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Sample records for afb2 g1 afg1

  1. Impairment of alveolar type-II cells involved in the toxicity of Aflatoxin G(1) in rat lung.

    PubMed

    Shen, Haitao; Lv, Ping; Xing, Xin; Xing, Lingxiao; Yan, Xia; Wang, Junling; Zhang, Xianghong

    2012-09-01

    Our previous studies showed intragastric administration of Aflatoxin G(1) (AFG(1)) could induce lung adenocarcinoma, which derived from alveolar type II cells (AT-II cells). AT-II cells contribute to Aflatoxin B(1) (AFB(1)) metabolism and also serve as the potential progenitor cells for AFB(1)-induced tumorigenesis. Thus, AT-II cells may constitute a target for AFG(1) exposure and serve as progenitor cells for tumorigenesis induced by AFG(1). The current experiment was designed to identify the acute toxicity of AFG(1) in AT-II cells following a single intratracheal administration of AFG(1). We observed inflammatory changes in the alveolar septum at days 3 and 7 after AFG(1) treatment, which resolved by 14days. We also found AFG(1) caused lamellar bodies damage in AT-II cells at days 3 and 7 post-treatment. Surfactant protein C (SP-C) expression, an AT-II cell-specific marker, was reduced at day 7 post-treatment. The structural and functional impairment in AT-II cells returned to normal by day 14. Moreover, we found that AFG(1) induced a elevation of intracellular calcium concentration [Ca(2+)]i in AT-II cells in vitro, which may contribute to the decreased SP-C expression. In conclusion, our results show AFG(1) induces structural and functional impairment in AT-II cells involved in the acute toxicity of AFG(1) in lung.

  2. Simultaneous analysis of aflatoxins B1, B2, G1, G2, M1 and ochratoxin A in breast milk by high-performance liquid chromatography/fluorescence after liquid-liquid extraction with low temperature purification (LLE-LTP).

    PubMed

    Andrade, Patricia Diniz; Gomes da Silva, Julyane Laine; Caldas, Eloisa Dutra

    2013-08-23

    The aims of this study were to optimize and validate a methodology for the simultaneous analysis of aflatoxins B1, B2, G1, G2, M1 (AFB1, AFB2, AFG1, AFG2, AFM1) and ochratoxin A (OTA) in breast milk, and to analyze these mycotoxins in samples obtained from human milk banks in the Federal District, Brazil. The optimized analytical method was based on liquid-liquid extraction with low temperature purification (3.25mL of acidified acetonitrile+0.75mL of ethyl acetate), followed by analysis by high-performance liquid chromatography with fluorescence detector (HPLC/FLD) and a photochemical post-column reactor. Limits of quantification (LOQ) ranged from 0.005 to 0.03ng/mL, recoveries from 73 to 99.5%, and relative standard deviations (RSD) from 1.8 to 17.3%. The LLE-LTP extraction method was shown to be simple and cost-effective, since no columns were needed for clean-up. Only 2 of the 224 breast milk samples analyzed were positive for the mycotoxins, both samples containing AFB2 at the LOQ level (0.005ng/mL). The identity of the mycotoxin detected was confirmed by liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). This result indicates that infants who are fed with breast milk from the milk banks are not at risk from aflatoxin and ochratoxin exposure.

  3. Aflatoxins B1, B2, G1, and G2 contamination in ground red peppers commercialized in Sanliurfa, Turkey.

    PubMed

    Karaaslan, Mehmet; Arslanğray, Yusuf

    2015-04-01

    Aflatoxins (AFs) are hepatogenic, teratogenic, imunosuppressive, and carcinogenic fungal metabolites found in feeds, nuts, wine-grapes, spices, and other grain crops. Humans are exposed to AFs via consumption of mycotoxin-contaminated foods. This study aimed to determine the prevalence of AF contamination in powdered red peppers sold in Sanliurfa. A total of 42 samples were randomly collected from retail shops, supermarkets, open bazaars, and apiaries and examined for the occurrence and levels of AFB1, AFB2, AFG1, and AFG2 toxins. AFs were determined by using an HPLC system after pre-separation utilizing immunoaffinity columns. AFs levels were below 2.5 μg/kg in 16 samples, between 2.5 and 10 μg/kg in 13 samples while 13 samples had AFs higher than the tolerable limit (10 μg/kg) according to the regulations of Turkish Food Codex and European Commission. The occurrence of AF fractions during powdered red pepper processing steps was also evaluated. According to the results obtained in this study, it was found that the highest AF accumulations in powdered red peppers start during perspiration and final drying of the products processed on soil contacted surfaces while there was no limit exceeding aflatoxin contamination in the samples produced on concrete surfaces. PMID:25773893

  4. Aflatoxins B1, B2, G1, and G2 contamination in ground red peppers commercialized in Sanliurfa, Turkey.

    PubMed

    Karaaslan, Mehmet; Arslanğray, Yusuf

    2015-04-01

    Aflatoxins (AFs) are hepatogenic, teratogenic, imunosuppressive, and carcinogenic fungal metabolites found in feeds, nuts, wine-grapes, spices, and other grain crops. Humans are exposed to AFs via consumption of mycotoxin-contaminated foods. This study aimed to determine the prevalence of AF contamination in powdered red peppers sold in Sanliurfa. A total of 42 samples were randomly collected from retail shops, supermarkets, open bazaars, and apiaries and examined for the occurrence and levels of AFB1, AFB2, AFG1, and AFG2 toxins. AFs were determined by using an HPLC system after pre-separation utilizing immunoaffinity columns. AFs levels were below 2.5 μg/kg in 16 samples, between 2.5 and 10 μg/kg in 13 samples while 13 samples had AFs higher than the tolerable limit (10 μg/kg) according to the regulations of Turkish Food Codex and European Commission. The occurrence of AF fractions during powdered red pepper processing steps was also evaluated. According to the results obtained in this study, it was found that the highest AF accumulations in powdered red peppers start during perspiration and final drying of the products processed on soil contacted surfaces while there was no limit exceeding aflatoxin contamination in the samples produced on concrete surfaces.

  5. Geothermal district G1

    SciTech Connect

    Not Available

    1988-12-01

    Geothermal District G1 includes 37 northeastern California counties and six geothermal fields: Lake City, Susanville, Litchfield, Wendel, Amedee, and Casa Diablo. Electrical generation from geothermal resources occurs in three of the fields: Wendel, Amedee, and Casa Diablo. Low-temperature geothermal projects are underway throughout the district and are described in a road log format. The ten projects described are located at Big Bend, Glass Mountain, Bieber, Alturas, Cedarville, Lake City, Honey Lake Valley, Greenville, and in Sierra and Mono Counties.

  6. Natural co-occurrence of ochratoxin A, ochratoxin B and aflatoxins in Sicilian red wines.

    PubMed

    Di Stefano, Vita; Avellone, Giuseppe; Pitonzo, Rosa; Capocchiano, Valentina Giusi; Mazza, Alessia; Cicero, Nicola; Dugo, Giacomo

    2015-01-01

    The natural occurrence of ochratoxin A, ochratoxin B, aflatoxin B1, aflatoxin B2, aflatoxin G1 and aflatoxin G2 (OTA, OTB, AFB1, AFB2, AFG1, AFG2) in red wines was investigated by HPLC/FLD after immunoaffinity column clean-up in 57 market samples produced in Sicily (Italy). The results showed a very low incidence of these mycotoxins in analysed samples, confirming the high degree of quality and safety of Sicilian red wines. The results indicated 71.9% and 64.9% positive samples for OTA and OTB respectively, with an average level of 0.13 μg l(-1), well below the European maximum permitted levels (MLs). The aflatoxin most frequently detected in the samples was AFG1, present in 57.9% of samples, while the other aflatoxins were rarely present. Recovery experiments were carried out on eight mycotoxin-free red wines spiked with OTA, OTB, AFB1, AFB2, AFG1 and AFG2 at two different levels. The limits of detection (LODs) in wines were 0.02 µg l(-1) for OTA, 0.04 µg l(-1) for OTB, 0.03 µg l(-1) for AFG1, AFG2 and AFB2, and 0.05 µg l(-1) for AFB1. A good correlation was found, with good performances in term of precision for the method.

  7. Cytochrome P450 2A13 is an efficient enzyme in metabolic activation of aflatoxin G1 in human bronchial epithelial cells.

    PubMed

    Zhang, Zhan; Yang, Xuejiao; Wang, Yun; Wang, Xichen; Lu, Huiyuan; Zhang, Xiaoming; Xiao, Xue; Li, Shushu; Wang, Xinru; Wang, Shou-Lin

    2013-09-01

    Cytochrome P450 2A13 (CYP2A13) is an extrahepatic enzyme that mainly expresses in human respiratory system, and it is reported to mediate the metabolic activation of aflatoxin B1. Due to the structural similarity, AFG1 is predicted to be metabolized by CYP2A13. However, the role of CYP2A13 in metabolic activation of AFG1 is unclear. In present study, human bronchial epithelial cells that stably express CYP2A13 (B-2A13) were used to conduct the effects of AFG1 on cytotoxicity, apoptosis, DNA damages, and their response protein expression. Low concentrations of AFG1 induced significant cytotoxicity and apoptosis, which was consistent with the increased expressions of pro-apoptotic proteins, such as C-PARP and C-caspase-3. In addition, AFG1 increased 8-OHdG and γH2AX in the nuclies and induced S phase arrest and DNA damage in B-2A13 cells, and the proteins related to DNA damage responses, such as ATM, ATR, Chk2, p53, BRCA1, and γH2AX, were activated. All the above effects were inhibited by nicotine (a substrate of CYP2A13) or 8-MOP (an inhibitor of CYP enzymes), confirming that CYP2A13 mediated the AFG1-induced cytotoxicity and DNA damages. Collectively, our findings first demonstrate that CYP2A13 might be an efficient enzyme in metabolic activation of AFG1 and helps provide a new insight into adverse effects of AFG1 in human respiratory system. PMID:23907605

  8. Monoclonal IgA Antibodies for Aflatoxin Immunoassays.

    PubMed

    Ertekin, Özlem; Pirinçci, Şerife Şeyda; Öztürk, Selma

    2016-01-01

    Antibody based techniques are widely used for the detection of aflatoxins which are potent toxins with a high rate of occurrence in many crops. We developed a murine monoclonal antibody of immunoglobulin A (IgA) isotype with a strong binding affinity to aflatoxin B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1), aflatoxin G2 (AFG2) and aflatoxin M1 (AFM1). The antibody was effectively used in immunoaffinity column (IAC) and ELISA kit development. The performance of the IACs was compatible with AOAC performance standards for affinity columns (Test Method: AOAC 991.31). The total binding capacity of the IACs containing our antibody was 111 ng, 70 ng, 114 ng and 73 ng for AFB1, AFB2, and AFG1 andAFG2, respectively. Furthermore, the recovery rates of 5 ng of each AF derivative loaded to the IACs were determined as 104.9%, 82.4%, 85.5% and 70.7% for AFB1, AFB2, AFG1 and AFG2, respectively. As for the ELISA kit developed using non-oriented, purified IgA antibody, we observed a detection range of 2-50 µg/L with 40 min total test time. The monoclonal antibody developed in this research is hitherto the first presentation of quadruple antigen binding IgA monoclonal antibodies in mycotoxin analysis and also the first study of their utilization in ELISA and IACs. IgA antibodies are valuable alternatives for immunoassay development, in terms of both sensitivity and ease of preparation, since they do not require any orientation effort. PMID:27187470

  9. Monoclonal IgA Antibodies for Aflatoxin Immunoassays

    PubMed Central

    Ertekin, Özlem; Pirinçci, Şerife Şeyda; Öztürk, Selma

    2016-01-01

    Antibody based techniques are widely used for the detection of aflatoxins which are potent toxins with a high rate of occurrence in many crops. We developed a murine monoclonal antibody of immunoglobulin A (IgA) isotype with a strong binding affinity to aflatoxin B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1), aflatoxin G2 (AFG2) and aflatoxin M1 (AFM1). The antibody was effectively used in immunoaffinity column (IAC) and ELISA kit development. The performance of the IACs was compatible with AOAC performance standards for affinity columns (Test Method: AOAC 991.31). The total binding capacity of the IACs containing our antibody was 111 ng, 70 ng, 114 ng and 73 ng for AFB1, AFB2, and AFG1 andAFG2, respectively. Furthermore, the recovery rates of 5 ng of each AF derivative loaded to the IACs were determined as 104.9%, 82.4%, 85.5% and 70.7% for AFB1, AFB2, AFG1 and AFG2, respectively. As for the ELISA kit developed using non-oriented, purified IgA antibody, we observed a detection range of 2–50 µg/L with 40 min total test time. The monoclonal antibody developed in this research is hitherto the first presentation of quadruple antigen binding IgA monoclonal antibodies in mycotoxin analysis and also the first study of their utilization in ELISA and IACs. IgA antibodies are valuable alternatives for immunoassay development, in terms of both sensitivity and ease of preparation, since they do not require any orientation effort. PMID:27187470

  10. Biotransformation of aflatoxin B1 and aflatoxin G1 in peanut meal by anaerobic solid fermentation of Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus.

    PubMed

    Chen, Yujie; Kong, Qing; Chi, Chen; Shan, Shihua; Guan, Bin

    2015-10-15

    The purpose of this study was to explore the ability of anaerobic solid fermentation of Streptococcus thermophilus and Lactobacillus delbrueckii subsp. bulgaricus to biotransform aflatoxins in peanut meal. The pH of the peanut meal was adjusted above 10, and then heated for 10 min at 100 °C, 115 °C and 121 °C. The S. thermophilus and L. delbrueckii subsp. bulgaricus were precultured together in MRS broth for 48 h at 37 °C. The heated peanut meal was mixed with precultured MRS broth containing 7.0×10(8) CFU/mL of S. thermophilus and 3.0×10(3) CFU/mL of L. delbrueckii subsp. bulgaricus with the ratio of 1 to 1 (weight to volume) and incubated in anaerobic jars at 37 °C for 3 days. The aflatoxin content in the peanut meal samples was determined by HPLC. The results showed that the peanut meal contained mainly aflatoxin B1 (AFB1) (10.5±0.64 μg/kg) and aflatoxin G1 (AFG1) (18.7±0.55 μg/kg). When heat treatment was combined with anaerobic solid fermentation, the biotransformation rate of aflatoxins in peanut meal could attain 100%. The cytotoxicity of fermented peanut meal to L929 mouse connective tissue fibroblast cells was determined by MTT assay and no significant toxicity was observed in the fermented peanut meal. Furthermore, heat treatment and anaerobic solid fermentation did not change the amino acid concentrations and profile in peanut meal.

  11. 26 CFR 1.167(g)-1 - Basis for depreciation.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 2 2010-04-01 2010-04-01 false Basis for depreciation. 1.167(g)-1 Section 1.167(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES (CONTINUED) Itemized Deductions for Individuals and Corporations § 1.167(g)-1...

  12. 26 CFR 301.6503(g)-1 - Suspension pending correction.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 18 2010-04-01 2010-04-01 false Suspension pending correction. 301.6503(g)-1 Section 301.6503(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED... Collection § 301.6503(g)-1 Suspension pending correction. The running of the periods of limitations...

  13. 26 CFR 1.475(g)-1 - Effective dates.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    .... (ii) A taxpayer may rely on the rules set out in § 1.475(c)-1T(b) (as contained in 26 CFR part 1... 26 Internal Revenue 6 2010-04-01 2010-04-01 false Effective dates. 1.475(g)-1 Section 1.475(g)-1...) INCOME TAXES Inventories § 1.475(g)-1 Effective dates. (a)-(b) (c) Section 1.475(a)-3...

  14. 26 CFR 1.280G-1 - Golden parachute payments.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ....280G-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Items Not Deductible § 1.280G-1 Golden parachute payments. The following questions... Revenue Code of 1986, as added by section 67 of the Tax Reform Act of 1984 (Pub. L. No. 98-369; 98...

  15. 26 CFR 1.514(g)-1 - Business lease indebtedness.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 26 Internal Revenue 7 2011-04-01 2009-04-01 true Business lease indebtedness. 1.514(g)-1 Section 1... (CONTINUED) INCOME TAXES (CONTINUED) Taxation of Business Income of Certain Exempt Organizations § 1.514(g)-1 Business lease indebtedness. (a) Definition. The term business lease indebtedness means, with respect...

  16. 26 CFR 1.514(g)-1 - Business lease indebtedness.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 26 Internal Revenue 7 2013-04-01 2013-04-01 false Business lease indebtedness. 1.514(g)-1 Section... TAX (CONTINUED) INCOME TAXES (CONTINUED) Taxation of Business Income of Certain Exempt Organizations § 1.514(g)-1 Business lease indebtedness. (a) Definition. The term business lease indebtedness...

  17. 26 CFR 1.514(g)-1 - Business lease indebtedness.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 7 2010-04-01 2010-04-01 true Business lease indebtedness. 1.514(g)-1 Section 1.514(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES (CONTINUED) Taxation of Business Income of Certain Exempt Organizations §...

  18. NARSTO SOS99NASH G-1 AIR CHEMISTRY DATA

    Atmospheric Science Data Center

    2014-04-25

    NARSTO SOS99NASH G-1 AIR CHEMISTRY DATA Project Title:  NARSTO ... Carbon Monoxide Ultraviolet Radiation Atmospheric Pressure Atmospheric Temperature Dew Point Upper Level Winds ... Data Guide Documents:  SOS99Nash G-1 Air Guide Related Data:  Southern Oxidants Study ...

  19. The effect of substrate, season, and agroecological zone on mycoflora and aflatoxin contamination of poultry feed from Khyber Pakhtunkhwa, Pakistan.

    PubMed

    Alam, Sahib; Shah, Hamid Ullah; Khan, Habibullah; Magan, Naresh

    2012-10-01

    To study the effects of and interactions among feed types, seasons, and agroecological zones on the total fungal viable count and aflatoxins B1 (AFB1), B2 (AFB2), G1 (AFG1), and G2 (AFG2) production in poultry feed, an experiment was conducted using three-factorial design. A total of 216 samples of poultry feed ingredients, viz. maize, wheat, rice, cotton seed meal (CSM), and finished products, that is, starter and finisher broilers' rations, were collected from Peshawar, Swat, and D. I. Khan districts of Khyber Pakhtunkhwa, Pakistan, during the winter, spring, summer, and autumn seasons of the year 2007/2008. Analysis of variance showed that there was a complex interaction among all these factors and that this influenced the total fungal viable count and relative concentrations of the aflatoxins produced. Minimum total culturable fungi (6.43 × 10³ CFUs/g) were counted in CSM from D. I. Khan region in winter season while maximum (26.68 × 10³ CFUs/g) in starter ration from Peshawar region in summer. Maximum concentrations of AFB1 (191.65 ng/g), AFB2 (86.85 ng/g), and AFG2 (89.90 ng/g) were examined during the summer season whereas the concentration of AFG1 was maximum (167.82 ng/g) in autumn in finisher ration from Peshawar region. Minimum aflatoxins were produced in the winter season across all the three agroecological zones.

  20. Aflatoxins and ochratoxin a reduction in black and white pepper by gamma radiation

    NASA Astrophysics Data System (ADS)

    Jalili, M.; Jinap, S.; Noranizan, M. A.

    2012-11-01

    Irradiation is an important means of decontamination of food commodities, especially spices. The aim of the current study was to investigate the efficacy of gamma radiation (60Co) for decontaminating ochratoxin A (OTA) and aflatoxins B1 (AFB1), B2 (AFB2), G1 (AFG1) and G2 (AFG2) residues in artificially contaminated black and white pepper samples. The moisture content of the pepper samples was set at 12% or 18%, and the applied gamma dose ranged from 5 to 30 kGy. Mycotoxin levels were determined by high-performance liquid chromatography (HPLC) after immunoaffinity column (IAC) chromatography. Both the gamma irradiation dose and moisture content showed significant effects (P<0.05) on mycotoxin reduction. The maximum toxin reductions, found at 18% moisture content and 30 kGy, were 55.2%, 50.6%, 39.2%, 47.7% and 42.9% for OTA, AFB1, AFB2, AFG1 and AFG2, respectively.

  1. Prevalence of aflatoxins in blood and urine of Egyptian infants with protein-energy malnutrition.

    PubMed

    Hatem, Nadia L; Hassab, Hoda M A; Abd Al-Rahman, Ehsan M; El-Deeb, Sami A; El-Sayed Ahmed, Rania L

    2005-03-01

    The aim of the present work was to study the presence of aflatoxins in blood and urine of infants with protein-energy malnutrition (PEM). The study was conducted on 60 infants, 30 with kwashiorkor and 30 with marasmus, with 10 age-matched healthy infants studied as a control group. Complete blood count, liver function tests, and determination of the level of aflatoxins (B1, B2, G1, G2, M1, M2, G2a, B3, GM1, P, and aflatoxicol R0) in blood and urine were carried out in all studied infants. Serum aflatoxins were detected in more infants with kwashiorkor (80%) than in those with marasmus (46.7%). The mean serum levels of total aflatoxins, AFB1, AFG1, and AFB2a, were significantly higher in infants with kwashiorkor (p <.001). Aflatoxin B1 (AFB1) was the most commonly detected type. The prevalence of aflatoxin excretion in the urine of infants with kwashiorkor was 80%, a higher value than that in infants with marasmus (46.7%). The mean urinary concentration of total aflatoxins followed the same pattern of distribution (p < .052). There were no significant differences between groups in the mean urinary concentrations of AFB1, AFG1, AFB2a, AFM1, and AFG2a. Aflatoxins were not detected in any of the serum or urine samples of the control group. Aflatoxins are highly prevalent in this study population and show a high degree of correlation with severe PEM.

  2. Functional overlap among distinct G1/S inhibitory pathways allows robust G1 arrest by yeast mating pheromones

    PubMed Central

    Pope, Patricia A.; Pryciak, Peter M.

    2013-01-01

    In budding yeast, mating pheromones arrest the cell cycle in G1 phase via a pheromone-activated Cdk-inhibitor (CKI) protein, Far1. Alternate pathways must also exist, however, because deleting the cyclin CLN2 restores pheromone arrest to far1∆ cells. Here we probe whether these alternate pathways require the G1/S transcriptional repressors Whi5 and Stb1 or the CKI protein Sic1, whose metazoan analogues (Rb or p27) antagonize cell cycle entry. Removing Whi5 and Stb1 allows partial escape from G1 arrest in far1∆ cln2∆ cells, along with partial derepression of G1/S genes, which implies a repressor-independent route for inhibiting G1/S transcription. This route likely involves pheromone-induced degradation of Tec1, a transcriptional activator of the cyclin CLN1, because Tec1 stabilization also causes partial G1 escape in far1∆ cln2∆ cells, and this is additive with Whi5/Stb1 removal. Deleting SIC1 alone strongly disrupts Far1-independent G1 arrest, revealing that inhibition of B-type cyclin-Cdk activity can empower weak arrest pathways. Of interest, although far1∆ cln2∆ sic1∆ cells escaped G1 arrest, they lost viability during pheromone exposure, indicating that G1 exit is deleterious if the arrest signal remains active. Overall our findings illustrate how multiple distinct G1/S-braking mechanisms help to prevent premature cell cycle commitment and ensure a robust signal-induced G1 arrest. PMID:24088572

  3. Strategic cell-cycle regulatory features that provide mammalian cells with tunable G1 length and reversible G1 arrest.

    PubMed

    Pfeuty, Benjamin

    2012-01-01

    Transitions between consecutive phases of the eukaryotic cell cycle are driven by the catalytic activity of selected sets of cyclin-dependent kinases (Cdks). Yet, their occurrence and precise timing is tightly scheduled by a variety of means including Cdk association with inhibitory/adaptor proteins (CKIs). Here we focus on the regulation of G1-phase duration by the end of which cells of multicelled organisms must decide whether to enter S phase or halt, and eventually then, differentiate, senesce or die to obey the homeostatic rules of their host. In mammalian cells, entry in and progression through G1 phase involve sequential phosphorylation and inactivation of the retinoblastoma Rb proteins, first, by cyclin D-Cdk4,6 with the help of CKIs of the Cip/Kip family and, next, by the cyclin E-Cdk2 complexes that are negatively regulated by Cip/Kip proteins. Using a dynamical modeling approach, we show that the very way how the Rb and Cip/Kip regulatory modules interact differentially with cyclin D-Cdk4,6 and cyclin E-Cdk2 provides to mammalian cells a powerful means to achieve an exquisitely-sensitive control of G1-phase duration and fully reversible G1 arrests. Consistently, corruption of either one of these two modules precludes G1 phase elongation and is able to convert G1 arrests from reversible to irreversible. This study unveils fundamental design principles of mammalian G1-phase regulation that are likely to confer to mammalian cells the ability to faithfully control the occurrence and timing of their division process in various conditions.

  4. [G1 and G2 chalones of the gastric mucosa].

    PubMed

    Aruin, L I; Smotrova, I A; Gorodinskaia, V S

    1984-04-01

    A study was made of the action of human gastric mucosa G1 and G2 chalones on cellular regeneration of mouse gastric mucosa and of the duration of their maximal effect. Chalone fractions were obtained from the mucous membranes of 21 stomachs resected for peptic ulcer by the method of fractional ethanol precipitation. The data indicate that the maximal inhibitory action of G1 chalone occurs in 3, whereas that of chalone G2 in 6 hours. Some specificity of the action of chalones was discovered depending on the part of the gastric mucosa from which they were obtained. PMID:6232965

  5. Rapid biodegradation of organophosphorus pesticides by Stenotrophomonas sp. G1.

    PubMed

    Deng, Shuyan; Chen, Yao; Wang, Daosheng; Shi, Taozhong; Wu, Xiangwei; Ma, Xin; Li, Xiangqiong; Hua, Rimao; Tang, Xinyun; Li, Qing X

    2015-10-30

    Organophosphorus insecticides have been widely used, which are highly poisonous and cause serious concerns over food safety and environmental pollution. A bacterial strain being capable of degrading O,O-dialkyl phosphorothioate and O,O-dialkyl phosphate insecticides, designated as G1, was isolated from sludge collected at the drain outlet of a chlorpyrifos manufacture plant. Physiological and biochemical characteristics and 16S rDNA gene sequence analysis suggested that strain G1 belongs to the genus Stenotrophomonas. At an initial concentration of 50 mg/L, strain G1 degraded 100% of methyl parathion, methyl paraoxon, diazinon, and phoxim, 95% of parathion, 63% of chlorpyrifos, 38% of profenofos, and 34% of triazophos in 24 h. Orthogonal experiments showed that the optimum conditions were an inoculum volume of 20% (v/v), a substrate concentration of 50 mg/L, and an incubation temperature in 40 °C. p-Nitrophenol was detected as the metabolite of methyl parathion, for which intracellular methyl parathion hydrolase was responsible. Strain G1 can efficiently degrade eight organophosphorus pesticides (OPs) and is a very excellent candidate for applications in OP pollution remediation.

  6. SNAILs promote G1 phase in selected cancer cells.

    PubMed

    Wu, Ya-Lan; Xue, Jian-Xin; Zhou, Lin; Deng, Lei; Shang, Yan-Na; Liu, Fang; Mo, Xian-Ming; Lu, You

    2015-11-01

    Cells can acquire a stem-like cell phenotype through epithelial-mesenchymal transition (EMT). However, it is not known which of the stem-like cancer cells are generated by these phenotype transitions. We studied the EMT-inducing roles of SNAILs (the key inducers for the onset of EMT) in selected cancer cells (lung cancer cell line with relatively stable genome), in order to provide more implications for the investigation of EMT-related phenotype transitions in cancer. However, SNAILs fail to induce completed EMT. In addition, we proved that Snail accelerates the early G1 phase whereas Slug accelerates the late G1 phase. Blocking G1 phase is one of the basic conditions for the onset of EMT-related phenotype transitions (e.g., metastasis, acquiring stemness). The discovery of this unexpected phenomenon (promoting G1 phase) typically reveals the heterogeneity of cancer cells. The onset of EMT-related phenotype transitions in cancer needs not only the induction and activation of SNAILs, but also some particular heredity alterations (genetic or epigenetic alterations, which cause heterogeneity). The new connection between heredity alteration (heterogeneity) and phenotype transition suggests a novel treatment strategy, the heredity alteration-directed specific target therapy. Further investigations need to be conducted to study the relevant heredity alterations.

  7. NARSTO EPA SS HOUSTON TEXAQS2000 DOE G-1 DATA

    Atmospheric Science Data Center

    2014-04-25

    NARSTO EPA SS HOUSTON TEXAQS2000 DOE G-1 DATA Project Title:  NARSTO ... Fluorometer Ion Chromatograph Location:  Houston, Texas Spatial Resolution:  Point Measurements ... Parameters:  Atmospheric Pressure Air Temperature Dew Point Flight Level Winds Nitrogen Oxides Ozone ...

  8. Rapid biodegradation of organophosphorus pesticides by Stenotrophomonas sp. G1.

    PubMed

    Deng, Shuyan; Chen, Yao; Wang, Daosheng; Shi, Taozhong; Wu, Xiangwei; Ma, Xin; Li, Xiangqiong; Hua, Rimao; Tang, Xinyun; Li, Qing X

    2015-10-30

    Organophosphorus insecticides have been widely used, which are highly poisonous and cause serious concerns over food safety and environmental pollution. A bacterial strain being capable of degrading O,O-dialkyl phosphorothioate and O,O-dialkyl phosphate insecticides, designated as G1, was isolated from sludge collected at the drain outlet of a chlorpyrifos manufacture plant. Physiological and biochemical characteristics and 16S rDNA gene sequence analysis suggested that strain G1 belongs to the genus Stenotrophomonas. At an initial concentration of 50 mg/L, strain G1 degraded 100% of methyl parathion, methyl paraoxon, diazinon, and phoxim, 95% of parathion, 63% of chlorpyrifos, 38% of profenofos, and 34% of triazophos in 24 h. Orthogonal experiments showed that the optimum conditions were an inoculum volume of 20% (v/v), a substrate concentration of 50 mg/L, and an incubation temperature in 40 °C. p-Nitrophenol was detected as the metabolite of methyl parathion, for which intracellular methyl parathion hydrolase was responsible. Strain G1 can efficiently degrade eight organophosphorus pesticides (OPs) and is a very excellent candidate for applications in OP pollution remediation. PMID:25938642

  9. 26 CFR 1.149(g)-1 - Hedge bonds.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ...) INCOME TAXES (CONTINUED) Tax Exemption Requirements for State and Local Bonds § 1.149(g)-1 Hedge bonds... of replacement proceeds (other than amounts in a bona fide debt service fund or a reasonably required... realize debt service savings or to relieve the issuer of significantly burdensome document provisions,...

  10. 26 CFR 1.149(g)-1 - Hedge bonds.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ...) INCOME TAXES (CONTINUED) Tax Exemption Requirements for State and Local Bonds § 1.149(g)-1 Hedge bonds... of replacement proceeds (other than amounts in a bona fide debt service fund or a reasonably required... realize debt service savings or to relieve the issuer of significantly burdensome document provisions,...

  11. Occurrence of Aflatoxins Contamination in Brown Rice from Pakistan

    PubMed Central

    ASGHAR, Muhammad Asif; IQBAL, Javed; AHMED, Aftab; KHAN, Mobeen Ahmed

    2014-01-01

    Abstract Background The objective of this study was to determine the distribution of an economically–important class of mycotoxins, the aflatoxins (AFs) in Pakistani Brown Rice. Methods A total of 262 of brown rice samples were collected from different vendors during July 2006 to June 2011. Samples were analyzed for the occurrence of aflatoxin B1 (AFB1), B2 (AFB2), G1 (AFG1) and G2 (AFG2) by thin layer chromatography (TLC) technique. Results AFB1 was detected in 250 (95.4%) samples, whereas AFB2 was detected in 20 (7.6%) samples. Furthermore, AFG1 and AFG2 were not found in any sample. The contamination range of AFB1 and AFB2 was found 1.07–24.65 μg/kg and 0.52–2.62 μg/kg, respectively. Total AFs were quantified in 250 (95.4%) samples with an average of 3.89 μg/kg and contamination range was noted to be between 1.07–27.27 μg/kg. The overall results indicated that in 12 (4.6%) samples, AFs were not found within detectable limits. Furthermore, in 188 (71.7%) samples, AFs level was found below than maximum tolerated levels (MTL) as recommended by the European Union (4 μg/kg). Moreover, in 61 (23.3%) samples, AFs range was found between 4–20 μg/kg, which were fit for human consumption as per MTL (20 μg/kg) assigned by USA (FDA and FAO) and Pakistan (PSQCA). While only one sample (27.27 μg/kg) exceeded the above mention regulation limits. Conclusion Low level of AFs occurs frequently in brown rice, and can be improved using proper harvesting practices, storage and transportation conditions. The small quantities of AFs warrant performing further investigation, monitoring and routine analysis on regular basis. PMID:25988088

  12. Survey of aflatoxins in retail samples of whole and ground black and white peppercorns.

    PubMed

    Adzahan, N; Jalili, M; Jinap, S

    2009-01-01

    A total of 126 local and imported samples of commercial white and black pepper in Malaysia were analysed for aflatoxins B1, B2, G1 and G2 (AFB1, AFB2, AFG1, AFG2) content using high-performance liquid chromatography (HPLC) with a fluorescence detector (FD). An acetonitrile-methanol-water (17 : 29 : 54; v/v) mixture was used as a mobile phase and clean-up was using an immunoaffinity column (IAC). Seventy out of 126 (55.5%) samples were contaminated with total aflatoxins, although only low levels of aflatoxins were found ranging from 0.1 to 4.9 ng g(-1). Aflatoxin B1 showed the highest incidence of contamination and was found in all contaminated samples. There was a significant difference between type of samples and different brands (p < 0.05). The results showed black peppers were more contaminated than white peppers. PMID:24785182

  13. Compass Measurement of g1 and QCD Fits

    NASA Astrophysics Data System (ADS)

    Kunne, Fabienne

    2016-02-01

    We present the latest COMPASS results on the proton spin structure function g1p(x) at 200GeV. The data improve the statistical precision by a factor of ˜2 at low x. A reevaluation of the Bjorken sum rule based on COMPASS proton and deuteron data confirms its validation to a 9% accuracy. Finally, results from a global NLO QCD fit of g1 world data are shown. The extracted spin singlet distribution leads to an integrated value of 0.26 < ΔΣ < 0.34 at Q2 = 3 (GeV/c)2. The large uncertainty is mainly driven by the unknown shape of the distribution.

  14. Crack azimuths on Europa: The G1 lineament sequence revisited

    USGS Publications Warehouse

    Sarid, A.R.; Greenberg, R.; Hoppa, G.V.; Brown, D.M.; Geissler, P.

    2005-01-01

    The tectonic sequence in the anti-jovian area covered by regional mapping images from Galileo's orbit E15 is determined from a study of cross-cutting relationships among lineament features. The sequence is used to test earlier results from orbit G1, based on lower resolution images, which appeared to display a progressive change in azimuthal orientation over about 90?? in a clockwise sense. Such a progression is consistent with expected stress variations that would accompany plausible non-synchronous rotation. The more recent data provide a more complete record than the G1 data did. We find that to fit the sequence into a continual clockwise change of orientation would require at least 1000?? (> 5 cycles) of azimuthal rotation. If due to non-synchronous rotation of Europa, this result implies that we are seeing back further into the tectonic record than the G1 results had suggested. The three sets of orientations found by Geissler et al. now appear to have been spaced out over several cycles, not during a fraction of one cycle. While our more complete sequence of lineament formation is consistent with non-synchronous rotation, a statistical test shows that it cannot be construed as independent evidence. Other lines of evidence do support non-synchronous rotation, but azimuths of crack sequences do not show it, probably because only a couple of cracks form in a given region in any given non-synchronous rotation period. ?? 2004 Elsevier Inc. All rights reserved.

  15. Centrosomal AKAP350 modulates the G1/S transition

    PubMed Central

    Mattaloni, Stella M; Ferretti, Anabela C; Tonucci, Facundo M; Favre, Cristián; Goldenring, James R; Larocca, M Cecilia

    2013-01-01

    AKAP350 (AKAP450/AKAP9/CG-NAP) is an A-kinase anchoring protein, which recruits multiple signaling proteins to the Golgi apparatus and the centrosomes. Several proteins recruited to the centrosomes by this scaffold participate in the regulation of the cell cycle. Previous studies indicated that AKAP350 participates in centrosome duplication. In the present study we specifically assessed the role of AKAP350 in the progression of the cell cycle. Our results showed that interference with AKAP350 expression inhibits G1/S transition, decreasing the initiation of both DNA synthesis and centrosome duplication. We identified an AKAP350 carboxyl-terminal domain (AKAP350CTD), which contained the centrosomal targeting domain of AKAP350 and induced the initiation of DNA synthesis. Nevertheless, AKAP350CTD expression did not induce centrosomal duplication. AKAP350CTD partially delocalized endogenous AKAP350 from the centrosomes, but increased the centrosomal levels of the cyclin-dependent kinase 2 (Cdk2). Accordingly, the expression of this AKAP350 domain increased the endogenous phosphorylation of nucleophosmin by Cdk2, which occurs at the G1/S transition and is a marker of the centrosomal activity of the cyclin E-Cdk2 complex. Cdk2 recruitment to the centrosomes is a necessary event for the development of the G1/S transition. Altogether, our results indicate that AKAP350 facilitates the initiation of DNA synthesis by scaffolding Cdk2 to the centrosomes, and enabling its specific activity at this organelle. Although this mechanism could also be involved in AKAP350-dependent modulation of centrosomal duplication, it is not sufficient to account for this process. PMID:24475373

  16. Cactin is essential for G1 progression in Toxoplasma gondii

    PubMed Central

    Szatanek, Tomasz; Anderson-White, Brooke R.; Faugno-Fusci, David M.; White, Michael; Saeij, Jeroen P.J.; Gubbels, Marc-Jan

    2012-01-01

    Toxoplasma gondii is an obligate intracellular protozoan parasite whose rapid lytic replication cycles define its pathogenicity. We identified a temperature sensitive growth mutant, FV-P6, which irreversibly arrests before the middle of the G1 stage of the tachyzoite cell cycle. This arrest is caused by a point mutation in a gene conserved across eukaryotes, Cactin, whose product localizes to the nucleus. To elucidate the role of TgCactin we performed genome-wide expression profiling. Besides the expected G1 expression profile, many genes associated with the extracellular state as well as with the bradyzoite cyst stage were identified. Consistent with these profiles were the expression of AP2 transcription factors typically associated with extracellular and bradyzoite stage parasites. This suggests a role for TgCactin in control of gene expression. Since TgCactin does not contain any functionally defined domains we reasoned TgCactin exerts its function through interactions with other proteins. In support of this model we demonstrated that TgCactin is present in a protein complex and can oligomerize. Taken together, these results suggest that TgCactin acts as a pivotal protein potentially regulating gene expression at several transition points in parasite development. PMID:22486860

  17. Development and validation of a high-performance liquid chromatography method with post-column derivatization for the detection of aflatoxins in cereals and grains.

    PubMed

    Asghar, Muhammad Asif; Iqbal, Javed; Ahmed, Aftab; Khan, Mobeen Ahmed; Shamsuddin, Zuzzer Ali; Jamil, Khalid

    2016-06-01

    A novel, reliable and rapid high-performance liquid chromatography (HPLC) method with post-column derivatization was developed and validated. The HPLC method was used for the simultaneous determination of aflatoxin B1 (AFB1), B2 (AFB2), G1 (AFG1) and G2 (AFG2) in various cereals and grains. Samples were extracted with 80:20 (v/v) methanol:water and purified using C18 (40-63 μm) solid-phase extraction cartridges. AFs were separated using a LiChroCART-RP-18 (5 μm, 250 × 4.0 mm(2)) column. The mobile phase consisted of methanol:acetonitrile:buffer (17.5:17.5:65 v/v) (pH 7.4) delivered at the flow rate of 1.0 mL min(-1) The fluorescence of each AF was detected at λex = 365 nm and λem = 435 nm. All four AFs were properly resolved within the total run time of 20 min. The established method was extensively validated as a final verification of the method development by the evaluation of selectivity (AFB1, AFB2, AFG1 and AFG2), linearity (R(2) ≥ 0.9994), precision (average SD ≤ 2.79), accuracy (relative mean error ≤ -5.51), robustness (p < 0.0080), ruggedness (p < 0.0100) and average recoveries (89.2-97.8%). The limits of quantification of AFB1, AFB2, AFG1 and AFG2 were 0.080, 0.073, 0.062 and 0.066 ng g(-1), respectively. Finally, the developed method was applied for the analysis of AFs in 45 samples comprising rice (n = 20), wheat (n = 15) and maize (n = 10). The results showed that 65% of rice, 20% of wheat and 80% of maize samples were found contaminated with AFs. Thus, according to the achieved results, it is suggested that the newly developed HPLC method could be effectively applied for the routine analysis of the AFs in different cereals and grains.

  18. LUMIX DMC-G1 - New Pleasantness of the Camera with Interchangeable Lenses That G1 Provides -

    NASA Astrophysics Data System (ADS)

    Ueda, Hiroshi; Hataji, Shinji; Morishita, Seiki; Inoue, Yoshiyuki

    Panasonic introduced in October 2008 the "LUMIX DMC-G1", which is adopting the Micro Four Thirds standard. This camera was a hot topic from the time of the announcement in September and after the sales start it was highly evaluated not only due to its small size and light weight, but also due to the compact camera like easy operation realized by the mirror-less construction and due to the performance, which is on the same level like conventional consumer SLR cameras. Within this chapter we will explain about the technology behind the high-speed AF, which was seen as difficult to realize in a system based on Live View, and the high resolution Live View Finder, as well as about the new challenge of color variations, presented for the first time for an interchangeable lens camera.

  19. G0/G1 Switch Gene 2 Regulates Cardiac Lipolysis*

    PubMed Central

    Heier, Christoph; Radner, Franz P. W.; Moustafa, Tarek; Schreiber, Renate; Grond, Susanne; Eichmann, Thomas O.; Schweiger, Martina; Schmidt, Albrecht; Cerk, Ines K.; Oberer, Monika; Theussl, H.-Christian; Wojciechowski, Jacek; Penninger, Josef M.; Zimmermann, Robert; Zechner, Rudolf

    2015-01-01

    The anabolism and catabolism of myocardial triacylglycerol (TAG) stores are important processes for normal cardiac function. TAG synthesis detoxifies and stockpiles fatty acids to prevent lipotoxicity, whereas TAG hydrolysis (lipolysis) remobilizes fatty acids from endogenous storage pools as energy substrates, signaling molecules, or precursors for complex lipids. This study focused on the role of G0/G1 switch 2 (G0S2) protein, which was previously shown to inhibit the principal TAG hydrolase adipose triglyceride lipase (ATGL), in the regulation of cardiac lipolysis. Using wild-type and mutant mice, we show the following: (i) G0S2 is expressed in the heart and regulated by the nutritional status with highest expression levels after re-feeding. (ii) Cardiac-specific overexpression of G0S2 inhibits cardiac lipolysis by direct protein-protein interaction with ATGL. This leads to severe cardiac steatosis. The steatotic hearts caused by G0S2 overexpression are less prone to fibrotic remodeling or cardiac dysfunction than hearts with a lipolytic defect due to ATGL deficiency. (iii) Conversely to the phenotype of transgenic mice, G0S2 deficiency results in a de-repression of cardiac lipolysis and decreased cardiac TAG content. We conclude that G0S2 acts as a potent ATGL inhibitor in the heart modulating cardiac substrate utilization by regulating cardiac lipolysis. PMID:26350455

  20. Early Evolution of Disrupted Asteroid P/2016 G1 (PANSTARRS)

    NASA Astrophysics Data System (ADS)

    Moreno, F.; Licandro, J.; Cabrera-Lavers, A.; Pozuelos, F. J.

    2016-08-01

    We present deep imaging observations of activated asteroid P/2016 G1 (PANSTARRS) using the 10.4 m Gran Telescopio Canarias (GTC) from 2016 late April to early June. The images are best interpreted as the result of a relatively short-duration event with an onset of about {350}-30+10 days before perihelion (i.e., around 2016 February 10), starting sharply and decreasing with {24}-7+10 days (HWHM). The results of the modeling imply that the emission of ˜1.7 × 107 kg of dust, if composed of particles of 1 μm to 1 cm in radius, is distributed following a power law of index -3 and having a geometric albedo of 0.15. A detailed fitting of a conspicuous westward feature in the head of the comet-like object indicates that a significant fraction of the dust was ejected along a privileged direction right at the beginning of the event, which suggests that the parent body has possibly suffered an impact followed by a partial or total disruption. From the limiting magnitude reachable with the instrumental setup, and assuming a geometric albedo of 0.15 for the parent body, an upper limit for the size of possible fragment debris of ˜50 m in radius is derived.

  1. Early Evolution of Disrupted Asteroid P/2016 G1 (PANSTARRS)

    NASA Astrophysics Data System (ADS)

    Moreno, F.; Licandro, J.; Cabrera-Lavers, A.; Pozuelos, F. J.

    2016-08-01

    We present deep imaging observations of activated asteroid P/2016 G1 (PANSTARRS) using the 10.4 m Gran Telescopio Canarias (GTC) from 2016 late April to early June. The images are best interpreted as the result of a relatively short-duration event with an onset of about {350}-30+10 days before perihelion (i.e., around 2016 February 10), starting sharply and decreasing with {24}-7+10 days (HWHM). The results of the modeling imply that the emission of ˜1.7 × 107 kg of dust, if composed of particles of 1 μm to 1 cm in radius, is distributed following a power law of index ‑3 and having a geometric albedo of 0.15. A detailed fitting of a conspicuous westward feature in the head of the comet-like object indicates that a significant fraction of the dust was ejected along a privileged direction right at the beginning of the event, which suggests that the parent body has possibly suffered an impact followed by a partial or total disruption. From the limiting magnitude reachable with the instrumental setup, and assuming a geometric albedo of 0.15 for the parent body, an upper limit for the size of possible fragment debris of ˜50 m in radius is derived.

  2. 26 CFR 1.904(g)-1 - Overall domestic loss and the overall domestic loss account.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... loss account. 1.904(g)-1 Section 1.904(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF... States § 1.904(g)-1 Overall domestic loss and the overall domestic loss account. For further guidance, see § 1.904(g)-1T....

  3. Preferential synthesis of the G1m(1) allotype of IgG1 in the central nervous system of multiple sclerosis patients.

    PubMed

    Salier, J P; Goust, J M; Pandey, J P; Fudenberg, H H

    1981-09-18

    Quantitations of the G1m(1) and G1m(3) allotypic determinants of human immunoglobulin G were performed by radioimmunoassay on cerebrospinal fluid and serum samples from patients with multiple sclerosis and from patients with other neurological disorders. In multiple sclerosis patients that were heterozygous for these determinants, G1m(1) concentration in the cerebrospinal fluid was greatly increased-reflected by an increased ratio of G1m(1)-in comparison with that of patients with other neurological disorders. These results suggest that in the heterozygous multiple sclerosis patients, most of the plasma cells in the central nervous system that secrete oligoclonal immunoglobulin G preferentially synthesize G1m(1) IgG1 molecules. PMID:6973823

  4. [Determination of aflatoxins in cereals and oils by liquid chromatography-triple quadrupole tandem mass spectrometry].

    PubMed

    Wang, Xiupin; Li, Peiwu; Yang, Yang; Zhang, Wen; Zhang, Qi; Fan, Sufang; Yu, Li; Wang, Lin; Chen, Xiaomei; Li, Ying; Jiang, Jun

    2011-06-01

    The high performance liquid chromatography (HPLC)-triple quadrupole tandem mass spectrometry was applied for the determination of the aflatoxins: B1 (AFB1), B2 (AFB2), G1 (AFG1) and G2 (AFG2), in cereals and oils. The samples were first extracted by ultrasonic method. The optimized conditions of ultrasonic extraction were as follows: temperature of 50 degrees C, extraction time of 3 min, methanol-water (containing 40 g/L NaCl) (80: 20, v/v) solution as the medium and a ratio of sample to solvent of 1 : 3 (g: mL). The extracts were then purified using an immunoaffinity column. The separation was performed on a C18 column with mobile phases of 10 mmol/L ammonium acetate and methanol in gradient elution. The sensitive detection of the four AFT compounds by electrospray ionization mass spectrometry (ESI-MS) was carried out in selected reaction monitoring (SRM) mode with aflatoxin M1 (AFM1) as the internal standard. Under the optimized conditions, the limits of detection of AFB1, AFB2, AFG1 and AFG2 were 0.002, 0.004, 0.004 and 0.012 microg/kg, respectively. The recoveries of aflatoxins in different spiked cereals and oils were in the range from 87% to 111%. The intra-day and inter-day precisions were not more than 6.7% and 5.6%, respectively. In comparison with the external standard method, this method can effectively inhibit the matrix effects, and greatly improve the accuracy.

  5. Determination for multiple mycotoxins in agricultural products using HPLC-MS/MS via a multiple antibody immunoaffinity column.

    PubMed

    Zhang, Zhaowei; Hu, Xiaofeng; Zhang, Qi; Li, Peiwu

    2016-05-15

    Mycotoxins usually found in agricultural products such as peanut, corn, and wheat, are a serious threat to human health and their detection requires multiplexed and sensitive analysis methods. Herein, a simultaneous determination for aflatoxin B1, B2, G1, G2, ochratoxin A, zearalanone and T-2 toxin was investigated using high performance liquid chromatography coupled with tandem mass spectrometry in a single run via a home-made multiple immunoaffinity column. Four monoclonal antibodies were produced in our lab against aflatoxins, ochratoxin A, zearalanone and T-2 toxin, respectively, then combined as a pool and bound to Sepharose-4B for affinity chromatography. Seven mycotoxins were effectively extracted from the agricultural product samples by using acetonitrile/water/acetic acid (80:19:1, v/v/v) Then, the extraction was cleanup by multiple immunoaffinity column. This method demonstrated a considerable linear range of 0.30-25, 0.12-20, 0.30-20, 0.12-20, 0.60-30, 0.30-25, and 1.2-40μgkg(-1)and lower limits of detection at 0.1, 0.04, 0.1, 0.04, 0.2, 0.1 and 0.4μgkg(-1) for AFB1, AFB2, AFG1, AFG2, OTA, ZEN and T-2, respectively, in comparison with previously reported methods, as well as excellent recoveries. The mIAC capacity for AFB1, AFB2, AFG1, AFG2, OTA, ZEN, and T-2 were 187, 181, 153, 151, 105, 130, 88ng, respectively. It was found that all of the 7 mycotoxins were present in 90 agricultural product samples. The proposed method meets the requirements for rapid sample preparation and highly sensitive identification of multiple mycotoxins in agricultural product and food safety. This method provides a promising alternative with high throughput and high sensitivity for rapid analysis of seven mycotoxins in the monitoring of food safety. PMID:26948441

  6. 26 CFR 1.143(g)-1 - Requirements related to arbitrage.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... in whole. Except as provided in paragraph (d)(4) of this section, issuers may apply § 1.143(g)-1, in... 26 Internal Revenue 2 2014-04-01 2014-04-01 false Requirements related to arbitrage. 1.143(g)-1 Section 1.143(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY...

  7. 16 CFR Appendix G1 to Part 305 - Furnaces-Gas

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 16 Commercial Practices 1 2014-01-01 2014-01-01 false Furnaces-Gas G1 Appendix G1 to Part 305... RULEâ) Appendix G1 to Part 305—Furnaces—Gas Furnace type Range of annual fuel utilization efficiencies (AFUEs) Low High Gas Furnaces Manufactured Before the Compliance Date of DOE Regional...

  8. 26 CFR 1.642(g)-1 - Disallowance of double deductions; in general.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ....642(g)-1 Section 1.642(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Estates, Trusts, and Beneficiaries § 1.642(g)-1 Disallowance..., after a statement is filed under section 642(g) with respect to a particular item or portion of an...

  9. 26 CFR 1.430(g)-1 - Valuation date and valuation of plan assets.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ....430(g)-1 Section 1.430(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Certain Stock Options § 1.430(g)-1 Valuation date and... valuation date and the valuation of a plan's assets for a plan year under section 430(g). Section 430...

  10. 26 CFR 25.2523(g)-1 - Special rule for charitable remainder trusts.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    .... 25.2523(g)-1 Section 25.2523(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE....2523(g)-1 Special rule for charitable remainder trusts. (a) In general. (1) With respect to gifts made... passing to the spouse qualifies for a marital deduction under section 2523(g) and the value of...

  11. 26 CFR 301.6511(g)-1 - Special rule for partnership items of federally registered partnerships.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... registered partnerships. 301.6511(g)-1 Section 301.6511(g)-1 Internal Revenue INTERNAL REVENUE SERVICE... Limitations on Assessment and Collection § 301.6511(g)-1 Special rule for partnership items of federally...(g) must also be taken into account in applying the various special periods of limitation...

  12. 26 CFR 301.6223(g)-1 - Responsibilities of the tax matters partner.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... contained in 26 CFR part 1, revised April 1, 2001. .... 301.6223(g)-1 Section 301.6223(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE....6223(g)-1 Responsibilities of the tax matters partner. (a) Notices described in section...

  13. 26 CFR 1.143(g)-1 - Requirements related to arbitrage.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 2 2010-04-01 2010-04-01 false Requirements related to arbitrage. 1.143(g)-1 Section 1.143(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED....143(g)-1 Requirements related to arbitrage. (a) In general. Under section 143, for an issue to be...

  14. 26 CFR 1.430(g)-1 - Valuation date and valuation of plan assets.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ....430(g)-1 Section 1.430(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES (CONTINUED) Certain Stock Options § 1.430(g)-1 Valuation date... plan's valuation date and the valuation of a plan's assets for a plan year under section...

  15. 26 CFR 25.2523(g)-1 - Special rule for charitable remainder trusts.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... passing to the spouse qualifies for a marital deduction under section 2523(g) and the value of the.... 25.2523(g)-1 Section 25.2523(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE....2523(g)-1 Special rule for charitable remainder trusts. (a) In general. (1) With respect to gifts...

  16. The flare activity of G1 718 = BD + 22 deg 3406

    NASA Astrophysics Data System (ADS)

    Chugainov, P. F.

    The results of 58.8 hours of photoelectric U-band observations of the red dwarf G1 718 are presented. The observations were carried out in order to confirm the conclusion of Mahmoud and Soliman (1980) that G1 718 is experiencing high flare activity. It is shown that the mean rate of energy release from G1 718 is approximately the same as that of G1 825. Both G1 718 and G1 825 show a deviation from the correlation between mean energy release rate and luminosity which has been established for young red dwarfs. No BY Dra variations were found for G1 718. The complete observational results are given in a table.

  17. Functional role of eukaryotic translation initiation factor 4 gamma 1 (EIF4G1) in NSCLC

    PubMed Central

    Cao, Yueyu; Wei, Mengdan; Li, Bing; Liu, Yali; Lu, Ying; Tang, Zhipeng; Lu, Tianbao; Yin, Yujiao; Qin, Zhiqiang; Xu, Zengguang

    2016-01-01

    Eukaryotic translation initiation factor 4 gamma 1(EIF4G1) is related to tumorigenesis and tumor progression. However, its role and the underlying mechanisms in the regulation of tumor development in non–small cell lung cancers (NSCLC) remain largely unknown. Here we report that the levels of EIF4G1 expression are much higher in NSCLC cell lines and tumor tissues than those in the normal lung cells and adjacent normal tissues from the same patients. Using shRNA to knock down EIF4G1 expression stably, we found EIF4G1 required for NSCLC cell proliferation, anchorage-independent growth, migration and invasion. Furthermore, silencing of EIF4G1 induces NSCLC cell apoptosis and causes G0/G1 cell cycle arrest. To identify the partner protein network of EIF4G1 in NSCLC cells, we found that Ubiquitin-specific protease 10 (USP10) can directly interacts with EIF4G1, while acting as a negative regulator for EIF4G1-mediated functions. Together, our results indicate that EIF4G1 functions as an oncoprotein during NSCLC development, which may represent a novel and promising therapeutic target in lung cancer. PMID:27003362

  18. Functional role of eukaryotic translation initiation factor 4 gamma 1 (EIF4G1) in NSCLC.

    PubMed

    Cao, Yueyu; Wei, Mengdan; Li, Bing; Liu, Yali; Lu, Ying; Tang, Zhipeng; Lu, Tianbao; Yin, Yujiao; Qin, Zhiqiang; Xu, Zengguang

    2016-04-26

    Eukaryotic translation initiation factor 4 gamma 1(EIF4G1) is related to tumorigenesis and tumor progression. However, its role and the underlying mechanisms in the regulation of tumor development in non-small cell lung cancers (NSCLC) remain largely unknown. Here we report that the levels of EIF4G1 expression are much higher in NSCLC cell lines and tumor tissues than those in the normal lung cells and adjacent normal tissues from the same patients. Using shRNA to knock down EIF4G1 expression stably, we found EIF4G1 required for NSCLC cell proliferation, anchorage-independent growth, migration and invasion. Furthermore, silencing of EIF4G1 induces NSCLC cell apoptosis and causes G0/G1 cell cycle arrest. To identify the partner protein network of EIF4G1 in NSCLC cells, we found that Ubiquitin-specific protease 10 (USP10) can directly interacts with EIF4G1, while acting as a negative regulator for EIF4G1-mediated functions. Together, our results indicate that EIF4G1 functions as an oncoprotein during NSCLC development, which may represent a novel and promising therapeutic target in lung cancer.

  19. Identification, expression and phylogenetic analysis of EgG1Y162 from Echinococcus granulosus

    PubMed Central

    Zhang, Fengbo; Ma, Xiumin; Zhu, Yuejie; Wang, Hongying; Liu, Xianfei; Zhu, Min; Ma, Haimei; Wen, Hao; Fan, Haining; Ding, Jianbing

    2014-01-01

    Objective: This study was to clone, identify and analyze the characteristics of egG1Y162 gene from Echinococcus granulosus. Methods: Genomic DNA and total RNAs were extracted from four different developmental stages of protoscolex, germinal layer, adult and egg of Echinococcus granulosus, respectively. Fluorescent quantitative PCR was used for analyzing the expression of egG1Y162 gene. Prokaryotic expression plasmid of pET41a-EgG1Y162 was constructed to express recombinant His-EgG1Y162 antigen. Western blot analysis was performed to detect antigenicity of EgG1Y162 antigen. Gene sequence, amino acid alignment and phylogenetic tree of EgG1Y162 were analyzed by BLAST, online Spidey and MEGA4 software, respectively. Results: EgG1Y162 gene was expressed in four developmental stages of Echinococcus granulosus. And, egG1Y162 gene expression was the highest in the adult stage, with the relative value of 19.526, significantly higher than other three stages. Additionally, Western blot analysis revealed that EgG1Y162 recombinant protein had good reaction with serum samples from Echinococcus granulosus infected human and dog. Moreover, EgG1Y162 antigen was phylogenetically closest to EmY162 antigen, with the similarity over 90%. Conclusion: Our study identified EgG1Y162 antigen in Echinococcus granulosus for the first time. EgG1Y162 antigen had a high similarity with EmY162 antigen, with the genetic differences mainly existing in the intron region. And, EgG1Y162 recombinant protein showed good antigenicity. PMID:25337206

  20. A reliable liquid chromatography-tandem mass spectrometry method for simultaneous determination of multiple mycotoxins in fresh fish and dried seafoods.

    PubMed

    Sun, Wenshuo; Han, Zheng; Aerts, Johan; Nie, Dongxia; Jin, Mengtong; Shi, Wen; Zhao, Zhiyong; De Saeger, Sarah; Zhao, Yong; Wu, Aibo

    2015-03-27

    A reliable liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed for simultaneous determination of nine mycotoxins, i.e., aflatoxin B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1), aflatoxin G2 (AFG2), ochratoxin A (OTA), zearalenone (ZEN), T-2 toxin, HT2 toxin and deoxynivalenol (DON), in fresh fish (muscle and entrails) as well as dried seafoods. Special focus was given to sample pretreatment which is crucial for an accurate and reliable analytical method. With regards to the high complexity of the matrices, extraction solvent, time, and temperature as well as clean-up cartridges were optimized to improve extraction efficiency and reduce matrix effects. The optimum procedure included ultrasound-assisted extraction with acetonitrile/water/acetic acid (79/20/1, v/v/v) at 40 °C for 30 min, defatting with n-hexane and purification by Oasis HLB cartridges. The method was further validated by determining the linearity (R(2) ≥ 0.9989), sensitivity (limit of detection ≤ 2 μg/kg, limit of quantitation ≤ 3 μg/kg), recovery (72.2-119.9%) and precision (≤ 18.3%) in muscle and entrails of fresh crucian carp (Carassius carassius) as well as dried fish products. The method was proven to be suitable for its intended purposes. Mycotoxins of OTA, ZEN and AFB2 have been found in fresh fish and dried seafoods for the first time.

  1. Diversity of VP7 genes of G1 rotaviruses isolated in Iran, 2009-2013.

    PubMed

    Jalilvand, Somayeh; Afchangi, Atefeh; Mohajel, Nasir; Roohvand, Farzin; Shoja, Zabihollah

    2016-01-01

    Genotype G1 of rotaviruses (RVs) is the most prevalent strain in human RV infections around the world. The present study evaluated genetic variations in the VP7 gene of RV G1 genotype isolates from Iran. Genetic and phylogenetic analyses indicated that RV strains from Iran clustered with G1 lineages IA, IC, and IIC, showing highest average of similarity versus reference sequences of the G1 lineages I and II. This study highlights the genetic pattern of G1 RV on the basis of distinct lineages and sublineages and indicates the importance of continuous monitoring on genetic variation and evolution pattern of G1 RV strains across the Iranian population for the final aim of RV vaccine introduction.

  2. Evidence of bovine immunoglobulin G1 (IgG1) protease activity in partially purified culture supernate of Pasteurella haemolytica A1.

    PubMed Central

    Lee, C W; Shewen, P E

    1996-01-01

    In the bovine respiratory tract, IgG1 is a major secretory immunoglobulin (Ig), and both IgG1 and IgG2 are believed to be important in defense against pneumonic pasteurellosis (shipping fever) in calves. Here we provide evidence for hydrolysis of IgG1 in the presence of partially purified culture supernate (ppCS) from the respiratory pathogen Pasteurella haemolytica A1. Bovine IgG1 was hydrolysed sequentially into three distinct bands (approximately 39, 12, and 7 kDa respectively). Furthermore, partial hydrolysis of bovine IgG2 was observed, but neither bovine IgA nor IgM were affected by incubation with ppCS. These findings suggest that the production of an IgG1-specific protease by P. haemolytica A1 may be a virulence mechanism contributing to the pathogenesis of bovine pneumonic pasteurellosis. Images Figure 1. Figure 2. Figure 3. Figure 4. Figure 5. PMID:8785718

  3. Endogenous production of immunoglobulin IgG1 in newborn calves.

    PubMed

    Devery, J E; Davis, C L; Larson, B L

    1979-11-01

    There is a decrease in the specific activity of labeled IgG1 of serum over 3 wk following the feeding of iodine-125 labeled immunoglobulin IgG1 in colostrum to calves at birth. This decrease indicated the appearance of new IgG1 from some source. To determine if this new IgG1 came from endogenous production in the calf or from continued small amount of intestinal absorption from milk, labeled IgG1 was added to normal milk and fed to calves of various ages up to 3 wk after an initial feeding of colostrum at birth. Labeled IgG1 was also added to colostrum fed to calves at birth, and the calves were maintained on a normal milk diet or fed a synthetic milk diet. Determination of iodine-125 in the serum protein fractions of these calves indicated that there was no apparent intestinal absorption of labeled IgG1 from the milk in the period from 2 days to 3 wk. Furthermore, comparable decreases occurred in the specific activity of labeled IgG1 in serum in the calves fed the labeled IgG1 in colostrum at birth and subsequently maintained either on a diet including milk or on the synthetic milk diet devoid of IgG1. The results support the conclusion that the origin of new IgG1 in the calf after about 36 h and up to about 3 wk of age arises from endogenous production at a rate of about 1 g of IgG1 per day.

  4. Most of the G1 period in hamster cells is eliminated by lengthening the S period.

    PubMed Central

    Stancel, G M; Prescott, D M; Liskay, R M

    1981-01-01

    Two Chinese hamster cell lines, G1+-1 and CHO, have been grown in the presence of low concentrations of hydroxyurea to determine how a slowing DNA synthesis (i.e., a lengthening of the S period) affects the length of the G1 period. Hydroxyurea concentrations of approximately 10 microM do not alter the generation times of these cell lines but do cause increases in S with corresponding decreases in G1. In both cell lines, 10 microM hydroxyurea reduces G1 to an absolute value of 1 hr, which represents decreases of 70% (G1+-1) and 60% (CHO) from control values. Higher concentrations of hydroxyurea increase the generation times and lengths of S for both cell lines but do not reduce G1 below the minimum value of 1 hr. These observations indicate that the majority of G1 is expendable and most of G1 therefore cannot contain specific events required for the initiation of DNA synthesis. This result supports the hypothesis that G1 is a portion of the cell growth cycle but not of the chromosome cycle. PMID:6947230

  5. FoxG1 and TLE2 act cooperatively to regulate ventral telencephalon formation

    PubMed Central

    Roth, Martin; Bonev, Boyan; Lindsay, Jennefer; Lea, Robert; Panagiotaki, Niki; Houart, Corinne; Papalopulu, Nancy

    2010-01-01

    FoxG1 is a conserved transcriptional repressor that plays a key role in the specification, proliferation and differentiation of the telencephalon, and is expressed from the earliest stages of telencephalic development through to the adult. How the interaction with co-factors might influence the multiplicity and diversity of FoxG1 function is not known. Here, we show that interaction of FoxG1 with TLE2, a Xenopus tropicalis co-repressor of the Groucho/TLE family, is crucial for regulating the early activity of FoxG1. We show that TLE2 is co-expressed with FoxG1 in the ventral telencephalon from the early neural plate stage and functionally cooperates with FoxG1 in an ectopic neurogenesis assay. FoxG1 has two potential TLE binding sites: an N-terminal eh1 motif and a C-terminal YWPMSPF motif. Although direct binding seems to be mediated by the N-terminal motif, both motifs appear important for functional synergism. In the neurogenesis assay, mutation of either motif abolishes functional cooperation of TLE2 with FoxG1, whereas in the forebrain deletion of both motifs renders FoxG1 unable to induce the ventral telencephalic marker Nkx2.1. Knocking down either FoxG1 or TLE2 disrupts the development of the ventral telencephalon, supporting the idea that endogenous TLE2 and FoxG1 work together to specify the ventral telencephalon. PMID:20356955

  6. Acrocentric Chromosomes in Cultured Leukocytes from Mothers of Children Affected With the G1- Trisomy Syndrome

    ERIC Educational Resources Information Center

    And Others; Cotton, James E.

    1973-01-01

    Analysis of venous blood samples from 24 mothers of G1-trisomy-affected (Down's Syndrome) children and 23 mothers of chromosomally normal children indicated that mothers of G1-trisomy-affected children had a greater than expected involvement of the G-chromosomes in associations of acrocentric satellited (chromosome configuration) chromosomes.…

  7. Enhanced HIV-1 neutralization by a CD4-VH3-IgG1 fusion protein

    SciTech Connect

    Meyuhas, Ronit; Noy, Hava; Fishman, Sigal; Margalit, Alon; Montefiori, David C.; Gross, Gideon

    2009-08-21

    HIV-1 gp120 is an alleged B cell superantigen, binding certain VH3+ human antibodies. We reasoned that a CD4-VH3 fusion protein could possess higher affinity for gp120 and improved HIV-1 inhibitory capacity. To test this we produced several human IgG1 immunoligands harboring VH3. Unlike VH3-IgG1 or VH3-CD4-IgG1, CD4-VH3-IgG1 bound gp120 considerably stronger than CD4-IgG1. CD4-VH3-IgG1 exhibited {approx}1.5-2.5-fold increase in neutralization of two T-cell laboratory-adapted strains when compared to CD4-IgG1. CD4-VH3-IgG1 improved neutralization of 7/10 clade B primary isolates or pseudoviruses, exceeding 20-fold for JR-FL and 13-fold for Ba-L. It enhanced neutralization of 4/8 clade C viruses, and had negligible effect on 1/4 clade A pseudoviruses. We attribute this improvement to possible pairing of VH3 with CD4 D1 and stabilization of an Ig Fv-like structure, rather than to superantigen interactions. These novel findings support the current notion that CD4 fusion proteins can act as better HIV-1 entry inhibitors with potential clinical implications.

  8. 16 CFR Appendix G1 to Part 305 - Furnaces-Gas

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 16 Commercial Practices 1 2013-01-01 2013-01-01 false Furnaces-Gas G1 Appendix G1 to Part 305 Commercial Practices FEDERAL TRADE COMMISSION REGULATIONS UNDER SPECIFIC ACTS OF CONGRESS RULE CONCERNING... Part 305—Furnaces—Gas Manufacturer's rated heating capacities (Btu's/hr.) Range of annual...

  9. 16 CFR Appendix G1 to Part 305 - Furnaces-Gas

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 16 Commercial Practices 1 2012-01-01 2012-01-01 false Furnaces-Gas G1 Appendix G1 to Part 305 Commercial Practices FEDERAL TRADE COMMISSION REGULATIONS UNDER SPECIFIC ACTS OF CONGRESS RULE CONCERNING... Part 305—Furnaces—Gas Manufacturer's rated heating capacities (Btu's/hr.) Range of annual...

  10. 16 CFR Appendix G1 to Part 305 - Furnaces-Gas

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 16 Commercial Practices 1 2010-01-01 2010-01-01 false Furnaces-Gas G1 Appendix G1 to Part 305 Commercial Practices FEDERAL TRADE COMMISSION REGULATIONS UNDER SPECIFIC ACTS OF CONGRESS RULE CONCERNING... Part 305—Furnaces—Gas Manufacturer's rated heating capacities (Btu's/hr.) Range of annual...

  11. 16 CFR Appendix G1 to Part 305 - Furnaces-Gas

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 16 Commercial Practices 1 2011-01-01 2011-01-01 false Furnaces-Gas G1 Appendix G1 to Part 305 Commercial Practices FEDERAL TRADE COMMISSION REGULATIONS UNDER SPECIFIC ACTS OF CONGRESS RULE CONCERNING... Part 305—Furnaces—Gas Manufacturer's rated heating capacities (Btu's/hr.) Range of annual...

  12. Excretion of serotype G1 rotavirus strains by asymptomatic staff: a possible source of nosocomial infection.

    PubMed

    Barnes, Graeme L; Callaghan, Sarah L; Kirkwood, Carl D; Bogdanovic-Sakran, Nada; Johnston, Linda J; Bishop, Ruth F

    2003-06-01

    This study supports the hypothesis that feces from asymptomatic adults may provide a vehicle for the transmission of rotavirus, in addition to aerosols, hands, and fomites. The observed preferential carriage of serotype G1 strains in the adult gastrointestinal tract may explain G1 predominance and persistence in epidemiologic studies worldwide.

  13. 26 CFR 1.402(g)-1 - Limitation on exclusion for elective deferrals.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 1, 2006 edition of 26 CFR Part 1). (ii) Method of allocating income. A plan may use any reasonable.... 1.402(g)-1 Section 1.402(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Pension, Profit-Sharing, Stock Bonus Plans, Etc. §...

