light. Part 2: Sequential INTFLATS in F439W with PC are obtained every 7 days to check for QEH.

  17. Rotenone induces KATP channel opening in PC12 cells in association with the expression of tyrosine hydroxylase.

    PubMed

    Bai, Qunhua; He, Junlin; Qiu, Jingfu; Wang, Yang; Wang, Shibo; Xiu, Yun; Yu, Chao

    2012-10-01

    The activation of ATP-sensitive potassium (KATP) channels in PC12 cells play a pivotal role in protection against the neurotoxic effect of rotenone. However, it remains unclear why rotenone seems to preferentially affect activation of KATP channels and if this could affect its physiological activity. In this study, we sought to determine how the different energy states caused by various doses of rotenone affect the KATP opening state and whether the KATP opening state influences the expression of tyrosine hydroxylase (TH) which is related with DA synthesis. With patch clamp technology, results showed that treatment of PC12 cells with rotenone (0.2-1 µg/ml) for 15 min can cause KATP channel opening with significantly increased intracellular ROS production. Treatment with rotenone (2-16 ng/ml) for 24 h also caused the channels to open with gently increased ROS. In order to study if the rather long-term action on KATP channel opening states could affect the specified function of PC12 cells, the KATP channel opener pinacidil and the inhibitor glibenclamide were used to treat cells for 24 h, and the expression of TH was detected. Our results showed that treatment of PC12 cells with glibenclamide for 24 h can notably promote TH expression and can also enhance the expression of TH which were reduced by rotenone. These data indicate that the energy states in PC12 induced by various doses of rotenone could significantly influence the opening states of KATP channels. However long-term energy stress may raise the opening rate and opening sensitivity of this channel. In addition, our results demonstrate for the first time that activation of plasma membrane KATP channels induced by rotenone inhibits TH expression which influences DA synthesis in PC12 cells.

  18. A dual function for Munc-18 in exocytosis of PC12 cells.

    PubMed

    Schütz, Dagmar; Zilly, Felipe; Lang, Thorsten; Jahn, Reinhard; Bruns, Dieter

    2005-05-01

    Munc-18 interacts with the SNARE protein syntaxin and is supposed to influence transmitter release by controlling the formation of exocytosis-relevant SNARE complexes. Here, we used combined biochemical and physiological analyses to study the role of the Munc-18/syntaxin interaction in large dense core vesicle (LDCV) exocytosis of neuroendocrine PC12 cells. We compared two Munc-18 mutants carrying mutations in the syntaxin-binding region and show that Munc-18's membrane association depends on direct binding to syntaxin. The data suggest that perturbation of syntaxin binding inhibits neurotransmitter release upstream of the individual fusion event implying an essential role of the Munc-18/syntaxin complex leading to exocytosis. Furthermore, we show that a Munc-18 mutant lacking any syntaxin binding has a stimulatory effect on secretion, and provide evidence that the Munc-18/Mint1 interaction may constitute a second pathway for Munc-18 to regulate exocytosis. We propose that Munc-18 represents a dynamic link between syntaxin-related and Mint1-related mechanisms, both involved in the control of LDCV exocytosis in neuroendocrine cells.

  19. Unraveling the neuroprotective mechanisms of PrPC in excitotoxicity

    PubMed Central

    Llorens, Franc; del Río, José Antonio

    2012-01-01

    Knowledge of the natural roles of cellular prion protein (PrPC) is essential to an understanding of the molecular basis of prion pathologies. This GPI-anchored protein has been described in synaptic contacts, and loss of its synaptic function in complex systems may contribute to the synaptic loss and neuronal degeneration observed in prionopathy. In addition, Prnp knockout mice show enhanced susceptibility to several excitotoxic insults, GABAA receptor-mediated fast inhibition was weakened, LTP was modified and cellular stress increased. Although little is known about how PrPC exerts its function at the synapse or the downstream events leading to PrPC-mediated neuroprotection against excitotoxic insults, PrPC has recently been reported to interact with two glutamate receptor subunits (NR2D and GluR6/7). In both cases the presence of PrPC blocks the neurotoxicity induced by NMDA and Kainate respectively. Furthermore, signals for seizure and neuronal cell death in response to Kainate in Prnp knockout mouse are associated with JNK3 activity, through enhancing the interaction of GluR6 with PSD-95. In combination with previous data, these results shed light on the molecular mechanisms behind the role of PrPC in excitotoxicity. Future experimental approaches are suggested and discussed. PMID:22437735

  20. Misfolded opsin mutants display elevated β -sheet structure

    SciTech Connect

    Miller, Lisa M.; Gragg, Megan; Kim, Tae Gyun; Park, Paul S. -H.

    2015-09-07

    Mutations in rhodopsin can cause misfolding and aggregation of the receptor, which leads to retinitis pigmentosa, a progressive retinal degenerative disease. The structure adopted by misfolded opsin mutants and the associated cell toxicity is poorly understood. Förster resonance energy transfer (FRET) and Fourier transform infrared (FTIR) microspectroscopy were utilized to probe within cells the structures formed by G188R and P23H opsins, which are misfolding mutants that cause autosomal dominant retinitis pigmentosa. Also, both mutants formed aggregates in the endoplasmic reticulum and exhibited altered secondary structure with elevated β-sheet and reduced α-helical content. The newly formed β-sheet structure may facilitate the aggregation of misfolded opsin mutants. In conclusion, the effects observed for the mutants were unrelated to retention of opsin molecules in the endoplasmic reticulum itself.

  1. Production of phytotoxin solanapyrones and generation of solanapyrone-deficient mutants in Ascochyta rabiei

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Ascochyta rabiei, the fungus that causes Ascochyta blight of chickpea, produces phytotoxin solanapyrones. Generating mutants deficient in the toxin biosynthesis would provide information on the role of the toxins during infection processes. Partial genomic sequences of the solanapyrone biosynthesis ...

  2. Epitaxial growth and electronic properties of well ordered phthalocyanine heterojunctions MnPc/F{sub 16}CoPc

    SciTech Connect

    Lindner, Susi; Mahns, Benjamin; Treske, Uwe; Knupfer, Martin; Vilkov, Oleg; Haidu, Francisc; Fronk, Michael; Zahn, Dietrich R. T.

    2014-09-07

    We have prepared phthalocyanine heterojunctions out of MnPc and F{sub 16}CoPc, which were studied by means of X-ray absorption spectroscopy. This heterojunction is characterized by a charge transfer at the interface, resulting in charged MnPc{sup δ} {sup +} and F{sub 16}CoPc{sup δ} {sup −} species. Our data reveal that the molecules are well ordered and oriented parallel to the substrate surface. Furthermore, we demonstrate the filling of the Co 3d{sub z{sup 2}} orbital due to the charge transfer, which supports the explanation of the density functional theory, that the charge transfer is local and affects the metal centers only.

  3. PC Tutor. Bericht uber ein PC-gestutzes Tutorensystem = PC Tutor. Report on a Tutoring System with Personal Computer. ZIFF Papiere 75.

    ERIC Educational Resources Information Center

    Fritsch, Helmut

    A project was conducted to increase as well as to professionalize communication between tutors and learners in a West German university's distance education program by the use of personal computers. Two tutors worked on the systematic development of a PC-based correcting system. The goal, apart from developing general language skills in English,…

  4. Detection of Cell Wall Chemical Variation in Zea Mays Mutants Using Near-Infrared Spectroscopy

    SciTech Connect

    Buyck, N.; Thomas, S.

    2001-01-01

    Corn stover is regarded as the prime candidate feedstock material for commercial biomass conversion in the United States. Variations in chemical composition of Zea mays cell walls can affect biomass conversion process yields and economics. Mutant lines were constructed by activating a Mu transposon system. The cell wall chemical composition of 48 mutant families was characterized using near-infrared (NIR) spectroscopy. NIR data were analyzed using a multivariate statistical analysis technique called Principal Component Analysis (PCA). PCA of the NIR data from 349 maize leaf samples reveals 57 individuals as outliers on one or more of six Principal Components (PCs) at the 95% confidence interval. Of these, 19 individuals from 16 families are outliers on either PC3 (9% of the variation) or PC6 (1% of the variation), the two PCs that contain information about cell wall polymers. Those individuals for which altered cell wall chemistry is confirmed with wet chemical analysis will then be subjected to fermentation analysis to determine whether or not biomass conversion process kinetics, yields and/or economics are significantly affected. Those mutants that provide indications for a decrease in process cost will be pursued further to identify the gene(s) responsible for the observed changes in cell wall composition and associated changes in process economics. These genes will eventually be incorporated into maize breeding programs directed at the development of a truly dual use crop.

  5. Multinucleation and Polykaryon Formation is Promoted by the EhPC4 Transcription Factor in Entamoeba histolytica

    PubMed Central

    Cruz, Olga Hernández de la; Marchat, Laurence A.; Guillén, Nancy; Weber, Christian; Rosas, Itzel López; Díaz-Chávez, José; Herrera, Luis; Rojo-Domínguez, Arturo; Orozco, Esther; López-Camarillo, César

    2016-01-01

    Entamoeba histolytica is the intestinal parasite responsible for human amoebiasis that is a leading cause of death in developing countries. In this protozoan, heterogeneity in DNA content, polyploidy and genome plasticity have been associated to alterations in mechanisms controlling DNA replication and cell division. Studying the function of the transcription factor EhPC4, we unexpectedly found that it is functionally related to DNA replication, and multinucleation. Site-directed mutagenesis on the FRFPKG motif revealed that the K127 residue is required for efficient EhPC4 DNA-binding activity. Remarkably, overexpression of EhPC4 significantly increased cell proliferation, DNA replication and DNA content of trophozoites. A dramatically increase in cell size resulting in the formation of giant multinucleated trophozoites (polykaryon) was also found. Multinucleation event was associated to cytokinesis failure leading to abortion of ongoing cell division. Consistently, genome-wide profiling of EhPC4 overexpressing trophozoites revealed the up-regulation of genes involved in carbohydrates and nucleic acids metabolism, chromosome segregation and cytokinesis. Forced overexpression of one of these genes, EhNUDC (nuclear movement protein), led to alterations in cytokinesis and partially recapitulated the multinucleation phenotype. These data indicate for the first time that EhPC4 is associated with events related to polyploidy and genome stability in E. histolytica. PMID:26792358

  6. Multinucleation and Polykaryon Formation is Promoted by the EhPC4 Transcription Factor in Entamoeba histolytica.

    PubMed

    Hernández de la Cruz, Olga; Marchat, Laurence A; Guillén, Nancy; Weber, Christian; López Rosas, Itzel; Díaz-Chávez, José; Herrera, Luis; Rojo-Domínguez, Arturo; Orozco, Esther; López-Camarillo, César

    2016-01-21

    Entamoeba histolytica is the intestinal parasite responsible for human amoebiasis that is a leading cause of death in developing countries. In this protozoan, heterogeneity in DNA content, polyploidy and genome plasticity have been associated to alterations in mechanisms controlling DNA replication and cell division. Studying the function of the transcription factor EhPC4, we unexpectedly found that it is functionally related to DNA replication, and multinucleation. Site-directed mutagenesis on the FRFPKG motif revealed that the K127 residue is required for efficient EhPC4 DNA-binding activity. Remarkably, overexpression of EhPC4 significantly increased cell proliferation, DNA replication and DNA content of trophozoites. A dramatically increase in cell size resulting in the formation of giant multinucleated trophozoites (polykaryon) was also found. Multinucleation event was associated to cytokinesis failure leading to abortion of ongoing cell division. Consistently, genome-wide profiling of EhPC4 overexpressing trophozoites revealed the up-regulation of genes involved in carbohydrates and nucleic acids metabolism, chromosome segregation and cytokinesis. Forced overexpression of one of these genes, EhNUDC (nuclear movement protein), led to alterations in cytokinesis and partially recapitulated the multinucleation phenotype. These data indicate for the first time that EhPC4 is associated with events related to polyploidy and genome stability in E. histolytica.

  7. Taurine inhibits 2,5-hexanedione-induced oxidative stress and mitochondria-dependent apoptosis in PC12 cells

    PubMed Central

    LI, Shuangyue; GUAN, Huai; QIAN, Zhiqiang; SUN, Yijie; GAO, Chenxue; LI, Guixin; YANG, Yi; PIAO, Fengyuan; HU, Shuhai

    2016-01-01

    2,5-hexanedione (HD) is the ultimate neurotoxic metabolite of hexane, causing the progression of nerve diseases in human. It was reported that HD induced apoptosis and oxidative stress. Taurine has been shown to be a potent antioxidant. In the present study, we investigated the protection of taurine against HD-induced apoptosis in PC12 cells and the underlying mechanism. Our results showed the decreased viability and increased apoptosis in HD-exposed PC12 cells. HD also induced the disturbance of Bax and Bcl-2 expression, the loss of MMP, the release of mitochondrial cytochrome c and caspase-3 activation in PC12 cells. Moreover, HD resulted in an increase in reactive oxygen species (ROS) level and a decline in the activities of superoxidedismutase and catalase in PC12 cells. However, taurine pretreatment ameliorated the increased apoptosis and the alterations in key regulators of mitochondria-dependent pathway in PC12 exposed to HD. The increased ROS level and the decreased activities of the antioxidant enzymes in HD group were attenuated by taurine. These results indicate that pretreatment of taurine may, at least partly, prevent HD-induced apoptosis via inhibiting mitochondria-dependent pathway. It is also suggested that the potential of taurine against HD-induced apoptosis may benefit from its anti-oxidative property. PMID:27840369

  8. Genetic interactions among homologous recombination mutants in Candida albicans.

    PubMed

    Bellido, Alberto; Andaluz, Encarnación; Gómez-Raja, Jonathan; Álvarez-Barrientos, Alberto; Larriba, Germán

    2015-01-01

    rad52-ΔΔ and, to a lesser extent, rad51-ΔΔ deletants of Candidaalbicans displayed slow growth and aberrant filamentous morphology whereas rad59-ΔΔ mutants, both by growth rate and morphology resembled wild type. In this study, we have constructed pair-wise double deletants to analyze genetic interactions among these homologous recombination (HR) proteins that affect growth and morphology traits. When grown in liquid YPD medium, double mutant rad51-ΔΔ rad59-ΔΔ exhibited growth rates, cell and colony morphologies, and plating efficiencies that were not significantly different from those observed for rad51-ΔΔ. The same was true for rad52-ΔΔ rad59-ΔΔ compared to rad52-ΔΔ. Slow growth and decreased plating efficiency were caused, at least in part, by a decreased viability, as deduced from FUN1 staining. Flow cytometry and microscopic studies of filamentous mutant populations revealed major changes in cell ploidy, size and morphology, whereas DAPI staining identified complex nuclear rearrangements in yeast and filamentous cells. These phenotypes were not observed in the rad59-ΔΔ mutant populations. Our results show that abolishing Rad51 functions induces the appearance of a subpopulation of aberrant yeast and filamentous forms with increased cell size and ploidy. The size of this complex subpopulation was exacerbated in rad52-ΔΔ mutants. The combination of filamentous cell morphology and viability phenotypes was reflected on the colony morphology of the respective mutants. We conclude that the rad52 mutation is epistatic to rad51 for all the morphological traits analyzed. We discuss these results in the light of the several functions of these recombination genes.

  9. Subunit interface mutants of rabbit muscle aldolase form active dimers.

    PubMed Central

    Beernink, P. T.; Tolan, D. R.

    1994-01-01

    We report the construction of subunit interface mutants of rabbit muscle aldolase A with altered quaternary structure. A mutation has been described that causes nonspherocytic hemolytic anemia and produces a thermolabile aldolase (Kishi H et al., 1987, Proc Natl Acad Sci USA 84:8623-8627). The disease arises from substitution of Gly for Asp-128, a residue at the subunit interface of human aldolase A. To elucidate the role of this residue in the highly homologous rabbit aldolase A, site-directed mutagenesis is used to replace Asp-128 with Gly, Ala, Asn, Gln, or Val. Rabbit aldolase D128G purified from Escherichia coli is found to be similar to human D128G by kinetic analysis, CD, and thermal inactivation assays. All of the mutant rabbit aldolases are similar to the wild-type rabbit enzyme in secondary structure and kinetic properties. In contrast, whereas the wild-type enzyme is a tetramer, chemical crosslinking and gel filtration indicate that a new dimeric species exists for the mutants. In sedimentation velocity experiments, the mutant enzymes as mixtures of dimer and tetramer at 4 degrees C. Sedimentation at 20 degrees C shows that the mutant enzymes are > 99.5% dimeric and, in the presence of substrate, that the dimeric species is active. Differential scanning calorimetry demonstrates that Tm values of the mutant enzymes are decreased by 12 degrees C compared to wild-type enzyme. The results indicate that Asp-128 is important for interface stability and suggest that 1 role of the quaternary structure of aldolase is to provide thermostability. PMID:7833800

  10. Selection of chemotaxis mutants of Dictyostelium discoideum

    PubMed Central

    1987-01-01

    A method has been developed for the efficient selection of chemotaxis mutants of Dictyostelium discoideum. Mutants defective in the chemotactic response to folate could be enriched up to 30-fold in one round of selection using a chamber in which a compartment that contained the chemoattractant was separated by a sandwich of four nitrocellulose filters from a compartment that contained buffer. Mutagenized cells were placed in the center of the filter layer and exposed to the attractant gradient built up between the compartments for a period of 3-4 h. While wild-type cells moved through the filters in a wave towards the compartment that contained attractant, mutant cells remained in the filter to which they were applied. After several repetitions of the selection procedure, mutants defective in chemotaxis made up 10% of the total cell population retained in that filter. Mutants exhibiting three types of alterations were collected: motility mutants with either reduced speed of movement, or altered rates of turning; a single mutant defective in production of the attractant- degrading enzyme, folate deaminase; and mutants with normal motility but reduced chemotactic responsiveness. One mutant showed drastically reduced sensitivity in folate-induced cGMP production. Morphogenetic alterations of mutants defective in folate chemotaxis are described. PMID:3793759

  11. Correction of hair shaft defects through allele-specific silencing of mutant Krt75

    PubMed Central

    Liu, Ying; Snedecor, Elizabeth R.; Zhang, Xu; Xu, Yan-Feng; Huang, Lan; Jones, Evan; Zhang, Lianfeng; Clark, Richard A.; Roop, Dennis R.; Qin, Chuan; Chen, Jiang

    2015-01-01

    Dominant mutations in keratin genes can cause a number of inheritable skin disorders characterized by intraepidermal blistering, epidermal hyperkeratosis, or abnormalities in skin appendages, such as nail plate dystrophy and structural defects in hair. Allele-specific silencing of mutant keratins through RNA interference is a promising therapeutic approach for suppressing the expression of mutant keratins and related phenotypes in the epidermis. However, its effectiveness on skin appendages remains to be confirmed in vivo. In this study, we developed allele specific siRNAs capable of selectively suppressing the expression of a mutant Krt75, which causes hair shaft structural defects characterized by the development of blebs along the hair shaft in mice. Hair regenerated from epidermal keratinocyte progenitor cells isolated from mutant Krt75 mouse models reproduced the blebbing phenotype when grafted in vivo. In contrast, mutant cells manipulated with a lentiviral vector expressing mutant Krt75-specific shRNA persistently suppressed this phenotype. The phenotypic correction was associated with significant reduction of mutant Krt75 mRNA in the skin grafts. Thus, data obtained from this study demonstrated the feasibility of utilizing RNA interference to achieve durable correction of hair structural phenotypes through allele-specific silencing of the mutant keratin genes. PMID:26763422

  12. Correction of Hair Shaft Defects through Allele-Specific Silencing of Mutant Krt75.

    PubMed

    Liu, Ying; Snedecor, Elizabeth R; Zhang, Xu; Xu, Yanfeng; Huang, Lan; Jones, Evan C; Zhang, Lianfeng; Clark, Richard A; Roop, Dennis R; Qin, Chuan; Chen, Jiang

    2016-01-01

    Dominant mutations in keratin genes can cause a number of inheritable skin disorders characterized by intraepidermal blistering, epidermal hyperkeratosis, or abnormalities in skin appendages, such as nail plate dystrophy and structural defects in hair. Allele-specific silencing of mutant keratins through RNA interference is a promising therapeutic approach for suppressing the expression of mutant keratins and related phenotypes in the epidermis. However, its effectiveness on skin appendages remains to be confirmed in vivo. In this study, we developed allele-specific small interfering RNAs capable of selectively suppressing the expression of a mutant Krt75, which causes hair shaft structural defects characterized by the development of blebs along the hair shaft in mice. Hair regenerated from epidermal keratinocyte progenitor cells isolated from mutant Krt75 mouse models reproduced the blebbing phenotype when grafted in vivo. In contrast, mutant cells manipulated with a lentiviral vector expressing mutant Krt75-specific short hairpin RNA (shRNA) persistently suppressed this phenotype. The phenotypic correction was associated with a significant reduction of mutant Krt75 mRNA in the skin grafts. Thus, data obtained from this study demonstrated the feasibility of utilizing RNA interference to achieve durable correction of hair structural phenotypes through allele-specific silencing of mutant keratin genes.

  13. HDAC1, HDAC4, and HDAC9 Bind to PC3/Tis21/Btg2 and Are Required for Its Inhibition of Cell Cycle Progression and Cyclin D1 Expression.

    PubMed

    Micheli, Laura; D'Andrea, Giorgio; Leonardi, Luca; Tirone, Felice

    2017-07-01

    PC3/Tis21 is a transcriptional cofactor that inhibits proliferation in several cell types, including neural progenitors. Here, we report that PC3/Tis21 associates with HDAC1, HDAC4, and HDAC9 in vivo, in fibroblast cells. Furthermore, when HDAC1, HDAC4, or HDAC9 are silenced in fibroblasts or in a line of cerebellar progenitor cells, the ability of PC3/Tis21 to inhibit proliferation is significantly reduced. Overexpression of HDAC1, HDAC4, or HDAC9 in fibroblasts and in cerebellar precursor cells synergizes with PC3/Tis21 in inhibiting the expression of cyclin D1, a cyclin selectively inhibited by PC3/Tis21. Conversely, the depletion of HDAC1 or HDAC4 (but not HDAC9) in fibroblasts and in cerebellar precursor cells significantly impairs the ability of PC3/Tis21 to inhibit cyclin D1 expression. An analysis of HDAC4 deletion mutants shows that both the amino-terminal moiety and the catalytic domain of HDAC4 associate to PC3/Tis21, but neither alone is sufficient to potentiate the inhibition of cyclin D1 by PC3/Tis21. As a whole, our findings indicate that PC3/Tis21 inhibits cell proliferation in a way dependent on the presence of HDACs, in fibroblasts as well as in neural cells. Considering that several reports have demonstrated that HDACs can act as transcriptional corepressors on the cyclin D1 promoter, our data suggest that the association of PC3/Tis21 to HDACs is functional to recruit them to target genes, such as cyclin D1, for repression of their expression. J. Cell. Physiol. 232: 1696-1707, 2017. © 2016 Wiley Periodicals, Inc.

  14. Oxidative stress-mediated cytotoxicity of zirconia nanoparticles on PC12 and N2a cells

    NASA Astrophysics Data System (ADS)

    Asadpour, Elham; Sadeghnia, Hamid R.; Ghorbani, Ahmad; Sedaghat, Mehran; Boroushaki, Mohammad T.

    2016-01-01

    In recent years, there is a growing interest in the application of nanoparticles like zirconium dioxide (zirconia <100 nm), for many purposes. Since a comprehensive study on the toxic effects of zirconia has not been done, we decided to investigate the effects of zirconia nanoparticles on cultured PC12 and N2a cells. In this study, cytotoxic effect of different concentrations of zirconia nanoparticles at three different time intervals were evaluated using MTT and ROS (reactive oxygen species) assays. Also, Lipid peroxidation, glutathione (GSH) content changes, and DNA damage were measured. Zirconia nanoparticles caused a significant reduction in cell viability and GSH content of the cells, and induce a significant increase in intracellular ROS and MDA content of PC12 and N2a cells. Moreover, it increases the percentage of DNA tail of treated cells as compared with control group. Zirconia nanoparticles have cytotoxic and genotoxic effects in PC12 and N2a cells in a time and concentration-dependent manner in concentration more than 31 µg/mL.

  15. Inhibition of ANO1 by luteolin and its cytotoxicity in human prostate cancer PC-3 cells

    PubMed Central

    Park, Jinhong; Jeon, Dong-kyu; Jo, Sungwoo; Lee, Ho K.