  14. 26 CFR 301.6323(g)-1 - Refiling of notice of tax lien.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 18 2010-04-01 2010-04-01 false Refiling of notice of tax lien. 301.6323(g)-1 Section 301.6323(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) PROCEDURE AND ADMINISTRATION PROCEDURE AND ADMINISTRATION Collection General Provisions §...

  15. 26 CFR 1.404(g)-1 - Deduction of employer liability payments.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... Termination of Sufficient Plans. See PBGC regulations, 29 CFR 2617.13(b) for rules concerning these...(g)-1 Section 1.404(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Pension, Profit-Sharing, Stock Bonus Plans, Etc. §...

  16. 26 CFR 1.414(g)-1 - Definition of plan administrator.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 5 2010-04-01 2010-04-01 false Definition of plan administrator. 1.414(g)-1 Section 1.414(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Pension, Profit-Sharing, Stock Bonus Plans, Etc. §...

  17. 26 CFR 1.415(g)-1 - Disqualification of plans and trusts.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 5 2010-04-01 2010-04-01 false Disqualification of plans and trusts. 1.415(g)-1 Section 1.415(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY (CONTINUED) INCOME TAX (CONTINUED) INCOME TAXES Pension, Profit-Sharing, Stock Bonus Plans, Etc. §...

  18. 26 CFR 25.2523(g)-1 - Special rule for charitable remainder trusts.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 26 Internal Revenue 14 2014-04-01 2013-04-01 true Special rule for charitable remainder trusts. 25.2523(g)-1 Section 25.2523(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY...(g). Accordingly, if the transfer to the trust is made prior to October 24, 1992, the spousal...

  19. A hyperactive transcriptional state marks genome reactivation at the mitosis-G1 transition.

    PubMed

    Hsiung, Chris C-S; Bartman, Caroline R; Huang, Peng; Ginart, Paul; Stonestrom, Aaron J; Keller, Cheryl A; Face, Carolyne; Jahn, Kristen S; Evans, Perry; Sankaranarayanan, Laavanya; Giardine, Belinda; Hardison, Ross C; Raj, Arjun; Blobel, Gerd A

    2016-06-15

    During mitosis, RNA polymerase II (Pol II) and many transcription factors dissociate from chromatin, and transcription ceases globally. Transcription is known to restart in bulk by telophase, but whether de novo transcription at the mitosis-G1 transition is in any way distinct from later in interphase remains unknown. We tracked Pol II occupancy genome-wide in mammalian cells progressing from mitosis through late G1. Unexpectedly, during the earliest rounds of transcription at the mitosis-G1 transition, ∼50% of active genes and distal enhancers exhibit a spike in transcription, exceeding levels observed later in G1 phase. Enhancer-promoter chromatin contacts are depleted during mitosis and restored rapidly upon G1 entry but do not spike. Of the chromatin-associated features examined, histone H3 Lys27 acetylation levels at individual loci in mitosis best predict the mitosis-G1 transcriptional spike. Single-molecule RNA imaging supports that the mitosis-G1 transcriptional spike can constitute the maximum transcriptional activity per DNA copy throughout the cell division cycle. The transcriptional spike occurs heterogeneously and propagates to cell-to-cell differences in mature mRNA expression. Our results raise the possibility that passage through the mitosis-G1 transition might predispose cells to diverge in gene expression states.

  20. 26 CFR 1.642(g)-1 - Disallowance of double deductions; in general.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 26 Internal Revenue 8 2013-04-01 2013-04-01 false Disallowance of double deductions; in general. 1.642(g)-1 Section 1.642(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY... Disallowance of double deductions; in general. Amounts allowable under section 2053(a)(2) (relating...

  1. 26 CFR 1.642(g)-1 - Disallowance of double deductions; in general.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 26 Internal Revenue 8 2012-04-01 2012-04-01 false Disallowance of double deductions; in general. 1.642(g)-1 Section 1.642(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY... Disallowance of double deductions; in general. Amounts allowable under section 2053(a)(2) (relating...

  2. 26 CFR 1.642(g)-1 - Disallowance of double deductions; in general.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 26 Internal Revenue 8 2014-04-01 2014-04-01 false Disallowance of double deductions; in general. 1.642(g)-1 Section 1.642(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY... Disallowance of double deductions; in general. Amounts allowable under section 2053(a)(2) (relating...

  3. 26 CFR 1.642(g)-1 - Disallowance of double deductions; in general.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 26 Internal Revenue 8 2011-04-01 2011-04-01 false Disallowance of double deductions; in general. 1.642(g)-1 Section 1.642(g)-1 Internal Revenue INTERNAL REVENUE SERVICE, DEPARTMENT OF THE TREASURY... Disallowance of double deductions; in general. Amounts allowable under section 2053(a)(2) (relating...

  4. Processing and membrane topology of the spike proteins G1 and G2 of Uukuniemi virus.

    PubMed Central

    Andersson, A M; Melin, L; Persson, R; Raschperger, E; Wikström, L; Pettersson, R F

    1997-01-01

    The membrane glycoproteins G1 and G2 of the members of the Bunyaviridae family are synthesized as a precursor from a single open reading frame. Here, we have analyzed the processing and membrane insertion of G1 and G2 of a member of the Phlebovirus genus, Uukuniemi virus. By expressing C-terminally truncated forms of the p10 precursor containing the whole of G1 and decreasing portions of G2, we found that processing in BHK21 cells occurred with an efficiency of about 50% if G1 was followed by 50 residues of G2, while complete processing occurred if 98, 150, or 200 residues of G2 were present. Surprisingly, processing of all truncated G2 forms was less efficient in HeLa cells. Proteinase K treatment of microsomes isolated from infected cells indicated that the C terminus of G1 is exposed on the cytoplasmic face. Using G1 tail peptide antisera, the tail was likewise found by immunofluorescence to be exposed on the cytoplasmic face in streptolysin O-permeabilized cells. By introducing stop codons at various positions of the G1 tail and at the natural cleavage site between G1 and G2 and expressing these mutants in BHK cells, we found that no further processing of the G1 C terminus occurred following cleavage of G2 by the signal peptidase. This was also supported by the finding that an antiserum raised against a peptide corresponding to the region immediately upstream from the G2 signal sequence reacted in immunoblotting with G1 from virions. Finally, we show that both G1 and G2 are palmitylated. Taken together, these results show that processing of p10 of Uukuniemi virus occurs cotranslationally at only one site, i.e., downstream of the internal G2 signal sequence. G1 and G2 are inserted as type I proteins into the lipid bilayer, leaving the G1 tail exposed on the cytoplasmic face of the membrane. Since the G2 tail is only 5 residues long, the G1 tail is likely to be responsible for the interaction with the nucleoproteins during the budding process, in addition to

  5. Role of HLA-G1 in trophoblast cell proliferation, adhesion and invasion

    SciTech Connect

    Jiang, Feng; Zhao, Hongxi; Wang, Li; Guo, Xinyu; Wang, Xiaohong; Yin, Guowu; Hu, Yunsheng; Li, Yi; Yao, Yuanqing

    2015-02-27

    Trophoblast cells are important in embryo implantation and fetomaternal tolerance. HLA-G is specifically expressed at the maternal–fetal interface and is a regulator in pregnancy. The aim of the present study was to detect the effect of HLA-G1 on trophoblast cell proliferation, adhesion, and invasion. Human trophoblast cell lines (JAR and HTR-8/SVneo cells) were infected with HLA-G1-expressing lentivirus. After infection, HLA-G1 expression of the cells was detected by western blotting. Cell proliferation was detected by the BrdU assay. The cell cycle and apoptosis of JAR and HTR-8/SVneo cells was measured by flow cytometry (FCM). The invasion of the cells under different conditions was detected by the transwell invasion chamber assay. HLA-G1 didn't show any significant influence on the proliferation, apoptosis, adhesion, and invasion of trophocytes in normal culture conditions. However, HLA-G1 inhibited JAR and HTR-8/SVneo cells invasion induced by hepatocyte growth factor (HGF) under normal oxygen conditions. In conditions of hypoxia, HLA-G1 couldn't inhibit the induction of cell invasion by HGF. HLA-G1 is not an independent factor for regulating the trophocytes. It may play an indirect role in embryo implantation and formation of the placenta. - Highlights: • HLA-G1 could not influence trophocytes under normal conditions. • HLA-G1 inhibited cell invasion induced by HGF under normal oxygen condition. • HLA-G1 could not influence cell invasion under hypoxia conditions.

  6. Msa1 and Msa2 Modulate G1-Specific Transcription to Promote G1 Arrest and the Transition to Quiescence in Budding Yeast

    PubMed Central

    Miles, Shawna; Croxford, Matthew W.; Abeysinghe, Amali P.; Breeden, Linda L.

    2016-01-01

    Yeast that naturally exhaust their glucose source can enter a quiescent state that is characterized by reduced cell size, and high cell density, stress tolerance and longevity. The transition to quiescence involves highly asymmetric cell divisions, dramatic reprogramming of transcription and global changes in chromatin structure and chromosome topology. Cells enter quiescence from G1 and we find that there is a positive correlation between the length of G1 and the yield of quiescent cells. The Swi4 and Swi6 transcription factors, which form the SBF transcription complex and promote the G1 to S transition in cycling cells, are also critical for the transition to quiescence. Swi6 forms a second complex with Mbp1 (MBF), which is not required for quiescence. These are the functional analogues of the E2F complexes of higher eukaryotes. Loss of the RB analogue, Whi5, and the related protein Srl3/Whi7, delays G1 arrest, but it also delays recovery from quiescence. Two MBF- and SBF-Associated proteins have been identified that have little effect on SBF or MBF activity in cycling cells. We show that these two related proteins, Msa1 and Msa2, are specifically required for the transition to quiescence. Like the E2F complexes that are quiescence-specific, Msa1 and Msa2 are required to repress the transcription of many SBF target genes, including SWI4, the CLN2 cyclin and histones, specifically after glucose is exhausted from the media. They also activate transcription of many MBF target genes. msa1msa2 cells fail to G1 arrest and rapidly lose viability upon glucose exhaustion. msa1msa2 mutants that survive this transition are very large, but they attain the same thermo-tolerance and longevity of wild type quiescent cells. This indicates that Msa1 and Msa2 are required for successful transition to quiescence, but not for the maintenance of that state. PMID:27272642

  7. Msa1 and Msa2 Modulate G1-Specific Transcription to Promote G1 Arrest and the Transition to Quiescence in Budding Yeast.

    PubMed

    Miles, Shawna; Croxford, Matthew W; Abeysinghe, Amali P; Breeden, Linda L

    2016-06-01

    Yeast that naturally exhaust their glucose source can enter a quiescent state that is characterized by reduced cell size, and high cell density, stress tolerance and longevity. The transition to quiescence involves highly asymmetric cell divisions, dramatic reprogramming of transcription and global changes in chromatin structure and chromosome topology. Cells enter quiescence from G1 and we find that there is a positive correlation between the length of G1 and the yield of quiescent cells. The Swi4 and Swi6 transcription factors, which form the SBF transcription complex and promote the G1 to S transition in cycling cells, are also critical for the transition to quiescence. Swi6 forms a second complex with Mbp1 (MBF), which is not required for quiescence. These are the functional analogues of the E2F complexes of higher eukaryotes. Loss of the RB analogue, Whi5, and the related protein Srl3/Whi7, delays G1 arrest, but it also delays recovery from quiescence. Two MBF- and SBF-Associated proteins have been identified that have little effect on SBF or MBF activity in cycling cells. We show that these two related proteins, Msa1 and Msa2, are specifically required for the transition to quiescence. Like the E2F complexes that are quiescence-specific, Msa1 and Msa2 are required to repress the transcription of many SBF target genes, including SWI4, the CLN2 cyclin and histones, specifically after glucose is exhausted from the media. They also activate transcription of many MBF target genes. msa1msa2 cells fail to G1 arrest and rapidly lose viability upon glucose exhaustion. msa1msa2 mutants that survive this transition are very large, but they attain the same thermo-tolerance and longevity of wild type quiescent cells. This indicates that Msa1 and Msa2 are required for successful transition to quiescence, but not for the maintenance of that state. PMID:27272642

  8. Phylogenetic inference of the porcine Rotavirus A origin of the human G1 VP7 gene.

    PubMed

    Do, Loan Phuong; Nakagomi, Toyoko; Otaki, Hiroki; Agbemabiese, Chantal Ama; Nakagomi, Osamu; Tsunemitsu, Hiroshi

    2016-06-01

    Rotavirus A (RVA) is an important cause of acute gastroenteritis in children worldwide. The most common VP7 genotype of human RVA is G1, but G1 is rarely detected in porcine strains. To understand the evolutionary relationships between human and porcine G1 VP7 genes, we sequenced the VP7 genes of three Japanese G1 porcine strains; the first two (PRV2, S80B) were isolated in 1980 and the third (Kyusyu-14) was isolated in 2001. Then, we performed phylogenetic and in-silico structural analyses. All three VP7 sequences clustered into lineage VI, and the mean nucleotide sequence identity between any pair of porcine G1 VP7 sequences belonging to lineage VI was 91.9%. In contrast, the mean nucleotide sequence identity between any pair of human G1 VP7 sequences belonging to lineages I-V was 95.5%. While the mean nucleotide sequence identity between any pair of porcine lineage VI strain and human lineage I-V strain was 85.4%, the VP7 genes of PRV2 and a rare porcine-like human G1P[6] strain (AU19) were 98% identical, strengthening the porcine RVA origin of AU19. The phylogenetic tree suggests that human G1 VP7 genes originated from porcine G1 VP7 genes. The time of their most recent common ancestor was estimated to be 1948, and human and porcine RVA strains evolved along independent pathways. In-silico structural analyses identified 7 amino acid residues within the known neutralisation epitopes that show differences in electric charges and shape between different porcine and human G1 strains. When compared with much divergent porcine G1 VP7 lineages, monophyletic, less divergent human G1 VP7 lineages support the hypothesis that all human G1 VP7 genes included in this study originated from a rare event of a porcine RVA transmitting to humans that was followed by successful adaptation to the human host. By contrast, AU19 represents interspecies transmission that terminated in dead-end infection. PMID:26961591

  9. Mitotic UV irradiation induces a DNA replication-licensing defect that potentiates G1 arrest response.

    PubMed

    Morino, Masayuki; Nukina, Kohei; Sakaguchi, Hiroki; Maeda, Takeshi; Takahara, Michiyo; Shiomi, Yasushi; Nishitani, Hideo

    2015-01-01

    Cdt1 begins to accumulate in M phase and has a key role in establishing replication licensing at the end of mitosis or in early G1 phase. Treatments that damage the DNA of cells, such as UV irradiation, induce Cdt1 degradation through PCNA-dependent CRL4-Cdt2 ubiquitin ligase. How Cdt1 degradation is linked to cell cycle progression, however, remains unclear. In G1 phase, when licensing is established, UV irradiation leads to Cdt1 degradation, but has little effect on the licensing state. In M phase, however, UV irradiation does not induce Cdt1 degradation. When mitotic UV-irradiated cells were released into G1 phase, Cdt1 was degraded before licensing was established. Thus, these cells exhibited both defective licensing and G1 cell cycle arrest. The frequency of G1 arrest increased in cells expressing extra copies of Cdt2, and thus in cells in which Cdt1 degradation was enhanced, whereas the frequency of G1 arrest was reduced in cell expressing an extra copy of Cdt1. The G1 arrest response of cells irradiated in mitosis was important for cell survival by preventing the induction of apoptosis. Based on these observations, we propose that mammalian cells have a DNA replication-licensing checkpoint response to DNA damage induced during mitosis.

  10. Fluorimetric measurements and chromatin condensation patterns of nuclei from 3T3 cells throughout G1.

    PubMed

    Moser, G C; Fallon, R J; Meiss, H K

    1981-02-01

    Using two cytological methods based on nuclear morphology, quinacrine dihydrochloride (QDH) staining and premature chromosome condensation (PCC), it has been possible to identify cell cycle positions within G1 of growing and arrested 3T3 cells. The fluorescent intensity of QDH-stained interphase cells appears to decrease as the cells pass from mitosis to S phase. Likewise, the length and thickness of prematurely condensed chromatids can be related to the cells; position within the G1 period. Data are presented that deal with three interrelated topics: 1) We determined by fluorometric measurements of nuclei from 3T3 cells that the visual observation of the decrease in QDH fluorescence during G1 reflects an actual decrease in total fluorescence and not a dispersion of the fluorescent chromatin in a larger nuclear area. 2) We correlated the results obtained by QDH staining with those of PCC on the same cell samples blocked in G1 by different conditions. Serum-starved and contact-inhibited cell nuclei had the highest intensity, hydroxyurea-treated ones had the lowest intensity, while that of isoleucine-deprived cells was in between. The same relative order of G1 positions was obtained based on PCC morphology. Thus, both methods monitor the state of chromatin condensation and can be used to identify cell cycle position within G1. 3) We showed with both methods that the states of chromatin resulting from the various G1 blocking conditions differ from each other.

  11. Quarter variation and correlations of colostrum albumin, immunoglobulin G1 and G2 in dairy cows.

    PubMed

    Samarütel, Jaak; Baumrucker, Craig R; Gross, Josef J; Dechow, Chad D; Bruckmaier, Rupert M

    2016-05-01

    A high variation in immunoglobulin G1 (IgG1) concentration in first milked quarter colostrum has been reported, but BSA quarter colostrum variation is not known. The occurrence of serum albumin in milk has been attributed to increased blood-milk barrier penetration. Reports of serum albumin binding to the Fc Receptor of the neonate, the receptor thought to be responsible for IgG1 transcytosis, suggested that a correlation with the appearance of IgG1 in colostrum of dairy cows was likely. The objective of the study was to establish the quarter colostrum concentration and mass of immunoglobulins and serum albumin. First colostrum was quarter collected within 4 h of parturition from healthy udders of 31 multiparous dairy cows. Individual quarter colostrum weight was determined and a sample of each was frozen for subsequent analysis. Concentrations of immunoglobulin G1, G2, and BSA were measured by ELISA and total mass of components was calculated. In addition, colostrum was also analysed for L-lactate dehydrogenase activity. Analysis of concentration and mass of BSA, immunoglobulin G1, G2 established that the quarter variations were different by cow, quarter and quarter within cow. Partial correlations corrected for colostrum weight indicated that BSA and IgG2 concentration and mass are closely correlated while that of BSA and IgG1 concentration and mass exhibited no correlation suggesting that BSA and IgG1 may have different transport mechanisms. Interestingly, immunoglobulin G1 and G2 concentration and mass exhibited strong correlations suggesting that also some unknown mechanism of immunoglobulin G2 appearance in colostrum is occurring. Finally, no measured protein exhibited any correlation with the activity of lactate dehydrogenase in colostrum.

  12. Transfection-mediated cell synchronization: acceleration of G1-S phase transition by gamma irradiation.

    PubMed

    Jung, E J; Flemington, E K

    2001-11-01

    We have previously provided evidence that the uptake of DNA into cells is cell cycle specific following transfection. We show here that, immediately after transfection, successfully transfected cells are greatly enriched for cells in early G1 or G0 phase and that, upon removal of the DNA precipitates, cells progress through G1 and enter S phase in a synchronous fashion. We also demonstrate that this approach can be utilized in meaningful cell-cycle experiments, and we show that gamma irradiation accelerates the G1-S phase transition in a cell line with a functionally inactive p53 protein. PMID:11730009

  13. Moments of the Spin Structure Functions g1p and g1d for 0.05 < Q2 < 3.0 GeV2

    SciTech Connect

    Prok, Yelena; Bosted, Peter; Burkert, Volker; Deur, Alexandre; Dharmawardane, Kahanawita; Dodge, Gail; Griffioen, Keith; Kuhn, Sebastian; Minehart, Ralph; Adams, Gary; Amaryan, Moscov; Amaryan, Moskov; Anghinolfi, Marco; Asryan, G.; Audit, Gerard; Avagyan, Harutyun; Baghdasaryan, Hovhannes; Baillie, Nathan; Ball, J.P.; Ball, Jacques; Baltzell, Nathan; Barrow, Steve; Battaglieri, Marco; Beard, Kevin; Bedlinskiy, Ivan; Bektasoglu, Mehmet; Bellis, Matthew; Benmouna, Nawal; Berman, Barry; Biselli, Angela; Blaszczyk, Lukasz; Boyarinov, Sergey; Bonner, Billy; Bouchigny, Sylvain; Bradford, Robert; Branford, Derek; Briscoe, William; Brooks, William; Bultmann, S.; Bueltmann, Stephen; Butuceanu, Cornel; Calarco, John; Careccia, Sharon; Carman, Daniel; Casey, Liam; Cazes, Antoine; Chen, Shifeng; Cheng, Lu; Cole, Philip; Collins, Patrick; Coltharp, Philip; Cords, Dieter; Corvisiero, Pietro; Crabb, Donald; Crede, Volker; Cummings, John; Dale, Daniel; Dashyan, Natalya; De Masi, Rita; De Vita, Raffaella; De Sanctis, Enzo; Degtiarenko, Pavel; Denizli, Haluk; Dennis, Lawrence; Dhuga, Kalvir; Dickson, Richard; Djalali, Chaden; Doughty, David; Dugger, Michael; Dytman, Steven; Dzyubak, Oleksandr; Egiyan, Hovanes; Egiyan, Kim; Elfassi, Lamiaa; Elouadrhiri, Latifa; Eugenio, Paul; Fatemi, Renee; Fedotov, Gleb; Feldman, Gerald; Fersch, Robert; Feuerbach, Robert; Forest, Tony; Fradi, Ahmed; Funsten, Herbert; Garcon, Michel; Gavalian, Gagik; Gevorgyan, Nerses; Gilfoyle, Gerard; Giovanetti, Kevin; Girod, Francois-Xavier; Goetz, John; Golovach, Evgeny; Gothe, Ralf; Guidal, Michel; Guillo, Matthieu; Guler, Nevzat; Guo, Lei; Gyurjyan, Vardan; Hadjidakis, Cynthia; Hafidi, Kawtar; Hakobyan, Hayk; Hanretty, Charles; Hardie, John; Hassall, Neil; Heddle, David; Hersman, F.; Hicks, Kenneth; Hleiqawi, Ishaq; Holtrop, Maurik; Huertas, Marco; Hyde, Charles; Ilieva, Yordanka; Ireland, David; Ishkhanov, Boris; Isupov, Evgeny; Ito, Mark; Jenkins, David; Jo, Hyon-Suk; Johnstone, John; Joo, Kyungseon; Juengst, Henry; Kalantarians, Narbe; Keith, Christopher; Kellie, James; Khandaker, Mahbubul; Kim, Kui; Kim, Kyungmo; Kim, Wooyoung; Klein, Andreas; Klein, Franz; Klusman, Mike; Kossov, Mikhail; Krahn, Zebulun; Kramer, Laird; Kubarovsky, Valery; Kuhn, Joachim; Kuleshov, Sergey; Kuznetsov, Viacheslav; Lachniet, Jeff; Laget, Jean; Langheinrich, Jorn; Lawrence, Dave; Lima, Ana; Livingston, Kenneth; Lu, Haiyun; Lukashin, K.; MacCormick, Marion; Marchand, Claude; Markov, Nikolai; Mattione, Paul; McAleer, Simeon; McKinnon, Bryan; McNabb, John; Mecking, Bernhard; Mestayer, Mac; Meyer, Curtis; Mibe, Tsutomu; Mikhaylov, Konstantin; Mirazita, Marco; Miskimen, Rory; Mokeev, Viktor; Morand, Ludyvine; Moreno, Brahim; Moriya, Kei; Morrow, Steven; Moteabbed, Maryam; Mueller, James; Munevar Espitia, Edwin; Mutchler, Gordon; Nadel-Turonski, Pawel; Nasseripour, Rakhsha; Niccolai, Silvia; Niculescu, Gabriel; Niculescu, Maria-Ioana; Niczyporuk, Bogdan; Niroula, Megh; Niyazov, Rustam; Nozar, Mina; O'Rielly, Grant; Osipenko, Mikhail; Ostrovidov, Alexander; Park, Kijun; Pasyuk, Evgueni; Paterson, Craig; Anefalos Pereira, S.; Philips, Sasha; Pierce, J.; Pivnyuk, Nikolay; Pocanic, Dinko; Pogorelko, Oleg; Popa, Iulian; Pozdnyakov, Sergey; Preedom, Barry; Price, John; Procureur, Sebastien; Protopopescu, Dan; Qin, Liming; Raue, Brian; Riccardi, Gregory; Ricco, Giovanni; Ripani, Marco; Ritchie, Barry; Rosner, Guenther; Rossi, Patrizia; Rowntree, David; Rubin, Philip; Sabatie, Franck; Salamanca, Julian; Salgado, Carlos; Santoro, Joseph; Sapunenko, Vladimir; Schumacher, Reinhard; Seely, Mikell; Serov, Vladimir; Sharabian, Youri; Sharov, Dmitri; Shaw, Jeffrey; Shvedunov, Nikolay; Skabelin, Alexander; Smith, Elton; Smith, Lee; Sober, Daniel; Sokhan, Daria; Stavinskiy, Aleksey; Stepanyan, Samuel; Stepanyan, Stepan; Stokes, Burnham; Stoler, Paul; Strakovski, Igor; Strauch, Steffen; Suleiman, Riad; Taiuti, Mauro; Tedeschi, David; Tkabladze, Avtandil; Tkachenko, Svyatoslav; Todor, Luminita; Ungaro, Maurizio; V

    2009-02-01

    The spin structure functions $g_1$ for the proton and the deuteron have been measured over a wide kinematic range in $x$ and \\Q2 using 1.6 and 5.7 GeV longitudinally polarized electrons incident upon polarized NH$_3$ and ND$_3$ targets at Jefferson Lab. Scattered electrons were detected in the CEBAF Large Acceptance Spectrometer, for $0.05 < Q^2 < 5 $\\ GeV$^2$ and $W < 3$ GeV. The first moments of $g_1$ for the proton and deuteron are presented -- both have a negative slope at low \\Q2, as predicted by the extended Gerasimov-Drell-Hearn sum rule. The first result for the generalized forward spin polarizability of the proton $\\gamma_0^p$ is also reported, and shows evidence of scaling above $Q^2$ = 1.5 GeV$^2$. Although the first moments of $g_1$ are consistent with Chiral Perturbation Theory (\\ChPT) calculations up to approximately $Q^2 = 0.06$ GeV$^2$, a significant discrepancy is observed between the $\\gamma_0^p$ data and \\ChPT\\ for $\\gamma_0^p$,even at the lowest \\Q2.

  14. IgG1 variations in the colostrum of Holstein dairy cows.

    PubMed

    Le Cozler, Y; Guatteo, R; Le Dréan, E; Turban, H; Leboeuf, F; Pecceu, K; Guinard-Flament, J

    2016-02-01

    High-immune quality colostrum (IgG1 concentration ⩾50 g/l) is crucial for the health and development of the young calf. Studies on colostrum quality tend to focus on external factors such as breed, parity or dry period length, but few have focused on within-cow variations. Here we ran experiments to gain a deeper insight into within-cow variation in IgG1 concentrations in dairy cow colostrum. Trials were performed in an experimental farm, located in the Western part of France. Colostrum from each quarter and a composite sample (mix of four quarters) were concomitantly collected on 77 Holstein dairy cows just after calving to assess the influence of sample type on IgG1 concentrations. Variation in IgG1 concentrations during the first milking was studied on samples from nine cows collected every minute from the start of milking. Repeatability of colostral IgG1 concentration was estimated from 2009 and 2010 data on 16 healthy cows. IgG1 concentrations were tested using a radial immunodiffusion method. Sensitivity and specificity were similar regardless of sample type tested (individual quarter or composite milk). Mean average IgG1 concentration was 54.1 g/l in composite colostrum, and was significantly higher in hind quarter teats (56.2 g/l) than front quarter teats (53.1 g/l). Average IgG1 concentration did not change significantly during colostrum milking, and the variations observed (15% or less) were likely due to the laboratory method (CV 15%). IgG1 concentrations in dam colostrum increased slightly from 2009 to 2010 due to BW and parity effects. In 56% of cases, colostrum quality could have been assessed on either individual or composite colostrum samples collected at any time during the first milking without affecting the reliability of the measurement. However, in other cases, differences were significant enough to mean that estimates of average IgG1 concentration in colostrum from any one quarter would not be reliable. It is concluded that colostrum quality

  15. Commercial geophysical well logs from the USW G-1 drill hole, Nevada Test Site, Nevada

    USGS Publications Warehouse

    Muller, D.C.; Kibler, J.E.

    1983-01-01

    Drill hole USW G-1 was drilled at Yucca Mountain, Nevada Test Site, Nevada, as part of the ongoing exploration program for the Nevada Nuclear Waste Storage Investigations. Contract geophysical well logs run at USW G-1 show only limited stratigraphic correlations, but correlate reasonably well with the welding of the ash-flow and ash-fall tuffs. Rocks in the upper part of the section have highly variable physical properties, but are more uniform and predictably lower in the section.

  16. Effects of Bordetella pertussis components on IgE and IgG1 responses.

    PubMed

    Sekiya, K

    1983-01-01

    The effect of dermonecrotic toxin (DNT), fimbrial hemagglutinin (FHA), K-agglutinogen, lipopolysaccharide (LPS), and pertussigen from Bordetella pertussis on the production of IgE and IgG1 antibodies to hen egg albumin (Ea) was investigated in C57BL/6 mice. The IgE antibody contents were determined by passive cutaneous anaphylaxis (PCA) in the skin of Lewis rats, while the IgG1 antibody contents were determined by PCA reactions on the skin of mice using sera that had been heated for 3 hr at 56 C to destroy the IgE antibodies. Among the B. pertussis components tested, pertussigen was the most effective adjuvant for increasing the IgE and IgG1 antibodies to Ea. LPS also moderately increased both types of antibodies, and FHA slightly increased the IgG1 titers. When LPS was given 5 days before Ea, it suppressed both IgE and IgG1 titers while FHA had only slight adjuvant action on both type of antibodies. When each of the components was tested for its ability to modify the adjuvant action of pertussigen, it was found that only DNT interfered significantly with the adjuvanticity of pertussigen when given on the day of immunization with Ea. When the components were given 5 days before Ea, DNT produced significant suppression of only the IgG1 response. LPS, FHA, and K-agglutinogen did not significantly affect the adjuvant action of pertussigen. PMID:6321910

  17. GPER agonist G-1 decreases adrenocortical carcinoma (ACC) cell growth in vitro and in vivo.

    PubMed

    Chimento, Adele; Sirianni, Rosa; Casaburi, Ivan; Zolea, Fabiana; Rizza, Pietro; Avena, Paola; Malivindi, Rocco; De Luca, Arianna; Campana, Carmela; Martire, Emilia; Domanico, Francesco; Fallo, Francesco; Carpinelli, Giulia; Cerquetti, Lidia; Amendola, Donatella; Stigliano, Antonio; Pezzi, Vincenzo

    2015-08-01

    We have previously demonstrated that estrogen receptor (ER) alpha (ESR1) increases proliferation of adrenocortical carcinoma (ACC) through both an estrogen-dependent and -independent (induced by IGF-II/IGF1R pathways) manner. Then, the use of tamoxifen, a selective estrogen receptor modulator (SERM), appears effective in reducing ACC growth in vitro and in vivo. However, tamoxifen not only exerts antiestrogenic activity, but also acts as full agonist on the G protein-coupled estrogen receptor (GPER). Aim of this study was to investigate the effect of a non-steroidal GPER agonist G-1 in modulating ACC cell growth. We found that G-1 is able to exert a growth inhibitory effect on H295R cells both in vitro and, as xenograft model, in vivo. Treatment of H295R cells with G-1 induced cell cycle arrest, DNA damage and cell death by the activation of the intrinsic apoptotic mechanism. These events required sustained extracellular regulated kinase (ERK) 1/2 activation. Silencing of GPER by a specific shRNA partially reversed G-1-mediated cell growth inhibition without affecting ERK activation. These data suggest the existence of G-1 activated but GPER-independent effects that remain to be clarified. In conclusion, this study provides a rational to further study G-1 mechanism of action in order to include this drug as a treatment option to the limited therapy of ACC.

  18. GPER agonist G-1 decreases adrenocortical carcinoma (ACC) cell growth in vitro and in vivo

    PubMed Central

    Zolea, Fabiana; Rizza, Pietro; Avena, Paola; Malivindi, Rocco; De Luca, Arianna; Campana, Carmela; Martire, Emilia; Domanico, Francesco; Fallo, Francesco; Carpinelli, Giulia; Cerquetti, Lidia; Amendola, Donatella; Stigliano, Antonio; Pezzi, Vincenzo

    2015-01-01

    We have previously demonstrated that estrogen receptor (ER) alpha (ESR1) increases proliferation of adrenocortical carcinoma (ACC) through both an estrogen-dependent and -independent (induced by IGF-II/IGF1R pathways) manner. Then, the use of tamoxifen, a selective estrogen receptor modulator (SERM), appears effective in reducing ACC growth in vitro and in vivo. However, tamoxifen not only exerts antiestrogenic activity, but also acts as full agonist on the G protein-coupled estrogen receptor (GPER). Aim of this study was to investigate the effect of a non-steroidal GPER agonist G-1 in modulating ACC cell growth. We found that G-1 is able to exert a growth inhibitory effect on H295R cells both in vitro and, as xenograft model, in vivo. Treatment of H295R cells with G-1 induced cell cycle arrest, DNA damage and cell death by the activation of the intrinsic apoptotic mechanism. These events required sustained extracellular regulated kinase (ERK) 1/2 activation. Silencing of GPER by a specific shRNA partially reversed G-1-mediated cell growth inhibition without affecting ERK activation. These data suggest the existence of G-1 activated but GPER-independent effects that remain to be clarified. In conclusion, this study provides a rational to further study G-1 mechanism of action in order to include this drug as a treatment option to the limited therapy of ACC. PMID:26131713

  19. GPER agonist G-1 decreases adrenocortical carcinoma (ACC) cell growth in vitro and in vivo.

    PubMed

    Chimento, Adele; Sirianni, Rosa; Casaburi, Ivan; Zolea, Fabiana; Rizza, Pietro; Avena, Paola; Malivindi, Rocco; De Luca, Arianna; Campana, Carmela; Martire, Emilia; Domanico, Francesco; Fallo, Francesco; Carpinelli, Giulia; Cerquetti, Lidia; Amendola, Donatella; Stigliano, Antonio; Pezzi, Vincenzo

    2015-08-01

    We have previously demonstrated that estrogen receptor (ER) alpha (ESR1) increases proliferation of adrenocortical carcinoma (ACC) through both an estrogen-dependent and -independent (induced by IGF-II/IGF1R pathways) manner. Then, the use of tamoxifen, a selective estrogen receptor modulator (SERM), appears effective in reducing ACC growth in vitro and in vivo. However, tamoxifen not only exerts antiestrogenic activity, but also acts as full agonist on the G protein-coupled estrogen receptor (GPER). Aim of this study was to investigate the effect of a non-steroidal GPER agonist G-1 in modulating ACC cell growth. We found that G-1 is able to exert a growth inhibitory effect on H295R cells both in vitro and, as xenograft model, in vivo. Treatment of H295R cells with G-1 induced cell cycle arrest, DNA damage and cell death by the activation of the intrinsic apoptotic mechanism. These events required sustained extracellular regulated kinase (ERK) 1/2 activation. Silencing of GPER by a specific shRNA partially reversed G-1-mediated cell growth inhibition without affecting ERK activation. These data suggest the existence of G-1 activated but GPER-independent effects that remain to be clarified. In conclusion, this study provides a rational to further study G-1 mechanism of action in order to include this drug as a treatment option to the limited therapy of ACC. PMID:26131713

  20. Evaluation of the hydrometer for testing immunoglobulin G1 concentrations in Holstein colostrum.

    PubMed

    Pritchett, L C; Gay, C C; Hancock, D D; Besser, T E

    1994-06-01

    Hydrometer measurement in globulin and IgG1 concentration measured by the radial immunodiffusion technique were compared for 915 samples of first milking colostrum from Holstein cows. Least squares analysis of the relationship between hydrometer measurement and IgG1 concentration was improved by log transformation of IgG1 concentration and resulted in a significant linear relationship between hydrometer measurement and log10 IgG1 concentration; r2 = .469. At 50 mg of globulin/ml of colostrum, the recommended hydrometer cutoff point for colostrum selection, the sensitivity of the hydrometer as a test of IgG1 concentration in Holstein colostrum was 26%, and the negative predictive value was 67%. The negative predictive value and sensitivity of the hydrometer as a test of IgG1 in Holstein colostrum was improved, and the cost of misclassification of colostrum was minimized, when the cutoff point for colostrum selection was increased above the recommended 50 mg/ml.

  1. IgG1 Fc N-glycan galactosylation as a biomarker for immune activation

    PubMed Central

    de Jong, Sanne E.; Selman, Maurice H. J.; Adegnika, Ayola A.; Amoah, Abena S.; van Riet, Elly; Kruize, Yvonne C. M.; Raynes, John G.; Rodriguez, Alejandro; Boakye, Daniel; von Mutius, Erika; Knulst, André C.; Genuneit, Jon; Cooper, Philip J.; Hokke, Cornelis H.; Wuhrer, Manfred; Yazdanbakhsh, Maria

    2016-01-01

    Immunoglobulin G (IgG) Fc N-glycosylation affects antibody-mediated effector functions and varies with inflammation rooted in both communicable and non-communicable diseases. Worldwide, communicable and non-communicable diseases tend to segregate geographically. Therefore, we studied whether IgG Fc N-glycosylation varies in populations with different environmental exposures in different parts of the world. IgG Fc N-glycosylation was analysed in serum/plasma of 700 school-age children from different communities of Gabon, Ghana, Ecuador, the Netherlands and Germany. IgG1 galactosylation levels were generally higher in more affluent countries and in more urban communities. High IgG1 galactosylation levels correlated with low total IgE levels, low C-reactive protein levels and low prevalence of parasitic infections. Linear mixed modelling showed that only positivity for parasitic infections was a significant predictor of reduced IgG1 galactosylation levels. That IgG1 galactosylation is a predictor of immune activation is supported by the observation that asthmatic children seemed to have reduced IgG1 galactosylation levels as well. This indicates that IgG1 galactosylation levels could be used as a biomarker for immune activation of populations, providing a valuable tool for studies examining the epidemiological transition from communicable to non-communicable diseases. PMID:27306703

  2. Evaluation of the hydrometer for testing immunoglobulin G1 concentrations in Holstein colostrum.

    PubMed

    Pritchett, L C; Gay, C C; Hancock, D D; Besser, T E

    1994-06-01

    Hydrometer measurement in globulin and IgG1 concentration measured by the radial immunodiffusion technique were compared for 915 samples of first milking colostrum from Holstein cows. Least squares analysis of the relationship between hydrometer measurement and IgG1 concentration was improved by log transformation of IgG1 concentration and resulted in a significant linear relationship between hydrometer measurement and log10 IgG1 concentration; r2 = .469. At 50 mg of globulin/ml of colostrum, the recommended hydrometer cutoff point for colostrum selection, the sensitivity of the hydrometer as a test of IgG1 concentration in Holstein colostrum was 26%, and the negative predictive value was 67%. The negative predictive value and sensitivity of the hydrometer as a test of IgG1 in Holstein colostrum was improved, and the cost of misclassification of colostrum was minimized, when the cutoff point for colostrum selection was increased above the recommended 50 mg/ml. PMID:8083433

  3. 26 CFR 6a.6652(g)-1 - Failure to make return or furnish statement required under section 6039C.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... required under section 6039C. 6a.6652(g)-1 Section 6a.6652(g)-1 Internal Revenue INTERNAL REVENUE SERVICE... OMNIBUS RECONCILIATION ACT OF 1980 § 6a.6652(g)-1 Failure to make return or furnish statement required... limitation under § 6a.6652(g)-1(b)(3) with respect to failure to meet the requirements of section 6039C(c),...

  4. IgG1 deficiency exacerbates experimental autoimmune myasthenia gravis in BALB/c mice.

    PubMed

    Huda, Ruksana; Strait, Richard T; Tüzün, Erdem; Finkelman, Fred D; Christadoss, Premkumar

    2015-04-15

    Myasthenia gravis is an autoimmune disease characterized by muscle weakness due to neuromuscular junction (NMJ) damage by anti-acetylcholine receptor (AChR) auto-antibodies and complement. In experimental autoimmune myasthenia gravis (EAMG), which is induced by immunization with Torpedo AChR in CFA, anti-AChR IgG2b and IgG1 are the predominant isotypes in the circulation. Complement activation by isotypes such as IgG2b plays a crucial role in EAMG pathogenesis; this suggested the possibility that IgG1, which does not activate complement through the classical pathway, may suppress EAMG. In this study, we show that AChR-immunized BALB/c mice genetically deficient for IgG1 produce higher levels of complement-activating isotypes of anti-AChR, especially IgG3 and IgG2a, and develop increased IgG3/IgG2a deposits at the NMJ, as compared to wild type (WT) BALB/c mice. Consistent with this, AChR-immunized IgG1(-/-) BALB/c mice lose muscle strength and muscle AChR to a greater extent than AChR-immunized WT mice. These observations demonstrate that IgG1 deficiency leads to increased severity of EAMG associated with an increase in complement activating IgG isotypes. Further studies are needed to dissect the specific role or mechanism of IgG1 in limiting EAMG and that of EAMG exacerbating role of complement activating IgG3 and IgG2a in IgG1 deficiency. PMID:25867470

  5. IgG1 deficiency exacerbates experimental autoimmune myasthenia gravis in BALB/c mice.

    PubMed

    Huda, Ruksana; Strait, Richard T; Tüzün, Erdem; Finkelman, Fred D; Christadoss, Premkumar

    2015-04-15

    Myasthenia gravis is an autoimmune disease characterized by muscle weakness due to neuromuscular junction (NMJ) damage by anti-acetylcholine receptor (AChR) auto-antibodies and complement. In experimental autoimmune myasthenia gravis (EAMG), which is induced by immunization with Torpedo AChR in CFA, anti-AChR IgG2b and IgG1 are the predominant isotypes in the circulation. Complement activation by isotypes such as IgG2b plays a crucial role in EAMG pathogenesis; this suggested the possibility that IgG1, which does not activate complement through the classical pathway, may suppress EAMG. In this study, we show that AChR-immunized BALB/c mice genetically deficient for IgG1 produce higher levels of complement-activating isotypes of anti-AChR, especially IgG3 and IgG2a, and develop increased IgG3/IgG2a deposits at the NMJ, as compared to wild type (WT) BALB/c mice. Consistent with this, AChR-immunized IgG1(-/-) BALB/c mice lose muscle strength and muscle AChR to a greater extent than AChR-immunized WT mice. These observations demonstrate that IgG1 deficiency leads to increased severity of EAMG associated with an increase in complement activating IgG isotypes. Further studies are needed to dissect the specific role or mechanism of IgG1 in limiting EAMG and that of EAMG exacerbating role of complement activating IgG3 and IgG2a in IgG1 deficiency.

  6. RESEARCH PAPER: Old stellar population synthesis: new age and mass estimates for Mayall II = G1

    NASA Astrophysics Data System (ADS)

    Ma, Jun; de Grijs, Richard; Fan, Zhou; Rey, Soo-Chang; Wu, Zhen-Yu; Zhou, Xu; Wu, Jiang-Hua; Jiang, Zhao-Ji; Chen, Jian-Sheng; Lee, Kyungsook; Sohn, Sangmo Tony

    2009-06-01

    Mayall II = G1 is one of the most luminous globular clusters (GCs) in M31. Here, we determine its age and mass by comparing multicolor photometry with theoretical stellar population synthesis models. Based on far- and near-ultraviolet GALEX photometry, broad-band UBVRI, and infrared JHKS 2MASS data, we construct the most extensive spectral energy distribution of G1 to date, spanning the wavelength range from 1538 to 20 000 Å. A quantitative comparison with a variety of simple stellar population (SSP) models yields a mean age which is consistent with G1 being among the oldest building blocks of M31 and having formed within ~1.7 Gyr after the Big Bang. Irrespective of the SSP model or stellar initial mass function adopted, the resulting mass estimates (of order 107 Modot) indicate that G1 is one of the most massive GCs in the Local Group. However, we speculate that the cluster's exceptionally high mass suggests that it may not be a genuine GC. Our results also suggest that G1 may contain, on average, (1.65±0.63) × 102 Lodot far-ultraviolet-bright, hot, extreme horizontal-branch stars, depending on the adopted SSP model. In addition, we demonstrate that extensive multi-passband photometry coupled with SSP analysis enables one to obtain age estimates for old SSPs that have similar accuracies as those from integrated spectroscopy or resolved stellar photometry, provided that some of the free parameters can be constrained independently.

  7. Laparoscopic and endoscopic cooperative surgery for duodenal neuroendocrine tumor (NET) G1: Report of a case

    PubMed Central

    Tsushimi, Takaaki; Mori, Hirohito; Harada, Takasuke; Nagase, Takashi; Iked, Yoshitaka; Ohnishi, Hiromo

    2014-01-01

    INTRODUCTION We report a case of duodenal neuroendocrine tumor (NET) G1 resected by laparoscopic and endoscopic cooperative surgery (LECS) technique. PRESENTATION OF CASE A 58-year-old woman underwent esophagastroduodenoscopy, revealing an 8-mm, gently rising tumor distal to the pylorus, on the anterior wall of the duodenal bulb. Endoscopic ultrasonography suggested the tumor might invade the submucosal layer. The tumor was pathologically diagnosed as a G1 duodenal NET, by biopsy. Endoscopic submucosal dissection was attempted, but was unsuccessful because of the difficulty of endoscopically performing an inversion operation in the narrow working space. The case was further complicated by the patient's duodenal ulcer scar. We performed a full-thickness local excision using laparoscopic and endoscopic cooperative surgery. The tumor was confirmed and endoscopically marked along the resection line. After full-thickness excision, using endoscopy and laparoscopy, interrupted full-thickness closure was performed laparoscopically. DISCUSSION Endoscopic treatment is generally recommended for G1 NETs <10 mm in diameter and extending only to the submucosal layer. However, some cases are difficult to resect endoscopically because the wall of duodenum is thinner than that of stomach, and endoscope maneuverability is limited within the narrow working space. LECS is appropriate for early duodenal G1 NETs because they are less invasive and resection of the lesion area is possible. CONCLUSION We demonstrated that LECS is a safe and feasible procedure for duodenal G1 NETs in the anterior wall of the first portion of the duodenum. PMID:25460463

  8. A Dynamical Framework for the All-or-None G1/S Transition.

    PubMed

    Barr, Alexis R; Heldt, Frank S; Zhang, Tongli; Bakal, Chris; Novák, Béla

    2016-01-27

    The transition from G1 into DNA replication (S phase) is an emergent behavior resulting from dynamic and complex interactions between cyclin-dependent kinases (Cdks), Cdk inhibitors (CKIs), and the anaphase-promoting complex/cyclosome (APC/C). Understanding the cellular decision to commit to S phase requires a quantitative description of these interactions. We apply quantitative imaging of single human cells to track the expression of G1/S regulators and use these data to parametrize a stochastic mathematical model of the G1/S transition. We show that a rapid, proteolytic, double-negative feedback loop between Cdk2:Cyclin and the Cdk inhibitor p27(Kip1) drives a switch-like entry into S phase. Furthermore, our model predicts that increasing Emi1 levels throughout S phase are critical in maintaining irreversibility of the G1/S transition, which we validate using Emi1 knockdown and live imaging of G1/S reporters. This work provides insight into the general design principles of the signaling networks governing the temporally abrupt transitions between cell-cycle phases. PMID:27136687

  9. A Dynamical Framework for the All-or-None G1/S Transition

    PubMed Central

    Barr, Alexis R.; Heldt, Frank S.; Zhang, Tongli; Bakal, Chris; Novák, Béla

    2016-01-01

    Summary The transition from G1 into DNA replication (S phase) is an emergent behavior resulting from dynamic and complex interactions between cyclin-dependent kinases (Cdks), Cdk inhibitors (CKIs), and the anaphase-promoting complex/cyclosome (APC/C). Understanding the cellular decision to commit to S phase requires a quantitative description of these interactions. We apply quantitative imaging of single human cells to track the expression of G1/S regulators and use these data to parametrize a stochastic mathematical model of the G1/S transition. We show that a rapid, proteolytic, double-negative feedback loop between Cdk2:Cyclin and the Cdk inhibitor p27Kip1 drives a switch-like entry into S phase. Furthermore, our model predicts that increasing Emi1 levels throughout S phase are critical in maintaining irreversibility of the G1/S transition, which we validate using Emi1 knockdown and live imaging of G1/S reporters. This work provides insight into the general design principles of the signaling networks governing the temporally abrupt transitions between cell-cycle phases. PMID:27136687

  10. A comparative genomic analysis of the alkalitolerant soil bacterium Bacillus lehensis G1.

    PubMed

    Noor, Yusuf Muhammad; Samsulrizal, Nurul Hidayah; Jema'on, Noor Azah; Low, Kheng Oon; Ramli, Aizi Nor Mazila; Alias, Noor Izawati; Damis, Siti Intan Rosdianah; Fuzi, Siti Fatimah Zaharah Mohd; Isa, Mohd Noor Mat; Murad, Abdul Munir Abdul; Raih, Mohd Firdaus Mohd; Bakar, Farah Diba Abu; Najimudin, Nazalan; Mahadi, Nor Muhammad; Illias, Rosli Md

    2014-07-25

    Bacillus lehensis G1 is a Gram-positive, moderately alkalitolerant bacterium isolated from soil samples. B. lehensis produces cyclodextrin glucanotransferase (CGTase), an enzyme that has enabled the extensive use of cyclodextrin in foodstuffs, chemicals, and pharmaceuticals. The genome sequence of B. lehensis G1 consists of a single circular 3.99 Mb chromosome containing 4017 protein-coding sequences (CDSs), of which 2818 (70.15%) have assigned biological roles, 936 (23.30%) have conserved domains with unknown functions, and 263 (6.55%) have no match with any protein database. Bacillus clausii KSM-K16 was established as the closest relative to B. lehensis G1 based on gene content similarity and 16S rRNA phylogenetic analysis. A total of 2820 proteins from B. lehensis G1 were found to have orthologues in B. clausii, including sodium-proton antiporters, transport proteins, and proteins involved in ATP synthesis. A comparative analysis of these proteins and those in B. clausii and other alkaliphilic Bacillus species was carried out to investigate their contributions towards the alkalitolerance of the microorganism. The similarities and differences in alkalitolerance-related genes among alkalitolerant/alkaliphilic Bacillus species highlight the complex mechanism of pH homeostasis. The B. lehensis G1 genome was also mined for proteins and enzymes with potential viability for industrial and commercial purposes. PMID:24811681

  11. PVP and G1.5 PAMAM dendrimer co-mediated synthesis of silver nanoparticles

    SciTech Connect

    Li Guoping; Luo Yunjun . E-mail: yjluo@bit.edu.cn; Tan Huimin

    2005-04-15

    PVP and G1.5 PAMAM dendrimer co-mediated silver nanoparticles of smaller than 5nm in diameter were prepared using H{sub 2} as reducing agent. With the TEM micrograph, it was found that the molar ratios of PVP and G1.5 PAMAM dendrimer have significant effect in the morphology and size distribution of silver nanoparticles. The reaction rate (fitting a first-order equation) was strongly influenced by the molar ratios of PVP and G1.5 PAMAM dendrimer and the reaction temperature. From the UV-Vis spectra of an aqueous solution of silver nanoparticles, they could be stored for at least 2 months without coagulation at room temperature.

  12. Acute appendicitis with a neuroendocrine tumor G1 (carcinoid): pitfalls of conservative treatment.

    PubMed

    Watanabe, Hiroyuki A; Fujimoto, Taketoshi; Kato, Yo; Sasaki, Mayumi; Ikusue, Toshikazu

    2016-08-01

    A man in his early thirties presented to our clinic with right lower abdominal pain. Computed tomography (CT) and ultrasonography (US) revealed a swollen appendix and an appendicolith. Abscess formation was not observed but ongoing appendiceal rupture was not ruled out. Three months after successful conservative therapy, the lumen of the apical portion was kept dilated and laparoscopic interval appendectomy was performed. No tumorous findings were observed macroscopically. However, histology revealed many tiny nests infiltrating the submucosa, muscular layer, and subserosa at the root of the appendix. An appendiceal neuroendocrine tumor G1 (NET G1; carcinoid) was diagnosed immunohistologically. Neither CT nor US visualized the tumor because of its non-tumor-forming but infiltrative growth. In conclusion, after successful conservative treatment, interval appendectomy should be considered to uncover a possible appendiceal NET G1 (carcinoid), particularly when dilatation of the distal lumen is kept under observation. PMID:27311320

  13. Detection of histidine oxidation in a monoclonal immunoglobulin gamma (IgG) 1 antibody.

    PubMed

    Amano, Masato; Kobayashi, Naoki; Yabuta, Masayuki; Uchiyama, Susumu; Fukui, Kiichi

    2014-08-01

    Although oxidation of methionine and tryptophan are known as popular chemical modifications that occur in monoclonal antibody (mAb) molecules, oxidation of other amino acids in mAbs has not been reported to date. In this study, oxidation of the histidine residue in a human immunoglobulin gamma (IgG) 1 molecule was discovered for the first time by mass spectrometry. The oxidation of a specific histidine located at the CH2 domain of IgG1 occurred under light stress, but it was not observed under heat stress. With the forced degradation study using several reactive oxygen species, the singlet oxygen was attributed to a reactive source of the histidine oxidation. The reaction mechanism of the histidine oxidation was proposed on the basis of the mass spectrometric analysis of IgG1 oxidized in deuterium oxide and hydrogen heavy oxide. PMID:24940720

  14. Doxazosin inhibits retinoblastoma protein phosphorylation and G(1)-->S transition in human coronary smooth muscle cells.

    PubMed

    Kintscher, U; Wakino, S; Kim, S; Jackson, S M; Fleck, E; Hsueh, W A; Law, R E

    2000-05-01

    Previous studies have demonstrated that the alpha(1)-adrenergic receptor antagonist doxazosin (Dox) inhibits multiple mitogenic signaling pathways in human vascular smooth muscle cells. This broad antiproliferative activity of Dox occurs through a novel mechanism unrelated to its blocking the alpha(1)-adrenergic receptor. Flow cytometry demonstrated that Dox prevents mitogen-induced G(1)-->S progression of human coronary artery smooth muscle cells (CASMCs) in a dose-dependent manner, with a maximal reduction of S-phase transition by 88+/-10.5% in 20 ng/mL platelet-derived growth factor and 1 micromol/L insulin (P+I)-stimulated cells (P<0.01 for 10 micromol/L Dox versus P+I alone) and 52+/-18.7% for 10% FBS-induced mitogenesis (P<0.05 for 10 micromol/L Dox versus 10% FBS alone). Inhibition of G(1) exit by Dox was accompanied by a significant blockade of retinoblastoma protein (Rb) phosphorylation. Hypophosphorylated Rb sequesters the E2F transcription factor, leading to G(1) arrest. Adenoviral overexpression of E2F-1 stimulated quiescent CASMCs to progress through G(1) and enter the S phase. E2F-mediated G(1) exit was not affected by Dox, suggesting that it targets events upstream from Rb hyperphosphorylation. Downregulation of the cyclin-dependent kinase inhibitory protein p27 is important for maximal activation of G(1) cyclin/cyclin-dependent kinase holoenzymes to overcome the cell cycle inhibitory activity of Rb. In Western blot analysis, p27 levels decreased after mitogenic stimulation (after P+I, 43+/-1.8% of quiescent cells [P<0.01 versus quiescent cells]; after 10% FBS, 55+/-7.7% of quiescent cells [P<0. 05 versus quiescent cells]), whereas the addition of Dox (10 micromol/L) markedly attenuated its downregulation (after P+I, 90+/-8.3% of quiescent cells [P<0.05 versus P+I alone]; after 10% FBS, 78+/-8.3% of quiescent cells [P<0.05 versus 10% FBS alone]). Furthermore, Dox inhibited cyclin A expression, an E2F regulated gene that is essential for cell cycle

  15. 17 CFR 240.12g-1 - Exemption from section 12(g).

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... pursuant to section 12(g)(1) if on the last day of its most recent fiscal year the issuer had total assets... quoted in an automated inter-dealer quotation system. (Secs. 6, 7, 8, 10, 19(a), 48 Stat. 78, 79, 81,...

  16. 26 CFR 301.6501(g)-1 - Certain income tax returns of corporations.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 26 Internal Revenue 18 2010-04-01 2010-04-01 false Certain income tax returns of corporations. 301... and Collection § 301.6501(g)-1 Certain income tax returns of corporations. (a) Trusts or partnerships... and additions to the tax, additional amounts and assessable penalties allowed by chapter 68 of...

  17. 26 CFR 301.6501(g)-1 - Certain income tax returns of corporations.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 26 Internal Revenue 18 2011-04-01 2011-04-01 false Certain income tax returns of corporations. 301... and Collection § 301.6501(g)-1 Certain income tax returns of corporations. (a) Trusts or partnerships... and additions to the tax, additional amounts and assessable penalties allowed by chapter 68 of...

  18. Human IgG1 antibodies suppress angiogenesis in a target-independent manner

    PubMed Central

    Bogdanovich, Sasha; Kim, Younghee; Mizutani, Takeshi; Yasuma, Reo; Tudisco, Laura; Cicatiello, Valeria; Bastos-Carvalho, Ana; Kerur, Nagaraj; Hirano, Yoshio; Baffi, Judit Z; Tarallo, Valeria; Li, Shengjian; Yasuma, Tetsuhiro; Arpitha, Parthasarathy; Fowler, Benjamin J; Wright, Charles B; Apicella, Ivana; Greco, Adelaide; Brunetti, Arturo; Ruvo, Menotti; Sandomenico, Annamaria; Nozaki, Miho; Ijima, Ryo; Kaneko, Hiroki; Ogura, Yuichiro; Terasaki, Hiroko; Ambati, Balamurali K; Leusen, Jeanette HW; Langdon, Wallace Y; Clark, Michael R; Armour, Kathryn L; Bruhns, Pierre; Verbeek, J Sjef; Gelfand, Bradley D; De Falco, Sandro; Ambati, Jayakrishna

    2016-01-01

    Aberrant angiogenesis is implicated in diseases affecting nearly 10% of the world’s population. The most widely used anti-angiogenic drug is bevacizumab, a humanized IgG1 monoclonal antibody that targets human VEGFA. Although bevacizumab does not recognize mouse Vegfa, it inhibits angiogenesis in mice. Here we show bevacizumab suppressed angiogenesis in three mouse models not via Vegfa blockade but rather Fc-mediated signaling through FcγRI (CD64) and c-Cbl, impairing macrophage migration. Other approved humanized or human IgG1 antibodies without mouse targets (adalimumab, alemtuzumab, ofatumumab, omalizumab, palivizumab and tocilizumab), mouse IgG2a, and overexpression of human IgG1-Fc or mouse IgG2a-Fc, also inhibited angiogenesis in wild-type and FcγR humanized mice. This anti-angiogenic effect was abolished by Fcgr1 ablation or knockdown, Fc cleavage, IgG-Fc inhibition, disruption of Fc-FcγR interaction, or elimination of FcRγ-initated signaling. Furthermore, bevacizumab’s Fc region potentiated its anti-angiogenic activity in humanized VEGFA mice. Finally, mice deficient in FcγRI exhibited increased developmental and pathological angiogenesis. These findings reveal an unexpected anti-angiogenic function for FcγRI and a potentially concerning off-target effect of hIgG1 therapies. PMID:26918197

  19. 17 CFR 275.202(a)(11)(G)-1 - Family offices.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 17 Commodity and Securities Exchanges 3 2012-04-01 2012-04-01 false Family offices. 275.202(a)(11... COMMISSION (CONTINUED) RULES AND REGULATIONS, INVESTMENT ADVISERS ACT OF 1940 § 275.202(a)(11)(G)-1 Family offices. (a) Exclusion. A family office, as defined in this section, shall not be considered to be...

  20. 17 CFR 275.202(a)(11)(G)-1 - Family offices.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 17 Commodity and Securities Exchanges 4 2014-04-01 2014-04-01 false Family offices. 275.202(a)(11... COMMISSION (CONTINUED) RULES AND REGULATIONS, INVESTMENT ADVISERS ACT OF 1940 § 275.202(a)(11)(G)-1 Family offices. (a) Exclusion. A family office, as defined in this section, shall not be considered to be...

  1. 17 CFR 275.202(a)(11)(G)-1 - Family offices.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 17 Commodity and Securities Exchanges 3 2013-04-01 2013-04-01 false Family offices. 275.202(a)(11... COMMISSION (CONTINUED) RULES AND REGULATIONS, INVESTMENT ADVISERS ACT OF 1940 § 275.202(a)(11)(G)-1 Family offices. (a) Exclusion. A family office, as defined in this section, shall not be considered to be...

  2. An Overview of Observations Made from the G-1 Aircraft during TexAQS 2000

    NASA Astrophysics Data System (ADS)

    Kleinman, L. I.; Daum, P. H.; Brechtel, F.; Buzorius, G.; Lee, Y.; Nunnermacker, L. J.; Springston, S. R.; Weinstein-Lloyd, J.; Zheng, J.

    2002-12-01

    As part of the Texas Air Quality Study, the DOE G-1 aircraft conducted 18 research flights in and around the Houston metropolitan area between August 19 and September 12, 2000. Overall objectives were to characterize emissions and their relation to the production of O3 and particulate matter. Chemical parameters measured on the G-1 included the concentrations of O3, NO, NO2, NOy, CO, speciated VOCs, HCHO, SO2, H2O2, organic peroxides, and aerosol anions and cations. Particle size distributions were obtained over the range 5nm to 15 um. This presentation is an overview of the G-1 data set and secondary quantities such as the production rate of O3 which can be calculated using trace gas observations from the G-1. In order to put into perspective the extraordinarily high concentrations of reactive VOCs and concomitantly high O3 production rates observed in Houston, we will rely on similar aircraft measurements from Nashville, New York City, Phoenix, and Philadelphia.

  3. H, G1, G2 photometric phase function extended to low-accuracy data

    NASA Astrophysics Data System (ADS)

    Penttilä, A.; Shevchenko, V. G.; Wilkman, O.; Muinonen, K.

    2016-04-01

    We introduce a constrained nonlinear least-squares algorithm to be used in estimating the parameters in the H, G1, G2 phase function. As the algorithm works directly in the magnitude space, it will surpass the possible bias problem that may be present in the existing H ,G1 ,G2 fit procedure when applied to low-accuracy observations with large magnitude variations. With constraints on the photometric phase-curve shape parameters G1 and G2, it guarantees a physically reasonable phase-curve estimate. With a new data set of 93 asteroids, we re-assess the two-parameter version of the H ,G1 ,G2 function. Finally, we introduce a one-parameter version of the phase function that can give a suggestion of the asteroids taxonomic group based only on its phase curve. A statistical model selection procedure is presented that can automatically select between the different versions of the photometric phase functions. An online tool that implements these algorithms is introduced.

  4. 26 CFR 301.6501(g)-1 - Certain income tax returns of corporations.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 26 Internal Revenue 18 2012-04-01 2012-04-01 false Certain income tax returns of corporations. 301... and Collection § 301.6501(g)-1 Certain income tax returns of corporations. (a) Trusts or partnerships... and additions to the tax, additional amounts and assessable penalties allowed by chapter 68 of...

  5. 26 CFR 301.6501(g)-1 - Certain income tax returns of corporations.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 26 Internal Revenue 18 2014-04-01 2014-04-01 false Certain income tax returns of corporations. 301... and Collection § 301.6501(g)-1 Certain income tax returns of corporations. (a) Trusts or partnerships... and additions to the tax, additional amounts and assessable penalties allowed by chapter 68 of...

  6. 26 CFR 301.6501(g)-1 - Certain income tax returns of corporations.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 26 Internal Revenue 18 2013-04-01 2013-04-01 false Certain income tax returns of corporations. 301... and Collection § 301.6501(g)-1 Certain income tax returns of corporations. (a) Trusts or partnerships... and additions to the tax, additional amounts and assessable penalties allowed by chapter 68 of...

  7. Overview of Shipyard coast line with Piers G1, G2, G3, ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    Overview of Shipyard coast line with Piers G-1, G-2, G-3, G-4, and G-5 in view, view facing east-southeast - U.S. Naval Base, Pearl Harbor, Pier & Quay Walls, Entrance to Dry Dock No. 2 & Repair Wharfs, east & west sides of Dry Dock No. 2 & west side of Dry Dock No. 3, Pearl City, Honolulu County, HI

  8. Learning Progression of Ecological System Reasoning for Lower Elementary (G1-4) Students

    ERIC Educational Resources Information Center

    Hokayem, Hayat Al

    2012-01-01

    In this study, I utilized a learning progression framework to investigate lower elementary students (G1-4) systemic reasoning in ecology and I related students reasoning to their sources of knowledge. I used semi-structured interviews with 44 students from first through fourth grade, four teachers, and eight parents. The results revealed that a…

  9. HPV-18 transformed cells fail to arrest in G1 in response to quercetin treatment.

    PubMed

    Beniston, R G; Campo, M S

    2005-05-01

    Previous work with primary human keratinocytes demonstrated that quercetin, a potent mutagen found in high levels in bracken fern (Pteridium aquilinum), arrested cells in G1 with concomitant elevation of the cyclin-dependent kinase inhibitor (cdki) p27Kip1. Expression of the human papillomavirus type 16 (HPV-16) E6 and E7 oncoproteins, under transcriptional control of a heterologous promoter, in transformed keratinocytes failed to abrogate this arrest [Beniston, R., Campo, M.S., 2003. Quercetin elevates p27Kip1 and arrests both primary and HPV-16 E6/E7 transformed human keratinocytes in G1. Oncogene 22, 5504-5514]. Given the link between papillomavirus infection, bracken fern in the diet and cancer of the oesophagus in humans, we wished to investigate further whether cells transformed by the whole genome of HPV-16 or HPV-18, with E6 and E7 under the transcriptional control of their respective homologous promoters, would be similarly arrested in G1 by quercetin. In agreement with earlier work, quercetin arrested HPV-16 transformed cells in G1 with an increase in the cyclin-dependent kinase inhibitor p27Kip1. However, HPV-18 transformed cells did not arrest after quercetin treatment. The failure of HPV-18 transformed cells to arrest in G1 was linked to the up-regulation of the HPV-18 long control region (LCR) by quercetin, maintaining high expression of the viral transforming proteins. Transcriptional up-regulation of the HPV-18 LCR was mediated by a "quercetin responsive element" homologous to the one identified previously in the bovine papillomavirus type 4 (BPV-4) LCR.

  10. A mechanism for the suppression of homologous recombination in G1 cells.

    PubMed

    Orthwein, Alexandre; Noordermeer, Sylvie M; Wilson, Marcus D; Landry, Sébastien; Enchev, Radoslav I; Sherker, Alana; Munro, Meagan; Pinder, Jordan; Salsman, Jayme; Dellaire, Graham; Xia, Bing; Peter, Matthias; Durocher, Daniel

    2015-12-17

    DNA repair by homologous recombination is highly suppressed in G1 cells to ensure that mitotic recombination occurs solely between sister chromatids. Although many homologous recombination factors are cell-cycle regulated, the identity of the events that are both necessary and sufficient to suppress recombination in G1 cells is unknown. Here we report that the cell cycle controls the interaction of BRCA1 with PALB2-BRCA2 to constrain BRCA2 function to the S/G2 phases in human cells. We found that the BRCA1-interaction site on PALB2 is targeted by an E3 ubiquitin ligase composed of KEAP1, a PALB2-interacting protein, in complex with cullin-3 (CUL3)-RBX1 (ref. 6). PALB2 ubiquitylation suppresses its interaction with BRCA1 and is counteracted by the deubiquitylase USP11, which is itself under cell cycle control. Restoration of the BRCA1-PALB2 interaction combined with the activation of DNA-end resection is sufficient to induce homologous recombination in G1, as measured by RAD51 recruitment, unscheduled DNA synthesis and a CRISPR-Cas9-based gene-targeting assay. We conclude that the mechanism prohibiting homologous recombination in G1 minimally consists of the suppression of DNA-end resection coupled with a multi-step block of the recruitment of BRCA2 to DNA damage sites that involves the inhibition of BRCA1-PALB2-BRCA2 complex assembly. We speculate that the ability to induce homologous recombination in G1 cells with defined factors could spur the development of gene-targeting applications in non-dividing cells. PMID:26649820

  11. 26 CFR 1.1402(g)-1 - Treatment of certain remuneration erroneously reported as net earnings from self-employment.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... reported as net earnings from self-employment. 1.1402(g)-1 Section 1.1402(g)-1 Internal Revenue INTERNAL...-Employment Income § 1.1402(g)-1 Treatment of certain remuneration erroneously reported as net earnings from... request, and should indicate clearly that it is a request that, pursuant to section 1402(g) of the...

  12. 26 CFR 1.904(g)-1T - Overall domestic loss and the overall domestic loss account (temporary).

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... loss account (temporary). 1.904(g)-1T Section 1.904(g)-1T Internal Revenue INTERNAL REVENUE SERVICE... United States § 1.904(g)-1T Overall domestic loss and the overall domestic loss account (temporary). (a... accounts for purposes of section 904(g). Section 1.904(g)-2T provides rules for recapturing the balance...

  13. 17 CFR 240.17g-1 - Application for registration as a nationally recognized statistical rating organization.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... as a nationally recognized statistical rating organization. 240.17g-1 Section 240.17g-1 Commodity and... Statistical Rating Organizations § 240.17g-1 Application for registration as a nationally recognized statistical rating organization. (a) Initial application. A credit rating agency applying to the Commission...

  14. 17 CFR 270.17g-1 - Bonding of officers and employees of registered management investment companies.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... employees of registered management investment companies. 270.17g-1 Section 270.17g-1 Commodity and... ACT OF 1940 § 270.17g-1 Bonding of officers and employees of registered management investment companies. (a) Each registered management investment company shall provide and maintain a bond which...

  15. 17 CFR 240.17g-1 - Application for registration as a nationally recognized statistical rating organization.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... as a nationally recognized statistical rating organization. 240.17g-1 Section 240.17g-1 Commodity and... Statistical Rating Organizations § 240.17g-1 Application for registration as a nationally recognized statistical rating organization. (a) Initial application. A credit rating agency applying to the Commission...

  16. 17 CFR 240.17g-1 - Application for registration as a nationally recognized statistical rating organization.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... as a nationally recognized statistical rating organization. 240.17g-1 Section 240.17g-1 Commodity and... Statistical Rating Organizations § 240.17g-1 Application for registration as a nationally recognized statistical rating organization. (a) Initial application. A credit rating agency applying to the Commission...

  17. 17 CFR 240.17g-1 - Application for registration as a nationally recognized statistical rating organization.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... as a nationally recognized statistical rating organization. 240.17g-1 Section 240.17g-1 Commodity and... Statistical Rating Organizations § 240.17g-1 Application for registration as a nationally recognized statistical rating organization. (a) Initial application. A credit rating agency applying to the Commission...

  18. 17 CFR 240.17g-1 - Application for registration as a nationally recognized statistical rating organization.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... as a nationally recognized statistical rating organization. 240.17g-1 Section 240.17g-1 Commodity and... Statistical Rating Organizations § 240.17g-1 Application for registration as a nationally recognized statistical rating organization. (a) Initial application. A credit rating agency applying to the Commission...