    2017-01-01

    Anoctamin 1 (ANO1), a calcium-activated chloride channel, is highly amplified in prostate cancer, the most common form of cancer and leading causes of cancer death in men, and downregulation of ANO1 expression or its functional activity is known to inhibit cell proliferation, migration and invasion in prostate cancer cells. Here, we performed a cell-based screening for the identification of ANO1 inhibitors as potential anticancer therapeutic agents for prostate cancer. Screening of ~300 selected bioactive natural products revealed that luteolin is a novel potent inhibitor of ANO1. Electrophysiological studies indicated that luteolin potently inhibited ANO1 chloride channel activity in a dose-dependent manner with an IC50 value of 9.8 μM and luteolin did not alter intracellular calcium signaling in PC-3 prostate cancer cells. Luteolin inhibited cell proliferation and migration of PC-3 cells expressing high levels of ANO1 more potently than that of ANO1-deficient PC-3 cells. Notably, luteolin not only inhibited ANO1 channel activity, but also strongly decreased protein expression levels of ANO1. Our results suggest that downregulation of ANO1 by luteolin is a potential mechanism for the anticancer effect of luteolin. PMID:28362855

  16. Hydrogen gas alleviates oxygen toxicity by reducing hydroxyl radical levels in PC12 cells

    PubMed Central

    Yu, Junchao; Yu, Qiuhong; Liu, Yaling; Zhang, Ruiyun; Xue, Lianbi

    2017-01-01

    Hyperbaric oxygen (HBO) therapy through breathing oxygen at the pressure of above 1 atmosphere absolute (ATA) is useful for varieties of clinical conditions, especially hypoxic-ischemic diseases. Because of generation of reactive oxygen species (ROS), breathing oxygen gas at high pressures can cause oxygen toxicity in the central nervous system, leading to multiple neurological dysfunction, which limits the use of HBO therapy. Studies have shown that Hydrogen gas (H2) can diminish oxidative stress and effectively reduce active ROS associated with diseases. However, the effect of H2 on ROS generated from HBO therapy remains unclear. In this study, we investigated the effect of H2 on ROS during HBO therapy using PC12 cells. PC12 cells cultured in medium were exposed to oxygen gas or mixed oxygen gas and H2 at 1 ATA or 5 ATA. Cells viability and oxidation products and ROS were determined. The data showed that H2 promoted the cell viability and inhibited the damage in the cell and mitochondria membrane, reduced the levels of lipid peroxidation and DNA oxidation, and selectively decreased the levels of •OH but not disturbing the levels of O2•-, H2O2, or NO• in PC12 cells during HBO therapy. These results indicated that H2 effectively reduced •OH, protected cells against oxygen toxicity resulting from HBO therapy, and had no effect on other ROS. Our data supported that H2 could be potentially used as an antioxidant during HBO therapy. PMID:28362819

  17. Ternary phase diagram of dipalmitoyl-PC/dilauroyl-PC/cholesterol: nanoscopic domain formation driven by cholesterol.

    PubMed Central

    Feigenson, G W; Buboltz, J T

    2001-01-01

    A ternary phase diagram is proposed for the hydrated lamellar lipid mixture dipalmitoylphosphatidylcholine/dilauroylphosphatidylcholine/cholesterol (DPPC/DLPC/cholesterol) at room temperature. The entire composition space has been thoroughly mapped by complementary experimental techniques, revealing interesting phase behavior that has not been previously described. Confocal fluorescence microscopy shows a regime of coexisting DPPC-rich ordered and DLPC-rich fluid lamellar phases, having an upper boundary at apparently constant cholesterol mole fraction chi(chol) approximately 0.16. Fluorescence resonance energy transfer experiments confirm the identification and extent of this two-phase regime and, furthermore, reveal a 1-phase regime between chi(chol) approximately 0.16 and 0.25, consisting of ordered and fluid nanoscopic domains. Dipyrene-PC excimer/monomer measurements confirm the new regime between chi(chol) approximately 0.16 and 0.25 and also show that rigidly ordered phases seem to disappear around chi(chol) approximately 0.25. This study should be considered as a step toward a more complete understanding of lateral heterogeneity within biomembranes. Cholesterol may play a role in domain separation on the nanometer scale. PMID:11371452

  18. Expression and properties of wild-type and mutant forms of the Drosophila sex comb on midleg (SCM) repressor protein.

    PubMed

    Bornemann, D; Miller, E; Simon, J

    1998-10-01

    The Sex comb on midleg (Scm) gene encodes a transcriptional repressor of the Polycomb group (PcG). Here we show that SCM protein is nuclear and that its expression is widespread during fly development. SCM protein contains a C-terminal domain, termed the SPM domain, which mediates protein-protein interactions. The biochemical function of another domain consisting of two 100-amino-acid-long repeats, termed "mbt" repeats, is unknown. We have determined the molecular lesions of nine Scm mutant alleles, which identify functional requirements for specific domains. The Scm alleles were tested for genetic interactions with mutations in other PcG genes. Intriguingly, three hypomorphic Scm mutations, which map within an mbt repeat, interact with PcG mutations more strongly than do Scm null alleles. The strongest interactions produce partial synthetic lethality that affects doubly heterozygous females more severely than males. We show that mbt repeat alleles produce stable SCM proteins that associate with normal sites in polytene chromosomes. We also analyzed progeny from Scm mutant germline clones to compare the effects of an mbt repeat mutation during embryonic vs. pupal development. We suggest that the mbt repeat alleles produce altered SCM proteins that incorporate into and impair function of PcG protein complexes.

  19. Fast-moving diffuse auroral patches: A new aspect of daytime Pc3 auroral pulsations

    NASA Astrophysics Data System (ADS)

    Motoba, Tetsuo; Ebihara, Yusuke; Kadokura, Akira; Engebretson, Mark J.; Lessard, Marc R.; Weatherwax, Allan T.; Gerrard, Andrew J.

    2017-02-01

    Auroral pulsations are a convenient diagnostic of wave-particle interactions in the magnetosphere. A case study of a daytime Pc3 (22-100 mHz) auroral pulsation event, measured with a 2 Hz sampling all-sky camera at South Pole Station (74.4°S magnetic latitude) on 17 May 2012, is presented. The daytime Pc3 auroral pulsations were most active in a closed field line region where the aurora was dominated by diffuse green-line emissions and within ±2 h of magnetic local noon. Usually, but not always, the corresponding periodic variations were recorded with a colocated search coil magnetometer. Of particular interest is the two-dimensional auroral signature, indicating that the temporal luminosity variations at a given point were due to repeated formation and horizontal motion of faint, nonpulsating auroral patches with scale sizes of 100 km. The individual patches propagated equatorward with speeds of 15 km s-1 up to 20-25 km s-1 one after another along the magnetic meridian through local magnetic zenith. These properties differ considerably from typical pulsating aurorae, being periodic on-off luminosity variations in a particular auroral patch and drifting in accordance with the convection electric field in the magnetosphere. We speculate that such repetitive patterns of the fast-moving auroral patches, being another aspect of the daytime Pc3 auroral pulsations, may be a visible manifestation of compressional Pc3 waves which propagate earthward and cause modulation of precipitating keV electron fluxes in the dayside outer magnetosphere.

  20. Targeting ESR1-Mutant Breast Cancer

    DTIC Science & Technology

    2015-09-01

    Award Number: W81XWH-14-1-0360 TITLE: Targeting ESR1- Mutant Breast Cancer PRINCIPAL INVESTIGATOR: Geoffrey L. Greene, Ph.D. CONTRACTING...ADDRESS. 1. REPORT DATE September 2015 2. REPORT TYPE Annual 3. DATES COVERED 1 Sep 2014 - 31 Aug 2015 4. TITLE AND SUBTITLE Targeting ESR1- Mutant ...approved hormonal therapies and that more potent, selective estrogen receptor degraders (SERDs) will enable complete inhibition of mutant ER signaling and

  1. Targeting ESR1-Mutant Breast Cancer

    DTIC Science & Technology

    2015-09-01

    AWARD NUMBER: W81XWH-14-1-0359 TITLE: Targeting ESR1- Mutant Breast Cancer PRINCIPAL INVESTIGATOR: Dr. Sarat Chandarlapaty CONTRACTING...31 Aug 2015 4. TITLE AND SUBTITLE Targeting ESR1- Mutant Breast Cancer 5a. CONTRACT NUMBER 5b. GRANT NUMBER W81XWH-14-1-0359 5c. PROGRAM ELEMENT...current FDA approved hormonal therapies and that more potent, selective estrogen receptor degraders (SERDs) will enable complete inhibition of mutant

  2. Demonstration of a PC 25 Fuel Cell in Russia

    SciTech Connect

    John C. Trocciola; Thomas N. Pompa; Linda S. Boyd

    2004-09-01

    This project involved the installation of a 200kW PC25C{trademark} phosphoric-acid fuel cell power plant at Orgenergogaz, a Gazprom industrial site in Russia. In April 1997, a PC25C{trademark} was sold by ONSI Corporation to Orgenergogaz, a subsidiary of the Russian company ''Gazprom''. Due to instabilities in the Russian financial markets, at that time, the unit was never installed and started by Orgenergogaz. In October of 2001 International Fuel Cells (IFC), now known as UTC Fuel Cells (UTCFC), received a financial assistance award from the United States Department of Energy (DOE) entitled ''Demonstration of PC 25 Fuel Cell in Russia''. Three major tasks were part of this award: the inspection of the proposed site and system, start-up assistance, and installation and operation of the powerplant.

  3. Aspartame-induced apoptosis in PC12 cells.

    PubMed

    Horio, Yukari; Sun, Yongkun; Liu, Chuang; Saito, Takeshi; Kurasaki, Masaaki

    2014-01-01

    Aspartame is an artificial sweetner added to many low-calorie foods. The safety of aspartame remains controversial even though there are many studies on its risks. In this study, to understand the physiological effects of trace amounts of artificial sweetners on cells, the effects of aspartame on apoptosis were investigated using a PC12 cell system. In addition, the mechanism of apoptosis induced by aspartame in PC12 cells and effects on apoptotic factors such as cytochrome c, apoptosis-inducing factor, and caspase family proteins were studied by Western blotting and RT-PCR. Aspartame-induced apoptosis in PC12 cells in a dose-dependent manner. In addition, aspartame exposure increased the expressions of caspases 8 and 9, and cytochrome c. These results indicate that aspartame induces apoptosis mainly via mitochondrial pathway involved in apoptosis due to oxigen toxicity.

  4. Expression of dystrophin Dp71 during PC12 cell differentiation.

    PubMed

    Cisneros, B; Rendon, A; Genty, V; Aranda, G; Marquez, F; Mornet, D; Montañez, C

    1996-08-02

    The expression of dystrophin-protein 71 (Dp71) was investigated during nerve growth factor (NGF) induced differentiation of PC12 cells. A semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) assay was designed to measure Dp71 mRNA, whereas the Dp71 protein amount was evaluated by immunoblot analysis using an anti-dystrophin monoclonal antibody. Comparison with control cultures showed that Dp71 mRNA and protein levels increased in parallel with NGF treatment peaking with increments of 60% and 1.4 times, respectively. The upregulation of Dp71 expression during PC12 cells differentiation point at PC12 cells as a suitable model for studying the function of Dp71 in neuronal cells.

  5. Modelling of Pc5 pulsation structure in the magnetosphere

    NASA Astrophysics Data System (ADS)

    Walker, A. D. M.

    1980-03-01

    Magnetohydrodynamic resonance theory is used to model the structure of the magnetospheric and ionospheric electric and magnetic fields associated with Pc5 geomagnetic pulsations. In this paper the variation of the fields across the invariant latitude of the resonance are computed. The results are combined with calculations of the variation along a field line to map the fields down to the ionosphere. In one case the results are compared with measurements obtained by the STARE auroral radar and show good agreement. The relationship between the width of the resonance region and ionospheric height-integrated Pedersen conductivity is computed and it is shown how auroral radar measurements of Pc5 oscillations could be used to determine ionospheric height-integrated Pedersen conductivity. It is pointed out that from these calculations it would be possible to identify the field line on which a satellite was located by comparing a Pc5 pulsation observed by the satellite, and the same pulsation observed by STARE.

  6. PC-1D installation manual and user's guide

    SciTech Connect

    Basore, P.A.

    1991-05-01

    PC-1D is a software package for personal computers that uses finite-element analysis to solve the fully-coupled two-carrier semiconductor transport equations in one dimension. This program is particularly useful for analyzing the performance of optoelectronic devices such as solar cells, but can be applied to any bipolar device whose carrier flows are primarily one-dimensional. This User's Guide provides the information necessary to install PC-1D, define a problem for solution, solve the problem, and examine the results. Example problems are presented which illustrate these steps. The physical models and numerical methods utilized are presented in detail. This document supports version 3.1 of PC-1D, which incorporates faster numerical algorithms with better convergence properties than previous versions of the program. 51 refs., 17 figs., 5 tabs.

  7. Tablet PC as a mobil PACS terminal using wireless LAN

    NASA Astrophysics Data System (ADS)

    Tsao, Bo-Shen; Ching, Yu-Tai; Lee, Wen-Jeng; Chen, Shyh-Jye; Chang, Chia-Hung; Chen, Chien-Jung; Yen, York; Lee, Yuan-Ten

    2003-05-01

    A PACS mobile terminal has applications in ward round, emergency room and remote teleradiology consultation. Personal Digital Assistants (PDAs) have the highest mobility and are used for many medical applications. However, their roles are limited in the field of radiology due to small screen size. In this study, we built a wireless PACS terminal using a hand-held tablet-PC. A tablet PC (X-pilot, LEO systems, Taiwan) running the WinCE operating systems was used as our mobile PACS terminal. This device is equipped with 800×600 resolution 10.4 inch TFT monitor. The network connection between the tablet PC and the server was linked via wireless LAN (IEEE 802.11b).

  8. Electrophysiological study of Drosophila rhodopsin mutants

    PubMed Central

    1986-01-01

    Electrophysiological investigations were carried out on several independently isolated mutants of the ninaE gene, which encodes opsin in R1-6 photoreceptors, and a mutant of the ninaD gene, which is probably important in the formation of the rhodopsin chromophore. In these mutants, the rhodopsin content in R1-6 photoreceptors is reduced by 10(2)-10(6)-fold. Light-induced bumps recorded from even the most severely affected mutants are physiologically normal. Moreover, a detailed noise analysis shows that photoreceptor responses of both a ninaE mutant and a ninaD mutant follow the adapting bump model. Since any extensive rhodopsin-rhodopsin interactions are not likely in these mutants, the above results suggest that such interactions are not needed for the generation and adaptation of light-induced bumps. Mutant bumps are strikingly larger in amplitude than wild-type bumps. This difference is observed both in ninaD and ninaE mutants, which suggests that it is due to severe depletion of rhodopsin content, rather than to any specific alterations in the opsin protein. Lowering or buffering the intracellular calcium concentration by EGTA injection mimics the effects of the mutations on the bump amplitude, but, unlike the mutations, it also affects the latency and kinetics of light responses. PMID:3097245

  9. Realizing broad-bandwidth visible wavelength photodiode based on solution-processed ZnPc/PC71BM dyad

    NASA Astrophysics Data System (ADS)

    Zafar, Qayyum; Fatima, Noshin; Karimov, Khasan S.; Ahmed, Muhammad M.; Sulaiman, Khaulah

    2017-02-01

    Herein, we demonstrate a solution-processed visible wavelength organic photodiode (OPD) using donor/acceptor dyad of zinc phthalocyanine (ZnPc) and [6,6]-phenyl-C71-butyric-acid methyl ester (PC71BM), respectively. The synergic absorption profiles of both ZnPc and PC71BM moieties have been exploited to realize broader (350 and 800 nm) and consistent absorption spectrum of the photoactive film. The optimum loading ratio (by volume) of D/A dyad has been estimated to be 1:0.8, via quenching phenomenon in ZnPc photoluminescence spectrum. The performance of the OPD has been evaluated by detecting the photocurrent density with respect to varied illumination levels (0-150 mW/cm2) of impinging light at different reverse bias conditions. Under identical reverse bias mode, the photocurrent density has shown significant upsurge as the incident intensity of light is increased; ultimately leading to the significantly higher responsivity (162.4 μA/W) of the fabricated diode. The light to dark current density ratio (Jph/Jd) of the device at 3 V reverse bias has been calculated to be ∼20.12. The transient photocurrent density response of the fabricated OPD has also been characterized at -4 V operational bias under switch ON/OFF illumination. The measured response and recovery time for the fabricated OPD are ∼200 and 300 ms, respectively.

  10. Abnormal grooming activity in Dab1(scm) (scrambler) mutant mice.

    PubMed

    Strazielle, C; Lefevre, A; Jacquelin, C; Lalonde, R

    2012-07-15

    Dab1(scm) mutant mice, characterized by cell ectopias and degeneration in cerebellum, hippocampus, and neocortex, were compared to non-ataxic controls for different facets of grooming caused by brief water immersions, as well as some non-grooming behaviors. Dab1(scm) mutants were strongly affected in their quantitative functional parameters, exhibiting higher starting latencies before grooming relative to non-ataxic littermates of the A/A strain, fewer grooming bouts, and grooming components of shorter duration, with an unequal regional distribution targeting almost totally the rostral part (head washing and forelimb licking) of the animal. Only bouts of a single grooming element were preserved. The cephalocaudal order of grooming elements appeared less disorganized, mutant and control mice initiating the grooming with head washing and forelimb licking prior to licking posterior parts. However, mutants differed from controls in that all their bouts were incomplete but uninterrupted, although intergroup difference for percentage of the incorrect transitions was not significant. In contrast to grooming, Dab1(scm) mice ambulated for a longer time. During walking episodes, they exhibited more body scratching than controls, possibly to compensate for the lack of licking different body parts. In conjunction with studies with other ataxic mice, these results indicate that the cerebellar cortex affects grooming activity and is consequently involved in executing various components, but not in its sequential organization, which requires other brain regions such as cerebral cortices or basal ganglia.

  11. Run-08 pC polarization analysis - October 16, 2008

    SciTech Connect

    Dharmawardane,V.; Bazilevsky,A.; Bunce, G.; Gill, R.; Huang, H.; Makdisi, Y.; Nakagawa, I.; Morozov, B.; Okada, H.; Sivertz, M.; Zelenski, A.; Alekseev, I.; Svirida, D.

    2009-03-01

    In this note we will discuss the analysis of RHIC run 08 pC data that were collected during February 14 - March 10, 2008. An analysis method that is similar to Run 05 and Run 06 was adopted for Run 08 analysis (except few minor changes, which are described below). A detailed analysis note and a NIM article that describe the pC analysis procedure (for run 05 and run 06) can be found elsewhere. In brief, the analysis consists of calibrating the detectors, determining energy corrections ('dead layers'), determining good runs and extracting the polarization from data.

  12. PC-Based Applications Programming on the SRS Control System.

    NASA Astrophysics Data System (ADS)

    Martlew, Brian; Owen, Hywel; Pugh, Martin; Rawlinson, Bill; Smith, Susan

    1997-05-01

    The CERN PC-based ISOLDE control system has been installed at the SRS electron storage ring at Daresbury Laboratory. The use of Windows NT for the control consoles together with PC and VME front-end computers running under several operating systems has resulted in a flexible and reliable system for accelerator control. The implementation and philosophy of control application programs, based around a suite of Microsoft Visual Basic and Excel programs, is described. In particular, the use of Excel to provide adaptable programs online allows rapid generation of new control functions; orbit correction and servoing at the application level are described as examples of this.

  13. Effect of morphine on PC12 cells with molecular radar

    NASA Astrophysics Data System (ADS)

    Shi, Chen; Yu, Xiaoli; Lu, Jiuyi; Zhang, Chunyang; Jin, Lei; Ma, Hui; Zhang, Dacheng; Chen, Die Yan

    2000-10-01

    Molecular Radar (MR) is a new method to detect biological processes in living cells at the level of molecular, it is also the newest means to get intracellular information. In this paper we study the effect of morphine on PC12 cells using MR. The results show that the effect of morphine on PC12 cells is time- and concentration-dependent. Morphine treating for short time induces the increase and fluctuation of intracellular (CA2+), while morphine treating for long time induces chromatin condensation, loss of mitochondria membrane potential apoptosis.

  14. PC Windows finite element modeling of landfill gas flow

    SciTech Connect

    Mull, S.R.; Lang, R.J.; Vigil, S.A.; Cota, H.

    1996-09-01

    A two dimensional demonstration program, GAS, has been developed for the solution of landfill gas (LFG) flow problems on a personal computer (PC). The program combines a Windows{trademark} graphical user interface, object oriented programming (OOP) techniques, and finite element modeling (FEM) to demonstrate the practicality of performing LFG flow modeling on the PC. GAS is demonstrated on a sample LFG problem consisting of a landfill, one gas extraction well, the landfill liner, cap, and surrounding soil. Analyses of the program results are performed for successively finer grid resolutions. Element flux imbalance, execution time, and required memory are characterized as a function of grid resolution.

  15. Wf/pc Cycle 1 Calibration: Internal Delta Flats

    NASA Astrophysics Data System (ADS)

    MacKenty, John

    1990-12-01

    This calibration takes internal flats periodically to permit the repair of external (earth-target) flat fields changed by pixel dependent QE changes which occur whenever the WF/PC is decontaminated. Note that the internal flat lamp is also useful with relatively wide filters in the red. This obtains intflats for: WFC: F439W, F547M, F555W, F622W, F702W, F785LP PC: F439W, F547M, F555W, F622W, F702W, F718M, F785LP

  16. Glial dysfunction causes age-related memory impairment in Drosophila.

    PubMed

    Yamazaki, Daisuke; Horiuchi, Junjiro; Ueno, Kohei; Ueno, Taro; Saeki, Shinjiro; Matsuno, Motomi; Naganos, Shintaro; Miyashita, Tomoyuki; Hirano, Yukinori; Nishikawa, Hiroyuki; Taoka, Masato; Yamauchi, Yoshio; Isobe, Toshiaki; Honda, Yoshiko; Kodama, Tohru; Masuda, Tomoko; Saitoe, Minoru

    2014-11-19

    Several aging phenotypes, including age-related memory impairment (AMI), are thought to be caused by cumulative oxidative damage. In Drosophila, age-related impairments in 1 hr memory can be suppressed by reducing activity of protein kinase A (PKA). However, the mechanism for this effect has been unclear. Here we show that decreasing PKA suppresses AMI by reducing activity of pyruvate carboxylase (PC), a glial metabolic enzyme whose amounts increase upon aging. Increased PC activity causes AMI through a mechanism independent of oxidative damage. Instead, increased PC activity is associated with decreases in D-serine, a glia-derived neuromodulator that regulates NMDA receptor activity. D-serine feeding suppresses both AMI and memory impairment caused by glial overexpression of dPC, indicating that an oxidative stress-independent dysregulation of glial modulation of neuronal activity contributes to AMI in Drosophila.

  17. Spontaneous white sectored-mutants in Streptomyces hygroscopicus 111-81: characterization and antibiotic productivity.

    PubMed

    Gesheva, Victoria

    2008-08-01

    Spontaneous white mutants from sectors of Streptomyces hygroscopicus 111-81 were isolated. The comparison of morphological, cultural, and biochemical properties of the mutants and ancestor showed the differences in colors of aerial, substrate mycelia, and sporulation. Changes in resistance to antibiotics and sensitivity to lysozyme indicated alterations in cell walls and cell membranes of the mutants. They showed antifungal activity close to that of the parent strain on fermentation medium FM2, with unchanged component composition of the AK-111-81 antibiotic complex. The cells of spontaneous white mutants are characterized with electron-transparent structures, vacuoles, aggregation of ribosomes, intrahyphal growth, and lack of multiple cell septa, which was established by transmission electron microscopy. The appearance of white sectored-mutants in S. hygroscopicus 111-81 is connected with exhausting of nutrients causing the substrate limitation and is a stress response to starvation.

  18. A non-cell autonomous mouse model of CNS haemangioblastoma mediated by mutant KRAS

    PubMed Central

    Bao, Leyuan; Al-Assar, Osama; Drynan, Lesley F.; Arends, Mark J.; Tyers, Pam; Barker, Roger A.; Rabbitts, Terence H.

    2017-01-01

    Haemangioblastoma is a rare malignancy of the CNS where vascular proliferation causes lesions due to endothelial propagation. We found that conditionally expressing mutant Kras, using Rag1-Cre, gave rise to CNS haemangioblastoma in the cortex and cerebellum in mice that present with highly vascular tumours with stromal cells similar to human haemangioblastomas. The aberrant haemangioblastoma endothelial cells do not express mutant Kras but rather the mutant oncogene is expressed in CNS interstitial cells, including neuronal cells and progeny. This demonstrates a non-cell autonomous origin of this disease that is unexpectedly induced via Rag1-Cre expression in CNS interstitial cells. This is the first time that mutant RAS has been shown to stimulate non-cell autonomous proliferation in malignancy and suggests that mutant RAS can control endothelial cell proliferation in neo-vascularisation when expressed in certain cells. PMID:28322325

  19. Mutant calreticulin requires both its mutant C-terminus and the thrombopoietin receptor for oncogenic transformation

    PubMed Central

    Elf, Shannon; Abdelfattah, Nouran S.; Chen, Edwin; Perales-Patón, Javier; Rosen, Emily A.; Ko, Amy; Peisker, Fabian; Florescu, Natalie; Giannini, Silvia; Wolach, Ofir; Morgan, Elizabeth A.; Tothova, Zuzana; Losman, Julie-Aurore; Schneider, Rebekka K.; Al-Shahrour, Fatima; Mullally, Ann

    2016-01-01

    Somatic mutations in calreticulin (CALR) are present in approximately 40% of patients with myeloproliferative neoplasms (MPN) but the mechanism by which mutant CALR is oncogenic remains unclear. Here, we demonstrate that expression of mutant CALR alone is sufficient to engender MPN in mice and recapitulates the disease phenotype of CALR-mutant MPN patients. We further show that the thrombopoietin receptor, MPL is required for mutant CALR-driven transformation through JAK-STAT pathway activation, thus rendering mutant CALR-transformed hematopoietic cells sensitive to JAK2 inhibition. Finally, we demonstrate that the oncogenicity of mutant CALR is dependent on the positive electrostatic charge of the C-terminus of the mutant protein, which is necessary for physical interaction between mutant CALR and MPL. Together, our findings elucidate a novel paradigm of cancer pathogenesis and reveal how CALR mutations induce MPN. PMID:26951227

  20. Catalytic properties of ADAM12 and its domain deletion mutants.

    PubMed

    Jacobsen, Jonas; Visse, Robert; Sørensen, Hans Peter; Enghild, Jan J; Brew, Keith; Wewer, Ulla M; Nagase, Hideaki

    2008-01-15

    Human ADAM12 (a disintegrin and metalloproteinase) is a multidomain zinc metalloproteinase expressed at high levels during development and in human tumors. ADAM12 exists as two splice variants: a classical type 1 membrane-anchored form (ADAM12-L) and a secreted splice variant (ADAM12-S) consisting of pro, catalytic, disintegrin, cysteine-rich, and EGF domains. Here we present a novel activity of recombinant ADAM12-S and its domain deletion mutants on S-carboxymethylated transferrin (Cm-Tf). Cleavage of Cm-Tf occurred at multiple sites, and N-terminal sequencing showed that the enzyme exhibits restricted specificity but a consensus sequence could not be defined as its subsite requirements are promiscuous. Kinetic analysis revealed that the noncatalytic C-terminal domains are important regulators of Cm-Tf activity and that ADAM12-PC consisting of the pro domain and catalytic domain is the most active on this substrate. It was also observed that NaCl inhibits ADAM12. Among the tissue inhibitors of metalloproteinases (TIMP) examined, the N-terminal domain of TIMP-3 (N-TIMP-3) inhibits ADAM12-S and ADAM12-PC with low nanomolar Ki(app) values while TIMP-2 inhibits them with a slightly lower affinity (9-44 nM). However, TIMP-1 is a much weaker inhibitor. N-TIMP-3 variants that lack MMP inhibitory activity but retained the ability to inhibit ADAM17/TACE failed to inhibit ADAM12. These results indicate unique enzymatic properties of ADAM12 among the members of the ADAM family of metalloproteinases.

  1. Immunocytochemical localization of prohormone convertases PC1 and PC2 in the anuran pituitary gland: subcellular localization in corticotrope and melanotrope cells.

    PubMed

    Kurabuchi, S; Tanaka, S

    1997-06-01

    Specific antisera against mammalian prohormone convertases PC1 and PC2 have been used to examine, light-immunocytochemically, the distribution of these enzymes in the pituitary gland of five different species of anuran amphibians (Rana catesbeiana, Bufo japonicus formosus, Xenopus laevis, Rana brevipoda porosa, and Buergeria japonica). A differential pattern of immunoreactivity of PC1 and PC2 was found among these species. Only PC1 was found in the corticotrope cells of the pars distalis in R. catesbeiana, B. japonicus formosus, and X. laevis. Only PC2 was observed in these cells in B. japonica, whereas both PC1 and PC2 were present in these cells in R. brevipoda porosa. PC2 immunoreactivity was always observed in melanotrope cells in the pars intermedia of all of the species, but it coexisted with PC1 immunoreactivity only in R. catesbeiana and X. laevis. The nerve fibers and terminals in the pars nervosa in all of the species were intensely immunopositive with both PC1 and PC2 antibodies. Immunoelectron microscopy on B. japonicus formosus and B. japonica, by means of double-labeling with gold particles of different sizes, revealed that almost all the adrenocorticotropin-positive secretory granules in the corticotrope cells and alpha-melanophore-stimulating-hormone-positive secretory granules in the melanotrope cells were also labeled with either PC1 or PC2 antibodies. This study suggests that PC1 and PC2 are involved in the intracellular proteolytic cleavage of proopiomelanocortin in amphibian pituitary glands, a situation similar to that proposed for mammals.

  2. Processing and intracellular localization of rice stripe virus Pc2 protein in insect cells

    SciTech Connect

    Zhao, Shuling; Zhang, Gaozhan; Dai, Xuejuan; Hou, Yanling; Li, Min; Liang, Jiansheng; Liang, Changyong

    2012-08-01

    Rice stripe virus (RSV) belongs to the genus Tenuivirus and its genome consists of four single-stranded RNAs encoding seven proteins. Here, we have analyzed the processing and membrane association of Pc2 encoded by vcRNA2 in insect cells. The enhanced green fluorescent protein (eGFP) was fused to the Pc2 and used for the detection of Pc2 fusion proteins. The results showed that Pc2 was cleaved to produce two proteins named Pc2-N and Pc2-C. When expressed alone, either Pc2-N or Pc2-C could transport to the Endoplasmic reticulum (ER) membranes independently. Further mutagenesis studies revealed that Pc2 contained three ER-targeting domains. The results led us to propose a model for the topology of the Pc2 in which an internal signal peptide immediately followed a cleavage site, and two transmembrane regions are contained.

  3. Na+/Ca2+ exchanger inhibitors inhibit neurite outgrowth in PC12 cells.

    PubMed

    Oda, Toru; Kume, Toshiaki; Izumi, Yasuhiko; Ishihara, Kumatoshi; Sugmimoto, Hachiro; Akaike, Akinori

    2011-01-01

    To elucidate the role of Na(+)/Ca(2+) exchanger (NCX) in neurite outgrowth, we investigated the effects of NCX inhibitors on neurite outgrowth in PC12 cells. KB-R7943 and 3',4'-dichlorobenzamil, NCX inhibitors, inhibited the neurite outgrowth caused by nerve growth factor (NGF). NCX inhibitors inhibited the neurite outgrowth caused by dibutylyl cAMP, which rapidly reorganizes the cytoskeleton. KB-R7943 inhibited the neurite outgrowth caused by Y-27632, an inhibitor of Rho kinase (ROCK) that regulates actin. However, NCX inhibitors did not inhibit NGF-induced phosphorylation of extracellular signal-regulated kinase. These results suggest that NCX inhibitor affects downstream of the Rho-ROCK signal transduction pathways in neurite outgrowth.

  4. Optimization of a nanomedicine-based silicon phthalocyanine 4 photodynamic therapy (Pc 4-PDT) strategy for targeted treatment of EGFR-overexpressing cancers.

    PubMed

    Master, Alyssa M; Livingston, Megan; Oleinick, Nancy L; Sen Gupta, Anirban

    2012-08-06

    The current clinical mainstays for cancer treatment, namely, surgical resection, chemotherapy, and radiotherapy, can cause significant trauma, systemic toxicity, and functional/cosmetic debilitation of tissue, especially if repetitive treatment becomes necessary due to tumor recurrence. Hence there is significant clinical interest in alternate treatment strategies like photodynamic therapy (PDT) which can effectively and selectively eradicate tumors and can be safely repeated if needed. We have previously demonstrated that the second-generation photosensitizer Pc 4 (silicon phthalocyanine 4) can be formulated within polymeric micelles, and these micelles can be specifically targeted to EGFR-overexpressing cancer cells using GE11 peptide ligands, to enhance cell-specific Pc 4 delivery and internalization. In the current study, we report on the in vitro optimization of the EGFR-targeting, Pc 4 loading of the micellar nanoformulation, along with optimization of the corresponding photoirradiation conditions to maximize Pc 4 delivery, internalization, and subsequent PDT-induced cytotoxicity in EGFR-overexpressing cells in vitro. In our studies, absorption and fluorescence spectroscopy were used to monitor the cell-specific uptake of the GE11-decorated Pc 4-loaded micelles and the cytotoxic singlet oxygen production from the micelle-encapsulated Pc 4, to determine the optimum ligand density and Pc 4 loading. It was found that the micelle formulations bearing 10 mol % of GE11-modified polymer component resulted in the highest cellular uptake in EGFR-overexpressing A431 cells within the shortest incubation periods. Also, the loading of ∼ 50 μg of Pc 4 per mg of polymer in these micellar formulations resulted in the highest levels of singlet oxygen production. When formulations bearing these optimized parameters were tested in vitro on A431 cells for PDT effect, a formulation dose containing 400 nM Pc 4 and photoirradiation duration of 400 s at a fluence of 200 mJ/cm(2

  5. Secretory phospholipases A2 induce neurite outgrowth in PC12 cells.

    PubMed Central

    Nakashima, Satoru; Ikeno, Yutaka; Yokoyama, Tatsuya; Kuwana, Masakazu; Bolchi, Angelo; Ottonello, Simone; Kitamoto, Katsuhiko; Arioka, Manabu

    2003-01-01

    sPLA(2)s (secretory phospholipases A(2)) belong to a broad and structurally diverse family of enzymes that hydrolyse the sn -2 ester bond of glycerophospholipids. We previously showed that a secreted fungal 15 kDa protein, named p15, as well as its orthologue from Streptomyces coelicolor (named Scp15) induce neurite outgrowth in PC12 cells at nanomolar concentrations. We report here that both p15 and Scp15 are members of a newly identified group of fungal/bacterial sPLA(2)s. The phospholipid-hydrolysing activity of p15 is absolutely required for neurite outgrowth induction. Mutants with a reduced PLA(2) activity exhibited a comparable reduction in neurite-inducing activity, and the ability to induce neurites closely matched the capacity of various p15 forms to promote fatty acid release from live PC12 cells. A structurally divergent member of the sPLA(2) family, bee venom sPLA(2), also induced neurites in a phospholipase activity-dependent manner, and the same effect was elicited by mouse group V and X sPLA(2)s, but not by group IB and IIA sPLA(2)s. Lysophosphatidylcholine, but not other lysophospholipids, nor arachidonic acid, elicited neurite outgrowth in an L-type Ca(2+) channel activity-dependent manner. In addition, p15-induced neuritogenesis was unaffected by various inhibitors that block arachidonic acid conversion into bioactive eicosanoids. Altogether, these results delineate a novel, Ca(2+)- and lysophosphatidylcholine-dependent neurotrophin-like role of sPLA(2)s in the nervous system. PMID:12967323

  6. Enhancers of Conidiation Mutants in Aspergillus Nidulans

    PubMed Central

    Gems, D. H.; Clutterbuck, A. J.

    1994-01-01

    Mutants at a number of loci, designated sthenyo, have been isolated as enhancers of the oligoconidial mutations at the medA locus. Two loci have been mapped: sthA on linkage group I, and sthB on linkage group V. Two probable alleles have been identified at each locus but two further mutants were unlinked to either sthA or sthB. Neither sthA nor sthB mutants have conspicuous effects on morphology on their own, nor could the sthA1 sthB2 double mutant be distinguished from wild type. Mutants at both loci also interact with the temperature-sensitive brlA42 mutant at the permissive temperature to give a phenotype described as ``Abacoid.'' sthA1 also induces a slight modification of the phenotype of an abaA mutant. We conclude that sthenyo genes act mainly at the phialide stage of conidiation. We also describe the isolation of new medA mutants arising spontaneously as outgrowths on brlA42 colonies. PMID:8056325

  7. Regulation of Mutant p53 Protein Expression.

    PubMed

    Vijayakumaran, Reshma; Tan, Kah Hin; Miranda, Panimaya Jeffreena; Haupt, Sue; Haupt, Ygal

    2015-01-01

    For several decades, p53 has been detected in cancer biopsies by virtue of its high protein expression level which is considered indicative of mutation. Surprisingly, however, mouse genetic studies revealed that mutant p53 is inherently labile, similar to its wild type (wt) counterpart. Consistently, in response to stress conditions, both wt and mutant p53 accumulate in cells. While wt p53 returns to basal level following recovery from stress, mutant p53 remains stable. In part, this can be explained in mutant p53-expressing cells by the lack of an auto-regulatory loop with Mdm2 and other negative regulators, which are pivotal for wt p53 regulation. Further, additional protective mechanisms are acquired by mutant p53, largely mediated by the co-chaperones and their paralogs, the stress-induced heat shock proteins. Consequently, mutant p53 is accumulated in cancer cells in response to chronic stress and this accumulation is critical for its oncogenic gain of functions (GOF). Building on the extensive knowledge regarding wt p53, the regulation of mutant p53 is unraveling. In this review, we describe the current understanding on the major levels at which mutant p53 is regulated. These include the regulation of p53 protein levels by microRNA and by enzymes controlling p53 proteasomal degradation.

  8. A halotolerant mutant of Saccharomyces cerevisiae.

    PubMed Central

    Gaxiola, R; Corona, M; Zinker, S

    1996-01-01

    FRD, a nuclear and dominant spontaneous mutant of Saccharomyces cerevisiae capable of growing in up to 2 M NaCl, was isolated. Compared with parental cells, the mutant cells have a lower intracellular Na+/K+ ratio, shorter generation times in the presence of 1 M NaCl, and alterations in gene expression. PMID:8631691

  9. Transcriptomic comparison of Drosophila snRNP biogenesis mutants reveals mutant-specific changes in pre-mRNA processing: implications for spinal muscular atrophy.

    PubMed

    Garcia, Eric L; Wen, Ying; Praveen, Kavita; Matera, A Gregory

    2016-08-01

    Survival motor neuron (SMN) functions in the assembly of spliceosomal small nuclear ribonucleoproteins (snRNPs) that catalyze pre-mRNA splicing. Here, we used disruptions in Smn and two additional snRNP biogenesis genes, Phax and Ars2, to classify RNA processing differences as snRNP-dependent or gene-specific in Drosophila Phax and Smn mutants exhibited comparable reductions in snRNAs, and comparison of their transcriptomes uncovered shared sets of RNA processing changes. In contrast, Ars2 mutants displayed only small decreases in snRNA levels, and RNA processing changes in these mutants were generally distinct from those identified in Phax and Smn animals. Instead, RNA processing changes in Ars2 mutants support the known interaction of Ars2 protein with the cap-binding complex, as splicing changes showed a clear bias toward the first intron. Bypassing disruptions in snRNP biogenesis, direct knockdown of spliceosomal proteins caused similar changes in the splicing of snRNP-dependent events. However, these snRNP-dependent events were largely unaltered in three Smn mutants expressing missense mutations that were originally identified in human spinal muscular atrophy (SMA) patients. Hence, findings here clarify the contributions of Phax, Smn, and Ars2 to snRNP biogenesis in Drosophila, and loss-of-function mutants for these proteins reveal differences that help disentangle cause and effect in SMA model flies.

  10. PC Board Layout and Electronic Drafting with CAD. Teacher Edition.

    ERIC Educational Resources Information Center

    Bryson, Jimmy

    This teacher's guide contains 11 units of instruction for a course on computer electronics and computer-assisted drafting (CAD) using a personal computer (PC). The course covers the following topics: introduction to electronic drafting with CAD; CAD system and software; basic electronic theory; component identification; basic integrated circuit…

  11. Audio Podcasting in a Tablet PC-Enhanced Biochemistry Course

    ERIC Educational Resources Information Center

    Lyles, Heather; Robertson, Brian; Mangino, Michael; Cox, James R.

    2007-01-01

    This report describes the effects of making audio podcasts of all lectures in a large, basic biochemistry course promptly available to students. The audio podcasts complement a previously described approach in which a tablet PC is used to annotate PowerPoint slides with digital ink to produce electronic notes that can be archived. The fundamentals…

  12. Using a PC as a Frequency Meter or a Counter.

    ERIC Educational Resources Information Center

    Sartori, J.; And Others

    1995-01-01

    Describes hardware that enables the use of an IBM PC microcomputer as a frequency meter or a counter by using the parallel printer port. Eliminates the 16-bit time-day counter through the use of an external time base that can be conveniently set depending on the desired frequency range. (JRH)

  13. Teaching Mathematics in the PC Lab--The Students' Viewpoints

    ERIC Educational Resources Information Center

    Schmidt, Karsten; Kohler, Anke

    2013-01-01

    The Matrix Algebra portion of the intermediate mathematics course at the Schmalkalden University Faculty of Business and Economics has been moved from a traditional classroom setting to a technology-based setting in the PC lab. A Computer Algebra System license was acquired that also allows its use on the students' own PCs. A survey was carried…

  14. A PC-based Workstation for Robotic Discectomy

    NASA Technical Reports Server (NTRS)

    Casadei, C.; Fiorini, P.; Martelli, S.; Montanari, M.; Morri, A.

    1998-01-01

    Ths paper describes a PC-based controller for robot-assisted minimally invasive surgery. The development is motivated by the need of reducing the exposure of operating room personnel to X-rays during surgical procedures such as percutanrous discectomy.

  15. An Apple for Your IBM PC--The Quadlink Board.

    ERIC Educational Resources Information Center

    Owen, G. Scott

    1984-01-01

    Describes nature and installation of the QUADLINK board which allows Apple software to be run on IBM PC microcomputers. Although programs tested ran without problems, users should test their own programs since there are some copy protection schemes that can baffle the board. (JN)

  16. What's Under Your PC's 'Hood': A Primer for Today's Machines.

    ERIC Educational Resources Information Center

    Chen, Li; Mills, Joyce White

    2002-01-01

    Explains how to decide what type of computer to buy for a small academic library, based on software requirements, staff and patron use, and PC technology. Topics include the central processing unit; types of memory; storage devices; ports; multimedia capabilities; modems; operating systems; and monitors. (LRW)

  17. Electronic States of IC60BA and PC71BM

    NASA Astrophysics Data System (ADS)

    Sheng, Chun-Qi; Wang, Peng; Shen, Ying; Li, Wen-Jie; Zhang, Wen-Hua; Zhu, Jun-Fa; Lai, Guo-Qiao; Li, Hong-Nian

    2013-11-01

    We investigate the electronic states of IC60BA and PC71BM using first-principles calculations and photoelectron spectroscopy (PES) measurements. The energy level structures for all possible isomers are reported and compared with those of C60, C70 and PC61BM. The attachment of the side chains can raise the LUMO energies and decrease the HOMO-LUMO gaps, and thus helps to increase the power-conversion efficiency of bulk heterojunction solar cells. In the PES studies, we prepared IC60BA and PC71BM films on Si:H(111) substrates to construct adsorbate/substrate interfaces describable with the integer charge-transfer (ICT) model. Successful measurements then revealed that one of the most important material properties for an electron acceptor, the energy of the negative integer charge-transfer state (EICT-), is 4.31 eV below the vacuum level for PC71BM. The EICT- of IC60BA is smaller than 4.14 eV.

  18. Wf/pc Cycle 2 Calib: Single Chip UV Calibration

    NASA Astrophysics Data System (ADS)

    MacKenty, John

    1991-07-01

    THIS PROGRAM CALIBRATES THE QE OF THE WFC AND PC IN THE ULTRAVIOLET (F194W, F230W, AND F284W). This calibration is done using exposures of a UV flux standard star. This program is intended for use only following a UV decontamination.

  19. Wf/pc Cycle 1 Calib: 4-CHIP UV Calibration

    NASA Astrophysics Data System (ADS)

    MacKenty, John

    1990-12-01

    THIS PROGRAM CALIBRATES THE QE OF THE WFC AND PC IN THE ULTRAVIOLET (F194W, F230W, AND F284W). This calibration is done for each CCD detector using exposures of a UV flux standard star. This program is intended for use only during a UV campaign.

  20. 39 CFR 501.16 - PC postage payment methodology.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... examination of its PC Postage system and any other applications and technology infrastructure that may have a... organization uses another service organization (sub-service provider), Postal Service Management should... contribution of the sub-service organization's processes and controls in the achievement of the Postal...

  1. 39 CFR 501.16 - PC postage payment methodology.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... examination of its PC Postage system and any other applications and technology infrastructure that may have a... organization uses another service organization (sub-service provider), Postal Service Management should... contribution of the sub-service organization's processes and controls in the achievement of the Postal...

  2. 39 CFR 501.16 - PC postage payment methodology.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... examination of its PC Postage system and any other applications and technology infrastructure that may have a... uses another service organization (sub-service provider), Postal Service management should consider the... of the sub-service organization's processes and controls in the achievement of the Postal...

  3. Multi-Purpose, Multi-Reservoir Simulation on a PC

    DTIC Science & Technology

    1988-08-01

    The methodology and difficulties in converting a large, general purpose, mainframe, batch oriented computer program (for reservoir simulation ) to...work effectively in the PC environment are described in this paper. A brief overview of the present capabilities of the general purpose reservoir ... simulation program (HEC-5), that works on mainframe and MS DOS compatible computers, is also presented. Keywords: Simulation, Personal computer, Reservior

  4. PC-CUBE: A Personal Computer Based Hypercube

    NASA Technical Reports Server (NTRS)

    Ho, Alex; Fox, Geoffrey; Walker, David; Snyder, Scott; Chang, Douglas; Chen, Stanley; Breaden, Matt; Cole, Terry

    1988-01-01

    PC-CUBE is an ensemble of IBM PCs or close compatibles connected in the hypercube topology with ordinary computer cables. Communication occurs at the rate of 115.2 K-band via the RS-232 serial links. Available for PC-CUBE is the Crystalline Operating System III (CrOS III), Mercury Operating System, CUBIX and PLOTIX which are parallel I/O and graphics libraries. A CrOS performance monitor was developed to facilitate the measurement of communication and computation time of a program and their effects on performance. Also available are CXLISP, a parallel version of the XLISP interpreter; GRAFIX, some graphics routines for the EGA and CGA; and a general execution profiler for determining execution time spent by program subroutines. PC-CUBE provides a programming environment similar to all hypercube systems running CrOS III, Mercury and CUBIX. In addition, every node (personal computer) has its own graphics display monitor and storage devices. These allow data to be displayed or stored at every processor, which has much instructional value and enables easier debugging of applications. Some application programs which are taken from the book Solving Problems on Concurrent Processors (Fox 88) were implemented with graphics enhancement on PC-CUBE. The applications range from solving the Mandelbrot set, Laplace equation, wave equation, long range force interaction, to WaTor, an ecological simulation.

  5. Wf/pc Cycle 1 CALIBRATION:UV Flats 1

    NASA Astrophysics Data System (ADS)

    MacKenty, John

    1990-12-01

    This program takes "uniform illuminated" pictures of the earth to obtain the "instrument signature" for flat field corrections for the UV filters. The camera - filter combinations done in this proposal are: WFC: F194W, F230W, F284W PC: F194W, F230W, F284W This calibration should be part of each UV campaign.

  6. Deciding when It's Time to Buy a New PC

    ERIC Educational Resources Information Center

    Goldsborough, Reid

    2004-01-01

    How to best decide when it's time to replace your PC, whether at home or at work, is always tricky. Spending on computers can make you more productive, but it's money you otherwise cannot spend, invest or save, and faster systems always await you in the future. What is clear is that the computer industry really wants you to buy, and the computer…

  7. 36. View of preset counter (PC) console and tracking console ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    36. View of preset counter (PC) console and tracking console on right, located in MWOC facility in transmitter building no. 102. - Clear Air Force Station, Ballistic Missile Early Warning System Site II, One mile west of mile marker 293.5 on Parks Highway, 5 miles southwest of Anderson, Anderson, Denali Borough, AK

  8. PC vs. Mac--Which Way Should You Go?

    ERIC Educational Resources Information Center

    Wodarz, Nan

    1997-01-01

    Outlines the factors in hardware, software, and administration to consider in developing specifications for choosing a computer operating system. Compares Microsoft Windows 95/NT that runs on PC/Intel-based systems and System 7.5 that runs on the Apple-based systems. Lists reasons why the Microsoft platform clearly stands above the Apple platform.…

  9. Privacy Impact Assessment for the PC Label System

    EPA Pesticide Factsheets

    The PC Label System collects contact information for individuals with an interest in EPA's Region 1. Learn how this data will be collected in the system, how it will be used, access to the data, the purpose of data collection, and record retention policies

  10. PLK1 inhibition enhances temozolomide efficacy in IDH1 mutant gliomas.

    PubMed

    Koncar, Robert F; Chu, Zhengtao; Romick-Rosendale, Lindsey E; Wells, Susanne I; Chan, Timothy A; Qi, Xiaoyang; Bahassi, El Mustapha

    2017-02-28

    Despite multimodal therapy with radiation and the DNA alkylating agent temozolomide (TMZ), malignant gliomas remain incurable. Up to 90% of grades II-III gliomas contain a single mutant isocitrate dehydrogenase 1 (IDH1) allele. IDH1 mutant-mediated transformation is associated with TMZ resistance; however, there is no clinically available means of sensitizing IDH1 mutant tumors to TMZ. In this study we sought to identify a targetable mechanism of TMZ resistance in IDH1 mutant tumors to enhance TMZ efficacy. IDH1 mutant astrocytes rapidly bypassed the G2 checkpoint with unrepaired DNA damage following TMZ treatment. Checkpoint adaptation was accompanied by PLK1 activation and IDH1 mutant astrocytes were more sensitive to treatment with BI2536 and TMZ in combination (<20% clonogenic survival) than either TMZ (~60%) or BI2536 (~75%) as single agents. In vivo, TMZ or BI2536 alone had little effect on tumor size. Combination treatment caused marked tumor shrinkage in all mice and complete tumor regression in 5 of 8 mice. Mutant IDH1 promotes checkpoint adaptation which can be exploited therapeutically with the combination of TMZ and a PLK1 inhibitor, indicating PLK1 inhibitors may be clinically valuable in the treatment of IDH1 mutant gliomas.

  11. Temperature-sensitive yeast mutants defective in mitochondrial inheritance.

    PubMed

    McConnell, S J; Stewart, L C; Talin, A; Yaffe, M P

    1990-09-01

    The distribution of mitochondria to daughter cells is an essential feature of mitotic cell growth, yet the molecular mechanisms facilitating this mitochondrial inheritance are unknown. We have isolated mutants of Saccharomyces cerevisiae that are temperature-sensitive for the transfer of mitochondria into a growing bud. Two of these mutants contain single, recessive, nuclear mutations, mdm1 and mdm2, that cause temperature-sensitive growth and aberrant mitochondrial distribution at the nonpermissive temperature. The absence of mitochondria from the buds of mutant cells was confirmed by indirect immunofluorescence microscopy and by transmission electron microscopy. The mdm1 lesion also retards nuclear division and prevents the transfer of nuclei into the buds. Cells containing the mdm2 mutation grown at the nonpermissive temperature sequentially form multiple buds, each receiving a nucleus but no mitochondria. Neither mdm1 or mdm2 affects the transfer of vacuolar material into the buds or causes apparent changes in the tubulin- or actin-based cytoskeletons. The mdm1 and mdm2 mutations are cell-cycle specific, displaying an execution point in late G1 or early S phase.

  12. Morphological Characterization of a New and Easily Recognizable Nuclear Male Sterile Mutant of Sorghum (Sorghum bicolor)

    PubMed Central

    Xin, Zhanguo; Huang, Jian; Smith, Ashley R.; Chen, Junping; Burke, John; Sattler, Scott E.