  19. Global Structure of HIV-1 Neutralizing Antibody IgG1 b12 is Asymmetric

    SciTech Connect

    Ashish, F.; Solanki, A; Boone, C; Krueger, J

    2010-01-01

    Human antibody IgG1 b12 is one of the four antibodies known to neutralize a broad range of human immunodeficiency virus-1. The crystal structure of this antibody displayed an asymmetric disposition of the Fab arms relative to its Fc portion. Comparison of structures solved for other IgG1 antibodies led to a notion that crystal packing forces entrapped a 'snap-shot' of different conformations accessible to this antibody. To elucidate global structure of this unique antibody, we acquired small-angle X-ray scattering data from its dilute solution. Data analysis indicated that b12 adopts a bilobal globular structure in solution with a radius of gyration and a maximum linear dimension of {approx}54 and {approx}180 {angstrom}, respectively. Extreme similarity between its solution and crystal structure concludes that non-flexible, asymmetric shape is an inherent property of this rare antibody.

  20. Permanent draft genome sequence of the gliding predator Saprospira grandis strain Sa g1 (= HR1)

    SciTech Connect

    Mavromatis, K; Chertkov, Olga; Lapidus, Alla L.; Nolan, Matt; Lucas, Susan; Tice, Hope; Glavina Del Rio, Tijana; Cheng, Jan-Fang; Han, Cliff; Tapia, Roxanne; Bruce, David; Goodwin, Lynne A.; Pitluck, Sam; Huntemann, Marcel; Liolios, Konstantinos; Pagani, Ioanna; Ivanova, N; Mikhailova, Natalia; Pati, Amrita; Chen, Amy; Palaniappan, Krishna; Land, Miriam L; Brambilla, Evelyne-Marie; Rohde, Manfred; Spring, Stefan; Goker, Markus; Detter, J. Chris; Bristow, James; Eisen, Jonathan; Markowitz, Victor; Hugenholtz, Philip; Kyrpides, Nikos C; Klenk, Hans-Peter; Woyke, Tanja

    2012-01-01

    Saprospira grandis Gross et al. 1911 is a member of the Saprospiraceae, a family in the class 'Sphingobacteria' that remains poorly characterized at the genomic level. The species is known for preying on other marine bacteria via 'ixotrophy'. S. grandis strain Sa g1 was isolated from decaying crab carapace in France and was selected for genome sequencing because of its isolated location in the tree of life. Only one type strain genome has been published so far from the Saprospiraceae, while the sequence of strain Sa g1 represents the second genome to be published from a non-type strain of S. grandis. Here we describe the features of this organism, together with the complete genome sequence and annotation. The 4,495,250 bp long Improved-High-Quality draft of the genome with its 3,536 protein-coding and 62 RNA genes is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

  1. Implications of intermediate mass black hole in globular cluster G1 on dark matter detection.

    SciTech Connect

    Zaharijas, G.; High Energy Physics

    2008-07-01

    Recently there has been growing evidence in favor of the presence of an intermediate mass black hole in the globular cluster G1, in Andromeda Galaxy. Under the assumption that formation of this globular cluster occurred within a dark matter halo, we explore whether the presence of a black hole could result in an observable gamma ray signal due to dark matter annihilation in this globular cluster. Starting from an initial Navarro-Frenk-White matter profile, with density parameters consistent with G1 observations, we find that indeed, if the spike in the density has been formed and has survived until the present, the signal could be observed by GLAST and current atmospheric Cerenkov telescope detectors.

  2. Implications of the intermediate mass black hole in globular cluster G1 on dark matter detection

    SciTech Connect

    Zaharijas, Gabrijela

    2008-07-15

    Recently there has been growing evidence in favor of the presence of an intermediate mass black hole in the globular cluster G1, in Andromeda Galaxy. Under the assumption that formation of this globular cluster occurred within a dark matter halo, we explore whether the presence of a black hole could result in an observable gamma ray signal due to dark matter annihilation in this globular cluster. Starting from an initial Navarro-Frenk-White matter profile, with density parameters consistent with G1 observations, we find that indeed, if the spike in the density has been formed and has survived until the present, the signal could be observed by GLAST and current atmospheric Cerenkov telescope detectors.

  3. The existence of inflection points for generalized log-aesthetic curves satisfying G1 data

    NASA Astrophysics Data System (ADS)

    Karpagavalli, R.; Gobithaasan, R. U.; Miura, K. T.; Shanmugavel, Madhavan

    2015-12-01

    Log-Aesthetic (LA) curves have been implemented in a CAD/CAM system for various design feats. LA curves possess linear Logarithmic Curvature Graph (LCG) with gradient (shape parameter) denoted as α. In 2009, a generalized form of LA curves called Generalized Log-Aesthetic Curves (GLAC) has been proposed which has an extra shape parameter as ν compared to LA curves. Recently, G1 continuous GLAC algorithm has been proposed which utilizes the extra shape parameter using four control points. This paper discusses on the existence of inflection points in a GLAC segment satisfying G1 Hermite data and the effect of inflection point on convex hull property. It is found that the existence of inflection point can be avoided by manipulating the value of α. Numerical experiments show that the increase of α may remove the inflection point (if any) in a GLAC segment.

  4. Echinococcus ortleppi and E. granulosus G1, G2 and G3 genotypes in Italian bovines.

    PubMed

    Casulli, Adriano; Manfredi, Maria Teresa; La Rosa, Giuseppe; Cerbo, Anna Rita Di; Genchi, Claudio; Pozio, Edoardo

    2008-08-01

    To increase the knowledge on Echinococcus genotypes infesting cattle and water buffaloes (Bubalus bubalis) born and bred in Italy, the germinal layer of hydatid cysts was collected from the liver and the lungs of 80 animals slaughtered in 2007. Two mitochondrial genes (the cytochrome c oxidase subunit I and the NADH subunit I) were tested by PCR. Four genotypes were identified: G1 (sheep strain), G2 (Tasmanian sheep strain), G3 (buffalo strain), and G5 (cattle strain). Fertile cysts were detected only in the lungs of 4.5% of the total G1 lung cysts, of 9.4% of the total G3 lung cysts, and in the only G5 infected animal. This is the first report of Echinococcus ortleppi (genotype G5) in Italy.

  5. Aircraft measurements of aerosol properties during GoAmazon - G1 and HALO inter-comparison

    NASA Astrophysics Data System (ADS)

    Mei, F.; Cecchini, M. A.; Wang, J.; Tomlinson, J. M.; Comstock, J. M.; Hubbe, J. M.; Pekour, M. S.; Machado, L.; Wendisch, M.; Longo, K.; Martin, S. T.; Schmid, B.; Weinzierl, B.; Krüger, M. L.; Zöger, M.

    2015-12-01

    Currently, the indirect effects of atmospheric aerosols remain the most uncertain components in forcing of climate change over the industrial period (IPCC, 2013). This large uncertainty is partially a result of our incomplete understanding of the ability of particles to form cloud droplets under atmospherically relevant supersaturations. One objective of the US Department of Energy (DOE) Green Ocean Amazon Project (GoAmazon2014/5) is to understand the influence of the emission from Manaus, a tropical megacity, on aerosol size, concentration, and chemical composition, and their impact on cloud condensation nuclei (CCN) spectrum. The GoAmazon2014/5 study was an international campaign with the collaboration efforts from US, Brazil and Germany. During the intensive operation period, in the dry season (Sep. 1st - Oct. 10th, 2014), aerosol concentration, size distributions, and CCN spectra, both under pristine conditions and inside the Manaus plume, were characterized in-situ from the DOE Gulfstream-1 (G-1) research aircraft and German HALO aircraft during 4 coordinated flights on Sep. 9th, Sep. 16th, Sep 21st and Oct. 1st, 2014. During those four flights, aerosol number concentrations and CCN concentrations at two supersaturations (0.25% and 0.5%) were measured by condensation particle counters (CPCs) and a DMT dual column CCN counter onboard both G-1 and HALO. Aerosol size distribution was also measured by a Fast Integrated Mobility Spectrometer (FIMS) aboard the G-1 and is compared with the size distribution from Ultra High Sensitivity Aerosol Spectrometer - Airborne (UHSAS-A, DMT), which were deployed both on the G-1 and the HALO. Good agreement between the aerosol properties measured from the two aircraft has been achieved. The vertical profiles of aerosol size distribution and CCN spectrum will be discussed.

  6. On the first G 1 stiff fluid spike solution in General Relativity

    NASA Astrophysics Data System (ADS)

    Coley, A. A.; Gregoris, D.; Lim, W. C.

    2016-11-01

    Using the Geroch transformation we obtain the first example of an exact stiff fluid spike solution to the Einstein field equations in a closed form exhibiting a spacelike G 1 group of symmetries (i.e., with a single isometry). This new solution is of Petrov type I and exhibits a spike crossing which persists to the past, which allows us to better understand spike crossings in the context of structure formation.

  7. Colostrogenesis: candidate genes for IgG1 transcytosis mechanisms in primary bovine mammary epithelial cells.

    PubMed

    Stark, A; Vachkova, E; Wellnitz, O; Bruckmaier, R; Baumrucker, C

    2013-12-01

    Bovine colostrogenesis is distinguished by the specific transfer of IgG1 from the blood to mammary secretions. The process has been shown to be initiated by hormones and occurs during the last weeks of pregnancy when steroid concentrations of estradiol (E2 ) and progesterone (P4 ) are highly elevated. Rodent intestinal uptake of immunoglobulin G is mediated by a receptor termed Fc fragment of IgG, Receptor, Transporter, alpha (FcGRT) and supported by light chain Beta-2-Microglobulin (β2M). We hypothesized that steroid hormone treatments (E2 and P4 ) of bovine mammary epithelial cells in vitro would induce up-regulation of IgG1 transcytosis candidate gene mRNA expression suggesting involvement in IgG1 transcytosis. Two different primary bovine mammary epithelial cell cultures were cultured on plastic and rat tail collagen and treated with hormonal combinations (steroids/lactogenic hormones). Evaluated mRNA components were bLactoferrin (bLf: a control), bFcGRT, β2M, and various small GTPases; the latter components are reported to direct endosomal movements in eukaryotic cells. All tested transcytosis components showed strong expression of mRNA in the cells. Expression of bFcGRT, bRab25 and bRhoB were significantly up-regulated (p < 0.05) by steroid hormones. bRab25 and bRhoB showed increased expression by steroid treatments, but also with lactogenic hormones. Analysis for the oestrogen receptor (ER) mRNA was mostly negative, but 25% of the cultures tested exhibited weak expression, while the progesterone receptor (PR) mRNA was always detected. bRab25 and bRhoB and likely bFcGRT are potential candidate genes for IgG1 transcytosis in bovine mammary cells. PMID:23279563

  8. Nonuniform Expansion of the Youngest Galactic Supernova Remnant G1.9+0.3

    NASA Astrophysics Data System (ADS)

    Borkowski, Kazimierz J.; Reynolds, Stephen P.; Green, David A.; Hwang, Una; Petre, Robert; Krishnamurthy, Kalyani; Willett, Rebecca

    2014-08-01

    We report measurements of the X-ray expansion of the youngest Galactic supernova remnant, G1.9+0.3, using Chandra observations in 2007, 2009, and 2011. The measured rates strongly deviate from uniform expansion, decreasing radially by about 60% along the X-ray bright SE-NW axis from 0.84% ± 0.06% yr-1 to 0.52% ± 0.03% yr-1. This corresponds to undecelerated ages of 120-190 yr, confirming the young age of G1.9+0.3 and implying a significant deceleration of the blast wave. The synchrotron-dominated X-ray emission brightens at a rate of 1.9% ± 0.4% yr-1. We identify bright outer and inner rims with the blast wave and reverse shock, respectively. Sharp density gradients in either the ejecta or ambient medium are required to produce the sudden deceleration of the reverse shock or the blast wave implied by the large spread in expansion ages. The blast wave could have been decelerated recently by an encounter with a modest density discontinuity in the ambient medium, such as may be found at a wind termination shock, requiring strong mass loss in the progenitor. Alternatively, the reverse shock might have encountered an order-of-magnitude density discontinuity within the ejecta, such as may be found in pulsating delayed-detonation Type Ia models. We demonstrate that the blast wave is much more decelerated than the reverse shock in these models for remnants at ages similar to G1.9+0.3. Similar effects may also be produced by dense shells possibly associated with high-velocity features in Type Ia spectra. Accounting for the asymmetry of G1.9+0.3 will require more realistic three-dimensional Type Ia models.

  9. NONUNIFORM EXPANSION OF THE YOUNGEST GALACTIC SUPERNOVA REMNANT G1.9+0.3

    SciTech Connect

    Borkowski, Kazimierz J.; Reynolds, Stephen P.; Green, David A.; Hwang, Una; Petre, Robert; Willett, Rebecca

    2014-08-01

    We report measurements of the X-ray expansion of the youngest Galactic supernova remnant, G1.9+0.3, using Chandra observations in 2007, 2009, and 2011. The measured rates strongly deviate from uniform expansion, decreasing radially by about 60% along the X-ray bright SE-NW axis from 0.84% ± 0.06% yr{sup –1} to 0.52% ± 0.03% yr{sup –1}. This corresponds to undecelerated ages of 120-190 yr, confirming the young age of G1.9+0.3 and implying a significant deceleration of the blast wave. The synchrotron-dominated X-ray emission brightens at a rate of 1.9% ± 0.4% yr{sup –1}. We identify bright outer and inner rims with the blast wave and reverse shock, respectively. Sharp density gradients in either the ejecta or ambient medium are required to produce the sudden deceleration of the reverse shock or the blast wave implied by the large spread in expansion ages. The blast wave could have been decelerated recently by an encounter with a modest density discontinuity in the ambient medium, such as may be found at a wind termination shock, requiring strong mass loss in the progenitor. Alternatively, the reverse shock might have encountered an order-of-magnitude density discontinuity within the ejecta, such as may be found in pulsating delayed-detonation Type Ia models. We demonstrate that the blast wave is much more decelerated than the reverse shock in these models for remnants at ages similar to G1.9+0.3. Similar effects may also be produced by dense shells possibly associated with high-velocity features in Type Ia spectra. Accounting for the asymmetry of G1.9+0.3 will require more realistic three-dimensional Type Ia models.

  10. A novel anticancer agent, decursin, induces G1 arrest and apoptosis in human prostate carcinoma cells.

    PubMed

    Yim, Dongsool; Singh, Rana P; Agarwal, Chapla; Lee, Sookyeon; Chi, Hyungjoon; Agarwal, Rajesh

    2005-02-01

    We isolated a coumarin compound decursin (C(19)H(20)O(5); molecular weight 328) from Korean angelica (Angelica gigas) root and characterized it by spectroscopy. Here, for the first time, we observed that decursin (25-100 micromol/L) treatment for 24 to 96 hours strongly inhibits growth and induces death in human prostate carcinoma DU145, PC-3, and LNCaP cells. Furthermore, we observed that decursinol [where (CH(3))(2)-C=CH-COO- side chain of decursin is substituted with -OH] has much lower effects compared with decursin, suggesting a possible structure-activity relationship. Decursin-induced growth inhibition was associated with a strong G(1) arrest (P < 0.001) in DU145 and LNCaP cells, and G(1), S as well as G(2)-M arrests depending upon doses and treatment times in PC-3 cells. Comparatively, decursin was nontoxic to human prostate epithelial PWR-1E cells and showed only moderate growth inhibition and G(1) arrest. Consistent with G(1) arrest in DU145 cells, decursin strongly increased protein levels of Cip1/p21 but showed a moderate increase in Kip1/p27 with a decrease in cyclin-dependent kinases (CDK); CDK2, CDK4, CDK6, and cyclin D1, and inhibited CDK2, CDK4, CDK6, cyclin D1, and cyclin E kinase activity, and increased binding of CDK inhibitor (CDKI) with CDK. Decursin-caused cell death was associated with an increase in apoptosis (P < 0.05-0.001) and cleaved caspase-9, caspase-3, and poly(ADP-ribose) polymerase; however, pretreatment with all-caspases inhibitor (z-VAD-fmk) only partially reversed decursin-induced apoptosis, suggesting the involvement of both caspase-dependent and caspase-independent pathways. These findings suggest the novel anticancer efficacy of decursin mediated via induction of cell cycle arrest and apoptosis selectively in human prostate carcinoma cells.

  11. MicroRNA-224 Induces G1/S Checkpoint Release in Liver Cancer

    PubMed Central

    An, Fangmei; Olaru, Alexandru V.; Mezey, Esteban; Xie, Qing; Li, Ling; Piontek, Klaus B.; Selaru, Florin M.

    2015-01-01

    Profound changes in microRNA (miR) expression levels are frequently found in liver cancers compared to the normal liver. In this study, we evaluate the expression of miR-224 in human HCC and CCA, as well as its downstream targets and affected pathways. We show that miR-224 is upregulated in a large cohort of human CCA, similar to its upregulation in human HCC. For the purpose of studying the roles of miR-224 in HCC and CCA, we enforced miR-224 expression in cells. mRNA arrays followed by Ingenuity Pathway Analysis (IPA)-identified putative molecules and pathways downstream of miR-224. Phenotypically, we report that enforced expression of miR-224 increases the growth rate of normal cholangiocytes, CCA cell lines, and HCC cell lines. In addition, we identified, in an unbiased fashion, that one of the major biologic processes affected by miR-224 is Gap1 (G1) to Synthesis (S) transition checkpoint release. We next identified p21, p15, and CCNE1 as downstream targets of miR-224 and confirmed the coordinated downregulation results in the increased phosphorylation of Retinoblastoma (Rb) with resulting G1/S checkpoint release. Our data suggest that miR-224 is a master regulator of cell cycle progression, and that its overexpression results in G1/S checkpoint release followed by accelerated cell growth. PMID:26343737

  12. Fate of aflatoxins B1 and G 1 residues during biscuit processing.

    PubMed

    Amra, H A; Mahmoud, S A; Taha, A H; El-Azab, M A

    1996-09-01

    Effect of biscuit processing on the destruction of aflatoxins B1 and G1 with and/or without some commonly leavening agents used namely sodium bicarbonate, ammonium bicarbonate and sodium bisulfite and sodium chloride.It was found that mixing step reduced the concentration of aflatoxins B1 and G1 by 80.7% and 82.7%, while the effect of baking step being 28.9% and 21.5%. The effect of mixing was found to be more pronounced than that baking step.The highest destruction effect on aflatoxin B1 was observed by adding a mixture composed of sodium and ammonium bicarbonate and sodium bisulfite followed by sodium chloride, sodium bisulfite, ammonium bicarbonate and/or sodium bicarbonate alone, where the reduction values of toxin after mixing were 93.4,91.9,91.7, 88.8 and 86.6% respectively, while the baking effect ranged 17.2 to 34.5% in the presence of different leaving agents added.Concerning aflatoxin G1; the highest destructive effect of toxin was adsorbed by adding a mixture of sodium and ammonium bicarbonate and sodium bisulfite followed by sodium bisulfite, sodium chloride, ammonium bicarbonate and/or sodium bicarbonate alone since the destruction values of such toxin after mixing were 96.2%, 92.8%, 92.6%, 89.0% and 87.7% respectively, while the baking effect ranged 20.9 to 34.5% in all leavening agents added.

  13. Tet1 is required for Rb phosphorylation during G1/S phase transition

    SciTech Connect

    Huang, Shengsong; Zhu, Ziqi; Wang, Yiqin; Wang, Yanru; Xu, Longxia; Chen, Xuemei; Xu, Qing; Zhang, Qimin; Zhao, Xin; Yu, Yi; Wu, Denglong

    2013-05-03

    Highlights: •Tet1 was required for NIT3T3 proliferation. •Tet1 depletion inhibited G1-S entry. •Cyclin D1 accumulation and Rb phosphorylation was blocked by Tet1 knockdown. -- Abstract: DNA methylation plays an important role in many biological processes, including regulation of gene expression, maintenance of chromatin conformation and genomic stability. TET-family proteins convert 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), which indicates that these enzymes may participate in DNA demethylation. The function of TET1 has not yet been well characterized in somatic cells. Here, we show that depletion of Tet1 in NIH3T3 cells inhibits cell growth. Furthermore, Tet1 knockdown blocks cyclin D1 accumulation in G1 phase, inhibits Rb phosphorylation and consequently delays entrance to G1/S phase. Taken together, this study demonstrates that Tet1 is required for cell proliferation and that this process is mediated through the Rb pathway.

  14. Dux4 induces cell cycle arrest at G1 phase through upregulation of p21 expression

    SciTech Connect

    Xu, Hongliang; Wang, Zhaoxia; Jin, Suqin; Hao, Hongjun; Zheng, Lemin; Zhou, Boda; Zhang, Wei; Lv, He; Yuan, Yun

    2014-03-28

    Highlights: • Dux4 induced TE671 cell proliferation defect and G1 phase arrest. • Dux4 upregulated p21 expression without activating p53. • Silencing p21 rescued Dux4 mediated proliferation defect and cell cycle arrest. • Sp1 binding site was required for Dux4-induced p21 promoter activation. - Abstract: It has been implicated that Dux4 plays crucial roles in development of facioscapulohumeral dystrophy. But the underlying myopathic mechanisms and related down-stream events of this retrogene were far from clear. Here, we reported that overexpression of Dux4 in a cell model TE671 reduced cell proliferation rate, and increased G1 phase accumulation. We also determined the impact of Dux4 on p53/p21 signal pathway, which controls the checkpoint in cell cycle progression. Overexpression of Dux4 increased p21 mRNA and protein level, while expression of p53, phospho-p53 remained unchanged. Silencing p21 rescued Dux4 mediated proliferation defect and cell cycle arrest. Furthermore, we demonstrated that enhanced Dux4 expression increased p21 promoter activity and elevated expression of Sp1 transcription factor. Mutation of Sp1 binding site decreased dux4 induced p21 promoter activation. Chromatin immunoprecipitation (ChIP) assays confirmed the Dux4-induced binding of Sp1 to p21 promoter in vivo. These results suggest that Dux4 might induce proliferation inhibition and G1 phase arrest through upregulation of p21.

  15. SUMOylation of Rb enhances its binding with CDK2 and phosphorylation at early G1 phase.

    PubMed

    Meng, Fengxi; Qian, Jiang; Yue, Han; Li, Xiaofeng; Xue, Kang

    2016-07-01

    Retinoblastoma protein (Rb) is a prototypical tumor suppressor that is vital to the negative regulation of the cell cycle and tumor progression. Hypo-phosphorylated Rb is associated with G0/G1 arrest by suppressing E2F transcription factor activity, whereas Rb hyper-phosphorylation allows E2F release and cell cycle progression from G0/G1 to S phase. However, the factors that regulate cyclin-dependent protein kinase (CDK)-dependent hyper-phosphorylation of Rb during the cell cycle remain obscure. In this study, we show that throughout the cell cycle, Rb is specifically small ubiquitin-like modifier (SUMO)ylated at early G1 phase. SUMOylation of Rb stimulates its phosphorylation level by recruiting a SUMO-interaction motif (SIM)-containing kinase CDK2, leading to Rb hyper-phosphorylation and E2F-1 release. In contrast, a SUMO-deficient Rb mutant results in reduced SUMOylation and phosphorylation, weakened CDK2 binding, and attenuated E2F-1 sequestration. Furthermore, we reveal that Rb SUMOylation is required for cell proliferation. Therefore, our study describes a novel mechanism that regulates Rb phosphorylation during cell cycle progression. PMID:27163259

  16. Optimized culture condition for enhancing lytic performance of waste activated sludge by Geobacillus sp. G1.

    PubMed

    Yang, Chunxue; Zhou, Aijuan; Hou, Yanan; Zhang, Xu; Guo, Zechong; Wang, Aijie; Liu, Wenzong

    2014-01-01

    Hydrolysis is known as the rate-limiting step during waste activated sludge (WAS) digestion. The optimization of the culture conditions of Geobacillus sp. G1 for enhancing WAS hydrolysis was conducted in this study with uniform design and response surface methodology. Taking the lysis rate of Escherichia coli as the response, the Plackett-Burman design was used to screen the most important variables. Experimental results showed that the maximum predicted lysis rate of E. coli was 50.9% for 4 h treatment time with concentrations of skim milk, NaCl and NH4SO4 at 10.78, 4.36 and 11.28 g/L, respectively. The optimized dosage ratio of Geobacillus sp. G1 to WAS was 35%:65% (VG1:VWAS). Under this condition, soluble protein was increased to 695 mg chemical oxygen demand (COD)/L, which was 5.0 times higher than that obtained in the control (140 mg COD/L). The corresponding protease activity reached 1.1 Eu/mL. Scanning electron microscopy showed that abundant cells were apparently lysed with treatment of Geobacillus sp. G1.

  17. SUMOylation of Rb enhances its binding with CDK2 and phosphorylation at early G1 phase

    PubMed Central

    Meng, Fengxi; Qian, Jiang; Yue, Han; Li, Xiaofeng; Xue, Kang

    2016-01-01

    ABSTRACT Retinoblastoma protein (Rb) is a prototypical tumor suppressor that is vital to the negative regulation of the cell cycle and tumor progression. Hypo-phosphorylated Rb is associated with G0/G1 arrest by suppressing E2F transcription factor activity, whereas Rb hyper-phosphorylation allows E2F release and cell cycle progression from G0/G1 to S phase. However, the factors that regulate cyclin-dependent protein kinase (CDK)-dependent hyper-phosphorylation of Rb during the cell cycle remain obscure. In this study, we show that throughout the cell cycle, Rb is specifically small ubiquitin-like modifier (SUMO)ylated at early G1 phase. SUMOylation of Rb stimulates its phosphorylation level by recruiting a SUMO-interaction motif (SIM)-containing kinase CDK2, leading to Rb hyper-phosphorylation and E2F-1 release. In contrast, a SUMO-deficient Rb mutant results in reduced SUMOylation and phosphorylation, weakened CDK2 binding, and attenuated E2F-1 sequestration. Furthermore, we reveal that Rb SUMOylation is required for cell proliferation. Therefore, our study describes a novel mechanism that regulates Rb phosphorylation during cell cycle progression. PMID:27163259

  18. G1/S Cell Cycle Checkpoint Defect in Lymphocytes from Patients with Alzheimer's Disease

    PubMed Central

    Song, Misun; Kwon, Young-Ah; Lee, Yujin; Kim, Hyeran; Yun, Ji Hea; Kim, Seonwoo

    2012-01-01

    Objective We compared the cell responsiveness of activated lymphocytes to rapamycin, which blocks the G1/S transition, between patients with Alzheimer's disease (AD) and normal controls to assess the early phase control defect in cell cycle. Methods Blood samples of 26 patients with AD and 28 normal controls were collected to separate peripheral lymphocytes. We measured the proportion of each cell cycle phase in activated lymphocytes using flow cytometry and evaluated the responsiveness of these lymphocytes to rapamycin. Results The patients with AD were older than the normal controls (AD 74.03±7.90 yr vs. control 68.28±6.21 yr, p=0.004). The proportion of G1 phase cells in the AD group was significantly lower than that in the control group (70.29±6.32% vs. 76.03±9.05%, p=0.01), and the proportion of S phase cells in the AD group was higher than that in control group (12.45±6.09% vs. 6.03±5.11%, p=0.001). Activated lymphocytes in patients with AD were not arrested in the G1 phase and they progressed to the late phase of the cell cycle despite rapamycin treatment, in contrast to those of normal subjects. Conclusion The patients with AD probably have a control defect of early phase cell cycle in peripheral lymphocytes that may be associated with the underlying pathology of neuronal death. PMID:23251208

  19. The human G1m1 allotype associates with CD4+ T-cell responsiveness to a highly conserved IgG1 constant region peptide and confers an asparaginyl endopeptidase cleavage site

    PubMed Central

    Stickler, M M; Reddy, A; Xiong, J M; Hinton, P R; DuBridge, R; Harding, F A

    2011-01-01

    The human G1m1 allotype comprises two amino acids, D12 and L14, in the CH3 domain of IGHG1. Although the G1m1 allotype is prevalent in human populations, ∼40% of Caucasiods are homozygous for the nG1m1 allotype corresponding to E12 and M14. Peptides derived from the G1m1 region were tested for their ability to induce CD4+ T-cell proliferative responses in vitro. A peptide immediately downstream from the G1m1 sequence was recognized by CD4+ T cells in a large percentage of donors (peptide CH315−29). CD4+ T-cell proliferative responses to CH315−29 were found at an increased frequency in nG1m1 homozygous donors. Homozygous nG1m1 donors possessing the HLA-DRB1*07 allele displayed the highest magnitudes of proliferation. CD4+ T cells from donors homozygous for nG1m1 proliferated to G1m1-carrying Fc-fragment proteins, whereas CD4+ T cells from G1m1 homozygous donors did not. The G1m1 sequence creates an enzymatic cleavage site for asparaginyl endopeptidase in vitro. Proteolytic activity at D12 may allow the presentation of the CH315−29 peptide, which in turn may result in the establishment of tolerance to this peptide in G1m1-positive donors. Homozygous nG1m1 patients may be more likely to develop CD4+ T-cell-mediated immune responses to therapeutic antibodies carrying the G1m1 allotype. PMID:21326320

  20. Characterization of recombinant β-fructofuranosidase from Bifidobacterium adolescentis G1

    PubMed Central

    2010-01-01

    Background We have previously reported on purification and characterization of β-fructofuranosidase (β-FFase) from Bifidobacterium adolescentis G1. This enzyme showed high activity of hydrolysis on fructo-oligosaccharides with a low degree of polymerization. Recently, genome sequences of B. longum NCC2705 and B. adolescentis ATCC 15703 were determined, and cscA gene in the both genome sequences encoding β-FFase was predicted. Here, cloning of cscA gene encoding putative β-FFase from B. adolescentis G1, its expression in E. coli and properties of the recombinant protein are described. Results Using the information of cscA gene from Bifidobacterium adolescentis ATCC 15703, cscA gene from B. adolescentis G1 was cloned and sequenced. The N-terminal amino acid sequence of purified β-FFase from B. adolescentis G1 was identical to the deduced amino acid sequences of cscA gene from B. adolescentis G1. To confirm the translated product of the cscA gene, the recombinant protein was expressed in Escherichia coli. Molecular mass of the purified recombinant enzyme was estimated to be about 66,000 by SDS-PAGE and 60,300 by MALDI TOF-MS. The optimum pH of the enzyme was 5.7 and the enzyme was stable at pH 5.0-8.6. The thermostability of the enzyme was up to 50°C. The Km (mM), Vmax (μmol/mg of protein/min), k0 (sec-1) and k0/Km(mM-1 sec-1) for 1-kestose, neokestose, nystose, fructosylnystose, sucrose and inulin were 1.7, 107, 107.5, 63.2, and 1.7, 142, 142.7, 83.9, and 3.9, 152, 152.8, 39.2, and 2.2, 75, 75.4, 34.3, and 38, 79, 79.4, 2.1, and 25.9, 77, 77.4, 3.0, respectively. The hydrolytic activity was strongly inhibited by AgNO3, SDS, and HgCl2. Conclusion The recombinant enzyme had similar specificity to the native enzyme, high affinity for 1-kestose, and low affinity for sucrose and inulin, although properties of the recombinant enzyme showed slight difference from those of the native one previously described. PMID:20380746

  1. Stabilization of IgG1 in spray-dried powders for inhalation.

    PubMed

    Schüle, S; Schulz-Fademrecht, T; Garidel, P; Bechtold-Peters, K; Frieb, W

    2008-08-01

    The protein stabilizing capabilities of spray-dried IgG1/mannitol formulations were evaluated. The storage stability was tested at different residual moisture levels prepared by vacuum-drying or equilibration prior to storage. Vacuum-drying at 32 degrees C/0.1mbar for 24h reduced the moisture level below 1%, constituting an optimal basis for improved storage stability. The crystalline IgG1/mannitol powders with a weight ratio of 20/80 up to 40/60 failed to prevent the antibody aggregation as assessed by size exclusion chromatography during storage. Ratios of 60/40 up to 80/20 IgG1/mannitol provided superior stability of the antibody and the powders could be produced with high yields. The lower the residual moisture, the better was the stabilizing capability. An amount of 20% mannitol provided the best stabilization. Storage stability of 60/40, 70/30, and 80/20 IgG1/mannitol formulations over one year was adequate at 2-8 degrees C and 25 degrees C. Closed storage (sealed in vials) at 40 degrees C/75% RH and open storage at 25 degrees C/60% RH revealed that the stability still required optimization. The lower the protein content, the better was the powder flowability. The aerodynamic properties of powders spray-dried with 10% solids content were inadequate, as the particle size ranged between 5.1 and 7.2 microm and the fine particle fraction accounted for only 4-11%. Reduction of the solids content to 2.5% did improve the aerodynamic properties as the mass mean aerodynamic diameter was reduced to 3.6 microm and the fine particle fraction was increased to about 14%. The reduction of the solids content did not influence the storage stability significantly. Also spray-drying at higher temperatures had no significant impact on the storage stability, despite a higher tendency to form amorphous systems. In order to improve the storage stability and to maintain the good flowability of 70/30 IgG1/mannitol powder or to keep the storage stability but to improve the flowability

  2. Contribution of variable domains to the stability of humanized IgG1 monoclonal antibodies.

    PubMed

    Ionescu, Roxana M; Vlasak, Josef; Price, Colleen; Kirchmeier, Marc

    2008-04-01

    Temperature-induced unfolding of three humanized IgG1 monoclonal antibodies and their Fab and Fc fragments was monitored by differential scanning calorimetry at neutral pH. With some exceptions, the thermogram of the intact antibody presents two peaks and the transition with the larger experimental enthalpy contains the contribution from the Fab fragments. Although the measured enthalpy was similar for all three Fab fragments studied, the apparent melting temperatures were found to vary significantly, even for Fab fragments originating from the same human germline. Therefore, we propose to use the measured enthalpy of unfolding as the key parameter to recognize the unfolding events in the melting profile of an intact IgG1 antibody. If the variable domain sequences, resulting from complementarity determining regions (CDRs) grafting and humanization, destabilize the Fab fragment with respect to the CH3 domain, the first transition represents the unfolding of the Fab fragment and the CH2 domain, while the second transition represents CH3 domain unfolding. Otherwise, the first transition represents CH2 domain unfolding, and the second transition represents the unfolding of the Fab fragment and the CH3 domain. In some cases, the DSC profile may present three transitions, with the Fab unfolding occurring at distinct temperatures compared to the melting of the CH2 and CH3 domains. If the DSC profile of a humanized IgG1 monoclonal antibody cannot be described by the model above, the result may be an indication of significant structural heterogeneity and/or of disruption of the Fab cooperative unfolding. Low stability or heterogeneity of the Fab fragment may prove problematic for long-term storage or consistency of production. Therefore, understanding the features of a DSC profile is important for clone selection and process maturation in the early stages of development of therapeutic monoclonal antibodies.

  3. JAZ mediates G1 cell cycle arrest by interacting with and inhibiting E2F1

    PubMed Central

    Yang, Mingli; Wu, Song; Jia, Jinghua

    2011-01-01

    We discovered and reported JAZ as a unique dsRNA binding zinc finger protein that functions as a direct, positive regulator of p53 transcriptional activity to mediate G1 cell cycle arrest in a mechanism involving upregulation of the p53 target gene, p21. We now find that JAZ can also negatively regulate the cell cycle in a novel, p53-independent mechanism resulting from the direct interaction with E2F1, a key intermediate in regulating cell proliferation and tumor suppression. JAZ associates with E2F1's central DNA binding/dimerization region and its C-terminal transactivation domain. Functionally, JAZ represses E2F1 transcriptional activity in association with repression of cyclin A expression and inhibition of G1/S transition. This mechanism involves JAZ-mediated inhibition of E2F1's specific DNA binding activity. JAZ directly binds E2F1 in vitro in a dsRNA-independent manner, and JAZ's dsRNA binding ZF domains, which are necessary for localizing JAZ to the nucleus, are required for repression of transcriptional activity in vivo. Importantly for specificity, siRNA-mediated “knockdown” of endogenous JAZ increases E2F transcriptional activity and releases cells from G1 arrest, indicating a necessary role for JAZ in this transition. Although JAZ can directly inhibit E2F1 activity independently of p53, if functional p53 is expressed, JAZ may exert a more potent inhibition of cell cycle following growth factor withdrawal. Therefore, JAZ plays a dual role in cell cycle regulation by both repressing E2F1 transcriptional activity and activating p53 to facilitate efficient growth arrest in response to cellular stress, which may potentially be exploited therapeutically for tumor growth inhibition. PMID:21715977

  4. Selenium Preconcentration in Geological Materials for Determination at sub-μ g\\ g-1 Concentration

    NASA Astrophysics Data System (ADS)

    Bedard, L. P.

    2004-05-01

    Selenium is important because it is a path finder element in economic geology. It has similar geochemical properties as sulfur, but slightly less mobile and less volatile in sulfides. Although its environmental cycle is better understood, its geological cycle is almost unknown. In geological samples, Se concentration ranges 0,02-1 μ g\\ g-1. Se has many spectral interefences in ICP-MS, rendering difficult to determine. INAA detection limit for geological samples is about 10 μ g\\ g-1. The analytical difficulties are one of the main reason why the geological cycle of Se is so poorly known. The preconcentration of Se with Thiol Cotton Fiber (TCF) followed by atomic absorption (AA) has been modified to be used with INAA. The modified technique involves sample dissolution (HF-HNO3) and evaporation to dryness at low temperature (55-60 oC) to keep Se in solution. Se is converted to SeIV by adding 5-6 mol l-1 HCl and heating covered in a boiling bath (95-100oC). Sample is diluted with deionized water to obtain 0,3 - 1 mol l-1 HCl and then collected on TCF. TCF is put in a polyethylene vial for irradiation in the SLOWPOKE II reactor for 10 seconds at a neutron flux of 1015 m-2 s-1. The 162 KeV peak of 77Se (half-life 17,36 sec) is read for 20 seconds after a decay of 7 seconds. Results for certified reference materials show the TCF preconcentration technique followed by INAA provides results comparable with AA with a detection limit of approximately 0,05 μ g\\ g-1. Moreover INAA provides many advantages such as eliminating the desorption step and is less time consuming than AA.

  5. The G1 restriction point as critical regulator of neocortical neuronogenesis

    NASA Technical Reports Server (NTRS)

    Caviness, V. S. Jr; Takahashi, T.; Nowakowski, R. S.

    1999-01-01

    Neuronogenesis in the pseudostratified ventricular epithelium is the initial process in a succession of histogenetic events which give rise to the laminate neocortex. Here we review experimental findings in mouse which support the thesis that the restriction point of the G1 phase of the cell cycle is the critical point of regulation of the overall neuronogenetic process. The neuronogenetic interval in mouse spans 6 days. In the course of these 6 days the founder population and its progeny execute 11 cell cycles. With each successive cycle there is an increase in the fraction of postmitotic cells which leaves the cycle (the Q fraction) and also an increase in the length of the cell cycle due to an increase in the length of the G1 phase of the cycle. Q corresponds to the probability that postmitotic cells will exit the cycle at the restriction point of the G1 phase of the cell cycle. Q increases non-linearly, but the rate of change of Q with cycle (i.e., the first derivative) over the course of the neuronogenetic interval is a constant, k, which appears to be set principally by cell internal mechanisms which are species specific. Q also seems to be modulated, but at low amplitude, by a balance of mitogenic and antimitogenic influences acting from without the cell. We suggest that intracellular signal transduction systems control a general advance of Q during development and thereby determine the general developmental plan (i.e., cell number and laminar composition) of the neocortex and that external mitogens and anti-mitogens modulate this advance regionally and temporally and thereby produce regional modifications of the general plan.

  6. Monoclonal Immunoglobulin G1 Directed against Aspergillus fumigatus Cell Wall Glycoprotein Protects against Experimental Murine Aspergillosis†

    PubMed Central

    Chaturvedi, Ashok K.; Kavishwar, A.; Keshava, G. B. Shiva; Shukla, P. K.

    2005-01-01

    Most of the biological functions related to pathogenicity and virulence reside in the fungal cell wall, which, being the outermost part of the cell, mediates the host-fungus interplay. For these reasons much effort has focused on the discovery of useful inhibitors of cell wall glucan, chitin, and mannoprotein biosynthesis. In the absence of a wide-spectrum, safe, and potent antifungal agent, a new strategy for antifungal therapy is directed towards the development of monoclonal antibodies (MAbs). In the present study the MAb A9 (immunoglobulin G1 [IgG1]) was identified from hybridomas raised in BALB/c mice immunized with cell wall antigen of Aspergillus fumigatus. The immunoreactive epitopes for this IgG1 MAb appeared to be associated with a peptide moiety, and indirect immunofluorescence microscopy revealed its binding to the cell wall surface of hyphae as well as with swollen conidia. MAb A9 inhibited hyphal development as observed by MTT [3-(4,5-dimethythiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay (25.76%), reduced the duration of spore germination, and exerted an in vitro cidal effect against Aspergillus fumigatus. The in vivo protective efficacy of MAb A9 was also evaluated in a murine model of invasive aspergillosis, where a reduction in CFU (>4 log10 units) was observed in kidney tissue of BALB/c mice challenged with A. fumigatus (2 × 105 CFU/ml) and where enhanced mean survival times (19.5 days) compared to the control (7.1 days) and an irrelevant MAb (6.1 days) were also observed. PMID:16148172

  7. Phospholipid hydroperoxide glutathione peroxidase induces a delay in G1 of the cell cycle.

    PubMed

    Wang, Hong P; Schafer, Freya Q; Goswami, Prabhat C; Oberley, Larry W; Buettner, Garry R

    2003-06-01

    Phospholipid hydroperoxide glutathione peroxidase (PhGPx) is an antioxidant enzyme that reduces cellular phospholipid hydroperoxides (PLOOHs) to alcohols. Cellular peroxide tone has been implicated in cell growth and differentiation. By reducing the PLOOH level in the cell membrane, PhGPx regulates the peroxide tone and thereby might be involved in cell growth. We hypothesized that overexpression of PhGPx in human breast cancer cells would decrease their growth rate. We stably transfected MCF-7 cells (Wt) with L-PhGPx and measured cell doubling time, plating efficiency, and cell cycle phase transit times. P-4 cells (8-fold increase in PhGPx activity) showed a 2-fold increase in doubling time; doubling time increased directly with PhGPx activity (r = 0.95). The higher the PhGPx activity, the lower the plating efficiency (r = -0.86). The profile of other antioxidant enzymes was unchanged. Overexpression of PhGPx lowered the steady-state level of PLOOH (by > 60%). Results from bromodeoxyuridine pulse-chase experiments and flow cytometry indicate that PhGPx induced a delay in MCF-7 proliferation that was primarily due to a slower progression from G1 to S. These results support the hypothesis that PhGPx plays a regulatory role in the progression of MCF-7 cells from G1 to S possibly by regulating the steady-state levels of PLOOH. These data suggest that PhGPx can lower the peroxide tone, which might change the cellular redox environment resulting in a delay in G1 transit. Thus, PhGPx could be an important factor in cell growth. PMID:12868489

  8. AAF G1 and Millimeter Wavelength ARM Radar Observations and Analysis from the ACAPEX Field Campaign

    NASA Astrophysics Data System (ADS)

    Matthews, A. A.; Hardin, J. C.; Comstock, J. M.; Bharadwaj, N.; Mei, F.

    2015-12-01

    From late January to early March of 2015, the ARM Cloud Aerosol Precipitation Experiment (ACAPEX) deployed a variety of instruments to study atmospheric rivers as they made landfall on the western United States. These atmospheric rivers are an important source of rainfall for the drought-ridden regions of California, so it is important to understand the cloud characteristics and processes that occur during the events. For this study, the KAZR zenith pointing millimeter wavelength radar aboard the Ron Brown research vessel as well as two cloud probes, namely the Two-Dimensional Stereo Probe (2DS) and High Volume Precipitation Spectrometer-3 (HVPS3), flown on the ARM Aerial Facility Gulfstream-1 (AAF G1) aircraft were used to analyze the clouds associated with the atmospheric rivers. PyDisdrometer, an open-source software, is used to calculate radar reflectivity values based on the drop size distributions observed by the cloud probes so that the aircraft data may be directly compared to the KAZR data. This research will focus on two flight days. First, a coordinated flight flown on February 5, where the G1 flew a spiral pattern over the Ron Brown while the KAZR was running will be used to examine the clouds over the ocean. This will also allow for a direct comparison between the calculated radar reflectivity from the cloud probes and the KAZR measured reflectivity. The second case was chosen to be the atmospheric river landfall event on February 6. This case will be analyzed over the ocean using the KAZR, then over the shoreline, Sacramento, and the inland mountains using the cloud probe data, as well as other instrumentations onboard the G1, such as those that measure cloud thickness and liquid water content. In this way, the changes that occur to the clouds as they move from open-ocean to land may be examined in greater detail.

  9. Some alkali and titania analyses of tektites before and after G-1 precision monitoring

    USGS Publications Warehouse

    Tatlock, D.B.

    1966-01-01

    A comparison of 55 older analyses of Australasian tektites with 110 modern precisely monitored analyses suggests that more than half of the older alkali and titania determinations are decidedly inaccurate and misleading. Deviations of the older analyses from the restricted values of the modern analyses are comparable to the imprecisions shown by early analyses of G-1 granite and W-1 diabase. This suggests that a high percentage of older alkali and titania analyses, such as those of Washington's tables, are of questionable quality. ?? 1966.

  10. THE ABSENCE OF RADIO EMISSION FROM THE GLOBULAR CLUSTER G1

    SciTech Connect

    Miller-Jones, J. C. A.; Wrobel, J. M.; Sivakoff, G. R.; Heinke, C. O.; Miller, R. E.; Plotkin, R. M.; Di Stefano, R.; Greene, J. E.; Ho, L. C.; Joseph, T. D.; Maccarone, T. J.; Kong, A. K. H.

    2012-08-10

    The detections of both X-ray and radio emission from the cluster G1 in M31 have provided strong support for existing dynamical evidence for an intermediate-mass black hole (IMBH) of mass (1.8 {+-} 0.5) Multiplication-Sign 10{sup 4} M{sub Sun} at the cluster center. However, given the relatively low significance and astrometric accuracy of the radio detection, and the non-simultaneity of the X-ray and radio measurements, this identification required further confirmation. Here we present deep, high angular resolution, strictly simultaneous X-ray and radio observations of G1. While the X-ray emission (L{sub X} = 1.74{sup +0.53}{sub -0.44} Multiplication-Sign 10{sup 36} (d/750 kpc){sup 2} erg s{sup -1} in the 0.5-10 keV band) remained fully consistent with previous observations, we detected no radio emission from the cluster center down to a 3{sigma} upper limit of 4.7 {mu}Jy beam{sup -1}. Our favored explanation for the previous radio detection is flaring activity from a black hole low-mass X-ray binary (LMXB). We performed a new regression of the 'Fundamental Plane' of black hole activity, valid for determining black hole mass from radio and X-ray observations of sub-Eddington black holes, finding log M{sub BH} = (1.638 {+-} 0.070)log L{sub R} - (1.136 {+-} 0.077)log L{sub X} - (6.863 {+-} 0.790), with an empirically determined uncertainty of 0.44 dex. This constrains the mass of the X-ray source in G1, if a black hole, to be <9.7 Multiplication-Sign 10{sup 3} M{sub Sun} at 95% confidence, suggesting that it is a persistent LMXB. This annuls what was previously the most convincing evidence from radiation for an IMBH in the Local Group, though the evidence for an IMBH in G1 from velocity dispersion measurements remains unaffected by these results.

  11. Asymmetric expansion of the youngest Galactic supernova remnant G1.9+0.3

    NASA Astrophysics Data System (ADS)

    Reynolds, Stephen P.

    2016-06-01

    The youngest Galactic supernova remnant (SNR) G1.9+0.3, produced by a (probable) Type Ia SN that exploded around CE 1900, is strongly asymmetric at radio wavelengths, with a single bright maximum in its shell, but exhibits a bilaterally symmetric morphology in X-rays. It has been difficult to understand the origin of these contrasting morphologies. We present the results of expansion measurements of G1.9+0.3 that illuminate the origin of the radio asymmetry. These measurements are based on a comparison of our 2015 400-ks Chandra observation with earlier Chandra observations, including a 1-Ms observation in 2011. The mean expansion rate from 2011 to 2015 is 0.58% per yr, in agreement with previous measurements. We also confirm that the expansion decreases radially away from the remnant's center along the major E-W axis, from 0.77% per yr to 0.53% per yr. Large variations in expansion are also present along the minor N-S axis, but expansion there is strongly asymmetric and varies on small spatial scales. We use the “Demons” method to study the complex motions within G1.9+0.3. This method provides a nonparametric way for measuring these motions globally. We find motions varying by a factor of 5, from 0.09" to 0.44" per year. The slowest shocks are in the north, at the outer boundary of the bright radio emission, with speeds there as low as 3,600 km/s (for an assumed distance of 8.5 kpc), much less than the average shock speed of 12,000 km/s. Such strong deceleration of the northern blast wave most likely arises from the collision of SN ejecta with a much denser than average ambient medium there. The presence of this asymmetric ambient medium naturally explains the radio asymmetry. The SN ejecta have also been strongly decelerated in the N, but they expand faster than the blast wave. In several locations, significant morphological changes and strongly nonradial motions are apparent. The spatially-integrated X-ray flux continues to increase with time. As with Kepler

  12. Nonuniform Expansion of the Youngest Galactic Supernova Remnant G1.9+0.3

    NASA Astrophysics Data System (ADS)

    Reynolds, Stephen P.; Borkowski, Kazimierz J.; Green, David; Hwang, Una; Petre, Robert

    2014-08-01

    G1.9+0.3 is the youngest known Galactic supernova remnant (SNR), about 100 yr old from global expansion measurements, and most likely the result of an asymmetric Type Ia supernova explosion. We smoothed a Chandra image from a 1 Ms observation in 2011 and fit the resulting model to unsmoothed images from 2007 and 2009, allowing for expansion and image shifts. The measured expansion rates strongly deviate from uniform expansion, increasing inward by about 60% along the X-ray bright SE-NW axis, from 0.52% +- 0.03% per yr to 0.84% +- 0.06% per yr. This corresponds to undecelerated ages of 120 - 190 yr, confirming the young age of G1.9 +0.3, and implying a significant (deceleration parameter m < 0.6) deceleration of the blast wave. The spatially-integrated X-ray flux, strongly dominated by synchrotron emission, increases at a rate of 1.9% +- 0.7% per year, in agreement with previous measurements. G1.9+0.3 is the only Galactic SNR brightening at X-ray and radio wavelengths. We identify the inner rims with the reverse shock and more slowly-expanding rims farther out with the blast wave. The large spread in expansion ages between the reverse shock and the blast wave requires abrupt density gradients in either the ejecta or the ambient medium, to suddenly decelerate the reverse shock or the blast wave. The blast wave could have been decelerated recently by an encounter with a modest (factor of several) density discontinuity in the ambient medium, such as found at a wind termination shock, implying a strong presupernova wind from the progenitor system. Alternatively, the reverse shock might have encountered a larger (factor of 10 or more) density discontinuity within the SN ejecta, such as found in pulsating delayed-detonation Type Ia SN models. Through 1D hydrodynamical simulations, we demonstrate that the blast wave is much more decelerated than the reverse shock in these models for remnants at ages similar to G1.9+0.3. The presence of strong density gradients in the outer

  13. Overexpression of c-Myc alters G(1)/S arrest following ionizing radiation.

    PubMed

    Sheen, Joon-Ho; Dickson, Robert B

    2002-03-01

    Study of the mechanism(s) of genomic instability induced by the c-myc proto-oncogene has the potential to shed new light on its well-known oncogenic activity. However, an underlying mechanism(s) for this phenotype is largely unknown. In the present study, we investigated the effects of c-Myc overexpression on the DNA damage-induced G(1)/S checkpoint, in order to obtain mechanistic insights into how deregulated c-Myc destabilizes the cellular genome. The DNA damage-induced checkpoints are among the primary safeguard mechanisms for genomic stability, and alterations of cell cycle checkpoints are known to be crucial for certain types of genomic instability, such as gene amplification. The effects of c-Myc overexpression were studied in human mammary epithelial cells (HMEC) as one approach to understanding the c-Myc-induced genomic instability in the context of mammary tumorigenesis. Initially, flow-cytometric analyses were used with two c-Myc-overexpressing, nontransformed immortal lines (184A1N4 and MCF10A) to determine whether c-Myc overexpression leads to alteration of cell cycle arrest following ionizing radiation (IR). Inappropriate entry into S phase was then confirmed with a bromodeoxyuridine incorporation assay measuring de novo DNA synthesis following IR. Direct involvement of c-Myc overexpression in alteration of the G(1)/S checkpoint was then confirmed by utilizing the MycER construct, a regulatable c-Myc. A transient excess of c-Myc activity, provided by the activated MycER, was similarly able to induce the inappropriate de novo DNA synthesis following IR. Significantly, the transient expression of full-length c-Myc in normal mortal HMECs also facilitated entry into S phase and the inappropriate de novo DNA synthesis following IR. Furthermore, irradiated, c-Myc-infected, normal HMECs developed a sub-G(1) population and a >4N population of cells. The c-Myc-induced alteration of the G(1)/S checkpoint was also compared to the effects of expression of MycS (N

  14. Micro-solid phase extraction with liquid chromatography-tandem mass spectrometry for the determination of aflatoxins in coffee and malt beverage.

    PubMed

    Khayoon, Wejdan Shakir; Saad, Bahruddin; Salleh, Baharuddin; Manaf, Normaliza Hj Abdul; Latiff, Aishah A

    2014-03-15

    A single step extraction-cleanup procedure using porous membrane-protected micro-solid phase extraction (μ-SPE) in conjunction with liquid chromatography-tandem mass spectrometry for the extraction and determination of aflatoxins (AFs) B1, B2, G1 and G2 from food was successfully developed. After the extraction, AFs were desorbed from the μ-SPE device by ultrasonication using acetonitrile. The optimum extraction conditions were: sorbent material, C8; sorbent mass, 20mg; extraction time, 90 min; stirring speed, 1,000 rpm; sample volume, 10 mL; desorption solvent, acetonitrile; solvent volume, 350 μL and ultrasonication period, 25 min without salt addition. Under the optimum conditions, enrichment factor of 11, 9, 9 and 10 for AFG2, AFG1, AFB2 and AFB1, respectively were achieved. Good linearity and correlation coefficient was obtained over the concentration range of 0.4-50 ng g(-1) (r(2) 0.9988-0.9999). Good recoveries for AFs ranging from 86.0-109% were obtained. The method was applied to 40 samples involving malt beverage (19) and canned coffee (21). No AFs were detected in the selected samples.

  15. Nonuniform Expansion of the Youngest Galactic Supernova Remnant G1.9+0.3

    NASA Technical Reports Server (NTRS)

    Borkowski, Kazimierz J.; Reynolds, Stephen P.; Green, David A.; Hwang, Una; Petre, Robert; Krishnamurthy, Kalyani; Willett, Rebecca

    2014-01-01

    We report measurements of the X-ray expansion of the youngest Galactic supernova remnant, G1.9+0.3, using Chandra observations in 2007, 2009, and 2011. The measured rates strongly deviate from uniform expansion, decreasing radially by about 60 along the X-ray bright SE-NW axis from 0.84 plus or minus 0.06% yr(exp -1) to 0.52% plus or minus 0.03 yr(exp -1). This corresponds to undecelerated ages of 120-190 yr, confirming the young age of G1.9+0.3 and implying a significant deceleration of the blast wave. The synchrotron-dominated X-ray emission brightens at a rate of 1.9% plus or minus 0.4% yr(exp -1). We identify bright outer and inner rims with the blast wave and reverse shock, respectively. Sharp density gradients in either the ejecta or ambient medium are required to produce the sudden deceleration of the reverse shock or the blast wave implied by the large spread in expansion ages. The blast wave could have been decelerated recently by an encounter with a modest density discontinuity in the ambient medium, such as may be found at a wind termination shock, requiring strong mass loss in the progenitor.

  16. Berberine inhibits growth and induces G1 arrest and apoptosis in human cholangiocarcinoma QBC939 cells.

    PubMed

    He, Wei; Wang, Bin; Zhuang, Yun; Shao, Dong; Sun, Kewen; Chen, Jianping

    2012-01-01

    The chemotherapeutic approach using non-toxic natural products may be one of the strategies for the management of the cholangiocarcinoma. Here we report that in vitro treatment of human cholangiocarcinoma QBC939 cells with berberine, a naturally occurring isoquinoline alkaloid, decreased cell viability and induced cell death in a dose-dependent manner, which was associated with an increase in G1 arrest. Our western blot analysis showed that berberine-induced G1 cell cycle arrest was mediated through the increased expression of cyclin-dependent kinase inhibitors (Cdki) proteins (Cip1/p21 and Kip1/p27); a simultaneous decrease in Cdk2 and Cdk4 and cyclins D1, and reduced activity of the Cyclins-Cdk complex. In additional studies, treatment of QBC939 cells with different concentrations (10, 40, 80 μM) of berberine for 48 h resulted in a significant dose-dependent increase in apoptosis compared to the non-berberine-treated control, which was associated with an increased expression of pro-apoptotic protein Bax and decreased expression of anti-apoptotic proteins Bcl-2 and Bcl-xL. Together, this study for the first time identified berberine as a chemotherapeutic agent against human cholangiocarcinoma cells QBC939 cells in vitro. Further in vivo studies are required to determine whether berberine could be an effective chemotherapeutic agent for the management of cholangiocarcinoma.

  17. Magnetic Shielding Accelerates the Proliferation of Human Neuroblastoma Cell by Promoting G1-Phase Progression

    PubMed Central

    Liu, Ying; Bartlett, Perry F.; He, Rong-qiao

    2013-01-01

    Organisms have been exposed to the geomagnetic field (GMF) throughout evolutionary history. Exposure to the hypomagnetic field (HMF) by deep magnetic shielding has recently been suggested to have a negative effect on the structure and function of the central nervous system, particularly during early development. Although changes in cell growth and differentiation have been observed in the HMF, the effects of the HMF on cell cycle progression still remain unclear. Here we show that continuous HMF exposure significantly increases the proliferation of human neuroblastoma (SH-SY5Y) cells. The acceleration of proliferation results from a forward shift of the cell cycle in G1-phase. The G2/M-phase progression is not affected in the HMF. Our data is the first to demonstrate that the HMF can stimulate the proliferation of SH-SY5Y cells by promoting cell cycle progression in the G1-phase. This provides a novel way to study the mechanism of cells in response to changes of environmental magnetic field including the GMF. PMID:23355897

  18. The extraction of the spin structure function, g2 (and g1) at low Bjorken x

    SciTech Connect

    Ndukum, Luwani Z.

    2015-08-01

    The Spin Asymmetries of the Nucleon Experiment (SANE) used the Continuous Electron Beam Accelerator Facility at Jefferson Laboratory in Newport News, VA to investigate the spin structure of the proton. The experiment measured inclusive double polarization electron asymmetries using a polarized electron beam, scattered off a solid polarized ammonia target with target polarization aligned longitudinal and near transverse to the electron beam, allowing the extraction of the spin asymmetries A1 and A2, and spin structure functions g1 and g2. Polarized electrons of energies of 4.7 and 5.9 GeV were used. The scattered electrons were detected by a novel, non-magnetic array of detectors observing a four-momentum transfer range of 2.5 to 6.5 GeV*V. This document addresses the extraction of the spin asymmetries and spin structure functions, with a focus on spin structure function, g2 (and g1) at low Bjorken x. The spin structure functions were measured as a function of x and W in four Q square bins. A full understanding of the low x region is necessary to get clean results for SANE and extend our understanding of the kinematic region at low x.

  19. G1-arrested newborn cells are the predominant infectious form of the pathogen Brucella abortus

    PubMed Central

    Deghelt, Michaël; Mullier, Caroline; Sternon, Jean-François; Francis, Nayla; Laloux, Géraldine; Dotreppe, Delphine; Van der Henst, Charles; Jacobs-Wagner, Christine; Letesson, Jean-Jacques; De Bolle, Xavier

    2014-01-01

    Several intracellular pathogens, such as Brucella abortus, display a biphasic infection process starting with a non-proliferative stage of unclear nature. Here, we study the cell cycle of B. abortus at the single-cell level, in culture and during infection of HeLa cells and macrophages. The localization of segregation and replication loci of the two bacterial chromosomes indicates that, immediately after being engulfed by host-cell endocytic vacuoles, most bacterial cells are newborn. These bacterial cells do not initiate DNA replication for the next 4 to 6 h, indicating a G1 arrest. Moreover, growth is completely stopped during that time, reflecting a global cell cycle block. Growth and DNA replication resume later, although bacteria still reside within endosomal-like compartments. We hypothesize that the predominance of G1-arrested bacteria in the infectious population, and the bacterial cell cycle arrest following internalization, may constitute a widespread strategy among intracellular pathogens to colonize new proliferation niches. PMID:25006695

  20. Revised direct radiocarbon dating of the Vindija G1 Upper Paleolithic Neandertals.

    PubMed

    Higham, Tom; Ramsey, Christopher Bronk; Karavanić, Ivor; Smith, Fred H; Trinkaus, Erik

    2006-01-17

    The 1998/1999 direct dating of two Neandertal specimens from level G(1) of Vindija Cave in Croatia to approximately 28,000 and approximately 29,000 radiocarbon ((14)C) years ago has led to interpretations concerning the late survival of Neandertals in south-central Europe, patterns of interaction between Neandertals and in-dispersing early modern humans in Europe, and complex biocultural scenarios for the earlier phases of the Upper Paleolithic. Given improvements, particularly in sample pretreatment techniques for bone radiocarbon samples, especially ultrafiltration of collagen samples, these Vindija G(1) Neandertal fossils are redated to approximately 32,000-33,000 (14)C years ago and possibly earlier. These results and the recent redating of a number of purportedly old modern human skeletal remains in Europe to younger time periods highlight the importance of fine chronological control when studying this biocultural time period and the tenuous nature of monolithic scenarios for the establishment of modern humans and earlier phases of the Upper Paleolithic in Europe.

  1. Revised direct radiocarbon dating of the Vindija G1 Upper Paleolithic Neandertals

    PubMed Central

    Higham, Tom; Ramsey, Christopher Bronk; Karavanić, Ivor; Smith, Fred H.; Trinkaus, Erik

    2006-01-01

    The 1998/1999 direct dating of two Neandertal specimens from level G1 of Vindija Cave in Croatia to ≈28,000 and ≈29,000 radiocarbon (14C) years ago has led to interpretations concerning the late survival of Neandertals in south-central Europe, patterns of interaction between Neandertals and in-dispersing early modern humans in Europe, and complex biocultural scenarios for the earlier phases of the Upper Paleolithic. Given improvements, particularly in sample pretreatment techniques for bone radiocarbon samples, especially ultrafiltration of collagen samples, these Vindija G1 Neandertal fossils are redated to ≈32,000–33,000 14C years ago and possibly earlier. These results and the recent redating of a number of purportedly old modern human skeletal remains in Europe to younger time periods highlight the importance of fine chronological control when studying this biocultural time period and the tenuous nature of monolithic scenarios for the establishment of modern humans and earlier phases of the Upper Paleolithic in Europe. PMID:16407102

  2. G-1-activated membrane estrogen receptors mediate increased contractility of the human myometrium.

    PubMed

    Maiti, K; Paul, J W; Read, M; Chan, E C; Riley, S C; Nahar, P; Smith, R

    2011-06-01

    Estrogens are key mediators of increased uterine contractility at labor. We sought to determine whether membrane-associated estrogen receptors, such as the recently described seven-transmembrane receptor G protein-coupled receptor 30 (GPR30), mediated some of this effect. Using human myometrium obtained at term cesarean section before or after the onset of labor, we demonstrated the presence of GPR30 mRNA and protein using quantitative RT-PCR and Western blotting. GPR30 receptor was localized to the cell membrane and often colocalized with calveolin-1. Using the specific estrogen membrane receptor agonist G-1 and myometrial explants, we showed that membrane receptor activation led to phosphorylation of MAPK and the actin-modifying small heat shock protein 27. Using myometrial strips incubated with G-1 or vehicle we demonstrated that estrogen membrane receptor activation increased the myometrial contractile response to oxytocin. These data suggest that activation of the plasma membrane estrogen receptor GPR30 likely participates in the physiology of the human myometrium during pregnancy and identifies it as a potential target to modify uterine activity. PMID:21427217

  3. Engineering upper hinge improves stability and effector function of a human IgG1.

    PubMed

    Yan, Boxu; Boyd, Daniel; Kaschak, Timothy; Tsukuda, Joni; Shen, Amy; Lin, Yuwen; Chung, Shan; Gupta, Priyanka; Kamath, Amrita; Wong, Anne; Vernes, Jean-Michel; Meng, Gloria Y; Totpal, Klara; Schaefer, Gabriele; Jiang, Guoying; Nogal, Bartek; Emery, Craig; Vanderlaan, Martin; Carter, Paul; Harris, Reed; Amanullah, Ashraf

    2012-02-17

    Upper hinge is vulnerable to radical attacks that result in breakage of the heavy-light chain linkage and cleavage of the hinge of an IgG1. To further explore mechanisms responsible for the radical induced hinge degradation, nine mutants were designed to determine the roles that the upper hinge Asp and His play in the radical reactions. The observation that none of these substitutions could inhibit the breakage of the heavy-light chain linkage suggests that the breakage may result from electron transfer from Cys(231) directly to the heavy-light chain linkage upon radical attacks, and implies a pathway separate from His(229)-mediated hinge cleavage. On the other hand, the substitution of His(229) with Tyr showed promising advantages over the native antibody and other substitutions in improving the stability and function of the IgG1. This substitution inhibited the hinge cleavage by 98% and suggests that the redox active nature of Tyr did not enable it to replicate the ability of His to facilitate radical induced degradation. We propose that the lower redox potential of Tyr, a residue that may be the ultimate sink for oxidizing equivalents in proteins, is responsible for the inhibition. More importantly, the substitution increased the antibody's binding to FcγRIII receptors by 2-3-fold, and improved ADCC activity by 2-fold, while maintaining a similar pharmacokinetic profile with respect to the wild type. Implications of these observations for antibody engineering and development are discussed.

  4. Magnetic shielding accelerates the proliferation of human neuroblastoma cell by promoting G1-phase progression.

    PubMed

    Mo, Wei-chuan; Zhang, Zi-jian; Liu, Ying; Bartlett, Perry F; He, Rong-qiao

    2013-01-01

    Organisms have been exposed to the geomagnetic field (GMF) throughout evolutionary history. Exposure to the hypomagnetic field (HMF) by deep magnetic shielding has recently been suggested to have a negative effect on the structure and function of the central nervous system, particularly during early development. Although changes in cell growth and differentiation have been observed in the HMF, the effects of the HMF on cell cycle progression still remain unclear. Here we show that continuous HMF exposure significantly increases the proliferation of human neuroblastoma (SH-SY5Y) cells. The acceleration of proliferation results from a forward shift of the cell cycle in G1-phase. The G2/M-phase progression is not affected in the HMF. Our data is the first to demonstrate that the HMF can stimulate the proliferation of SH-SY5Y cells by promoting cell cycle progression in the G1-phase. This provides a novel way to study the mechanism of cells in response to changes of environmental magnetic field including the GMF. PMID:23355897

  5. Dectin-1 agonist selectively induces IgG1 class switching by LPS-activated mouse B cells.

    PubMed

    Seo, Beom-Seok; Park, Ha-Yan; Yoon, Hee-Kyung; Yoo, Yung-Choon; Lee, Junglim; Park, Seok-Rae

    2016-10-01

    Heat-killed Saccharomyces cerevisiae (HKSC) is an agonist for Dectin-1, a major fungal cell wall β-glucan receptor. We previously reported that HKSC selectively enhances IgG1 production by LPS-activated mouse B cells. To determine if this IgG1 selectivity is caused by selective IgG1 class switching, we performed RT-PCRs for measuring germline transcripts (GLTs), flow cytometric analyses for detecting Ig-expressing cells, and ELISPOT assays for measuring the number of Ig-secreting cells in HKSC/LPS-stimulated mouse B cell cultures. HKSC selectively enhanced expression of GLTγ1, the number of IgG1-expressing cells, and the number of IgG1-secreting B cells in the presence of LPS stimulation. In addition, HKSC induced the expression of CD69, an activation marker for B lymphocytes, and the expression of surface Dectin-1. Two Dectin-1 antagonists, laminarin and a neutralizing Dectin-1 antibody, selectively diminished HKSC-reinforced IgG1 production by LPS-stimulated B cells. Furthermore, depleted zymosan (dzn), a Dectin-1 agonist with increased selectivity, also selectively enhanced GLTγ1 transcription. The Dectin-1 antagonists blocked dzn-induced IgG1 production by LPS-activated B cells. Collectively, these results suggest that Dectin-1 agonists selectively induce IgG1 class switching by direct stimulation of Dectin-1 on LPS-activated B cells resulting in selective production of IgG1. PMID:27568820

  6. Dectin-1 agonist selectively induces IgG1 class switching by LPS-activated mouse B cells.

    PubMed

    Seo, Beom-Seok; Park, Ha-Yan; Yoon, Hee-Kyung; Yoo, Yung-Choon; Lee, Junglim; Park, Seok-Rae

    2016-10-01

    Heat-killed Saccharomyces cerevisiae (HKSC) is an agonist for Dectin-1, a major fungal cell wall β-glucan receptor. We previously reported that HKSC selectively enhances IgG1 production by LPS-activated mouse B cells. To determine if this IgG1 selectivity is caused by selective IgG1 class switching, we performed RT-PCRs for measuring germline transcripts (GLTs), flow cytometric analyses for detecting Ig-expressing cells, and ELISPOT assays for measuring the number of Ig-secreting cells in HKSC/LPS-stimulated mouse B cell cultures. HKSC selectively enhanced expression of GLTγ1, the number of IgG1-expressing cells, and the number of IgG1-secreting B cells in the presence of LPS stimulation. In addition, HKSC induced the expression of CD69, an activation marker for B lymphocytes, and the expression of surface Dectin-1. Two Dectin-1 antagonists, laminarin and a neutralizing Dectin-1 antibody, selectively diminished HKSC-reinforced IgG1 production by LPS-stimulated B cells. Furthermore, depleted zymosan (dzn), a Dectin-1 agonist with increased selectivity, also selectively enhanced GLTγ1 transcription. The Dectin-1 antagonists blocked dzn-induced IgG1 production by LPS-activated B cells. Collectively, these results suggest that Dectin-1 agonists selectively induce IgG1 class switching by direct stimulation of Dectin-1 on LPS-activated B cells resulting in selective production of IgG1.

  7. Analysis of aflatoxins in nonalcoholic beer using liquid-liquid extraction and ultraperformance LC-MS/MS.

    PubMed

    Khan, Mohammad R; Alothman, Zeid A; Ghfar, Ayman A; Wabaidur, Saikh M

    2013-02-01

    Aflatoxins AFB1, AFB2, AFG1, and AFG2 are toxic secondary metabolites produced by Aspergillus flavus and Aspergillus parasiticus and posses a potential threat to food safety. In the present work, liquid-liquid extraction and ultraperformance LC-MS/MS method has been applied for the determination of four naturally occurring aflatoxins AFB1, AFB2, AFG1, and AFG2 in nonalcoholic beer. Aflatoxins extraction from nonalcoholic beer was carried out using liquid-liquid extraction procedure. The effects of solvent-types were studied to obtain maximum recovery of the target analytes with minimum contamination. Among different solvents, the aflatoxins extraction was best achieved using ethyl acetate. The obtained recoveries were ranged from 85 to 96% with good quality parameters: LOD values between 0.001 and 0.003 ng/mL, linearity of the calibration curve (r(2) > 0.999), and repeatability (run-to-run) and reproducibility (day-to-day) precisions with RSDs lower than 5% (n = 5) achieved at 0.50 ng/mL concentration. The optimized liquid-liquid extraction in combination with ultraperformance LC-MS/MS was applied successfully to the analysis of AFB1, AFB2, AFG1, and AFG2 aflatoxins in 11 nonalcoholic beers and were detected up to 15.31 ng/L in some of the samples.