    2017-01-01

    Sorghum (Sorghum bicolor L. Moench) is one of the most important grain crops in the world. The nuclear male sterility (NMS) trait, which is caused by mutations on the nuclear gene, is valuable for hybrid breeding and genetic studies. Several NMS mutants have been reported previously, but none of them were well characterized. Here, we present our detailed morphological characterization of a new and easily recognizable NMS sorghum mutant male sterile 8 (ms8) isolated from an elite inbred BTx623 mutagenized by ethyl methane sulfonate (EMS). Our results show that the ms8 mutant phenotype was caused by a mutation on a single recessive nuclear gene that is different from all available NMS loci reported in sorghum. In fertile sorghum plants, yellow anthers appeared first during anthesis, while in the ms8 mutant, white hairy stigma emerged first and only small white anthers were observed, making ms8 plants easily recognizable when flowering. The ovary development and seed production after manual pollination are normal in the ms8 mutant, indicating it is female fertile and male sterile only. We found that ms8 anthers did not produce pollen grains. Further analysis revealed that ms8 anthers were defective in tapetum development, which led to the arrest of pollen formation. As a stable male sterile mutant across different environments, greenhouses, and fields in different locations, the ms8 mutant could be a useful breeding tool. Moreover, ms8 might be an important for elucidating male gametophyte development in sorghum and other plants. PMID:28052078

  13. Mutant CAG Repeats Effectively Targeted by RNA Interference in SCA7 Cells

    PubMed Central

    Fiszer, Agnieszka; Wroblewska, Joanna P.; Nowak, Bartosz M.; Krzyzosiak, Wlodzimierz J.

    2016-01-01

    Spinocerebellar ataxia type 7 (SCA7) is a human neurodegenerative polyglutamine (polyQ) disease caused by a CAG repeat expansion in the open reading frame of the ATXN7 gene. The allele-selective silencing of mutant transcripts using a repeat-targeting strategy has previously been used for several polyQ diseases. Herein, we demonstrate that the selective targeting of a repeat tract in a mutant ATXN7 transcript by RNA interference is a feasible approach and results in an efficient decrease of mutant ataxin-7 protein in patient-derived cells. Oligonucleotides (ONs) containing specific base substitutions cause the downregulation of the ATXN7 mutant allele together with the upregulation of its normal allele. The A2 ON shows high allele selectivity at a broad range of concentrations and also restores UCHL1 expression, which is downregulated in SCA7. PMID:27999335

  14. Induction of cytoprotective autophagy in PC-12 cells by cadmium

    SciTech Connect

    Wang, Qiwen; Zhu, Jiaqiao; Zhang, Kangbao; Jiang, Chenyang; Wang, Yi; Yuan, Yan; Bian, Jianchun; Liu, Xuezhong; Gu, Jianhong; Liu, Zongping

    2013-08-16

    Highlights: •Cadmium can promote early upregulation of autophagy in PC-12 cells. •Autophagy precedes apoptosis in cadmium-treated PC-12 cells. •Cadmium-induced autophagy is cytoprotective in PC-12 cells. •Class III PI3K/beclin-1/Bcl-2 signaling pathway plays a positive role in cadmium-triggered autophagy. -- Abstract: Laboratory data have demonstrated that cadmium (Cd) may induce neuronal apoptosis. However, little is known about the role of autophagy in neurons. In this study, cell viability decreased in a dose- and time-dependent manner after treatment with Cd in PC-12 cells. As cells were exposed to Cd, the levels of LC3-II proteins became elevated, specific punctate distribution of endogenous LC3-II increased, and numerous autophagosomes appeared, which suggest that Cd induced a high level of autophagy. In the late stages of autophagy, an increase in the apoptosis ratio was observed. Likewise, pre-treatment with chloroquine (an autophagic inhibitor) and rapamycin (an autophagic inducer) resulted in an increased and decreased percentage of apoptosis in contrast to other Cd-treated groups, respectively. The results indicate that autophagy delayed apoptosis in Cd-treated PC-12 cells. Furthermore, co-treatment of cells with chloroquine reduced autophagy and cell activity. However, rapamycin had an opposite effect on autophagy and cell activity. Moreover, class III PI3 K/beclin-1/Bcl-2 signaling pathways served a function in Cd-induced autophagy. The findings suggest that Cd can induce cytoprotective autophagy by activating class III PI3 K/beclin-1/Bcl-2 signaling pathways. In sum, this study strongly suggests that autophagy may serve a positive function in the reduction of Cd-induced cytotoxicity.

  15. IPEG- IMPROVED PRICE ESTIMATION GUIDELINES (IBM PC VERSION)

    NASA Technical Reports Server (NTRS)

    Aster, R. W.

    1994-01-01

    The Improved Price Estimation Guidelines, IPEG, program provides a simple yet accurate estimate of the price of a manufactured product. IPEG facilitates sensitivity studies of price estimates at considerably less expense than would be incurred by using the Standard Assembly-line Manufacturing Industry Simulation, SAMIS, program (COSMIC program NPO-16032). A difference of less than one percent between the IPEG and SAMIS price estimates has been observed with realistic test cases. However, the IPEG simplification of SAMIS allows the analyst with limited time and computing resources to perform a greater number of sensitivity studies than with SAMIS. Although IPEG was developed for the photovoltaics industry, it is readily adaptable to any standard assembly line type of manufacturing industry. IPEG estimates the annual production price per unit. The input data includes cost of equipment, space, labor, materials, supplies, and utilities. Production on an industry wide basis or a process wide basis can be simulated. Once the IPEG input file is prepared, the original price is estimated and sensitivity studies may be performed. The IPEG user selects a sensitivity variable and a set of values. IPEG will compute a price estimate and a variety of other cost parameters for every specified value of the sensitivity variable. IPEG is designed as an interactive system and prompts the user for all required information and offers a variety of output options. The IPEG/PC program is written in TURBO PASCAL for interactive execution on an IBM PC computer under DOS 2.0 or above with at least 64K of memory. The IBM PC color display and color graphics adapter are needed to use the plotting capabilities in IPEG/PC. IPEG/PC was developed in 1984. The original IPEG program is written in SIMSCRIPT II.5 for interactive execution and has been implemented on an IBM 370 series computer with a central memory requirement of approximately 300K of 8 bit bytes. The original IPEG was developed in 1980.

  16. crl mutants of Saccharomyces cerevisiae resemble both mutants affecting general control of amino acid biosynthesis and omnipotent translational suppressor mutants.

    PubMed

    McCusker, J H; Haber, J E

    1988-06-01

    Cyocloheximide resistant lethal (crl) mutants of Saccharomyces cerevisiae, defining 22 unlinked complementation groups, are unable to grow at 37 degrees. They are also highly pleiotropic at their permissive temperature of 25 degrees. The mutants are all unable to arrest at the G1 stage of the cell cycle when grown to stationary phase or when starved for a single amino acid, though they do arrest at G1 when deprived of all nitrogen. The crl mutants are also hypersensitive to various amino acid analogs and to 3-aminotriazole. These mutants also "tighten" leaky auxotrophic mutations that permit wild-type cells to grow in the absence of the appropriate amino acid. All of these phenotypes are also exhibited by gcn mutants affecting general control of amino acid biosynthesis. In addition, the crl mutants are all hypersensitive to hygromycin B, an aminoglycoside antibiotic that stimulates translational misreading. The crl mutations also suppress one nonsense mutation which is phenotypically suppressed by hygromycin B. Many crl mutants are also osmotically sensitive. These are phenotypes which the crl mutations have in common with previously isolated omnipotent suppressors. We suggest that the the crl mutations all affect the fidelity of protein translation.

  17. Nucleophosmin leukemogenic mutant activates Wnt signaling during zebrafish development

    PubMed Central

    Barbieri, Elisa; Deflorian, Gianluca; Pezzimenti, Federica; Valli, Debora; Saia, Marco; Meani, Natalia

    2016-01-01

    Nucleophosmin (NPM1) is a ubiquitous multifunctional phosphoprotein with both oncogenic and tumor suppressor functions. Mutations of the NPM1 gene are the most frequent genetic alterations in acute myeloid leukemia (AML) and result in the expression of a mutant protein with aberrant cytoplasmic localization, NPMc+. Although NPMc+ causes myeloproliferation and AML in animal models, its mechanism of action remains largely unknown. Here we report that NPMc+ activates canonical Wnt signaling during the early phases of zebrafish development and determines a Wnt-dependent increase in the number of progenitor cells during primitive hematopoiesis. Coherently, the canonical Wnt pathway is active in AML blasts bearing NPMc+ and depletion of the mutant protein in the patient derived OCI-AML3 cell line leads to a decrease in the levels of active β-catenin and of Wnt target genes. Our results reveal a novel function of NPMc+ and provide insight into the molecular pathogenesis of AML bearing NPM1 mutations. PMID:27486814

  18. Surface-type nonvolatile electric memory elements based on organic-on-organic CuPc-H2Pc heterojunction

    NASA Astrophysics Data System (ADS)

    Khasan, S. Karimov; Zubair, Ahmad; Farid, Touati; Mahroof-Tahir, M.; M. Muqeet, Rehman; S. Zameer, Abbas

    2015-11-01

    A novel surface-type nonvolatile electric memory elements based on organic semiconductors CuPc and H2Pc are fabricated by vacuum deposition of the CuPc and H2Pc films on preliminary deposited metallic (Ag and Cu) electrodes. The gap between Ag and Cu electrodes is 30-40 μm. For the current-voltage (I-V) characteristics the memory effect, switching effect, and negative differential resistance regions are observed. The switching mechanism is attributed to the electric-field-induced charge transfer. As a result the device switches from a low to a high-conductivity state and then back to a low conductivity state if the opposite polarity voltage is applied. The ratio of resistance at the high resistance state to that at the low resistance state is equal to 120-150. Under the switching condition, the electric current increases ˜ 80-100 times. A comparison between the forward and reverse I-V characteristics shows the presence of rectifying behavior. Project supported by the GIK Institute of Engineering Science and Technology, Pakistan and Physical Technical Institute of Academy of Sciences of Tajikistan.

  19. Chromosomal assignment of the genes for proprotein convertases PC4, PC5, and PACE 4 in mouse and human.

    PubMed

    Mbikay, M; Seidah, N G; Chrétien, M; Simpson, E M

    1995-03-01

    The genes for three subtilisin/kexin-like proprotein convertases, PC4, PC5, and PACE4, were mapped in the mouse by RFLP analysis of a DNA panel from a (C57BL/6JEi x SPRET/Ei)F1 x SPRET/Ei backcross. The chromosomal locations of the human homologs were determined by Southern blot analysis of a DNA panel from human-rodent somatic cell hybrids, most of which contained a single human chromosome each. The gene for PC4 (Pcsk4 locus) mapped to mouse chromosome 10, close to the Adn (adipsin, a serine protease) locus and near the Amh (anti-müllerian hormone) locus; in human, the gene was localized to chromosome 19. The gene for PC5 (Pcsk5 locus) mapped to mouse chromosome 19 close to the Lpc1 (lipocortin-1) locus and, in human, was localized to chromosome 9. The gene for PACE4 (Pcsk6 locus) mapped to mouse chromosome 7, at a distance of 13 cM from the Pcsk3 locus, which specifies furin, another member of this family of enzymes previously mapped to this chromosome. This is in concordance with the known close proximity of these two loci in the homologous region on human chromosome 15q25-qter. Pcsk3 and Pcsk6 mapped to a region of mouse chromosome 7 that has been associated cytogenetically with postnatal lethality in maternal disomy, suggesting that these genes might be candidates for imprinting.

  20. Isolation and characterization of a low phytic acid rice mutant reveals a mutation in the rice orthologue of maize mik.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Using a forward genetics approach, we isolated two independent low phytic acid (lpa) rice mutants, N15-186 and N15-375. Both mutants are caused by single gene, recessive non-lethal mutations which result in approximately 75% (N15-186) and 43% (N15-375) reductions in seed phytic acid (inositol hexaki...

  1. METHANE de-NOX for Utility PC Boilers

    SciTech Connect

    Bruce Bryan; Serguei Nester; Joseph Rabovitser; Stan Wohadlo

    2005-09-30

    The overall project objective is the development and validation of an innovative combustion system, based on a novel coal preheating concept prior to combustion, that can reduce NO{sub x} emissions to 0.15 lb/million Btu or less on utility pulverized coal (PC) boilers. This NO{sub x} reduction should be achieved without loss of boiler efficiency or operating stability, and at more than 25% lower levelized cost than state-of-the-art SCR technology. A further objective is to ready technology for full-scale commercial deployment to meet the market demand for NO{sub x} reduction technologies. Over half of the electric power generated in the U.S. is produced by coal combustion, and more than 80% of these units utilize PC combustion technology. Conventional measures for NOx reduction in PC combustion processes rely on combustion and post-combustion modifications. A variety of combustion-based NO{sub x} reduction technologies are in use today, including low-NO{sub x} burners (LNBs), flue gas recirculation (FGR), air staging, and natural gas or other fuel reburning. Selective non-catalytic reduction (SNCR) and selective catalytic reduction (SCR) are post-combustion techniques. NO{sub x} reduction effectiveness from these technologies ranges from 30 to 60% and up to 90-93% for SCR. Typically, older wall-fired PC burner units produce NO{sub x} emissions in the range of 0.8-1.6 lb/million Btu. Low-NO{sub x} burner systems, using combinations of fuel staging within the burner and air staging by introduction of overfire air in the boiler, can reduce NO{sub x} emissions by 50-60%. This approach alone is not sufficient to meet the desired 0.15 lb/million Btu NO{sub x} standard with a range of coals and boiler loads. Furthermore, the heavy reliance on overfire air can lead to increased slagging and corrosion in furnaces, particularly with higher-sulfur coals, when LNBs are operated at sub-stoichiometric conditions to reduce fuel-derived NOx in the flame. Therefore, it is desirable

  2. The secretory proprotein convertases furin, PC5, and PC7 activate VEGF-C to induce tumorigenesis

    PubMed Central

    Siegfried, Geraldine; Basak, Ajoy; Cromlish, James A.; Benjannet, Suzanne; Marcinkiewicz, Jadwiga; Chrétien, Michel; Seidah, Nabil G.; Khatib, Abdel-Majid

    2003-01-01

    The secretory factor VEGF-C has been directly implicated in various physiological processes during embryogenesis and human cancers. However, the importance of the conversion of its precursor proVEGF-C to mature VEGF-C in tumorigenesis, and vessel formation and the identity of the protease(s) that regulate these processes is/are not known. The intracellular processing of proVEGF-C that occurs within the dibasic motif HSIIRR227SL suggests the involvement of the proprotein convertases (PCs) in this process. In addition, furin and VEGF-C were found to be coordinately expressed in adult mouse tissues. Cotransfection of the furin-deficient colon carcinoma cell line LoVo with proVEGF-C and different PC members revealed that furin, PC5, and PC7 are candidate VEGF-C convertases. This finding is consistent with the in vitro digestions of an internally quenched synthetic fluorogenic peptide mimicking the cleavage site of proVEGF-C (220Q-VHSIIRR↓SLP230). The processing of proVEGF-C is blocked by the inhibitory prosegments of furin, PC5, and PACE4, as well as by furin-motif variants of α2-macroglobulin and α1-antitrypsin. Subcutaneous injection of CHO cells stably expressing VEGF-C into nude mice enhanced angiogenesis and lymphangiogenesis, but not tumor growth. In contrast, expression of proVEGF-C obtained following mutation of the cleavage site (HSIIRR227SL to HSIISS227SL) inhibits angiogenesis and lymphangiogenesis as well as tumor growth. Our findings demonstrate the processing of proVEGF-C by PCs and highlight the potential use of PC inhibitors as agents for inhibiting malignancies induced by VEGF-C. PMID:12782675

  3. Establishment and characterization of methylmercury-resistant PC12 cell line.

    PubMed Central

    Miura, K; Clarkson, T W; Ikeda, K; Naganuma, A; Imura, N

    1994-01-01

    Methylmercury (MeHg)-resistant sublines of rat pheochromocytoma (PC12) cells were isolated by repeated exposure to stepwise increased concentrations of MeHg. One of the sublines (PC12/TM) showed an 8- to 10-fold increase in resistance to MeHg compared with parent PC12 cells on the basis of the concentration required for 50% inhibition (IC50) of growth. PC12/TM cells accumulated smaller amounts of MeHg than parent PC12 cells. This reduction in MeHg accumulation in PC12/TM cells resulted from slow uptake and rapid efflux. The intracellular glutathione (GSH) level in PC12/TM cells was four times higher than that of PC12 cells. Pretreatment of PC12/TM cells with buthionine sulfoximine, which decreased the GSH level to that of the parent PC12 cells, increased the sensitivity of PC12/TM cells to MeHg. A close correlation between the MeHg accumulation and MeHg sensitivity was found among seven sublines of PC12 cells and parent PC12 cell line. The GSH level in PC12 sublines was also correlated with their sensitivity to MeHg. PMID:7843125

  4. Compromised mutant EFEMP1 secretion associated with macular dystrophy remedied by proteostasis network alteration.

    PubMed

    Hulleman, John D; Kaushal, Shalesh; Balch, William E; Kelly, Jeffery W

    2011-12-01

    An Arg345Trp (R345W) mutation in epidermal growth factor-containing, fibulin-like extracellular matrix protein 1 (EFEMP1) causes its inefficient secretion and the macular dystrophy malattia leventinese/Doyne honeycomb retinal dystrophy (ML/DHRD). To understand the influence of the protein homeostasis (or proteostasis) network in rescuing mutant EFEMP1 misfolding and inefficient secretion linked to ML/DHRD, we developed a convenient and sensitive cell-based luminescence assay to monitor secretion versus intracellular accumulation. Fusing EFEMP1 to Gaussia luciferase faithfully recapitulates mutant EFEMP1 secretion defects observed previously using more cumbersome methodology. To understand what governs mutant intracellular retention, we generated a series of R345 mutants. These mutants revealed that aromatic residue substitutions (i.e., Trp, Tyr, and Phe) at position 345 cause significant EFEMP1 secretion deficiencies. These secretion defects appear to be caused, in part, by reduced native disulfide bonding in domain 6 harboring the 345 position. Finally, we demonstrate that mutant EFEMP1 secretion and proper disulfide formation are enhanced by adaptation of the cellular environment by a reduced growth temperature and/or translational attenuation. This study highlights the mechanisms underlying the inefficient secretion of R345W EFEMP1 and demonstrates that alteration of the proteostasis network may provide a strategy to alleviate or delay the onset of this macular dystrophy.

  5. Compromised mutant EFEMP1 secretion associated with macular dystrophy remedied by proteostasis network alteration

    PubMed Central

    Hulleman, John D.; Kaushal, Shalesh; Balch, William E.; Kelly, Jeffery W.

    2011-01-01

    An Arg345Trp (R345W) mutation in epidermal growth factor–containing, fibulin-like extracellular matrix protein 1 (EFEMP1) causes its inefficient secretion and the macular dystrophy malattia leventinese/Doyne honeycomb retinal dystrophy (ML/DHRD). To understand the influence of the protein homeostasis (or proteostasis) network in rescuing mutant EFEMP1 misfolding and inefficient secretion linked to ML/DHRD, we developed a convenient and sensitive cell-based luminescence assay to monitor secretion versus intracellular accumulation. Fusing EFEMP1 to Gaussia luciferase faithfully recapitulates mutant EFEMP1 secretion defects observed previously using more cumbersome methodology. To understand what governs mutant intracellular retention, we generated a series of R345 mutants. These mutants revealed that aromatic residue substitutions (i.e., Trp, Tyr, and Phe) at position 345 cause significant EFEMP1 secretion deficiencies. These secretion defects appear to be caused, in part, by reduced native disulfide bonding in domain 6 harboring the 345 position. Finally, we demonstrate that mutant EFEMP1 secretion and proper disulfide formation are enhanced by adaptation of the cellular environment by a reduced growth temperature and/or translational attenuation. This study highlights the mechanisms underlying the inefficient secretion of R345W EFEMP1 and demonstrates that alteration of the proteostasis network may provide a strategy to alleviate or delay the onset of this macular dystrophy. PMID:22031286

  6. Determination of the Mutant Prevention Concentration and the Mutant Selection Window of Topical Antimicrobial Agents against Propionibacterium acnes.

    PubMed

    Nakase, Keisuke; Nakaminami, Hidemasa; Toda, Yuta; Noguchi, Norihisa

    2017-01-01

    Determination of the mutant prevention concentration (MPC) and the mutant selection window (MSW) of antimicrobial agents used to treat pathogenic bacteria is important in order to apply effective antimicrobial therapies. Here, we determined the MPCs of the major topical antimicrobial agents against Propionibacterium acnes and Staphylococcus aureus which cause skin infections and compared their MSWs. Among the MPCs of nadifloxacin and clindamycin, the clindamycin MPC was determined to be the lowest against P. acnes. In contrast, the nadifloxacin MPC was the lowest against S. aureus. Calculations based on the minimum inhibitory concentrations and MPCs showed that clindamycin has the lowest MSW against both P. acnes and S. aureus. Nadifloxacin MSWs were 4-fold higher against P. acnes than against S. aureus. It is more likely for P. acnes to acquire resistance to fluoroquinolones than S. aureus. Therefore, topical application of clindamycin contributes very little to the emergence of resistant P. acnes and S. aureus strains.

  7. A role for SIRT1 in cell growth and chemoresistance in prostate cancer PC3 and DU145 cells

    SciTech Connect

    Kojima, Keitaro; Ohhashi, Riyako; Fujita, Yasunori; Hamada, Nanako; Akao, Yukihiro; Nozawa, Yoshinori; Deguchi, Takashi; Ito, Masafumi

    2008-08-29

    SIRT1, which belongs to the family of type III histone deacetylase, is implicated in diverse cellular processes. We have determined the expression levels of SIRT1 in human prostate cancer cell lines and have examined the roles of SIRT1 in cell growth and chemoresistance. SIRT1 expression was markedly up-regulated in androgen-refractory PC3 and DU145 cells compared with androgen-sensitive LNCaP cells and its expression level was correlated with cell growth in PC3 cells. Treatment with a SIRT1 inhibitor, sirtinol, inhibited cell growth and increased sensitivity to camptothecin and cisplatin. Silencing of SIRT1 expression by siRNA also suppressed cell proliferation and reduced camptothecin resistance in PC3 cells, mimicking the chemosensitizing effect caused by sirtinol. Also in DU145 cells, sirtinol treatment enhanced sensitivity to camptothecin and cisplatin. These results suggest that up-regulation of SIRT1 expression may play an important role in promoting cell growth and chemoresistance in androgen-refractory PC3 and DU145 cells.

  8. Salmonella typhimurium mutants lacking NAD pyrophosphatase.

    PubMed Central

    Park, U E; Roth, J R; Olivera, B M

    1988-01-01

    NAD can serve as both a purine and a pyridine source for Salmonella typhimurium. Exogenous NAD is rapidly broken down into nicotinamide mononucleotide and AMP by an NAD pyrophosphatase, the first step in the pathway for the assimilation of exogenous NAD. We isolated and characterized mutants of S. typhimurium lacking NAD pyrophosphatase activity; such mutants were identified by their failure to use exogenous NAD as a purine source. These mutants carry mutations that map at a new locus, designated pnuE, between 86 and 87 min on the Salmonella chromosome. PMID:2841298

  9. Autosomal mutants of proton-exposed kidney cells display frequent loss of heterozygosity on nonselected chromosomes.

    PubMed

    Grygoryev, Dmytro; Dan, Cristian; Gauny, Stacey; Eckelmann, Bradley; Ohlrich, Anna P; Connolly, Marissa; Lasarev, Michael; Grossi, Gianfranco; Kronenberg, Amy; Turker, Mitchell S

    2014-05-01

    High-energy protons found in the space environment can induce mutations and cancer, which are inextricably linked. We hypothesized that some mutants isolated from proton-exposed kidneys arose through a genome-wide incident that causes loss of heterozygosity (LOH)-generating mutations on multiple chromosomes (termed here genomic LOH). To test this hypothesis, we examined 11 pairs of nonselected chromosomes for LOH events in mutant cells isolated from the kidneys of mice exposed to 4 or 5 Gy of 1 GeV protons. The mutant kidney cells were selected for loss of expression of the chromosome 8-encoded Aprt gene. Genomic LOH events were also assessed in Aprt mutants isolated from isogenic cultured kidney epithelial cells exposed to 5 Gy of protons in vitro. Control groups were spontaneous Aprt mutants and clones isolated without selection from the proton-exposed kidneys or cultures. The in vivo results showed significant increases in genomic LOH events in the Aprt mutants from proton-exposed kidneys when compared with spontaneous Aprt mutants and when compared with nonmutant (i.e., nonselected) clones from the proton-exposed kidneys. A bias for LOH events affecting chromosome 14 was observed in the proton-induced Aprt mutants, though LOH for this chromosome did not confer increased radiation resistance. Genomic LOH events were observed in Aprt mutants isolated from proton-exposed cultured kidney cells; however the incidence was fivefold lower than in Aprt mutants isolated from exposed intact kidneys, suggesting a more permissive environment in the intact organ and/or the evolution of kidney clones prior to their isolation from the tissue. We conclude that proton exposure creates a subset of viable cells with LOH events on multiple chromosomes, that these cells form and persist in vivo, and that they can be isolated from an intact tissue by selection for a mutation on a single chromosome.