  8. Asymmetric Expansion of the Youngest Galactic Supernova Remnant G1.9+0.3

    NASA Astrophysics Data System (ADS)

    Borkowski, Kazimierz J.; Green, David; Gwynne, Peter; Hwang, Una; Petre, Robert; Reynolds, Stephen P.; Willett, Rebecca

    2016-04-01

    The youngest Galactic supernova remnant (SNR) G1.9+0.3, produced by a (likely) Type Ia SN that exploded around CE 1900, is strongly asymmetric at radio wavelengths but exhibits a bilaterally symmetric morphology in X-rays. It has been difficult to understand the origin of these contrasting morphologies. We present results of X-ray expansion measurements of G1.9+0.3 that illuminate the origin of the radio asymmetry. These measurements are based on comparing recent (2015), 400 ks-long Chandra observations with earlier Chandra observations that include 1 Ms-long 2011 observations. The mean expansion rate from 2011 to 2015 is 0.58% yr-1, in agreement with previous measurements. We also confirm that expansion decreases radially away from the remnant's center along the major E-W axis, from 0.77% yr-1 to 0.53% yr-1. Large variations in expansion are also present along the minor N-S axis. Expansion of the faint S rim and the outermost faint N rim is comparable to the mean expansion. But the prominent X-ray rim in the N, coincident with the outer edge of the bright radio rim that marks the primary blast wave there, is expanding more slowly. Its expansion relative to the S rim is only 0.47% yr-1. At 8.5 kpc, this corresponds to a speed of about 5000 km/s, less than half of the overall blast wave speed of 12,000 km/s. Such strong deceleration of the northern blast wave most likely arises from the collision of SN ejecta with a much denser than average ambient medium there. The presence of the asymmetric ambient medium naturally explains the radio asymmetry. The SN ejecta have also been strongly decelerated in the N, but they expand faster than the blast wave. In several locations, significant morphological changes and strongly nonradial motions are apparent. The spatially-integrated X-ray flux continues to increase with time. As with Kepler's SN - the most recent historical SN in the Galaxy - the SN ejecta are likely colliding with the asymmetric circumstellar medium (CSM

  9. Birth of Archaeal Cells: Molecular Phylogenetic Analyses of G1P Dehydrogenase, G3P Dehydrogenases, and Glycerol Kinase Suggest Derived Features of Archaeal Membranes Having G1P Polar Lipids

    PubMed Central

    2016-01-01

    Bacteria and Eukarya have cell membranes with sn-glycerol-3-phosphate (G3P), whereas archaeal membranes contain sn-glycerol-1-phosphate (G1P). Determining the time at which cells with either G3P-lipid membranes or G1P-lipid membranes appeared is important for understanding the early evolution of terrestrial life. To clarify this issue, we reconstructed molecular phylogenetic trees of G1PDH (G1P dehydrogenase; EgsA/AraM) which is responsible for G1P synthesis and G3PDHs (G3P dehydrogenase; GpsA and GlpA/GlpD) and glycerol kinase (GlpK) which is responsible for G3P synthesis. Together with the distribution of these protein-encoding genes among archaeal and bacterial groups, our phylogenetic analyses suggested that GlpA/GlpD in the Commonote (the last universal common ancestor of all extant life with a cellular form, Commonote commonote) acquired EgsA (G1PDH) from the archaeal common ancestor (Commonote archaea) and acquired GpsA and GlpK from a bacterial common ancestor (Commonote bacteria). In our scenario based on this study, the Commonote probably possessed a G3P-lipid membrane synthesized enzymatically, after which the archaeal lineage acquired G1PDH followed by the replacement of a G3P-lipid membrane with a G1P-lipid membrane. PMID:27774041

  10. Production of Group Specific Monoclonal Antibody to Aflatoxins and its Application to Enzyme-linked Immunosorbent Assay

    PubMed Central

    Kim, Sung-Hee; Cha, Sang-Ho; Karyn, Bischoff; Park, Sung-Won; Son, Seong-Wan

    2011-01-01

    Through the present study, we produced a monoclonal antibody against aflatoxin B1 (AFB1) using AFB1- carboxymethoxylamine BSA conjugates. One clone showing high binding ability was selected and it was applied to develop a direct competitive ELISA system. The epitope densities of AFB1-CMO against BSA and KLH were about 1 : 6 and 1 : 545, respectively. The monoclonal antibody (mAb) from cloned hybridoma cell was the IgG1 subclass with λ-type light chains. The IC50s of the monoclonal antibody developed for AFB1, AFB2, AFG1 and AFG2 were 4.36, 7.22, 6.61 and 29.41 ng/ml, respectively, based on the AFB1-KLH coated ELISA system and 15.28, 26.62, 32.75 and 56.67 ng/ml, respectively, based on the mAb coated ELISA. Cross-relativities of mAb to AFB1 for AFB2, AFG1 and AFG2 were 60.47, 65.97 and 14.83% in the AFB1-KLH coated ELISA, and 59.41, 46.66 and 26.97% in the mAb coated ELISA, respectively. Quantitative calculations for AFB1 from the AFB1-Ab ELISA and AFB1-Ag ELISA ranged from 0.25 to 25 ng/ml (R2 > 0.99) and from 1 to 100 ng/ml (R2 > 0.99), respectively. The intra- and inter-assay precision CVs were < 10% in both ELISA assay, representing good reproducibility of developed assay. Recoveries ranged from 79.18 to 91.27%, CVs ranged from 3.21 to 7.97% after spiking AFB1 at concentrations ranging from 5 to 50 ng/ml and following by extraction with 70% methanol solution in the Ab-coated ELISA. In conclusion, we produced a group specific mAb against aflatoxins and developed two direct competitive ELISAs for the detection of AFB1 in feeds based on a monoclonal antibody developed. PMID:24278561

  11. Simultaneous multi-mycotoxin determination in nutmeg by ultrasound-assisted solid-liquid extraction and immunoaffinity column clean-up coupled with liquid chromatography and on-line post-column photochemical derivatization-fluorescence detection.

    PubMed

    Kong, Wei-Jun; Liu, Shu-Yu; Qiu, Feng; Xiao, Xiao-He; Yang, Mei-Hua

    2013-05-01

    A simple and sensitive analytical method based on ultrasound-assisted solid-liquid extraction and immunoaffinity column clean-up coupled with high performance liquid chromatography and on-line post-column photochemical derivatization-fluorescence detection (USLE-IAC-HPLC-PCD-FLD) has been developed for simultaneous multi-mycotoxin determination of aflatoxins B1, B2, G1, G2 (AFB1, AFB2, AFG1, AFG2) and ochratoxin A (OTA) in 13 edible and medicinal nutmeg samples marketed in China. AFs and OTA were extracted from nutmeg samples by ultrasonication using a methanol : water (80 : 20, v/v) solution, followed by an IAC clean-up step. Different USL extraction conditions, pre-processing ways for nutmeg sample and clean-up columns for mycotoxins, as well as HPLC-PCD-FLD parameters (mobile phase, column temperature, elution procedure, excitation and emission wavelengths) were optimized. This method, which was appraised for analyzing nutmeg samples, showed satisfactory results with reference to limits of detection (LODs) (from 0.02 to 0.25 μg kg(-1)), limits of quantification (LOQs) (from 0.06 to 0.8 μg kg(-1)), linear ranges (up to 30 ng mL(-1) for AFB1, AFG1 and OTA and 9 ng mL(-1) for AFB2 and AFG2), intra- and inter-day variability (all <2%) and average recoveries (from 79.6 to 90.8% for AFs and from 93.6 to 97.3% for OTA, respectively). The results of the application of developed method in nutmeg samples have elucidated that four samples were detected with contamination of AFs and one with OTA. AFB1 was the most frequently found mycotoxin in 30.8% of nutmeg samples at contamination levels of 0.73-16.31 μg kg(-1). At least two different mycotoxins were co-occurred in three samples, and three AFs were simultaneously detected in one sample.

  12. Simultaneous multi-mycotoxin determination in nutmeg by ultrasound-assisted solid-liquid extraction and immunoaffinity column clean-up coupled with liquid chromatography and on-line post-column photochemical derivatization-fluorescence detection.

    PubMed

    Kong, Wei-Jun; Liu, Shu-Yu; Qiu, Feng; Xiao, Xiao-He; Yang, Mei-Hua

    2013-05-01

    A simple and sensitive analytical method based on ultrasound-assisted solid-liquid extraction and immunoaffinity column clean-up coupled with high performance liquid chromatography and on-line post-column photochemical derivatization-fluorescence detection (USLE-IAC-HPLC-PCD-FLD) has been developed for simultaneous multi-mycotoxin determination of aflatoxins B1, B2, G1, G2 (AFB1, AFB2, AFG1, AFG2) and ochratoxin A (OTA) in 13 edible and medicinal nutmeg samples marketed in China. AFs and OTA were extracted from nutmeg samples by ultrasonication using a methanol : water (80 : 20, v/v) solution, followed by an IAC clean-up step. Different USL extraction conditions, pre-processing ways for nutmeg sample and clean-up columns for mycotoxins, as well as HPLC-PCD-FLD parameters (mobile phase, column temperature, elution procedure, excitation and emission wavelengths) were optimized. This method, which was appraised for analyzing nutmeg samples, showed satisfactory results with reference to limits of detection (LODs) (from 0.02 to 0.25 μg kg(-1)), limits of quantification (LOQs) (from 0.06 to 0.8 μg kg(-1)), linear ranges (up to 30 ng mL(-1) for AFB1, AFG1 and OTA and 9 ng mL(-1) for AFB2 and AFG2), intra- and inter-day variability (all <2%) and average recoveries (from 79.6 to 90.8% for AFs and from 93.6 to 97.3% for OTA, respectively). The results of the application of developed method in nutmeg samples have elucidated that four samples were detected with contamination of AFs and one with OTA. AFB1 was the most frequently found mycotoxin in 30.8% of nutmeg samples at contamination levels of 0.73-16.31 μg kg(-1). At least two different mycotoxins were co-occurred in three samples, and three AFs were simultaneously detected in one sample. PMID:23486692

  13. Echinococcus canadensis (G7) and Echinococcus granulosus sensu stricto (G1) in swine of southern Brazil.

    PubMed

    Monteiro, D U; Botton, S A; Tonin, A A; Azevedo, M I; Graichen, D A S; Noal, C B; de la Rue, M L

    2014-05-28

    The cystic echinococcosis (CE) is an important zoonotic disease caused by the parasite Echinococcus spp. In Brazil, this parasite is present in Rio Grande do Sul (RS) state, border with Argentina and Uruguay, causing several damages to human and animal health. This study aimed to identify Echinococcus spp. in hydatid cysts of swine and evaluate the similarity of the genotypes through the phylogenetic analysis. A total of 3,101,992 swine were slaughtered in the central/northern region of RS/Brazil, during 2008-2012. Five isolates were characterized as hydatid cyst by molecular analysis, based on the mitochondrial gene cytochrome c oxidase subunit I (cox-I). The genotypes E. granulosus sensu stricto (G1) (n=2) and E. canadensis (G7) (n=3) were identified in the hydatid cysts. The swine represents a potential intermediate host for different genotypes of Echinococcus spp., besides it can contribute to the perpetuation of the parasite's life cycle in rural areas.

  14. Cdi1, a human G1 and S phase protein phosphatase that associates with Cdk2.

    PubMed

    Gyuris, J; Golemis, E; Chertkov, H; Brent, R

    1993-11-19

    We used the interaction trap, a yeast genetic selection for interacting proteins, to isolate human cyclin-dependent kinase interactor 1 (Cdi1). In yeast, Cdi1 interacts with cyclin-dependent kinases, including human Cdc2, Cdk2, and Cdk3, but not with Ckd4. In HeLa cells, Cdi1 is expressed at the G1 to S transition, and the protein forms stable complexes with Cdk2. Cdi1 bears weak sequence similarity to known tyrosine and dual specificity phosphatases. In vitro, Cdi1 removes phosphate from tyrosine residues in model substrates, but a mutant protein that bears a lesion in the putative active site cysteine does not. Overexpression of wild-type Cdi1 delays progression through the cell cycle in yeast and HeLa cells; delay is dependent on Cdi1 phosphatase activity. These experiments identify Cdi1 as a novel type of protein phosphatase that forms complexes with cyclin-dependent kinases. PMID:8242750

  15. The impact of geoengineering on vegetation in experiment G1 of the GeoMIP

    NASA Astrophysics Data System (ADS)

    Glienke, Susanne; Irvine, Peter J.; Lawrence, Mark G.

    2015-10-01

    Solar Radiation Management (SRM) has been proposed as a mean to partly counteract global warming. The Geoengineering Model Intercomparison Project (GeoMIP) has simulated the climate consequences of a number of SRM techniques. Thus far, the effects on vegetation have not yet been thoroughly analyzed. Here the vegetation response to the idealized GeoMIP G1 experiment from eight fully coupled Earth system models (ESMs) is analyzed, in which a reduction of the solar constant counterbalances the radiative effects of quadrupled atmospheric CO2 concentrations (abrupt4 × CO2). For most models and regions, changes in net primary productivity (NPP) are dominated by the increase in CO2, via the CO2 fertilization effect. As SRM will reduce temperatures relative to abrupt4 × CO2, in high latitudes this will offset increases in NPP. In low latitudes, this cooling relative to the abrupt4 × CO2 simulation decreases plant respiration while having little effect on gross primary productivity, thus increasing NPP. In Central America and the Mediterranean, generally dry regions which are expected to experience increased water stress with global warming, NPP is highest in the G1 experiment for all models due to the easing of water limitations from increased water use efficiency at high-CO2 concentrations and the reduced evaporative demand in a geoengineered climate. The largest differences in the vegetation response are between models with and without a nitrogen cycle, with a much smaller CO2 fertilization effect for the former. These results suggest that until key vegetation processes are integrated into ESM predictions, the vegetation response to SRM will remain highly uncertain.

  16. Supernova Ejecta in the Youngest Galactic Supernova Remnant G1.9+0.3

    NASA Technical Reports Server (NTRS)

    Borkowski, Kazimierz J.; Reynolds, Stephen P.; Hwang, Una; Green, David A.; Petre, Robert; Krishnamurthy, Kalyani; Willett, Rebecca

    2013-01-01

    G1.9+0.3 is the youngest known Galactic supernova remnant (SNR), with an estimated supernova (SN) explosion date of approximately 1900, and most likely located near the Galactic Center. Only the outermost ejecta layers with free-expansion velocities (is) approximately greater than 18,000 km s-1 have been shocked so far in this dynamically young, likely Type Ia SNR. A long (980 ks) Chandra observation in 2011 allowed spatially-resolved spectroscopy of heavy-element ejecta. We denoised Chandra data with the spatio-spectral method of Krishnamurthy et al., and used a wavelet based technique to spatially localize thermal emission produced by intermediate-mass elements (IMEs: Si and S) and iron. The spatial distribution of both IMEs and Fe is extremely asymmetric, with the strongest ejecta emission in the northern rim. Fe K alpha emission is particularly prominent there, and fits with thermal models indicate strongly oversolar Fe abundances. In a localized, outlying region in the northern rim, IMEs are less abundant than Fe, indicating that undiluted Fe-group elements (including 56Ni) with velocities greater than 18,000 km s-1 were ejected by this SN. But in the inner west rim, we find Si- and S-rich ejecta without any traces of Fe, so high-velocity products of O-burning were also ejected. G1.9+0.3 appears similar to energetic Type Ia SNe such as SN 2010jn where iron-group elements at such high free-expansion velocities have been recently detected. The pronounced asymmetry in the ejecta distribution and abundance inhomogeneities are best explained by a strongly asymmetric SN explosion, similar to those produced in some recent 3D delayed-detonation Type Ia models.

  17. Production of α2,6-sialylated IgG1 in CHO cells

    PubMed Central

    Raymond, Céline; Robotham, Anna; Spearman, Maureen; Butler, Michael; Kelly, John; Durocher, Yves

    2015-01-01

    The presence of α2,6-sialic acids on the Fc N-glycan provides anti-inflammatory properties to the IgGs through a mechanism that remains unclear. Fc-sialylated IgGs are rare in humans as well as in industrial host cell lines such as Chinese hamster ovary (CHO) cells. Facilitated access to well-characterized α2,6-sialylated IgGs would help elucidate the mechanism of this intriguing IgG's effector function. This study presents a method for the efficient Fc glycan α2,6-sialylation of a wild-type and a F243A IgG1 mutant by transient co-expression with the human α2,6-sialyltransferase 1 (ST6) and β1,4-galactosyltransferase 1 (GT) in CHO cells. Overexpression of ST6 alone only had a moderate effect on the glycoprofiles, whereas GT alone greatly enhanced Fc-galactosylation, but not sialylation. Overexpression of both GT and ST6 was necessary to obtain a glycoprofile dominated by α2,6-sialylated glycans in both antibodies. The wild-type was composed of the G2FS(6)1 glycan (38%) with remaining unsialylated glycans, while the mutant glycoprofile was essentially composed of G2FS(6)1 (25%), G2FS(3,6)2 (16%) and G2FS(6,6)2 (37%). The α2,6-linked sialic acids represented over 85% of all sialic acids in both antibodies. We discuss how the limited sialylation level in the wild-type IgG1 expressed alone or with GT results from the glycan interaction with Fc's amino acid residues or from intrinsic galactosyl- and sialyl-transferases substrate specificities. PMID:25875452

  18. SUPERNOVA EJECTA IN THE YOUNGEST GALACTIC SUPERNOVA REMNANT G1.9+0.3

    SciTech Connect

    Borkowski, Kazimierz J.; Reynolds, Stephen P.; Hwang, Una; Green, David A.; Petre, Robert

    2013-07-01

    G1.9+0.3 is the youngest known Galactic supernova remnant (SNR), with an estimated supernova (SN) explosion date of {approx}1900, and most likely located near the Galactic center. Only the outermost ejecta layers with free-expansion velocities {approx}>18,000 km s{sup -1} have been shocked so far in this dynamically young, likely Type Ia SNR. A long (980 ks) Chandra observation in 2011 allowed spatially resolved spectroscopy of heavy-element ejecta. We denoised Chandra data with the spatio-spectral method of Krishnamurthy et al., and used a wavelet-based technique to spatially localize thermal emission produced by intermediate-mass elements (IMEs; Si and S) and iron. The spatial distribution of both IMEs and Fe is extremely asymmetric, with the strongest ejecta emission in the northern rim. Fe K{alpha} emission is particularly prominent there, and fits with thermal models indicate strongly oversolar Fe abundances. In a localized, outlying region in the northern rim, IMEs are less abundant than Fe, indicating that undiluted Fe-group elements (including {sup 56}Ni) with velocities >18,000 km s{sup -1} were ejected by this SN. However, in the inner west rim, we find Si- and S-rich ejecta without any traces of Fe, so high-velocity products of O-burning were also ejected. G1.9+0.3 appears similar to energetic Type Ia SNe such as SN 2010jn where iron-group elements at such high free-expansion velocities have been recently detected. The pronounced asymmetry in the ejecta distribution and abundance inhomogeneities are best explained by a strongly asymmetric SN explosion, similar to those produced in some recent three-dimensional delayed-detonation Type Ia models.

  19. D-type cyclins and G1 progression during liver development in the rat

    SciTech Connect

    Boylan, Joan M. . E-mail: Joan_Boylan@brown.edu; Gruppuso, Philip A. . E-mail: Philip_Gruppuso@brown.edu

    2005-05-13

    Initiation and progression through G1 requires the activity of signaling complexes containing cyclins (D- or E-type) and cyclin-dependent kinases (CDK4/6 and CDK2, respectively). We set out to identify the G1-phase cyclins and CDKs that are operative during late gestation liver development in the rat. This is a period during which hepatocytes show a high rate of proliferation that is, at least in part, independent of the mitogenic signaling pathways that are functional in mature hepatocytes. RNase protection assay and Western immunoblotting indicated that cyclin D1 is expressed at similar levels in fetal and adult liver. When cyclin D1 was induced after partial hepatectomy, its predominant CDK-binding partner was CDK4. In contrast, cyclins D2 and D3 predominated in fetal liver and were complexed with both CDK4 and CDK6. Little CDK6 protein was expressed in quiescent or regenerating adult liver. Cyclins E1 and E2 were both transcriptionally up-regulated in fetal liver. Activity of complexes containing cyclins E1 and E2 was higher in fetal liver, as was content of the cell cycle regulator, Rb. In fetal liver, Rb was highly phosphorylated at both cyclin D- and cyclin E-dependent sites. In conclusion, liver development is associated with a switch from cyclin D2/D3-containing complexes to cyclin D1:CDK4 complexes. We speculate that the switch in D-type cyclins may be associated with the dependence on mitogenic signaling that develops as hepatocytes mature.

  20. Nuclear vasohibin-2 promotes cell proliferation by inducing G0/G1 to S phase progression.

    PubMed

    Ge, Qianqian; Zhou, Jia; Tu, Min; Xue, Xiaofeng; Li, Zhanjun; Lu, Zipeng; Wei, Jishu; Song, Guoxin; Chen, Jianmin; Guo, Feng; Jiang, Kuirong; Miao, Yi; Gao, Wentao

    2015-09-01

    As a member of the vasohibin (VASH2) family, VASH2 is localized intracellularly as a nuclear and cytoplasmic type. Cytoplasmic VASH2 is associated with carcinoma angiogenesis and malignant transformation and promotes cancer growth. However, the function of nuclear VASH2 has yet to be investigated. The aim of the present study was to detect the nuclear VASH2 expression profile in human organs and tissues by protein microarray technique. To examine the function of nuclear VASH2, we analyzed the relationship between nuclear VASH2 and Ki-67, and stably constructed VASH2 overexpression and knockdown in LO2 and HepG2 cell lines, based on a previous study in hepatic cells. The study was conducted using bromodeoxyuridine, immunofluorescent staining, western blot analysis and flow cytometry. Nuclear VASH2 was highly expressed in actively dividing cells in normal and cancer tissues. There was a significant positive correlation between nuclear VASH2 and Ki-67, indicating that nuclear VASH2 positively correlated with cell proliferation in normal and cancer tissues. The bromodeoxyuridine (BrdU) proliferation test showed that nuclear VASH2 increased the S-phase population and promoted cell proliferation, while VASH2 knockdown reduced BrdU absorbance. Cell cycle analysis revealed that nuclear VASH2 overexpression increased the S-phase population in LO2 and HepG2 cells, while nuclear VASH2 knockdown reduced the S-phase population and increased the G0/G1 population. The findings of this study challenge the classic view of VASH2, which was previously reported as an angiogenesis factor. Furthermore, to the best of our knowledge, these results are the first clinical data indicating that nuclear VASH2, but not cytoplasmic VASH2, promotes cell proliferation by driving the cell cycle from the G0/G1 to S phase.

  1. A prominent lack of IgG1-Fc fucosylation of platelet alloantibodies in pregnancy.

    PubMed

    Kapur, Rick; Kustiawan, Iwan; Vestrheim, Anne; Koeleman, Carolien A M; Visser, Remco; Einarsdottir, Helga K; Porcelijn, Leendert; Jackson, Dave; Kumpel, Belinda; Deelder, André M; Blank, Dennis; Skogen, Björn; Killie, Mette Kjaer; Michaelsen, Terje E; de Haas, Masja; Rispens, Theo; van der Schoot, C Ellen; Wuhrer, Manfred; Vidarsson, Gestur

    2014-01-23

    Immunoglobulin G (IgG) formed during pregnancy against human platelet antigens (HPAs) of the fetus mediates fetal or neonatal alloimmune thrombocytopenia (FNAIT). Because antibody titer or isotype does not strictly correlate with disease severity, we investigated by mass spectrometry variations in the glycosylation at Asn297 in the IgG Fc because the composition of this glycan can be highly variable, affecting binding to phagocyte IgG-Fc receptors (FcγR). We found markedly decreased levels of core fucosylation of anti-HPA-1a-specific IgG1 from FNAIT patients (n = 48), but not in total serum IgG1. Antibodies with a low amount of fucose displayed higher binding affinity to FcγRIIIa and FcγRIIIb, but not to FcγRIIa, compared with antibodies with a high amount of Fc fucose. Consequently, these antibodies with a low amount of Fc fucose showed enhanced phagocytosis of platelets using FcγRIIIb(+) polymorphonuclear cells or FcγRIIIa(+) monocytes as effector cells, but not with FcγRIIIa(-) monocytes. In addition, the degree of anti-HPA-1a fucosylation correlated positively with the neonatal platelet counts in FNAIT, and negatively to the clinical disease severity. In contrast to the FNAIT patients, no changes in core fucosylation were observed for anti-HLA antibodies in refractory thrombocytopenia (post platelet transfusion), indicating that the level of fucosylation may be antigen dependent and/or related to the immune milieu defined by pregnancy.

  2. A prominent lack of IgG1-Fc fucosylation of platelet alloantibodies in pregnancy

    PubMed Central

    Kapur, Rick; Kustiawan, Iwan; Vestrheim, Anne; Koeleman, Carolien A. M.; Visser, Remco; Einarsdottir, Helga K.; Porcelijn, Leendert; Jackson, Dave; Kumpel, Belinda; Deelder, André M.; Blank, Dennis; Skogen, Björn; Killie, Mette Kjaer; Michaelsen, Terje E.; de Haas, Masja; Rispens, Theo; van der Schoot, C. Ellen; Wuhrer, Manfred

    2014-01-01

    Immunoglobulin G (IgG) formed during pregnancy against human platelet antigens (HPAs) of the fetus mediates fetal or neonatal alloimmune thrombocytopenia (FNAIT). Because antibody titer or isotype does not strictly correlate with disease severity, we investigated by mass spectrometry variations in the glycosylation at Asn297 in the IgG Fc because the composition of this glycan can be highly variable, affecting binding to phagocyte IgG-Fc receptors (FcγR). We found markedly decreased levels of core fucosylation of anti-HPA-1a–specific IgG1 from FNAIT patients (n = 48), but not in total serum IgG1. Antibodies with a low amount of fucose displayed higher binding affinity to FcγRIIIa and FcγRIIIb, but not to FcγRIIa, compared with antibodies with a high amount of Fc fucose. Consequently, these antibodies with a low amount of Fc fucose showed enhanced phagocytosis of platelets using FcγRIIIb+ polymorphonuclear cells or FcγRIIIa+ monocytes as effector cells, but not with FcγRIIIa– monocytes. In addition, the degree of anti-HPA-1a fucosylation correlated positively with the neonatal platelet counts in FNAIT, and negatively to the clinical disease severity. In contrast to the FNAIT patients, no changes in core fucosylation were observed for anti-HLA antibodies in refractory thrombocytopenia (post platelet transfusion), indicating that the level of fucosylation may be antigen dependent and/or related to the immune milieu defined by pregnancy. PMID:24243971

  3. Retinoic acid receptor agonists regulate expression of ATP-binding cassette transporter G1 in macrophages.

    PubMed

    Ayaori, Makoto; Yakushiji, Emi; Ogura, Masatsune; Nakaya, Kazuhiro; Hisada, Tetsuya; Uto-Kondo, Harumi; Takiguchi, Shunichi; Terao, Yoshio; Sasaki, Makoto; Komatsu, Tomohiro; Iizuka, Maki; Yogo, Makiko; Uehara, Yoshinari; Kagechika, Hiroyuki; Nakanishi, Tsuyoshi; Ikewaki, Katsunori

    2012-04-01

    ABC transporter G1 (ABCG1) plays a pivotal role in HDL-mediated cholesterol efflux and atherogenesis. We investigated whether, and how, retinoic acid receptors (RARs) regulate ABCG1 expression in macrophages. All-trans retinoic acid (ATRA), an RAR ligand, increased ABCG1 protein levels and apoA-I/HDL-mediated cholesterol efflux from the macrophages. Both ATRA and other RAR agonists, TTNPB and Am580, increased major transcripts driven by promoter B upstream of exon 5, though minor transcripts driven by promoter A upstream of exon 1 were only increased by ATRA. The stimulatory effects of ATRA on ABCG1 expression were completely abolished in the presence of RAR/RXR antagonists but were only partially canceled in the presence of an LXR antagonist. Adenovirus with overexpressed oxysterol sulfotransferase abolished the LXR pathway, as previously reported, and ATRA-responsiveness in ABCA1/ABCG1 expressions were respectively attenuated by 38 and 22% compared to the control virus. Promoter assays revealed that ABCG1 levels were regulated more by promoter B than promoter A, and ATRA activated promoter B in a liver X receptor-responsive element (LXRE)-dependent manner. Further, LXRE-B in intron 7, but not LXRE-A in intron 5, enhanced ATRA responsiveness under overexpression of all RAR isoforms-RARα/β/γ. In contrast, the activation of promoter B by TTNPB depended on LXRE-B and RARα, but not on RARβ/γ. Finally, chromatin immunoprecipitation and gel-shift assays revealed a specific and direct repeat 4-dependent binding of RARα to LXRE-B. In conclusion, RAR ligands increase ABCA1/G1 expression and apoA-I/HDL-mediated cholesterol efflux from macrophages, and modulate ABCG1 promoter activity via LXRE-dependent mechanisms.

  4. Phosphorylation-triggered CUEDC2 degradation promotes UV-induced G1 arrest through APC/C(Cdh1) regulation.

    PubMed

    Zhang, Wei-Na; Zhou, Jie; Zhou, Tao; Li, Ai-Ling; Wang, Na; Xu, Jin-Jing; Chang, Yan; Man, Jiang-Hong; Pan, Xin; Li, Tao; Li, Wei-Hua; Mu, Rui; Liang, Bing; Chen, Liang; Jin, Bao-Feng; Xia, Qing; Gong, Wei-Li; Zhang, Xue-Min; Wang, Li; Li, Hui-Yan

    2013-07-01

    DNA damage triggers cell cycle arrest to provide a time window for DNA repair. Failure of arrest could lead to genomic instability and tumorigenesis. DNA damage-induced G1 arrest is generally achieved by the accumulation of Cyclin-dependent kinase inhibitor 1 (p21). However, p21 is degraded and does not play a role in UV-induced G1 arrest. The mechanism of UV-induced G1 arrest thus remains elusive. Here, we have identified a critical role for CUE domain-containing protein 2 (CUEDC2) in this process. CUEDC2 binds to and inhibits anaphase-promoting complex/cyclosome-Cdh1 (APC/C(Cdh1)), a critical ubiquitin ligase in G1 phase, thereby stabilizing Cyclin A and promoting G1-S transition. In response to UV irradiation, CUEDC2 undergoes ERK1/2-dependent phosphorylation and ubiquitin-dependent degradation, leading to APC/C(Cdh1)-mediated Cyclin A destruction, Cyclin-dependent kinase 2 inactivation, and G1 arrest. A nonphosphorylatable CUEDC2 mutant is resistant to UV-induced degradation. Expression of this stable mutant effectively overrides UV-induced G1-S block. These results establish CUEDC2 as an APC/C(Cdh1) inhibitor and indicate that regulated CUEDC2 degradation is critical for UV-induced G1 arrest.

  5. Antitumor Efficacy of Anti-GD2 IgG1 Is Enhanced by Fc Glyco-Engineering.

    PubMed

    Xu, Hong; Guo, Hongfen; Cheung, Irene Y; Cheung, Nai-Kong V

    2016-07-01

    The affinity of therapeutic antibodies for Fcγ receptors (FcγRs) strongly influences their antitumor potency. To generate antibodies with optimal binding and immunologic efficacy, we compared the affinities of different versions of an IgG1 Fc region that had an altered peptide backbone, altered glycans, or both. To produce IgG1 with glycans that lacked α1,6-fucose, we used CHO cells that were deficient in the enzyme UDP-N-acetylglucosamine: α-3-d-mannoside-β-1,2-N-acetylglucosaminyltransferase I (GnT1), encoded by the MGAT1 gene. Mature N-linked glycans require this enzyme, and without it, CHO cells synthesize antibodies carrying only Man5-GlcNAc2, which were more effective in antibody-dependent cell-mediated cytotoxicity (ADCC). Our engineered IgG1, hu3F8-IgG1, is specific for GD2, a neuroendocrine tumor ganglioside. Its peptide mutant is IgG1-DEL (S239D/I332E/A330L), both produced in wild-type CHO cells. When produced in GnT1-deficient CHO cells, we refer to them as IgG1n and IgG1n-DEL, respectively. Affinities for human FcγRs were measured using Biacore T-100 (on CD16 and CD32 polymorphic alleles), their immunologic properties compared for ADCC and complement-mediated cytotoxicity (CMC) in vitro, and pharmacokinetics and antitumor effects were compared in vivo in humanized mice. IgG1n and IgG1n-DEL contained only mannose and acetylglucosamine and had preferential affinity for activating CD16s, over inhibitory CD32B, receptors. In vivo, the antitumor effects of IgG1, IgG1-DEL, and IgG1n-DEL were similar but modest, whereas IgG1n was significantly more effective (P < 0.05). Thus, IgG1n antibodies produced in GnT1-deficient CHO cells may have potential as improved anticancer therapeutics. Cancer Immunol Res; 4(7); 631-8. ©2016 AACR. PMID:27197064

  6. G1/ELE Functions in the Development of Rice Lemmas in Addition to Determining Identities of Empty Glumes

    PubMed Central

    Liu, Mengjia; Li, Haifeng; Su, Yali; Li, Wenqiang; Shi, Chunhai

    2016-01-01

    Rice empty glumes, also named sterile lemmas or rudimentary lemmas according to different interpretations, are distinct from lemmas in morphology and cellular pattern. Consistently, the molecular mechanism to control the development of lemmas is different from that of empty glumes. Rice LEAFY HULL STERILE1(OsLHS1) and DROOPING LEAF(DL) regulate the cellular pattern and the number of vascular bundles of lemmas respectively, while LONG STERILE LEMMA1 (G1)/ELONGATED EMPTY GLUME (ELE) and PANICLE PHYTOMER2 (PAP2)/OsMADS34 determine identities of empty glumes. Though some progress has been made, identities of empty glumes remain unclear, and genetic interactions between lemma genes and glume genes have been rarely elucidated. In this research, a new G1/ELE mutant g1–6 was identified and the phenotype was analyzed. Similar to previously reported mutant lines of G1/ELE, empty glumes of g1–6 plants transform into lemma-like organs. Furthermore, Phenotypes of single and double mutant plants suggest that, in addition to their previously described gene-specific functions, G1/ELE and OsLHS1 play redundant roles in controlling vascular bundle number, cell volume, and cell layer number of empty glumes and lemmas. Meanwhile, expression patterns of G1/ELE in osmads1-z flowers and OsLHS1 in g1–6 flowers indicate they do not regulate each other at the level of transcription. Finally, down-regulation of the empty glume gene OsMADS34/PAP2 and ectopic expression of the lemma gene DL, in the g1–6 plants provide further evidence that empty glumes are sterile lemmas. Generally, our findings provided valuable information for better understanding functions of G1 and OsLHS1 in flower development and identities of empty glumes. PMID:27462334

  7. Isolation, purification, and characterization of a thermostable xylanase from a novel strain, Paenibacillus campinasensis G1-1.

    PubMed

    Zheng, Hongchen; Liu, Yihan; Liu, Xiaoguang; Wang, Jianling; Han, Ying; Lu, Fuping

    2012-07-01

    High levels of xylanase activity (143.98 IU/ml) produced by the newly isolated Paenibacillus campinasensis G1-1 were detected when it was cultivated in a synthetic medium. A thermostable xylanase, designated XynG1-1, from P. campinasensis G1-1 was purified to homogeneity by Octyl-Sepharose hydrophobic-interaction chromatography, Sephadex G75 gel-filter chromatography, and Q-Sepharose ion-exchange chromatography, consecutively. By multistep purification, the specific activity of XynG1-1 was up to 1,865.5 IU/mg with a 9.1-fold purification. The molecular mass of purified XynG1-1 was about 41.3 kDa as estimated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Sequence analysis revealed that XynG1-1 containing 377 amino acids encoded by 1,134 bp genomic sequences of P. campinasensis G1-1 shared 96% homology with XylX from Paenibacillus campinasensis BL11 and 77%~78% homology with xylanases from Bacillus sp. YA- 335 and Bacillus sp. 41M-1, respectively. The activity of XynG1-1 was stimulated by Ca2+, Ba2+, DTT, and beta- mercaptoethanol, but was inhibited by Ni2+, Fe2+, Fe3+, Zn2+, SDS, and EDTA. The purified XynG1-1 displayed a greater affinity for birchwood xylan, with an optimal temperature of 60 degrees C and an optimal pH of 7.5. The fact that XynG1-1 is cellulose-free, thermostable (stability at high temperature of 70 degrees C~80 degrees C), and active over a wide pH range (pH 5.0~9.0) suggests that the enzyme is potentially valuable for various industrial applications, especially for pulp bleaching pretreatment.

  8. G1/ELE Functions in the Development of Rice Lemmas in Addition to Determining Identities of Empty Glumes.

    PubMed

    Liu, Mengjia; Li, Haifeng; Su, Yali; Li, Wenqiang; Shi, Chunhai

    2016-01-01

    Rice empty glumes, also named sterile lemmas or rudimentary lemmas according to different interpretations, are distinct from lemmas in morphology and cellular pattern. Consistently, the molecular mechanism to control the development of lemmas is different from that of empty glumes. Rice LEAFY HULL STERILE1(OsLHS1) and DROOPING LEAF(DL) regulate the cellular pattern and the number of vascular bundles of lemmas respectively, while LONG STERILE LEMMA1 (G1)/ELONGATED EMPTY GLUME (ELE) and PANICLE PHYTOMER2 (PAP2)/OsMADS34 determine identities of empty glumes. Though some progress has been made, identities of empty glumes remain unclear, and genetic interactions between lemma genes and glume genes have been rarely elucidated. In this research, a new G1/ELE mutant g1-6 was identified and the phenotype was analyzed. Similar to previously reported mutant lines of G1/ELE, empty glumes of g1-6 plants transform into lemma-like organs. Furthermore, Phenotypes of single and double mutant plants suggest that, in addition to their previously described gene-specific functions, G1/ELE and OsLHS1 play redundant roles in controlling vascular bundle number, cell volume, and cell layer number of empty glumes and lemmas. Meanwhile, expression patterns of G1/ELE in osmads1-z flowers and OsLHS1 in g1-6 flowers indicate they do not regulate each other at the level of transcription. Finally, down-regulation of the empty glume gene OsMADS34/PAP2 and ectopic expression of the lemma gene DL, in the g1-6 plants provide further evidence that empty glumes are sterile lemmas. Generally, our findings provided valuable information for better understanding functions of G1 and OsLHS1 in flower development and identities of empty glumes. PMID:27462334

  9. Heavy-Element Ejecta in G1.9+0.3

    NASA Astrophysics Data System (ADS)

    Borkowski, Kazimierz J.; Reynolds, S. P.; Green, D.; Hwang, U.; Petre, R.; Krishnamurthy, K.; Willett, R.

    2013-04-01

    G1.9+0.3 is the youngest known Galactic supernova remnant (SNR), with an estimated supernova (SN) explosion date of about 1900, most likely located near the Galactic center. Only the outermost ejecta layers with free-expansion velocities in excess of 18,000 km/s have been shocked so far in this dynamically-young, likely Type Ia SNR. A long (980 ks) Chandra observation in 2011 allowed for spatially-resolved spectroscopy of heavy-element ejecta. We denoised Chandra data with the spatio-spectral method of Krishnamurthy, Raginsky, & Willett, and then used a wavelet-based technique to spatially localize thermal emission produced by intermediate-mass elements (IMEs: Si, S, Ar, and Ca) and by iron. The spatial distribution of both IMEs and Fe is extremely asymmetric and inhomogeneous, with the strongest ejecta emission in the northern limb. Fe K emission is particularly prominent there, and fits with a thermal plane-shock model indicate strongly oversolar Fe abundances. In a localized, outlying region in the northern shell, IMEs are at least 5 times less abundant than Fe (by mass), indicating that undiluted Fe-group elements (including radioactive Ni) with velocities > 18,000 km/s were ejected by this SN. More modest (up to a factor of 2) Fe overabundances with respect to IMEs are present in other locations within the northern limb. There are several thousandths of a solar mass of shocked Fe in G1.9+0.3. In several locations within the remnant, including the (inner) west limb, we also find Si- and S-rich ejecta without any traces of Fe, so high-velocity, presumably undiluted products of O-burning were also ejected by the SN. If the underlying continuum is thermal, with plasma temperatures of 3-4 keV, then it must be produced by lighter elements such as O that comprise the bulk of the shocked gas. We discuss these findings in the context of Type Ia SNe such as SN 2010jn where iron-group elements at such high free-expansion velocities have been recently detected. We also

  10. Occurrence of mycotoxins in refrigerated pizza dough and risk assessment of exposure for the Spanish population.

    PubMed

    Quiles, Juan Manuel; Saladino, Federica; Mañes, Jordi; Fernández-Franzón, Mónica; Meca, Giuseppe

    2016-08-01

    Mycotoxins are toxic metabolites produced by filamentous fungi, as Aspergillus, Penicillium and Fusarium. The first objective of this research was to study the presence of mycotoxins in 60 samples of refrigerated pizza dough, by extraction with methanol and determination by liquid chromatography associated with tandem mass spectrometry (LC-MS/MS). Then, the estimated dietary intakes (EDIs) of these mycotoxins, among the Spanish population, was calculated and the health risk assessment was performed, comparing the EDIs data with the tolerable daily intake values (TDIs). The mycotoxins detected in the analyzed samples were aflatoxin B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1), zearalenone (ZEA), enniatin A (ENA), enniatin A1 (ENA1), enniatin (ENB), enniatin B1 (ENB1) and BEA (beauvericin) with average concentration of the positive samples of 4.09 μg/kg, 0.50 μg/kg, 0.79 μg/kg, 77.78 μg/kg, 14.96 μg/kg, 4.54 μg/kg, 3.37 μg/kg, 1.69 μg/kg and 22.39 μg/kg, respectively. The presence of ZEA, ENA1, ENB and ENB1 was detected in 100% of the samples, AFB2 in 32%, AFB1 in 23%, ENA in 8% and BEA in 3%. Twelve percent of the samples contaminated with AFB1 and 12% of the doughs contaminated with ZEA exceeded the EU legislated maximum limits. The dietary intakes were estimated considering three different age groups of population, and the EDIs calculated for the mycotoxins detected in the samples were all below the established TDI.

  11. Emergence and Characterization of Unusual DS-1-Like G1P[8] Rotavirus Strains in Children with Diarrhea in Thailand

    PubMed Central

    Komoto, Satoshi; Tacharoenmuang, Ratana; Guntapong, Ratigorn; Ide, Tomihiko; Haga, Kei; Katayama, Kazuhiko; Kato, Takema; Ouchi, Yuya; Kurahashi, Hiroki; Tsuji, Takao; Sangkitporn, Somchai; Taniguchi, Koki

    2015-01-01

    The emergence and rapid spread of unusual DS-1-like G1P[8] rotaviruses in Japan have been recently reported. During rotavirus surveillance in Thailand, three DS-1-like G1P[8] strains (RVA/Human-wt/THA/PCB-180/2013/G1P[8], RVA/Human-wt/THA/SKT-109/2013/G1P[8], and RVA/Human-wt/THA/SSKT-41/2013/G1P[8]) were identified in stool specimens from hospitalized children with severe diarrhea. In this study, we sequenced and characterized the complete genomes of strains PCB-180, SKT-109, and SSKT-41. On whole genomic analysis, all three strains exhibited a unique genotype constellation including both genogroup 1 and 2 genes: G1-P[8]-I2-R2-C2-M2-A2-N2-T2-E2-H2. This novel genotype constellation is shared with Japanese DS-1-like G1P[8] strains. Phylogenetic analysis revealed that the G/P genes of strains PCB-180, SKT-109, and SSKT-41 appeared to have originated from human Wa-like G1P[8] strains. On the other hand, the non-G/P genes of the three strains were assumed to have originated from human DS-1-like strains. Thus, strains PCB-180, SKT-109, and SSKT-41 appeared to be derived through reassortment event(s) between Wa-like G1P[8] and DS-1-like human rotaviruses. Furthermore, strains PCB-180, SKT-109, and SSKT-41 were found to have the 11-segment genome almost indistinguishable from one another in their nucleotide sequences and phylogenetic lineages, indicating the derivation of the three strains from a common origin. Moreover, all the 11 genes of the three strains were closely related to those of Japanese DS-1-like G1P[8] strains. Therefore, DS-1-like G1P[8] strains that have emerged in Thailand and Japan were assumed to have originated from a recent common ancestor. To our knowledge, this is the first report on whole genome-based characterization of DS-1-like G1P[8] strains that have emerged in an area other than Japan. Our observations will provide important insights into the evolutionary dynamics of emerging DS-1-like G1P[8] rotaviruses. PMID:26540260

  12. Fangchinoline induces G1 arrest in breast cancer cells through cell-cycle regulation.

    PubMed

    Xing, Zhibo; Zhang, Youxue; Zhang, Xianyu; Yang, Yanmei; Ma, Yuyan; Pang, Da

    2013-12-01

    Fangchinoline, an alkaloid derived from the dry roots of Stephaniae tetrandrine S. Moore (Menispermaceae), has been shown to possess cytotoxic, anti-inflammatory, and antioxidant properties. In this study, we used Fangchinoline to inhibit breast cancer cell proliferation and to investigate its underlying molecular mechanisms. Human breast cancer cell lines, MCF-7 and MDA-MB-231, were both used in this study. We found that Fangchinoline significantly decreased cell proliferation in a dose-dependent manner and induced G1-phase arrest in both cell lines. In addition, upon analysis of expression of cell cycle-related proteins, we found that Fangchinoline reduced expression of cyclin D1, cyclin D3, and cyclin E, and increased expression of the cyclin-dependent kinase (CDK) inhibitors, p21/WAF1, and p27/KIP1. Moreover, Fangchinoline also inhibited the kinase activities of CDK2, CDK4, and CDK6. These results suggest that Fangchinoline can inhibit human breast cancer cell proliferation and thus may have potential applications in cancer therapy. PMID:23401195

  13. 3D structural fluctuation of IgG1 antibody revealed by individual particle electron tomography

    DOE PAGES

    Zhang, Xing; Zhang, Lei; Tong, Huimin; Peng, Bo; Rames, Matthew J.; Zhang, Shengli; Ren, Gang

    2015-05-05

    Commonly used methods for determining protein structure, including X-ray crystallography and single-particle reconstruction, often provide a single and unique three-dimensional (3D) structure. However, in these methods, the protein dynamics and flexibility/fluctuation remain mostly unknown. Here, we utilized advances in electron tomography (ET) to study the antibody flexibility and fluctuation through structural determination of individual antibody particles rather than averaging multiple antibody particles together. Through individual-particle electron tomography (IPET) 3D reconstruction from negatively-stained ET images, we obtained 120 ab-initio 3D density maps at an intermediate resolution (~1–3 nm) from 120 individual IgG1 antibody particles. Using these maps as a constraint, wemore » derived 120 conformations of the antibody via structural flexible docking of the crystal structure to these maps by targeted molecular dynamics simulations. Statistical analysis of the various conformations disclosed the antibody 3D conformational flexibility through the distribution of its domain distances and orientations. This blueprint approach, if extended to other flexible proteins, may serve as a useful methodology towards understanding protein dynamics and functions.« less

  14. Long noncoding RNA linc00598 regulates CCND2 transcription and modulates the G1 checkpoint.

    PubMed

    Jeong, Oh-Seok; Chae, Yun-Cheol; Jung, Hyeonsoo; Park, Soon Cheol; Cho, Sung-Jin; Kook, Hyun; Seo, SangBeom

    2016-01-01

    Data derived from genomic and transcriptomic analyses have revealed that long noncoding RNAs (lncRNAs) have important roles in the transcriptional regulation of various genes. Recent studies have identified the mechanism underlying this function. To date, a variety of noncoding transcripts have been reported to function in conjunction with epigenetic regulator proteins. In this study, we investigated the function of linc00598, which is transcribed by a genomic sequence on chromosome 13, downstream of FoxO1 and upstream of COG6. Microarray analysis showed that linc00598 regulates the transcription of specific target genes, including those for cell cycle regulators. We discovered that linc00598 regulates CCND2 transcription through modulation of the transcriptional regulatory effect of FoxO1 on the CCND2 promoter. Moreover, we observed that knockdown of linc00598 induced G0/G1 cell cycle arrest and inhibited proliferation. These data indicate that linc00598 plays an important role in cell cycle regulation and proliferation through its ability to regulate the transcription of CCND2. PMID:27572135

  15. A gradient in the duration of the G1 phase in the murine neocortical proliferative epithelium

    NASA Technical Reports Server (NTRS)

    Miyama, S.; Takahashi, T.; Nowakowski, R. S.; Caviness, V. S. Jr

    1997-01-01

    Neuronogenesis in the neocortical pseudostratified ventricular epithelium (PVE) is initiated rostrolaterally and progresses caudo-medially as development progresses. Here we have measured the cytokinetic parameters and the fractional neuronal output parameter, Q, of laterally located early-maturing regions over the principal embryonic days (E12-E15) of neocortical neuronogenesis in the mouse. These measures are compared with ones previously made of a medial, late-maturing portion of the PVE. Laterally, as medially, the duration of the neuronogenetic interval is 6 days and comprises 11 integer cell cycles. Also, in both lateral and medial areas the length of G1 phase (TG1) increases nearly 4-fold and is the only cell cycle parameter to change. Q progresses essentially identically laterally and medially with respect to the succession of integer cell cycles. Most importantly, from E12 to E13 there is a steeply declining lateral to medial gradient in TG1. The gradient is due both to the lateral to medial graded stage of neuronogenesis and to the stepwise increase in TG1 with each integer cycle during the neuronogenetic interval. To our knowledge this gradient in TG1 of the cerebral PVE is the first cell biological gradient to be demonstrated experimentally in such an extensive proliferative epithelial sheet. We suggest that this gradient in TG1 is the cellular mechanism for positionally encoding a protomap of the neocortex within the PVE.

  16. Biosimilarity Assessments of Model IgG1-Fc Glycoforms Using a Machine Learning Approach.

    PubMed

    Kim, Jae Hyun; Joshi, Sangeeta B; Tolbert, Thomas J; Middaugh, C Russell; Volkin, David B; Smalter Hall, Aaron

    2016-02-01

    Biosimilarity assessments are performed to decide whether 2 preparations of complex biomolecules can be considered "highly similar." In this work, a machine learning approach is demonstrated as a mathematical tool for such assessments using a variety of analytical data sets. As proof-of-principle, physical stability data sets from 8 samples, 4 well-defined immunoglobulin G1-Fragment crystallizable glycoforms in 2 different formulations, were examined (see More et al., companion article in this issue). The data sets included triplicate measurements from 3 analytical methods across different pH and temperature conditions (2066 data features). Established machine learning techniques were used to determine whether the data sets contain sufficient discriminative power in this application. The support vector machine classifier identified the 8 distinct samples with high accuracy. For these data sets, there exists a minimum threshold in terms of information quality and volume to grant enough discriminative power. Generally, data from multiple analytical techniques, multiple pH conditions, and at least 200 representative features were required to achieve the highest discriminative accuracy. In addition to classification accuracy tests, various methods such as sample space visualization, similarity analysis based on Euclidean distance, and feature ranking by mutual information scores are demonstrated to display their effectiveness as modeling tools for biosimilarity assessments.

  17. G1 phase regulation, area-specific cell cycle control, and cytoarchitectonics in the primate cortex

    PubMed Central

    Lukaszewicz, Agnès; Savatier, Pierre; Cortay, Véronique; Giroud, Pascale; Huissoud, Cyril; Berland, Michel; Kennedy, Henry; Dehay, Colette

    2005-01-01

    We have investigated the cell-cycle related mechanisms that lead to the emergence of the primate areas 17 and 18. These areas are characterized by striking differences in cytoarchitectonics and neuron number. We show in vivo that (i) area 17 precursors of supragranular neurons exhibit a shorter cell-cycle duration, a reduced G1 phase and a higher rate of cell-cycle re-entry than area 18 precursors (ii) area-specific levels of expression of Cyclin E (high in area 17, low in area 18) and p27Kip1 (low in area 17, high in area 18) (iii) Ex vivo up and down modulation of Cyclin E and p27Kip1 show that both regulators influence cell-cycle kinetics by modifying rates of cell-cycle progression and cell-cycle re-entry (iv) Modeling the areal differences in cell-cycle parameters suggests that they contribute to areal differences in numbers of precursors and neuron production. PMID:16055060

  18. Long noncoding RNA linc00598 regulates CCND2 transcription and modulates the G1 checkpoint

    PubMed Central

    Jeong, Oh-Seok; Chae, Yun-Cheol; Jung, Hyeonsoo; Park, Soon Cheol; Cho, Sung-Jin; Kook, Hyun; Seo, SangBeom

    2016-01-01

    Data derived from genomic and transcriptomic analyses have revealed that long noncoding RNAs (lncRNAs) have important roles in the transcriptional regulation of various genes. Recent studies have identified the mechanism underlying this function. To date, a variety of noncoding transcripts have been reported to function in conjunction with epigenetic regulator proteins. In this study, we investigated the function of linc00598, which is transcribed by a genomic sequence on chromosome 13, downstream of FoxO1 and upstream of COG6. Microarray analysis showed that linc00598 regulates the transcription of specific target genes, including those for cell cycle regulators. We discovered that linc00598 regulates CCND2 transcription through modulation of the transcriptional regulatory effect of FoxO1 on the CCND2 promoter. Moreover, we observed that knockdown of linc00598 induced G0/G1 cell cycle arrest and inhibited proliferation. These data indicate that linc00598 plays an important role in cell cycle regulation and proliferation through its ability to regulate the transcription of CCND2. PMID:27572135

  19. Bacillus amyloliquefaciens G1: A Potential Antagonistic Bacterium against Eel-Pathogenic Aeromonas hydrophila

    PubMed Central

    Cao, Haipeng; He, Shan; Wei, Ruopeng; Diong, Marek; Lu, Liqun

    2011-01-01

    Recent studies have revealed that the use of probiotics is an alternative to control marine aeromonas. However, few probiotics are available against Aeromonas hydrophila infections in eels. In the present study, a potential antagonistic strain G1 against the eel-pathogenic A. hydrophila was isolated from sediment underlying brackish water. Its extracellular products with antibacterial activities were shown to be stable under wide range of pH, temperature, and proteinase K. It was initially identified as Bacillus amyloliquefaciens using API identification kits and confirmed to be B. amyloliquefaciens strain (GenBank accession number DQ422953) by phylogenetic analysis. In addition, it was shown to be safe for mammalians, had a wide anti-A. hydrophila spectrum, and exhibited significant effects on inhibiting the growth of the eel-pathogenic A. hydrophila both in vitro and in vivo. To the best of our knowledge, this is the first report on a promising antagonistic Bacillus amyloliquefaciens strain from brackish water sediment against eel-pathogenic A. hydrophila. PMID:21754944

  20. Screening of recombinant glycosyltransferases reveals the broad acceptor specificity of stevia UGT-76G1.

    PubMed

    Dewitte, Griet; Walmagh, Maarten; Diricks, Margo; Lepak, Alexander; Gutmann, Alexander; Nidetzky, Bernd; Desmet, Tom

    2016-09-10

    UDP-glycosyltransferases (UGTs) are a promising class of biocatalysts that offer a sustainable alternative for chemical glycosylation of natural products. In this study, we aimed to characterize plant-derived UGTs from the GT-1 family with an emphasis on their acceptor promiscuity and their potential application in glycosylation processes. Recombinant expression in E. coli provided sufficient amounts of enzyme for the in-depth characterization of the salicylic acid UGT from Capsella rubella (UGT-SACr) and the stevia UGT from Stevia rebaudiana (UGT-76G1Sr). The latter was found to have a remarkably broad specificity with activities on a wide diversity of structures, from aliphatic and branched alcohols, over small phenolics to larger flavonoids, terpenoids and even higher glycoside compounds. As an example for its industrial potential, the glycosylation of curcumin was thoroughly evaluated. Under optimized conditions, 96% of curcumin was converted within 24h into the corresponding curcumin β-glycosides. In addition, the reaction was performed in a coupled system with sucrose synthase from Glycine max, to enable the cost-efficient (re)generation of UDP-Glc from sucrose as abundant and renewable resource.

  1. Screening of recombinant glycosyltransferases reveals the broad acceptor specificity of stevia UGT-76G1.

    PubMed

    Dewitte, Griet; Walmagh, Maarten; Diricks, Margo; Lepak, Alexander; Gutmann, Alexander; Nidetzky, Bernd; Desmet, Tom

    2016-09-10

    UDP-glycosyltransferases (UGTs) are a promising class of biocatalysts that offer a sustainable alternative for chemical glycosylation of natural products. In this study, we aimed to characterize plant-derived UGTs from the GT-1 family with an emphasis on their acceptor promiscuity and their potential application in glycosylation processes. Recombinant expression in E. coli provided sufficient amounts of enzyme for the in-depth characterization of the salicylic acid UGT from Capsella rubella (UGT-SACr) and the stevia UGT from Stevia rebaudiana (UGT-76G1Sr). The latter was found to have a remarkably broad specificity with activities on a wide diversity of structures, from aliphatic and branched alcohols, over small phenolics to larger flavonoids, terpenoids and even higher glycoside compounds. As an example for its industrial potential, the glycosylation of curcumin was thoroughly evaluated. Under optimized conditions, 96% of curcumin was converted within 24h into the corresponding curcumin β-glycosides. In addition, the reaction was performed in a coupled system with sucrose synthase from Glycine max, to enable the cost-efficient (re)generation of UDP-Glc from sucrose as abundant and renewable resource. PMID:27378621

  2. 3D structural fluctuation of IgG1 antibody revealed by individual particle electron tomography

    SciTech Connect

    Zhang, Xing; Zhang, Lei; Tong, Huimin; Peng, Bo; Rames, Matthew J.; Zhang, Shengli; Ren, Gang

    2015-05-05

    Commonly used methods for determining protein structure, including X-ray crystallography and single-particle reconstruction, often provide a single and unique three-dimensional (3D) structure. However, in these methods, the protein dynamics and flexibility/fluctuation remain mostly unknown. Here, we utilized advances in electron tomography (ET) to study the antibody flexibility and fluctuation through structural determination of individual antibody particles rather than averaging multiple antibody particles together. Through individual-particle electron tomography (IPET) 3D reconstruction from negatively-stained ET images, we obtained 120 ab-initio 3D density maps at an intermediate resolution (~1–3 nm) from 120 individual IgG1 antibody particles. Using these maps as a constraint, we derived 120 conformations of the antibody via structural flexible docking of the crystal structure to these maps by targeted molecular dynamics simulations. Statistical analysis of the various conformations disclosed the antibody 3D conformational flexibility through the distribution of its domain distances and orientations. This blueprint approach, if extended to other flexible proteins, may serve as a useful methodology towards understanding protein dynamics and functions.

  3. c-myc can induce expression of G0/G1 transition genes.

    PubMed Central

    Schweinfest, C W; Fujiwara, S; Lau, L F; Papas, T S

    1988-01-01

    The human c-myc oncogene was linked to the heat shock-inducible Drosophila hsp70 promoter and used to stably transfect mouse BALB/c 3T3 cells. Heat shock of the transfectants at 42 degrees C followed by recovery at 37 degrees C resulted in the appearance of the human c-myc protein which was appropriately localized to the nuclear fraction. Two-dimensional analysis of the proteins of density-arrested cells which had been heat shock treated revealed the induction of eight protein species and the repression of five protein species. All of the induced and repressed proteins were nonabundant. cDNA clones corresponding to genes induced during the G0/G1 transition were used as probes to assay for c-myc inducibility of these genes. Two anonymous sequences previously identified as serum inducible (3CH77 and 3CH92) were induced when c-myc was expressed. In response to serum stimulation, 3CH77 and 3CH92 were expressed before c-myc mRNA levels increased. However, in response to specific induction of c-myc by heat shock of serum arrested cells, 3CH77 and 3CH92 mRNA levels increased after the rise in c-myc mRNA. Therefore, we hypothesize that abnormal expression of c-myc can induce genes involved in the proliferative response. Images PMID:3211137

  4. First passage times in M2[X ]|G |1 |R queue with hysteretic overload control policy

    NASA Astrophysics Data System (ADS)

    Pechinkin, Alexander V.; Razumchik, Rostislav R.; Zaryadov, Ivan S.

    2016-06-01

    One of the reported approaches towards the solution of overload problem in networks of SIP servers is the implementation of multi-level hysteretic control of arrivals in SIP servers. Each level, being the parameter of the policy, specifies operation mode of SIP server i.e. it implicitly indicates what SIP server must do with the arriving packets. The choice of parameters' values is not guided by standards and is usually left for the network owner. In general, all operation modes of the considered policy can be grouped into two groups: normal mode (when all arriving packets are accepted) and congested mode (when part or all arriving packets are being dropped). Such grouping may serve as the criteria for choosing parameters' values of the policy: pick those values which minimize SIP server sojourn time in congested mode. In this short note we propose some analytical results which facilitate the solution of stated minimization problem. The considered mathematical model of SIP server is the queueing system M2[X ]|G |1 |R with batch arrivals and bi-level hysteretic control policy, which specifies three operation modes: normal (customers both flows are accepted), overload (only customers from one flow are accepted), discard (customers from both flows are blocked/lost)). The switching between modes can occur only on service completions. Analytical method allowing computation of stationary sojourn times in different operation modes (as well as first passage times between modes) is presented in brief. Numerical example is given.

  5. Whole genomic analysis of human G1P[8] rotavirus strains from different age groups in China.

    PubMed

    Shintani, Tsuzumi; Ghosh, Souvik; Wang, Yuan-Hong; Zhou, Xuan; Zhou, Dun-Jin; Kobayashi, Nobumichi

    2012-08-01

    G1P[8] rotaviruses are an important cause of diarrhea in humans in China. To date, there are no reports on the whole genomic analysis of the Chinese G1P[8] rotaviruses. To determine the origin and overall genetic makeup of the recent Chinese G1P[8] strains, the whole genomes of three strains, RVA/Human-wt/CHN/E1911/2009/G1P[8], RVA/Human-tc/CHN/R588/2005/G1P[8] and RVA/Human-tc/CHN/Y128/2004/G1P[8], detected in an infant, a child and an adult, respectively, were analyzed. Strains E1911, R588 and Y128 exhibited a typical Wa-like genotype constellation. Except for the NSP3 gene of E1911, the whole genomes of strains E1911, R588 and Y128 were found to be more closely related to those of the recent Wa-like common human strains from different countries than those of the prototype G1P[8] strain, or other old strains. On the other hand, the NSP3 gene of E1911 was genetically distinct from those of Y128, R588, or other Wa-like common human strains, and appeared to share a common origin with those of the porcine-like human G9 strains, providing evidence for intergenotype reassortment events. Comparisons of the amino acid residues defining the VP7 and VP4 antigenic domains revealed several mismatches between these Chinese G1P[8] strains and the G1 and P[8] strains contained in the currently licensed rotavirus vaccines Rotarix(TM )and RotaTeq(TM). PMID:23012626

  6. The impact of geoengineering on vegetation in experiment G1 of the Geoengineering Model Intercomparison Project

    NASA Astrophysics Data System (ADS)

    Irvine, Peter; Glienke, Susanne; Lawrence, Mark

    2015-04-01

    Solar Radiation Management (SRM) has been proposed as a means to partly counteract global warming. The Geoengineering Model Intercomparison Project (GeoMIP) simulated the climate consequences of a number of SRM techniques, but the effects on vegetation have not yet been thoroughly studied. Here, the vegetation response to the idealized GeoMIP G1 experiment is analyzed, in which a reduction of the solar constant counterbalances the radiative effects of quadrupled atmospheric CO2 concentrations; the results from eight fully coupled earth system models (ESMs) are included. For most models and regions, changes in net primary productivity (NPP) are dominated by the increase in CO2, via the CO2 fertilization effect. As SRM will lower temperatures, in high latitudes this will reverse gains in NPP from the lifting of temperature limitations. In low latitudes this cooling relative to the 4xCO2 simulation decreases plant respiration whilst having little effect on gross primary productivity, increasing NPP. Despite reductions in precipitation in most regions in response to SRM, runoff and NPP increase in many regions including those previously highlighted as potentially being at risk of drought under SRM. This is due to simultaneous reductions in evaporation and increases in water use efficiency by plants due to higher CO2 concentrations. The relative differences between models in the vegetation response are substantially larger than the differences in their climate responses. The largest differences between models are for those with and without a nitrogen-cycle, with a much smaller CO2 fertilization effect for the former. These results suggest that until key vegetation processes are integrated into ESM predictions, the vegetation response to SRM will remain highly uncertain.

  7. Increased Expression of Bcl11b Leads to Chemoresistance Accompanied by G1 Accumulation

    PubMed Central

    Grabarczyk, Piotr; Nähse, Viola; Delin, Martin; Przybylski, Grzegorz; Depke, Maren; Hildebrandt, Petra; Völker, Uwe; Schmidt, Christian A.

    2010-01-01

    Background The expression of BCL11B was reported in T-cells, neurons and keratinocytes. Aberrations of BCL11B locus leading to abnormal gene transcription were identified in human hematological disorders and corresponding animal models. Recently, the elevated levels of Bcl11b protein have been described in a subset of squameous cell carcinoma cases. Despite the rapidly accumulating knowledge concerning Bcl11b biology, the contribution of this protein to normal or transformed cell homeostasis remains open. Methodology/Principal Findings Here, by employing an overexpression strategy we revealed formerly unidentified features of Bcl11b. Two different T-cell lines were forced to express BCL11B at levels similar to those observed in primary T-cell leukemias. This resulted in markedly increased resistance to radiomimetic drugs while no influence on death-receptor apoptotic pathway was observed. Apoptosis resistance triggered by BCL11B overexpression was accompanied by a cell cycle delay caused by accumulation of cells at G1. This cell cycle restriction was associated with upregulation of CDKN1C (p57) and CDKN2C (p18) cyclin dependent kinase inhibitors. Moreover, p27 and p130 proteins accumulated and the SKP2 gene encoding a protein of the ubiquitin-binding complex responsible for their degradation was repressed. Furthermore, the expression of the MYCN oncogene was silenced which resulted in significant depletion of the protein in cells expressing high BCL11B levels. Both cell cycle restriction and resistance to DNA-damage-induced apoptosis coincided and required the histone deacetylase binding N-terminal domain of Bcl11b. The sensitivity to genotoxic stress could be restored by the histone deacetylase inhibitor trichostatine A. Conclusions The data presented here suggest a potential role of BCL11B in tumor survival and encourage developing Bcl11b-inhibitory approaches as a potential tool to specifically target chemoresistant tumor cells. PMID:20824091

  8. Induction of G1 cell cycle arrest and apoptosis by berberine in bladder cancer cells.

    PubMed

    Yan, Keqiang; Zhang, Cheng; Feng, Jinbo; Hou, Lifang; Yan, Lei; Zhou, Zunlin; Liu, Zhaoxu; Liu, Cheng; Fan, Yidon; Zheng, Baozhong; Xu, Zhonghua

    2011-07-01

    Bladder cancer is the ninth most common type of cancer, and its surgery is always followed by chemotherapy to prevent recurrence. Berberine is non-toxic to normal cells but has anti-cancer effects in many cancer cell lines. This study was aimed to determine whether berberine inhibits the cell proliferation and induces cell cycle arrest and apoptosis in BIU-87 and T24 bladder cancer cell line. The superficial bladder cancer cell line BIU-87 and invasive T24 bladder cancer cells were treated with different concentrations of berberine. MTT assay was used to determine the effects of berberine on the viability of these cells. The cell cycle arrest was detected through propidium iodide (PI) staining. The induction of apoptosis was determined through Annexin V-conjugated Alexa Fluor 488 (Alexa488) staining. Berberine inhibited the viability of BIU-87 and T24 cells in a dose- and time-dependent manner. It also promoted cell cycle arrest at G0/G1 in a dose-dependent manner and induced apoptosis. We observed that H-Ras and c-fos mRNA and protein expressionswere dose-dependently and time-dependently decreased by berberine treatment. Also, we investigated the cleaved caspase-3 and caspase-9 protein expressions increased in a dose-dependent manner. Berberine inhibits the cell proliferation and induces cell cycle arrest and apoptosis in BIU-87, bladder cancer cell line and T24, invasive bladder cancer cell line. Berberine can inhibit the oncogentic H-Ras and c-fos in T24 cells, and can induce the activation of the caspase-3 and caspase-9 apoptosis. Therefore, berberine has the potential to be a novel chemotherapy drug to treat the bladder cancer by suppressing tumor growth.

  9. Nicotine overrides DNA damage-induced G1/S restriction in lung cells.

    PubMed

    Nishioka, Takashi; Yamamoto, Daisuke; Zhu, Tongbo; Guo, Jinjin; Kim, Sung-Hoon; Chen, Chang Yan

    2011-01-01

    As an addictive substance, nicotine has been suggested to facilitate pro-survival activities (such as anchorage-independent growth or angiogenesis) and the establishment of drug resistance to anticancer therapy. Tobacco smoking consists of a variety of carcinogens [such as benzopyrene (BP) and nitrosamine derivatives] that are able to cause DNA double strand breaks. However, the effect of nicotine on DNA damage-induced checkpoint response induced by genotoxins remains unknown. In this study, we investigated the events occurred during G(1) arrest induced by γ-radiation or BP in nicotine-treated murine or human lung epithelial cells. DNA synthesis was rapidly inhibited after exposure to γ-radiation or BP treatment, accompanied with the activation of DNA damage checkpoint. When these cells were co-treated with nicotine, the growth restriction was compromised, manifested by upregulation of cyclin D and A, and attenuation of Chk2 phosphorylation. Knockdown of cyclin D or Chk2 by the siRNAs blocked nicotine-mediated effect on DNA damage checkpoint activation. However, nicotine treatment appeared to play no role in nocodazole-induced mitotic checkpoint activation. Overall, our study presented a novel observation, in which nicotine is able to override DNA damage checkpoint activated by tobacco-related carcinogen BP or γ-irradiation. The results not only indicates the potentially important role of nicotine in facilitating the establishment of genetic instability to promote lung tumorigenesis, but also warrants a dismal prognosis for cancer patients who are smokers, heavily exposed second-hand smokers or nicotine users. PMID:21559516

  10. Downregulation of CREB Promotes Cell Proliferation by Mediating G1/S Phase Transition in Hodgkin Lymphoma.

    PubMed

    Lu, Fangjin; Zheng, Ying; Donkor, Paul Owusu; Zou, Peng; Mu, Ping

    2016-01-01

    The cyclic-AMP response element-binding protein (CREB), a well-known nuclear transcription factor, has been shown to play an essential role in many cellular processes, including differentiation, cell survival, and cell proliferation, by regulating the expression of downstream genes. Recently, increased expression of CREB was frequently found in various tumors, indicating that CREB is implicated in the process of tumorigenesis. However, the effects of CREB on Hodgkin lymphoma (HL) remain unknown. To clarify the role of CREB in HL, we performed knockdown experiments in HL. We found that downregulation of CREB by short hairpin RNA (shRNA) resulted in enhancement of cell proliferation and promotion of G1/S phase transition, and these effects can be rescued by expression of shRNA-resistant CREB. Meanwhile, the expression level of cell cycle-related proteins, such as cyclin D1, cyclin E1, cyclin-dependent kinase 2 (CDK2), and CDK4, was elevated in response to depletion of CREB. Furthermore, we performed chromatin immunoprecipitation (ChIP) assay and confirmed that CREB directly bound to the promoter regions of these genes, which consequently contributed to the regulation of cell cycle. Consistent with our results, a clinical database showed that high expression of CREB correlates with favorable prognosis in B-cell lymphoma patients, which is totally different from the function of CREB in other cancers such as colorectal cancer, acute myeloid leukemia, and some endocrine cancers. Taken together, all of these features of CREB in HL strongly support its role as a tumor suppressor gene that can decelerate cell proliferation by inhibiting the expression of several cell cycle-related genes. Our results provide new evidence for prognosis prediction of HL and a promising therapeutic strategy for HL patients. PMID:27458098

  11. Aflatoxins, ochratoxins and zearalenone in sorghum and sorghum products in Sudan.

    PubMed

    Elbashir, Abdalla A; Ali, Salah Eldeen A

    2014-01-01

    This survey examined 60 samples of sorghum and 30 samples of sorghum products from three states (Khartoum, Kordofan and Gadarif) of Sudan for aflatoxin B1, B2, G1 and G2 (AFB1, AFB2, AFG1, AFG2), ochratoxin A and B (OTA, OTB) and zearalenone (ZEN), using high performance liquid chromatography with fluorescence detection. The limits of detection and limits of quantification were in the range 0.01-0.6 µg kg(-1) and 0.03-2.0 µg kg(-1), respectively. The frequency of contaminated samples with AFB1 from Khartoum, Gadarif and Kordofan state was 38.1%, 22.2% and 23.8%, respectively. Only two samples of sorghum from Khartoum state were contaminated with OTA (3.3%). Concentrations of OTA and OTB were low and may not cause problems. No sample of sorghum or sorghum products was contaminated with ZEN. Some sorghum samples contained AFB1 concentrations above the European Union regulatory limits. The highest contaminated samples were found in Khartoum state.

  12. Estimated exposure to EU regulated mycotoxins and risk characterization of aflatoxin-induced hepatic toxicity through the consumption of the toasted cereal flour called "gofio", a traditional food of the Canary Islands (Spain).

    PubMed

    Luzardo, Octavio P; Bernal-Suárez, María Del Mar; Camacho, María; Henríquez-Hernández, Luis Alberto; Boada, Luis D; Rial-Berriel, Cristian; Almeida-González, Maira; Zumbado, Manuel; Díaz-Díaz, Ricardo

    2016-07-01

    "Gofio" is a type of flour made from toasted grain, which is part of the staple food in the Canary Islands, Spain, in which the occurrence of Aflatoxins B1, B2, G1 and G2 (AFB1, AFB2, AFG1, AFG2), Fumonisins B1 and B2 (FB1 and FB2) Ochratoxin A (OTA), Deoxynivalenol (DNV) and Zearalenone (ZEA) was evaluated. 83% of the samples were contaminated with at least one mycotoxin and 69.2% of the analyzed samples showed co-occurrence of mycotoxins (range 2 to 8). All the concentrations were well below the established limits (maximum values of AFs=0.42 μg/kg; FBs=178.3 μg/kg; OTA=0.3 μg/kg; DON=92.5 μg/kg; and ZEA=9.9 μg/kg). The daily dietary exposure to total AFs was estimated to be 7.1% of the TDI. This value was almost double in children, and considering the upper-bound approach could reach 35% of the TDI. For the rest of mycotoxins, the consumers would be exposed to less than 2% of their TDIs. The risk characterization indicates that there is a potential risk in developing aflatoxin induced liver cancer due to gofio consumption in the subpopulation which is simultaneously exposed to other hepatocarcinogens, such as the hepatitis B virus. PMID:27132021

  13. Aflatoxin levels in chronic hepatitis B patients with cirrhosis or hepatocellular carcinoma in Balıkesir, Turkey.

    PubMed

    Aydın, M; Aydın, S; Bacanlı, M; Başaran, N

    2015-11-01

    Aflatoxins, the secondary metabolites produced by species of naturally occurring Aspergilli, are commonly found in food such as cereals, dried fruits and juice, wine, beer and spices. They are hepatotoxic and are well known human carcinogens based on evidence from human studies. Aflatoxins are an environmental risk factor for the development of hepatocellular carcinoma (HCC). Chronic hepatitis B-infected patients are at increased risk of cirrhosis, hepatic failure and liver cancer. This study was designed to determine the serum aflatoxin B1 (AFB1 ), aflatoxin B2 (AFB2 ), aflatoxin G1 (AFG1 ) and aflatoxin G2 (AFG2 ) concentrations using high-pressure liquid chromatography (HPLC) in hepatitis B-infected patients with or without cirrhosis and liver cancer, alongside healthy controls in Balıkesir, Turkey. The mean AFB1 and total AF levels in patients without liver cancer and cirrhosis were significantly higher than healthy controls. The mean AFB1 and total AF levels in patients with chronic hepatitis B and HCC were significantly higher than infected patients with or without cirrhosis. These results suggest that patients with chronic hepatitis B who are exposed to AFs are at increased risk for developing HCC, which might be prevented by reducing consumption of contaminated foods.

  14. The potential effects of Zataria multiflora Boiss essential oil on growth, aflatoxin production and transcription of aflatoxin biosynthesis pathway genes of toxigenic Aspergillus parasiticus

    PubMed Central

    Yahyaraeyat, R.; Khosravi, A.R.; Shahbazzadeh, D.; Khalaj, V.

    2013-01-01

    This study aims at evaluating the effects of Zataria multiflora (Z. multiflora) essential oil (EO) on growth, aflatoxin production and transcription of aflatoxin biosynthesis pathway genes. Total RNAs of Aspergillus parasiticus (A.parasiticus) ATCC56775 grown in yeast extract sucrose (YES) broth medium treated with Z. multiflora EO were subjected to reverse transcription- polymerase chain reaction (RT-PCR). Specific primers of nor-1, ver-1, omt-A and aflR genes were used. In parallel mycelial dry weight of samples were measured and all the media were assayed by high-pressure liquid chromatography (HPLC) for aflatoxinB1 (AFB1), aflatoxinB2 (AFB2), aflatoxinG1 (AFG1), aflatoxinG2 (AFG2) and aflatoxin total (AFTotal) production. The results showed that mycelial dry weight and aflatoxin production reduce in the presence of Z. multiflora EO (100 ppm) on day 5 of growth. It was found that the expression of nor-1, ver-1, omt-A and aflR genes was correlated with the ability of fungus to produce aflatoxins on day 5 in YES medium. RT-PCR showed that in the presence of Z.multiflora EO (100 ppm) nor-1, ver-1 and omtA genes expression was reduced. It seems that toxin production inhibitory effects of Z. multiflora EO on day 5 may be at the transcription level and this herb may cause reduction in aflatoxin biosynthesis pathway genes activity. PMID:24294264

  15. Activity in mice of recombinant BCG-EgG1Y162 vaccine for Echinococcus granulosus infection.

    PubMed

    Ma, Xiumin; Zhao, Hui; Zhang, Fengbo; Zhu, Yuejie; Peng, Shanshan; Ma, Haimei; Cao, Chunbao; Xin, Yan; Yimiti, Delixiati; Wen, Hao; Ding, Jianbing

    2016-01-01

    Cystic hydatid disease is a zoonotic parasitic disease caused by Echinococcus granulosus which is distributed worldwide. The disease is difficult to treat with surgery removal is the only cure treatment. In the high endemic areas, vaccination of humans is believed a way to protect communities from the disease. In this study we vaccinated BALB/c mice with rBCG-EgG1Y162, and then detected the level of IgG and IgE specifically against the recombinant protein by ELISA, rBCG-EgG1Y162 induced strong and specific cellular and humoral immune responses. In vitro study showed that rBCG-EgG1Y162 vaccine not only promote splenocytes proliferation but also active T cell. In addition, the rBCG-EgG1Y162 induced a protection in the mice against secondary infection of Echinococcus granulosus.

  16. Activity in mice of recombinant BCG-EgG1Y162 vaccine for Echinococcus granulosus infection

    PubMed Central

    Ma, Xiumin; Zhao, Hui; Zhang, Fengbo; Zhu, Yuejie; Peng, Shanshan; Ma, Haimei; Cao, Chunbao; Xin, Yan; Yimiti, Delixiati; Wen, Hao; Ding, Jianbing

    2016-01-01

    Cystic hydatid disease is a zoonotic parasitic disease caused by Echinococcus granulosus which is distributed worldwide. The disease is difficult to treat with surgery removal is the only cure treatment. In the high endemic areas, vaccination of humans is believed a way to protect communities from the disease. In this study we vaccinated BALB/c mice with rBCG-EgG1Y162, and then detected the level of IgG and IgE specifically against the recombinant protein by ELISA, rBCG-EgG1Y162 induced strong and specific cellular and humoral immune responses. In vitro study showed that rBCG-EgG1Y162 vaccine not only promote splenocytes proliferation but also active T cell. In addition, the rBCG-EgG1Y162 induced a protection in the mice against secondary infection of Echinococcus granulosus. PMID:26266551

  17. Review of geochemical reference sample programs since G-1 and W-1: progress to date and remaining challenges

    USGS Publications Warehouse

    Kane, J.S.

    1991-01-01

    A brief history of programs to develop geochemical reference samples and certified reference samples for use in geochemical analysis is presented. While progress has been made since G-1 and W-1 were issued, many challenges remain. ?? 1991.

  18. Galactosylation of IgG1 modulates FcγRIIB-mediated inhibition of murine autoimmune hemolytic anemia.

    PubMed

    Yamada, Kazunori; Ito, Kiyoaki; Furukawa, Jun-Ichi; Nakata, Junichiro; Alvarez, Montserrat; Verbeek, J Sjef; Shinohara, Yasuro; Izui, Shozo

    2013-12-01

    Murine immune effector cells express three different stimulatory FcγRs (FcγRI, FcγRIII and FcγRIV) and one inhibitory receptor, FcγRIIB. Competitive engagement of stimulatory and inhibitory FcγRs has been shown to be critical for the development of immune complex-mediated inflammatory disorders. Because of the previous demonstration that FcγRIIB was unable to inhibit FcγRIII-mediated autoimmune hemolytic anemia induced by 105-2H IgG1 anti-RBC mAb, we reevaluated the regulatory role of FcγRIIB on the development of anemia using two additional IgG1 anti-RBC mAbs (34-3C and 3H5G1) and different 34-3C IgG subclass-switch variants. We were able to induce a more severe anemia in FcγRIIB-deficient mice than in FcγRIIB-sufficient mice after injection of 34-3C and 3H5G1 IgG1, but not 105-2H IgG1. Structural analysis of N-linked oligosaccharides attached to the CH2 domain revealed that 105-2H was poorly galactosylated as compared with the other mAbs, while the extent of sialylation was comparable between all mAbs. In addition, we observed that a more galactosylated 105-2H variant provoked more severe anemia in FcγRIIB-deficient mice than FcγRIIB-sufficient mice. In contrast, the development of anemia induced by three non-IgG1 subclass variants of the 34-3C mAb was not down-regulated by FcγRIIB, although they were more galactosylated than its IgG1 variant. These data indicate that FcγRIIB-mediated inhibition of autoimmune hemolytic anemia is restricted to the IgG1 subclass and that galactosylation, but not sialylation, of IgG1 (but not other IgG subclasses) is critical for the interaction with FcγR, thereby determining the pathogenic potential of IgG1 autoantibodies.

  19. Fc Engineering of Human IgG1 for Altered Binding to the Neonatal Fc Receptor Affects Fc Effector Functions.

    PubMed

    Grevys, Algirdas; Bern, Malin; Foss, Stian; Bratlie, Diane Bryant; Moen, Anders; Gunnarsen, Kristin Støen; Aase, Audun; Michaelsen, Terje Einar; Sandlie, Inger; Andersen, Jan Terje

    2015-06-01

    Engineering of the constant Fc part of monoclonal human IgG1 (hIgG1) Abs is an approach to improve effector functions and clinical efficacy of next-generation IgG1-based therapeutics. A main focus in such development is tailoring of in vivo half-life and transport properties by engineering the pH-dependent interaction between IgG and the neonatal Fc receptor (FcRn), as FcRn is the main homeostatic regulator of hIgG1 half-life. However, whether such engineering affects binding to other Fc-binding molecules, such as the classical FcγRs and complement factor C1q, has not been studied in detail. These effector molecules bind to IgG1 in the lower hinge-CH2 region, structurally distant from the binding site for FcRn at the CH2-CH3 elbow region. However, alterations of the structural composition of the Fc may have long-distance effects. Indeed, in this study we show that Fc engineering of hIgG1 for altered binding to FcRn also influences binding to both the classical FcγRs and complement factor C1q, which ultimately results in alterations of cellular mechanisms such as Ab-dependent cell-mediated cytotoxicity, Ab-dependent cellular phagocytosis, and Ab-dependent complement-mediated cell lysis. Thus, engineering of the FcRn-IgG1 interaction may greatly influence effector functions, which has implications for the therapeutic efficacy and use of Fc-engineered hIgG1 variants.

  20. Fc Engineering of Human IgG1 for Altered Binding to the Neonatal Fc Receptor Affects Fc Effector Functions

    PubMed Central

    Grevys, Algirdas; Bern, Malin; Foss, Stian; Bratlie, Diane Bryant; Moen, Anders; Gunnarsen, Kristin Støen; Aase, Audun; Michaelsen, Terje Einar; Sandlie, Inger

    2015-01-01

    Engineering of the constant Fc part of monoclonal human IgG1 (hIgG1) Abs is an approach to improve effector functions and clinical efficacy of next-generation IgG1-based therapeutics. A main focus in such development is tailoring of in vivo half-life and transport properties by engineering the pH-dependent interaction between IgG and the neonatal Fc receptor (FcRn), as FcRn is the main homeostatic regulator of hIgG1 half-life. However, whether such engineering affects binding to other Fc-binding molecules, such as the classical FcγRs and complement factor C1q, has not been studied in detail. These effector molecules bind to IgG1 in the lower hinge–CH2 region, structurally distant from the binding site for FcRn at the CH2–CH3 elbow region. However, alterations of the structural composition of the Fc may have long-distance effects. Indeed, in this study we show that Fc engineering of hIgG1 for altered binding to FcRn also influences binding to both the classical FcγRs and complement factor C1q, which ultimately results in alterations of cellular mechanisms such as Ab-dependent cell-mediated cytotoxicity, Ab-dependent cellular phagocytosis, and Ab-dependent complement-mediated cell lysis. Thus, engineering of the FcRn–IgG1 interaction may greatly influence effector functions, which has implications for the therapeutic efficacy and use of Fc-engineered hIgG1 variants. PMID:25904551

  1. Whi5 phosphorylation embedded in the G1/S network dynamically controls critical cell size and cell fate

    PubMed Central

    Palumbo, Pasquale; Vanoni, Marco; Cusimano, Valerio; Busti, Stefano; Marano, Francesca; Manes, Costanzo; Alberghina, Lilia

    2016-01-01

    In budding yeast, overcoming of a critical size to enter S phase and the mitosis/mating switch—two central cell fate events—take place in the G1 phase of the cell cycle. Here we present a mathematical model of the basic molecular mechanism controlling the G1/S transition, whose major regulatory feature is multisite phosphorylation of nuclear Whi5. Cln3–Cdk1, whose nuclear amount is proportional to cell size, and then Cln1,2–Cdk1, randomly phosphorylate both decoy and functional Whi5 sites. Full phosphorylation of functional sites releases Whi5 inhibitory activity, activating G1/S transcription. Simulation analysis shows that this mechanism ensures coherent release of Whi5 inhibitory action and accounts for many experimentally observed properties of mitotically growing or conjugating G1 cells. Cell cycle progression and transcriptional analyses of a Whi5 phosphomimetic mutant verify the model prediction that coherent transcription of the G1/S regulon and ensuing G1/S transition requires full phosphorylation of Whi5 functional sites. PMID:27094800

  2. DNA resection proteins Sgs1 and Exo1 are required for G1 checkpoint activation in budding yeast

    PubMed Central

    Balogun, Fiyinfolu O.; Truman, Andrew W.; Kron, Stephen J.

    2013-01-01

    Double-strand breaks (DSBs) in budding yeast trigger activation of DNA damage checkpoints, allowing repair to occur. Although resection is necessary for initiating damage-induced cell cycle arrest in G2, no role has been assigned to it in the activation of G1 checkpoint. Here we demonstrate for the first time that the resection proteins Sgs1 and Exo1 are required for efficient G1 checkpoint activation. We find in G1 arrested cells that histone H2A phosphorylation in response to ionizing radiation is independent of Sgs1 and Exo1. In contrast, these proteins are required for damage-induced recruitment of Rfa1 to the DSB sites, phosphorylation of the Rad53 effector kinase, cell cycle arrest and RNR3 expression. Checkpoint activation in G1 requires the catalytic activity of Sgs1, suggesting that it is DNA resection mediated by Sgs1 that stimulates the damage response pathway rather than protein-protein interactions with other DDR proteins. Together, these results implicate DNA resection, which is thought to be minimal in G1, as necessary for activation of the G1 checkpoint. PMID:23835406

  3. Characterization of a G1 inhibitor from old JB-1 ascites tumor fluid. Interaction with polyions and ion exchangers.

    PubMed

    Barfod, N M; Bichel, P

    1976-09-17

    In most experimental ascites tumors the growth rate decreases with increasing age and cell number. This decrease is caused by a prolongation of the cell cycle and an increasing accumulation of noncycling cells in resting (or quiescent) G1 and G2 compartments. In cell-free ascitic fluid from the JB-1 ascites tumor in the plateau phase of growth, low molecular weight substances have been found which reversibly and specifically arrest JB-1 cells in G1 and G2. In order to characterize the JB-1 G1 inhibitor we have investigated the effect of ion exchangers and polyions on the activity of this inhibitor assayed in vitro by means of a partially synchronized JB-1 cell population analyzed by flow microfluorometry. The results indicate that polyanions and cation exchangers (immobilized polyanions) bind and abolish the G1-inhibitory activity. From this it is suggested that the G1 inhibitor is of a basic or polycationic nature. Since anion exchangers (immobilized polycations) are without effect on this activity it was surprising to find that polycations also neutralize the activity. The results indicate that this occurs by blocking an anionic G2-inhibitor receptor on the cell, thus preventing the polycationic G1 inhibitor from being bound to this receptor.

  4. Canonical Wnt activity regulates trunk neural crest delamination linking BMP/noggin signaling with G1/S transition.

    PubMed

    Burstyn-Cohen, Tal; Stanleigh, Jonathan; Sela-Donenfeld, Dalit; Kalcheim, Chaya

    2004-11-01

    Delamination of premigratory neural crest cells depends on a balance between BMP/noggin and on successful G1/S transition. Here, we report that BMP regulates G1/S transition and consequent crest delamination through canonical Wnt signaling. Noggin overexpression inhibits G1/S transition and blocking G1/S abrogates BMP-induced delamination; moreover, transcription of Wnt1 is stimulated by BMP and by the developing somites, which concomitantly inhibit noggin production. Interfering with beta-catenin and LEF/TCF inhibits G1/S transition, neural crest delamination and transcription of various BMP-dependent genes, which include Cad6B, Pax3 and Msx1, but not that of Slug, Sox9 or FoxD3. Hence, we propose that developing somites inhibit noggin transcription in the dorsal tube, resulting in activation of BMP and consequent Wnt1 production. Canonical Wnt signaling in turn stimulates G1/S transition and generation of neural crest cell motility independently of its proposed role in earlier neural crest specification. PMID:15456730

  5. Whi5 phosphorylation embedded in the G1/S network dynamically controls critical cell size and cell fate.

    PubMed

    Palumbo, Pasquale; Vanoni, Marco; Cusimano, Valerio; Busti, Stefano; Marano, Francesca; Manes, Costanzo; Alberghina, Lilia

    2016-01-01

    In budding yeast, overcoming of a critical size to enter S phase and the mitosis/mating switch--two central cell fate events--take place in the G1 phase of the cell cycle. Here we present a mathematical model of the basic molecular mechanism controlling the G1/S transition, whose major regulatory feature is multisite phosphorylation of nuclear Whi5. Cln3-Cdk1, whose nuclear amount is proportional to cell size, and then Cln1,2-Cdk1, randomly phosphorylate both decoy and functional Whi5 sites. Full phosphorylation of functional sites releases Whi5 inhibitory activity, activating G1/S transcription. Simulation analysis shows that this mechanism ensures coherent release of Whi5 inhibitory action and accounts for many experimentally observed properties of mitotically growing or conjugating G1 cells. Cell cycle progression and transcriptional analyses of a Whi5 phosphomimetic mutant verify the model prediction that coherent transcription of the G1/S regulon and ensuing G1/S transition requires full phosphorylation of Whi5 functional sites. PMID:27094800

  6. Netrin-G1 regulates fear-like and anxiety-like behaviors in dissociable neural circuits

    PubMed Central

    Zhang, Qi; Sano, Chie; Masuda, Akira; Ando, Reiko; Tanaka, Mika; Itohara, Shigeyoshi

    2016-01-01

    In vertebrate mammals, distributed neural circuits in the brain are involved in emotion-related behavior. Netrin-G1 is a glycosyl-phosphatidylinositol-anchored synaptic adhesion molecule whose deficiency results in impaired fear-like and anxiety-like behaviors under specific circumstances. To understand the cell type and circuit specificity of these responses, we generated netrin-G1 conditional knockout mice with loss of expression in cortical excitatory neurons, inhibitory neurons, or thalamic neurons. Genetic deletion of netrin-G1 in cortical excitatory neurons resulted in altered anxiety-like behavior, but intact fear-like behavior, whereas loss of netrin-G1 in inhibitory neurons resulted in attenuated fear-like behavior, but intact anxiety-like behavior. These data indicate a remarkable double dissociation of fear-like and anxiety-like behaviors involving netrin-G1 in excitatory and inhibitory neurons, respectively. Our findings support a crucial role for netrin-G1 in dissociable neural circuits for the modulation of emotion-related behaviors, and provide genetic models for investigating the mechanisms underlying the dissociation. The results also suggest the involvement of glycosyl-phosphatidylinositol-anchored synaptic adhesion molecules in the development and pathogenesis of emotion-related behavior. PMID:27345935

  7. Binding of fusion protein FLSC IgG1 to CCR5 is enhanced by CCR5 antagonist Maraviroc.

    PubMed

    Latinovic, Olga; Schneider, Kate; Szmacinski, Henryk; Lakowicz, Joseph R; Heredia, Alonso; Redfield, Robert R

    2014-12-01

    The CCR5 chemokine receptor is crucial for human immunodeficiency virus type 1 (HIV-1) infection, acting as the principal coreceptor for HIV-1 entry and transmission and is thus an attractive target for antiviral therapy. Studies have suggested that CCR5 surface density and its conformational changes subsequent to virion engagement are rate limiting for entry, and consequently, infection. Not all CCR5 antibodies inhibit HIV-1 infection, suggesting a need for more potent reagents. Here we evaluated full length single chain (FLSC) IgG1, a novel IgG-CD4-gp120(BAL) fusion protein with several characteristics that make it an attractive candidate for treatment of HIV-1 infections, including bivalency and a potentially increased serum half-life over FLSC, the parental molecule. FLSC IgG1 binds two domains on CCR5, the N-terminus and the second extracellular loop, lowering the levels of available CCR5 viral attachment sites. Furthermore, FLSC IgG1 synergizes with Maraviroc (MVC), the only licensed CCR5 antagonist. In this study, we used both microscopy and functional assays to address the mechanistic aspects of the interactions of FLSC IgG1 and MVC in the context of CCR5 conformational changes and viral infection. We used a novel stochastic optical reconstruction microscopy (STORM), based on high resolution localization of photoswitchable dyes to visualize direct contacts between FLSC IgG1 and CCR5. We compared viral entry inhibition by FLSC IgG1 with that of other CCR5 blockers and showed FLSC IgG1 to be the most potent. We also showed that lower CCR5 surface densities in HIV-1 infected primary cells result in lower FLSC IgG1 EC50 values. In addition, CCR5 binding by FLSC IgG1, but not CCR5 Ab 2D7, was significantly increased when cells were treated with MVC, suggesting MVC allosterically increases exposure of the FLSC IgG1 binding site. These data have implications for future antiviral therapy development.

  8. Mitotic phosphorylation of eukaryotic initiation factor 4G1 (eIF4G1) at Ser1232 by Cdk1:cyclin B inhibits eIF4A helicase complex binding with RNA.

    PubMed

    Dobrikov, Mikhail I; Shveygert, Mayya; Brown, Michael C; Gromeier, Matthias

    2014-02-01

    During mitosis, global translation is suppressed, while synthesis of proteins with vital mitotic roles must go on. Prior evidence suggests that the mitotic translation shift involves control of initiation. Yet, no signals specifically targeting translation initiation factors during mitosis have been identified. We used phosphoproteomics to investigate the central translation initiation scaffold and "ribosome adaptor," eukaryotic initiation factor 4G1 (eIF4G1) in interphase or nocodazole-arrested mitotic cells. This approach and kinase inhibition assays, in vitro phosphorylation with recombinant kinase, and kinase depletion-reconstitution experiments revealed that Ser1232 in eIF4G1 is phosphorylated by cyclin-dependent kinase 1 (Cdk1):cyclin B during mitosis. Ser1232 is located in an unstructured region of the C-terminal portion of eIF4G1 that coordinates assembly of the eIF4G/-4A/-4B helicase complex and binding of the mitogen-activated protein kinase (MAPK) signal-integrating kinase, Mnk. Intense phosphorylation of Ser1232 in mitosis strongly enhanced the interactions of eIF4A with HEAT domain 2 of eIF4G and decreased association of eIF4G/-4A with RNA. Our findings implicate phosphorylation of eIF4G1(Ser1232) by Cdk1:cyclin B and its inhibitory effects on eIF4A helicase activity in the mitotic translation initiation shift.

  9. Staurosporine is chemoprotective by inducing G1 arrest in a Chk1- and pRb-dependent manner.

    PubMed

    Murray, Mollianne McGahren; Bui, Tuyen; Smith, Michelle; Bagheri-Yarmand, Rozita; Wingate, Hannah; Hunt, Kelly K; Keyomarsi, Khandan

    2013-10-01

    Chemotherapeutic agents have been the mainstay of cancer therapy for years. However, their effectiveness has been limited by toxicities they impart on normal cells. Staurosporine (ST) has been shown to arrest normal, but not breast cancer, cells in G1. Therefore, ST may become a chemoprotective agent, arresting normal cells while allowing tumor cells to enter cell cycle phases where they are sensitive to chemotherapeutic agents. Understanding the mechanism of ST-mediated G1 arrest may allow for a beneficial chemoprotective treatment strategy for patients. We utilized 76NE6 (pRb+/p53-), 76NF2V (pRb+/p53+) and 76NE7 (pRb-/P53+) non-tumorigenic human mammary epithelial cell lines to understand the role of the Rb and p53 pathways in ST-directed G1 arrest. CDK4 was downregulated by ST in Rb+ cells, but its presence could not reverse the arrest, neither did its stable downregulation alter ST-mediated cellular response. ST-mediated G1 arrest required pRb, which in turn initiated a cascade of events leading to inhibition of CDK4. Further assessment of this pathway revealed that Chk1 expression and activity were required for the Rb-dependent arrest. For example, pRb+ cells with small interfering RNA to Chk1 had approximately 60% less cells in G1 phase compared with controls and pRb- cells do not arrest upon ST. Furthermore, Chk1 expression facilitates the release of the Rb+ cells from G1 arrest. Collectively, our data suggest that pRb cooperates with Chk1 to mediate a G1 arrest only in pRb+ cells. The elucidation of this pathway can help identify novel agents to protect cancer patients against the debilitating effects of chemotherapy.

  10. Reduced vasorelaxation to estradiol and G-1 in aged female and adult male rats is associated with GPR30 downregulation.

    PubMed

    Lindsey, Sarah H; da Silva, Ariel S; Silva, Mauro S; Chappell, Mark C

    2013-07-01

    Previously, we reported that chronic activation of the estrogen receptor GPR30 by its selective agonist G-1 decreases blood pressure in ovariectomized hypertensive mRen2.Lewis (mRen2) rats but not intact male littermates. Furthermore, G-1 relaxes female mesenteric resistance arteries via both endothelium-dependent and -independent mechanisms. Because of the lack of a blood pressure-lowering effect by G-1 in males and the potential influence of aging on estrogen receptor expression, we hypothesized that GPR30-dependent vasodilation and receptor expression are altered in males and aged females. Thus, we assessed the response to 17β-estradiol or G-1 in mesenteric arteries obtained from 15-wk-old normotensive Lewis and hypertensive mRen2 females and males as well as 52-wk-old Lewis females. Vasodilation to 17β-estradiol (E₂) and G-1 was significantly attenuated in 15-wk-old Lewis and mRen2 males compared with age-matched females. Pretreatment of male vessels with the nitric oxide synthase inhibitor L-NAME had no significant effect on the estradiol or G-1 response. In aged females, E₂ and G-1 vasorelaxation was also significantly blunted; however, L-NAME essentially abolished the response. Associated with the reduced vascular responses, GPR30 expression in mesenteric arteries was approximately 50% lower in males and aged females compared with young females. We conclude that alterations in GPR30 expression and signaling may contribute to vascular dysfunction in aging females and a greater blood pressure in hypertensive males.

  11. Isoform-specific toxicity of Mecp2 in postmitotic neurons: suppression of neurotoxicity by FoxG1.

    PubMed

    Dastidar, Somasish Ghosh; Bardai, Farah H; Ma, Chi; Price, Valerie; Rawat, Varun; Verma, Pragya; Narayanan, Vinodh; D'Mello, Santosh R

    2012-02-22

    The methyl-CpG binding protein 2 (MeCP2) is a widely expressed protein, the mutations of which cause Rett syndrome. The level of MeCP2 is highest in the brain where it is expressed selectively in mature neurons. Its functions in postmitotic neurons are not known. The MeCP2 gene is alternatively spliced to generate two proteins with different N termini, designated as MeCP2-e1 and MeCP2-e2. The physiological significance of these two isoforms has not been elucidated, and it is generally assumed they are functionally equivalent. We report that in cultured cerebellar granule neurons induced to die by low potassium treatment and in Aβ-treated cortical neurons, Mecp2-e2 expression is upregulated whereas expression of the Mecp2-e1 isoform is downregulated. Knockdown of Mecp2-e2 protects neurons from death, whereas knockdown of the e1 isoform has no effect. Forced expression of MeCP2-e2, but not MeCP2-e1, promotes apoptosis in otherwise healthy neurons. We find that MeCP2-e2 interacts with the forkhead protein FoxG1, mutations of which also cause Rett syndrome. FoxG1 has been shown to promote neuronal survival and its downregulation leads to neuronal death. We find that elevated FoxG1 expression inhibits MeCP2-e2 neurotoxicity. MeCP2-e2 neurotoxicity is also inhibited by IGF-1, which prevents the neuronal death-associated downregulation of FoxG1 expression, and by Akt, the activation of which is necessary for FoxG1-mediated neuroprotection. Finally, MeCP2-e2 neurotoxicity is enhanced if FoxG1 expression is suppressed or in neurons cultured from FoxG1-haplodeficient mice. Our results indicate that Mecp2-e2 promotes neuronal death and that this activity is normally inhibited by FoxG1. Reduced FoxG1 expression frees MecP2-e2 to promote neuronal death. PMID:22357867

  12. Cell cycle transition from S-phase to G1 in Caulobacter is mediated by ancestral virulence regulators

    PubMed Central

    Fumeaux, Coralie; Radhakrishnan, Sunish Kumar; Ardissone, Silvia; Théraulaz, Laurence; Frandi, Antonio; Martins, Daniel; Nesper, Jutta; Abel, Sören; Jenal, Urs; Viollier, Patrick H.

    2014-01-01

    Zinc-finger domain transcriptional regulators regulate a myriad of functions in eukaryotes. Interestingly, ancestral versions (MucR) from Alpha-proteobacteria control bacterial virulence/symbiosis. Whether virulence regulators can also control cell cycle transcription is unknown. Here we report that MucR proteins implement a hitherto elusive primordial S→G1 transcriptional switch. After charting G1-specific promoters in the cell cycle model Caulobacter crescentus by comparative ChIP-seq, we use one such promoter as genetic proxy to unearth two MucR paralogs, MucR1/2, as constituents of a quadripartite and homeostatic regulatory module directing the S→G1 transcriptional switch. Surprisingly, MucR orthologues that regulate virulence and symbiosis gene transcription in Brucella, Agrobacterium or Sinorhizobium support this S→G1 switch in Caulobacter. Pan-genomic ChIP-seq analyses in Sinorhizobium and Caulobacter show that this module indeed targets orthologous genes. We propose that MucR proteins and possibly other virulence regulators primarily control bacterial cell cycle (G1-phase) transcription, rendering expression of target (virulence) genes periodic and in tune with the cell cycle. PMID:24939058

  13. Copy Number Variation and Transposable Elements Feature in Recent, Ongoing Adaptation at the Cyp6g1 Locus

    PubMed Central

    Schmidt, Joshua M.; Good, Robert T.; Appleton, Belinda; Sherrard, Jayne; Raymant, Greta C.; Bogwitz, Michael R.; Martin, Jon; Daborn, Phillip J.; Goddard, Mike E.; Batterham, Philip; Robin, Charles

    2010-01-01

    The increased transcription of the Cyp6g1 gene of Drosophila melanogaster, and consequent resistance to insecticides such as DDT, is a widely cited example of adaptation mediated by cis-regulatory change. A fragment of an Accord transposable element inserted upstream of the Cyp6g1 gene is causally associated with resistance and has spread to high frequencies in populations around the world since the 1940s. Here we report the existence of a natural allelic series at this locus of D. melanogaster, involving copy number variation of Cyp6g1, and two additional transposable element insertions (a P and an HMS-Beagle). We provide evidence that this genetic variation underpins phenotypic variation, as the more derived the allele, the greater the level of DDT resistance. Tracking the spatial and temporal patterns of allele frequency changes indicates that the multiple steps of the allelic series are adaptive. Further, a DDT association study shows that the most resistant allele, Cyp6g1-[BP], is greatly enriched in the top 5% of the phenotypic distribution and accounts for ∼16% of the underlying phenotypic variation in resistance to DDT. In contrast, copy number variation for another candidate resistance gene, Cyp12d1, is not associated with resistance. Thus the Cyp6g1 locus is a major contributor to DDT resistance in field populations, and evolution at this locus features multiple adaptive steps occurring in rapid succession. PMID:20585622

  14. Molecular Dynamics Simulation of the Crystallizable Fragment of IgG1—Insights for the Design of Fcabs

    PubMed Central

    Lai, Balder; Hasenhindl, Christoph; Obinger, Christian; Oostenbrink, Chris

    2014-01-01

    An interesting format in the development of therapeutic monoclonal antibodies uses the crystallizable fragment of IgG1 as starting scaffold. Engineering of its structural loops allows generation of an antigen binding site. However, this might impair the molecule’s conformational stability, which can be overcome by introducing stabilizing point mutations in the CH3 domains. These point mutations often affect the stability and unfolding behavior of both the CH2 and CH3 domains. In order to understand this cross-talk, molecular dynamics simulations of the domains of the Fc fragment of human IgG1 are reported. The structure of human IgG1-Fc obtained from X-ray crystallography is used as a starting point for simulations of the wild-type protein at two different pH values. The stabilizing effect of a single point mutation in the CH3 domain as well as the impact of the hinge region and the glycan tree structure connected to the CH2 domains is investigated. Regions of high local flexibility were identified as potential sites for engineering antigen binding sites. Obtained data are discussed with respect to the available X-ray structure of IgG1-Fc, directed evolution approaches that screen for stability and use of the scaffold IgG1-Fc in the design of antigen binding Fc proteins. PMID:24451126

  15. Expanding the phenotype associated with FOXG1 mutations and in vivo FoxG1 chromatin-binding dynamics.

    PubMed

    De Filippis, R; Pancrazi, L; Bjørgo, K; Rosseto, A; Kleefstra, T; Grillo, E; Panighini, A; Cardarelli, F; Meloni, I; Ariani, F; Mencarelli, M A; Hayek, J; Renieri, A; Costa, M; Mari, F

    2012-10-01

    Mutations in the Forkhead box G1 (FOXG1) gene, a brain specific transcriptional factor, are responsible for the congenital variant of Rett syndrome. Until now FOXG1 point mutations have been reported in 12 Rett patients. Recently seven additional patients have been reported with a quite homogeneous severe phenotype designated as the FOXG1 syndrome. Here we describe two unrelated patients with a de novo FOXG1 point mutation, p.Gln46X and p.Tyr400X, respectively, having a milder phenotype and sharing a distinctive facial appearance. Although FoxG1 action depends critically on its binding to chromatin, very little is known about the dynamics of this process. Using fluorescence recovery after photobleaching, we showed that most of the GFP-FoxG1 fusion protein associates reversibly to chromatin whereas the remaining fraction is bound irreversibly. Furthermore, we showed that the two pathologic derivatives of FoxG1 described in this paper present a dramatic alteration in chromatin affinity and irreversibly bound fraction in comparison with Ser323fsX325 mutant (associated with a severe phenotype) and wild type Foxg1 protein. Our observations suggest that alterations in the kinetics of FoxG1 binding to chromatin might contribute to the pathological effects of FOXG1 mutations.

  16. The spin structure function g1p of the proton and a test of the Bjorken sum rule

    NASA Astrophysics Data System (ADS)

    Adolph, C.; Akhunzyanov, R.; Alexeev, M. G.; Alexeev, G. D.; Amoroso, A.; Andrieux, V.; Anosov, V.; Austregesilo, A.; Azevedo, C.; Badełek, B.; Balestra, F.; Barth, J.; Baum, G.; Beck, R.; Bedfer, Y.; Bernhard, J.; Bicker, K.; Bielert, E. R.; Birsa, R.; Bisplinghoff, J.; Bodlak, M.; Boer, M.; Bordalo, P.; Bradamante, F.; Braun, C.; Bressan, A.; Büchele, M.; Burtin, E.; Capozza, L.; Chang, W.-C.; Chiosso, M.; Choi, I.; Chung, S. U.; Cicuttin, A.; Crespo, M. L.; Curiel, Q.; Dalla Torre, S.; Dasgupta, S. S.; Dasgupta, S.; Denisov, O. Yu.; Dhara, L.; Donskov, S. V.; Doshita, N.; Duic, V.; Dziewiecki, M.; Efremov, A.; Eversheim, P. D.; Eyrich, W.; Ferrero, A.; Finger, M.; Finger, M.; Fischer, H.; Franco, C.; du Fresne von Hohenesche, N.; Friedrich, J. M.; Frolov, V.; Fuchey, E.; Gautheron, F.; Gavrichtchouk, O. P.; Gerassimov, S.; Giordano, F.; Gnesi, I.; Gorzellik, M.; Grabmüller, S.; Grasso, A.; Grosse-Perdekamp, M.; Grube, B.; Grussenmeyer, T.; Guskov, A.; Haas, F.; Hahne, D.; von Harrach, D.; Hashimoto, R.; Heinsius, F. H.; Herrmann, F.; Hinterberger, F.; Horikawa, N.; d'Hose, N.; -Yu Hsieh, C.; Huber, S.; Ishimoto, S.; Ivanov, A.; Ivanshin, Yu.; Iwata, T.; Jahn, R.; Jary, V.; Jörg, P.; Joosten, R.; Kabuß, E.; Ketzer, B.; Khaustov, G. V.; Khokhlov, Yu. A.; Kisselev, Yu.; Klein, F.; Klimaszewski, K.; Koivuniemi, J. H.; Kolosov, V. N.; Kondo, K.; Königsmann, K.; Konorov, I.; Konstantinov, V. F.; Kotzinian, A. M.; Kouznetsov, O.; Krämer, M.; Kremser, P.; Krinner, F.; Kroumchtein, Z. V.; Kuchinski, N.; Kunne, F.; Kurek, K.; Kurjata, R. P.; Lednev, A. A.; Lehmann, A.; Levillain, M.; Levorato, S.; Lichtenstadt, J.; Longo, R.; Maggiora, A.; Magnon, A.; Makins, N.; Makke, N.; Mallot, G. K.; Marchand, C.; Martin, A.; Marzec, J.; Matousek, J.; Matsuda, H.; Matsuda, T.; Meshcheryakov, G.; Meyer, W.; Michigami, T.; Mikhailov, Yu. V.; Miyachi, Y.; Nagaytsev, A.; Nagel, T.; Nerling, F.; Neyret, D.; Nikolaenko, V. I.; Novy, J.; Nowak, W.-D.; Nunes, A. S.; Olshevsky, A. G.; Orlov, I.; Ostrick, M.; Panzieri, D.; Parsamyan, B.; Paul, S.; Peng, J.-C.; Pereira, F.; Pesek, M.; Peshekhonov, D. V.; Platchkov, S.; Pochodzalla, J.; Polyakov, V. A.; Pretz, J.; Quaresma, M.; Quintans, C.; Ramos, S.; Regali, C.; Reicherz, G.; Riedl, C.; Rocco, E.; Rossiyskaya, N. S.; Ryabchikov, D. I.; Rychter, A.; Samoylenko, V. D.; Sandacz, A.; Santos, C.; Sarkar, S.; Savin, I. A.; Sbrizzai, G.; Schiavon, P.; Schmidt, K.; Schmieden, H.; Schönning, K.; Schopferer, S.; Selyunin, A.; Shevchenko, O. Yu.; Silva, L.; Sinha, L.; Sirtl, S.; Slunecka, M.; Sozzi, F.; Srnka, A.; Stolarski, M.; Sulc, M.; Suzuki, H.; Szabelski, A.; Szameitat, T.; Sznajder, P.; Takekawa, S.; ter Wolbeek, J.; Tessaro, S.; Tessarotto, F.; Thibaud, F.; Tosello, F.; Tskhay, V.; Uhl, S.; Veloso, J.; Virius, M.; Weisrock, T.; Wilfert, M.; Windmolders, R.; Zaremba, K.; Zavertyaev, M.; Zemlyanichkina, E.; Ziembicki, M.; Zink, A.

    2016-02-01

    New results for the double spin asymmetry A1p and the proton longitudinal spin structure function g1p are presented. They were obtained by the COMPASS Collaboration using polarised 200 GeV muons scattered off a longitudinally polarised NH3 target. The data were collected in 2011 and complement those recorded in 2007 at 160 GeV, in particular at lower values of x. They improve the statistical precision of g1p (x) by about a factor of two in the region x ≲ 0.02. A next-to-leading order QCD fit to the g1 world data is performed. It leads to a new determination of the quark spin contribution to the nucleon spin, ΔΣ, ranging from 0.26 to 0.36, and to a re-evaluation of the first moment of g1p. The uncertainty of ΔΣ is mostly due to the large uncertainty in the present determinations of the gluon helicity distribution. A new evaluation of the Bjorken sum rule based on the COMPASS results for the non-singlet structure function g1NS (x ,Q2) yields as ratio of the axial and vector coupling constants |gA /gV | = 1.22 ± 0.05 (stat.) ± 0.10 (syst.), which validates the sum rule to an accuracy of about 9%.

  17. Impacts, Effectiveness and Regional Inequalities of the GeoMIP G1 to G4 Solar Radiation Management Scenarios

    SciTech Connect

    Yu, Xiaoyong; Moore, John; Cui, Xuefeng; Rinke, Annette; Ji, Duoying; Kravitz, Benjamin S.; Yoon, Jin-Ho

    2015-06-01

    We evaluate the regional effectiveness of solar radiation management (SRM) to compensate for simultaneous changes in temperature and precipitation induced by increased greenhouse gas concentrations. We analyze results from multiple earth system models under four Geoengineering Model Intercomparison Project(GeoMIP) experiments with a modified form of the Residual Climate Response approach. Under the solar dimming geoengineering experiments G1(4xCO2) and G2(increasing CO2 by 1% per year), global average temperature is successfully restored to pre-industrial level over 50 years simulations. However, these two SRM experiments also produce a robust global precipitation decrease. The stratospheric aerosol GeoMIP geoengineering experiment, G4 has significantly greater regional inequality and lower effectiveness for compensating temperature change than G1 and G2. G4 also has significantly larger regional inequality for compensating precipitation change than G1and G2. However, there is no significant difference between precipitation change compensation effectiveness of G4 and G2, though there is much larger across model variability in G4 results. G3 has significant greater regional inequality for compensating temperature change than G1 and G2, and has significant lower effectiveness than G1. The effectiveness of four SRMs to compensate for temperature change is much higher than for precipitation. The large cross-model variation in adjustment percentage of compensated SAT and precipitation change by SRM to achieve optimal compensation effectiveness shed a light on the uncertainty accumulation effect in optimizing compensation effectiveness of SRM.

  18. Expression of the NS5 (VPg) Protein of Murine Norovirus Induces a G1/S Phase Arrest

    PubMed Central

    Davies, Colin; Ward, Vernon K.

    2016-01-01

    Murine norovirus-1 (MNV-1) is known to subvert host cell division inducing an accumulation of cells in the G0/G1 phase, creating conditions where viral replication is favored. This study identified that NS5 (VPg), is capable of inducing cell cycle arrest in the absence of viral replication or other viral proteins in an analogous manner to MNV-1 infection. NS5 expression induced an accumulation of cells in the G0/G1 phase in an asynchronous population by inhibiting progression at the G1/S restriction point. Furthermore, NS5 expression resulted in a down-regulation of cyclin A expression in asynchronous cells and inhibited cyclin A expression in cells progressing from G1 to S phase. The activity of NS5 on the host cell cycle occurs through an uncharacterized function. Amino acid substitutions of NS5(Y26A) and NS5(F123A) that inhibit the ability for NS5 to attach to RNA and recruit host eukaryotic translation initiation factors, respectively, retained the ability to induce an accumulation of cells in the G0/G1 phase as identified for wild-type NS5. To the best of our knowledge, this is the first report of a VPg protein manipulating the host cell cycle. PMID:27556406

  19. HLA-DR-dependent variation of intrathecal IgG1 (Gm) allotype synthesis in multiple sclerosis.

    PubMed

    Salier, J P; Martin-Mondiere, C; Sesboüé, R; Daveau, M; Goust, J M; Govaerts, A; Schuller, E; Degos, J D

    1985-03-01

    Genetic susceptibility to multiple sclerosis (MS) in Caucasians was previously shown to be correlated to the presence of given alleles at the HLA-DR and Gm loci. We now demonstrate that the humoral immune response in MS central nervous system (CNS) is modulated by both loci: the levels of IgG1 subclass and IgG1 allotypes in cerebrospinal fluid of MS patients depend on both their Gm genotype and their HLA-DR2 or HLA-DR7 phenotype. That HLA-DR molecules may either participate in a preferential recruitment of IgG1 allotype-producing B cells in MS CNS or act after such a selective homing is discussed. These results demonstrate that both HLA and Gm loci are synergistically involved in the modulation of the humoral immune response. PMID:3855432

  20. Precision measurements of g1 of the proton and the deuteron with 6 GeV electrons

    SciTech Connect

    Prok, Yelena; Bosted, Peter; Kvaltine, Nicholas; Adhikari, Krishna; Adikaram-Mudiyanselage, Dasuni; Aghasyan, Mher; Amaryan, Moskov; Anderson, Mark; Anefalos Pereira, Sergio; Avagyan, Harutyun; Baghdasaryan, Hovhannes; Ball, Jacques; Baltzell, Nathan; Battaglieri, Marco; Biselli, Angela; Bono, Jason; Briscoe, William; Brock, Joseph; Brooks, William; Bueltmann, Stephen; Burkert, Volker; Carlin, Christopher; Carman, Daniel; Celentano, Andrea; Chandavar, Shloka; Colaneri, Luca; Cole, Philip; Contalbrigo, Marco; Cortes, Olga; Crabb, Donald; Crede, Volker; D'Angelo, Annalisa; Dashyan, Natalya; De Vita, Raffaella; De Sanctis, Enzo; Deur, Alexandre; Djalali, Chaden; Dodge, Gail; Doughty, David; Dupre, Raphael; El Alaoui, Ahmed; El Fassi, Lamiaa; Elouadrhiri, Latifa; Fedotov, Gleb; Fegan, Stuart; Fersch, Robert; Fleming, Jamie; Forest, Tony; Garcon, Michel; Gevorgyan, Nerses; Ghandilyan, Yeranuhi; Gilfoyle, Gerard; Girod-Gard, Francois-Xavier; Giovanetti, Kevin; Goetz, John; Gohn, Wesley; Gothe, Ralf; Griffioen, Keith; Guegan, Baptiste; Guler, Nevzat; Hafidi, Kawtar; Hanretty, Charles; Harrison, Nathan; Hattawy, Mohammad; Hicks, Kenneth; Ho, Dao; Holtrop, Maurik; Ilieva, Yordanka; Ireland, David; Ishkhanov, Boris; Isupov, Evgeny; Jawalkar, Sucheta; Jiang, Xiaodong; Jo, Hyon-Suk; Joo, Kyungseon; Kalantarians, Narbe; Keith, Christopher; Keller, Daniel; Khandaker, Mahbubul; Kim, Andrey; Kim, Wooyoung; Klein, Andreas; Klein, Franz; Koirala, Suman; Kubarovsky, Valery; Kuhn, Sebastian; Kuleshov, Sergey; Lenisa, Paolo; Livingston, Kenneth; Lu, Haiyun; MacGregor, Ian; Markov, Nikolai; Mayer, Michael; McKinnon, Bryan; Meekins, David; Mineeva, Taisiya; Mirazita, Marco; Mokeev, Viktor; Montgomery, Rachel; MOUTARDE, Herve; Movsisyan, Aram; Munevar Espitia, Edwin; Munoz Camacho, Carlos; Nadel-Turonski, Pawel; Niccolai, Silvia; Niculescu, Gabriel; Niculescu, Maria; Osipenko, Mikhail; Ostrovidov, Alexander; Pappalardo, Luciano; Paremuzyan, Rafayel; Park, K; Peng, Peng; Phillips, J J; Pierce, Joshua; Pisano, Silvia; Pogorelko, Oleg; Pozdniakov, Serguei; Price, John; Procureur, Sebastien; Protopopescu, Dan; Puckett, Andrew; Raue, Brian; Rimal, Dipak; Ripani, Marco; Rizzo, Alessandro; Rosner, Guenther; Rossi, Patrizia; Roy, Priyashree; Sabatie, Franck; Saini, Mukesh; Salgado, Carlos; Schott, Diane; Schumacher, Reinhard; Seder, Erin; Sharabian, Youri; Simonyan, Ani; Smith, Claude; Smith, Gregory; Sober, Daniel; Sokhan, Daria; Stepanyan, Stepan; Stepanyan, Samuel; Strakovski, Igor; Strauch, Steffen; Sytnik, Valeriy; Taiuti, Mauro; Tang, Wei; Tkachenko, Svyatoslav; Ungaro, Maurizio; Vernarsky, Brian; Vlasov, Alexander; Voskanyan, Hakob; Voutier, Eric; Walford, Natalie; Watts, Daniel; Weinstein, Lawrence; Zachariou, Nicholas; Zana, Lorenzo; Zhang, Jixie; Zhao, Bo; Zhao, Zhiwen; Zonta, Irene

    2014-08-01

    The inclusive polarized structure functions of the proton and deuteron, g1p and g1d, were measured with high statistical precision using polarized 6 GeV electrons incident on a polarized ammonia target in Hall B at Jefferson Laboratory. Electrons scattered at lab angles between 18 and 45 degrees were detected using the CEBAF Large Acceptance Spectrometer (CLAS). For the usual DIS kinematics, Q^2>1 GeV^2 and the final-state invariant mass W>2 GeV, the ratio of polarized to unpolarized structure functions g1/F1 is found to be nearly independent of Q^2 at fixed x. Significant resonant structure is apparent at values of W up to 2.3 GeV. In the framework of perturbative QCD, the high-W results can be used to better constrain the polarization of quarks and gluons in the nucleon, as well as high-twist contributions.

  1. cDNA, genomic sequence cloning and overexpression of giant panda (Ailuropoda melanoleuca) mitochondrial ATP synthase ATP5G1.

    PubMed

    Hou, W-R; Hou, Y-L; Ding, X; Wang, T

    2012-09-03

    The ATP5G1 gene is one of the three genes that encode mitochondrial ATP synthase subunit c of the proton channel. We cloned the cDNA and determined the genomic sequence of the ATP5G1 gene from the giant panda (Ailuropoda melanoleuca) using RT-PCR technology and touchdown-PCR, respectively. The cloned cDNA fragment contains an open reading frame of 411 bp encoding 136 amino acids; the length of the genomic sequence is of 1838 bp, containing three exons and two introns. Alignment analysis revealed that the nucleotide sequence and the deduced protein sequence are highly conserved compared to Homo sapiens, Mus musculus, Rattus norvegicus, Bos taurus, and Sus scrofa. The homologies for nucleotide sequences of the giant panda ATP5G1 to those of these species are 93.92, 92.21, 92.46, 93.67, and 92.46%, respectively, and the homologies for amino acid sequences are 90.44, 95.59, 93.38, 94.12, and 91.91%, respectively. Topology prediction showed that there is one protein kinase C phosphorylation site, one casein kinase II phosphorylation site, five N-myristoylation sites, and one ATP synthase c subunit signature in the ATP5G1 protein of the giant panda. The cDNA of ATP5G1 was transfected into Escherichia coli, and the ATP5G1 fused with the N-terminally GST-tagged protein gave rise to accumulation of an expected 40-kDa polypeptide, which had the characteristics of the predicted protein.

  2. Tracking the evolution of the G1/RHDVb recombinant strains introduced from the Iberian Peninsula to the Azores islands, Portugal.

    PubMed

    Almeida, Tereza; Lopes, Ana M; Magalhães, Maria J; Neves, Fabiana; Pinheiro, Ana; Gonçalves, David; Leitão, Manuel; Esteves, Pedro J; Abrantes, Joana

    2015-08-01

    Previous genetic characterization of rabbit haemorrhagic disease virus (RHDV) from Azores, Portugal, revealed the presence of genogroup 3-5 (G3-G5) like strains. These strains differed from the genogroup 1 (G1) strains circulating in mainland Portugal, suggesting an independent evolution of RHDV in Azores. More recently, the new variant RHDV (RHDVb) was detected in Azores. In mainland Portugal, current circulating strains resulted from recombination events between RHDVb and non-pathogenic or pathogenic G1 strains. To characterize the RHDVb strains from Azores, a ∼2.5 kb fragment of the RHDV genome (nucleotide positions 4873-7323), including the complete sequence of the capsid gene VP60 (nucleotide positions 5305-7044), was amplified and sequenced. Samples were obtained from rabbits found dead in the field between December 2014 and March 2015 in the Azorean islands Flores, Graciosa, São Jorge, Terceira, Faial, Pico, São Miguel and Santa Maria. For VP60, the highest homology was found with Iberian RHDVb strains, while the upstream fragment revealed high similarity (∼95%) with Iberian G1 strains. Phylogenetic reconstruction based either on VP60 or VP10 grouped the Azorean strains with Iberian RHDVb strains. For the fragment upstream of VP60, the Azorean strains grouped with G1. Our results show that the RHDVb strains circulating in Azores are G1/RHDVb recombinants and we hypothesize that such strains had their origin in Iberian strains. The geographic isolation of Azores suggests that arrival of RHDVb was man-mediated. A network analysis further allowed us to trace virus dispersion in Azores: from an initial outbreak in Graciosa, RHDVb spread to São Jorge and Faial, to Terceira, Flores and Santa Maria, and finally to Pico; dispersion to São Miguel occurred later from Terceira. As the consequences of the presence of G1/RHDVb strains in Azores are unpredictable, we suggest a continued monitoring and characterization of RHD outbreaks.

  3. cDNA, genomic sequence cloning and overexpression of giant panda (Ailuropoda melanoleuca) mitochondrial ATP synthase ATP5G1.

    PubMed

    Hou, W-R; Hou, Y-L; Ding, X; Wang, T

    2012-01-01

    The ATP5G1 gene is one of the three genes that encode mitochondrial ATP synthase subunit c of the proton channel. We cloned the cDNA and determined the genomic sequence of the ATP5G1 gene from the giant panda (Ailuropoda melanoleuca) using RT-PCR technology and touchdown-PCR, respectively. The cloned cDNA fragment contains an open reading frame of 411 bp encoding 136 amino acids; the length of the genomic sequence is of 1838 bp, containing three exons and two introns. Alignment analysis revealed that the nucleotide sequence and the deduced protein sequence are highly conserved compared to Homo sapiens, Mus musculus, Rattus norvegicus, Bos taurus, and Sus scrofa. The homologies for nucleotide sequences of the giant panda ATP5G1 to those of these species are 93.92, 92.21, 92.46, 93.67, and 92.46%, respectively, and the homologies for amino acid sequences are 90.44, 95.59, 93.38, 94.12, and 91.91%, respectively. Topology prediction showed that there is one protein kinase C phosphorylation site, one casein kinase II phosphorylation site, five N-myristoylation sites, and one ATP synthase c subunit signature in the ATP5G1 protein of the giant panda. The cDNA of ATP5G1 was transfected into Escherichia coli, and the ATP5G1 fused with the N-terminally GST-tagged protein gave rise to accumulation of an expected 40-kDa polypeptide, which had the characteristics of the predicted protein. PMID:23007995

  4. Tracking the evolution of the G1/RHDVb recombinant strains introduced from the Iberian Peninsula to the Azores islands, Portugal.

    PubMed

    Almeida, Tereza; Lopes, Ana M; Magalhães, Maria J; Neves, Fabiana; Pinheiro, Ana; Gonçalves, David; Leitão, Manuel; Esteves, Pedro J; Abrantes, Joana

    2015-08-01

    Previous genetic characterization of rabbit haemorrhagic disease virus (RHDV) from Azores, Portugal, revealed the presence of genogroup 3-5 (G3-G5) like strains. These strains differed from the genogroup 1 (G1) strains circulating in mainland Portugal, suggesting an independent evolution of RHDV in Azores. More recently, the new variant RHDV (RHDVb) was detected in Azores. In mainland Portugal, current circulating strains resulted from recombination events between RHDVb and non-pathogenic or pathogenic G1 strains. To characterize the RHDVb strains from Azores, a ∼2.5 kb fragment of the RHDV genome (nucleotide positions 4873-7323), including the complete sequence of the capsid gene VP60 (nucleotide positions 5305-7044), was amplified and sequenced. Samples were obtained from rabbits found dead in the field between December 2014 and March 2015 in the Azorean islands Flores, Graciosa, São Jorge, Terceira, Faial, Pico, São Miguel and Santa Maria. For VP60, the highest homology was found with Iberian RHDVb strains, while the upstream fragment revealed high similarity (∼95%) with Iberian G1 strains. Phylogenetic reconstruction based either on VP60 or VP10 grouped the Azorean strains with Iberian RHDVb strains. For the fragment upstream of VP60, the Azorean strains grouped with G1. Our results show that the RHDVb strains circulating in Azores are G1/RHDVb recombinants and we hypothesize that such strains had their origin in Iberian strains. The geographic isolation of Azores suggests that arrival of RHDVb was man-mediated. A network analysis further allowed us to trace virus dispersion in Azores: from an initial outbreak in Graciosa, RHDVb spread to São Jorge and Faial, to Terceira, Flores and Santa Maria, and finally to Pico; dispersion to São Miguel occurred later from Terceira. As the consequences of the presence of G1/RHDVb strains in Azores are unpredictable, we suggest a continued monitoring and characterization of RHD outbreaks. PMID:26165506

  5. HLA-G1, but Not HLA-G3, Suppresses Human Monocyte/Macrophage-mediated Swine Endothelial Cell Lysis.

    PubMed

    Eguchi, H; Maeda, A; Lo, P C; Matsuura, R; Esquivel, E L; Asada, M; Sakai, R; Nakahata, K; Yamamichi, T; Umeda, S; Deguchi, K; Ueno, T; Okuyama, H; Miyagawa, S

    2016-05-01

    The inhibitory function of HLA-G1, a class Ib molecule, on monocyte/macrophage-mediated cytotoxicity was examined. The expression of inhibitory receptors that interact with HLA-G, immunoglobulin-like transcript 2 (ILT2), ILT4, and KIR2DL4 (CD158d) on in vitro-generated macrophages obtained from peripheral blood mononuclear cells and the phorbol 12-myristate 13-acetate (PMA)-activated THP-1 cells were examined by flow cytometry. cDNAs of HLA-G1, HLA-G3, HLA-E, and human β2-microglobulin were prepared, transfected into pig endothelial cells (PECs), and macrophage- and the THP-1 cell-mediated PEC cytolysis was then assessed. In vitro-generated macrophages expressed not only ILT2 and ILT4 but CD158d as well. The transgenic HLA-G1 on PEC indicated a significant suppression in macrophage-mediated cytotoxicity, which was equivalent to that of transgenic HLA-E. HLA-G1 was clearly expressed on the cell surface of PEC, whereas the levels of HLA-G3 were much lower and remained in the intracellular space. On the other hand, the PMA-activated THP-1 cell was less expressed these inhibitory molecules than in vitro-generated macrophages. Therefore, the HLA-G1 on PECs showed a significant but relatively smaller suppression to THP-1 cell-mediated cytotoxicity compared to in vitro-generated macrophages. These results indicate that by generating HLA-G1, but not HLA-G3, transgenic pigs can protect porcine grafts from monocyte/macrophage-mediated cytotoxicity. PMID:27320605

  6. A Novel Single Virus Infection System Reveals That Influenza Virus Preferentially Infects Cells in G1 Phase

    PubMed Central

    Ueda, Ryuta; Sugiura, Tadao; Kume, Shinichiro; Ichikawa, Akihiko; Larsen, Steven; Miyoshi, Hideaki; Hiramatsu, Hiroaki; Nagatsuka, Yasuko; Arai, Fumihito; Suzuki, Yasuo; Hirabayashi, Yoshio; Fukuda, Toshio; Honda, Ayae

    2013-01-01

    Background Influenza virus attaches to sialic acid residues on the surface of host cells via the hemagglutinin (HA), a glycoprotein expressed on the viral envelope, and enters into the cytoplasm by receptor-mediated endocytosis. The viral genome is released and transported in to the nucleus, where transcription and replication take place. However, cellular factors affecting the influenza virus infection such as the cell cycle remain uncharacterized. Methods/Results To resolve the influence of cell cycle on influenza virus infection, we performed a single-virus infection analysis using optical tweezers. Using this newly developed single-virus infection system, the fluorescence-labeled influenza virus was trapped on a microchip using a laser (1064 nm) at 0.6 W, transported, and released onto individual H292 human lung epithelial cells. Interestingly, the influenza virus attached selectively to cells in the G1-phase. To clarify the molecular differences between cells in G1- and S/G2/M-phase, we performed several physical and chemical assays. Results indicated that: 1) the membranes of cells in G1-phase contained greater amounts of sialic acids (glycoproteins) than the membranes of cells in S/G2/M-phase; 2) the membrane stiffness of cells in S/G2/M-phase is more rigid than those in G1-phase by measurement using optical tweezers; and 3) S/G2/M-phase cells contained higher content of Gb3, Gb4 and GlcCer than G1-phase cells by an assay for lipid composition. Conclusions A novel single-virus infection system was developed to characterize the difference in influenza virus susceptibility between G1- and S/G2/M-phase cells. Differences in virus binding specificity were associated with alterations in the lipid composition, sialic acid content, and membrane stiffness. This single-virus infection system will be useful for studying the infection mechanisms of other viruses. PMID:23874406

  7. Glut1 deficiency (G1D): Epilepsy and metabolic dysfunction in a mouse model of the most common human phenotype

    PubMed Central

    Marin-Valencia, Isaac; Good, Levi B.; Ma, Qian; Duarte, Joao; Bottiglieri, Teodoro; Sinton, Christopher M.; Heilig, Charles W.; Pascual, Juan M.

    2012-01-01

    Brain glucose supplies most of the carbon required for acetyl-coenzyme A (acetyl-CoA) generation (an important step for myelin synthesis) and for neurotransmitter production via further metabolism of acetyl-CoA in the tricarboxylic acid (TCA) cycle. However, it is not known whether reduced brain glucose transporter type I (GLUT-1) activity, the hallmark of the GLUT-1 deficiency (G1D) syndrome, leads to acetyl-CoA, TCA or neurotransmitter depletion. This question is relevant because, in its most common form in man, G1D is associated with cerebral hypomyelination (manifested as microcephaly) and epilepsy, suggestive of acetyl-CoA depletion and neurotransmitter dysfunction, respectively. Yet, brain metabolism in G1D remains underexplored both theoretically and experimentally, partly because computational models of limited brain glucose transport are subordinate to metabolic assumptions and partly because current hemizygous G1D mouse models manifest a mild phenotype not easily amenable to investigation. In contrast, adult antisense G1D mice replicate the human phenotype of spontaneous epilepsy associated with robust thalamocortical electrical oscillations. Additionally, and in consonance with human metabolic imaging observations, thalamus and cerebral cortex display the lowest GLUT-1 expression and glucose uptake in the mutant mouse. This depletion of brain glucose is associated with diminished plasma fatty acids and elevated ketone body levels, and with decreased brain acetyl-CoA and fatty acid contents, consistent with brain ketone body consumption and with stimulation of brain beta-oxidation and/or diminished cerebral lipid synthesis. In contrast with other epilepsies, astrocyte glutamine synthetase expression, cerebral TCA cycle intermediates, amino acid and amine neurotransmitter contents are also intact in G1D. The data suggest that the TCA cycle is preserved in G1D because reduced glycolysis and acetyl-CoA formation can be balanced by enhanced ketone body

  8. Base substitution mutations in uridinediphosphate-dependent glycosyltransferase 76G1 gene of Stevia rebaudiana causes the low levels of rebaudioside A: mutations in UGT76G1, a key gene of steviol glycosides synthesis.

    PubMed

    Yang, Yong-Heng; Huang, Su-Zhen; Han, Yu-Lin; Yuan, Hai-Yan; Gu, Chun-Sun; Zhao, Yan-Hai

    2014-07-01

    Steviol glycosides, extracted from the leaves of Stevia rebaudiana (Bert) Bertoni, are calorie-free sugar substitute of natural origin with intensely sweet (Boileau et al., 2012). Stevioside and rebaudioside A are the two main kinds of the diterpenic glycosides. We analyzed the concentration of stevioside and rebaudioside A in Stevia leaves of about 500 samples (hybrid progenies) and discovered a mutation plant "Z05" with very low levels of rebaudioside A. Because UGT76G1, a uridinediphosphate-dependent glycosyltransferases, is responsible for the conversion from stevioside to rebaudioside A (Richman et al., 2005), so mutation identification was done by sequencing the candidate gene, UGT76G1. In this study molecular analysis of two strains revealed a heterozygotic nonsense mutation of c.389T > G (p.L121X) in UGT76G1. Meanwhile, we found some amino acid substitutions significant change the protein structure. And the difference of enzyme activity between two strains proved the lack of functionality of UGT76G1 of the mutation "Z05". So the nonsense mutation and amino acid substitution mutation resulted in the low levels of rebaudioside A.

  9. IgG1 adsorption to siliconized glass vials-influence of pH, ionic strength, and nonionic surfactants.

    PubMed

    Höger, Kerstin; Mathes, Johannes; Frieß, Wolfgang

    2015-01-01

    In this study, the adsorption of an IgG1 antibody to siliconized vials was investigated with focus on the formulation parameters pH, ionic strength, and nonionic surfactants. Electrophoretic mobility measurements were performed to investigate the charge characteristics of protein and siliconized glass particles at different pH values. Calculation of the electrokinetic charge density allowed further insight into the energetic conditions in the protein-sorbent interface. Maximum adsorption of IgG1 was found at acidic pH values and could be correlated with energetically favorable minimal ion incorporation into the interface. The importance of electrostatic interactions for IgG1 adsorption at acidic pH values was also confirmed by the efficient adsorption reduction at decreased solution ionic strength. A second adsorption maximum around the pI of the protein was assigned to hydrophobic interactions with the siliconized surface. Addition of the nonionic surfactants poloxamer 188 or polysorbate 80 resulted in almost complete suppression of adsorption at pH 7.2, and a strong but less efficient effect at pH 4 on siliconized glass vials. This adsorption suppression was much less pronounced on borosilicate glass vials. From these results, it can be concluded that electrostatic interactions contribute substantially to IgG1 adsorption to siliconized glass vials especially at acidic formulation pH.

  10. Identification of O-methylsterigmatocystin as an aflatoxin B1 and G1 precursor in Aspergillus parasiticus.

    PubMed Central

    Bhatnagar, D; McCormick, S P; Lee, L S; Hill, R A

    1987-01-01

    An isolate of Aspergillus parasiticus CP461 (SRRC 2043) produced no detectable aflatoxins, but accumulated O-methylsterigmatocystin (OMST). When sterigmatocystin (ST) was fed to this isolate in a low-sugar medium, there was an increase in the accumulation of OMST, without aflatoxin synthesis. When radiolabeled [14C]OMST was fed to resting mycelia of a non-aflatoxin-, non-ST-, and non-OMST-producing mutant of A. parasiticus AVN-1 (SRRC 163), 14C-labeled aflatoxins B1 and G1 were produced; 10 nmol of OMST produced 7.8 nmol of B1 and 1.0 nmol of G1, while 10 nmol of ST produced 6.4 nmol of B1 and 0.6 nmol of G1. A time course study of aflatoxin synthesis in ST feeding experiments with AVN-1 revealed that OMST is synthesized by the mold during the onset of aflatoxin synthesis. The total amount of aflatoxins recovered from OMST feeding experiments was higher than from experiments in which ST was fed to the resting mycelia. These results suggest that OMST is a true metabolite in the aflatoxin biosynthetic pathway between sterigmatocystin and aflatoxins B1 and G1 and is not a shunt metabolite, as thought previously. PMID:3111363

  11. Ethanol fermentation driven by elevated expression of the G1 cyclin gene CLN3 in sake yeast.

    PubMed

    Watanabe, Daisuke; Nogami, Satoru; Ohya, Yoshikazu; Kanno, Yoichiro; Zhou, Yan; Akao, Takeshi; Shimoi, Hitoshi

    2011-12-01

    Cellular and subcellular morphology reflects the physiological state of a cell. To determine the physiological nature of sake yeast with superior fermentation properties, we quantitatively analyzed the morphology of sake yeast cells by using the CalMorph system. All the sake strains examined here exhibited common morphological traits that are typically observed in the well-characterized whiskey (whi) mutants that show accelerated G(1)/S transition. In agreement with this finding, the sake strain showed less efficient G(0)/G(1) arrest and elevated expression of the G(1) cyclin gene CLN3 throughout the fermentation period. Furthermore, deletion of CLN3 remarkably impaired the fermentation rate in both sake and laboratory strains. Disruption of the SWI6 gene, a transcriptional coactivator responsible for Cln3p-mediated G(1)/S transition, also resulted in a decreased fermentation rate, whereas whi mutants exhibited significant improvement in the fermentation rate, demonstrating positive roles of Cln3p and its downstream signalling pathway in facilitating ethanol fermentation. The combined results indicate that enhanced induction of CLN3 contributes to the high fermentation rate of sake yeast, which are natural whi mutants.