  10. Suppression of pleiotropic phenotypes of a Burkholderia multivorans fur mutant by oxyR mutation.

    PubMed

    Kimura, Akane; Yuhara, Satoshi; Ohtsubo, Yoshiyuki; Nagata, Yuji; Tsuda, Masataka

    2012-05-01

    Fur (ferric uptake regulator) is an iron-responsive transcriptional regulator in many bacterial species, and the fur mutant of Burkholderia multivorans ATCC 17616 exhibits pleiotropic phenotypes, such as an inability to efficiently use several carbon sources, as well as high sensitivity to hydrogen peroxide (H(2)O(2)), paraquat (a superoxide-producing compound) and nitric oxide (NO). To gain more insight into the pleiotropic role of the Fur protein of ATCC 17616, spontaneous suppressor mutants of the ATCC 17616 fur mutant that restored tolerance to NO were isolated and characterized in this study. The microarray-based comparative genomic analysis and subsequent sequencing analysis indicated that such suppressor mutants had a 2 bp deletion in the oxyR gene, whose orthologues encode H(2)O(2)-responsive transcriptional regulators in other bacterial species. The suppressor mutants and the reconstructed fur-oxyR double-deletion mutant showed indistinguishable phenotypes in that they were all (i) more resistant than the fur mutant to H(2)O(2), superoxide, NO and streptonigrin (an iron-activated antibiotic) and (ii) able to use carbon sources that cannot efficiently support the growth of the fur mutant. These results clearly indicate that the oxyR mutation suppressed the pleiotropic effect of the B. multivorans fur mutant. The fur-oxyR double mutants were found to overexpress the KatG (catalase/peroxidase) and AhpC1 and AhpD (alkyl hydroperoxide reductase subunits C and D) proteins, and their enzymic activities to remove reactive oxygen and nitrogen species were suggested to be responsible for the suppression of phenotypes caused by the fur mutation.

  11. Isolation and characterization of Saccharomyces cerevisiae mutants resistant to Calcofluor white.

    PubMed Central

    Roncero, C; Valdivieso, M H; Ribas, J C; Durán, A

    1988-01-01

    Calcofluor is a fluorochrome that exhibits antifungal activity and a high affinity for yeast cell wall chitin. We isolated Saccharomyces cerevisiae mutants resistant to Calcofluor. The resistance segregated in a Mendelian fashion and behaved as a recessive character in all the mutants analyzed. Five loci were defined by complementation analysis. The abnormally thick septa between mother and daughter cells caused by Calcofluor in wild-type cells were absent in the mutants. The Calcofluor-binding capacity, observed by fluorescence microscopy, in a S. cerevisiae wild-type cells during alpha-factor treatment was also absent in some mutants and reduced in others. Staining of cell walls with wheat germ agglutinin-fluorescein complex indicated that the chitin uniformly distributed over the whole cell wall in vegetative or in alpha-factor-treated cells was almost absent in three of the mutants and reduced in the two others. Cell wall analysis evidenced a five- to ninefold reduction in the amount of chitin in mutants compared with that in the wild-type strain. The total amounts of cell wall mannan and beta-glucan in wild-type and mutant strains were similar; however, the percentage of beta-glucan that remained insoluble after alkali extraction was considerably reduced in mutant cells. The susceptibilities of the mutants and the wild-type strains to a cell wall enzymic lytic complex were rather similar. The in vitro levels of chitin synthase 2 detected in all mutants were similar to that in the wild type. The significance of these results is discussed in connection with the mechanism of chitin synthesis and cell wall morphogenesis in S. cerevisiae. Images PMID:3280554

  12. Fused 3-Hydroxy-3-trifluoromethylpyrazoles Inhibit Mutant Huntingtin Toxicity

    PubMed Central

    2013-01-01

    Here, we describe the selection and optimization of a chemical series active in both a full-length and a fragment-based Huntington’s disease (HD) assay. Twenty-four thousand small molecules were screened in a phenotypic HD assay, identifying a series of compounds bearing a 3-hydroxy-3-trifluoromethylpyrazole moiety as able to revert the toxicity induced by full-length mutant Htt by up to 50%. A chemical exploration around the series led to the identification of compound 4f, which demonstrated to be active in a Htt171–82Q rat primary striatal neuron assay and a PC12-Exon-1 based assay. This compound was selected for testing in R6/2 mice, in which it was well-tolerated and showed a positive effect on body weight and a positive trend in preventing ventricular volume enlargment. These studies provide strong rationale for further testing the potential benefits of 3-hydroxy-3-trifluoromethylpyrazoles in treating HD. PMID:24900595

  13. Pancreatic Cancer-derived Exosomes Causes Paraneoplastic β-cell Dysfunction

    PubMed Central

    Javeed, Naureen; Sagar, Gunisha; Dutta, Shamit K.; Smyrk, Thomas C.; Lau, Julie S.; Bhattacharya, Santanu; Truty, Mark; Petersen, Gloria M.; Kaufman, Randal J.; Chari, Suresh T.; Mukhopadhyay, Debabrata

    2014-01-01

    Background & Aims Pancreatic cancer (PC) frequently causes diabetes. We recently proposed adrenomedullin (AM) as a candidate mediator of pancreatic β-cell dysfunction in PC. How PC-derived AM reaches β-cells remote from the cancer to induce β-cell dysfunction is unknown. We tested a novel hypothesis that PC sheds AM-containing exosomes into circulation which are transported to β-cells and impair insulin secretion. Methods We characterized exosomes from conditioned media of PC-cell lines (n=5) and portal/peripheral venous blood of PC patients (n=20). Western blot analysis showed the presence of AM in PC-Exosomes. We determined the effect of AM-containing PC-Exosomes on insulin secretion from INS-1 β-cells and human islets, and showed how exosomes internalize into β-cells. We studied the interaction between β-cell AM receptors and AM present in PC-Exosomes. In addition, we studied the effect of AM on endoplasmic reticulum (ER) stress response genes and reactive oxygen/nitrogen species generation in β-cells. Results Exosomes were found to be the predominant extracellular vesicles secreted by PC into culture media and human plasma. PC-Exosomes contained AM and CA19-9, readily entered β-cells through caveolin-mediated endocytosis or macropinocytosis, and inhibited insulin secretion. AM in PC-Exosomes interacted with its receptor on β-cells. AM receptor blockade abrogated the inhibitory effect of exosomes on insulin secretion. β-cells exposed to AM or PC-Exosomes showed upregulation of ER stress genes and increased reactive oxygen/nitrogen species. Conclusions Pancreatic cancer causes paraneoplastic β-cell dysfunction by shedding AM+/CA19-9+ exosomes into circulation that inhibit insulin secretion, likely through AM-induced ER stress and failure of the UPR. PMID:25355928

  14. Flexible missile autopilot design studies with PC-MATLAB/386

    NASA Technical Reports Server (NTRS)

    Ruth, Michael J.

    1989-01-01

    Development of a responsive, high-bandwidth missile autopilot for airframes which have structural modes of unusually low frequency presents a challenging design task. Such systems are viable candidates for modern, state-space control design methods. The PC-MATLAB interactive software package provides an environment well-suited to the development of candidate linear control laws for flexible missile autopilots. The strengths of MATLAB include: (1) exceptionally high speed (MATLAB's version for 80386-based PC's offers benchmarks approaching minicomputer and mainframe performance); (2) ability to handle large design models of several hundred degrees of freedom, if necessary; and (3) broad extensibility through user-defined functions. To characterize MATLAB capabilities, a simplified design example is presented. This involves interactive definition of an observer-based state-space compensator for a flexible missile autopilot design task. MATLAB capabilities and limitations, in the context of this design task, are then summarized.

  15. Intraocular pressure following ECCE, phacoemulsification, and PC-IOL implantation.

    PubMed

    Kooner, K S; Cooksey, J C; Perry, P; Zimmerman, T J

    1988-09-01

    Abnormal intraocular pressure (IOP), either transient or permanent, may follow extracapsular cataract extraction (ECCE) with phacoemulsification (PE) and posterior chamber intraocular lens (PC-IOL) implantation. We retrospectively studied IOP measurements at different intervals post ECCE and PE in 242 eyes of 211 patients: 105 males, 106 females, 198 Caucasians and 13 blacks. Elevated IOP (greater than 23 mm Hg) was observed in 20 eyes (8.2%). Only two patients (0.8%) had persistent (greater than 3 months) IOP elevation and needed antiglaucoma therapy. Six more eyes (2.5%), however, developed glaucoma after 1 year. Hence, the incidence of secondary pseudophakic glaucoma at the conclusion of this study was 3.3%. No patient required laser or other mechanical surgery for IOP control. ECCE and PE with PC-IOL does not appear to adversely affect IOP. Patients, however, must be followed closely, as some may develop glaucoma months after surgery.

  16. Implementation of WASP-III on an IBM PC

    SciTech Connect

    Buehring, W.A.; Huber, C.C.; Hamilton, B.P.

    1985-01-01

    Argonne National Laboratory (ANL) is currently developing a microcomputer-based energy planning package entitled ENergy and Power Evaluation Program (ENPEP). As presently constructed, ENPEP consists of six analytic modules that can be run sequentially or independently: (1) macroeconomic analysis, (2) energy demand forecasting, (3) integrated energy demand/supply analysis, (4) electric load forecasting, (5) expansion planning for electrical generating systems, and (6) impacts of alternative supply systems. The Wien Automatic System Planning Package (WASP-III) is the fifth module of ENPEP and forms the heart of the ENPEP analytic capability. The experience in converting WASP-III to the IBM PC is reviewed and some recommendations for good performance on the PC are given. Finally, a few suggestions are presented for WASP improvements from the point of view of the user of the personal computer version of WASP.

  17. Structure of pentaquarks Pc+ in the chiral quark model

    NASA Astrophysics Data System (ADS)

    Yang, Gang; Ping, Jialun; Wang, Fan

    2017-01-01

    The recent experimental results of the LHCb Collaboration suggested the existence of pentaquark states with a charmonium. To understand the structure of the states, a dynamical calculation of 5-quark systems with quantum numbers I JP=1/2 (1/2 )±,1/2 (3/2 )±and1/2 (5/2 )±is performed in the framework of the chiral quark model with the help of the Gaussian expansion method. The results show that there are several negative parity resonance states while all of the positive parity states are the scattering states. The Pc(4380 ) state is suggested to be the pentaquark state of Σc*D ¯. Although the energy of ΣcD ¯* is very close to the mass of Pc(4450 ), the inconsistent parity prevents the assignment. The calculated distances between quarks confirm the molecular nature of the states.

  18. Dystrophin Dp71 in PC12 cell adhesion

    PubMed Central

    Enríquez-Aragón, Jose Arturo; Cerna-Cortés, Joel; Bermúdez de León, Mario; García-Sierra, Francisco; González, Everardo; Mornet, Dominique; Cisneros, Bulmaro

    2005-01-01

    Previously, we reported that PC12 cells with decreased Dp71 expression (antisense-Dp71 cells) display deficient nerve-growth-factor-induced neurite outgrowth. In this study, we show that disturbed neurite outgrowth of antisense-Dp71 cells is accompanied by decreased adhesion activity on laminin, collagen and fibronectin. In wild-type cells, the immunostaining of Dp71 and _1-integrin overlaps in the basal area contacting the substrate, but staining of both proteins decrease in the antisense-Dp71 cells. Morphology of antisense-Dp71 cells at the electron microscopic level is characterized by the lack of filopodia, cellular projections involved in adhesion. Our findings suggest that Dp71 is required for the efficient PC12 cell attachment to b1-integrin-dependent substrata and that decreased adhesion activity of the anti-sense-Dp71 cells could determine their deficiency to extend neurites. PMID:15706226

  19. Tablet PC Enabled Body Sensor System for Rural Telehealth Applications

    PubMed Central

    Panicker, Nitha V.; Kumar, A. Sukesh

    2016-01-01

    Telehealth systems benefit from the rapid growth of mobile communication technology for measuring physiological signals. Development and validation of a tablet PC enabled noninvasive body sensor system for rural telehealth application are discussed in this paper. This system includes real time continuous collection of physiological parameters (blood pressure, pulse rate, and temperature) and fall detection of a patient with the help of a body sensor unit and wireless transmission of the acquired information to a tablet PC handled by the medical staff in a Primary Health Center (PHC). Abnormal conditions are automatically identified and alert messages are given to the medical officer in real time. Clinical validation is performed in a real environment and found to be successful. Bland-Altman analysis is carried out to validate the wrist blood pressure sensor used. The system works well for all measurements. PMID:26884757

  20. An update to the list of mouse mutants with neural tube closure defects and advances toward a complete genetic perspective of neural tube closure.

    PubMed

    Harris, Muriel J; Juriloff, Diana M

    2010-08-01

    The number of mouse mutants and strains with neural tube defects (NTDs) now exceeds 240, including 205 representing specific genes, 30 for unidentified genes, and 9 multifactorial strains. These mutants identify genes needed for embryonic neural tube closure. Reports of 50 new NTD mutants since our 2007 review (Harris and Juriloff, 2007) were considered in relation to the previously reviewed mutants to obtain new insights into mechanisms of NTD etiology. In addition to null mutations, some are hypomorphs or conditional mutants. Some mutations do not cause NTDs on their own, but do so in digenic, trigenic, and oligogenic combinations, an etiology that likely parallels the nature of genetic etiology of human NTDs. Mutants that have only exencephaly are fourfold more frequent than those that have spina bifida aperta with or without exencephaly. Many diverse cellular functions and biochemical pathways are involved; the NTD mutants draw new attention to chromatin modification (epigenetics), the protease-activated receptor cascade, and the ciliopathies. Few mutants directly involve folate metabolism. Prevention of NTDs by maternal folate supplementation has been tested in 13 mutants and reduces NTD frequency in six diverse mutants. Inositol reduces spina bifida aperta frequency in the curly tail mutant, and three new mutants involve inositol metabolism. The many NTD mutants are the foundation for a future complete genetic understanding of the processes of neural fold elevation and fusion along mechanistically distinct cranial-caudal segments of the neural tube, and they point to several candidate processes for study in human NTD etiology.

  1. Direct Comparison of the Efficacy and Safety of Oral Treatments with Oleylphosphocholine (OlPC) and Miltefosine in a Mouse Model of L. major Cutaneous Leishmaniasis

    PubMed Central

    Fortin, Anny; Caridha, Diana P.; Leed, Susan; Ngundam, Franklyn; Sena, Jenell; Bosschaerts, Tom; Parriott, Sandi; Hickman, Mark R.; Hudson, Thomas H.; Grogl, Max

    2014-01-01

    Background Cutaneous leishmaniasis (CL) represents a range of skin diseases caused by infection with Leishmania parasites and associated with tissue inflammation and skin ulceration. CL is clinically widespread in both the Old and New World but lacks treatments that are well tolerated, effective and inexpensive. Oleylphosphocholine (OlPC) is a new orally bioavailable drug of the alkylphosphocholine family with potent antileishmanial activity against a broad range of Leishmania species/strains. Methodology/principal findings The potential of OlPC against Old World CL was evaluated in a mouse model of Leishmania (L.) major infection in BALB/c mice. Initial dose-response experiments showed that an oral daily dose of 40 mg/kg of OlPC was needed to impact time to cure and lesion sizes. This dose was then used to directly compare the efficacy of OlPC to the efficacy of the antileishmanial drugs miltefosine (40 mg/kg/day), fluconazole (160 mg/kg/day) and amphotericin B (25 mg/kg/day). OlPC, miltefosine and fluconazole were given orally for 21 days while amphotericin B was administered intraperitoneally for 10 days. Ulcer sizes and animal weights were followed up on a weekly basis and parasitemia was determined by means of a real-time in vivo imaging system which detects luminescence emitted from luciferase-expressing infecting L. major parasites. Amphotericin B and OlPC showed excellent efficacy against L. major lesions in terms of reduction of parasitic loads and by inducing complete healing of established lesions. In contrast, treatment with miltefosine did not significantly affect parasitemia and lesion sizes, while fluconazole was completely ineffective at the dose regimen tested. Conclusions/Significance Given the data showing the outstanding efficacy and tolerability of OlPC, our results suggest that OlPC is a promising new drug candidate to improve and simplify current clinical management of L. major CL. PMID:25210745

  2. Practical Pocket PC Application w/Biometric Security

    NASA Technical Reports Server (NTRS)

    Logan, Julian

    2004-01-01

    I work in the Flight Software Engineering Branch, where we provide design and development of embedded real-time software applications for flight and supporting ground systems to support the NASA Aeronautics and Space Programs. In addition, this branch evaluates, develops and implements new technologies for embedded real-time systems, and maintains a laboratory for applications of embedded technology. The majority of microchips that are used in modern society have been programmed using embedded technology. These small chips can be found in microwaves, calculators, home security systems, cell phones and more. My assignment this summer entails working with an iPAQ HP 5500 Pocket PC. This top-of-the-line hand-held device is one of the first mobile PC's to introduce biometric security capabilities. Biometric security, in this case a fingerprint authentication system, is on the edge of technology as far as securing information. The benefits of fingerprint authentication are enormous. The most significant of them are that it is extremely difficult to reproduce someone else's fingerprint, and it is equally difficult to lose or forget your own fingerprint as opposed to a password or pin number. One of my goals for this summer is to integrate this technology with another Pocket PC application. The second task for the summer is to develop a simple application that provides an Astronaut EVA (Extravehicular Activity) Log Book capability. The Astronaut EVA Log Book is what an astronaut would use to report the status of field missions, crew physical health, successes, future plans, etc. My goal is to develop a user interface into which these data fields can be entered and stored. The applications that I am developing are created using eMbedded Visual C++ 4.0 with the Pocket PC 2003 Software Development Kit provided by Microsoft.

  3. The PC Dugout Micro-Mainframe Integration System: PAX Implementation

    DTIC Science & Technology

    1991-05-01

    System Mainframe Implementation Language PC Implementation Language Communications Package and File Transfer Protocol Requirements 5 DUGOUT DESIGN...remember any commands or language syntax. In some cases users are asked to answer a few simple, nontechnical prompts. To download or upload files, a...problem. The database management systems Focus and Structured Query Language /Data System (SQL/DS) successfully circumvent this restriction in the PAX VM/CMS

  4. Falling PC Solitaire Cards: An Open-Inquiry Approach

    ERIC Educational Resources Information Center

    Gonzalez-Espada, Wilson J.

    2012-01-01

    Many of us have played the PC Solitaire game that comes as standard software in many computers. Although I am not a great player, occasionally I win a game or two. The game celebrates my accomplishment by pushing the cards forward, one at a time, falling gracefully in what appears to look like a parabolic path in a drag-free environment. One day,…

  5. The photodynamic antibacterial effects of silicon phthalocyanine (Pc) 4.

    PubMed

    Dimaano, Matthew L; Rozario, Chantal; Nerandzic, Michelle M; Donskey, Curtis J; Lam, Minh; Baron, Elma D

    2015-04-08

    The emergence of antibiotic-resistant strains in facultative anaerobic Gram-positive coccal bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA), is a global health issue. Typically, MRSA strains are found associated with institutions like hospitals but recent data suggest that they are becoming more prevalent in community-acquired infections. It is thought that the incidence and prevalence of bacterial infections will continue to increase as (a) more frequent use of broad-spectrum antibiotics and immunosuppressive medications; (b) increased number of invasive medical procedures; and (c) higher incidence of neutropenia and HIV infections. Therefore, more optimal treatments, such as photodynamic therapy (PDT), are warranted. PDT requires the interaction of light, a photosensitizing agent, and molecular oxygen to induce cytotoxic effects. In this study, we investigated the efficacy and characterized the mechanism of cytotoxicity induced by photodynamic therapy sensitized by silicon phthalocyanine (Pc) 4 on (a) methicillin-sensitive Staphylococcus aureus (MSSA) (ATCC 25923); (b) community acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) (ATCC 43300); and (c) hospital acquired methicillin-resistant Staphylococcus aureus (HA-MRSA) (PFGE type 300). Our data include confocal image analysis, which confirmed that Pc 4 is taken up by all S. aureus strains, and viable cell recovery assay, which showed that concentrations as low as 1.0 μM Pc 4 incubated for 3 h at 37 °C followed by light at 2.0 J/cm2 can reduce cell survival by 2-5 logs. These results are encouraging, but before PDT can be utilized as an alternative treatment for eradicating resistant strains, we must first characterize the mechanism of cell death that Pc 4-based PDT employs in eliminating these pathogens.

  6. The Photodynamic Antibacterial Effects of Silicon Phthalocyanine (Pc) 4

    PubMed Central

    Dimaano, Matthew L.; Rozario, Chantal; Nerandzic, Michelle M.; Donskey, Curtis J.; Lam, Minh; Baron, Elma D.

    2015-01-01

    The emergence of antibiotic-resistant strains in facultative anaerobic Gram-positive coccal bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA), is a global health issue. Typically, MRSA strains are found associated with institutions like hospitals but recent data suggest that they are becoming more prevalent in community-acquired infections. It is thought that the incidence and prevalence of bacterial infections will continue to increase as (a) more frequent use of broad-spectrum antibiotics and immunosuppressive medications; (b) increased number of invasive medical procedures; and (c) higher incidence of neutropenia and HIV infections. Therefore, more optimal treatments, such as photodynamic therapy (PDT), are warranted. PDT requires the interaction of light, a photosensitizing agent, and molecular oxygen to induce cytotoxic effects. In this study, we investigated the efficacy and characterized the mechanism of cytotoxicity induced by photodynamic therapy sensitized by silicon phthalocyanine (Pc) 4 on (a) methicillin-sensitive Staphylococcus aureus (MSSA) (ATCC 25923); (b) community acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) (ATCC 43300); and (c) hospital acquired methicillin-resistant Staphylococcus aureus (HA-MRSA) (PFGE type 300). Our data include confocal image analysis, which confirmed that Pc 4 is taken up by all S. aureus strains, and viable cell recovery assay, which showed that concentrations as low as 1.0 μM Pc 4 incubated for 3 h at 37 °C followed by light at 2.0 J/cm2 can reduce cell survival by 2–5 logs. These results are encouraging, but before PDT can be utilized as an alternative treatment for eradicating resistant strains, we must first characterize the mechanism of cell death that Pc 4-based PDT employs in eliminating these pathogens. PMID:25856680

  7. The phenotype of a phospholipase C (plc-1) mutant in a filamentous fungus, Neurospora crassa.

    PubMed

    Lew, Roger R; Giblon, Rachel E; Lorenti, Miranda S H

    2015-09-01

    In the filamentous fungus Neurospora crassa, phospholipase C may play a role in hyphal extension at the growing tips as part of a growth-sensing mechanism that activates calcium release from internal stores to mediate continued expansion of the hyphal tip. One candidate for a tip-localized phospholipase C is PLC-1. We characterized morphology and growth characteristics of a knockout mutant (KO plc-1) and a RIP mutated strain (RIP plc-1) (missense mutations and a nonsense mutation render the gene product non-functional). Growth and hyphal cytology of wildtype and KO plc-1 were similar, but the RIP plc-1 mutant grew slower and exhibited abnormal membrane structures at the hyphal tip, imaged using the fluorescence dye FM4-64. To test for causes of the slower growth of the RIP plc-1 mutant, we examined its physiological poise compared to wildtype and the KO plc-1 mutant. The electrical properties of all three strains and the electrogenic contribution of the plasma membrane H(+)-ATPase (identified by cyanide inhibition) were the same. Responses to high osmolarity were also similar. However, the RIP plc-1 mutant had a significantly lower turgor, a possible cause of its slower growth. While growth of all three strains was inhibited by the phospholipase C inhibitor 3-nitrocoumarin, the RIP plc-1 mutant did not exhibit hyphal bursting after addition of the inhibitor, observed in both wildtype and the KO plc-1 mutant. Although the plc-1 gene is not obligatory for tip growth, the phenotype of the RIP plc-1 mutant - abnormal tip cytology, lower turgor and resistance to inhibitor-induced hyphal bursting - suggest it does play a role in tip growth. The expression of a dysfunctional plc-1 gene may cause a shift to alternative mechanism(s) of growth sensing in hyphal extension.

  8. Characterization of rag1 mutant zebrafish leukocytes

    PubMed Central

    Petrie-Hanson, Lora; Hohn, Claudia; Hanson, Larry

    2009-01-01

    Background Zebrafish may prove to be one of the best vertebrate models for innate immunology. These fish have sophisticated immune components, yet rely heavily on innate immune mechanisms. Thus, the development and characterization of mutant and/or knock out zebrafish are critical to help define immune cell and immune gene functions in the zebrafish model. The use of Severe Combined Immunodeficient (SCID) and recombination activation gene 1 and 2 mutant mice has allowed the investigation of the specific contribution of innate defenses in many infectious diseases. Similar zebrafish mutants are now being used in biomedical and fish immunology related research. This report describes the leukocyte populations in a unique model, recombination activation gene 1-/- mutant zebrafish (rag1 mutants). Results Differential counts of peripheral blood leukocytes (PBL) showed that rag1 mutants had significantly decreased lymphocyte-like cell populations (34.7%) compared to wild-types (70.5%), and significantly increased granulocyte populations (52.7%) compared to wild-types (17.6%). Monocyte/macrophage populations were similar between mutants and wild-types, 12.6% and 11.3%, respectively. Differential leukocyte counts of rag1 mutant kidney hematopoietic tissue showed a significantly reduced lymphocyte-like cell population (8%), a significantly increased myelomonocyte population (57%), 34.8% precursor cells, and 0.2% thrombocytes, while wild-type hematopoietic kidney tissue showed 29.4% lymphocytes/lymphocyte-like cells, 36.4% myelomonocytes, 33.8% precursors and 0.5% thrombocytes. Flow cytometric analyses of kidney hematopoietic tissue revealed three leukocyte populations. Population A was monocytes and granulocytes and comprised 34.7% of the gated cells in rag1 mutants and 17.6% in wild-types. Population B consisted of hematopoietic precursors, and comprised 50% of the gated cells for rag1 mutants and 53% for wild-types. Population C consisted of lymphocytes and lymphocyte

  9. Characteristics of Agrobacterium tumefaciens auxotrophic mutant infectivity.

    PubMed

    Lippincott, B B; Lippincott, J A

    1966-10-01

    Lippincott, Barbara B. (Northwestern University, Evanston, Ill.), and James A. Lippincott. Characteristics of Agrobacterium tumefaciens auxotrophic mutant infectivity. J. Bacteriol. 92:937-945. 166.-Mutants of Agrobacterium tumefaciens auxotrophic for adenine, methionine, or asparagine are less infectious than the wild-type strain B6 from which they were derived and show increased infectivity on pinto bean leaves when the specific compounds required for growth of the mutants are added to the infected leaf. Reversion to a prototrophic form of nutrition is accompanied by increased infectivity. Tumors initiated by these auxotrophic mutants are shown to arise only at large wound sites where nutritional conditions may be less restricting. The data indicate that, after inoculation, the bacteria pass through a phase in which host-supplied nutrients are utilized for the production of one or more factors necessary for successful tumor initiation.

  10. Kinetics of alpha-PcCu --> beta-PcCu isothermal conversion in air and thermal behavior of beta-PcCu from in situ real-time laboratory parallel-beam X-ray powder diffraction.

    PubMed

    Ballirano, Paolo; Caminiti, Ruggero

    2009-07-09

    The kinetics of the alpha-PcCu --> beta-PcCu conversion in air has been followed, under isothermal conditions, in situ real-time in the 423-443 K temperature range. Data have been fitted following the JMAK model. The reaction order of the kinetics at 423 K is consistent with a diffusion controlled, deceleratory nucleation rate process for 2D laminar particles, whereas at higher temperatures it is consistent with a phase boundary controlled, deceleratory nucleation rate process for 2D laminar particles. At 423 K, the overall transformation mechanism implies three steps: growth of the alpha-PcCu phase, disordering of adjacent columns of molecules of phthalocyanine, and nucleation and growth of the beta-PcCu phase. The calculated empirical activation energy is of 187 kJ/mol significantly greater than that for the alpha-PcCo --> beta-PcCo conversion. This fact seems to support the reported different structures of alpha-PcCo and alpha-PcCu. Investigation of the thermal behavior of beta-PcCu indicates a strongly anisotropic thermal expansion that follows the alpha(c) > alpha(a) approximately = alpha(b) trend. Moreover, the beta angle decreases with increasing temperature. Such anisotropy is consistent with the geometry of the very weak N3...H3 hydrogen bond which acts mainly along the c axis.

  11. Design and research of retrofitting PC boiler into CFB boiler

    SciTech Connect

    Chen, H.P.; Lu, J.D.; Huang, L.; Liu, H.; Lin, Z.; Liu, D.C.

    1997-12-31

    In China, there are a large number of aged pulverized coal (PC) boilers at aging utility power plants. Many of them are beyond their reasonable working life or in a condition of unreliable operation, low combustion efficiency, and serious air pollution. It is very important and urgent to retrofit the aged PC boilers, and repower the aging utility power plants in China. Circulating fluidized bed (CFB) boilers have been developed rapidly, and paid great attention to in China. There are many striking advantages to retrofit an aged boiler with a CFB boiler. The retrofitting is suitable to meet the needs of effective utilization of low-grade coal, reducing SO{sub 2} and NO{sub x} emissions and repowering an aging power plant. The cost is much lower than that of building a new CFB unit. The National Laboratory of Coal Combustion (NLCC) has always paid great attention to studying and developing CFB combustion technology in connection with Chinese national conditions, and has evolved distinguishing technology features of its own. This paper introduces a new design concept of retrofitting PC boiler into Pi ({Pi}-shaped) CFB boiler with downward exhaust cyclone, and relevant research work and results of design and calculation.

  12. Electronic transitions and band offsets in C60:SubPc and C60:MgPc on MoO3 studied by modulated surface photovoltage spectroscopy

    NASA Astrophysics Data System (ADS)

    Fengler, S.; Dittrich, Th.; Rusu, M.

    2015-07-01

    Electronic transitions at interfaces between MoO3 layers and organic layers of C60, SubPc, MgPc, and nano-composite layers of SubPc:C60 and MgPc:C60 have been studied by modulated surface photovoltage (SPV) spectroscopy. For all systems, time dependent and modulated SPV signals pointed to dissociation of excitons at the MoO3/organic layer interfaces with a separation of holes towards MoO3. The highest occupied molecular orbital (HOMO)-lowest unoccupied molecular orbital (LUMO) gaps (EHL) of C60, SubPc, and MgPc and the effective EHL of SubPc:C60 and MgPc:C60 were measured. The offsets between the LUMO (ΔEL) or HOMO (ΔEH) bands were obtained with high precision and amounted to 0.33 or 0.73 eV for SubPc:C60, respectively, and to -0.33 or 0.67 eV for MgPc:C60, respectively. Exponential tails below EHL and most pronounced sub-bandgap transitions were characterized and ascribed to disorder and transitions from HOMO bands to unoccupied defect states.

  13. Insights into prevention of human neural tube defects by folic acid arising from consideration of mouse mutants.

    PubMed

    Harris, Muriel J

    2009-04-01

    Almost 30 years after the initial study by Richard W. Smithells and coworkers, it is still unknown how maternal periconceptional folic acid supplementation prevents human neural tube defects (NTDs). In this article, questions about human NTD prevention are considered in relation to three groups of mouse models: NTD mutants that respond to folate, NTD mutants and strains that do not respond to folate, and mutants involving folate-pathway genes. Of the 200 mouse NTD mutants, only a few have been tested with folate; half respond and half do not. Among responsive mutants, folic acid supplementation reduces exencephaly and/or spina bifida aperta frequency in the Sp(2H), Sp, Cd, Cited2, Cart1, and Gcn5 mutants. Prevention ranges from 35 to 85%. The responsive Sp(2H) (Pax3) mutant has abnormal folate metabolism, but the responsive Cited2 mutant does not. Neither folic nor folinic acid reduces NTD frequency in Axd, Grhl3, Fkbp8, Map3k4, or Nog mutants or in the curly tail or SELH/Bc strains. Spina bifida frequency is reduced in Axd by methionine and in curly tail by inositol. Exencephaly frequency is reduced in SELH/Bc by an alternative commercial ration. Mutations in folate-pathway genes do not cause NTDs, except for 30% exencephaly in folate-treated Folr1. Among folate-pathway mutants, neural tube closure is normal in Cbs, Folr2, Mthfd1, Mthfd2, Mthfr, and Shmt1 mutants. Embryos die by midgestation in Folr1, Mtr, Mtrr, and RFC1 mutants. The mouse models point to genetic heterogeneity in the ability to respond to folic acid and also to heterogeneity in genetic cause of NTDs that can be prevented by folic acid.

  14. Fatty acid regulates gene expression and growth of human prostate cancer PC-3 cells

    NASA Technical Reports Server (NTRS)

    Hughes-Fulford, M.; Chen, Y.; Tjandrawinata, R. R.

    2001-01-01

    It has been proposed that the omega-6 fatty acids increase the rate of tumor growth. Here we test that hypothesis in the PC-3 human prostate tumor. We found that the essential fatty acids, linoleic acid (LA) and arachidonic acid (AA), and the AA metabolite PGE(2) stimulate tumor growth while oleic acid (OA) and the omega-3 fatty acid, eicosapentaenoic acid (EPA) inhibited growth. In examining the role of AA in growth response, we extended our studies to analyze changes in early gene expression induced by AA. We demonstrate that c-fos expression is increased within minutes of addition in a dose-dependent manner. Moreover, the immediate early gene cox-2 is also increased in the presence of AA in a dose-dependent manner, while the constitutive cox-1 message was not increased. Three hours after exposure to AA, the synthesis of PGE(2) via COX-2 was also increased. Previous studies have demonstrated that AA was primarily delivered by low density lipoprotein (LDL) via its receptor (LDLr). Since it is known that hepatomas, acute myelogenous leukemia and colorectal tumors lack normal cholesterol feedback, we examined the role of the LDLr in growth regulation of the PC-3 prostate cancer cells. Analysis of ldlr mRNA expression and LDLr function demonstrated that human PC-3 prostate cancer cells lack normal feedback regulation. While exogenous LDL caused a significant stimulation of cell growth and PGE(2) synthesis, no change was seen in regulation of the LDLr by LDL. Taken together, these data show that normal cholesterol feedback of ldlr message and protein is lost in prostate cancer. These data suggest that unregulated over-expression of LDLr in tumor cells would permit increased availability of AA, which induces immediate early genes c-fos and cox-2 within minutes of uptake.

  15. Predicting flow in aortic dissection: comparison of computational model with PC-MRI velocity measurements.

    PubMed

    Cheng, Z; Juli, C; Wood, N B; Gibbs, R G J; Xu, X Y

    2014-09-01

    Aortic dissection is a life-threatening process in which the weakened wall develops a tear, causing separation of wall layers. The dissected layers separate the original true aortic lumen and a newly created false lumen. If untreated, the condition can be fatal. Flow rate in the false lumen is a key feature for false lumen patency, which has been regarded as one of the most important predictors of adverse early and later outcomes. Detailed flow analysis in the dissected aorta may assist vascular surgeons in making treatment decisions, but computational models to simulate flow in aortic dissections often involve several assumptions. The purpose of this study is to assess the computational models adopted in previous studies by comparison with in vivo velocity data obtained by means of phase-contrast magnetic resonance imaging (PC-MRI). Aortic dissection geometry was reconstructed from computed tomography (CT) images, while PC-MRI velocity data were used to define inflow conditions and to provide distal velocity components for comparison with the simulation results. The computational fluid dynamics (CFD) simulation incorporated a laminar-turbulent transition model, which is necessary for adequate flow simulation in aortic conditions. Velocity contours from PC-MRI and CFD in the two lumens at the distal plane were compared at four representative time points in the pulse cycle. The computational model successfully captured the complex regions of flow reversal and recirculation qualitatively, although quantitative differences exist. With a rigid wall assumption and exclusion of arch branches, the CFD model over-predicted the false lumen flow rate by 25% at peak systole. Nevertheless, an overall good agreement was achieved, confirming the physiological relevance and validity of the computational model for type B aortic dissection with a relatively stiff dissection flap.

  16. Mutant IDH1 and thrombosis in gliomas.

    PubMed

    Unruh, Dusten; Schwarze, Steven R; Khoury, Laith; Thomas, Cheddhi; Wu, Meijing; Chen, Li; Chen, Rui; Liu, Yinxing; Schwartz, Margaret A; Amidei, Christina; Kumthekar, Priya; Benjamin, Carolina G; Song, Kristine; Dawson, Caleb; Rispoli, Joanne M; Fatterpekar, Girish; Golfinos, John G; Kondziolka, Douglas; Karajannis, Matthias; Pacione, Donato; Zagzag, David; McIntyre, Thomas; Snuderl, Matija; Horbinski, Craig

    2016-12-01

    Mutant isocitrate dehydrogenase 1 (IDH1) is common in gliomas, and produces D-2-hydroxyglutarate (D-2-HG). The full effects of IDH1 mutations on glioma biology and tumor microenvironment are unknown. We analyzed a discovery cohort of 169 World Health Organization (WHO) grade II-IV gliomas, followed by a validation cohort of 148 cases, for IDH1 mutations, intratumoral microthrombi, and venous thromboemboli (VTE). 430 gliomas from The Cancer Genome Atlas were analyzed for mRNAs associated with coagulation, and 95 gliomas in a tissue microarray were assessed for tissue factor (TF) protein. In vitro and in vivo assays evaluated platelet aggregation and clotting time in the presence of mutant IDH1 or D-2-HG. VTE occurred in 26-30 % of patients with wild-type IDH1 gliomas, but not in patients with mutant IDH1 gliomas (0 %). IDH1 mutation status was the most powerful predictive marker for VTE, independent of variables such as GBM diagnosis and prolonged hospital stay. Microthrombi were far less common within mutant IDH1 gliomas regardless of WHO grade (85-90 % in wild-type versus 2-6 % in mutant), and were an independent predictor of IDH1 wild-type status. Among all 35 coagulation-associated genes, F3 mRNA, encoding TF, showed the strongest inverse relationship with IDH1 mutations. Mutant IDH1 gliomas had F3 gene promoter hypermethylation, with lower TF protein expression. D-2-HG rapidly inhibited platelet aggregation and blood clotting via a novel calcium-dependent, methylation-independent mechanism. Mutant IDH1 glioma engraftment in mice significantly prolonged bleeding time. Our data suggest that mutant IDH1 has potent antithrombotic activity within gliomas and throughout the peripheral circulation. These findings have implications for the pathologic evaluation of gliomas, the effect of altered isocitrate metabolism on tumor microenvironment, and risk assessment of glioma patients for VTE.

  17. Quantitative Analysis of Triple Mutant Genetic Interactions

    PubMed Central

    Braberg, Hannes; Alexander, Richard; Shales, Michael; Xu, Jiewei; Franks-Skiba, Kathleen E.; Wu, Qiuqin; Haber, James E.; Krogan, Nevan J.

    2014-01-01

    The quantitative analysis of genetic interactions between pairs of gene mutations has proven effective for characterizing cellular functions but can miss important interactions for functionally redundant genes. To address this limitation, we have developed an approach termed Triple Mutant Analysis (TMA). The procedure relies on a query strain that contains two deletions in a pair of redundant or otherwise related genes, that is crossed against a panel of candidate deletion strains to isolate triple mutants and measure their growth. A central feature of TMA is to interrogate mutants that are synthetically sick when two other genes are deleted but interact minimally with either single deletion. This approach has been valuable for discovering genes that restore critical functions when the principle actors are deleted. TMA has also uncovered double mutant combinations that produce severe defects because a third protein becomes deregulated and acts in a deleterious fashion, and it has revealed functional differences between proteins presumed to act together. The protocol is optimized for Singer ROTOR pinning robots, takes 3 weeks to complete, and measures interactions for up to 30 double mutants against a library of 1536 single mutants. PMID:25010907

  18. High-Throughput Parallel Sequencing to Measure Fitness of Leptospira interrogans Transposon Insertion Mutants during Acute Infection

    PubMed Central

    Matsunaga, James; Haake, David A.

    2016-01-01

    Pathogenic species of Leptospira are the causative agents of leptospirosis, a zoonotic disease that causes mortality and morbidity worldwide. The understanding of the virulence mechanisms of Leptospira spp is still at an early stage due to the limited number of genetic tools available for this microorganism. The development of random transposon mutagenesis in pathogenic strains a decade ago has contributed to the identification of several virulence factors. In this study, we used the transposon sequencing (Tn-Seq) technique, which combines transposon mutagenesis with massive parallel sequencing, to study the in vivo fitness of a pool of Leptospira interrogans mutants. We infected hamsters with a pool of 42 mutants (input pool), which included control mutants with insertions in four genes previously analyzed by virulence testing (loa22, ligB, flaA1, and lic20111) and 23 mutants with disrupted signal transduction genes. We quantified the mutants in different tissues (blood, kidney and liver) at 4 days post-challenge by high-throughput sequencing and compared the frequencies of mutants recovered from tissues to their frequencies in the input pool. Control mutants that were less fit in the Tn-Seq experiment were attenuated for virulence when tested separately in the hamster model of lethal leptospirosis. Control mutants with unaltered fitness were as virulent as the wild-type strain. We identified two mutants with the transposon inserted in the same putative adenylate/guanylate cyclase gene (lic12327) that had reduced in vivo fitness in blood, kidney and liver. Both lic12327 mutants were attenuated for virulence when tested individually in hamsters. Growth of the control mutants and lic12327 mutants in culture medium were similar to that of the wild-type strain. These results demonstrate the feasibility of screening large pools of L. interrogans transposon mutants for those with altered fitness, and potentially attenuated virulence, by transposon sequencing. PMID

  19. High-Throughput Parallel Sequencing to Measure Fitness of Leptospira interrogans Transposon Insertion Mutants during Acute Infection.

    PubMed

    Lourdault, Kristel; Matsunaga, James; Haake, David A

    2016-11-01

    Pathogenic species of Leptospira are the causative agents of leptospirosis, a zoonotic disease that causes mortality and morbidity worldwide. The understanding of the virulence mechanisms of Leptospira spp is still at an early stage due to the limited number of genetic tools available for this microorganism. The development of random transposon mutagenesis in pathogenic strains a decade ago has contributed to the identification of several virulence factors. In this study, we used the transposon sequencing (Tn-Seq) technique, which combines transposon mutagenesis with massive parallel sequencing, to study the in vivo fitness of a pool of Leptospira interrogans mutants. We infected hamsters with a pool of 42 mutants (input pool), which included control mutants with insertions in four genes previously analyzed by virulence testing (loa22, ligB, flaA1, and lic20111) and 23 mutants with disrupted signal transduction genes. We quantified the mutants in different tissues (blood, kidney and liver) at 4 days post-challenge by high-throughput sequencing and compared the frequencies of mutants recovered from tissues to their frequencies in the input pool. Control mutants that were less fit in the Tn-Seq experiment were attenuated for virulence when tested separately in the hamster model of lethal leptospirosis. Control mutants with unaltered fitness were as virulent as the wild-type strain. We identified two mutants with the transposon inserted in the same putative adenylate/guanylate cyclase gene (lic12327) that had reduced in vivo fitness in blood, kidney and liver. Both lic12327 mutants were attenuated for virulence when tested individually in hamsters. Growth of the control mutants and lic12327 mutants in culture medium were similar to that of the wild-type strain. These results demonstrate the feasibility of screening large pools of L. interrogans transposon mutants for those with altered fitness, and potentially attenuated virulence, by transposon sequencing.

  20. PRRT2 Mutant Leads to Dysfunction of Glutamate Signaling

    PubMed Central

    Li, Ming; Niu, Fenghe; Zhu, Xilin; Wu, Xiaopan; Shen, Ning; Peng, Xiaozhong; Liu, Ying

    2015-01-01

    Paroxysmal kinesigenic choreoathetosis (PKC) is an inherited disease of the nervous system. We previously identified PRRT2 as the causative gene of PKC. However, as little is known about the function of PRRT2, elucidating its function will benefit not only PKC studies, but also many other related disorders. Here, we reveal higher levels of glutamate in the plasma of PKC patients and the culture medium of neurons following knock-out Prrt2 expression. Using double immunostaining assays we confirm Prrt2 is located at the glutamatergic neurons in accordance with its function. Our co-immunoprecipitation assays reveal mutant PRRT2 interferes with SNAP25 and GRIA1 interactions, respectively. Furthermore, using live-labeling techniques, we confirmed co-transfection with mutant PRRT2 caused an increase in GRIA1 distribution on the cell surface. Therefore, our results suggest that mutant PRRT2, probably through its weakened interaction with SNAP25, affects glutamate signaling and glutamate receptor activity, resulting in the increase of glutamate release and subsequent neuronal hyperexcitability. PMID:25915028

  1. Mutant p53 protein localized in the cytoplasm inhibits autophagy.

    PubMed

    Morselli, Eugenia; Tasdemir, Ezgi; Maiuri, Maria Chiara; Galluzzi, Lorenzo; Kepp, Oliver; Criollo, Alfredo; Vicencio, José Miguel; Soussi, Thierry; Kroemer, Guido

    2008-10-01

    The knockout, knockdown or chemical inhibition of p53 stimulates autophagy. Moreover, autophagy-inducing stimuli such as nutrient depletion, rapamycin or lithium cause the depletion of cytoplasmic p53, which in turn is required for the induction of autophagy. Here, we show that retransfection of p53(-/-) HCT 116 colon carcinoma cells with wild type p53 decreases autophagy down to baseline levels. Surprisingly, one third among a panel of 22 cancer-associated p53 single amino acid mutants also inhibited autophagy when transfected into p53(-/-) cells. Those variants of p53 that preferentially localize to the cytoplasm effectively repressed autophagy, whereas p53 mutants that display a prominently nuclear distribution failed to inhibit autophagy. The investigation of a series of deletion mutants revealed that removal of the DNA-binding domain from p53 fails to interfere with its role in the regulation of autophagy. Altogether, these results identify the cytoplasmic localization of p53 as the most important feature for p53-mediated autophagy inhibition. Moreover, the structural requirements for the two biological activities of extranuclear p53, namely induction of apoptosis and inhibition of autophagy, are manifestly different.

  2. Yeast mutants affecting possible quality control of plasma membrane proteins.

    PubMed

    Li, Y; Kane, T; Tipper, C; Spatrick, P; Jenness, D D

    1999-05-01

    Mutations gef1, stp22, STP26, and STP27 in Saccharomyces cerevisiae were identified as suppressors of the temperature-sensitive alpha-factor receptor (mutation ste2-3) and arginine permease (mutation can1(ts)). These suppressors inhibited the elimination of misfolded receptors (synthesized at 34 degrees C) as well as damaged surface receptors (shifted from 22 to 34 degrees C). The stp22 mutation (allelic to vps23 [M. Babst and S. Emr, personal communication] and the STP26 mutation also caused missorting of carboxypeptidase Y, and ste2-3 was suppressed by mutations vps1, vps8, vps10, and vps28 but not by mutation vps3. In the stp22 mutant, both the mutant and the wild-type receptors (tagged with green fluorescent protein [GFP]) accumulated within an endosome-like compartment and were excluded from the vacuole. GFP-tagged Stp22p also accumulated in this compartment. Upon reaching the vacuole, cytoplasmic domains of both mutant and wild-type receptors appeared within the vacuolar lumen. Stp22p and Gef1p are similar to tumor susceptibility protein TSG101 and voltage-gated chloride channel, respectively. These results identify potential elements of plasma membrane quality control and indicate that cytoplasmic domains of membrane proteins are translocated into the vacuolar lumen.

  3. Studies on Electron-Transport Reactions of Photosynthesis in Plastome Mutants of Oenothera

    PubMed Central

    Fork, David C.; Heber, Ulrich W.

    1968-01-01

    Fluorescence characteristics and light-induced absorbance changes of 5 plastome mutants of Oenothera, all having a defect in photosynthesis, were investigated to localize the site of the block in their photosynthetic mechanism and to relate mutational changes in the plastome to specific biochemical events in photosynthesis. In 4 of the mutants examined photosystem 2 was largely, or completely, nonfunctional. Excitation of system 2 did not cause reduction of oxidized cytochrome f in these mutants. The system-2 dependent absorbance change at 518 mμ seen in normal leaves was absent in the mutants. Moreover, the mutants had a high initial fluorescence in the presence and in the absence of 3- (3,4-dichlorophenyl)-1,1-dimethylurea, which did not change during illumination, indicating that the reaction centers of system 2 were affected by the mutations. Photosystem 1 functioned normally. A fifth mutant had an impaired photosystem 1. Even high intensity far-red light did not lead to an accumulation of oxidized cytochrome f as was seen in normal plants. Photosystem 2 was functioning, as evidenced by the fast reduction of the primary system-2 oxidant, and by the characteristics of the 518-mμ absorbance change. Because 1 of the 2 photosystems is functional in all mutants, and because they all have the enzymes of the photosynthetic carbon cycle, it appears that the effect of the mutation is specific. The results suggest that the plastome controls reactions within the electron-transport chain of photosynthesis. PMID:16656813

  4. Static and Dynamic Energetic Disorders in the C60, PC61BM, C70, and PC71BM Fullerenes.

    PubMed

    Tummala, Naga Rajesh; Zheng, Zilong; Aziz, Saadullah G; Coropceanu, Veaceslav; Brédas, Jean-Luc

    2015-09-17

    We use a combination of molecular dynamics simulations and density functional theory calculations to investigate the energetic disorder in fullerene systems. We show that the energetic disorder evaluated from an ensemble average contains contributions of both static origin (time-independent, due to loose packing) and dynamic origin (time-dependent, due to electron-vibration interactions). In order to differentiate between these two contributions, we compare the results obtained from an ensemble average approach with those derived from a time average approach. It is found that in both amorphous C60 and C70 bulk systems, the degrees of static and dynamic disorder are comparable, while in the amorphous PC61BM and PC71BM systems, static disorder is about twice as large as dynamic disorder.

  5. An overview of the evaluation plan for PC/MISI: PC-based Multiple Information System Interface

    NASA Technical Reports Server (NTRS)

    Dominick, Wayne D. (Editor); Lim, Bee Lee; Hall, Philip P.

    1985-01-01

    An initial evaluation plan for the personal computer multiple information system interface (PC/MISI) project is discussed. The document is intend to be used as a blueprint for the evaluation of this system. Each objective of the design project is discussed along with the evaluation parameters and methodology to be used in the evaluation of the implementation's achievement of those objectives. The potential of the system for research activities related to more general aspects of information retrieval is also discussed.