  12. A two-stage approach in solving the state probabilities of the multi-queue M/G/1 model

    NASA Astrophysics Data System (ADS)

    Chen, Mu-Song; Yen, Hao-Wei

    2016-04-01

    The M/G/1 model is the fundamental basis of the queueing system in many network systems. Usually, the study of the M/G/1 is limited by the assumption of single queue and infinite capacity. In practice, however, these postulations may not be valid, particularly when dealing with many real-world problems. In this paper, a two-stage state-space approach is devoted to solving the state probabilities for the multi-queue finite-capacity M/G/1 model, i.e. q-M/G/1/Ki with Ki buffers in the ith queue. The state probabilities at departure instants are determined by solving a set of state transition equations. Afterward, an embedded Markov chain analysis is applied to derive the state probabilities with another set of state balance equations at arbitrary time instants. The closed forms of the state probabilities are also presented with theorems for reference. Applications of Little's theorem further present the corresponding results for queue lengths and average waiting times. Simulation experiments have demonstrated the correctness of the proposed approaches.

  13. Experimental infection with Cryptosporidium parvum IIaA21G1R1 subtype in immunosuppressed mice

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cryptosporidium parvum subtype IIaA21G1R1 oocysts were used to infect dexamethasone immunosuppressed N: NIH Swiss mice. Histology showed developmental stages in the duodenum, proximal and distal jejunum, ileum, cecum and colon, with the small intestine remaining infected until day 35 post infection....

  14. 26 CFR 1.904(g)-1T - Overall domestic loss and the overall domestic loss account (temporary).

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... Without the United States § 1.904(g)-1T Overall domestic loss and the overall domestic loss account..., net capital losses, U.S. source losses, and separate limitation losses, and the recapture of separate... any capital gains or losses, the amount of the taxpayer's domestic loss shall be determined by...

  15. Infection of humans and animals with Echinococcus granulosus (G1 and G3 strains) and E. ortleppi in Southern Brazil.

    PubMed

    de la Rue, Mario L; Takano, Keishi; Brochado, Joaquim F; Costa, Carmem V; Soares, Antonio G; Yamano, Kimiaki; Yagi, Kinpei; Katoh, Yoshinobu; Takahashi, Kenichi

    2011-04-19

    The Rio Grande do Sul state, in Southern Brazil, is one of the foci of human cystic echinococcosis (CE). The sheep strain (G1) of Echinococcus granulosus and Echinococcus ortleppi (also known as cattle strain G5) have been reported before to infect livestock. However, up to the present, no molecular data are available on isolates of the E. granulosus complex from humans and dogs. The present study analyzed hydatid cysts from 6 CE patients and adult worms from 12 dogs. Sequencing of the mitochondrial cox1 and 12S rRNA genes detected the E. granulosus G1 genotype from four human cases, the G3 genotype (or buffalo strain) from one human case and E. ortleppi from another human case, respectively. Ten of the twelve dogs were found infected with the G1 genotype, and one dog each harbored worms of the G3 genotype and E. ortleppi. Obvious morphological differences were recognized between the G1 and E. ortleppi adult worms from dogs in this region. The buffalo strain (G3) is for the first time reported from South America.

  16. Association of Cdk2/cyclin E and NF-kappa B complexes at G1/S phase.

    PubMed

    Chen, E; Li, C C

    1998-08-28

    NF-kappa B/Rel family plays a pivotal role in a wide variety of cellular functions including growth, development, apoptosis and stress responses. Recent studies indicated that NF-kappa B is also involved in the cell cycle regulation, and high expression of c-Rel results in a cell cycle arrest at the G1/S-phase transition (Bash, J., Zong, W,-X., and Gelinas, C. (1997) Mol. Cell. Biol. 17, 6526-6536). Here we report the detection of Cdk2, a critical kinase responsible for the G1/S-phase transition, in immune complexes precipitated by the NF-kappa B antisera. Cdk2 and NF-kappa B association was detected by co-precipitation in the nuclear lysates of the G1/S-phase cells, and was found in cultured cell lines and in T cells purified from human peripheral blood. Using an affinity column containing the C-terminal peptide of human c-Rel, we isolated cyclin E, the regulatory subunit of the Cdk2 complex, as a c-Rel-binding protein. These findings support and provide physical basis for the involvement of NF-kappa B in the G1/S-phase cell cycle control, and suggest an important role played by the C-terminal sequence of c-Rel. PMID:9731206

  17. Antigenic differences between the EG95-related proteins from Echinococcus granulosus G1 and G6 genotypes: implications for vaccination.

    PubMed

    Alvarez Rojas, C A; Gauci, C G; Lightowlers, M W

    2013-02-01

    Cystic echinococcosis caused by Echinococcus granulosus remains an important and neglected issue in public health. The study of the likely efficacy of the currently available EG95 vaccine against other genotypes of the parasite is important to improve the vaccine as a potential tool to be used in control programmes. The recombinant vaccine EG95-1G1 was developed based on the G1 genotype of E. granulosus. Characterization of the eg95 gene family in the G6 genotype by genomic DNA cloning previously produced the first unequivocal information about the composition of the gene family in a different genotype. The information was used in this study to predict and express two EG95-related proteins from the G6 genotype as recombinants, for assessment of their capacity to bind antibodies raised in sheep vaccinated with the EG95-1G1 vaccine. The proteins (EG95-1G6 and EG95-5G6) from the G6 genotype of E. granulosus were unable to bind all the antibodies raised by sheep vaccinated with EG95-1G1. Differences in the amino acid sequence of EG95-related proteins from G6 and likely the differences in the encoded FnIII domain may be responsible for changes in the conformation of these epitopes.

  18. Ethanol fermentation driven by elevated expression of the G1 cyclin gene CLN3 in sake yeast.

    PubMed

    Watanabe, Daisuke; Nogami, Satoru; Ohya, Yoshikazu; Kanno, Yoichiro; Zhou, Yan; Akao, Takeshi; Shimoi, Hitoshi

    2011-12-01

    Cellular and subcellular morphology reflects the physiological state of a cell. To determine the physiological nature of sake yeast with superior fermentation properties, we quantitatively analyzed the morphology of sake yeast cells by using the CalMorph system. All the sake strains examined here exhibited common morphological traits that are typically observed in the well-characterized whiskey (whi) mutants that show accelerated G(1)/S transition. In agreement with this finding, the sake strain showed less efficient G(0)/G(1) arrest and elevated expression of the G(1) cyclin gene CLN3 throughout the fermentation period. Furthermore, deletion of CLN3 remarkably impaired the fermentation rate in both sake and laboratory strains. Disruption of the SWI6 gene, a transcriptional coactivator responsible for Cln3p-mediated G(1)/S transition, also resulted in a decreased fermentation rate, whereas whi mutants exhibited significant improvement in the fermentation rate, demonstrating positive roles of Cln3p and its downstream signalling pathway in facilitating ethanol fermentation. The combined results indicate that enhanced induction of CLN3 contributes to the high fermentation rate of sake yeast, which are natural whi mutants. PMID:21906996

  19. Whole-genomic analysis of a human G1P[9] rotavirus strain reveals intergenogroup-reassortment events.

    PubMed

    Ghosh, Souvik; Shintani, Tsuzumi; Urushibara, Noriko; Taniguchi, Koki; Kobayashi, Nobumichi

    2012-08-01

    Group A rotavirus (RVA) strain K8 (RVA/Human-tc/JPN/K8/1977/G1P[9]) was found to have Wa-like VP7 and NSP1 genes and AU-1-like VP4 and NSP5 genes. To determine the exact origin and overall genetic makeup of this unusual RVA strain, the remaining genes (VP1-VP3, VP6 and NSP2-NSP4) of K8 were analysed in this study. Strain K8 exhibited a G1-P[9]-I1-R3-C3-M3-A1-N1-T3-E3-H3 genotype constellation, not reported previously. The VP6 and NSP2 genes of strain K8 were related closely to those of common human Wa-like G1P[8] and/or G3P[8] strains, whilst its VP1-VP3, NSP3 and NSP4 genes were related more closely to those of AU-1-like RVAs and/or AU-1-like genes of multi-reassortant strains than to those of other RVAs. Therefore, strain K8 might have originated from intergenogroup-reassortment events involving acquisition of four Wa-like genes, possibly from G1P[8] RVAs, by an AU-1-like P[9] strain. Whole-genomic analysis of strain K8 has provided important insights into the complex genetic diversity of RVAs. PMID:22592265

  20. Dynamic FoxG1 expression coordinates the integration of multipolar pyramidal neuron precursors into the cortical plate.

    PubMed

    Miyoshi, Goichi; Fishell, Gord

    2012-06-21

    Pyramidal cells of the cerebral cortex are born in the ventricular zone and migrate through the intermediate zone to enter into the cortical plate. In the intermediate zone, these migrating precursors move tangentially and initiate the extension of their axons by transiently adopting a characteristic multipolar morphology. We observe that expression of the forkhead transcription factor FoxG1 is dynamically regulated during this transitional period. By utilizing conditional genetic strategies, we show that the downregulation of FoxG1 at the beginning of the multipolar cell phase induces Unc5D expression, the timing of which ultimately determines the laminar identity of pyramidal neurons. In addition, we demonstrate that the re-expression of FoxG1 is required for cells to transit out of the multipolar cell phase and to enter into the cortical plate. Thus, the dynamic expression of FoxG1 during migration within the intermediate zone is essential for the proper assembly of the cerebral cortex.

  1. Thrombus imaging: A comparison of radiolabeled GC4 and T2G1s fibrin-specific monoclonal antibodies

    SciTech Connect

    Rosebrough, S.F.; McAfee, J.G.; Grossman, Z.D.; Kudryk, B.J.; Ritter-Hrncirik, C.A.; Witanowski, L.S.; Maley, B.L.; Bertrand, E.A.; Gagne, G.M. )

    1990-06-01

    Radioimmunoimaging of experimentally-induced canine thrombi has previously been achieved with iodine-131- and indium-111-labeled ({sup 131}I and {sup 111}In) anti-fibrin T2G1s monoclonal antibody (MAb). We now compare T2G1s to another anti-fibrin MAb, designated GC4, for imaging fresh and aged canine thrombi. GC4 is specific for a neoepitope exposed on fibrin later in the thrombolytic process after plasmin digestion. Femoral venous thrombi were induced in six groups of dogs, each containing three dogs. In two groups, the MAbs were compared when the thrombi were 3-hr or 3-days old at the time of injection, and the dogs were killed at 48 hr. In thrombi 3-hr-old, the GC4/T2G1s concentration ratio averaged 0.53 compared to 1.9 in 3-day-old thrombi. Two groups of dogs with thrombi 1- or 3-days-old were heparinized before MAb injection and were killed at 24 hr. The heparinized dogs with thrombi 1- or 3-days-old had GC4/T2G1s mean ratios of 2.3 and 2.9, respectively. In the unheparinized groups, the corresponding ratios were 1.1 and 1.9. GC4 may be more useful for clinical thrombus imaging than T2G1s because spontaneous venous thrombi are usually several days old at the time of presentation and patients are often heparinized immediately.

  2. Overestimated Oncologic Significance of Lymph Node Metastasis in G1 Nonfunctioning Neuroendocrine Tumor in the Left Side of the Pancreas.

    PubMed

    Yoo, Young Jin; Yang, Seok Jeong; Hwang, Ho Kyoung; Kang, Chang Moo; Kim, Hogeun; Lee, Woo Jung

    2015-09-01

    Recent studies have expounded on the oncologic significance of lymph node metastasis in nonfunctioning (NF) neuroendocrine tumors (NETs) of the pancreas and suggest regional lymph node dissection for treating pancreatic NET. We tested this recommendation in NF pancreatic NET-G1, as these tumors are generally small and suitable for function-preserving minimally invasive pancreatectomy.From January 2005 to December 2014, medical records of patients who underwent pancreatectomy for pathologically confirmed NF NET-G1 of the left side of the pancreas were retrospectively reviewed. Oncologic outcomes were compared between limited pancreatectomy and distal pancreatosplenectomy.Thirty-five patients (14 males and 21 females) with a mean age of 55.9 ± 11.4 years were enrolled in this study. Six patients (17.1%) underwent distal pancreatosplenectomy. Limited pancreatectomies comprised 15 spleen-preserving distal pancreatectomies (42.8%), 10 enucleations (28.6%), and 4 central pancreatectomies (11.4%). Lymph node metastasis was not found in 6 patients who underwent distal pancreatectomy with a splenectomy; meanwhile, the others were regarded as pNx since no lymph node retrieval was attempted during the limited pancreatectomy. Overall disease-free survival was 36.5 months (95% confidence interval [CI]: 25.9-47.1) and no tumor-related mortality was noted. Minimally invasive pancreatectomy (P = 0.557) and limited pancreatectomy (P = 0.758) showed no adverse impact in treating NF NET-G1 of the left side of the pancreas.The oncologic significance of lymph node metastasis is overestimated in NF NET-G1 of the left side of the pancreas. Routine conventional distal pancreatosplenectomy to retrieve regional lymph nodes may be too excessive in treating NF NET-G1 of the distal pancreas.

  3. Whole exome sequencing of rare variants in EIF4G1 and VPS35 in Parkinson disease

    PubMed Central

    Nuytemans, Karen; Bademci, Guney; Inchausti, Vanessa; Dressen, Amy; Kinnamon, Daniel D.; Mehta, Arpit; Wang, Liyong; Züchner, Stephan; Beecham, Gary W.; Martin, Eden R.; Scott, William K.

    2013-01-01

    Objective: Recently, vacuolar protein sorting 35 (VPS35) and eukaryotic translation initiation factor 4 gamma 1 (EIF4G1) have been identified as 2 causal Parkinson disease (PD) genes. We used whole exome sequencing for rapid, parallel analysis of variations in these 2 genes. Methods: We performed whole exome sequencing in 213 patients with PD and 272 control individuals. Those rare variants (RVs) with <5% frequency in the exome variant server database and our own control data were considered for analysis. We performed joint gene-based tests for association using RVASSOC and SKAT (Sequence Kernel Association Test) as well as single-variant test statistics. Results: We identified 3 novel VPS35 variations that changed the coded amino acid (nonsynonymous) in 3 cases. Two variations were in multiplex families and neither segregated with PD. In EIF4G1, we identified 11 (9 nonsynonymous and 2 small indels) RVs including the reported pathogenic mutation p.R1205H, which segregated in all affected members of a large family, but also in 1 unaffected 86-year-old family member. Two additional RVs were found in isolated patients only. Whereas initial association studies suggested an association (p = 0.04) with all RVs in EIF4G1, subsequent testing in a second dataset for the driving variant (p.F1461) suggested no association between RVs in the gene and PD. Conclusions: We confirm that the specific EIF4G1 variation p.R1205H seems to be a strong PD risk factor, but is nonpenetrant in at least one 86-year-old. A few other select RVs in both genes could not be ruled out as causal. However, there was no evidence for an overall contribution of genetic variability in VPS35 or EIF4G1 to PD development in our dataset. PMID:23408866

  4. Characterization of IgG1 Conformation and Conformational Dynamics by Hydrogen/Deuterium Exchange Mass Spectrometry

    SciTech Connect

    Houde, Damian; Arndt, Joseph; Domeier, Wayne; Berkowitz, Steven; Engen, John R.

    2009-04-22

    Protein function is dictated by protein conformation. For the protein biopharmaceutical industry, therefore, it is important to have analytical tools that can detect changes in protein conformation rapidly, accurately, and with high sensitivity. In this paper we show that hydrogen/deuterium exchange mass spectrometry (H/DX-MS) can play an important role in fulfilling this need within the industry. H/DX-MS was used to assess both global and local conformational behavior of a recombinant monoclonal IgG1 antibody, a major class of biopharmaceuticals. Analysis of exchange into the intact, glycosylated IgG1 (and the Fab and Fc regions thereof) showed that the molecule was folded, highly stable, and highly amenable to analysis by this method using less than a nanomole of material. With improved chromatographic methods, peptide identification algorithms and data-processing steps, the analysis of deuterium levels in peptic peptides produced after labeling was accomplished in 1--2 days. On the basis of peptic peptide data, exchange was localized to specific regions of the antibody. Changes to IgG1 conformation as a result of deglycosylation were determined by comparing exchange into the glycosylated and deglycosylated forms of the antibody. Two regions of the IgG1 (residues 236-253 and 292-308) were found to have altered exchange properties upon deglycosylation. These results are consistent with previous findings concerning the role of glycosylation in the interaction of IgG1 with Fc receptors. Moreover, the data clearly illustrate how H/DX-MS can provide important characterization information on the higher order structure of antibodies and conformational changes that these molecules may experience upon modification.

  5. Role of Androgen Receptor in Progression of LNCaP Prostate Cancer Cells from G1 to S Phase

    PubMed Central

    Murthy, Shalini; Wu, Min; Bai, V. Uma; Hou, Zizheng; Menon, Mani; Barrack, Evelyn R.; Kim, Sahn-Ho; Reddy, G. Prem-Veer

    2013-01-01

    Background The androgen receptor (AR) plays a critical role in the proliferation of prostate cancer cells. However, its mechanism of action in proliferation remains unknown. An understanding of the mechanism of AR action in proliferation may lead to the development of effective strategies for the treatment of prostate cancer. Methodology/Principal Findings In this study we report that pulse treatment of synchronized LNCaP cells with Casodex, an AR-antagonist, for 4 hours in mid-G1 phase was sufficient to prevent cells from entering S phase. Since the assembly of pre-replication complex (pre-RC) in G1 is required for the progression of cells from G1 to S phase, the effect of Casodex during mid-G1 suggested that the role of AR in proliferation might be to regulate the assembly of pre-RC. To test this possibility, we investigated the interaction between AR and Cdc6, an essential component of pre-RC in LNCaP cells. AR co-localized and co-immunoprecipitated with Cdc6, and Casodex treatment disrupted this interaction. AR-immunoprecipitate (AR-IP) also contained cyclin E and cyclin A, which play a critical role in pre-RC assembly and cell cycle entry into S phase, and DNA polymerase-α, PCNA, and ribonucleotide reductase, which are essential for the initiation of DNA synthesis. In addition, in cells in S phase, AR co-sedimented with components of the DNA replication machinery of cells that entered S phase. Conclusions/Significance Together, these observations suggest a novel role of AR as a component of the pre-RC to exert control over progression of LNCaP cells from G1 to S phase through a mechanism that is independent of its role as a transcription factor. PMID:23437213

  6. Genome-Wide Evolutionary Analyses of G1P[8] Strains Isolated Before and After Rotavirus Vaccine Introduction.

    PubMed

    Zeller, Mark; Donato, Celeste; Trovão, Nídia Sequeira; Cowley, Daniel; Heylen, Elisabeth; Donker, Nicole C; McAllen, John K; Akopov, Asmik; Kirkness, Ewen F; Lemey, Philippe; Van Ranst, Marc; Matthijnssens, Jelle; Kirkwood, Carl D

    2015-08-08

    Rotaviruses are the most important etiological agent of acute gastroenteritis in young children worldwide. Among the first countries to introduce rotavirus vaccines into their national immunization programs were Belgium (November 2006) and Australia (July 2007). Surveillance programs in Belgium (since 1999) and Australia (since 1989) offer the opportunity to perform a detailed comparison of rotavirus strains circulating pre- and postvaccine introduction. G1P[8] rotaviruses are the most prominent genotype in humans, and a total of 157 G1P[8] rotaviruses isolated between 1999 and 2011 were selected from Belgium and Australia and their complete genomes were sequenced. Phylogenetic analysis showed evidence of frequent reassortment among Belgian and Australian G1P[8] rotaviruses. Although many different phylogenetic subclusters were present before and after vaccine introduction, some unique clusters were only identified after vaccine introduction, which could be due to natural fluctuation or the first signs of vaccine-driven evolution. The times to the most recent common ancestors for the Belgian and Australian G1P[8] rotaviruses ranged from 1846 to 1955 depending on the gene segment, with VP7 and NSP4 resulting in the most recent estimates. We found no evidence that rotavirus population size was affected after vaccine introduction and only six amino acid sites in VP2, VP3, VP7, and NSP1 were identified to be under positive selective pressure. Continued surveillance of G1P[8] strains is needed to determine long-term effects of vaccine introductions, particularly now rotavirus vaccines are implemented in the national immunization programs of an increasing number of countries worldwide.

  7. Genome-Wide Evolutionary Analyses of G1P[8] Strains Isolated Before and After Rotavirus Vaccine Introduction.

    PubMed

    Zeller, Mark; Donato, Celeste; Trovão, Nídia Sequeira; Cowley, Daniel; Heylen, Elisabeth; Donker, Nicole C; McAllen, John K; Akopov, Asmik; Kirkness, Ewen F; Lemey, Philippe; Van Ranst, Marc; Matthijnssens, Jelle; Kirkwood, Carl D

    2015-09-01

    Rotaviruses are the most important etiological agent of acute gastroenteritis in young children worldwide. Among the first countries to introduce rotavirus vaccines into their national immunization programs were Belgium (November 2006) and Australia (July 2007). Surveillance programs in Belgium (since 1999) and Australia (since 1989) offer the opportunity to perform a detailed comparison of rotavirus strains circulating pre- and postvaccine introduction. G1P[8] rotaviruses are the most prominent genotype in humans, and a total of 157 G1P[8] rotaviruses isolated between 1999 and 2011 were selected from Belgium and Australia and their complete genomes were sequenced. Phylogenetic analysis showed evidence of frequent reassortment among Belgian and Australian G1P[8] rotaviruses. Although many different phylogenetic subclusters were present before and after vaccine introduction, some unique clusters were only identified after vaccine introduction, which could be due to natural fluctuation or the first signs of vaccine-driven evolution. The times to the most recent common ancestors for the Belgian and Australian G1P[8] rotaviruses ranged from 1846 to 1955 depending on the gene segment, with VP7 and NSP4 resulting in the most recent estimates. We found no evidence that rotavirus population size was affected after vaccine introduction and only six amino acid sites in VP2, VP3, VP7, and NSP1 were identified to be under positive selective pressure. Continued surveillance of G1P[8] strains is needed to determine long-term effects of vaccine introductions, particularly now rotavirus vaccines are implemented in the national immunization programs of an increasing number of countries worldwide. PMID:26254487

  8. Genome-Wide Evolutionary Analyses of G1P[8] Strains Isolated Before and After Rotavirus Vaccine Introduction

    PubMed Central

    Zeller, Mark; Donato, Celeste; Trovão, Nídia Sequeira; Cowley, Daniel; Heylen, Elisabeth; Donker, Nicole C.; McAllen, John K.; Akopov, Asmik; Kirkness, Ewen F.; Lemey, Philippe; Van Ranst, Marc; Matthijnssens, Jelle; Kirkwood, Carl D.

    2015-01-01

    Rotaviruses are the most important etiological agent of acute gastroenteritis in young children worldwide. Among the first countries to introduce rotavirus vaccines into their national immunization programs were Belgium (November 2006) and Australia (July 2007). Surveillance programs in Belgium (since 1999) and Australia (since 1989) offer the opportunity to perform a detailed comparison of rotavirus strains circulating pre- and postvaccine introduction. G1P[8] rotaviruses are the most prominent genotype in humans, and a total of 157 G1P[8] rotaviruses isolated between 1999 and 2011 were selected from Belgium and Australia and their complete genomes were sequenced. Phylogenetic analysis showed evidence of frequent reassortment among Belgian and Australian G1P[8] rotaviruses. Although many different phylogenetic subclusters were present before and after vaccine introduction, some unique clusters were only identified after vaccine introduction, which could be due to natural fluctuation or the first signs of vaccine-driven evolution. The times to the most recent common ancestors for the Belgian and Australian G1P[8] rotaviruses ranged from 1846 to 1955 depending on the gene segment, with VP7 and NSP4 resulting in the most recent estimates. We found no evidence that rotavirus population size was affected after vaccine introduction and only six amino acid sites in VP2, VP3, VP7, and NSP1 were identified to be under positive selective pressure. Continued surveillance of G1P[8] strains is needed to determine long-term effects of vaccine introductions, particularly now rotavirus vaccines are implemented in the national immunization programs of an increasing number of countries worldwide. PMID:26254487

  9. Improved ENSO simulation from climate system model FGOALS-g1.0 to FGOALS-g2

    NASA Astrophysics Data System (ADS)

    Chen, Lin; Yu, Yongqiang; Zheng, Weipeng

    2016-10-01

    This study presents an overview of the improvement in the simulation of El Niño-Southern Oscillation (ENSO) in the latest generation of the Institute of Atmospheric Physics' coupled general circulation model (CGCM), the Flexible Global Ocean-Atmosphere-Land System model Grid-point Version 2 (FGOALS-g2; hereafter referred to as "g2") from its predecessor FGOALS-g1.0 (referred to as "g1"), including the more realistic amplitude, irregularity, and ENSO cycle. The changes have been analyzed quantitatively based on the Bjerknes stability index, which serves as a measure of ENSO growth rate. The improved simulation of ENSO amplitude is mainly due to the reasonable representation of the thermocline and thermodynamic feedbacks: On the one hand, the deeper mean thermocline results in a weakened thermocline response to the zonal wind stress anomaly, and the looser vertical stratification of mean temperature leads to a weakened response of anomalous subsurface temperature to anomalous thermocline depth, both of which cause the reduced thermocline feedback in g2; on the other hand, the alleviated cold bias of mean sea surface temperature leads to more reasonable thermodynamic feedback in g2. The regular oscillation of ENSO in g1 is associated with its unsuccessful representation of the role of atmospheric noise over the western-central equatorial Pacific (WCEP) in triggering ENSO events, which arises from the weak synoptic-intraseasonal variability of zonal winds over the WCEP in g1. The asymmetric transition of ENSO in g1 is attributed to the asymmetric effect of thermocline feedback, which is due to the annual cycle of mean upwelling in the eastern Pacific. This study highlights the great impact of improving the representation of mean states on the improved simulation of air-sea feedback processes and ultimately more reasonable depiction of ENSO behaviors in CGCMs.

  10. Cytoplasmic-nuclear trafficking of G1/S cell cycle molecules and adult human β-cell replication: a revised model of human β-cell G1/S control.

    PubMed

    Fiaschi-Taesch, Nathalie M; Kleinberger, Jeffrey W; Salim, Fatimah G; Troxell, Ronnie; Wills, Rachel; Tanwir, Mansoor; Casinelli, Gabriella; Cox, Amy E; Takane, Karen K; Srinivas, Harish; Scott, Donald K; Stewart, Andrew F

    2013-07-01

    Harnessing control of human β-cell proliferation has proven frustratingly difficult. Most G1/S control molecules, generally presumed to be nuclear proteins in the human β-cell, are in fact constrained to the cytoplasm. Here, we asked whether G1/S molecules might traffic into and out of the cytoplasmic compartment in association with activation of cell cycle progression. Cdk6 and cyclin D3 were used to drive human β-cell proliferation and promptly translocated into the nucleus in association with proliferation. In contrast, the cell cycle inhibitors p15, p18, and p19 did not alter their location, remaining cytoplasmic. Conversely, p16, p21, and p27 increased their nuclear frequency. In contrast once again, p57 decreased its nuclear frequency. Whereas proliferating β-cells contained nuclear cyclin D3 and cdk6, proliferation generally did not occur in β-cells that contained nuclear cell cycle inhibitors, except p21. Dynamic cytoplasmic-nuclear trafficking of cdk6 was confirmed using green fluorescent protein-tagged cdk6 and live cell imaging. Thus, we provide novel working models describing the control of cell cycle progression in the human β-cell. In addition to known obstacles to β-cell proliferation, cytoplasmic-to-nuclear trafficking of G1/S molecules may represent an obstacle as well as a therapeutic opportunity for human β-cell expansion. PMID:23493571

  11. Moelcular Phylogenetic Analyses of G1P Dehydrogenase and G3P Dehydrogenase Suggest the Late Origin of Archaea-Type Membrane

    NASA Astrophysics Data System (ADS)

    Yokobori, S.; Nakajima, Y.; Akanuma, S.; Yamagishi, A.

    2013-11-01

    Phylogenetic analyses of G1PDH and G3PDH suggested that the common ancestor of Bacteria/Archaea had cellular membrane with G3P formed by G3PDH. The archaeal ancestry acquired G1PDH, and then the membrane with G3P was replaced with that with G1P.

  12. 26 CFR 1.1033(g)-1 - Condemnation of real property held for productive use in trade or business or for investment.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... productive use in trade or business or for investment. 1.1033(g)-1 Section 1.1033(g)-1 Internal Revenue... Nontaxable Exchanges § 1.1033(g)-1 Condemnation of real property held for productive use in trade or business... advertising displays as real property—(1) In general. Under section 1033(g)(3) of the Code, a taxpayer...

  13. Reassortment of Human and Animal Rotavirus Gene Segments in Emerging DS-1-Like G1P[8] Rotavirus Strains.

    PubMed

    Komoto, Satoshi; Tacharoenmuang, Ratana; Guntapong, Ratigorn; Ide, Tomihiko; Tsuji, Takao; Yoshikawa, Tetsushi; Tharmaphornpilas, Piyanit; Sangkitporn, Somchai; Taniguchi, Koki

    2016-01-01

    The emergence and rapid spread of novel DS-1-like G1P[8] human rotaviruses in Japan were recently reported. More recently, such intergenogroup reassortant strains were identified in Thailand, implying the ongoing spread of unusual rotavirus strains in Asia. During rotavirus surveillance in Thailand, three DS-1-like intergenogroup reassortant strains having G3P[8] (RVA/Human-wt/THA/SKT-281/2013/G3P[8] and RVA/Human-wt/THA/SKT-289/2013/G3P[8]) and G2P[8] (RVA/Human-wt/THA/LS-04/2013/G2P[8]) genotypes were identified in fecal samples from hospitalized children with acute gastroenteritis. In this study, we sequenced and characterized the complete genomes of strains SKT-281, SKT-289, and LS-04. On whole genomic analysis, all three strains exhibited unique genotype constellations including both genogroup 1 and 2 genes: G3-P[8]-I2-R2-C2-M2-A2-N2-T2-E2-H2 for strains SKT-281 and SKT-289, and G2-P[8]-I2-R2-C2-M2-A2-N2-T2-E2-H2 for strain LS-04. Except for the G genotype, the unique genotype constellation of the three strains (P[8]-I2-R2-C2-M2-A2-N2-T2-E2-H2) is commonly shared with DS-1-like G1P[8] strains. On phylogenetic analysis, nine of the 11 genes of strains SKT-281 and SKT-289 (VP4, VP6, VP1-3, NSP1-3, and NSP5) appeared to have originated from DS-1-like G1P[8] strains, while the remaining VP7 and NSP4 genes appeared to be of equine and bovine origin, respectively. Thus, strains SKT-281 and SKT-289 appeared to be reassortant strains as to DS-1-like G1P[8], animal-derived human, and/or animal rotaviruses. On the other hand, seven of the 11 genes of strain LS-04 (VP7, VP6, VP1, VP3, and NSP3-5) appeared to have originated from locally circulating DS-1-like G2P[4] human rotaviruses, while three genes (VP4, VP2, and NSP1) were assumed to be derived from DS-1-like G1P[8] strains. Notably, the remaining NSP2 gene of strain LS-04 appeared to be of bovine origin. Thus, strain LS-04 was assumed to be a multiple reassortment strain as to DS-1-like G1P[8], locally circulating

  14. Isolation and partial identification of eight endogenous G1 inhibitors of JB-1 ascites tumor cell proliferation.

    PubMed

    Barfod, N M

    1982-06-01

    Eight endogenous G1 inhibitors of the proliferation of JB-1 ascites tumor cells have been isolated and characterized. The activity of the inhibitors has been analyzed on synchronized JB-1 (murine plasmacytoma) and L1A2 (murine sarcoma) cells in vitro using flow cytometry. The purified inhibitors have been tested for in vivo activity on partially synchronized JB-1 and L1A2 ascites tumors in situ. Four of the inhibitors exhibited a high degree of cell specificity (chalone-like inhibitors) and were chemically related, whereas the other four showed no cell specificity. In most extractions, the amount of cell-specific activity is more than 50% of the total G1-inhibitory activity. Most of the inhibitors are low-molecular-weight peptides and glycopeptides.

  15. Mainstream cigarette smoke exposure alters cytochrome P4502G1 expression in F344 rat olfactory mucosa

    SciTech Connect

    Hotchkiss, J.A.; Nikula, K.J.; Lewis, J.L.; Finch, G.L.; Belinsky, S.A.; Dahl, A.R.

    1994-11-01

    Inhalation of mainstream cigarette smoke (MCS) by rats results in multifocal rhinitis, mucous hypersecretion, nasal epithelial hyperplasia and metaplasia, and focal olfactory mucosal atrophy. In humans, cigarette smoking causes long-term, dose-related alterations in olfactory function in both current and former smokers. An olfactory-specific cytochrome P450 has been identified in rabbits and rats. The presence of olfactory-specific P450s, as well as relatively high levels of other biotransformation enzymes, such as NADPH-cytochrome P450 reductase and UDP-glucuronosyl transferase, in the olfactory neuroepithelium suggest that these enzyme systems may play a role in olfaction. This hypothesis is strengthened by the observation that, in rats, the temporal gene activation of P4502G1 coincides with the postnatal increase in the sensitivity of olfactory response to odorants. The purpose of this investigation was to examine the effect of MCS exposure on P4502G1 protein expression.

  16. [Role of the ABC transporters A1 and G1, key reverse cholesterol transport proteins, in atherosclerosis].

    PubMed

    Demina, E P; Miroshnikova, V V; Schwarzman, A L

    2016-01-01

    Atherosclerosis is one of the most common causes of death worldwide. Epidemiology studies firmly established an inverse relationship between atherogenesis and distorted lipid metabolism, in particular, higher levels of total cholesterol, an accumulation of CH-laden macrophages (foam cells), and lower plasma levels of antiatherogenic high density lipoprotein (HDL). It is believed that the reverse cholesterol transport, a process that removes excess cholesterol from peripheral tissues/cells including macrophages to circulating HDL, is one of the main mechanisms responsible for anti-atherogenic properties of HDL. The key proteins of reverse cholesterol transport-ATP-binding cassette transporters A1 (ABCA1) and G1 (ABCG1)-mediate the cholesterol efflux from macrophages and prevent their transformation into foam cells. This review focuses on the role of ABC transporters A1 and G1 in the pathogenesis of atherosclerosis.

  17. Reassortment of Human and Animal Rotavirus Gene Segments in Emerging DS-1-Like G1P[8] Rotavirus Strains

    PubMed Central

    Komoto, Satoshi; Tacharoenmuang, Ratana; Guntapong, Ratigorn; Ide, Tomihiko; Tsuji, Takao; Yoshikawa, Tetsushi; Tharmaphornpilas, Piyanit; Sangkitporn, Somchai; Taniguchi, Koki

    2016-01-01

    The emergence and rapid spread of novel DS-1-like G1P[8] human rotaviruses in Japan were recently reported. More recently, such intergenogroup reassortant strains were identified in Thailand, implying the ongoing spread of unusual rotavirus strains in Asia. During rotavirus surveillance in Thailand, three DS-1-like intergenogroup reassortant strains having G3P[8] (RVA/Human-wt/THA/SKT-281/2013/G3P[8] and RVA/Human-wt/THA/SKT-289/2013/G3P[8]) and G2P[8] (RVA/Human-wt/THA/LS-04/2013/G2P[8]) genotypes were identified in fecal samples from hospitalized children with acute gastroenteritis. In this study, we sequenced and characterized the complete genomes of strains SKT-281, SKT-289, and LS-04. On whole genomic analysis, all three strains exhibited unique genotype constellations including both genogroup 1 and 2 genes: G3-P[8]-I2-R2-C2-M2-A2-N2-T2-E2-H2 for strains SKT-281 and SKT-289, and G2-P[8]-I2-R2-C2-M2-A2-N2-T2-E2-H2 for strain LS-04. Except for the G genotype, the unique genotype constellation of the three strains (P[8]-I2-R2-C2-M2-A2-N2-T2-E2-H2) is commonly shared with DS-1-like G1P[8] strains. On phylogenetic analysis, nine of the 11 genes of strains SKT-281 and SKT-289 (VP4, VP6, VP1-3, NSP1-3, and NSP5) appeared to have originated from DS-1-like G1P[8] strains, while the remaining VP7 and NSP4 genes appeared to be of equine and bovine origin, respectively. Thus, strains SKT-281 and SKT-289 appeared to be reassortant strains as to DS-1-like G1P[8], animal-derived human, and/or animal rotaviruses. On the other hand, seven of the 11 genes of strain LS-04 (VP7, VP6, VP1, VP3, and NSP3-5) appeared to have originated from locally circulating DS-1-like G2P[4] human rotaviruses, while three genes (VP4, VP2, and NSP1) were assumed to be derived from DS-1-like G1P[8] strains. Notably, the remaining NSP2 gene of strain LS-04 appeared to be of bovine origin. Thus, strain LS-04 was assumed to be a multiple reassortment strain as to DS-1-like G1P[8], locally circulating

  18. Molecular characterization of cystic echinococcosis: First record of G7 in Egypt and G1 in Yemen.

    PubMed

    Alam-Eldin, Yosra H; Abdel Aaty, Heba E; Ahmed, Mona A

    2015-12-01

    Few molecular studies have identified the current status of cystic echinococcosis in Egypt. The present study aimed to ascertain the genotype(s) of Echinococcus granulosus responsible for human hydatidosis in different Egyptian governorates (regions). Animal isolates were collected from 40 camels, 5 pigs and 44 sheep. 27 human isolates were included in the present study. Specific PCR was performed and followed by DNA sequencing for mitochondrial 12S ribosomal RNA gene and BLAST analysis.The sheep cysts were not hydatid cysts. G6 genotype (camel starin) predominates in human, camel and pig isolates. G7 genotype (pig strain) was detected in two human isolates and one pig isolate. G1 genotype (sheep strain) was detected in one human isolate from Yemen and in no animal isolates. This is the first record of G7 in Egypt and G1 in Yemen.

  19. Toxocara canis glycans influence antigen recognition by mouse IgG1 and IgM antibodies.

    PubMed

    Długosz, Ewa; Wiśniewski, Marcin

    2016-01-01

    The impact of sugar moieties of Toxocara canis glycoprotein antigens on their recognition by infected mouse antibodies was investigated in this study. Native TES and recombinant Toxocara mucins generated in Pichia pastoris yeast as well as their deglycosylated forms were used in ELISA. TES and recombinant mucins were equally recognized by T. canis infected mouse IgG1 antibodies. IgM immunoglobulins predominantly recognized TES antigens. Among mucins recognition of Tc-MUC-4 was the most significant. Deglycosylation of antigens resulted in significant loss of IgM and IgG1 reactivity to TES, mucins, Tc-MUC-3 and Tc-MUC-4. The presence of sugar moieties had no influence on IgE binding to native or recombinant T. canis antigens. Our results suggest that glycans are involved in epitope formation what should be taken into consideration in production of recombinant helminth antigens for diagnostic purposes. PMID:26751891

  20. Crystal Structures of Glycosyltransferase UGT78G1 Reveal the Molecular Basis for Glycosylation and Deglycosylation of (Iso)flavonoids

    SciTech Connect

    Modolo, Luzia V.; Li, Lenong; Pan, Haiyun; Blount, Jack W.; Dixon, Richard A.; Wang, Xiaoqiang

    2010-09-21

    The glycosyltransferase UGT78G1 from Medicago truncatula catalyzes the glycosylation of various (iso)flavonoids such as the flavonols kaempferol and myricetin, the isoflavone formononetin, and the anthocyanidins pelargonidin and cyanidin. It also catalyzes a reverse reaction to remove the sugar moiety from glycosides. The structures of UGT78G1 bound with uridine diphosphate or with both uridine diphosphate and myricetin were determined at 2.1 {angstrom} resolution, revealing detailed interactions between the enzyme and substrates/products and suggesting a distinct binding mode for the acceptor/product. Comparative structural analysis and mutagenesis identify glutamate 192 as a key amino acid for the reverse reaction. This information provides a basis for enzyme engineering to manipulate substrate specificity and to design effective biocatalysts with glycosylation and/or deglycosylation activity.

  1. Comparison of carbohydrate and peptide biotinylation on the immunological activity of IgG1 murine monoclonal antibodies.

    PubMed

    Miralles, F; Takeda, Y; Escribano, M J

    1991-07-01

    When the classical amino acid esterification procedure was used for the biotinylation of the IgG1 monoclonal antibody J28 it resulted in a loss of immunological activity. This antibody recognizes the fetoacinar pancreatic (FAP) antigen and the decrease in reactivity was directly proportional to the molar biotin/antibody ratio indicating substitutions at or near the antibody combining site. This effect was specific to J28 since the IgG1 Mab F22 which recognises the same antigen was not damaged by this procedure. Active Mab J28 conjugates were obtained using biotinylation via oligosaccharide moieties. The biotinylation efficiency using this method was dependent on the previous degree of antibody periodate oxidation and the maximal substitution was 3 mol biotin per mol of antibody. Using these conditions the sensitivity of the biotinylated J28 for the FAP antigen was similar to that obtained when using non-substituted antibody in the two antibodies technique.

  2. A novel peptide sansalvamide analogue inhibits pancreatic cancer cell growth through G0/G1 cell-cycle arrest

    SciTech Connect

    Ujiki, Michael B. |; Milam, Ben; Ding Xianzhong |; Roginsky, Alexandra B.; Salabat, M. Reza; Talamonti, Mark S.; Bell, Richard H. |; Gu Wenxin; Silverman, Richard B. ||; Adrian, Thomas E. |. E-mail: tadrian@northwestern.edu

    2006-02-24

    Patients with pancreatic cancer have little hope for cure because no effective therapies are available. Sansalvamide A is a cyclic depsipeptide produced by a marine fungus. We investigated the effect of a novel sansalvamide A analogue on growth, cell-cycle phases, and induction of apoptosis in human pancreatic cancer cells in vitro. The sansalvamide analogue caused marked time- and concentration-dependent inhibition of DNA synthesis and cell proliferation of two human pancreatic cancer cell lines (AsPC-1 and S2-013). The analogue induced G0/G1 phase cell-cycle arrest and morphological changes suggesting induction of apoptosis. Apoptosis was confirmed by annexin V binding. This novel sansalvamide analogue inhibits growth of pancreatic cancer cells through G0/G1 arrest and induces apoptosis. Sansalvamide analogues may be valuable for the treatment of pancreatic cancer.

  3. G-1 exerts neuroprotective effects through G protein-coupled estrogen receptor 1 following spinal cord injury in mice.

    PubMed

    Cheng, Qiang; Meng, Jia; Wang, Xin-Shang; Kang, Wen-Bo; Tian, Zhen; Zhang, Kun; Liu, Gang; Zhao, Jian-Ning

    2016-08-01

    Spinal cord injury (SCI) always occurs accidently and leads to motor dysfunction because of biochemical and pathological events. Estrogen has been shown to be neuroprotective against SCI through estrogen receptors (ERs), but the underlying mechanisms have not been fully elucidated. In the present study, we investigated the role of a newly found membrane ER, G protein-coupled estrogen receptor 1 (GPR30 or GPER1), and discussed the feasibility of a GPR30 agonist as an estrogen replacement. Forty adult female C57BL/6J mice (10-12 weeks old) were divided randomly into vehicle, G-1, E2, G-1 + G-15 and E2 + G-15 groups. All mice were subjected to SCI using a crushing injury approach. The specific GPR30 agonist, G-1, mimicked the effects of E2 treatment by preventing SCI-induced apoptotic cell death and enhancing motor functional recovery after injury. GPR30 activation regulated phosphatidylinositol 3-kinase (PI3K)/Akt and MAPK/extracellular signal-regulated kinase (ERK) signalling pathways, increased GPR30 and anti-apoptosis proteins Bcl-2 and brain derived neurotrophic factor (BDNF), but decreased the pro-apoptosis factor Bax and cleaved caspase-3. However, the neuroprotective effects of G-1 and E2 were blocked by the specific GPR30 antagonist, G-15. Thus, GPR30 rather than classic ERs is required to induce estrogenic neuroprotective effects. Given that estrogen replacement therapy may cause unexpected side effects, especially on the reproductive system, GPR30 agonists may represent a potential therapeutic approach for treating SCI. PMID:27407175

  4. Detailed petrographic descriptions and microprobe data for tertiary silicic volcanic rocks in drill hole USW G-1, Yucca Mountain, Nevada

    SciTech Connect

    Caporuscio, F.A.; Warren, R.G.; Broxton, D.E.

    1985-12-01

    This report contains detailed petrographic descriptions of 74 thin sections from drill hole USW G-1 at Yucca Mountain, Nevada. These descriptions are keyed to the distinctions between devitrified, vitrophyre, vitric, and zeolitized intervals below the Topopah Spring Member repository horizon. The petrographic features of the zeolitized intervals down through the Crater Flat tuff, as well as the sorption properties determined from these intervals, suggest that these zeolite occurrences may each have comparable sorptive capability.

  5. Selective Subnormal IgG1 in 54 Adult Index Patients with Frequent or Severe Bacterial Respiratory Tract Infections

    PubMed Central

    Barton, James C.; Bertoli, Luigi F.; Barton, J. Clayborn; Acton, Ronald T.

    2016-01-01

    We characterized 54 adult index patients with reports of frequent or severe bacterial respiratory tract infections at diagnosis of selective subnormal IgG1. Mean age was 50 ± 13 (SD) y; 87.0% were women. Associated disorders included the following: autoimmune conditions 50.0%; hypothyroidism 24.1%; atopy 38.9%; and other allergy 31.5%. In 35.5%, proportions of protective S. pneumoniae serotype-specific IgG levels did not increase after polyvalent pneumococcal polysaccharide vaccination (PPPV). Blood lymphocyte subset levels were within reference limits in most patients. Regressions on IgG1 and IgG3 revealed no significant association with age, sex, autoimmune conditions, hypothyroidism, atopy, other allergy, corticosteroid therapy, or lymphocyte subsets. Regression on IgG2 revealed significant associations with PPPV response (negative) and CD19+ lymphocytes (positive). Regression on IgG4 revealed significant positive associations with episodic corticosteroid use and IgA. Regression on IgA revealed positive associations with IgG2 and IgG4. Regression on IgM revealed negative associations with CD56+/CD16+ lymphocytes. Regressions on categories of infection revealed a negative association of urinary tract infections and IgG1. HLA-A⁎03, HLA-B⁎55 and HLA-A⁎24, HLA-B⁎35 haplotype frequencies were greater in 38 patients than 751 controls. We conclude that nonprotective S. pneumoniae IgG levels and atopy contribute to increased susceptibility to respiratory tract infections in patients with selective subnormal IgG1. PMID:27123464

  6. Androgen receptor regulates Cdc6 in synchronized LNCaP cells progressing from G1 to S phase.

    PubMed

    Bai, V Uma; Cifuentes, Eugenia; Menon, Mani; Barrack, Evelyn R; Reddy, G Prem-Veer

    2005-08-01

    We have shown previously that androgen receptor (AR) activity is required for the progression of cells from G(1) to S phase. In an attempt to elucidate the mechanism of androgen- and androgen-receptor-mediated proliferation of prostate cancer cells, we studied the effect of anti-androgen bicalutamide (Casodex) on the expression of cell-cycle regulatory genes in synchronized LNCaP cells progressing from G(1) to S phase. LNCaP cells were synchronized by isoleucine-deprivation. Expression of cell-cycle regulatory genes in S phase control cells versus Casodex-treated cells that fail to enter S phase was studied using a microarray containing cDNA probes for 111 cell-cycle specific genes. RT-PCR and Western-blots were used to validate microarray data. Casodex blocked synchronized LNCaP cells from entering S phase. Microarrays revealed downregulation of eight genes in cells prevented from entering into S phase by Casodex. Of these eight genes, only Cdc6, cyclin A, and cyclin B were downregulated at both the mRNA and protein level in Casodex treated cells as compared to control cells. The mRNA and protein levels of Cdc6 increased as synchronized LNCaP cells progressed from G(1) to S phase, and were attenuated in Casodex-treated cells failed to enter S phase. Cyclins A and B were detected when cells entered S phase, but not when they were in G(1) phase. Like Cdc6, the levels of both cyclins A and B were attenuated in Casodex-treated cells. AR may play an important role in the onset of DNA synthesis in prostate cancer cells by regulating the expression and stability of Cdc6, which is critically required for the assembly of the pre-replication complex(pre-RC).

  7. Echinococcus ortleppi (G5) and Echinococcus granulosus sensu stricto (G1) loads in cattle from Southern Brazil.

    PubMed

    Balbinotti, Helier; Santos, Guilherme B; Badaraco, Jeferson; Arend, Ana C; Graichen, Daniel Ângelo S; Haag, Karen L; Zaha, Arnaldo

    2012-09-10

    Echinococcus granulosus sensu stricto (G1) and Echinococcus ortleppi (G5) are haplotypes of the parasite formerly known as Echinococcus granulosus sensu lato, which in its larval stage causes cystic hydatid disease, endemic in Southern Brazil. Epidemiological and molecular knowledge about the haplotypes occurring in a region is essential to control the spread of the disease. The aim of this work was to analyze the haplotype frequency and fertility of hydatid cysts in cattle from the state of Rio Grande do Sul. Cysts were collected and classified according to their fertility status. DNA was extracted from protoscoleces and germinal layers and then used as template for the amplification of the cytochrome c oxidase subunit 1 gene by PCR. Amplicons were purified and sequenced, and the sequences were analyzed for haplotype identification. A total of 638 fertile cysts collected in the last ten years were genotyped. On average, G1 (56.6%) was more frequent than G5 (43.4%). In lungs, the G5 haplotype exhibited a higher parasite load (52.8%), whereas in the liver, G1 was more frequent (90.4%). The analysis revealed an increase in the frequency of G5 haplotype cysts during the period of sampling, and an increase in the abundance of fertile cysts has also been observed in the last several years. Most infertile cysts were genotyped as G1. The possible factors involved in the increase in the proportion of E. ortleppi (G5) and the consequences of this increase are discussed. This study suggests that the proportion of E. ortleppi (G5) loads in cattle may be increasing overtime.

  8. G-1 exerts neuroprotective effects through G protein-coupled estrogen receptor 1 following spinal cord injury in mice

    PubMed Central

    Cheng, Qiang; Meng, Jia; Wang, Xin-shang; Kang, Wen-bo; Tian, Zhen; Zhang, Kun; Liu, Gang; Zhao, Jian-ning

    2016-01-01

    Spinal cord injury (SCI) always occurs accidently and leads to motor dysfunction because of biochemical and pathological events. Estrogen has been shown to be neuroprotective against SCI through estrogen receptors (ERs), but the underlying mechanisms have not been fully elucidated. In the present study, we investigated the role of a newly found membrane ER, G protein-coupled estrogen receptor 1 (GPR30 or GPER1), and discussed the feasibility of a GPR30 agonist as an estrogen replacement. Forty adult female C57BL/6J mice (10–12 weeks old) were divided randomly into vehicle, G-1, E2, G-1 + G-15 and E2 + G-15 groups. All mice were subjected to SCI using a crushing injury approach. The specific GPR30 agonist, G-1, mimicked the effects of E2 treatment by preventing SCI-induced apoptotic cell death and enhancing motor functional recovery after injury. GPR30 activation regulated phosphatidylinositol 3-kinase (PI3K)/Akt and MAPK/extracellular signal-regulated kinase (ERK) signalling pathways, increased GPR30 and anti-apoptosis proteins Bcl-2 and brain derived neurotrophic factor (BDNF), but decreased the pro-apoptosis factor Bax and cleaved caspase-3. However, the neuroprotective effects of G-1 and E2 were blocked by the specific GPR30 antagonist, G-15. Thus, GPR30 rather than classic ERs is required to induce estrogenic neuroprotective effects. Given that estrogen replacement therapy may cause unexpected side effects, especially on the reproductive system, GPR30 agonists may represent a potential therapeutic approach for treating SCI. PMID:27407175

  9. Expression of CAR in SW480 and HepG2 cells during G1 is associated with cell proliferation

    SciTech Connect

    Osabe, Makoto; Sugatani, Junko Takemura, Akiko; Yamazaki, Yasuhiro; Ikari, Akira; Kitamura, Naomi; Negishi, Masahiko; Miwa, Masao

    2008-05-16

    Constitutive androstane receptor (CAR) is a transcription factor to regulate the expression of several genes related to drug-metabolism. Here, we demonstrate that CAR protein accumulates during G1 in human SW480 and HepG2 cells. After the G1/S phase transition, CAR protein levels decreased, and CAR was hardly detected in cells by the late M phase. CAR expression in both cell lines was suppressed by RNA interference-mediated suppression of CDK4. Depletion of CAR by RNA interference in both cells and by hepatocyte growth factor treatment in HepG2 cells resulted in decreased MDM2 expression that led to p21 upregulation and repression of HepG2 cell growth. Thus, our results demonstrate that CAR expression is an early G1 event regulated by CDK4 that contributes to MDM2 expression; these findings suggest that CAR may influence the expression of genes involved in not only the metabolism of endogenous and exogenous substances but also in the cell proliferation.

  10. Downregulation of FOXP1 Inhibits Cell Proliferation in Hepatocellular Carcinoma by Inducing G1/S Phase Cell Cycle Arrest

    PubMed Central

    Wang, Xin; Sun, Ji; Cui, Meiling; Zhao, Fangyu; Ge, Chao; Chen, Taoyang; Yao, Ming; Li, Jinjun

    2016-01-01

    Forkhead box P1 (FOXP1) belongs to a family of winged-helix transcription factors that are involved in the processes of cellular proliferation, differentiation, metabolism, and longevity. FOXP1 can affect cell proliferation and migratory ability in hepatocellular carcinoma (HCC) in vitro. However, little is known about the mechanism of FOXP1 in the proliferation of HCC cells. This study aimed to further explore the function of FOXP1 on the proliferation of HCC cells as well as the relevant mechanism involved. Western blot analysis, tumor xenograft models, and flow cytometry analysis were performed to elucidate the function of FOXP1 in the regulation of cell proliferation in human HCC. We observed that silencing FOXP1 significantly suppressed the growth ability of HCC cells both in vitro and in vivo. In addition, knockdown of FOXP1 induced G1/S phase arrest, and the expression of total and phosphorylated Rb (active type) as well as the levels of E2F1 were markedly decreased at 24 h; however, other proteins, including cyclin-dependent kinase (CDK) 4 and 6 and cyclin D1 did not show noticeable changes. In conclusion, downregulation of FOXP1 inhibits cell proliferation in hepatocellular carcinoma by inducing G1/S phase cell cycle arrest, and the decrease in phosphorylated Rb is the main contributor to this G1/S phase arrest. PMID:27618020

  11. Downregulation of FOXP1 Inhibits Cell Proliferation in Hepatocellular Carcinoma by Inducing G1/S Phase Cell Cycle Arrest.

    PubMed

    Wang, Xin; Sun, Ji; Cui, Meiling; Zhao, Fangyu; Ge, Chao; Chen, Taoyang; Yao, Ming; Li, Jinjun

    2016-01-01

    Forkhead box P1 (FOXP1) belongs to a family of winged-helix transcription factors that are involved in the processes of cellular proliferation, differentiation, metabolism, and longevity. FOXP1 can affect cell proliferation and migratory ability in hepatocellular carcinoma (HCC) in vitro. However, little is known about the mechanism of FOXP1 in the proliferation of HCC cells. This study aimed to further explore the function of FOXP1 on the proliferation of HCC cells as well as the relevant mechanism involved. Western blot analysis, tumor xenograft models, and flow cytometry analysis were performed to elucidate the function of FOXP1 in the regulation of cell proliferation in human HCC. We observed that silencing FOXP1 significantly suppressed the growth ability of HCC cells both in vitro and in vivo. In addition, knockdown of FOXP1 induced G1/S phase arrest, and the expression of total and phosphorylated Rb (active type) as well as the levels of E2F1 were markedly decreased at 24 h; however, other proteins, including cyclin-dependent kinase (CDK) 4 and 6 and cyclin D1 did not show noticeable changes. In conclusion, downregulation of FOXP1 inhibits cell proliferation in hepatocellular carcinoma by inducing G1/S phase cell cycle arrest, and the decrease in phosphorylated Rb is the main contributor to this G1/S phase arrest. PMID:27618020

  12. MKP1 phosphatase mediates G1-specific dephosphorylation of H3Serine10P in response to DNA damage.

    PubMed

    Sharma, Ajit K; Khan, Shafqat A; Sharda, Asmita; Reddy, Divya V; Gupta, Sanjay

    2015-08-01

    Histone mark, H3S10 phosphorylation plays a dual role in a cell by maintaining relaxed chromatin for active transcription in interphase and condensed chromatin state in mitosis. The level of H3S10P has also been shown to alter on DNA damage; however, its cell cycle specific behavior and regulation during DNA damage response is largely unexplored. In the present study, we demonstrate G1 cell cycle phase specific reversible loss of H3S10P in response to IR-induced DNA damage is mediated by opposing activities of phosphatase, MKP1 and kinase, MSK1 of the MAP kinase pathway. We also show that the MKP1 recruits to the chromatin in response to DNA damage and correlates with the decrease of H3S10P, whereas MKP1 is released from chromatin during recovery phase of DDR. Furthermore, blocking of H3S10 dephosphorylation by MKP1 inhibition impairs DNA repair process and results in poor survival of WRL68 cells. Collectively, our data proposes a pathway regulating G1 cell cycle phase specific reversible reduction of H3S10P on IR induced DNA damage and also raises the possibility of combinatorial modulation of H3S10P with specific inhibitors to target the cancer cells in G1-phase of cell cycle.

  13. Cysteine protease of the nematode Nippostrongylus brasiliensis preferentially evokes an IgE/IgG1 antibody response in rats.

    PubMed Central

    Kamata, I; Yamada, M; Uchikawa, R; Matsuda, S; Arizono, N

    1995-01-01

    Some cysteine proteases such as papain and those of mites and schistosomes have potent allergenic properties. To clarify the allergenicity of nematode cysteine proteases, the enzyme was purified from the intestinal nematode Nippostrongylus brasiliensis using cation exchange chromatography and gel filtration chromatography. The purified protease, of 16 kD and pI 8.5, showed maximum enzyme activity at pH 5.5 and substrate preference for Z-Phe-Arg-MCA. The specific inhibitors of cysteine protease leupeptin, iodoacetic acid, and E-64, completely suppressed the activity, indicating that the purified enzyme belongs to the cysteine protease family. Cysteine protease activity was found not only in somatic extract, but also in the excretory-secretory (ES) product of the nematode. When anti-cysteine protease immunoglobulin isotypes were examined in sera from rats infected with N. brasiliensis, a high level of IgG1 and a lower level of IgE antibody were detected. Depletion of IgG antibodies from the sera using protein G affinity columns resulted in a marked increase in reactivity of anti-cysteine protease IgE with the antigen, possibly due to the removal of competing IgG antibodies. In contrast to IgE and IgG1, production of anti-cysteine protease IgG2a was negligible. These results indicate that the nematode cysteine protease preferentially evokes an IgE/IgG1 antibody response. Images Fig. 2 PMID:7554403

  14. PTEN/MMAC1/TEP1 suppresses the tumorigenicity and induces G1 cell cycle arrest in human glioblastoma cells

    PubMed Central

    Li, Da-Ming; Sun, Hong

    1998-01-01

    PTEN/MMAC1/TEP1 is a tumor suppressor that possesses intrinsic phosphatase activity. Deletions or mutations of its encoding gene are associated with a variety of human cancers. However, very little is known about the molecular mechanisms by which this important tumor suppressor regulates cell growth. Here, we show that PTEN expression potently suppressed the growth and tumorigenicity of human glioblastoma U87MG cells. The growth suppression activity of PTEN was mediated by its ability to block cell cycle progression in the G1 phase. Such an arrest correlated with a significant increase of the cell cycle kinase inhibitor p27KIP1 and a concomitant decrease in the activities of the G1 cyclin-dependent kinases. PTEN expression also led to the inhibition of Akt/protein kinase B, a serine-threonine kinase activated by the phosphatidylinositol 3-kinase (PI 3-kinase) signaling pathway. In addition, the effect of PTEN on p27KIP1 and the cell cycle can be mimicked by treatment of U87MG cells with LY294002, a selective inhibitor of PI 3-kinase. Taken together, our studies suggest that the PTEN tumor suppressor modulates G1 cell cycle progression through negatively regulating the PI 3-kinase/Akt signaling pathway, and one critical target of this signaling process is the cyclin-dependent kinase inhibitor p27KIP1. PMID:9860981

  15. Singularity-conquering ZG controllers of z2g1 type for tracking control of the IPC system

    NASA Astrophysics Data System (ADS)

    Zhang, Yunong; Yu, Xiaotian; Yin, Yonghua; Peng, Chen; Fan, Zhengping

    2014-09-01

    With wider investigations and applications of autonomous robotics and intelligent vehicles, the inverted pendulum on a cart (IPC) system has become more attractive for numerous researchers due to its concise and representative structure. In this article, the tracking-control problem of the IPC system is considered and investigated. Based on Zhang dynamics (ZD) and gradient dynamics (GD), a novel kind of ZG controllers are developed and investigated for achieving the tracking-control purpose, which contains controllers of z2g0 and z2g1 types according to the number of times of using the ZD and GD methods. Besides, theoretical analyses are presented to guarantee the global and exponential convergence performance of both z2g0 and z2g1 controllers. Computer simulations are further performed to substantiate the feasibility and effectiveness of ZG controllers. More importantly, comparative simulation results demonstrate that controllers of z2g1 type can conquer the singularity problem (i.e. the division-by-zero problem).

  16. Development of hyperbranched polymers with non-covalent interactions for extraction and determination of aflatoxins in cereal samples.

    PubMed

    Liu, Xiaoyan; Li, Huihui; Xu, Zhigang; Peng, Jialin; Zhu, Shuqiang; Zhang, Haixia

    2013-10-01

    A novel approach for assembling homogeneous hyperbranched polymers based on non-covalent interactions with aflatoxins was developed; the polymers were used to evaluate the extraction of aflatoxins B1, B2, G1 and G2 (AFB1, AFB2, AFG1 and AFG2) in simulant solutions. The results showed that the extraction efficiencies of three kinds of synthesized polymers for the investigated analytes were not statistically different; as a consequence, one of the representative polymers (polymer I) was used as the solid-phase extraction (SPE) sorbent to evaluate the influences of various parameters, such as desorption conditions, pH, ionic strength, concentration of methanol in sample solutions, and the mass of the sorbent on the extraction efficiency. In addition, the extraction efficiencies for these aflatoxins were compared between the investigated polymer and the traditional sorbent C18. The results showed that the investigated polymer had superior extraction efficiencies. Subsequently, the proposed polymer for the SPE packing material was employed to enrich and analyze four aflatoxins in the cereal powder samples. The limits of detection (LODs) at a signal-to-noise (S/N) ratio of 3 were in the range of 0.012-0.120 ng g(-1) for four aflatoxins, and the limits of quantification (LOQs) calculated at S/N=10 were from 0.04 to 0.40 ng g(-1) for four aflatoxins. The recoveries of four aflatoxins from cereal powder samples were in the range of 82.7-103% with relative standard deviations (RSDs) lower than 10%. The results demonstrate the suitability of the SPE approach for the analysis of trace aflatoxins in cereal powder samples.

  17. Asteroid phase curve analysis with the H, G 1, G 2 photometric phase function: application to the PTF survey observations

    NASA Astrophysics Data System (ADS)

    Penttilä, Antti; Cellino, Alberto; Lu, Xiaoping; Shevchenko, Vasilij G.; Muinonen, Karri

    2016-10-01

    Estimation of an asteroid's absolute magnitude H from its photometry is extremely important. The absolute magnitude relates the brightness of the asteroid to its size, if the geometric albedo is known. The shape of the phase curve can serve as a proxy for the taxonomic type of the asteroid in cases with no spectral information available [1,2].In 2012, the IAU adopted the H,G1,G2 function to replace the H,G function for phase curve analysis [3]. This new function improves the backscattering behavior of the curve with high- and low-albedo asteroids. The phase function (PF) can be applied to asteroids with multiple high-quality observations. If the number of observations is small, or their accuracy is low, problems may arise. The most apparent problem is that the parameter G or the parameters G1, G2 might be poorly estimated. The solution has been to fix to value of G or values of G1, G2 and estimate only the H. In our recent work [4], we offer a solution that can improve the current situation with the photometric fits with a small number of low-accuracy observations. We present a constrained nonlinear least-squares method for fitting the H,G1,G2 function that can improve the possible bias with low-accuracy data. Then, we revisit the two-parameter PF with new data and offer a new version, the H,G12* PF. Finally, we assess the problem with fixed G or G1, G2 parameters by introducing one-parameter models that relate to five taxonomic asteroid groups. We tie all the models together with three or two parameters, or a single parameter, with a statistical model selection procedure to select the best version for a particular data set.We have developed practical tools for the abovementioned algorithms. We apply the tools to a dataset of 8,900 asteroids with almost 500,000 photometric observations from the Palomar Transient Factory survey [5]. We report the effect of the revised H estimates on the geometric albedos in cases where WISE-mission size estimates are available.[1] D

  18. Hepatitis C virus G1b infection decreases the number of small low-density lipoprotein particles

    PubMed Central

    Kinoshita, Chika; Nagano, Tomohisa; Seki, Nobuyoshi; Tomita, Yoichi; Sugita, Tomonori; Aida, Yuta; Itagaki, Munenori; Satoh, Kenichi; Sutoh, Satoshi; Abe, Hiroshi; Tsubota, Akihito; Aizawa, Yoshio

    2016-01-01

    AIM: To investigate how hepatitis C virus (HCV) G1b infection influences the particle number of lipoproteins. METHODS: The numbers of lipoprotein particles in fasting sera from 173 Japanese subjects, 82 with active HCV G1b infection (active HCV group) and 91 with cleared HCV infection (SVR group), were examined. Serum lipoprotein was fractionated by high-performance liquid chromatography into twenty fractions. The cholesterol and triglyceride concentrations in each fraction were measured using LipoSEARCH. The number of lipoprotein particles in each fraction was calculated using a newly developed algorithm, and the relationship between chronic HCV G1b infection and the lipoprotein particle number was determined by multiple linear regression analysis. RESULTS: The median number of low-density lipoprotein (LDL) particles was significantly lower in the active HCV group [1182 nmol/L, interquartile range (IQR): 444 nmol/L] than in the SVR group (1363 nmol/L, IQR: 472 nmol/L, P < 0.001), as was that of high-density lipoprotein (HDL) particles (14168 nmol/L vs 15054 nmol/L, IQR: 4114 nmol/L vs 3385 nmol/L, P = 0.042). The number of very low-density lipoprotein (VLDL) particles was similar between the two groups. Among the four LDL sub-fractions, the number of large LDL particles was similar between the two groups. However, the numbers of medium (median: 533.0 nmol/L, IQR: 214.7 nmol/L vs median: 633.5 nmol/L, IQR: 229.6 nmol/L, P < 0.001), small (median: 190.9 nmol/L, IQR: 152.4 nmol/L vs median: 263.2 nmol/L, IQR: 159.9 nmol/L; P < 0.001), and very small LDL particles (median: 103.5 nmol/L, IQR: 66.8 nmol/L vs median: 139.3 nmol/L, IQR: 67.3 nmol/L, P < 0.001) were significantly lower in the active HCV group than in the SVR group, respectively. Multiple linear regression analysis indicated an association between HCV G1b infection and the decreased numbers of medium, small, and very small LDL particles. However, active HCV infection did not affect the number of large LDL

  19. Mayall II=G1 in M31: Giant Globular Cluster or Core of a Dwarf Elliptical Galaxy?

    NASA Astrophysics Data System (ADS)

    Meylan, G.; Sarajedini, A.; Jablonka, P.; Djorgovski, S. G.; Bridges, T.; Rich, R. M.

    2001-08-01

    Mayall II=G1 is one of the brightest globular clusters belonging to M31, the Andromeda galaxy. Our observations with the Wide Field and Planetary Camera (WFPC2) on board the Hubble Space Telescope (HST) provide photometric data for the I versus V-I and V versus V-I color-magnitude diagrams. They reach stars with magnitudes fainter than V=27 mag, with a well populated red horizontal branch at about V=25.3 mag. From model fitting, we determine a rather high mean metallicity of [Fe/H]=-0.95+/-0.09, somewhat similar to 47 Tucanae. In order to determine our true measurement errors, we have carried out artificial star experiments. We find a larger spread in V-I than can be explained by the measurement errors, and we attribute this to an intrinsic metallicity dispersion amongst the stars of G1; this may be the consequence of self-enrichment during the early stellar/dynamical evolutionary phases of this cluster. So far, only ω Centauri, the giant Galactic globular cluster, has been known to exhibit such an intrinsic metallicity dispersion, a phenomenon certainly related to the deep potential wells of these two star clusters. We determine, from the same HST/WFPC2 data, the structural parameters of G1. Its surface brightness profile provides its core radius rc=0.14"=0.52 pc, its tidal radius rt~=54''=200 pc, and its concentration c=log(rt/rc)~=2.5. Such a high concentration indicates the probable collapse of the core of G1. KECK/HIRES observations provide the central velocity dispersion σobs=25.1 km s-1, with σp(0)=27.8 km s-1 once aperture corrected. Three estimates of the total mass of this globular cluster can be obtained. The King-model mass is MK=15×106 Msolar with M/LV~=7.5, and the virial mass is MVir=7.3×106 Msolar with M/LV~=3.6. By using a King-Michie model fitted simultaneously to the surface brightness profile and the central velocity dispersion value, mass estimates range from MKM=14×106 Msolar to 17×106 Msolar. Although uncertain, all of these mass

  20. Galectin-3 binds highly galactosylated IgG1 and is crucial for the IgG1 complex mediated inhibition of C5aReceptor induced immune responses.