  6. The Aqueous Extract of Rhizome of Gastrodia elata Protected Drosophila and PC12 Cells against Beta-Amyloid-Induced Neurotoxicity

    PubMed Central

    Ng, Chun-Fai; Ko, Chun-Hay; Koon, Chi-Man; Xian, Jia-Wen; Leung, Ping-Chung; Fung, Kwok-Pui; Chan, Ho Yin Edwin; Lau, Clara Bik-San

    2013-01-01

    This study aims to investigate the neuroprotective effect of the rhizome of Gastrodia elata (GE) aqueous extract on beta-amyloid(Aβ)-induced toxicity in vivo and in vitro. Transgenic Drosophila mutants with Aβ-induced neurodegeneration in pan-neuron and ommatidia were used to determine the efficacy of GE. The antiapoptotic and antioxidative mechanisms of GE were also studied in Aβ-treated pheochromocytoma (PC12) cells. In vivo studies demonstrated that GE (5 mg/g Drosophila media)-treated Drosophila possessed a longer lifespan, better locomotor function, and less-degenerated ommatidia when compared with the Aβ-expressing control (all P < 0.05). In vitro studies illustrated that GE increased the cell viability of Aβ-treated PC12 cells in dose-dependent manner, probably through attenuation of Aβ-induced oxidative and apoptotic stress. GE also significantly upregulated the enzymatic activities of catalase, superoxide dismutase, and glutathione peroxidase, leading to the decrease of reactive oxidation species production and apoptotic marker caspase-3 activity. In conclusion, our current data presented the first evidence that the aqueous extract of GE was capable of reducing the Aβ-induced neurodegeneration in Drosophila, possibly through inhibition of apoptosis and reduction of oxidative stress. GE aqueous extract could be developed as a promising herbal agent for neuroprotection and novel adjuvant therapies for Alzheimer's disease. PMID:24174977

  7. Is the Metallic Phosphorus Carbide (β0-PC) Monolayer Stable? An Answer from a Theoretical Perspective.

    PubMed

    Rajbanshi, Biplab; Sarkar, Pranab

    2017-02-16

    Phosphorus carbide (PC) has been the subject of major research efforts in recent years. In this regard, very recently, a stoichiometric metallic phosphorus carbide (β0-PC) monolayer has been proposed as locally stable with one lone nonbonding electron in each C atom. Therefore, the ambiguity of coexistence of a nonbonding electron with metallic properties for β0-PC is reported and hence deserves further explanation. Herein, using first-principles calculations, we have explored the stability and electronic properties of β0-PC to resolve this ambiguity. The metallic behavior of β0-PC is explained on the basis of electron delocalization involving P and C atoms along a zigzag chain of β0-PC. We have also explored the possibility of getting a β0-PC monolayer via homogeneous doping of C (P) into phosphorene (graphene) and layer exfoliation of 3D bulk PC with β-InS-like structure, which has been experimentally synthesized.

  8. In vivo pathogenic role of mutant SOD1 localized in the mitochondrial intermembrane space

    PubMed Central

    Igoudjil, Anissa; Magrané, Jordi; Fischer, Lindsey R.; Kim, Hyun Jeong; Hervias, Isabel; Dumont, Magali; Cortez, Czrina; Glass, Jonathan D.; Starkov, Anatoly A.; Manfredi, Giovanni

    2011-01-01

    Mutations in Cu,Zn superoxide dismutase (SOD1) are associated with familial amyotrophic lateral sclerosis (ALS). Mutant SOD1 causes a complex array of pathological events, through toxic gain of function mechanisms, leading to selective motor neuron degeneration. Mitochondrial dysfunction is among the well-established toxic effects of mutant SOD1, but its mechanisms are just starting to be elucidated. A portion of mutant SOD1 is localized in mitochondria, where it accumulates mostly on the outer membrane and inside the intermembrane space (IMS). Evidence in cultured cells suggests that mutant SOD1 in the IMS causes mitochondrial dysfunction and compromises cell viability. Therefore, to test its pathogenic role in vivo we generated transgenic mice expressing G93A mutant or wild type (WT) human SOD1 targeted selectively to the mitochondrial IMS (mito-SOD1). We show that mito-SOD1 is correctly localized in the IMS, where it oligomerizes and acquires enzymatic activity. Mito-G93ASOD1 mice, but not mito-WTSOD1 mice, develop a progressive disease characterized by body weight loss, muscle weakness, brain atrophy, and motor impairment, which is more severe in females. These symptoms are associated with reduced spinal motor neuron counts and impaired mitochondrial bioenergetics, characterized by decreased cytochrome oxidase activity and defective calcium handling. However, there is no evidence of muscle denervation, a cardinal pathological feature of ALS. Taken together, our findings indicate that mutant SOD1 in the mitochondrial IMS causes mitochondrial dysfunction and neurodegeneration, but per se it is not sufficient to cause a full-fledged ALS phenotype, which requires the participation of mutant SOD1 localized in other cellular compartments. PMID:22049426

  9. In vivo pathogenic role of mutant SOD1 localized in the mitochondrial intermembrane space.

    PubMed

    Igoudjil, Anissa; Magrané, Jordi; Fischer, Lindsey R; Kim, Hyun Jeong; Hervias, Isabel; Dumont, Magali; Cortez, Czrina; Glass, Jonathan D; Starkov, Anatoly A; Manfredi, Giovanni

    2011-11-02

    Mutations in Cu,Zn superoxide dismutase (SOD1) are associated with familial amyotrophic lateral sclerosis (ALS). Mutant SOD1 causes a complex array of pathological events, through toxic gain of function mechanisms, leading to selective motor neuron degeneration. Mitochondrial dysfunction is among the well established toxic effects of mutant SOD1, but its mechanisms are just starting to be elucidated. A portion of mutant SOD1 is localized in mitochondria, where it accumulates mostly on the outer membrane and inside the intermembrane space (IMS). Evidence in cultured cells suggests that mutant SOD1 in the IMS causes mitochondrial dysfunction and compromises cell viability. Therefore, to test its pathogenic role in vivo we generated transgenic mice expressing G93A mutant or wild-type (WT) human SOD1 targeted selectively to the mitochondrial IMS (mito-SOD1). We show that mito-SOD1 is correctly localized in the IMS, where it oligomerizes and acquires enzymatic activity. Mito-G93ASOD1 mice, but not mito-WTSOD1 mice, develop a progressive disease characterized by body weight loss, muscle weakness, brain atrophy, and motor impairment, which is more severe in females. These symptoms are associated with reduced spinal motor neuron counts and impaired mitochondrial bioenergetics, characterized by decreased cytochrome oxidase activity and defective calcium handling. However, there is no evidence of muscle denervation, a cardinal pathological feature of ALS. Together, our findings indicate that mutant SOD1 in the mitochondrial IMS causes mitochondrial dysfunction and neurodegeneration, but per se it is not sufficient to cause a full-fledged ALS phenotype, which requires the participation of mutant SOD1 localized in other cellular compartments.

  10. Mutant TDP-43 within motor neurons drives disease onset but not progression in amyotrophic lateral sclerosis.

    PubMed

    Ditsworth, Dara; Maldonado, Marcus; McAlonis-Downes, Melissa; Sun, Shuying; Seelman, Amanda; Drenner, Kevin; Arnold, Eveline; Ling, Shuo-Chien; Pizzo, Donald; Ravits, John; Cleveland, Don W; Da Cruz, Sandrine

    2017-03-29

    Mutations in TDP-43 cause amyotrophic lateral sclerosis (ALS), a fatal paralytic disease characterized by degeneration and premature death of motor neurons. The contribution of mutant TDP-43-mediated damage within motor neurons was evaluated using mice expressing a conditional allele of an ALS-causing TDP-43 mutant (Q331K) whose broad expression throughout the central nervous system mimics endogenous TDP-43. TDP-43(Q331K) mice develop age- and mutant-dependent motor deficits from degeneration and death of motor neurons. Cre-recombinase-mediated excision of the TDP-43(Q331K) gene from motor neurons is shown to delay onset of motor symptoms and appearance of TDP-43-mediated aberrant nuclear morphology, and abrogate subsequent death of motor neurons. However, reduction of mutant TDP-43 selectively in motor neurons did not prevent age-dependent degeneration of axons and neuromuscular junction loss, nor did it attenuate astrogliosis or microgliosis. Thus, disease mechanism is non-cell autonomous with mutant TDP-43 expressed in motor neurons determining disease onset but progression defined by mutant acting within other cell types.

  11. Rhes, a Striatal Specific Protein, Mediates Mutant-Huntingtin Cytotoxicity

    PubMed Central

    Subramaniam, Srinivasa; Sixt, Katherine M.; Barrow, Roxanne; Snyder, Solomon H.

    2009-01-01

    Huntington's disease (HD) is caused by a polyglutamine repeat in the protein huntingtin (Htt) with mutant Htt (mHtt) expressed throughout the body and similarly in all brain regions. Yet, HD neuropathology is largely restricted to the corpus striatum. We report that the small guanine nucleotide–binding protein Rhes, which is localized very selectively to the striatum, binds physiologically to mHtt. Using cultured cells, we found Rhes induces sumoylation of mHtt, which leads to cytotoxicity. Thus, Rhes-mHtt interactions can account for the localized neuropathology of HD. PMID:19498170

  12. The Rec102 Mutant of Yeast Is Defective in Meiotic Recombination and Chromosome Synapsis

    PubMed Central

    Bhargava, J.; Engebrecht, J. A.; Roeder, G. S.

    1992-01-01

    A mutation at the REC102 locus was identified in a screen for yeast mutants that produce inviable spores. rec102 spore lethality is rescued by a spo13 mutation, which causes cells to bypass the meiosis I division. The rec102 mutation completely eliminates meiotically induced gene conversion and crossing over but has no effect on mitotic recombination frequencies. Cytological studies indicate that the rec102 mutant makes axial elements (precursors to the synaptonemal complex), but homologous chromosomes fail to synapse. In addition, meiotic chromosome segregation is significantly delayed in rec102 strains. Studies of double and triple mutants indicate that the REC102 protein acts before the RAD52 gene product in the meiotic recombination pathway. The REC102 gene was cloned based on complementation of the mutant defect and the gene was mapped to chromosome XII between CDC25 and STE11. PMID:1732169

  13. A Novel Positive Selection for Identifying Cold-Sensitive Myosin II Mutants in Dictyostelium

    PubMed Central

    Patterson, B.; Spudich, J. A.

    1995-01-01

    We developed a positive selection for myosin heavy chain mutants in Dictyostelium. This selection is based on the fact that brief exposure to azide causes wild-type cells to release from the substrate, whereas myosin null cells remain adherent. This procedure assays myosin function on a time scale of minutes and has therefore allowed us to select rapid-onset cold-sensitive mutants after random chemical mutagenesis of Dictyostelium cells. We developed a rapid technique for determining which mutations lie in sequences of the myosin gene that encode the head (motor) domain and localized 27 of 34 mutants to this domain. We recovered the appropriate sequences from five of the mutants and demonstrated that they retain their cold-sensitive properties when expressed from extrachromosomal plasmids. PMID:7498732

  14. Reduced Accumulation of ABA during Water Stress in a Molybdenum Cofactor Mutant of Barley 1

    PubMed Central

    Walker-Simmons, Mary; Kudrna, David A.; Warner, Robert L.

    1989-01-01

    A barley (Hordeum vulgare L.) mutant (Az34) has been identified with low basal levels of abscisic acid (ABA) and with reduced capacity for producing ABA in response to water stress. The mutation is in a gene controlling the molybdenum cofactor resulting in a pleiotropic deficiency in at least three molybdoenzymes, nitrate reductase, xanthine dehydrogenase, and aldehyde oxidase. The mutant was found to lack aldehyde oxidase activity with several substrates including: (a) ABA aldehyde, a putative precursor of ABA; (b) an acetylenic analog of ABA aldehyde; and (c) heptaldehyde. Elevating the growth temperature from 18 to 26°C caused mutant leaves to wilt and brown. Desiccation of mutant leaves was prevented by applying ABA. These results indicate that ABA biosynthesis at some developmental stages is dependent upon a molybdoenzyme which may be an aldehyde oxidase. Images Figure 5 PMID:16666835

  15. Protective Effects of Costunolide against Hydrogen Peroxide-Induced Injury in PC12 Cells.

    PubMed

    Cheong, Chong-Un; Yeh, Ching-Sheng; Hsieh, Yi-Wen; Lee, Ying-Ray; Lin, Mei-Ying; Chen, Chung-Yi; Lee, Chien-Hsing

    2016-07-09

    Oxidative stress-mediated cellular injury has been considered as a major cause of neurodegenerative diseases including Alzheimer's and Parkinson's diseases. The scavenging of reactive oxygen species (ROS) mediated by antioxidants may be a potential strategy for retarding the diseases' progression. Costunolide (CS) is a well-known sesquiterpene lactone, used as a popular herbal remedy, which possesses anti-inflammatory and antioxidant activity. This study aimed to investigate the protective role of CS against the cytotoxicity induced by hydrogen peroxide (H₂O₂) and to elucidate potential protective mechanisms in PC12 cells. The results showed that the treatment of PC12 cells with CS prior to H₂O₂ exposure effectively increased the cell viability. Furthermore, it decreased the intracellular ROS, stabilized the mitochondria membrane potential (MMP), and reduced apoptosis-related protein such as caspase 3. In addition, CS treatment attenuated the cell injury by H₂O₂ through the inhibition of phosphorylation of p38 and the extracellular signal-regulated kinase (ERK). These results demonstrated that CS is promising as a potential therapeutic candidate for neurodegenerative diseases resulting from oxidative damage and further research on this topic should be encouraged.

  16. K6PC-5, a novel sphingosine kinase 1 (SphK1) activator, alleviates dexamethasone-induced damages to osteoblasts through activating SphK1-Akt signaling.

    PubMed

    Ji, Feng; Mao, Li; Liu, Yuanyuan; Cao, Xiaojian; Xie, Yue; Wang, Shouguo; Fei, Haodong

    2015-03-13

    Long-term glucocorticoid usage is a common cause of non-traumatic femoral head osteonecrosis. Glucocorticoids (i.e. dexamethasone (Dex)) could directly induce damages to osteoblasts. In the current study, we investigated the potential activity of K6PC-5 [N-(1,3-dihydroxyisopropyl)-2-hexyl-3-oxo-decanamide], a novel sphingosine kinase 1 (SphK1) activator, against this process. Our data revealed that both osteoblastic-like MC3T3-E1 cells and primary murine osteoblasts were responsible to K6PC-5. K6PC-5 activated SphK1, increased sphingosine-1-phosphate (S1P) production and induced Akt phosphorylation in cultured osteoblasts. Functionally, K6PC-5 protected osteoblasts from Dex-induced apoptosis and necrosis. Such signaling and functional effects by K6PC-5 were prevented by the SphK1 inhibitor N,N-dimethylsphingosine (DMS), and by SphK1-siRNAs. On the other hand, exogenously-added S1P activated Akt and reduced Dex-induced osteoblast damages. LY294002 and MK-2206, two established Akt inhibitors, alleviated K6PC-5- or S1P-mediated osteoblast protection against Dex. Together, our results suggest that K6PC-5 alleviates Dex-induced osteoblast injuries through activating SphK1-Akt signaling. K6PC-5 might be further investigated in animal or clinical studies for its anti-glucocorticoids-associated osteonecrosis potential.

  17. Characterization of μ-oxo-(BsubPc)2 in Multiple Organic Photovoltaic Device Architectures: Comparing against and Combining with Cl-BsubPc.

    PubMed

    Castrucci, Jeffrey S; Garner, Richard K; Dang, Jeremy D; Thibau, Emmanuel; Lu, Zheng-Hong; Bender, Timothy P

    2016-09-21

    We demonstrate the first application of a unique boron subphthalocyanine (BsubPc) derivative, the oxygen bridged dimer μ-oxo-(BsubPc)2, as a multifunctional material within planar heterojunction organic photovoltaic (OPV) devices. We first explored the pairing of μ-oxo-(BsubPc)2 with well-known electron accepting and electron donating materials to explore its basic functionality. These preliminary device structures and metrics indicated that μ-oxo-(BsubPc)2 is best applied as an electron donating material when used in simple bilayer structures, as it yielded comparable OPV device efficiencies to that of the more well-established and highly optimized chloro-boron subphthalocyanine (Cl-BsubPc) OPV device structures. Thereafter we established that the HOMO/LUMO energy levels of μ-oxo-(BsubPc)2 are well-placed to apply it as a bifunctional donor/acceptor interlayer material in both energy and charge cascade OPV device architectures. Within this context, we found that μ-oxo-(BsubPc)2 was particularly effective in a charge cascade device as an interlayer between Cl-BsubPc and C70. We finally found evidence of an alloying-like effect for devices with mixed electron donor layers of (Cl-BsubPc) and μ-oxo-(BsubPc)2, achieved through co-deposition. The overarching conclusion is therefore that μ-oxo-(BsubPc)2 has the ability to improve the performance of Cl-BsubPc OPV devices and is a multifunctional material worthy of further study.

  18. Low Ubiquinone Content in Escherichia coli Causes Thiol Hypersensitivity

    PubMed Central

    Zeng, H.; Snavely, I.; Zamorano, P.; Javor, G. T.

    1998-01-01

    Thiol hypersensitivity in a mutant of Escherichia coli (IS16) was reversed by complementation with a plasmid that carried the ubiX gene. The mutant had low ubiquinone content. Complementation elevated the ubiquinone level and eliminated thiol hypersensitivity. Analysis of chromosomal ubiX genes indicated that both parent and mutant strains were ubiX mutants. The low ubiquinone content of IS16 was possibly caused by a ubiD ubiX genotype. A ubiA mutant also exhibited thiol hypersensitivity. Neither IS16 nor the ubiA mutant strain could produce alkaline phosphatase (in contrast to their parent strains) after 2 h of induction, thus showing Dsb− phenotypes. The phenomena of thiol hypersensitivity and low ubiquinone content may be linked by their connections to the periplasmic disulfide bond redox machinery. PMID:9658014

  19. Comparison microstructure and sliding wear properties of nickel-cobalt/CNT composite coatings by DC, PC and PRC current electrodeposition

    NASA Astrophysics Data System (ADS)

    Karslioglu, Ramazan; Akbulut, Hatem

    2015-10-01

    Nickel-cobalt (Ni-Co) alloys and Ni-Co/multiwalled carbon nanotube (MWCNT) composite coatings were prepared under direct current (DC), pulse current (PC) and pulse reverse current (PRC) methods. The effect of different deposition currents on the surface microstructure, crystallographic structure, microhardness, and reciprocating sliding wear behavior were investigated. MWCNT co-deposition caused to modify Ni-Co surface morphology, decrease in grain size, and increase in surface roughness, since MWCNTs effected the deposition mechanisms of Ni-Co alloy. The nanocomposite coatings deposited using PC and PRC deposition exhibited significant improvement in microhardness and wear resistance due to unique enhanced reinforcement of MWCNTs in Ni-Co coatings. Reciprocating sliding wear tests evidenced that co-deposition of MWCNTs provided effective load bearing ability and self-lubrication between the friction surfaces. However, the friction coefficient increases for all the nanocomposites produced with DC, PC and PRC methods showed to be increased. In the Ni-Co alloy coatings, the predominant wear mechanisms was delamination caused by fatigue micro cracking whereas in the MWCNT co-deposited composites wear mechanism showed abrasive grooves and plastic deformation due to decreased real contact area.

  20. The USL NASA PC R and D project: General specifications of objectives

    NASA Technical Reports Server (NTRS)

    Dominick, Wayne D. (Editor)

    1984-01-01

    Given here are the general specifications of the objectives of the University of Southwestern Louisiana Data Base Management System (USL/DBMS) NASA PC R and D Project, a project initiated to address future R and D issues related to PC-based processing environments acquired pursuant to the NASA contract work; namely, the IBM PC/XT systems.

  1. 21 CFR 868.2480 - Cutaneous carbon dioxide (PcCO2) monitor.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Cutaneous carbon dioxide (PcCO2) monitor. 868.2480... (CONTINUED) MEDICAL DEVICES ANESTHESIOLOGY DEVICES Monitoring Devices § 868.2480 Cutaneous carbon dioxide (PcCO2) monitor. (a) Identification. A cutaneous carbon dioxide (PcCO2) monitor is a noninvasive...

  2. 21 CFR 868.2480 - Cutaneous carbon dioxide (PcCO2) monitor.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Cutaneous carbon dioxide (PcCO2) monitor. 868.2480... (CONTINUED) MEDICAL DEVICES ANESTHESIOLOGY DEVICES Monitoring Devices § 868.2480 Cutaneous carbon dioxide (PcCO2) monitor. (a) Identification. A cutaneous carbon dioxide (PcCO2) monitor is a noninvasive...

  3. 21 CFR 868.2480 - Cutaneous carbon dioxide (PcCO 2) monitor.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Cutaneous carbon dioxide (PcCO 2) monitor. 868... SERVICES (CONTINUED) MEDICAL DEVICES ANESTHESIOLOGY DEVICES Monitoring Devices § 868.2480 Cutaneous carbon dioxide (PcCO 2) monitor. (a) Identification. A cutaneous carbon dioxide (PcCO2) monitor is a...

  4. 21 CFR 868.2480 - Cutaneous carbon dioxide (PcCO2) monitor.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Cutaneous carbon dioxide (PcCO2) monitor. 868.2480... (CONTINUED) MEDICAL DEVICES ANESTHESIOLOGY DEVICES Monitoring Devices § 868.2480 Cutaneous carbon dioxide (PcCO2) monitor. (a) Identification. A cutaneous carbon dioxide (PcCO2) monitor is a noninvasive...

  5. 21 CFR 868.2480 - Cutaneous carbon dioxide (PcCO 2) monitor.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Cutaneous carbon dioxide (PcCO 2) monitor. 868... SERVICES (CONTINUED) MEDICAL DEVICES ANESTHESIOLOGY DEVICES Monitoring Devices § 868.2480 Cutaneous carbon dioxide (PcCO 2) monitor. (a) Identification. A cutaneous carbon dioxide (PcCO2) monitor is a...

  6. 21 CFR 868.2500 - Cutaneous oxygen (PcO2) monitor.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Cutaneous oxygen (PcO2) monitor. 868.2500 Section... (CONTINUED) MEDICAL DEVICES ANESTHESIOLOGY DEVICES Monitoring Devices § 868.2500 Cutaneous oxygen (PcO2) monitor. (a) Identification. A cutaneous oxygen (PcO2) monitor is a noninvasive, heated sensor (e.g.,...

  7. 21 CFR 868.2500 - Cutaneous oxygen (PcO 2) monitor.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 8 2013-04-01 2013-04-01 false Cutaneous oxygen (PcO 2) monitor. 868.2500 Section... (CONTINUED) MEDICAL DEVICES ANESTHESIOLOGY DEVICES Monitoring Devices § 868.2500 Cutaneous oxygen (PcO 2) monitor. (a) Identification. A cutaneous oxygen (PcO2) monitor is a noninvasive, heated sensor (e.g.,...

  8. 21 CFR 868.2500 - Cutaneous oxygen (PcO2) monitor.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 8 2011-04-01 2011-04-01 false Cutaneous oxygen (PcO2) monitor. 868.2500 Section... (CONTINUED) MEDICAL DEVICES ANESTHESIOLOGY DEVICES Monitoring Devices § 868.2500 Cutaneous oxygen (PcO2) monitor. (a) Identification. A cutaneous oxygen (PcO2) monitor is a noninvasive, heated sensor (e.g.,...

  9. 21 CFR 868.2500 - Cutaneous oxygen (PcO 2) monitor.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 8 2014-04-01 2014-04-01 false Cutaneous oxygen (PcO 2) monitor. 868.2500 Section... (CONTINUED) MEDICAL DEVICES ANESTHESIOLOGY DEVICES Monitoring Devices § 868.2500 Cutaneous oxygen (PcO 2) monitor. (a) Identification. A cutaneous oxygen (PcO2) monitor is a noninvasive, heated sensor (e.g.,...

  10. 21 CFR 868.2500 - Cutaneous oxygen (PcO2) monitor.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 8 2012-04-01 2012-04-01 false Cutaneous oxygen (PcO2) monitor. 868.2500 Section... (CONTINUED) MEDICAL DEVICES ANESTHESIOLOGY DEVICES Monitoring Devices § 868.2500 Cutaneous oxygen (PcO2) monitor. (a) Identification. A cutaneous oxygen (PcO2) monitor is a noninvasive, heated sensor (e.g.,...

  11. Oxidative stress is involved in age-dependent spermatogenic damage of Immp2l mutant mice.

    PubMed

    George, Sunil K; Jiao, Yan; Bishop, Colin E; Lu, Baisong

    Mitochondrial reactive oxygen species (ROS) have been implicated in spermatogenic damage, although direct in vivo evidence is lacking. We recently generated a mouse in which the inner mitochondrial membrane peptidase 2-like (Immp2l) gene is mutated. This Immp2l mutation impairs the processing of signal peptide sequences from mitochondrial cytochrome c₁ and glycerol phosphate dehydrogenase 2. The mitochondria from mutant mice generate elevated levels of superoxide ion, which causes age-dependent spermatogenic damage. Here we confirm age-dependent spermatogenic damage in a new cohort of mutants, which started at the age of 10.5 months. Compared with age-matched controls, protein carbonyl content was normal in testes of 2- to 5-month-old mutants, but significantly elevated in testes of 13-month-old mutants, indicating elevated oxidative stress in the testes at the time of impaired spermatogenesis. Testicular expression of superoxide dismutases was not different between control and mutant mice, whereas that of catalase was increased in young and old mutants. The expression of cytosolic glutathione peroxidase 4 (phospholipid hydroperoxidase) in testes was significantly reduced in 13-month-old mutants, concomitant with impaired spermatogenesis. Apoptosis of all testicular populations was increased in mutant mice with spermatogenic damage. The mitochondrial DNA (mtDNA) mutation rate in germ cells of mutant mice with impaired spermatogenesis was unchanged, excluding a major role of mtDNA mutation in ROS-mediated spermatogenic damage. Our data show that increased mitochondrial ROS are one of the driving forces for spermatogenic impairment.

  12. Characterization and Complementation of a Chlorophyll-Less Dominant Mutant GL1 in Lagerstroemia indica.

    PubMed

    Wang, Shu'an; Wang, Peng; Gao, Lulu; Yang, Rutong; Li, Linfang; Zhang, Enliang; Wang, Qing; Li, Ya; Yin, Zengfang

    2017-03-09

    Crape myrtle (Lagerstroemia indica) is a woody ornamental plant popularly grown because of its long-lasting, midsummer blooms and beautiful colors. The GL1 dominant mutant is the first chlorophyll-less mutant identified in crape myrtle. It was obtained from a natural yellow leaf bud mutation. We previously revealed that leaf color of the GL1 mutant is affected by light intensity. However, the mechanism of the GL1 mutant on light response remained unclear. The acclimation response of mutant and wild-type (WT) plants was assessed in a time series after transferring from low light (LL) to high light (HL) by analyzing chlorophyll synthesis precursor content, photosynthetic performance, and gene expression. In LL conditions, coproporphyrinogen III (Coprogen III) content had the greatest amount of accumulation in the mutant compared with WT, increasing by 100%. This suggested that the yellow leaf phenotype of the GL1 dominant mutant might be caused by disruption of coproporphyrinogen III oxidase (CPO) biosynthesis. Furthermore, the candidate gene, oxygen-independent CPO (HEMN), might only affect expression of upstream genes involved in chlorophyll metabolism in the mutant. Moreover, two genes, photosystem II (PSII) 10 kDa protein (psbR) and chlorophyll a/b binding protein gene (CAB1), had decreased mRNA levels in the GL1 mutant within the first 96 h following LL/HL transfer compared with the WT. Hierarchical clustering revealed that these two genes shared a similar expression trend as the oxygen-dependent CPO (HEMF). These findings provide evidence that GL1 is highly coordinated with PSII stability and chloroplast biogenesis.

  13. Activated T cells exhibit increased uptake of silicon phthalocyanine Pc 4 and increased susceptibility to Pc 4-photodynamic therapy-mediated cell death.

    PubMed

    Soler, David C; Ohtola, Jennifer; Sugiyama, Hideaki; Rodriguez, Myriam E; Han, Ling; Oleinick, Nancy L; Lam, Minh; Baron, Elma D; Cooper, Kevin D; McCormick, Thomas S

    2016-06-08

    Photodynamic therapy (PDT) is an emerging treatment for malignant and inflammatory dermal disorders. Photoirradiation of the silicon phthalocyanine (Pc) 4 photosensitizer with red light generates singlet oxygen and other reactive oxygen species to induce cell death. We previously reported that Pc 4-PDT elicited cell death in lymphoid-derived (Jurkat) and epithelial-derived (A431) cell lines in vitro, and furthermore that Jurkat cells were more sensitive than A431 cells to treatment. In this study, we examined the effectiveness of Pc 4-PDT on primary human CD3(+) T cells in vitro. Fluorometric analyses of lysed T cells confirmed the dose-dependent uptake of Pc 4 in non-stimulated and stimulated T cells. Flow cytometric analyses measuring annexin V and propidium iodide (PI) demonstrated a dose-dependent increase of T cell apoptosis (6.6-59.9%) at Pc 4 doses ranging from 0-300 nM. Following T cell stimulation through the T cell receptor using a combination of anti-CD3 and anti-CD28 antibodies, activated T cells exhibited increased susceptibility to Pc 4-PDT-induced apoptosis (10.6-81.2%) as determined by Pc 4 fluorescence in each cell, in both non-stimulated and stimulated T cells, Pc 4 uptake increased with Pc 4 dose up to 300 nM as assessed by flow cytometry. The mean fluorescence intensity (MFI) of Pc 4 uptake measured in stimulated T cells was significantly increased over the uptake of resting T cells at each dose of Pc 4 tested (50, 100, 150 and 300 nM, p < 0.001 between 50 and 150 nM, n = 8). Treg uptake was diminished relative to other T cells. Cutaneous T cell lymphoma (CTCL) T cells appeared to take up somewhat more Pc 4 than normal resting T cells at 100 and 150 nm Pc 4. Confocal imaging revealed that Pc 4 localized in cytoplasmic organelles, with approximately half of the Pc 4 co-localized with mitochondria in T cells. Thus, Pc 4-PDT exerts an enhanced apoptotic effect on activated CD3(+) T cells that may be exploited in targeting T cell-mediated skin

  14. Chemotactic properties of Escherichia coli mutants having abnormal Ca2+ content.

    PubMed Central

    Tisa, L S; Adler, J

    1995-01-01

    The calA, calC, and calD mutants of Escherichia coli are known to be sensitive to Ca2+ (R. N. Brey and B. P. Rosen, J. Bacteriol. 139:824-834, 1979). In the absence of any added stimuli for chemotaxis, both the calC and the calD mutants swam with a tumbly bias. Both the calC and the calD mutants were defective in chemotaxis as measured by computer analysis, use of swarm plates, and capillary assays. The calA mutant was only slightly defective in motility and only slightly impaired in chemotaxis. Chemotactically wild-type cells had an intra-cellular free-Ca2+ level of about 105 nM. The intracellular free-Ca2+ levels of the mutants, as determined by use of the fluorescent Ca2+ indicator dye fura-2 or fluo-3, were about 90, about 1,130, and about 410 nM for calA, calC, and calD, respectively. Lowering the intracellular free-Ca2+ levels in wild-type cells and in the tumbly cal mutants by use of Ca2+ chelators promoted running (smooth swimming). Overexpression of CheZ (which causes dephosphorylation of CheY-phosphate) in the wild type and in the tumbly cal mutants decreased the level of tumbliness (which is caused by CheY-phosphate). The calA mutant was 4- to 10-fold more resistant than the wild type to the inhibitory effect of omega-conotoxin on chemotaxis. omega-Conotoxin had no effect on Ca2+ extrusion by wild-type E. coli; that result suggests that omega-conotoxin affects Ca2+ transport at the point of entry instead of exit. PMID:8522517

  15. Differential inhibitory effects of 2-azafluorenones on PI-PLC activation but not on PC-PLC- or PC-PLD-activation induced by histamine, PAF, PMA or A23187 in C6 glioma cells.

    PubMed

    Wang, Hai-Long; Wang, Li-Chuan; Wei, Jiann-Wu

    2013-02-28

    In this study, C6 glioma cells were used to test the effects of 2-azafluorenone and its related compounds on membrane phosphatidylinositol (PI) and phosphatidylcholine (PC) turnover. An increase of [³H]-labeled inositol phosphate (IP1) formation by histamine (100 μM) or A23187 (100 nM) via the activation of phosphatidylinositol-specific phospholipase C (PI-PLC) to breakdown labeled substrate was observed, and this effect could be partially blocked by about half at 100 μM of 2-azafluorenones. Histamine induced the increase of IP1 formation, but failed to cause an increase in extracellularly releasing of [3H]choline metabolites, or intracellular accumulation of [³H]phosphscholine. However, platelet activation factor (PAF) from 0.2 to 1 μM, and phorbol 12-myristate-13-acetate (PMA) at 1 μM caused an increase in extracellularly releasing of [³H]choline metabolites, and intracellular accumulation of [³H]phosphocholine via the activation on phosphatidylcholine (PC)-PLC. These responses of PAF and PMA were not affected by 2-azafluorenone or 4-methyl-2-azafluorenone even at high concentration (10⁻⁴ M). A23187 induced an increase of intracellular [³H]choline release via the activation of PCphospholipase D (PLD). This increasing effect of 100 nM A23187 was not affected by 2-azafluorenone or 4-methyl-2-azafluorenone even at a high concentration of 10⁻⁴ M. In summary, the inhibitory effect of 2-azafluorenone and its related compound 4-methyl-2-azafluorenone was observed selectively on PIPLC, but not on PC-PLC or PC-PLD based on changes of products after the activation of these enzymes.

  16. Triptolide Inhibited Cytotoxicity of Differentiated PC12 Cells Induced by Amyloid-Beta25–35 via the Autophagy Pathway

    PubMed Central

    Xu, Pengjuan; Li, Zhigui; Wang, Hui; Zhang, Xiaochen; Yang, Zhuo

    2015-01-01

    Evidence shows that an abnormal deposition of amyloid beta-peptide25–35 (Aβ25–35) was the primary cause of the pathogenesis of Alzheimer’s disease (AD). And the elimination of Aβ25–35 is considered an important target for the treatment of AD. Triptolide (TP), isolated from Tripterygium wilfordii Hook.f. (TWHF), has been shown to possess a broad spectrum of biological profiles, including neurotrophic and neuroprotective effects. In our study investigating the effect and potential mechanism of triptolide on cytotoxicity of differentiated rat pheochromocytoma cell line (the PC12 cell line is often used as a neuronal developmental model) induced by Amyloid-Beta25–35 (Aβ25–35), we used 3-(4, 5-dimethylthiazol-2-yl)-2, 5- diphenyltetrazolium bromide (MTT) assay, flow cytometry, Western blot, and acridine orange staining to detect whether triptolide could inhibit Aβ25–35–induced cell apoptosis. We focused on the potential role of the autophagy pathway in Aβ25–35-treated differentiated PC12 cells. Our experiments show that cell viability is significantly decreased, and the apoptosis increased in Aβ25–35-treated differentiated PC12 cells. Meanwhile, Aβ25–35 treatment increased the expression of microtubule-associated protein light chain 3 II (LC3 II), which indicates an activation of autophagy. However, triptolide could protect differentiated PC12 cells against Aβ25–35-induced cytotoxicity and attenuate Aβ25–35-induced differentiated PC12 cell apoptosis. Triptolide could also suppress the level of autophagy. In order to assess the effect of autophagy on the protective effects of triptolide in differentiated PC12 cells treated with Aβ25–35, we used 3-Methyladenine (3-MA, an autophagy inhibitor) and rapamycin (an autophagy activator). MTT assay showed that 3-MA elevated cell viability compared with the Aβ25–35-treated group and rapamycin inhibits the protection of triptolide. These results suggest that triptolide will repair the

  17. CLIPS - C LANGUAGE INTEGRATED PRODUCTION SYSTEM (IBM PC VERSION)

    NASA Technical Reports Server (NTRS)

    Riley, G.

    1994-01-01

    The C Language Integrated Production System, CLIPS, is a shell for developing expert systems. It is designed to allow artificial intelligence research, development, and delivery on conventional computers. The primary design goals for CLIPS are portability, efficiency, and functionality. For these reasons, the program is written in C. CLIPS meets or outperforms most micro- and minicomputer based artificial intelligence tools. CLIPS is a forward chaining rule-based language. The program contains an inference engine and a language syntax that provide a framework for the construction of an expert system. It also includes tools for debugging an application. CLIPS is based on the Rete algorithm, which enables very efficient pattern matching. The collection of conditions and actions to be taken if the conditions are met is constructed into a rule network. As facts are asserted either prior to or during a session, CLIPS pattern-matches the number of fields. Wildcards and variables are supported for both single and multiple fields. CLIPS syntax allows the inclusion of externally defined functions (outside functions which are written in a language other than CLIPS). CLIPS itself can be embedded in a program such that the expert system is available as a simple subroutine call. Advanced features found in CLIPS version 4.3 include an integrated microEMACS editor, the ability to generate C source code from a CLIPS rule base to produce a dedicated executable, binary load and save capabilities for CLIPS rule bases, and the utility program CRSV (Cross-Reference, Style, and Verification) designed to facilitate the development and maintenance of large rule bases. Five machine versions are available. Each machine version includes the source and the executable for that machine. The UNIX version includes the source and binaries for IBM RS/6000, Sun3 series, and Sun4 series computers. The UNIX, DEC VAX, and DEC RISC Workstation versions are line oriented. The PC version and the Macintosh

  18. Phanerochaete mutants with enhanced ligninolytic activity

    SciTech Connect

    Kakar, S.N.; Perez, A.; Gonzales, J.

    1993-06-01

    In addition to lignin, the white rot fungus Phanerochaete chrysosporium has the ability to degrade a wide spectrum of recalcitrant organopollutants in soils and aqueous media. Although some of the organic compounds are degraded under nonligninolytic conditions, most are degraded under ligninolytic conditions with the involvement of the extracellular enzymes, lignin peroxidases, and manganese-dependent peroxidases, which are produced as secondary metabolites triggered by conditions of nutrient starvation (e.g., nitrogen limitation). The fungus and its enzymes can thus provide alternative technologies for bioremediation, biopulping, biobleaching, and other industrial applications. The efficiency and effectiveness of the fungus can be enhanced by increasing production and secretion of the important enzymes in large quantities and as primary metabolites under enriched conditions. One way this can be achieved is through isolation of mutants that are deregulated or are hyperproducers or supersecretors of key enzymes under enriched conditions. Through ultraviolet-light and gamma-rays mutagenesis we have isolated a variety of mutants, some of which produce key enzymes of the ligninolytic system under high-nitrogen growth conditions. One of the mutants produced 272 units (U) of lignin peroxidases enzyme activity per liter after nine days under high nitrogen. The mutant and the parent strains produced up to 54 U/L and 62 U/L, respectively, of the enzyme activity under low-nitrogen growth conditions during this period. In some experiments the mutant showed 281 U/L of enzyme activity under high nitrogen after 17 days.

  19. Computing border bases using mutant strategies

    NASA Astrophysics Data System (ADS)

    Ullah, E.; Abbas Khan, S.

    2014-01-01

    Border bases, a generalization of Gröbner bases, have actively been addressed during recent years due to their applicability to industrial problems. In cryptography and coding theory a useful application of border based is to solve zero-dimensional systems of polynomial equations over finite fields, which motivates us for developing optimizations of the algorithms that compute border bases. In 2006, Kehrein and Kreuzer formulated the Border Basis Algorithm (BBA), an algorithm which allows the computation of border bases that relate to a degree compatible term ordering. In 2007, J. Ding et al. introduced mutant strategies bases on finding special lower degree polynomials in the ideal. The mutant strategies aim to distinguish special lower degree polynomials (mutants) from the other polynomials and give them priority in the process of generating new polynomials in the ideal. In this paper we develop hybrid algorithms that use the ideas of J. Ding et al. involving the concept of mutants to optimize the Border Basis Algorithm for solving systems of polynomial equations over finite fields. In particular, we recall a version of the Border Basis Algorithm which is actually called the Improved Border Basis Algorithm and propose two hybrid algorithms, called MBBA and IMBBA. The new mutants variants provide us space efficiency as well as time efficiency. The efficiency of these newly developed hybrid algorithms is discussed using standard cryptographic examples.

  20. Reactivation of ERK and Akt confers resistance of mutant BRAF colon cancer cells to the HSP90 inhibitor AUY922.

    PubMed

    Wang, Chun Yan; Guo, Su Tang; Wang, Jia Yu; Yan, Xu Guang; Farrelly, Margaret; Zhang, Yuan Yuan; Liu, Fen; Yari, Hamed; La, Ting; Lei, Fu Xi; Jin, Lei; Zhang, Xu Dong; Jiang, Chen Chen

    2016-08-02

    Oncogenic mutations of BRAF occur in approximately 10% of colon cancers and are associated with their resistance to clinically available therapeutic drugs and poor prognosis of the patients. Here we report that colon cancer cells with mutant BRAF are also resistant to the heat shock protein 90 (HSP90) inhibitor AUY922, and that this is caused by rebound activation of ERK and Akt. Although AUY922 triggered rapid reduction in ERK and Akt activation in both wild-type and mutant BRAF colon cancer cells, activation of ERK and Akt rebounded shortly in the latter leading to resistance of the cells to AUY922-induced apoptosis. Reactivation of ERK was associated with the persistent expression of mutant BRAF, which, despite being a client of HSP90, was only partially degraded by AUY922, whereas reactivation of Akt was related to the activity of the HSP90 co-chaperone, cell division cycle 37 (CDC37), in that knockdown of CDC37 inhibited Akt reactivation in mutant colon cancer cells treated with AUY922. In support, as a HSP90 client protein, Akt was only diminished by AUY922 in wild-type but not mutant BRAF colon cancer cells. Collectively, these results reveal that reactivation of ERK and Akt associated respectively with the activity of mutant BRAF and CDC37 renders mutant BRAF colon cancer cells resistant to AUY922, with implications of co-targeting mutant BRAF and/or CDC37 and HSP90 in the treatment of mutant BRAF colon cancers.

  1. Ionospheric signatures of cusp latitude Pc 3 pulsations

    SciTech Connect

    Engebretson, M.J.; Anderson, B.J. ); Cahill, L.J. Jr. ); Arnoldy, R.L. ); Rosenberg, T.J. ); Carpenter, D.L. ); Gail, W.B. ); Eather, R.H. )

    1990-03-01

    The authors have compared search coil magnetometer, riometer, photometer, and ELF-VLF receiver data obtained at South Pole Station and McMurdo, Antarctica, during selected days in March and April 1986. Narrow-band magnetic pulsations in the Pc 3 period range are observed simultaneously at both stations in the dayside sector during times of low interplanetary magnetic field (IMF) cone angle, but are considerably stronger at South Pole, which is located at a latitude near the nominal foot point of the daysie cusp/cleft region. Pulsations in auroral light a 427.8 nm wavelength are often observed with magnetic pulsations at South Pole, but such optical pulsations are not observed at McMurdo. When Pc 3 pulsations are present, they exhibit nearly identical frequencies, proportional to the magnitude of the IMF, in magnetometer, photometer, and ELF-VLF receiver signals at South Pole Station and in magnetometer signals at McMurdo. Singals from the 30-MHz riometer at South Pole are modulated in concert with the magnetic and optical variations during periods of broadband pulsation activity, but no riometer variations are noted during periods of narrow-band activity. Because riometers are sensitive to electrons of auroral energies (several keV and above), while the 427.8-nm photometer is sensitive to precipitation with much lower energies, they interpret these observatons as showing that precipitating magnetosheathlike electrons (with energies {le} 1 keV) at nominal dayside cleft latitudes are at times modulated with frequencies similar to those of upstream waves. They suggest that these particles may play an important role, via modification of ionospheric currents and conductivities, in the transmission of upstream wave signals into the magnetosphere and in the generation of dayside high-latitude Pc 3 pulsations.

  2. Effect of low temperature upon vitality of Saccharomyces cerevisiae phospholipid mutants.

    PubMed

    Redón, Marian; Borrull, Anna; López, Maria; Salvadó, Zoel; Cordero, Ricardo; Mas, Albert; Guillamón, José Manuel; Rozès, Nicolas

    2012-10-01

    The phospholipid metabolism of Saccharomyces cerevisiae plays a central role in its adaptation to low temperatures. In order to detect the key genes in this adaptation, various phospholipid mutants from the EUROSCARF collection of Saccharomyces cerevisiae BY4742 were tested to ascertain whether the suppression of some genes could improve the fermentation vitality of the cells at low temperature. The cell vitality and phospholipid composition of these mutants were analysed. Some knockouts improved (hmn1Δ) or impaired (cho2Δ and psd1Δ) their vitality at low temperature (13 ° C) but were not affected at optimum temperature (25 ° C). A common trait of the mutants that had some defect in vitality was a lower concentration of phosphatidylcholine and/or phosphatidylethanolamine. The supplementation with choline allowed them to recover viability, probably by synthesis through the Kennedy pathway. Hmn1Δ showed a lower concentration of phosphatidylcholine, which explains the dominant role of the de novo pathway in cellular phosphatidylethanolamine and phosphatidylcholine vs the Kennedy pathway. The absence of such genes as CRD1 or OPI3 produced important changes in phospholipid composition. Cardiolipin was not detected in crd1Δ but phosphatidylglycerol circumvents most of the functions assigned to CL. The considerable reduction in PC diminished the cell vitality of opi3Δ at both temperatures, although the decrease at 13 ° C was more marked.

  3. A versatile, PC-based gamma ray monitor.

    PubMed

    Drndarevic, V; Jevtic, N

    2008-01-01

    This paper describes a new computer-based gamma ray monitor for laboratory and field measurements. The monitor consists of a universal peripheral device, and GM counter gamma ray probe, connected via USB to the PC. The peripheral device is generic; in addition to the GM probe, it accepts sensors for spectrometric measurements as well as sensors for the measurement of other environmental parameters. Owing to its low-power consumption, it is used with laptop or palmtop computer, as a portable field measurement instrument. The proposed solution has advantages over conventional instrument: (1) open architecture; (2) user-defined functions and (3) easy network connection.

  4. Wf/pc Cycle 2 Calib: Measles Monitor

    NASA Astrophysics Data System (ADS)

    MacKenty, John

    1991-07-01

    This program takes "uniform illuminated" pictures of the earth to monitor the measles effect observed in WFPC observations. The goals of this program are: 1. Look for changes in measle numbers/characteristics 2. Have before/after images in place for decontaminations. This program will also build up a high quality flat field. The observations are done only with PC with a frequency of once every two weeks. The sequence (during a single earth occultation) is: F517N+OPEN, F517N+F122M, F517N+OPEN

  5. Wf/pc Cycle 1 CALIB:RECIPROCITY Monitor

    NASA Astrophysics Data System (ADS)

    MacKenty, John

    1990-12-01

    This program takes "uniform illuminated" pictures of the earth to monitor the "reciprocity" effect observed in earth calib observations. The goals of this program are: 1. Watch for discrete changes in this behavior 2. Look for long term variability 3. Have before/after checks in place for decontaminations safings, or UVflood. This program will also build up a high quality flat field. The observations are done only with PC with a frequency of once every week. The sequence (during a single earth occultation) is: F517N+OPEN, F517N+F122M, F517N+OPEN

  6. Casing centralization calculation in horizontal wells using PC's

    SciTech Connect

    Chen, P.; Wu, J. )

    1991-09-01

    In this paper the three-moments-equation method is used to calculate casing deflection and centralizer spacing in horizontal wells. This method, when used with a PC, makes it possible to study in detail the deflection of a casing string as a continuous beam in a horizontal wellbore. In particular, this approach makes it possible to study the near-end effect of the casing shoe on the deflection of the other spans in the casing string. Casing deflection in the transition zone where the centralizer spacing changes from one value to another and the casing deflection for an overhang at the casing shoe can also be studied. Examples are presented.

  7. Regulation of chloroplast biogenesis: the immutans mutant of Arabidopsis

    SciTech Connect

    Rodermel, Steven

    2015-11-16

    The immutans (im) variegation mutant of Arabidopsis is an ideal model to gain insight into factors that control chloroplast biogenesis. im defines the gene for PTOX, a plastoquinol terminal oxidase that participates in control of thylakoid redox. Here, we report that the im defect can be suppressed during the late stages of plant development by gigantea (gi2), which defines the gene for GIGANTEA (GI), a central component of the circadian clock that plays a poorly-understood role in diverse plant developmental processes. imgi2 mutants are late-flowering and display other well-known phenotypes associated with gi2, such as starch accumulation and resistance to oxidative stress. We show that the restoration of chloroplast biogenesis in imgi2 is caused by a developmental-specific de-repression of cytokinin signaling that involves crosstalk with signaling pathways mediated by gibberellin (GA) and SPINDLY (SPY), a GA response inhibitor. Suppression of the plastid defect in imgi2 is likely caused by a relaxation of excitation pressures in developing plastids by factors contributed by gi2, including enhanced rates of photosynthesis and increased resistance to oxidative stress. Interestingly, the suppression phenotype of imgi can be mimicked by crossing im with the starch accumulation mutant, sex1, perhaps because sex1 utilizes pathways similar to gi. We conclude that our studies provide a direct genetic linkage between GIGANTEA and chloroplast biogenesis, and we construct a model of interactions between signaling pathways mediated by gi, GA, SPY, cytokinins, and sex1 that are required for chloroplast biogenesis.

  8. Cause of Death in Korean Men with Prostate Cancer: an Analysis of Time Trends in a Nationwide Cohort

    PubMed Central

    Park, Jinsung; Suh, Beomseok; Shin, Dong Wook; Hong, Jun Hyuk

    2016-01-01

    Despite rapid increase in incidence of prostate cancer (PC) and PC survivors, there are few studies regarding competing causes of death and time trends in Asian population. We conducted a cohort study of 2% nationwide random sample of Korean National Health Insurance employees. A total of 873 patients who had received active treatments, including surgery, radiation therapy (RT) and androgen deprivation therapy (ADT), for newly diagnosed PC between 2003 and 2010 were included. The cause of death was categorized as PC, other cancers, cardiovascular disease, and other causes. During a median follow-up of 4.75 years, 29.4% (257/873) of the study population died. PC, other cancers, cardiovascular disease, and other causes were responsible for 46.3%, 35.4%, 6.6%, and 11.7%, respectively, of the decedents. Significant differences existed in the cause of death among treatment groups (P < 0.001). Only 20% and 9.5% of surgery and RT group died of PC, whereas 63.9% of ADT group died of PC. Other cancers were responsible for 56%, 74.6% and 17.8% of death in the surgery, RT and ADT group, respectively, while cardiovascular disease accounted for 4%, 6.3%, and 7.1% of death in the treatment groups. Analysis of time trends showed that PC-specific death tended to decrease (from 42.9% in 2003 to 23.1% in 2010), whereas non-PC causes tended to increase over the 8 years. Our results are valuable in overviewing causes of death and time trends in Korean PC patients, and planning future health policy for PC. PMID:27709860

  9. Heterozygous inactivation of tsc2 enhances tumorigenesis in p53 mutant zebrafish.

    PubMed

    Kim, Seok-Hyung; Kowalski, Marie L; Carson, Robert P; Bridges, L Richard; Ess, Kevin C

    2013-07-01

    Tuberous sclerosis complex (TSC) is a multi-organ disorder caused by mutations of the TSC1 or TSC2 genes. A key function of these genes is to inhibit mTORC1 (mechanistic target of rapamycin complex 1) kinase signaling. Cells deficient for TSC1 or TSC2 have increased mTORC1 signaling and give rise to benign tumors, although, as a rule, true malignancies are rarely seen. In contrast, other disorders with increased mTOR signaling typically have overt malignancies. A better understanding of genetic mechanisms that govern the transformation of benign cells to malignant ones is crucial to understand cancer pathogenesis. We generated a zebrafish model of TSC and cancer progression by placing a heterozygous mutation of the tsc2 gene in a p53 mutant background. Unlike tsc2 heterozygous mutant zebrafish, which never exhibited cancers, compound tsc2;p53 mutants had malignant tumors in multiple organs. Tumorigenesis was enhanced compared with p53 mutant zebrafish. p53 mutants also had increased mTORC1 signaling that was further enhanced in tsc2