    PubMed

    Heyl, Kerstin A; Karsten, Christian M; Slevogt, Hortense

    2016-10-01

    Changes in the glycosylation of immunoglobulins have been shown to modulate immune homeostasis and disease pathology. In this sense it has been shown that highly galactosylated but not agalactosylated IgG1 immune complexes (ICs) inhibit C5aR-mediated pro-inflammatory immune responses via the assembly of FcγRIIB-Dectin-1 receptor complexes. In this study we demonstrated that Galectin-3, a galactose-binding lectin that is known to cross-link proteins on cell-surfaces via binding their N-glycans, bound to highly-galactosylated, but not agalactosylated IgG1. Further, Galectin-3 was essential for the IC-mediated inhibition of C5a-induced neutrophil chemotaxis in vitro. Taken together our results indicate that Galectin-3 mediates the interaction of ICs with the FcγRIIB-Dectin-1 receptor complex for delivering immunoregulatory signals to inhibit C5aR-mediated immune responses. PMID:27620493

  1. Impacts, effectiveness and regional inequalities of the GeoMIP G1 to G4 solar radiation management scenarios

    NASA Astrophysics Data System (ADS)

    Yu, Xiaoyong; Moore, John C.; Cui, Xuefeng; Rinke, Annette; Ji, Duoying; Kravitz, Ben; Yoon, Jin-Ho

    2015-06-01

    We evaluate the effectiveness and the regional inequalities of solar radiation management (SRM) in compensating for simultaneous changes in temperature and precipitation caused by increased greenhouse gas concentrations. We analyze the results from Earth System Models under four Geoengineering Model Intercomparison Project (GeoMIP) experiments with a modified form of the Residual Climate Response approach. Each experiment produces 50 model yrs of simulations: 13 models completed experiment G1 (offsetting 4 × CO2 via solar reduction); 12 models completed experiment G2 (offsetting CO2 that increased by 1% per year); 3 models completed experiment G3 (offsetting increasing radiative forcing under RCP4.5 with increasing stratospheric aerosol); and 7 models completed experiment G4 (injection of 5 Tg SO2 a- 1 into the stratosphere). The regional inequalities in temperature and precipitation compensation for experiments G1, G3 and G4 are significantly different from their corresponding noise backgrounds for most models, but for G2 they are not significantly different from noise. Differences in the regional inequalities and the actual effectiveness among the four SRM scenarios are not significant for many models. However, in more than half of the models, the effectiveness for temperature in the solar dimming geoengineering scenarios (G1 and G2) is significantly higher than that in the SO2 geoengineering scenarios (G3 and G4). The effectiveness of the four SRM experiments in compensating for temperature change is considerably higher than for precipitation. The methodology used highlights that a large across-model variation in the treatment of key geoengineering processes (such as stratospheric aerosols) and the quantification of damage caused by climate change creates significant uncertainties in any strategies to achieve optimal compensation effectiveness across different regions.

  2. Cyclin D1 is dispensable for G1 control in retinoblastoma gene-deficient cells independently of cdk4 activity.

    PubMed Central

    Lukas, J; Bartkova, J; Rohde, M; Strauss, M; Bartek, J

    1995-01-01

    To elucidate the regulator-versus-target relationship in the cyclin D1/cdk4/retinoblastoma protein (pRB) pathway, we examined fibroblasts from RB-1 gene-deficient and RB-1 wild-type littermate mouse embryos (ME) and in human tumor cell lines that differed in the status of the RB-1 gene. The RB+/+ and RB-/- ME fibroblasts expressed similar protein levels of D-type cyclins, cdk4, and cdk6, showed analogous spectra and abundance of cellular proteins complexed with cdk4 and/or cyclins D1 and D2, and exhibited comparable associated kinase activities. Of the two human cell lines established from the same sarcoma biopsy, the RB-positive SKUT1B cells contained cdk4 that was mainly associated with D-type cyclins, contrary to a predominant cdk4-p16INK4 complex in the RB-deficient SKUT1A cells. Antibody-mediated neutralization of cyclin D1 arrested the RB-positive ME and SKUT1B cells in G1, whereas this cyclin appeared dispensable in the RB-deficient ME and SKUT1A cells. Lack of requirement for cyclin D1 therefore correlated with absence of functional pRB, regardless of whether active cyclin D1/cdk4 holoenzyme was present in the cells under study. Consistent with a potential role of cyclin D/cdk4 in phosphorylation of pRB, monoclonal anti-cyclin D1 antibodies supporting the associated kinase activity failed to significantly affect proliferation of RB-positive cells, whereas the antibody DCS-6, unable to coprecipitate cdk4, efficiently inhibited G1 progression and prevented pRB phosphorylation in vivo. These data provide evidence for an upstream control function of cyclin D1/cdk4, and a downstream role for pRB, in the order of events regulating transition through late G1 phase of the mammalian cell division cycle. PMID:7739541

  3. Interaction of KIR genes and G1M immunoglobulin allotypes confer susceptibility to type 2 diabetes in Puerto Rican Americans.

    PubMed

    Zuniga, Joaquin; Romero, Viviana; Azocar, Jose; Stern, Joel N H; Clavijo, Olga; Almeciga, Ingrid; Encinales, Liliana; Avendano, Angel; Fridkis-Hareli, Masha; Pandey, Janardan P; Yunis, Edmond J

    2006-11-01

    The susceptibility to type 2 diabetes (T2D) involves genetic factors. We studied the distribution of KIR and MHC class I ligands phenotype and genotype frequencies, as well as immunoglobulin KM and GM allotype frequencies in a group of patients (N = 95) with T2D and ethnically matched healthy controls (N = 74) with Puerto Rican ethnic background. We found a slight increase of the 2DL3/2DL3 homozygous genotype in T2D. Moreover, the association between 2DL3/2DL3 genotype was significant in the presence of 2DS4 (pC = 0.01). Also, we observed an epistatic effect of the interaction of 2DL3/2DL3, 2DS4 with allele z of G1M in T2D (pC = 0.004, OR = 3.60, 95% CI, 1.62-8.10). This genetic interaction between KIR and G1M allotypes, associated with T2D, was also significant by multiple logistic regression analysis (p < 0.0001, OR = 4.90, 95% CI, 2.12-11.3). We did not detect population stratification using unlinked short tandem repeat (STR) markers, demonstrating that the patients and controls were ethnically matched. Hence, we have demonstrated in this study an epistatic interaction between KIR genes and the G1M allotype that influences the susceptibility to T2D in Puerto Rican Americans. Our findings are important for understanding the autoimmune or innate immune inflammatory-mediated mechanisms involved in the pathogenesis of T2D.

  4. Jacalin interaction with human immunoglobulin A1 and bovine immunoglobulin G1: affinity constant determined by piezoelectric biosensoring.

    PubMed

    Pedroso, Mariele M; Pesquero, Naira C; Thomaz, Sandra M O; Roque-Barreira, Maria C; Faria, Ronaldo C; Bueno, Paulo R

    2012-03-01

    The affinity of the D-galactose-binding lectin from Artocarpus heterophyllus lectin, known as jacalin, with immonuglobulins (Igs) was determined by biofunctionalization of a piezoelectric transducer. This piezoelectric biofunctionalized transducer was used as a mass-sensitive analytical tool, allowing the real-time binding analysis of jacalin-human immunoglobulin A1 (IgA(1)) and jacalin-bovine IgG(1) interactions from which the apparent affinity constant was calculated. The strategy was centered in immobilizing jacalin on the gold electrode's surface of the piezoelectric crystal resonator using appropriate procedures based on self-assembling of 11-mercaptoundecanoic acid and 2-mercaptoethanol thiol's mixture, a particular immobilization strategy by which it was possible to avoid cross-interaction between the proteins over electrode's surface. The apparent affinity constants obtained between jacalin-human IgA(1) and jacalin-bovine IgG(1) differed by 1 order of magnitude [(8.0 ± 0.9) 10(5) vs (8.3 ± 0.1) 10(6) L mol(-1)]. On the other hand, the difference found between human IgA(1) and human IgA(2) interaction with jacalin, eight times higher for IgA(1), was attributed to the presence of O-linked glycans in the IgA(1) hinge region, which is absent in IgA(2). Specific interaction of jacalin with O-glycans, proved to be present in the human IgA(1) and hypothetically present in bovine IgG(1) structures, is discussed as responsible for the obtained affinity values.

  5. Transient Behaviour of Batch Arrival Queue with N-Policy and Single Vacation (Mx/G/1/N-POLICY)

    NASA Astrophysics Data System (ADS)

    Solanki, Anjana

    2009-07-01

    In this paper Mx/G/1 queuing system with N-policy and single vacation is considered. As soon as the system becomes empty, the server leaves the system for a vacation of random length V. When he returns from the vacation, if the system size is greater then or equal to predetermined value N (threshold), he begins to serve the customers. If not, the server waits in the system until the system size reaches or exceeds N. Here the time dependent system size distribution is obtained.

  6. Comparative Diagnosis of Serum IgG1 and Coproantigen ELISA for Fasciolosis Detection of Goats in Mexico

    PubMed Central

    Molina-Mendoza, Pedro; Hernández-Guzmán, Karina; Olivares-Pérez, Jaime; Sarracent-Pérez, Jorge; Zumaquero-Ríos, José

    2016-01-01

    The objective of present study was to determine the prevalence of natural caprine fasciolosis in the Mixteca region of Mexico using coproantigen and serum IgG1 ELISA tests for comparative purposes. A total of 1070 serum and faecal samples were analyzed for IgG1 antibodies and coproantigens, using ELISA with E/S products as antigen and a monoclonal antibody-based sandwich ELISA. Prevalence of 73.46% was found using the serological ELISA and a percentage of 77.20 was found for coproantigen ELISA. The diagnostic sensitivity and specificity for serum ELISA were 86.7% and 96.4%, and for the coproantigen ELISA they were 93.1% and 97.8%, respectively. The seropositive samples were further categorized as low, medium, or high positivity. Results show a great proportion of low and medium positive goats when the serum ELISA test was used. Correlation coefficients between coproantigens and seropositivity were statistically significant (P < 0.01) for low seropositivity (r = 0.93) and medium seropositivity (r = 0.84). The accuracy of faecal antigen ELISA was higher compared to indirect ELISA serological test. Two ELISAs were shown to be useful for demonstrating the current status of F. hepatica infection in the endemic areas and can be employed in studies on epidemiology as well as anthelmintics treatment for preventing economic loss and the risk of transmission to humans. PMID:27563665

  7. Sphingosine Kinase Regulates Microtubule Dynamics and Organelle Positioning Necessary for Proper G1/S Cell Cycle Transition in Trypanosoma brucei

    PubMed Central

    Pasternack, Deborah A.; Sharma, Aabha I.; Olson, Cheryl L.

    2015-01-01

    ABSTRACT Sphingolipids are important constituents of cell membranes and also serve as mediators of cell signaling and cell recognition. Sphingolipid metabolites such as sphingosine-1-phosphate and ceramide regulate signaling cascades involved in cell proliferation and differentiation, autophagy, inflammation, and apoptosis. Little is known about how sphingolipids and their metabolites function in single-celled eukaryotes. In the present study, we investigated the role of sphingosine kinase (SPHK) in the biology of the protozoan parasite Trypanosoma brucei, the agent of African sleeping sickness. T. brucei SPHK (TbSPHK) is constitutively but differentially expressed during the life cycle of T. brucei. Depletion of TbSPHK in procyclic-form T. brucei causes impaired growth and attenuation in the G1/S phase of the cell cycle. TbSPHK-depleted cells also develop organelle positioning defects and an accumulation of tyrosinated α-tubulin at the elongated posterior end of the cell, known as the “nozzle” phenotype, caused by other molecular perturbations in this organism. Our studies indicate that TbSPHK is involved in G1-to-S cell cycle progression, organelle positioning, and maintenance of cell morphology. Cytotoxicity assays using TbSPHK inhibitors revealed a favorable therapeutic index between T. brucei and human cells, suggesting TbSPHK to be a novel drug target. PMID:26443455

  8. p27Kip1 Is Required to Mediate a G1 Cell Cycle Arrest Downstream of ATM following Genotoxic Stress

    PubMed Central

    Cassimere, Erica K.; Mauvais, Claire; Denicourt, Catherine

    2016-01-01

    The DNA damage response (DDR) is a coordinated signaling network that ensures the maintenance of genome stability under DNA damaging stress. In response to DNA lesions, activation of the DDR leads to the establishment of cell cycle checkpoints that delay cell-cycle progression and allow repair of the defects. The tumor suppressor p27Kip1 is a cyclin-CDK inhibitor that plays an important role in regulating quiescence in a variety of tissues. Several studies have suggested that p27Kip1 also plays a role in the maintenance of genomic integrity. Here we demonstrate that p27Kip1 is essential for the establishment of a G1 checkpoint arrest after DNA damage. We also uncovered that ATM phosphorylates p27Kip1 on a previously uncharacterized residue (Ser-140), which leads to its stabilization after induction of DNA double-strand breaks. Inhibition of this stabilization by replacing endogenous p27Kip1 with a Ser-140 phospho-mutant (S140A) significantly sensitized cells to IR treatments. Our findings reveal a novel role for p27Kip1 in the DNA damage response pathway and suggest that part of its tumor suppressing functions relies in its ability to mediate a G1 arrest after the induction of DNA double strand breaks. PMID:27611996

  9. Comparative Diagnosis of Serum IgG1 and Coproantigen ELISA for Fasciolosis Detection of Goats in Mexico.

    PubMed

    Villa-Mancera, Abel; Molina-Mendoza, Pedro; Hernández-Guzmán, Karina; Olivares-Pérez, Jaime; Sarracent-Pérez, Jorge; Zumaquero-Ríos, José

    2016-01-01

    The objective of present study was to determine the prevalence of natural caprine fasciolosis in the Mixteca region of Mexico using coproantigen and serum IgG1 ELISA tests for comparative purposes. A total of 1070 serum and faecal samples were analyzed for IgG1 antibodies and coproantigens, using ELISA with E/S products as antigen and a monoclonal antibody-based sandwich ELISA. Prevalence of 73.46% was found using the serological ELISA and a percentage of 77.20 was found for coproantigen ELISA. The diagnostic sensitivity and specificity for serum ELISA were 86.7% and 96.4%, and for the coproantigen ELISA they were 93.1% and 97.8%, respectively. The seropositive samples were further categorized as low, medium, or high positivity. Results show a great proportion of low and medium positive goats when the serum ELISA test was used. Correlation coefficients between coproantigens and seropositivity were statistically significant (P < 0.01) for low seropositivity (r = 0.93) and medium seropositivity (r = 0.84). The accuracy of faecal antigen ELISA was higher compared to indirect ELISA serological test. Two ELISAs were shown to be useful for demonstrating the current status of F. hepatica infection in the endemic areas and can be employed in studies on epidemiology as well as anthelmintics treatment for preventing economic loss and the risk of transmission to humans.

  10. H2O2 scavenging inhibits G1/S transition by increasing nuclear levels of p27KIP1.

    PubMed

    Ibañez, Irene L; Policastro, Lucía L; Tropper, Ivanna; Bracalente, Candelaria; Palmieri, Mónica A; Rojas, Paola A; Molinari, Beatriz L; Durán, Hebe

    2011-06-01

    The aim of the present study was to evaluate cell cycle regulation by scavenging H(2)O(2) in tumor cells. A significant arrest in the G1 phase of the cell cycle was demonstrated in CH72-T4 carcinoma cells exposed to catalase, associated with a decrease in cyclin D1 and an increase in the CDK inhibitory protein p27(KIP1). Moreover, we found a differential intracellular distribution of p27(KIP1), which remained in the nucleus after catalase treatment. In vivo experiments showed an increase in nuclear levels of p27(KIP1) associated with the inhibition of tumor growth by H(2)O(2) scavenging, confirming in vitro results. To conclude, H(2)O(2) scavenging may induce cell cycle arrest through the modulation of cyclin D1 and p27(KIP1) levels and nuclear localization of p27(KIP1). To our knowledge, this is the first report that demonstrates that the modulation of ROS alters the intracellular localization of a key regulatory protein of G1/S transition.

  11. Young Remnants of Type Ia Supernovae and Their Progenitors: A Study of SNR G1.9+0.3

    NASA Astrophysics Data System (ADS)

    Chakraborti, Sayan; Childs, Francesca; Soderberg, Alicia

    2016-03-01

    SNe Ia, with their remarkably homogeneous light curves and spectra, have been used as standardizable candles to measure the accelerating expansion of the universe. Yet, their progenitors remain elusive. Common explanations invoke a degenerate star (white dwarf) that explodes upon almost reaching the Chandrasekhar limit, by either steadily accreting mass from a companion star or violently merging with another degenerate star. We show that circumstellar interaction in young Galactic supernova remnants can be used to distinguish between these single and double degenerate (DD) progenitor scenarios. Here we propose a new diagnostic, the surface brightness index, which can be computed from theory and compared with Chandra and Very Large Array (VLA) observations. We use this method to demonstrate that a DD progenitor can explain the decades-long flux rise and size increase of the youngest known galactic supernova remnant (SNR), G1.9+0.3. We disfavor a single degenerate scenario for SNR G1.9+0.3. We attribute the observed properties to the interaction between a steep ejecta profile and a constant density environment. We suggest using the upgraded VLA, ASKAP, and MeerKAT to detect circumstellar interaction in the remnants of historical SNe Ia in the Local Group of galaxies. This may settle the long-standing debate over their progenitors.

  12. Comparative Diagnosis of Serum IgG1 and Coproantigen ELISA for Fasciolosis Detection of Goats in Mexico.

    PubMed

    Villa-Mancera, Abel; Molina-Mendoza, Pedro; Hernández-Guzmán, Karina; Olivares-Pérez, Jaime; Sarracent-Pérez, Jorge; Zumaquero-Ríos, José

    2016-01-01

    The objective of present study was to determine the prevalence of natural caprine fasciolosis in the Mixteca region of Mexico using coproantigen and serum IgG1 ELISA tests for comparative purposes. A total of 1070 serum and faecal samples were analyzed for IgG1 antibodies and coproantigens, using ELISA with E/S products as antigen and a monoclonal antibody-based sandwich ELISA. Prevalence of 73.46% was found using the serological ELISA and a percentage of 77.20 was found for coproantigen ELISA. The diagnostic sensitivity and specificity for serum ELISA were 86.7% and 96.4%, and for the coproantigen ELISA they were 93.1% and 97.8%, respectively. The seropositive samples were further categorized as low, medium, or high positivity. Results show a great proportion of low and medium positive goats when the serum ELISA test was used. Correlation coefficients between coproantigens and seropositivity were statistically significant (P < 0.01) for low seropositivity (r = 0.93) and medium seropositivity (r = 0.84). The accuracy of faecal antigen ELISA was higher compared to indirect ELISA serological test. Two ELISAs were shown to be useful for demonstrating the current status of F. hepatica infection in the endemic areas and can be employed in studies on epidemiology as well as anthelmintics treatment for preventing economic loss and the risk of transmission to humans. PMID:27563665

  13. An APC/C-Cdh1 Biosensor Reveals the Dynamics of Cdh1 Inactivation at the G1/S Transition

    PubMed Central

    Ondracka, Andrej; Robbins, Jonathan A.; Cross, Frederick R.

    2016-01-01

    B-type cyclin-dependent kinase activity must be turned off for mitotic exit and G1 stabilization. B-type cyclin degradation is mediated by the anaphase-promoting complex/cyclosome (APC/C); during and after mitotic exit, APC/C is dependent on Cdh1. Cdh1 is in turn phosphorylated and inactivated by cyclin-CDK at the Start transition of the new cell cycle. We developed a biosensor to assess the cell cycle dynamics of APC/C-Cdh1. Nuclear exit of the G1 transcriptional repressor Whi5 is a known marker of Start; APC/C-Cdh1 is inactivated 12 min after Whi5 nuclear exit with little measurable cell-to-cell timing variability. Multiple phosphorylation sites on Cdh1 act in a redundant manner to repress its activity. Reducing the number of phosphorylation sites on Cdh1 can to some extent be tolerated for cell viability, but it increases variability in timing of APC/C-Cdh1 inactivation. Mutants with minimal subsets of phosphorylation sites required for viability exhibit striking stochasticity in multiple responses including budding, nuclear division, and APC/C-Cdh1 activity itself. Multiple cyclin-CDK complexes, as well as the stoichiometric inhibitor Acm1, contribute to APC/C-Cdh1 inactivation; this redundant control is likely to promote rapid and reliable APC/C-Cdh1 inactivation immediately following the Start transition. PMID:27410035

  14. Structural Characterization of IgG1 mAb Aggregates and Particles Generated under Various Stress Conditions

    PubMed Central

    Telikepalli, Srivalli N.; Kumru, Ozan S.; Kalonia, Cavan; Esfandiary, Reza; Joshi, Sangeeta B.; Middaugh, C. Russell; Volkin, David B.

    2014-01-01

    IgG1 mAb solutions were prepared with and without sodium chloride and subjected to different environmental stresses. Formation of aggregates and particles of varying size was monitored by a combination of size exclusion chromatography (SEC), Nanosight Tracking Analysis (NTA), Micro-flow Imaging (MFI), turbidity, and visual assessments. Stirring and heating induced the highest concentration of particles. In general, the presence of NaCl enhanced this effect. The morphology of the particles formed from mAb samples exposed to different stresses was analyzed from TEM and MFI images. Shaking samples without NaCl generated the most fibrillar particles, while stirring created largely spherical particles. The composition of the particles was evaluated for covalent cross-linking by SDS-PAGE, overall secondary structure by FTIR microscopy, and surface apolarity by extrinsic fluorescence spectroscopy. Freeze-thaw and shaking led to particles containing protein with native-like secondary structure. Heating and stirring produced IgG1 containing aggregates and particles with some non-native disulfide crosslinks, varying levels of intermolecular beta sheet content, and increased surface hydrophobicity. These results highlight the importance of evaluating protein particle morphology and composition, in addition to particle number and size distributions, to better understand the effect of solution conditions and environmental stresses on the formation of protein particles in mAb solutions. PMID:24452866

  15. TopBP1 is required at mitosis to reduce transmission of DNA damage to G1 daughter cells

    PubMed Central

    Pedersen, Rune Troelsgaard; Kruse, Thomas; Nilsson, Jakob

    2015-01-01

    Genome integrity is critically dependent on timely DNA replication and accurate chromosome segregation. Replication stress delays replication into G2/M, which in turn impairs proper chromosome segregation and inflicts DNA damage on the daughter cells. Here we show that TopBP1 forms foci upon mitotic entry. In early mitosis, TopBP1 marks sites of and promotes unscheduled DNA synthesis. Moreover, TopBP1 is required for focus formation of the structure-selective nuclease and scaffold protein SLX4 in mitosis. Persistent TopBP1 foci transition into 53BP1 nuclear bodies (NBs) in G1 and precise temporal depletion of TopBP1 just before mitotic entry induced formation of 53BP1 NBs in the next cell cycle, showing that TopBP1 acts to reduce transmission of DNA damage to G1 daughter cells. Based on these results, we propose that TopBP1 maintains genome integrity in mitosis by controlling chromatin recruitment of SLX4 and by facilitating unscheduled DNA synthesis. PMID:26283799

  16. TopBP1 is required at mitosis to reduce transmission of DNA damage to G1 daughter cells.

    PubMed

    Pedersen, Rune Troelsgaard; Kruse, Thomas; Nilsson, Jakob; Oestergaard, Vibe H; Lisby, Michael

    2015-08-17

    Genome integrity is critically dependent on timely DNA replication and accurate chromosome segregation. Replication stress delays replication into G2/M, which in turn impairs proper chromosome segregation and inflicts DNA damage on the daughter cells. Here we show that TopBP1 forms foci upon mitotic entry. In early mitosis, TopBP1 marks sites of and promotes unscheduled DNA synthesis. Moreover, TopBP1 is required for focus formation of the structure-selective nuclease and scaffold protein SLX4 in mitosis. Persistent TopBP1 foci transition into 53BP1 nuclear bodies (NBs) in G1 and precise temporal depletion of TopBP1 just before mitotic entry induced formation of 53BP1 NBs in the next cell cycle, showing that TopBP1 acts to reduce transmission of DNA damage to G1 daughter cells. Based on these results, we propose that TopBP1 maintains genome integrity in mitosis by controlling chromatin recruitment of SLX4 and by facilitating unscheduled DNA synthesis.

  17. p27Kip1 Is Required to Mediate a G1 Cell Cycle Arrest Downstream of ATM following Genotoxic Stress.

    PubMed

    Cassimere, Erica K; Mauvais, Claire; Denicourt, Catherine

    2016-01-01

    The DNA damage response (DDR) is a coordinated signaling network that ensures the maintenance of genome stability under DNA damaging stress. In response to DNA lesions, activation of the DDR leads to the establishment of cell cycle checkpoints that delay cell-cycle progression and allow repair of the defects. The tumor suppressor p27Kip1 is a cyclin-CDK inhibitor that plays an important role in regulating quiescence in a variety of tissues. Several studies have suggested that p27Kip1 also plays a role in the maintenance of genomic integrity. Here we demonstrate that p27Kip1 is essential for the establishment of a G1 checkpoint arrest after DNA damage. We also uncovered that ATM phosphorylates p27Kip1 on a previously uncharacterized residue (Ser-140), which leads to its stabilization after induction of DNA double-strand breaks. Inhibition of this stabilization by replacing endogenous p27Kip1 with a Ser-140 phospho-mutant (S140A) significantly sensitized cells to IR treatments. Our findings reveal a novel role for p27Kip1 in the DNA damage response pathway and suggest that part of its tumor suppressing functions relies in its ability to mediate a G1 arrest after the induction of DNA double strand breaks. PMID:27611996

  18. DNA damage during the G0/G1 phase triggers RNA-templated, Cockayne syndrome B-dependent homologous recombination.

    PubMed

    Wei, Leizhen; Nakajima, Satoshi; Böhm, Stefanie; Bernstein, Kara A; Shen, Zhiyuan; Tsang, Michael; Levine, Arthur S; Lan, Li

    2015-07-01

    Damage repair mechanisms at transcriptionally active sites during the G0/G1 phase are largely unknown. To elucidate these mechanisms, we introduced genome site-specific oxidative DNA damage and determined the role of transcription in repair factor assembly. We find that KU and NBS1 are recruited to damage sites independent of transcription. However, assembly of RPA1, RAD51C, RAD51, and RAD52 at such sites is strictly governed by active transcription and requires both wild-type Cockayne syndrome protein B (CSB) function and the presence of RNA in the G0/G1 phase. We show that the ATPase activity of CSB is indispensable for loading and binding of the recombination factors. CSB counters radiation-induced DNA damage in both cells and zebrafish models. Taken together, our results have uncovered a novel, RNA-based recombination mechanism by which CSB protects genome stability from strand breaks at transcriptionally active sites and may provide insight into the clinical manifestations of Cockayne syndrome. PMID:26100862

  19. G1/S Inhibitors and the SWI/SNF Complex Control Cell-Cycle Exit during Muscle Differentiation.

    PubMed

    Ruijtenberg, Suzan; van den Heuvel, Sander

    2015-07-16

    The transition from proliferating precursor cells to post-mitotic differentiated cells is crucial for development, tissue homeostasis, and tumor suppression. To study cell-cycle exit during differentiation in vivo, we developed a conditional knockout and lineage-tracing system for Caenorhabditis elegans. Combined lineage-specific gene inactivation and genetic screening revealed extensive redundancies between previously identified cell-cycle inhibitors and the SWI/SNF chromatin-remodeling complex. Muscle precursor cells missing either SWI/SNF or G1/S inhibitor function could still arrest cell division, while simultaneous inactivation of these regulators caused continued proliferation and a C. elegans tumor phenotype. Further genetic analyses support that SWI/SNF acts in concert with hlh-1 MyoD, antagonizes Polycomb-mediated transcriptional repression, and suppresses cye-1 Cyclin E transcription to arrest cell division of muscle precursors. Thus, SWI/SNF and G1/S inhibitors provide alternative mechanisms to arrest cell-cycle progression during terminal differentiation, which offers insight into the frequent mutation of SWI/SNF genes in human cancers.

  20. Cellular expression of human centromere protein C demonstrates a cyclic behavior with highest abundance in the G1 phase.

    PubMed Central

    Knehr, M; Poppe, M; Schroeter, D; Eickelbaum, W; Finze, E M; Kiesewetter, U L; Enulescu, M; Arand, M; Paweletz, N

    1996-01-01

    Centromere proteins are localized within the centromere-kinetochore complex, which can be proven by means of immunofluorescence microscopy and immunoelectron microscopy. In consequence, their putative functions seem to be related exclusively to mitosis, namely to the interaction of the chromosomal kinetochores with spindle microtubules. However, electron microscopy using immune sera enriched with specific antibodies against human centromere protein C (CENP-C) showed that it occurs not only in mitosis but during the whole cell cycle. Therefore, we investigated the cell cycle-specific expression of CENP-C systematically on protein and mRNA levels applying HeLa cells synchronized in all cell cycle phases. Immunoblotting confirmed protein expression during the whole cell cycle and revealed an increase of CENP-C from the S phase through the G2 phase and mitosis to highest abundance in the G1 phase. Since this was rather surprising, we verified it by quantifying phase-specific mRNA levels of CENP-C, paralleled by the amplification of suitable internal standards, using the polymerase chain reaction. The results were in excellent agreement with abundant protein amounts and confirmed the cyclic behavior of CENP-C during the cell cycle. In consequence, we postulate that in addition to its role in mitosis, CENP-C has a further role in the G1 phase that may be related to cell cycle control. Images Fig. 1 Fig. 2 Fig. 4 PMID:8816782

  1. An APC/C-Cdh1 Biosensor Reveals the Dynamics of Cdh1 Inactivation at the G1/S Transition.

    PubMed

    Ondracka, Andrej; Robbins, Jonathan A; Cross, Frederick R

    2016-01-01

    B-type cyclin-dependent kinase activity must be turned off for mitotic exit and G1 stabilization. B-type cyclin degradation is mediated by the anaphase-promoting complex/cyclosome (APC/C); during and after mitotic exit, APC/C is dependent on Cdh1. Cdh1 is in turn phosphorylated and inactivated by cyclin-CDK at the Start transition of the new cell cycle. We developed a biosensor to assess the cell cycle dynamics of APC/C-Cdh1. Nuclear exit of the G1 transcriptional repressor Whi5 is a known marker of Start; APC/C-Cdh1 is inactivated 12 min after Whi5 nuclear exit with little measurable cell-to-cell timing variability. Multiple phosphorylation sites on Cdh1 act in a redundant manner to repress its activity. Reducing the number of phosphorylation sites on Cdh1 can to some extent be tolerated for cell viability, but it increases variability in timing of APC/C-Cdh1 inactivation. Mutants with minimal subsets of phosphorylation sites required for viability exhibit striking stochasticity in multiple responses including budding, nuclear division, and APC/C-Cdh1 activity itself. Multiple cyclin-CDK complexes, as well as the stoichiometric inhibitor Acm1, contribute to APC/C-Cdh1 inactivation; this redundant control is likely to promote rapid and reliable APC/C-Cdh1 inactivation immediately following the Start transition.

  2. Effects of furanodiene on 95-D lung cancer cells: apoptosis, autophagy and G1 phase cell cycle arrest.

    PubMed

    Xu, Wen-Shan; Li, Ting; Wu, Guo-Sheng; Dang, Yuan-Ye; Hao, Wen-Hui; Chen, Xiu-Ping; Lu, Jin-Jian; Wang, Yi-Tao

    2014-01-01

    Furanodiene (FUR) is a natural terpenoid isolated from Rhizoma curcumae, a well-known Chinese medicinal herb that presents anti-proliferative activities in several cancer cell lines. Herein, we systematically investigated the effects of FUR on the significant processes of tumor progression with the relatively low concentrations in 95-D lung cancer cells. FUR concentration-dependently inhibited cell proliferation and blocked the cell cycle progressions in G1 phase by down-regulating the protein levels of cyclin D1 and CDK6, and up-regulating those of p21 and p27 in 95-D cells. FUR also affected the signaling molecules that regulate apoptosis in 95-D cells revealed by the down-regulation of the protein levels of full PARP, pro-caspase-7, survivin, and Bcl-2, and the up-regulation of cleaved PARP. Further studies showed that FUR enhanced the expression of light chain 3-II (LC3-II) in the protein level, indicating that autophagy is involved in this process. Besides, the adhesion ability of 95-D cells to matrigel and fibronectin was slightly inhibited after FUR treatment for 1 h in our experimental condition. FUR also slightly suppressed cell migration and invasion in 95-D cells according to the data from wound healing and Transwell assays, respectively. Taken together, FUR activated the signal molecules regulating G1 cell cycle arrest, apoptosis and autophagy, while slightly affecting the key steps of cell metastasis in 95-D lung cancer cells in the relatively low concentrations.

  3. Ethanol extract of Innotus obliquus (Chaga mushroom) induces G1 cell cycle arrest in HT-29 human colon cancer cells

    PubMed Central

    Lee, Hyun Sook; Kim, Eun Ji

    2015-01-01

    BACKGROUND/OBJECTIVES Inonotus obliquus (I. obliquus, Chaga mushroom) has long been used as a folk medicine to treat cancer. In the present study, we examined whether or not ethanol extract of I. obliquus (EEIO) inhibits cell cycle progression in HT-29 human colon cancer cells, in addition to its mechanism of action. MATERIALS/METHODS To examine the effects of Inonotus obliquus on the cell cycle progression and the molecular mechanism in colon cancer cells, HT-29 human colon cancer cells were cultured in the presence of 2.5 - 10 µg/mL of EEIO, and analyzed the cell cycle arrest by flow cytometry and the cell cycle controlling protein expression by Western blotting. RESULTS Treatment cells with 2.5 - 10 µg/mL of EEIO reduced viable HT-29 cell numbers and DNA synthesis, increased the percentage of cells in G1 phase, decreased protein expression of CDK2, CDK4, and cyclin D1, increased expression of p21, p27, and p53, and inhibited phosphorylation of Rb and E2F1 expression. Among I. obliquus fractions, fraction 2 (fractionated by dichloromethane from EEIO) showed the same effect as EEIO treatment on cell proliferation and cell cycle-related protein levels. CONCLUSIONS These results demonstrate that fraction 2 is the major fraction that induces G1 arrest and inhibits cell proliferation, suggesting I. obliquus could be used as a natural anti-cancer ingredient in the food and/or pharmaceutical industry. PMID:25861415

  4. Expression patterns of cyclin D1 and related proteins regulating G1-S phase transition in uveal melanoma and retinoblastoma

    PubMed Central

    Coupland, S; Bechrakis, N; Schuler, A; Anagnostopoulos, I; Hummel, M; Bornfeld, N; Stein, H

    1998-01-01

    BACKGROUND/AIMS—A checkpoint mechanism in late G1, whose regulation via loss of retinoblastoma protein (pRB) or p16, or overexpression of cyclin D1 or cyclin dependent kinase 4 (CDK4), has been proposed to constitute a common pathway to malignancy. The aims of this study were (a) to compare markers of cell cycle G1-S phase transition in an intraocular tumour with known pRB deficiency (retinoblastoma) and compare it with one with an apparently functional pRB (uveal melanoma); (b) to determine if one of these markers may have a role in the pathogenesis of uveal melanoma; and (c) to determine if there is a difference in cell cycle marker expression following treatment of uveal melanoma and retinoblastoma.
METHODS—90 eyes were enucleated from 89 patients for retinoblastoma (n=24) or for choroidal or ciliary body melanoma (n=66). Conventional paraffin sections were assessed for cell type and degree of differentiation. Additional slides were investigated applying standard immunohistochemical methods with antibodies specific for cyclin D1 protein, pRB, p53, p21, p16, BCL-2, and MIB-1.
RESULTS—Cyclin D1 protein and pRB were negative in retinoblastoma using the applied antibodies. In contrast, cyclin D1 protein expression was observed in 65% of uveal melanomas; a positive correlation between cyclin D1 cell positivity and tumour cell type, location, growth fraction, as well as with pRB positivity was observed. p53, p21, and p16 could be demonstrated in both tumours. An inverse relation between p53 and p21 expression was demonstrated in most choroidal melanomas and in some retinoblastomas. Apart from a decrease in the growth fractions of the tumours as determined by MIB-1, a significant difference in the expression of G1-S phase transition markers in vital areas of uveal melanoma and retinoblastoma following treatment with radiotherapy and/or chemotherapy was not observed.
CONCLUSION—Retinoblastomas and uveal melanomas, two tumours of differing pRB status

  5. Human IgG1 Responses to Surface Localised Schistosoma mansoni Ly6 Family Members Drop following Praziquantel Treatment

    PubMed Central

    Chalmers, Iain W.; Fitzsimmons, Colin M.; Brown, Martha; Pierrot, Christine; Jones, Frances M.; Wawrzyniak, Jakub M.; Fernandez-Fuentes, Narcis; Tukahebwa, Edridah M.; Dunne, David W.; Khalife, Jamal; Hoffmann, Karl F.

    2015-01-01

    Background The heptalaminate-covered, syncytial tegument is an important anatomical adaptation that enables schistosome parasites to maintain long-term, intravascular residence in definitive hosts. Investigation of the proteins present in this surface layer and the immune responses elicited by them during infection is crucial to our understanding of host/parasite interactions. Recent studies have revealed a number of novel tegumental surface proteins including three (SmCD59a, SmCD59b and Sm29) containing uPAR/Ly6 domains (renamed SmLy6A SmLy6B and SmLy6D in this study). While vaccination with SmLy6A (SmCD59a) and SmLy6D (Sm29) induces protective immunity in experimental models, human immunoglobulin responses to representative SmLy6 family members have yet to be thoroughly explored. Methodology/Principal Findings Using a PSI-BLAST-based search, we present a comprehensive reanalysis of the Schistosoma mansoni Ly6 family (SmLy6A-K). Our examination extends the number of members to eleven (including three novel proteins) and provides strong evidence that the previously identified vaccine candidate Sm29 (renamed SmLy6D) is a unique double uPAR/Ly6 domain-containing representative. Presence of canonical cysteine residues, signal peptides and GPI-anchor sites strongly suggest that all SmLy6 proteins are cell surface-bound. To provide evidence that SmLy6 members are immunogenic in human populations, we report IgG1 (as well as IgG4 and IgE) responses against two surface-bound representatives (SmLy6A and SmLy6B) within a cohort of S. mansoni-infected Ugandan males before and after praziquantel treatment. While pre-treatment IgG1 prevalence for SmLy6A and SmLy6B differs amongst the studied population (7.4% and 25.3% of the cohort, respectively), these values are both higher than IgG1 prevalence (2.7%) for a sub-surface tegumental antigen, SmTAL1. Further, post-treatment IgG1 levels against surface-associated SmLy6A and SmLy6B significantly drop (p = 0.020 and p < 0

  6. Association between thyroid microsomal antibodies of subclass IgG-1 and hypothyroidism in autoimmune postpartum thyroiditis.

    PubMed Central

    Jansson, R; Thompson, P M; Clark, F; McLachlan, S M

    1986-01-01

    The potential role of thyroid microsomal (Mic) antibodies in the development of postpartum hypothyroidism was investigated in 34 euthyroid women, whose sera were found to contain Mic antibodies in pregnancy. Additional serum samples were obtained 2. 5 and 10-12 months after delivery and analysed for IgG class and IgG subclass levels of Mic antibodies by ELISA techniques. Characteristically, Mic antibodies decreased from early pregnancy to 2 months postpartum, increased two-fold 5 months postpartum and had returned 10-12 months postpartum to the early pregnancy level. Mic antibodies were predominantly subclass IgG-1 or IgG-4 with only minor contributions from IgG-2 and IgG-3. In each individual the percentage contribution made by each IgG subclass to Mic antibody was essentially similar in early pregnancy and the postpartum period despite changes in total IgG class Mic antibody. During the year following delivery, thyrotoxicosis alone (Graves' disease) developed in 5 women. In the remaining 29 patients the absolute levels of Mic antibodies of IgG-4 subclass were similar 5 months postpartum in women with maximal serum thyrotropin (TSH) greater than 20 mU/1 (mean optical density in ELISA +/- s.d.; 0.84 +/- 0.538; n = 13) and in women with maximal TSH less than 10 mU/l (0.69 +/- 0.457; n = 16). In contrast, significantly higher values were observed for Mic antibody of IgG-1 subclass in patients with TSH greater than 20 mU/l (1.14 +/- 0.440) compared with women with maximal TSH less than 10 mU/l (0.65 +/- 0.289) (P less than 0.001 by t-test for groups). These results imply that the magnitude of Mic antibody levels of subclass IgG-1 but not IgG-4 is associated with the development of postpartum hypothyroidism and possibly with tissue destruction in autoimmune thyroid disease in general. PMID:3754185

  7. Serum Immunoglobulin G4 and Immunoglobulin G1 for Distinguishing Immunoglobulin G4-Associated Cholangitis From Primary Sclerosing Cholangitis

    PubMed Central

    Boonstra, Kirsten; Culver, Emma L; de Buy Wenniger, Lucas Maillette; van Heerde, Marianne J; van Erpecum, Karel J; Poen, Alexander C; van Nieuwkerk, Karin MJ; Spanier, BW Marcel; Witteman, Ben JM; Tuynman, Hans ARE; van Geloven, Nan; van Buuren, Henk; Chapman, Roger W; Barnes, Eleanor; Beuers, Ulrich; Ponsioen, Cyriel Y

    2014-01-01

    The recent addition of immunoglobulin (Ig)G4-associated cholangitis (IAC), also called IgG4-related sclerosing cholangitis (IRSC), to the spectrum of chronic cholangiopathies has created the clinical need for reliable methods to discriminate between IAC and the more common cholestatic entities, primary (PSC) and secondary sclerosing cholangitis. The current American Association for the Study of Liver Diseases practice guidelines for PSC advise on the measurement of specific Ig (sIg)G4 in PSC patients, but interpretation of elevated sIgG4 levels remains unclear. We aimed to provide an algorithm to distinguish IAC from PSC using sIgG analyses. We measured total IgG and IgG subclasses in serum samples of IAC (n = 73) and PSC (n = 310) patients, as well as in serum samples of disease controls (primary biliary cirrhosis; n = 22). sIgG4 levels were elevated above the upper limit of normal (ULN = >1.4 g/L) in 45 PSC patients (15%; 95% confidence interval [CI]: 11-19). The highest specificity and positive predictive value (PPV; 100%) for IAC were reached when applying the 4× ULN (sIgG4 > 5.6 g/L) cutoff with a sensitivity of 42% (95% CI: 31-55). However, in patients with a sIgG4 between 1× and 2× ULN (n = 38/45), the PPV of sIgG4 for IAC was only 28%. In this subgroup, the sIgG4/sIgG1 ratio cutoff of 0.24 yielded a sensitivity of 80% (95% CI: 51-95), a specificity of 74% (95% CI: 57-86), a PPV of 55% (95% CI: 33-75), and a negative predictive value of 90% (95% CI: 73-97). Conclusion: Elevated sIgG4 (>1.4 g/L) occurred in 15% of patients with PSC. In patients with a sIgG4 >1.4 and <2.8 g/L, incorporating the IgG4/IgG1 ratio with a cutoff at 0.24 in the diagnostic algorithm significantly improved PPV and specificity. We propose a new diagnostic algorithm based on IgG4/IgG1 ratio that may be used in clinical practice to distinguish PSC from IAC. (Hepatology 2014;59:1954–1963) PMID:24375491

  8. [Scientific connotation of processing Bombyx Batryticatus under high temperature].

    PubMed

    Ma, Li; Wang, Xuan; Ma, Lin; Wang, Man-yuan; Qiu, Feng

    2015-12-01

    The aim of this study was to elucidate the scientific connotation of Bombyx Batryticatus processing with wheat bran under high temperature. The contents of soluble protein extracted from Bombyx Batryticatus and its processed products and the limited content of AFT in Bombyx Batryticatus and the processed one were compared. The concentration of protein was measured with the Bradford methods and the difference of protein between Bombyx Batryticatus and its processed products was compared by SDS-PAGE analysis. Aflatoxin B1, B2, G1, and G2 were determined by reversed-phase HPLC. The results showed that the soluble protein content of Bombyx Batryticatus and its processed products were (47.065 +/- 0.249), (29.756 +/- 1.961) mg x g(-1), correspondingly. Analysis of protein gel electrophoresis showed that there were no significant differences between the crude and processed one in protein varieties. 6 bands were detected: 31.90, 26.80, 18.71, 15.00, 10.18, 8.929 kDa. Below 10 kDa, the color of bands of the processed one was deeper than the crude one, which demonstrate that macromolecular protein was degradated into micromolecule. The content of AFG1, AFB1, AFG2, AFB2 were 0.382, 0.207, 0.223, 0.073 g x kg(-1), not exceeded 5 microg x kg(-1) while the processed one was not detected. Through processing with wheat bran under high temperature, the content of soluble protein in Bombyx Batryticatus decreased, the processing purpose for alleviating drug property was achieved. Meanwhile, the limited content of aflatoxins were reduced or cleared by processing procedure or absorbed by processing auxillary material, adding the safety of the traditional Chinese Medicine. In conclusion, as a traditional processing method, bran frying Bombyx Batryticatus was scientific and reasonable. PMID:27141674

  9. Identification and characterization of asparagine deamidation in the light chain CDR1 of a humanized IgG1 antibody.

    PubMed

    Vlasak, Josef; Bussat, Marie C; Wang, Shiyi; Wagner-Rousset, Elsa; Schaefer, Mark; Klinguer-Hamour, Christine; Kirchmeier, Marc; Corvaïa, Nathalie; Ionescu, Roxana; Beck, Alain

    2009-09-15

    Despite technological advances, detection of deamidation in large proteins remains a challenge and the use of orthogonal methods is needed for unequivocal assignment. By a combination of cation-exchange separation, papain digestion, and a panel of mass spectrometry techniques we identified asparagine deamidation in light chain complementarity determining region 1 (CDR1) of a humanized IgG1 monoclonal antibody. The reaction yields both Asp and isoAsp, which were assigned by Edman degradation and by isoAsp detection using protein isoaspartate methyltransferase. The deamidated antibody variants were less potent in antigen binding compared to the nondegraded antibody. Changes in near-UV CD spectra, susceptibility to papain cleavage in an adjacent CDR2 loop, and the tendency of the newly formed isoAsp to undergo isomerization suggest local perturbations in the structure of the isoAsp-containing antibody.

  10. Detailed mineralogical characterization of the Bullfrog and Tram members USW-G1, with emphasis on clay mineralogy

    SciTech Connect

    Bish, D.L.

    1981-10-01

    The detailed mineralogy of the Bullfrog and Tram Members of the Crater Flat Tuff from drill hole USW-G1 has been examined, primarily to characterize fully the amounts and types of clay minerals in the tuffs and the possible effects clay minerals have on rock properties. Results of bulk sample x-ray diffraction analyses agree closely with previous determinations, although slightly higher clay mineral contents were found in this study. X-ray diffraction analysis of fine fractions revealed that the clay minerals in the tuffs are sodium-saturated montmorillonite-beidellites with typical layer charges and no high-charge layers. These smectites are found in virtually all samples of the Bullfrog and Tram, and there is no correlation between the amounts of smectites and the amounts of zeolite, quartz, and feldspar. Smectites are present in both welded and nonwelded horizons and are scarce in some zones with slight-to-absent welding.

  11. Optimal management for infinite capacity N-policy M/G/1 queue with a removable service station

    NASA Astrophysics Data System (ADS)

    Chang, Y. C.; Pearn, W. L.

    2011-07-01

    In this article, we consider an infinite capacity N-policy M/G/1 queueing system with a single removable server. Poisson arrivals and general distribution service times are assumed. The server is controllable that may be turned on at arrival epochs or off at service completion epochs. We apply a differential technique to study system sensitivity, which examines the effect of different system input parameters on the system. A cost model for infinite capacity queueing system under steady-state condition is developed, to determine the optimal management policy at minimum cost. Analytical results for sensitivity analysis are derived. We also provide extensive numerical computations to illustrate the analytical sensitivity properties obtained. Finally, an application example is presented to demonstrate how the model could be used in real applications to obtain the optimal management policy.

  12. Human RAD6 promotes G1-S transition and cell proliferation through upregulation of cyclin D1 expression.

    PubMed

    Cai, Fengfeng; Chen, Ping; Chen, Li; Biskup, Ewelina; Liu, Yan; Chen, Pei-Chao; Chang, Jian-Feng; Jiang, Wenjie; Jing, Yuanya; Chen, Youwei; Jin, Hui; Chen, Su

    2014-01-01

    Protein ubiquitinylation regulates protein stability and activity. RAD6, an E2 ubiquitin-conjugating enzyme, which that has been substantially biochemically characterized, functions in a number of biologically relevant pathways, including cell cycle progression. In this study, we show that RAD6 promotes the G1-S transition and cell proliferation by regulating the expression of cyclin D1 (CCND1) in human cells. Furthermore, our data indicate that RAD6 influences the transcription of CCND1 by increasing monoubiquitinylation of histone H2B and trimethylation of H3K4 in the CCND1 promoter region. Our study presents, for the first time, an evidence for the function of RAD6 in cell cycle progression and cell proliferation in human cells, raising the possibility that RAD6 could be a new target for molecular diagnosis and prognosis in cancer therapeutics.

  13. Human RAD6 Promotes G1-S Transition and Cell Proliferation through Upregulation of Cyclin D1 Expression

    PubMed Central

    Biskup, Ewelina; Liu, Yan; Chen, Pei-Chao; Chang, Jian-Feng; Jiang, Wenjie; Jing, Yuanya; Chen, Youwei; Jin, Hui; Chen, Su

    2014-01-01

    Protein ubiquitinylation regulates protein stability and activity. RAD6, an E2 ubiquitin-conjugating enzyme, which that has been substantially biochemically characterized, functions in a number of biologically relevant pathways, including cell cycle progression. In this study, we show that RAD6 promotes the G1-S transition and cell proliferation by regulating the expression of cyclin D1 (CCND1) in human cells. Furthermore, our data indicate that RAD6 influences the transcription of CCND1 by increasing monoubiquitinylation of histone H2B and trimethylation of H3K4 in the CCND1 promoter region. Our study presents, for the first time, an evidence for the function of RAD6 in cell cycle progression and cell proliferation in human cells, raising the possibility that RAD6 could be a new target for molecular diagnosis and prognosis in cancer therapeutics. PMID:25409181

  14. A Peptide Derived from G0/G1 Switch Gene 2 Acts as Noncompetitive Inhibitor of Adipose Triglyceride Lipase*

    PubMed Central

    Cerk, Ines K.; Salzburger, Barbara; Boeszoermenyi, Andras; Heier, Christoph; Pillip, Christoph; Romauch, Matthias; Schweiger, Martina; Cornaciu, Irina; Lass, Achim; Zimmermann, Robert; Zechner, Rudolf; Oberer, Monika

    2014-01-01

    The protein G0/G1 switch gene 2 (G0S2) is a small basic protein that functions as an endogenous inhibitor of adipose triglyceride lipase (ATGL), a key enzyme in intracellular lipolysis. In this study, we identified a short sequence covering residues Lys-20 to Ala-52 in G0S2 that is still fully capable of inhibiting mouse and human ATGL. We found that a synthetic peptide corresponding to this region inhibits ATGL in a noncompetitive manner in the nanomolar range. This peptide is highly selective for ATGL and does not inhibit other lipases, including hormone-sensitive lipase, monoacylglycerol lipase, lipoprotein lipase, and patatin domain-containing phospholipases 6 and 7. Because increased lipolysis is linked to the development of metabolic disorders, the inhibition of ATGL by G0S2-derived peptides may represent a novel therapeutic tool to modulate lipolysis. PMID:25258314

  15. Do comets C/1861 G1 (Thatcher) and C/1861 J1 (Great comet) have a common origin?

    NASA Astrophysics Data System (ADS)

    Branham, R. L.

    2015-10-01

    A new orbit is calculated for Comet C/1861 G1 (Thatcher), associated with the Lyrid meteor shower, to replace Oppolzer's orbit of 1864. The new orbit is based upon 649 observations, 326 in right ascension and 323 in declination, made between 11 April 1861 and 7 Sept. 1861. The final orbit uses residuals calculated with the Welsch weighting function. The comet's period of 416.87 ± 0.56 yr agrees with Oppolzer's period of 415 yr athough other elements such as the inclination differ. Although the post-perihelion residuals are relatively random, 52.1% probability of randomness, pre-perihelion residuals lack randomness indicating possible deviations from Keplerian motion caused by ejection of meteoritic material. Comet Thatcher is unrelated to the Great comet of 1861.

  16. Human RAD6 promotes G1-S transition and cell proliferation through upregulation of cyclin D1 expression.

    PubMed

    Cai, Fengfeng; Chen, Ping; Chen, Li; Biskup, Ewelina; Liu, Yan; Chen, Pei-Chao; Chang, Jian-Feng; Jiang, Wenjie; Jing, Yuanya; Chen, Youwei; Jin, Hui; Chen, Su

    2014-01-01

    Protein ubiquitinylation regulates protein stability and activity. RAD6, an E2 ubiquitin-conjugating enzyme, which that has been substantially biochemically characterized, functions in a number of biologically relevant pathways, including cell cycle progression. In this study, we show that RAD6 promotes the G1-S transition and cell proliferation by regulating the expression of cyclin D1 (CCND1) in human cells. Furthermore, our data indicate that RAD6 influences the transcription of CCND1 by increasing monoubiquitinylation of histone H2B and trimethylation of H3K4 in the CCND1 promoter region. Our study presents, for the first time, an evidence for the function of RAD6 in cell cycle progression and cell proliferation in human cells, raising the possibility that RAD6 could be a new target for molecular diagnosis and prognosis in cancer therapeutics. PMID:25409181

  17. Short report: the use of a polymerase chain reaction to detect Echinococcus granulosus (G1 strain) eggs in soil samples.

    PubMed

    Shaikenov, B S; Rysmukhambetova, A T; Massenov, B; Deplazes, P; Mathis, A; Torgerson, P R

    2004-10-01

    Cystic echinococcosis is a re-emerging disease in central Asia. A total of 120 soil samples taken from 30 gardens of rural homesteads in southern Kazakhstan were analyzed for the presence of taeniid eggs using a concentration technique. Of these, 21 (17.5%) were shown to be contaminated with taeniid eggs. These isolated taeniid eggs were further analyzed using a polymerase chain reaction specific for the G1 (sheep) strain of the tapeworm Echinococcus granulosus, and five samples were shown to be positive. This study demonstrates the widespread contamination of the environment with E. granulosus eggs in an Echinococcus-endemic area and thus the potential for indirect transmission of E. granulosus to humans from such sources.

  18. Matrine promotes G0/G1 arrest and down-regulates cyclin D1 expression in human rhabdomyosarcoma cells.

    PubMed

    Guo, L; Xue, T Y; Xu, W; Gao, J Z

    2013-09-01

    Matrine has a broad-spectrum of anti-cancer effects and is efficient in the inhibition of proliferation of hepatoma cells, leukemia cells and neuroblastoma cell. However, its efficacy and tentative mechanisms in rhabdomyosarcoma have not been addressed before. This study aimed to investigate the effects of Matrine on cell cycle and expression of cyclin D1 in human rhabdomyosarcoma cells (RD cell line). RD cell line was treated with different concentrations (0, 0.5, 1.0, and 1.5 mg/mL) of Matrine, and cell proliferation and cell cycle were evaluated using, respectively, MTT assay and flow cytometry. The effect of Matrine on cyclin D1 mRNA levels was measured by RT-PCR. There was a dose-dependent inhibition of proliferation in the matrine-treated group (inhibition of proliferation rate in control cells 12.70 ± 0.35%; Matrine-treated cells [0.5, 1.0, and 1.5 mg/mL]: 31.16 ± 0.11%, 42.96 ± 0.9%, and 57.26 ± 0.8%). The G0 / G1 ratio in study groups were, respectively, 58.44 ± 3.57%, 64.79 ± 2.03%, 69.97 ± 2.89% and 75.03 ± 1.23%.Cyclin D1 mRNA levels progressively diminished (control group ratio of cyclin D1 / β-actin: 0.59 ± 0.06; Matrine: 0.35 ± 0.05, 0.27 ± 0.02 and 0.04 ± 0.03). All aforementioned changes were significant (P<0.05). In conclusion, Matrine markedly suppresses cell proliferation in RD cells by decreasing expression of cyclin D1 mRNA and blocking the cell cycle at the G0 / G1 stage.

  19. Mechanisms of G1 cell cycle arrest and apoptosis in myeloma cells induced by hybrid-compound histone deacetylase inhibitor

    SciTech Connect

    Fujii, Seiko; Okinaga, Toshinori; Ariyoshi, Wataru; Takahashi, Osamu; Iwanaga, Kenjiro; Nishino, Norikazu; Tominaga, Kazuhiro; Nishihara, Tatsuji

    2013-05-10

    Highlights: •Novel histone deacetylase inhibitor Ky-2, remarkably inhibits myeloma cell growth. •Ky-2 demonstrates no cytotoxicity against normal lymphocytic cells. •Ky-2 induces cell cycle arrest through the cell cycle-associated proteins. •Ky-2 induces Bcl-2-inhibitable apoptosis through a caspase-dependent cascade. -- Abstract: Objectives: Histone deacetylase (HDAC) inhibitors are new therapeutic agents, used to treat various types of malignant cancers. In the present study, we investigated the effects of Ky-2, a hybrid-compound HDAC inhibitor, on the growth of mouse myeloma cells. Materials and methods: Myeloma cells, HS-72, P3U1, and mouse normal cells were used in this study. Effect of HDAC inhibitors on cell viability was determined by WST-assay and trypan blue assay. Cell cycle was analyzed using flow cytometer. The expression of cell cycle regulatory and the apoptosis associated proteins were examined by Western blot analysis. Hoechst’s staining was used to detect apoptotic cells. Results: Our findings showed that Ky-2 decreased the levels of HDACs, while it enhanced acetylation of histone H3. Myeloma cell proliferation was inhibited by Ky-2 treatment. Interestingly, Ky-2 had no cytotoxic effects on mouse normal cells. Ky-2 treatment induced G1-phase cell cycle arrest and accumulation of a sub-G1 phase population, while Western blotting analysis revealed that expressions of the cell cycle-associated proteins were up-regulated. Also, Ky-2 enhanced the cleavage of caspase-9 and -3 in myeloma cells, followed by DNA fragmentation. In addition, Ky-2 was not found to induce apoptosis in bcl-2 overexpressing myeloma cells. Conclusion: These findings suggest that Ky-2 induces apoptosis via a caspase-dependent cascade and Bcl-2-inhibitable mechanism in myeloma cells.

  20. Correlating the Impact of Well-Defined Oligosaccharide Structures on Physical Stability Profiles of IgG1-Fc Glycoforms.

    PubMed

    More, Apurva S; Toprani, Vishal M; Okbazghi, Solomon Z; Kim, Jae H; Joshi, Sangeeta B; Middaugh, C Russell; Tolbert, Thomas J; Volkin, David B

    2016-02-01

    As part of a series of articles in this special issue describing 4 well-defined IgG1-Fc glycoforms as a model system for biosimilarity analysis (high mannose-Fc, Man5-Fc, GlcNAc-Fc and N297Q-Fc aglycosylated), the focus of this work is comparisons of their physical properties. A trend of decreasing apparent solubility (thermodynamic activity) by polyethylene glycol precipitation (pH 4.5, 6.0) and lower conformational stability by differential scanning calorimetry (pH 4.5) was observed with reducing size of the N297-linked oligosaccharide structures. Using multiple high-throughput biophysical techniques, the physical stability of the Fc glycoproteins was then measured in 2 formulations (NaCl and sucrose) across a wide range of temperatures (10°C-90°C) and pH (4.0-7.5) conditions. The data sets were used to construct 3-index empirical phase diagrams and radar charts to visualize the regions of protein structural stability. Each glycoform showed improved stability in the sucrose (vs. salt) formulation. The HM-Fc and Man5-Fc displayed the highest relative stability, followed by GlcNAc-Fc, with N297Q-Fc being the least stable. Thus, the overall physical stability profiles of the 4 IgG1-Fc glycoforms also show a correlation with oligosaccharide structure. These data sets are used to develop a mathematical model for biosimilarity analysis (as described in a companion article by Kim et al. in this issue).

  1. Altered Signaling in the G1 Phase Deregulates Chondrocyte Growth in a Mouse Model With Proteoglycan Undersulfation

    PubMed Central

    Leonardis, Fabio De; Monti, Luca; Gualeni, Benedetta; Tenni, Ruggero; Forlino, Antonella; Rossi, Antonio

    2014-01-01

    In several skeletal dysplasias defects in extracellular matrix molecules affect not only the structural and mechanical properties of cartilage, but also the complex network of signaling pathways involved in cell proliferation and differentiation. Sulfated proteoglycans, besides playing an important structural role in cartilage, are crucial in modulating the transport, diffusion, and interactions of growth factors with their specific targets, taking part in the regulation of signaling pathways involved in skeletal development and growth. In this work, we investigated by real time PCR and Western blots of the microdissected growth plate and by immunohistochemistry the molecular basis of reduced chondrocyte proliferation in the growth plate of the dtd mouse, a chondrodysplastic model with defective chondroitin sulfate proteoglycan sulfation of articular and growth plate cartilage. We detected activation of the Wnt pathway, leading to an increase in the non-phosphorylated form of nuclear β-catenin and subsequent up-regulation of cyclin D1 expression in the G1 phase of the cell cycle. β-Catenin was further stabilized by up-regulation of Smad3 expression through TGF-β pathway synergistic activation. We demonstrate that notwithstanding cyclin D1 expression increase, cell cycle progression is compromised in the G1 phase due to reduced phosphorylation of the pocket protein p130 leading to inhibition of transcription factors of the E2F family which are crucial for cell cycle progression and DNA replication. These data, together with altered Indian hedgehox signaling detected previously, explain at the molecular level the reduced chondrocyte proliferation rate of the dtd growth plate leading to reduced skeletal growth. J. Cell. Biochem. 115: 1779–1786, 2014. PMID:24820054

  2. Downregulated miR-45 Inhibits the G1-S Phase Transition by Targeting Bmi-1 in Breast Cancer

    PubMed Central

    Wang, Lan; Liu, Jun-Ling; Yu, Liang; Liu, Xiang-Xia; Wu, Hong-Mei; Lei, Fang-Yong; Wu, Shu; Wang, Xi

    2015-01-01

    Abstract Bmi-1 (B cell-specific Moloney murine leukemia virus integration site 1) is upregulated in breast cancer and was involved in many malignant progressions of breast cells, including cell proliferation, stem cell pluripotency, and cancer initiation. However, the epigenetic regulatory mechanism of Bmi-1 in breast cancer remains unclear. After analysis of the ArrayExpress dataset GSE45666, we comparatively detected the expression levels of miR-495 in 9 examined breast cancer cell lines, normal breast epithelial cells and 8 pairs of fresh clinical tumor samples. Furthermore, to evaluate the effect of miR-495 on the progression of breast cancer, MCF-7 and MDA-MB-231 were transduced to stably overexpress miR-495. The 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide assay, colony formation assays, 5-Bromo-2-deoxyUridine labeling and immunofluorescence, anchorage-independent growth ability assay, flow cytometry analysis, and luciferase assays were used to test the effect of miR-495 in MCF-7 and MDA-MB-231 cells in vitro. Xenografted tumor model was also used to evaluate the effect of miR-495 in breast cancer. Herein, we found that miR-495, a predicted regulator of Bmi-1, was frequently downregulated in malignant cells and tissues of breast. Upregulation of miR-495 significantly suppressed breast cancer cell proliferation and tumorigenicity via G1-S arrest. Further analysis revealed that miR-495 targeted Bmi-1 through its 3′ untranslated region. Moreover, Bmi-1 could neutralize the suppressive effect of miR-495 on cell proliferation and tumorigenicity of breast cancer in vivo. These data suggested that miR-495 could inhibit the G1-S phase transition that leads to proliferation and tumorigenicity inhibition by targeting and suppressing Bmi-1 in breast cancer. PMID:26020378

  3. Impact of postharvest processing on the fungal population contaminating African walnut shells (Tetracarpidium conophorum Mull. Arg) at different maturity stages and potential mycotoxigenic implications.

    PubMed

    Nkwonta, Chikere G; Medina, Angel; Del Carmen Alamar, Maria; Terry, Leon A

    2015-02-01

    ), and B2 (AFB2) were produced by 20 isolates with both AFG1 and AFB1 being predominant at concentration ranges 4.33-32,200 and 4.20-22,700ng/g plug weight, respectively. No ochratoxin A (OTA) was detected out of 23 isolates analysed. From these findings, it is suggested that toasting of nuts, preferably at early maturity is a safer processing option than boiling in terms of prevention of possible fungal growth on nut shells and risk of mycotoxin contamination.

  4. Prolonged early G1 arrest by selective CDK4/CDK6 inhibition sensitizes myeloma cells to cytotoxic killing through cell cycle–coupled loss of IRF4

    PubMed Central

    Huang, Xiangao; Di Liberto, Maurizio; Jayabalan, David; Liang, Jun; Ely, Scott; Bretz, Jamieson; Shaffer, Arthur L.; Louie, Tracey; Chen, Isan; Randolph, Sophia; Hahn, William C.; Staudt, Louis M.; Niesvizky, Ruben; Moore, Malcolm A. S.

    2012-01-01

    Dysregulation of cyclin-dependent kinase 4 (CDK4) and CDK6 by gain of function or loss of inhibition is common in human cancer, including multiple myeloma, but success in targeting CDK with broad-spectrum inhibitors has been modest. By selective and reversible inhibition of CDK4/CDK6, we have developed a strategy to both inhibit proliferation and enhance cytotoxic killing of cancer cells. We show that induction of prolonged early-G1 arrest (pG1) by CDK4/CDK6 inhibition halts gene expression in early-G1 and prevents expression of genes programmed for other cell-cycle phases. Removal of the early-G1 block leads to S-phase synchronization (pG1-S) but fails to completely restore scheduled gene expression. Consequently, the IRF4 protein required to protect myeloma cells from apoptosis is markedly reduced in pG1 and further in pG1-S in response to cytotoxic agents, such as the proteasome inhibitor bortezomib. The coordinated loss of IRF4 and gain of Bim sensitize myeloma tumor cells to bortezomib-induced apoptosis in pG1 in the absence of Noxa and more profoundly in pG1-S in cooperation with Noxa in vitro. Induction of pG1 and pG1-S by reversible CDK4/CDK6 inhibition further augments tumor-specific bortezomib killing in myeloma xenografts. Reversible inhibition of CDK4/CDK6 in sequential combination therapy thus represents a novel mechanism-based cancer therapy. PMID:22718837

  5. [Comparative visual field study using the Octopus 2,000R with the global analysis program G1 with the grid pattern].

    PubMed

    Kieselbach, G F; Juen, S

    1988-01-01

    At the Universitäts-Augenklinik Innsbruck patients with diabetic macular edema are treated with grid pattern since February 1986. Thirty-two eyes from 18 patients were tested with the program G1 on perimeter Octopus 2000R before and after grid pattern. The evaluated parameters presented by the program G1 are discussed. PMID:3362513

  6. Asteroid observations at low phase angles. IV. Average parameters for the new H, G1, G2 magnitude system

    NASA Astrophysics Data System (ADS)

    Shevchenko, Vasilij G.; Belskaya, Irina N.; Muinonen, Karri; Penttilä, Antti; Krugly, Yurij N.; Velichko, Feodor P.; Chiorny, Vasilij G.; Slyusarev, Ivan G.; Gaftonyuk, Ninel M.; Tereschenko, Igor A.

    2016-04-01

    We present new observational data for selected main-belt asteroids of different compositional types. The detailed magnitude-phase dependences including small phase angles (<1°) were obtained for these asteroids, namely: (10) Hygiea (down to the phase angle of 0.3°, C-type), (176) Iduna (0.2°, G-type), (214) Aschera (0.2°, E-type), (218) Bianca (0.3°, S-type), (250) Bettina (0.3°, M-type), (419) Aurelia (0.1°, F-type), (596) Scheila (0.2°, D-type), (635) Vundtia (0.2°, B-type), (671) Carnegia (0.2°, P-type), (717) Wisibada (0.1°, T-type), (1021) Flammario (0.6°, B-type), and (1279) Uganda (0.5°, E-type). For several asteroids, the dependences of brightness on the phase angle were investigated in the BVRI bands. We found a great diversity in the opposition-effect behavior both in the magnitude and the width of the opposition surges, especially for low-albedo asteroids. Some low-albedo asteroids (e.g., (10) Hygiea) display a broad opposition effect with an amplitude of 0.15-0.20 mag relative to the extrapolation of the linear part of the phase curve. Other asteroids (e.g., (596) Scheila, (1021) Flammario) show linear magnitude-phase dependences down to small phase angles (0.1-0.2°). Using numerous data sets on the magnitude-phase dependences with extensive phase-angle coverage, we examined in more detail the new three-parameter H, G1, G2 magnitude system. We determined the values of the G1 and G2 parameters for magnitude phase dependences of individual asteroids and obtained the average parameters for main asteroid compositional types. The values obtained can be used for the estimation of the absolute magnitude of an asteroid from a single observed magnitude when the magnitude-phase dependency is unknown and/or to calculate a visible magnitude for the ephemerides.

  7. An Old Story Retold: Loss of G1 Control Defines A Distinct Genomic Subtype of Esophageal Squamous Cell Carcinoma

    PubMed Central

    Wang, Qiyan; Bai, Jian; Abliz, Amir; Liu, Ying; Gong, Kenan; Li, Jingjing; Shi, Wenjie; Pan, Yaqi; Liu, Fangfang; Lai, Shujuan; Yang, Haijun; Lu, Changdong; Zhang, Lixin; Chen, Wei; Xu, Ruiping; Cai, Hong; Ke, Yang; Zeng, Changqing

    2015-01-01

    Esophageal squamous cell carcinoma (ESCC) has a high mortality rate. To determine the molecular basis of ESCC development, this study sought to identify characteristic genome-wide alterations in ESCC, including exonic mutations and structural alterations. The clinical implications of these genetic alterations were also analyzed. Exome sequencing and verification were performed for nine pairs of ESCC and the matched blood samples, followed by validation with additional samples using Sanger sequencing. Whole-genome SNP arrays were employed to detect copy number alteration (CNA) and loss of heterozygosity (LOH) in 55 cases, including the nine ESCC samples subjected to exome sequencing. A total of 108 non-synonymous somatic mutations (NSSMs) in 102 genes were verified in nine patients. The chromatin modification process was found to be enriched in our gene ontology (GO) analysis. Tumor genomes with TP53 mutations were significantly more unstable than those without TP53 mutations. In terms of the landscape of genomic alterations, deletion of 9p21.3 covering CDKN2A/2B (30.9%), amplification of 11q13.3 covering CCND1 (30.9%), and TP53 point mutation (50.9%) occurred in two-thirds of the cases. These results suggest that the deregulation of the G1 phase during the cell cycle is a key event in ESCC. Furthermore, six minimal common regions were found to be significantly altered in ESCC samples and three of them, 9p21.3, 7p11.2, and 3p12.1, were associated with lymph node metastasis. With the high correlation of TP53 mutation and genomic instability in ESCC, the amplification of CCND1, the deletion of CDKN2A/2B, and the somatic mutation of TP53 appear to play pivotal roles via G1 deregulation and therefore helps to classify this cancer into different genomic subtypes. These findings provide clinical significance that could be useful in future molecular diagnoses and therapeutic targeting. PMID:26386145

  8. Characterization of the Expression of the RNA Binding Protein eIF4G1 and Its Clinicopathological Correlation with Serous Ovarian Cancer

    PubMed Central

    Xie, Zhe; Li, Guiqin; Mao, Chengyi; Liu, Yi; Wen, Xin; Yin, Na; Cao, Jianzhong; Wang, Jing; Li, Li; Yu, Jianhua; Wang, Fang; Yi, Ping

    2016-01-01

    Background Ovarian cancer is the most lethal type of malignant tumor in gynecological cancers and is associated with a high percentage of late diagnosis and chemotherapy resistance. Thus, it is urgent to identify a tumor marker or a molecular target that allows early detection and effective treatment. RNA-binding proteins (RBPs) are crucial in various cellular processes at the post-transcriptional level. The eukaryotic translation initiation factor 4 gamma, 1(eIF4G1), an RNA-binding protein, facilitates the recruitment of mRNA to the ribosome, which is a rate-limiting step during the initiation phase of protein synthesis. However, little is known regarding the characteristics of eIF4G1 expression and its clinical significance in ovarian cancer. Therefore, we propose to investigate the expression and clinicopathological significance of eIF4G1 in ovarian cancer patients. Methods We performed Real-time PCR in 40 fresh serous ovarian cancer tissues and 27 normal ovarian surface epithelial cell specimens to assess eIF4G1mRNA expression. Immunohistochemistry (IHC) was used to examine the expression of eIF4G1 at the protein level in 134 patients with serous ovarian cancer and 18 normal ovarian tissues. Statistical analysis was conducted to determine the correlation of the eIF4G1 protein levels with the clinicopathological characteristics and prognosis in ovarian cancer. Results The expression of eIF4G1 was upregulated in serous ovarian cancer tissues at both the mRNA (P = 0.0375) and the protein (P = 0.0007) levels. The eIF4G1 expression was significantly correlated with the clinical tumor stage (P = 0.0004) and omentum metastasis (P = 0.024). Moreover, patients with low eIF4G1 protein expression had a longer overall survival time (P = 0.026). Conclusions These data revealed that eIF4G1 is markedly expressed in serous ovarian cancer and that upregulation of the eIF4G1 protein expression is significantly associated with an advanced tumor stage. Besides, the patients with

  9. Impact of methionine oxidation in human IgG1 Fc on serum half-life of monoclonal antibodies.

    PubMed

    Wang, Weirong; Vlasak, Josef; Li, Yunsong; Pristatsky, Pavlo; Fang, Yulin; Pittman, Tamara; Roman, Jeanette; Wang, Yang; Prueksaritanont, Thomayant; Ionescu, Roxana

    2011-03-01

    IgG monoclonal antibodies (mAbs) consist of two Fab fragments and one Fc fragment. The Fab fragments contain the variable regions and are responsible for drug specificity (via antigen binding); the Fc fragment contains constant regions and is responsible for effector functions (via interactions with Fcγ receptors) and extended serum half-life (via interaction with the neonatal Fc receptor, FcRn). There are two conserved methionine (Met) residues located in the FcRn binding site of the Fc fragment. It has been shown previously that oxidation of these two Met residues decreases the binding affinity to FcRn. We have further evaluated the impact of Met oxidation on serum half-lives of two humanized IgG1 mAbs in transgenic mice with human FcRn. Variable oxidation levels were obtained by several procedures: exposure to an oxidizing agent, accumulation during extended refrigerated storage, or chromatographic separation. Our results show that Met oxidation can result in a significant reduction of the serum circulation half-life and the magnitude of the change correlates well with the extent of Met oxidation and changes in FcRn binding affinities. The relatively low levels of Met oxidation accumulated during 3 years of refrigerated storage had minimal impact on FcRn binding and no detectable impact on the serum half-life.

  10. Casz1 is required for cardiomyocyte G1-to-S phase progression during mammalian cardiac development.

    PubMed

    Dorr, Kerry M; Amin, Nirav M; Kuchenbrod, Lauren M; Labiner, Hanna; Charpentier, Marta S; Pevny, Larysa H; Wessels, Andy; Conlon, Frank L

    2015-06-01

    Organ growth occurs through the integration of external growth signals during the G1 phase of the cell cycle to initiate DNA replication. Although numerous growth factor signals have been shown to be required for the proliferation of cardiomyocytes, genetic studies have only identified a very limited number of transcription factors that act to regulate the entry of cardiomyocytes into S phase. Here, we report that the cardiac para-zinc-finger protein CASZ1 is expressed in murine cardiomyocytes. Genetic fate mapping with an inducible Casz1 allele demonstrates that CASZ1-expressing cells give rise to cardiomyocytes in the first and second heart fields. We show through the generation of a cardiac conditional null mutation that Casz1 is essential for the proliferation of cardiomyocytes in both heart fields and that loss of Casz1 leads to a decrease in cardiomyocyte cell number. We further report that the loss of Casz1 leads to a prolonged or arrested S phase, a decrease in DNA synthesis, an increase in phospho-RB and a concomitant decrease in the cardiac mitotic index. Taken together, these studies establish a role for CASZ1 in mammalian cardiomyocyte cell cycle progression in both the first and second heart fields.

  11. Oleanolic acid induces mitochondrial-dependent apoptosis and G0/G1 phase arrest in gallbladder cancer cells

    PubMed Central

    Li, Huai-Feng; Wang, Xu-An; Xiang, Shan-Shan; Hu, Yun-Ping; Jiang, Lin; Shu, Yi-Jun; Li, Mao-Lan; Wu, Xiang-Song; Zhang, Fei; Ye, Yuan-Yuan; Weng, Hao; Bao, Run-Fa; Cao, Yang; Lu, Wei; Dong, Qian; Liu, Ying-Bin

    2015-01-01

    Oleanolic acid (OA), a naturally occurring triterpenoid, exhibits potential antitumor activity in many tumor cell lines. Gallbladder carcinoma is the most common malignancy of the biliary tract, and is a highly aggressive tumor with an extremely poor prognosis. Unfortunately, the effects of OA on gallbladder carcinoma are unknown. In this study, we investigated the effects of OA on gallbladder cancer cells and the underlying mechanism. The results showed that OA inhibits proliferation of gallbladder cancer cells in a dose-dependent and time-dependent manner on MTT and colony formation assay. A flow cytometry assay revealed apoptosis and G0/G1 phase arrest in GBC-SD and NOZ cells. Western blot analysis and a mitochondrial membrane potential assay demonstrated that OA functions through the mitochondrial apoptosis pathway. Moreover, this drug inhibited tumor growth in nude mice carrying subcutaneous NOZ tumor xenografts. These data suggest that OA inhibits proliferation of gallbladder cancer cells by regulating apoptosis and the cell cycle process. Thus, OA may be a promising drug for adjuvant chemotherapy in gallbladder carcinoma. PMID:26109845

  12. Direct radiocarbon dates for Vindija G1 and Velika Pećina Late Pleistocene hominid remains

    PubMed Central

    Smith, Fred H.; Trinkaus, Erik; Pettitt, Paul B.; Karavanić, Ivor; Paunović, Maja

    1999-01-01

    New accelerator mass spectrometry radiocarbon dates taken directly on human remains from the Late Pleistocene sites of Vindija and Velika Pećina in the Hrvatsko Zagorje of Croatia are presented. Hominid specimens from both sites have played critical roles in the development of current perspectives on modern human evolutionary emergence in Europe. Dates of ≈28 thousand years (ka) before the present (B.P.) and ≈29 ka B.P. for two specimens from Vindija G1 establish them as the most recent dated Neandertals in the Eurasian range of these archaic humans. The human frontal bone from Velika Pećina, generally considered one of the earliest representatives of modern humans in Europe, dated to ≈5 ka B.P., rendering it no longer pertinent to discussions of modern human origins. Apart from invalidating the only radiometrically based example of temporal overlap between late Neandertal and early modern human fossil remains from within any region of Europe, these dates raise the question of when early modern humans first dispersed into Europe and have implications for the nature and geographic patterning of biological and cultural interactions between these populations and the Neandertals. PMID:10535913

  13. The G1/S Specific Cyclin D2 Is a Regulator of HIV-1 Restriction in Non-proliferating Cells

    PubMed Central

    Badia, Roger; Pujantell, Maria; Riveira-Muñoz, Eva; Puig, Teresa; Torres-Torronteras, Javier; Martí, Ramón; Clotet, Bonaventura; Ampudia, Rosa M.; Ballana, Ester

    2016-01-01

    Macrophages are a heterogeneous cell population strongly influenced by differentiation stimuli that become susceptible to HIV-1 infection after inactivation of the restriction factor SAMHD1 by cyclin-dependent kinases (CDK). Here, we have used primary human monocyte-derived macrophages differentiated through different stimuli to evaluate macrophage heterogeneity on cell activation and proliferation and susceptibility to HIV-1 infection. Stimulation of monocytes with GM-CSF induces a non-proliferating macrophage population highly restrictive to HIV-1 infection, characterized by the upregulation of the G1/S-specific cyclin D2, known to control early steps of cell cycle progression. Knockdown of cyclin D2, enhances HIV-1 replication in GM-CSF macrophages through inactivation of SAMHD1 restriction factor by phosphorylation. Co-immunoprecipitation experiments show that cyclin D2 forms a complex with CDK4 and p21, a factor known to restrict HIV-1 replication by affecting the function of the downstream cascade that leads to SAMHD1 deactivation. Thus, we demonstrate that cyclin D2 acts as regulator of cell cycle proteins affecting SAMHD1-mediated HIV-1 restriction in non-proliferating macrophages. PMID:27541004

  14. Histone acetyltransferase HAT4 modulates navigation across G2/M and re-entry into G1 in Leishmania donovani

    PubMed Central

    Yadav, Aarti; Chandra, Udita; Saha, Swati

    2016-01-01

    Histone acetyltransferases impact multiple processes. This study investigates the role of histone acetyltransferase HAT4 in Leishmania donovani. Though HAT4 was dispensable for survival, its elimination decreased cell viability and caused cell cycle defects, with HAT4-nulls experiencing an unusually long G2/M. Survival of HAT4-nulls in macrophages was also substantially compromised. DNA microarray analysis revealed that HAT4 modestly regulated the expression of only a select number of genes, thus not being a major modulator of global gene expression. Significantly, cdc20 was among the downregulated genes. To ascertain if decreased expression of cdc20 was responsible for HAT4-null growth and cell cycle defects we expressed LdCdc20 ectopically in HAT4-nulls. We found this to alleviate the aberrant growth and cell cycle progression patterns displayed by HAT4-nulls, with cells navigating G2/M phase and re-entering G1 phase smoothly. HAT4-nulls expressing LdCdc20 ectopically showed survival rates comparable to wild type within macrophages, suggesting that G2/M defects were responsible for poor survival of HAT4-nulls within host cells also. These are the first data analyzing the in vivo functional role of HAT4 in any trypanosomatid. Our results directly demonstrate for the first time a role for Cdc20 in regulating trypanosomatid G2/M events, opening avenues for further research in this area. PMID:27272906

  15. A short G1 phase imposes constitutive replication stress and fork remodelling in mouse embryonic stem cells

    PubMed Central

    Ahuja, Akshay K.; Jodkowska, Karolina; Teloni, Federico; Bizard, Anna H.; Zellweger, Ralph; Herrador, Raquel; Ortega, Sagrario; Hickson, Ian D.; Altmeyer, Matthias; Mendez, Juan; Lopes, Massimo

    2016-01-01

    Embryonic stem cells (ESCs) represent a transient biological state, where pluripotency is coupled with fast proliferation. ESCs display a constitutively active DNA damage response (DDR), but its molecular determinants have remained elusive. Here we show in cultured ESCs and mouse embryos that H2AX phosphorylation is dependent on Ataxia telangiectasia and Rad3 related (ATR) and is associated with chromatin loading of the ssDNA-binding proteins RPA and RAD51. Single-molecule analysis of replication intermediates reveals massive ssDNA gap accumulation, reduced fork speed and frequent fork reversal. All these marks of replication stress do not impair the mitotic process and are rapidly lost at differentiation onset. Delaying the G1/S transition in ESCs allows formation of 53BP1 nuclear bodies and suppresses ssDNA accumulation, fork slowing and reversal in the following S-phase. Genetic inactivation of fork slowing and reversal leads to chromosomal breakage in unperturbed ESCs. We propose that rapid cell cycle progression makes ESCs dependent on effective replication-coupled mechanisms to protect genome integrity. PMID:26876348

  16. Cystic echinococcosis in water buffaloes: epidemiological survey and molecular evidence of ovine (G1) and buffalo (G3) strains.

    PubMed

    Capuano, F; Rinaldi, L; Maurelli, M P; Perugini, A G; Veneziano, V; Garippa, G; Genchi, C; Musella, V; Cringoli, G

    2006-04-30

    A survey of cystic echinococcosis (CE) in the water buffalo (Bubalus bubalis) of the Italian Mediterranean breed was carried out in Campania, a region of southern Italy. In addition, a molecular study was performed on 48 hydatid cysts coming from 48 water buffaloes in order to determine the Echinococcus granulosus strain(s) present in this host. Out of a total of 722 water buffaloes examined for CE, 76 (10.5%) were found infected. The average number of cysts per buffalo was 4.3 (minimum 1, maximum 45). Seventeen buffaloes had hydatid cysts only in the liver (with an average of 5 cysts/liver), 34 only in the lungs (with an average of 1.8 cysts/lungs), and 25 buffaloes had cysts both in the liver and in the lungs. Fertile cysts were found in 10 (13.2%) out of the 76 positive buffaloes. The sequencing of the mitochondrial cytochrome C oxidase subunit 1 (CO1) gene of the 48 hydatid cysts produced sequences of 419 bp for each sample analysed. For 33 samples, alignment of the obtained sequences with those present in GenBank showed a total homology with the common domestic sheep strain G1; for 15 samples, sequences obtained showed 100% homology with buffalo strain G3. The findings of the present survey represent the first epidemiological and molecular comprehensive studies on CE in water buffalo from an endemic area for E. granulosus.

  17. CDK1-Cyclin B1 Activates RNMT, Coordinating mRNA Cap Methylation with G1 Phase Transcription.

    PubMed

    Aregger, Michael; Kaskar, Aneesa; Varshney, Dhaval; Fernandez-Sanchez, Maria Elena; Inesta-Vaquera, Francisco A; Weidlich, Simone; Cowling, Victoria H

    2016-03-01

    The creation of translation-competent mRNA is dependent on RNA polymerase II transcripts being modified by addition of the 7-methylguanosine (m7G) cap. The factors that mediate splicing, nuclear export, and translation initiation are recruited to the transcript via the cap. The cap structure is formed by several activities and completed by RNMT (RNA guanine-7 methyltransferase), which catalyzes N7 methylation of the cap guanosine. We report that CDK1-cyclin B1 phosphorylates the RNMT regulatory domain on T77 during G2/M phase of the cell cycle. RNMT T77 phosphorylation activates the enzyme both directly and indirectly by inhibiting interaction with KPNA2, an RNMT inhibitor. RNMT T77 phosphorylation results in elevated m7G cap methyltransferase activity at the beginning of G1 phase, coordinating mRNA capping with the burst of transcription that occurs following nuclear envelope reformation. RNMT T77 phosphorylation is required for the production of cohort of proteins, and inhibiting T77 phosphorylation reduces the cell proliferation rate. PMID:26942677

  18. Ponicidin suppresses HT29 cell growth via the induction of G1 cell cycle arrest and apoptosis.

    PubMed

    Du, Jie; Chen, Chunyou; Sun, Yiqun; Zheng, Lin; Wang, Wanchen

    2015-10-01

    Ponicidin is a diterpenoid extracted from the Chinese herb Isodon adenolomus, which has been reported as a therapeutic cytotoxic drug that may be used to treat various types of human cancer. The present study aimed to determine the antitumor effects of ponicidin, and to investigate its underlying mechanisms in colorectal cancer. The HT29 colorectal cancer cell line was used to detect the cytotoxicity of various doses of ponicidin. Cell proliferation was measured using a Cell Counting kit‑8 assay. Cell cycle and apoptosis analyses were performed using flow cytometry and fluorescent microscopy. Western blot analysis was used to measure the expression levels of apoptosis‑associated proteins following treatment with ponicidin. Treatment with ponicidin significantly suppressed HT29 cell growth by inducing G1 cell cycle arrest and apoptosis. The AKT and MEK signaling pathways were also suppressed by ponicidin; however, the p38 signaling pathway was significantly activated. The expression levels of caspase 3 and Bax protein were markedly upregulated following treatment with ponicidin. These results suggest that ponicidin exerts significant antitumor effects via the induction of cell cycle arrest and apoptosis in colorectal cells. In conclusion, ponicidin acted as an inducer of apoptosis, and may be used as a therapeutic cytotoxic drug to treat human cancer, including colorectal cancer. PMID:26239027

  19. Ponicidin suppresses HT29 cell growth via the induction of G1 cell cycle arrest and apoptosis

    PubMed Central

    DU, JIE; CHEN, CHUNYOU; SUN, YIQUN; ZHENG, LIN; WANG, WANCHEN

    2015-01-01

    Ponicidin is a diterpenoid extracted from the Chinese herb Isodon adenolomus, which has been reported as a therapeutic cytotoxic drug that may be used to treat various types of human cancer. The present study aimed to determine the antitumor effects of ponicidin, and to investigate its underlying mechanisms in colorectal cancer. The HT29 colorectal cancer cell line was used to detect the cytotoxicity of various doses of ponicidin. Cell proliferation was measured using a Cell Counting kit-8 assay. Cell cycle and apoptosis analyses were performed using flow cytometry and fluorescent microscopy. Western blot analysis was used to measure the expression levels of apoptosis-associated proteins following treatment with ponicidin. Treatment with ponicidin significantly suppressed HT29 cell growth by inducing G1 cell cycle arrest and apoptosis. The AKT and MEK signaling pathways were also suppressed by ponicidin; however, the p38 signaling pathway was significantly activated. The expression levels of caspase 3 and Bax protein were markedly upregulated following treatment with ponicidin. These results suggest that ponicidin exerts significant antitumor effects via the induction of cell cycle arrest and apoptosis in colorectal cells. In conclusion, ponicidin acted as an inducer of apoptosis, and may be used as a therapeutic cytotoxic drug to treat human cancer, including colorectal cancer. PMID:26239027

  20. CDK1-Cyclin B1 Activates RNMT, Coordinating mRNA Cap Methylation with G1 Phase Transcription

    PubMed Central

    Aregger, Michael; Kaskar, Aneesa; Varshney, Dhaval; Fernandez-Sanchez, Maria Elena; Inesta-Vaquera, Francisco A.; Weidlich, Simone; Cowling, Victoria H.

    2016-01-01

    Summary The creation of translation-competent mRNA is dependent on RNA polymerase II transcripts being modified by addition of the 7-methylguanosine (m7G) cap. The factors that mediate splicing, nuclear export, and translation initiation are recruited to the transcript via the cap. The cap structure is formed by several activities and completed by RNMT (RNA guanine-7 methyltransferase), which catalyzes N7 methylation of the cap guanosine. We report that CDK1-cyclin B1 phosphorylates the RNMT regulatory domain on T77 during G2/M phase of the cell cycle. RNMT T77 phosphorylation activates the enzyme both directly and indirectly by inhibiting interaction with KPNA2, an RNMT inhibitor. RNMT T77 phosphorylation results in elevated m7G cap methyltransferase activity at the beginning of G1 phase, coordinating mRNA capping with the burst of transcription that occurs following nuclear envelope reformation. RNMT T77 phosphorylation is required for the production of cohort of proteins, and inhibiting T77 phosphorylation reduces the cell proliferation rate. PMID:26942677

  1. Construction of pH-sensitive Her2-binding IgG1-Fc by directed evolution.

    PubMed

    Traxlmayr, Michael W; Lobner, Elisabeth; Hasenhindl, Christoph; Stadlmayr, Gerhard; Oostenbrink, Chris; Rüker, Florian; Obinger, Christian

    2014-08-01

    For most therapeutic proteins, a long serum half-life is desired. Studies have shown that decreased antigen binding at acidic pH can increase serum half-life. In this study, we aimed to investigate whether pH-dependent binding sites can be introduced into antigen binding crystallizable fragments of immunoglobulin G1 (Fcab). The C-terminal structural loops of an Fcab were engineered for reduced binding to the extracellular domain of human epidermal growth factor receptor 2 (Her2-ECD) at pH 6 compared to pH 7.4. A yeast-displayed Fcab-library was alternately selected for binding at pH 7.4 and non-binding at pH 6.0. Selected Fcab variants showed clear pH-dependent binding to soluble Her2-ECD (decrease in affinity at pH 6.0 compared to pH 7.4) when displayed on yeast. Additionally, some solubly expressed variants exhibited pH-dependent interactions with Her2-positive cells whereas their conformational and thermal stability was pH-independent. Interestingly, two of the three Fcabs did not contain a single histidine mutation but all of them contained variations next to histidines that already occurred in loops of the lead Fcab. The study demonstrates that yeast surface display is a valuable tool for directed evolution of pH-dependent binding sites in proteins. PMID:24964247

  2. Effect of ambient light on IgG1 monoclonal antibodies during drug product processing and development.

    PubMed

    Sreedhara, Alavattam; Yin, Jian; Joyce, Michael; Lau, Kimberly; Wecksler, Aaron T; Deperalta, Galahad; Yi, Li; John Wang, Y; Kabakoff, Bruce; Kishore, Ravuri S K

    2016-03-01

    Photostability studies are standard stress testing conducted during drug product development of various pharmaceutical compounds, including small molecules and proteins. These studies as recommended by ICH Q1B are carried out using no less than 1.2× 10(6)lux-hours in the visible region and no less than 200Wh/m(2) in UV light. However, normal drug product processing is carried out under fluorescent lamps that emit white light almost exclusively in the >400nm region with a small UV quotient. We term these as ambient or mild light conditions. We tested several IgG1 monoclonal antibodies (mAbs 1-5) under these ambient light conditions and compared them to the ICH light conditions. All the mAbs were significantly degraded under the ICH light but several mAbs (mAbs 3-5) were processed without impacting any product quality attributes under ambient or mild light conditions. Interestingly we observed site-specific Trp oxidation in mAb1, while higher aggregation and color change were observed for mAb2 under mild light conditions. The recommended ICH light conditions have a high UV component and hence may not help to rank order photosensitivity under normal protein DP processing conditions.

  3. Inhibition of G1P3 expression found in the differential display study on respiratory syncytial virus infection

    PubMed Central

    Zhao, Dongchi; Peng, Dan; Li, Lei; Zhang, Qiwei; Zhang, Chuyu

    2008-01-01

    Background Respiratory syncytial virus (RSV) is the leading viral pathogen associated with bronchiolitis and lower respiratory tract disease in infants and young children worldwide. The respiratory epithelium is the primary initiator of pulmonary inflammation in RSV infections, which cause significant perturbations of global gene expression controlling multiple cellular processes. In this study, differential display reverse transcription polymerase chain reaction amplification was performed to examine mRNA expression in a human alveolar cell line (SPC-A1) infected with RSV. Results Of the 2,500 interpretable bands on denaturing polyacrylamide gels, 40 (1.6%) cDNA bands were differentially regulated by RSV, in which 28 (70%) appeared to be upregulated and another 12 (30%) appeared to be downregulated. Forty of the expressed sequence tags (EST) were isolated, and 20 matched homologs in GenBank. RSV infection upregulated the mRNA expression of chemokines CC and CXC and interfered with type α/β interferon-inducible gene expression by upregulation of MG11 and downregulation of G1P3. Conclusion RSV replication could induce widespread changes in gene expression including both positive and negative regulation and play a different role in the down-regulation of IFN-α and up-regulation of IFN-γ inducible gene expression, which suggests that RSV interferes with the innate antiviral response of epithelial cells by multiple mechanisms. PMID:18838000

  4. The trefoil factor 1 participates in gastrointestinal cell differentiation by delaying G1-S phase transition and reducing apoptosis

    PubMed Central

    Bossenmeyer-Pourié, Carine; Kannan, Rama; Ribieras, Stéphane; Wendling, Corinne; Stoll, Isabelle; Thim, Lars; Tomasetto, Catherine; Rio, Marie-Christine

    2002-01-01

    Trefoil factor (TFF)1 is synthesized and secreted by the normal stomach mucosa and by the gastrointestinal cells of injured tissues. The link between mouse TFF1 inactivation and the fully penetrant antropyloric tumor phenotype prompted the classification of TFF1 as a gastric tumor suppressor gene. Accordingly, altered expression, deletion, and/or mutations of the TFF1 gene are frequently observed in human gastric carcinomas. The present study was undertaken to address the nature of the cellular and molecular mechanisms targeted by TFF1 signalling. TFF1 effects were investigated in IEC18, HCT116, and AGS gastrointestinal cells treated with recombinant human TFF1, and in stably transfected HCT116 cells synthesizing constitutive or doxycycline-induced human TFF1. We observed that TFF1 triggers two types of cellular responses. On one hand, TFF1 lowers cell proliferation by delaying G1-S cell phase transition. This results from a TFF1-mediated increase in the levels of cyclin-dependent kinase inhibitors of both the INK4 and CIP subfamilies, leading to lower E2F transcriptional activity. On the other hand, TFF1 protects cells from chemical-, anchorage-free–, or Bad-induced apoptosis. In this process, TFF1 signalling targets the active form of caspase-9. Together, these results provide the first evidence of a dual antiproliferative and antiapoptotic role for TFF1. Similar paradoxical functions have been reported for tumor suppressor genes involved in cell differentiation, a function consistent with TFF1. PMID:12034770

  5. MicroRNA-206 induces G1 arrest in melanoma by inhibition of CDK4 and Cyclin D.

    PubMed

    Georgantas, Robert W; Streicher, Katie; Luo, Xiaobing; Greenlees, Lydia; Zhu, Wei; Liu, Zheng; Brohawn, Philip; Morehouse, Christopher; Higgs, Brandon W; Richman, Laura; Jallal, Bahija; Yao, Yihong; Ranade, Koustubh

    2014-03-01

    Expression profiling of microRNAs in melanoma lesional skin biopsies compared with normal donor skin biopsies, as well as melanoma cell lines compared with normal melanocytes, revealed that hsa-miR-206 was down-regulated in melanoma (-75.4-fold, P = 1.7 × 10(-4)). MiR-206 has been implicated in a large number of cancers, including breast, lung, colorectal, ovarian, and prostate cancers; however, its role in tumor development remains largely unknown, its biologic function is poorly characterized, and its targets affecting cancer cells are largely unknown. MiR-206 reduced growth and migration/invasion of multiple melanoma cell lines. Bioinformatics identified cell cycle genes CDK2, CDK4, Cyclin C, and Cyclin D1 as strong candidate targets. Western blots and 3'UTR reporter gene assays revealed that miR-206 inhibited translation of CDK4, Cyclin D1, and Cyclin C. Additionally, hsa-miR-206 transfection induced G1 arrest in multiple melanoma cell lines. These observations support hsa-miR-206 as a tumor suppressor in melanoma and identify Cyclin C, Cyclin D1, and CDK4 as miR-206 targets.

  6. Cystic echinococcosis in water buffaloes: epidemiological survey and molecular evidence of ovine (G1) and buffalo (G3) strains.

    PubMed

    Capuano, F; Rinaldi, L; Maurelli, M P; Perugini, A G; Veneziano, V; Garippa, G; Genchi, C; Musella, V; Cringoli, G

    2006-04-30

    A survey of cystic echinococcosis (CE) in the water buffalo (Bubalus bubalis) of the Italian Mediterranean breed was carried out in Campania, a region of southern Italy. In addition, a molecular study was performed on 48 hydatid cysts coming from 48 water buffaloes in order to determine the Echinococcus granulosus strain(s) present in this host. Out of a total of 722 water buffaloes examined for CE, 76 (10.5%) were found infected. The average number of cysts per buffalo was 4.3 (minimum 1, maximum 45). Seventeen buffaloes had hydatid cysts only in the liver (with an average of 5 cysts/liver), 34 only in the lungs (with an average of 1.8 cysts/lungs), and 25 buffaloes had cysts both in the liver and in the lungs. Fertile cysts were found in 10 (13.2%) out of the 76 positive buffaloes. The sequencing of the mitochondrial cytochrome C oxidase subunit 1 (CO1) gene of the 48 hydatid cysts produced sequences of 419 bp for each sample analysed. For 33 samples, alignment of the obtained sequences with those present in GenBank showed a total homology with the common domestic sheep strain G1; for 15 samples, sequences obtained showed 100% homology with buffalo strain G3. The findings of the present survey represent the first epidemiological and molecular comprehensive studies on CE in water buffalo from an endemic area for E. granulosus. PMID:16480832

  7. MicroRNA-206 induces G1 arrest in melanoma by inhibition of CDK4 and Cyclin D.

    PubMed

    Georgantas, Robert W; Streicher, Katie; Luo, Xiaobing; Greenlees, Lydia; Zhu, Wei; Liu, Zheng; Brohawn, Philip; Morehouse, Christopher; Higgs, Brandon W; Richman, Laura; Jallal, Bahija; Yao, Yihong; Ranade, Koustubh

    2014-03-01

    Expression profiling of microRNAs in melanoma lesional skin biopsies compared with normal donor skin biopsies, as well as melanoma cell lines compared with normal melanocytes, revealed that hsa-miR-206 was down-regulated in melanoma (-75.4-fold, P = 1.7 × 10(-4)). MiR-206 has been implicated in a large number of cancers, including breast, lung, colorectal, ovarian, and prostate cancers; however, its role in tumor development remains largely unknown, its biologic function is poorly characterized, and its targets affecting cancer cells are largely unknown. MiR-206 reduced growth and migration/invasion of multiple melanoma cell lines. Bioinformatics identified cell cycle genes CDK2, CDK4, Cyclin C, and Cyclin D1 as strong candidate targets. Western blots and 3'UTR reporter gene assays revealed that miR-206 inhibited translation of CDK4, Cyclin D1, and Cyclin C. Additionally, hsa-miR-206 transfection induced G1 arrest in multiple melanoma cell lines. These observations support hsa-miR-206 as a tumor suppressor in melanoma and identify Cyclin C, Cyclin D1, and CDK4 as miR-206 targets. PMID:24289491

  8. CSBF/C10orf99, a novel potential cytokine, inhibits colon cancer cell growth through inducing G1 arrest.

    PubMed

    Pan, Wen; Cheng, Yingying; Zhang, Heyu; Liu, Baocai; Mo, Xiaoning; Li, Ting; Li, Lin; Cheng, Xiaojing; Zhang, Lianhai; Ji, Jiafu; Wang, Pingzhang; Han, Wenling

    2014-01-01

    Cytokines are soluble proteins that exert their functions by binding specific receptors. Many cytokines play essential roles in carcinogenesis and have been developed for the treatment of cancer. In this study, we identified a novel potential cytokine using immunogenomics designated colon-derived SUSD2 binding factor (CSBF), also known as chromosome 10 open reading frame 99 (C10orf99). CSBF/C10orf99 is a classical secreted protein with predicted molecular mass of 6.5 kDa, and a functional ligand of Sushi Domain Containing 2 (SUSD2). CSBF/C10orf99 has the highest expression level in colon tissue. Both CSBF/C10orf99 and SUSD2 are down-regulated in colon cancer tissues and cell lines with different regulation mechanisms. CSBF/C10orf99 interacts with SUSD2 to inhibit colon cancer cell growth and induce G1 cell cycle arrest by down-regulating cyclin D and cyclin-dependent kinase 6 (CDK6). CSBF/C10orf99 displays a bell-shaped activity curve with the optimal effect at ~10 ng/ml. Its growth inhibitory effects can be blocked by sSUSD2-Fc soluble protein. Our results suggest that CSBF/C10orf99 is a novel potential cytokine with tumor suppressor functions. PMID:25351403

  9. Construction of pH-sensitive Her2-binding IgG1-Fc by directed evolution.

    PubMed

    Traxlmayr, Michael W; Lobner, Elisabeth; Hasenhindl, Christoph; Stadlmayr, Gerhard; Oostenbrink, Chris; Rüker, Florian; Obinger, Christian

    2014-08-01

    For most therapeutic proteins, a long serum half-life is desired. Studies have shown that decreased antigen binding at acidic pH can increase serum half-life. In this study, we aimed to investigate whether pH-dependent binding sites can be introduced into antigen binding crystallizable fragments of immunoglobulin G1 (Fcab). The C-terminal structural loops of an Fcab were engineered for reduced binding to the extracellular domain of human epidermal growth factor receptor 2 (Her2-ECD) at pH 6 compared to pH 7.4. A yeast-displayed Fcab-library was alternately selected for binding at pH 7.4 and non-binding at pH 6.0. Selected Fcab variants showed clear pH-dependent binding to soluble Her2-ECD (decrease in affinity at pH 6.0 compared to pH 7.4) when displayed on yeast. Additionally, some solubly expressed variants exhibited pH-dependent interactions with Her2-positive cells whereas their conformational and thermal stability was pH-independent. Interestingly, two of the three Fcabs did not contain a single histidine mutation but all of them contained variations next to histidines that already occurred in loops of the lead Fcab. The study demonstrates that yeast surface display is a valuable tool for directed evolution of pH-dependent binding sites in proteins.

  10. Metabolism of methoxychlor by the P450-monooxygenase CYP6G1 involved in insecticide resistance of Drosophila melanogaster after expression in cell cultures of Nicotiana tabacum.

    PubMed

    Joussen, Nicole; Schuphan, Ingolf; Schmidt, Burkhard

    2010-03-01

    Cytochrome P450 monooxygenase CYP6G1 of Drosophila melanogaster was heterologously expressed in a cell suspension culture of Nicotiana tabacum. This in vitro system was used to study the capability of CYP6G1 to metabolize the insecticide methoxychlor (=1,1,1-trichloro-2,2-bis(4-methoxyphenyl)ethane, 1) against the background of endogenous enzymes of the corresponding non-transgenic culture. The Cyp6g1-transgenic cell culture metabolized 96% of applied methoxychlor (45.8 microg per assay) within 24 h by demethylation and hydroxylation mainly to trishydroxy and catechol methoxychlor (16 and 17%, resp.). About 34% of the metabolism and the distinct formation of trishydroxy and catechol methoxychlor were due to foreign enzyme CYP6G1. Furthermore, methoxychlor metabolism was inhibited by 43% after simultaneous addition of piperonyl butoxide (458 microg), whereas inhibition in the non-transgenic culture amounted to 92%. Additionally, the rate of glycosylation was reduced in both cultures. These results were supported by the inhibition of the metabolism of the insecticide imidacloprid (6; 20 microg, 24 h) in the Cyp6g1-transgenic culture by 82% in the presence of piperonyl butoxide (200 microg). Due to CYP6G1 being responsible for imidacloprid resistance of Drosophila or being involved in DDT resistance, it is likely that CYP6G1 conveys resistance to methoxychlor (1). Furthermore, treating Drosophila with piperonyl butoxide could weaken the observed resistance phenomena.

  11. Transformed mammalian cells are deficient in kinase-mediated control of progression through the G1 phase of the cell cycle.

    PubMed Central

    Crissman, H A; Gadbois, D M; Tobey, R A; Bradbury, E M

    1991-01-01

    To investigate the role of kinase-mediated mechanisms in regulating mammalian cell proliferation, we determined the effects of the general protein kinase inhibitor staurosporine on the proliferation of a series of nontransformed and transformed cultured rodent and human cells. Levels of staurosporine as low as 1 ng/ml prevented nontransformed cells from entering S phase (i.e., induced G1 arrest), indicating that kinase-mediated processes are essential for commitment to DNA replication in normal cells. At higher concentrations of staurosporine (50-75 ng/ml), nontransformed mammalian cells were arrested in both G1 and G2. The period of sensitivity of nontransformed human diploid fibroblasts to low levels of the drug commenced 3 hr later than the G0/G1 boundary and extended through the G1/S boundary. Interference with activity of the G1-essential kinase(s) caused nontransformed human cells traversing mid-to-late G1 at the time of staurosporine addition to be "set back" to the initial staurosporine block point, suggesting the existence of a kinase-dependent "G1 clock" mechanism that must function continuously throughout the early cycle in normal cells. The initial staurosporine block point at 3 hr into G1 corresponds to neither the serum nor the amino acid restriction point. In marked contrast to the behavior of nontransformed cells, neither low nor high concentrations of staurosporine affected G1 progression in transformed cultures; high drug concentrations caused transformed cells to be arrested solely in G2. These results indicate that kinase-mediated regulation of DNA replication is lost as the result of neoplastic transformation, but the G2-arrest mechanism remains intact. Images PMID:1652754

  12. Evolutionary changes in gene expression, coding sequence and copy-number at the Cyp6g1 locus contribute to resistance to multiple insecticides in Drosophila.

    PubMed

    Harrop, Thomas W R; Sztal, Tamar; Lumb, Christopher; Good, Robert T; Daborn, Phillip J; Batterham, Philip; Chung, Henry

    2014-01-01

    Widespread use of insecticides has led to insecticide resistance in many populations of insects. In some populations, resistance has evolved to multiple pesticides. In Drosophila melanogaster, resistance to multiple classes of insecticide is due to the overexpression of a single cytochrome P450 gene, Cyp6g1. Overexpression of Cyp6g1 appears to have evolved in parallel in Drosophila simulans, a sibling species of D. melanogaster, where it is also associated with insecticide resistance. However, it is not known whether the ability of the CYP6G1 enzyme to provide resistance to multiple insecticides evolved recently in D. melanogaster or if this function is present in all Drosophila species. Here we show that duplication of the Cyp6g1 gene occurred at least four times during the evolution of different Drosophila species, and the ability of CYP6G1 to confer resistance to multiple insecticides exists in D. melanogaster and D. simulans but not in Drosophila willistoni or Drosophila virilis. In D. virilis, which has multiple copies of Cyp6g1, one copy confers resistance to DDT and another to nitenpyram, suggesting that the divergence of protein sequence between copies subsequent to the duplication affected the activity of the enzyme. All orthologs tested conferred resistance to one or more insecticides, suggesting that CYP6G1 had the capacity to provide resistance to anthropogenic chemicals before they existed. Finally, we show that expression of Cyp6g1 in the Malpighian tubules, which contributes to DDT resistance in D. melanogaster, is specific to the D. melanogaster-D. simulans lineage. Our results suggest that a combination of gene duplication, regulatory changes and protein coding changes has taken place at the Cyp6g1 locus during evolution and this locus may play a role in providing resistance to different environmental toxins in different Drosophila species.

  13. Evolutionary Changes in Gene Expression, Coding Sequence and Copy-Number at the Cyp6g1 Locus Contribute to Resistance to Multiple Insecticides in Drosophila

    PubMed Central

    Harrop, Thomas W. R.; Sztal, Tamar; Lumb, Christopher; Good, Robert T.; Daborn, Phillip J.; Batterham, Philip; Chung, Henry

    2014-01-01

    Widespread use of insecticides has led to insecticide resistance in many populations of insects. In some populations, resistance has evolved to multiple pesticides. In Drosophila melanogaster, resistance to multiple classes of insecticide is due to the overexpression of a single cytochrome P450 gene, Cyp6g1. Overexpression of Cyp6g1 appears to have evolved in parallel in Drosophila simulans, a sibling species of D. melanogaster, where it is also associated with insecticide resistance. However, it is not known whether the ability of the CYP6G1 enzyme to provide resistance to multiple insecticides evolved recently in D. melanogaster or if this function is present in all Drosophila species. Here we show that duplication of the Cyp6g1 gene occurred at least four times during the evolution of different Drosophila species, and the ability of CYP6G1 to confer resistance to multiple insecticides exists in D. melanogaster and D. simulans but not in Drosophila willistoni or Drosophila virilis. In D. virilis, which has multiple copies of Cyp6g1, one copy confers resistance to DDT and another to nitenpyram, suggesting that the divergence of protein sequence between copies subsequent to the duplication affected the activity of the enzyme. All orthologs tested conferred resistance to one or more insecticides, suggesting that CYP6G1 had the capacity to provide resistance to anthropogenic chemicals before they existed. Finally, we show that expression of Cyp6g1 in the Malpighian tubules, which contributes to DDT resistance in D. melanogaster, is specific to the D. melanogaster–D. simulans lineage. Our results suggest that a combination of gene duplication, regulatory changes and protein coding changes has taken place at the Cyp6g1 locus during evolution and this locus may play a role in providing resistance to different environmental toxins in different Drosophila species. PMID:24416303

  14. UVB-Induced Cell Death Signaling Is Associated with G1-S Progression and Transcription Inhibition in Primary Human Fibroblasts

    PubMed Central

    Ortolan, Tatiana Grohmann; Menck, Carlos Frederico M.

    2013-01-01

    DNA damage induced by ultraviolet (UV) radiation can be removed by nucleotide excision repair through two sub-pathways, one general (GGR) and the other specific for transcribed DNA (TCR), and the processing of unrepaired lesions trigger signals that may lead to cell death. These signals involve the tumor suppressor p53 protein, a central regulator of cell responses to DNA damage, and the E3 ubiquitin ligase Mdm2, that forms a feedback regulatory loop with p53. The involvement of cell cycle and transcription on the signaling to apoptosis was investigated in UVB-irradiated synchronized, DNA repair proficient, CS-B (TCR-deficient) and XP-C (GGR-deficient) primary human fibroblasts. Cells were irradiated in the G1 phase of the cell cycle, with two doses with equivalent levels of apoptosis (low and high), defined for each cell line. In the three cell lines, the low doses of UVB caused only a transient delay in progression to the S phase, whereas the high doses induced permanent cell cycle arrest. However, while accumulation of Mdm2 correlated well with the recovery from transcription inhibition at the low doses for normal and CS-B fibroblasts, for XP-C cells this protein was shown to be accumulated even at UVB doses that induced high levels of apoptosis. Thus, UVB-induced accumulation of Mdm2 is critical for counteracting p53 activation and apoptosis avoidance, but its effect is limited due to transcription inhibition. However, in the case of XP-C cells, an excess of unrepaired DNA damage would be sufficient to block S phase progression, which would signal to apoptosis, independent of Mdm2 accumulation. The data clearly discriminate DNA damage signals that lead to cell death, depending on the presence of UVB-induced DNA damage in replicating or transcribing regions. PMID:24155908

  15. New analytical techniques for mycotoxins in complex organic matrices. [Aflatoxins B1, B2, G1, and G2

    SciTech Connect

    Bicking, M.K.L.

    1982-07-01

    Air samples are collected for analysis from the Ames Solid Waste Recovery System. The high level of airborne fungi within the processing area is of concern due to the possible presence of toxic mycotoxins, and carcinogenic fungal metabolites. An analytical method has been developed to determine the concentration of aflatoxins B1, B2, G1, and G2 in the air of the plant which produces Refuse Derived Fuel (RDF). After extraction with methanol, some components in the matrix are precipitated by dissolving the sample in 30% acetonitrile/chloroform. An aliquot of this solution is injected onto a Styragel column where the sample components undergo simultaneous size exclusion and reverse phase partitioning. Additional studies have provided a more thorough understanding of solvent related non-exclusion effects on size exclusion gels. The Styragel column appears to have a useable lifetime of more than six months. After elution from Styragel, the sample is diverted to a second column containing Florisil which has been modified with oxalic acid and deactivated with water. Aflatoxins are eluted with 5% water/acetone. After removal of this solvent, the sample is dissolved in 150 ..mu..L of a spotting solvent and the entire sample applied to a thin layer chromatography (TLC) plate using a unique sample applicator developed here. The aflatoxins on the TLC plate are analyzed by laser fluorescence. A detection limit of 10 pg is possible for aflatoxin standards using a nitrogen laser as the excitation source. Sample concentrations are determined by comparing with an internal standard, a specially synthesized aflatoxin derivative. In two separate RDF samples, aflatoxin B1 was found at levels of 6.5 and 17.0 ppB. The analytical method has also proven useful in the analysis of contaminated corn and peanut meal samples. 42 figures, 8 tables.

  16. Galbanic acid decreases androgen receptor abundance and signaling and induces G1 arrest in prostate cancer cells

    PubMed Central

    Zhang, Yong; Kim, Kwan-Hyun; Zhang, Wei; Guo, Yinglu; Kim, Sung-Hoon; Lü, Junxuan

    2011-01-01

    Androgen receptor (AR) signaling is crucial for the genesis and progression of prostate cancer (PCa). We compared the growth responses of AR(+) LNCaP and LNCaP C4-2 vs. AR(−) DU145 and PC-3 PCa cell lines to galbanic acid (GBA) isolated from the resin of medicinal herb Ferula assafoetida and assessed their connection to AR signaling and cell cycle regulatory pathways. Our results showed that GBA preferentially suppressed AR(+) PCa cell growth than AR(−) PCa cells. GBA induced a caspase-mediated apoptosis that was attenuated by a general caspase inhibitor. Subapoptotic GBA down-regulated AR protein in LNCaP cells primarily through promoting its proteasomal degradation, and inhibited AR-dependent transcription without affecting AR nuclear translocation. Whereas docking simulations predicted binding of GBA to the AR ligand binding domain with similarities and differences with the AR antagonist drug bicalutamide, LNCaP cell culture assays did not detect agonist activity of GBA. GBA and bicalutamide exerted greater than additive inhibitory effect on cell growth when used together. Subapoptotic GBA induced G1 arrest associated with an inhibition of cyclin/CDK4/6 pathway, especially cyclin D1 without the causal involvement of CDK inhibitory proteins P21Cip1 and P27Kip1. In summary, the novelty of GBA as an anti-AR compound resides in the distinction between GBA and bicalutamide with respect to AR protein turnover and a lack of agonist effect. Our observations of anti-AR and cell cycle arrest actions plus the anti-angiogenesis effect reported elsewhere suggest GBA as a multi-targeting drug candidate for the prevention and therapy of PCa. PMID:21328348

  17. PKCeta enhances cell cycle progression, the expression of G1 cyclins and p21 in MCF-7 cells.

    PubMed

    Fima, E; Shtutman, M; Libros, P; Missel, A; Shahaf, G; Kahana, G; Livneh, E

    2001-10-11

    Protein kinase C encodes a family of enzymes implicated in cellular differentiation, growth control and tumor promotion. However, not much is known with respect to the molecular mechanisms that link protein kinase C to cell cycle control. Here we report that the expression of PKCeta in MCF-7 cells, under the control of a tetracycline-responsive inducible promoter, enhanced cell growth and affected the cell cycle at several points. The induced expression of another PKC isoform, PKCdelta, in MCF-7 cells had opposite effects and inhibited their growth. PKCeta expression activated cellular pathways in these cells that resulted in the increased expression of the G1 phase cyclins, cyclin D and cyclin E. Expression of the cyclin-dependent kinase inhibitor p21(WAF1) was also specifically elevated in PKCeta expressing cells, but its overall effects were not inhibitory. Although, the protein levels of the cyclin-dependent kinase inhibitor p27(KIP1) were not altered by the induced expression of PKCeta, the cyclin E associated Cdk2 kinase activity was in correlation with the p27(KIP1) bound to the cyclin E complex and not by p21(WAF1) binding. PKCeta expression enhanced the removal of p27(KIP1) from this complex, and its re-association with the cyclin D/Cdk4 complex. Reduced binding of p27(KIP1) to the cyclin D/Cdk4 complex at early time points of the cell cycle also enhanced the activity of this complex, while at later time points the decrease in bound p21(WAF1) correlated with its increased activity in PKCeta-expressing cells. Thus, PKCeta induces altered expression of several cell cycle functions, which may contribute to its ability to affect cell growth.

  18. Design and Validation of a Novel Generic Platform for the Production of Tetravalent IgG1-like Bispecific Antibodies.

    PubMed

    Golay, Josée; Choblet, Sylvie; Iwaszkiewicz, Justyna; Cérutti, Pierre; Ozil, Annick; Loisel, Séverine; Pugnière, Martine; Ubiali, Greta; Zoete, Vincent; Michielin, Olivier; Berthou, Christian; Kadouche, Jean; Mach, Jean-Pierre; Duonor-Cérutti, Martine

    2016-04-01

    We have designed and validated a novel generic platform for production of tetravalent IgG1-like chimeric bispecific Abs. The VH-CH1-hinge domains of mAb2 are fused through a peptidic linker to the N terminus of mAb1 H chain, and paired mutations at the CH1-CL interface mAb1 are introduced that force the correct pairing of the two different free L chains. Two different sets of these CH1-CL interface mutations, called CR3 and MUT4, were designed and tested, and prototypic bispecific Abs directed against CD5 and HLA-DR were produced (CD5xDR). Two different hinge sequences between mAb1 and mAb2 were also tested in the CD5xDR-CR3 or -MUT4 background, leading to bispecific Ab (BsAbs) with a more rigid or flexible structure. All four Abs produced bound with good specificity and affinity to CD5 and HLA-DR present either on the same target or on different cells. Indeed, the BsAbs were able to efficiently redirect killing of HLA-DR(+) leukemic cells by human CD5(+) cytokine-induced killer T cells. Finally, all BsAbs had a functional Fc, as shown by their capacity to activate human complement and NK cells and to mediate phagocytosis. CD5xDR-CR3 was chosen as the best format because it had overall the highest functional activity and was very stable in vitro in both neutral buffer and in serum. In vivo, CD5xDR-CR3 was shown to have significant therapeutic activity in a xenograft model of human leukemia. PMID:26921308

  19. A haploid genetic screen identifies the G1/S regulatory machinery as a determinant of Wee1 inhibitor sensitivity.

    PubMed

    Heijink, Anne Margriet; Blomen, Vincent A; Bisteau, Xavier; Degener, Fabian; Matsushita, Felipe Yu; Kaldis, Philipp; Foijer, Floris; van Vugt, Marcel A T M

    2015-12-01

    The Wee1 cell cycle checkpoint kinase prevents premature mitotic entry by inhibiting cyclin-dependent kinases. Chemical inhibitors of Wee1 are currently being tested clinically as targeted anticancer drugs. Wee1 inhibition is thought to be preferentially cytotoxic in p53-defective cancer cells. However, TP53 mutant cancers do not respond consistently to Wee1 inhibitor treatment, indicating the existence of genetic determinants of Wee1 inhibitor sensitivity other than TP53 status. To optimally facilitate patient selection for Wee1 inhibition and uncover potential resistance mechanisms, identification of these currently unknown genes is necessary. The aim of this study was therefore to identify gene mutations that determine Wee1 inhibitor sensitivity. We performed a genome-wide unbiased functional genetic screen in TP53 mutant near-haploid KBM-7 cells using gene-trap insertional mutagenesis. Insertion site mapping of cells that survived long-term Wee1 inhibition revealed enrichment of G1/S regulatory genes, including SKP2, CUL1, and CDK2. Stable depletion of SKP2, CUL1, or CDK2 or chemical Cdk2 inhibition rescued the γ-H2AX induction and abrogation of G2 phase as induced by Wee1 inhibition in breast and ovarian cancer cell lines. Remarkably, live cell imaging showed that depletion of SKP2, CUL1, or CDK2 did not rescue the Wee1 inhibition-induced karyokinesis and cytokinesis defects. These data indicate that the activity of the DNA replication machinery, beyond TP53 mutation status, determines Wee1 inhibitor sensitivity, and could serve as a selection criterion for Wee1-inhibitor eligible patients. Conversely, loss of the identified S-phase genes could serve as a mechanism of acquired resistance, which goes along with development of severe genomic instability. PMID:26598692

  20. Kinetic partitioning during de novo septin filament assembly creates a critical G1 "window of opportunity" for mutant septin function.

    PubMed

    Schaefer, Rachel M; Heasley, Lydia R; Odde, David J; McMurray, Michael A

    2016-09-16

    Septin proteins form highly conserved cytoskeletal filaments composed of hetero-oligomers with strict subunit stoichiometry. Mutations within one hetero-oligomerization interface (the "G" interface) bias the mutant septin toward conformations that are incompatible with filament assembly, causing disease in humans and, in budding yeast cells, temperature-sensitive defects in cytokinesis. We previously found that, when the amount of other hetero-oligomerization partners is limiting, wild-type and G interface-mutant alleles of a given yeast septin "compete" along parallel but distinct folding pathways for occupancy of a limited number of positions within septin hetero-octamers. Here, we synthesize a mathematical model that outlines the requirements for this phenomenon: if a wild-type septin traverses a folding pathway that includes a single rate-limiting folding step, the acquisition by a mutant septin of additional slow folding steps creates an initially large disparity between wild-type and mutant in the cellular concentrations of oligomerization-competent monomers. When the 2 alleles are co-expressed, this kinetic disparity results in mutant exclusion from hetero-oligomers, even when the folded mutant monomer is oligomerization-competent. To test this model experimentally, we first visualize the kinetic delay in mutant oligomerization in living cells, and then narrow or widen the "window of opportunity" for mutant septin oligomerization by altering the length of the G1 phase of the yeast cell cycle, and observe the predicted exacerbation or suppression, respectively, of mutant cellular phenotypes. These findings reveal a fundamental kinetic principle governing in vivo assembly of multiprotein complexes, independent of the ability of the subunits to associate with each other.

  1. Torsion-Inversion Tunneling Patterns in the Ch-Stretch Vibrationally Excited States of the G{_1}{_2} Molecules

    NASA Astrophysics Data System (ADS)

    Dawadi, Mahesh B.; Bhatta, Ram S.; Perry, David S.

    2013-06-01

    Torsion-inversion tunneling models have been developed for CH-stretch vibrationally excited states in G{_1}{_2} molecules, including 2-methylmalonaldehyde (2-MMA), 5-methyltropolone (5-MT), and methylamine. These models are extensions of the group theoretical approach of Hougen and the internal coordinate model of Wang and Perry in which the inversion motion is included in addition to the torsion and the small-amplitude (e.g., CH stretch) vibrations. The present models incorporate torsion-inversion tunneling parameters {_2}{_V} and {_3}{_V}, respectively and a number of low-order terms couplings to the CH-stretch vibrations. Of the three methyl CH stretch vibrations, Model I includes only the two asymmetric stretches that correlate to the E-type degenerate CH stretch in a symmetric rotor; Model II includes all three. The models yield the torsion-inversion tunneling patterns of the four symmetry species, A, B, E{_1} and E{_2}, in the CH-stretch excited states. The principal results are as follows. (i) Both models and each of the coupling terms considered yield the same tunneling patterns, which are different in the asymmetric CH stretch excited states as compared to those in the ground state. (ii) In Model I, the magnitude of the tunneling splittings in the two asymmetric CH stretch excited states is exactly half of that in the ground state. (iii) In Model II, the relative magnitude of these splittings depends on the ratio \\vertμ\\vert/(\\vert{_2}{_V}\\vert+\\vert{_3}{_V}\\vert) where μ is the torsion-inversion-vibration coupling parameter. This ratio varies from 3 to 308 across the series methanol, methylamine, 2-methylmalonaldehyde and 5-methyltropolone, with a consequent variation in the magnitude of the tunneling splittings. J. T. HougenJ. Mol. Spectrosc. {207}, 60, (2001). X. Wang and D. S. PerryJ. Chem. Phys. {109}, 10795, (1998).

  2. PKCeta enhances cell cycle progression, the expression of G1 cyclins and p21 in MCF-7 cells.

    PubMed

    Fima, E; Shtutman, M; Libros, P; Missel, A; Shahaf, G; Kahana, G; Livneh, E

    2001-10-11

    Protein kinase C encodes a family of enzymes implicated in cellular differentiation, growth control and tumor promotion. However, not much is known with respect to the molecular mechanisms that link protein kinase C to cell cycle control. Here we report that the expression of PKCeta in MCF-7 cells, under the control of a tetracycline-responsive inducible promoter, enhanced cell growth and affected the cell cycle at several points. The induced expression of another PKC isoform, PKCdelta, in MCF-7 cells had opposite effects and inhibited their growth. PKCeta expression activated cellular pathways in these cells that resulted in the increased expression of the G1 phase cyclins, cyclin D and cyclin E. Expression of the cyclin-dependent kinase inhibitor p21(WAF1) was also specifically elevated in PKCeta expressing cells, but its overall effects were not inhibitory. Although, the protein levels of the cyclin-dependent kinase inhibitor p27(KIP1) were not altered by the induced expression of PKCeta, the cyclin E associated Cdk2 kinase activity was in correlation with the p27(KIP1) bound to the cyclin E complex and not by p21(WAF1) binding. PKCeta expression enhanced the removal of p27(KIP1) from this complex, and its re-association with the cyclin D/Cdk4 complex. Reduced binding of p27(KIP1) to the cyclin D/Cdk4 complex at early time points of the cell cycle also enhanced the activity of this complex, while at later time points the decrease in bound p21(WAF1) correlated with its increased activity in PKCeta-expressing cells. Thus, PKCeta induces altered expression of several cell cycle functions, which may contribute to its ability to affect cell growth. PMID:11709714

  3. UVB-induced cell death signaling is associated with G1-S progression and transcription inhibition in primary human fibroblasts.

    PubMed

    Ortolan, Tatiana Grohmann; Menck, Carlos Frederico M

    2013-01-01

    DNA damage induced by ultraviolet (UV) radiation can be removed by nucleotide excision repair through two sub-pathways, one general (GGR) and the other specific for transcribed DNA (TCR), and the processing of unrepaired lesions trigger signals that may lead to cell death. These signals involve the tumor suppressor p53 protein, a central regulator of cell responses to DNA damage, and the E3 ubiquitin ligase Mdm2, that forms a feedback regulatory loop with p53. The involvement of cell cycle and transcription on the signaling to apoptosis was investigated in UVB-irradiated synchronized, DNA repair proficient, CS-B (TCR-deficient) and XP-C (GGR-deficient) primary human fibroblasts. Cells were irradiated in the G1 phase of the cell cycle, with two doses with equivalent levels of apoptosis (low and high), defined for each cell line. In the three cell lines, the low doses of UVB caused only a transient delay in progression to the S phase, whereas the high doses induced permanent cell cycle arrest. However, while accumulation of Mdm2 correlated well with the recovery from transcription inhibition at the low doses for normal and CS-B fibroblasts, for XP-C cells this protein was shown to be accumulated even at UVB doses that induced high levels of apoptosis. Thus, UVB-induced accumulation of Mdm2 is critical for counteracting p53 activation and apoptosis avoidance, but its effect is limited due to transcription inhibition. However, in the case of XP-C cells, an excess of unrepaired DNA damage would be sufficient to block S phase progression, which would signal to apoptosis, independent of Mdm2 accumulation. The data clearly discriminate DNA damage signals that lead to cell death, depending on the presence of UVB-induced DNA damage in replicating or transcribing regions. PMID:24155908

  4. Determination of aflatoxins B1, B2, G1, and G2 in olive oil, peanut oil, and sesame oil.

    PubMed

    Bao, Lei; Trucksess, Mary W; White, Kevin D

    2010-01-01

    Edible oils are consumed directly, and used as ingredients in food, soaps, and skin products. However, oils such as olive oil, peanut oil, and sesame oil could be contaminated with aflatoxins, which are detrimental to human and animal health. A method using immunoaffinity column cleanup with RPLC separation and fluorescence detection (FLD) for determination of aflatoxins (AF) B1, B2, G1, and G2 in olive oil, peanut oil, and sesame oil was developed and validated. Test samples were extracted with methanol-water (55 + 45, v/v). After shaking and centrifuging, the lower layer was filtered, diluted with water, and filtered through glass microfiber filter paper. The filtrate was then passed through an immunoaffinity column, and the toxins were eluted with methanol. The toxins were then subjected to RPLC/FLD analysis after postcolumn UV photochemical derivatization. The accuracy and repeatability characteristics of the method were determined. Recoveries of AFB1 spiked at levels from 1.0 to 10.0 microg/kg in olive oil, peanut oil, and sesame oil ranged from 82.9 to 98.6%. RSDs ranged from 0.6 to 8.9%. HorRat values were < 0.2 for all of the matrixes tested. Recoveries of AF spiked at levels from 2.0 to 20.0 microg/kg ranged from 87.7 to 102.2%. RSDs ranged from 1.3 to 12.6%. HorRat values were < 0.4 for all of the matrixes tested. LC/MS/MS with multiple-reaction monitoring was used to confirm the identities of aflatoxins in a naturally contaminated peanut oil.

  5. Dendrimer-Based Responsive MRI Contrast Agents (G1-G4) for Biosensor Imaging of Redundant Deviation in Shifts (BIRDS).

    PubMed

    Huang, Yuegao; Coman, Daniel; Hyder, Fahmeed; Ali, Meser M

    2015-12-16

    Biosensor imaging of redundant deviation in shifts (BIRDS) is a molecular imaging platform for magnetic resonance that utilizes unique properties of low molecular weight paramagnetic monomers by detecting hyperfine-shifted nonexchangeable protons and transforming the chemical shift information to reflect its microenvironment (e.g., via temperature, pH, etc.). To optimize translational biosensing potential of BIRDS we examined if this detection scheme observed with monomers can be extended onto dendrimers, which are versatile and biocompatible macromolecules with modifiable surface for molecular imaging and drug delivery. Here we report on feasibility of paramagnetic dendrimers for BIRDS. The results show that BIRDS is resilient with paramagnetic dendrimers up to the fourth generation (i.e., G1-G4), where the model dendrimer and chelate were based on poly(amido amine) (PAMAM) and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA(4-)) complexed with thulium ion (Tm(3+)). Temperature sensitivities of two prominent signals of Gn-PAMAM-(TmDOTA(-))x (where n = 1-4, x = 6-39) were comparable to that of prominent signals in TmDOTA(-). Transverse relaxation times of the coalesced nonexchangeable protons on Gn-PAMAM-(TmDOTA(-))x were relatively short to provide signal-to-noise ratio that was comparable to or better than that of TmDOTA(-). A fluorescent dye, rhodamine, was conjugated to a G2-PAMAM-(TmDOTA)12 to create a dual-modality nanosized contrast agent. BIRDS properties of the dendrimer were unaltered with rhodamine conjugation. Purposely designed paramagnetic dendrimers for BIRDS in conjunction with novel macromolecular surface modification for functional ligands/drugs could potentially be used for biologically compatible theranostic sensors. PMID:26497087

  6. Array comparative genomic hybridization reveals frequent alterations of G1/S checkpoint genes in undifferentiated pleomorphic sarcoma of bone.

    PubMed

    Niini, Tarja; Lahti, Leo; Michelacci, Francesca; Ninomiya, Shinsuke; Hattinger, Claudia Maria; Guled, Mohamed; Böhling, Tom; Picci, Piero; Serra, Massimo; Knuutila, Sakari

    2011-05-01

    Undifferentiated pleomorphic sarcoma of bone (UPSb) is a rare tumor often difficult to differentiate from fibrosarcoma of bone (FSb), diagnostically. We applied array comparative genomic hybridization (array CGH) to screen for genes with potential importance in the tumor and compared the results with alterations seen in FSb. Twenty-two fresh frozen tissue specimens from 20 patients (18 primary tumors and 4 local recurrences) with UPSb were studied. DNA was isolated and hybridized onto Agilent 244K CGH oligoarrays. The hybridization data were analyzed using Agilent DNA Analytics Software. The number of changes ranged from 2 to 168 (average = 66). Losses were most frequently seen at 8p, 9p, 10, 13q, and 18q, and gains at 4q, 5p, 6p, 7p, 8q, 12p, 14q, 17q, 19p, 20q, 22q, and X. Homozygous deletions of CDKN2A, RB1, TP53, and ING1 were seen in 8/20, 7/20, 3/20, and 2/20 cases, respectively. Hypermethylation of both p16(INK4a) and p14(ARF) was found in two cases with loss at CDKN2A. Inactivation either of CDKN2A, RB1, or TP53 was detected in 18/20 cases. One case showed high level gains of CDK4 and MDM2. Frequent gains were seen at MYC, PDGFRA, KIT, and KDR. Immunohistochemical positivity of KIT, PDGFRA, KDR, and PDGFRB was found in 8/14, 5/14, 4/14, and 4/14 cases, respectively. The regions most significantly discriminating between UPSb and FSb included RB1 and MYC. No homozygous deletions of RB1 were found in FSb. In conclusion, our analysis showed the disruption of G1/S checkpoint regulation to be crucial for the oncogenesis of UPSb.

  7. Physiological characterization of Saccharomyces cerevisiae mutants supersensitive to G1 arrest by a factor and. cap alpha. factor pheromones

    SciTech Connect

    Chan, R.K.; Otte, C.A.

    1982-01-01

    Saccharomyces cerevisiae MATa cells carrying mutations in either sst1 or sst2 are supersensitive to the G1 arrest induced by ..cap alpha.. factor pheromone. When sst1 mutants were mixed with normal SST/sup +/ cells, the entire population recovered together from ..cap alpha.. factor arrest, suggesting that SST/sup +/ cells helped sst1 mutants to recover. Complementation tests and linkage analysis showed that sst1 and bar1, a mutation which eliminates the ability of MATa cells to act as a ''barrier'' to the diffusion of ..cap alpha.. factor, were lesions in the same genes. These findings suggest that sst1 mutants are defective in recovery from ..cap alpha.. factor arrest because they are unable to degrade the pheromone. In contrast, recovery of sst2 mutants was not potentiated by the presence of SST/sup +/ cells in mixing experiments. When either normal MATa cells or mutant cells carrying defects in sst1 or sst2 were exposed to ..cap alpha.. factor for 1 h and then washed free of the pheromone, the sst2 cells subsequently remained arrested in the absence of ..cap alpha.. factor for a much longer time than SST/sup +/ or sst1 cells. These observations suggest that the defect in sst2 mutants is intrinsic to the cell and is involved in the mechanism of ..cap alpha.. factor action at some step after the initial interaction of the pheromone with the cell. The presence of an sst2 mutation appears to cause a growth debility, since repeated serial subculture of haploid sst2-1 strains led to the accumulation of faster-growing revertants that were pheromone resistant and were mating defective (''sterile'').

  8. Dendrimer-Based Responsive MRI Contrast Agents (G1-G4) for Biosensor Imaging of Redundant Deviation in Shifts (BIRDS)

    PubMed Central

    Huang, Yuegao; Coman, Daniel; Hyder, Fahmeed; Ali, Meser M.

    2016-01-01

    Biosensor imaging of redundant deviation in shifts (BIRDS) is a molecular imaging platform for magnetic resonance that utilizes unique properties of low molecular weight paramagnetic monomers by detecting hyperfine-shifted nonexchangeable protons and transforming the chemical shift information to reflect its microenvironment (e.g., via temperature, pH, etc.). To optimize translational biosensing potential of BIRDS we examined if this detection scheme observed with monomers can be extended onto dendrimers, which are versatile and biocompatible macromolecules with modifiable surface for molecular imaging and drug delivery. Here we report on feasibility of paramagnetic dendrimers for BIRDS. The results show that BIRDS is resilient with paramagnetic dendrimers up to the fourth generation (i.e., G1-G4), where the model dendrimer and chelate were based on poly(amido amine) (PAMAM) and 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA4−) complexed with thulium ion (Tm3+). Temperature sensitivities of two prominent signals of Gn-PAMAM-(TmDOTA−)x (where n = 1–4, x = 6–39) were comparable to that of prominent signals in TmDOTA−. Transverse relaxation times of the coalesced nonexchangeable protons on Gn-PAMAM-(TmDOTA−)x were relatively short to provide signal-to-noise ratio that was comparable to or better than that of TmDOTA−. A fluorescent dye, rhodamine, was conjugated to a G2-PAMAM-(TmDOTA)12 to create a dual-modality nanosized contrast agent. BIRDS properties of the dendrimer were unaltered with rhodamine conjugation. Purposely designed paramagnetic dendrimers for BIRDS in conjunction with novel macromolecular surface modification for functional ligands/drugs could potentially be used for biologically compatible theranostic sensors. PMID:26497087

  9. Lipolytic inhibitor G0/G1 switch gene 2 inhibits reactive oxygen species production and apoptosis in endothelial cells.

    PubMed

    Wang, Yinfang; Zhang, Yahui; Zhu, Yichun; Zhang, Peng

    2015-03-15

    G0/G1 switch gene 2 (G0S2), a novel target gene of peroxisome proliferator-activated receptor, is highly expressed in fat tissues. G0S2 acts as proapoptotic factor toward human cancer cells. Endothelial cell (EC) apoptosis may be an initiating event in the development of atherosclerosis. However, the expression and function of G0S2 in vascular ECs remain unknown. Here, we reported for the first time that G0S2 is expressed in arterial ECs. Ectopic expression of G0S2 increased neutral lipid accumulation in cultured ECs. However, G0S2 prevented ECs from serum-free starvation stress- and hydrogen peroxide (H2O2)-induced apoptosis. G0S2 blocked the H2O2-induced dissipation of mitochondrial membrane potential. G0S2 decreased the release of cytochrome c from mitochondria into the cytosol, followed by activation of caspase-9 and caspase-3. The anti-apoptotic effect of G0S2 was Bcl-2 and adipose triglyceride lipase independent. In contrast, gene silence of G0S2 increased serum-free starvation stress-induced EC apoptosis and decreased the formation of capillary-like structures. We further found that G0S2 couples with the F0F1-ATP synthase in ECs. Levels of ATP were elevated, whereas reactive oxygen species levels were reduced in G0S2-expressing ECs. G0S2 can inhibit endothelial denudation secondary to H2O2-induced injury to ECs in vivo. These results indicate that G0S2 acts as a prosurvival molecule in ECs. Taken together, our results indicate that G0S2 has a protective function in ECs and may be a potential target for the treatment of cardiovascular diseases associated with reactive oxygen species-induced EC injury, such as atherosclerosis and restenosis. PMID:25588877

  10. A haploid genetic screen identifies the G1/S regulatory machinery as a determinant of Wee1 inhibitor sensitivity

    PubMed Central

    Heijink, Anne Margriet; Blomen, Vincent A.; Bisteau, Xavier; Degener, Fabian; Matsushita, Felipe Yu; Foijer, Floris; van Vugt, Marcel A. T. M.

    2015-01-01

    The Wee1 cell cycle checkpoint kinase prevents premature mitotic entry by inhibiting cyclin-dependent kinases. Chemical inhibitors of Wee1 are currently being tested clinically as targeted anticancer drugs. Wee1 inhibition is thought to be preferentially cytotoxic in p53-defective cancer cells. However, TP53 mutant cancers do not respond consistently to Wee1 inhibitor treatment, indicating the existence of genetic determinants of Wee1 inhibitor sensitivity other than TP53 status. To optimally facilitate patient selection for Wee1 inhibition and uncover potential resistance mechanisms, identification of these currently unknown genes is necessary. The aim of this study was therefore to identify gene mutations that determine Wee1 inhibitor sensitivity. We performed a genome-wide unbiased functional genetic screen in TP53 mutant near-haploid KBM-7 cells using gene-trap insertional mutagenesis. Insertion site mapping of cells that survived long-term Wee1 inhibition revealed enrichment of G1/S regulatory genes, including SKP2, CUL1, and CDK2. Stable depletion of SKP2, CUL1, or CDK2 or chemical Cdk2 inhibition rescued the γ-H2AX induction and abrogation of G2 phase as induced by Wee1 inhibition in breast and ovarian cancer cell lines. Remarkably, live cell imaging showed that depletion of SKP2, CUL1, or CDK2 did not rescue the Wee1 inhibition-induced karyokinesis and cytokinesis defects. These data indicate that the activity of the DNA replication machinery, beyond TP53 mutation status, determines Wee1 inhibitor sensitivity, and could serve as a selection criterion for Wee1-inhibitor eligible patients. Conversely, loss of the identified S-phase genes could serve as a mechanism of acquired resistance, which goes along with development of severe genomic instability. PMID:26598692

  11. Quercetin reduces cyclin D1 activity and induces G1 phase arrest in HepG2 cells

    PubMed Central

    ZHOU, JIN; LI, LU; FANG, LI; XIE, HUA; YAO, WENXIU; ZHOU, XIANG; XIONG, ZHUJUAN; WANG, LI; LI, ZHIXI; LUO, FENG

    2016-01-01

    Quercetin is able to inhibit proliferation of malignant tumor cells; however, the exact mechanism involved in this biological process remains unclear. The current study utilized a quantitative proteomic analysis to explore the antitumor mechanisms of quercetin. The leucine of HepG2 cells treated with quercetin was labeled as d3 by stable isotope labeling by amino acids in cell culture (SILAC). The isotope peaks of control HepG2 cells were compared with the d3-labeled HepG2 cells by mass spectrometry (MS) to identify significantly altered proteins. Reverse transcription-polymerase chain reaction (RT-PCR) and western blot analyses were subsequently employed to verify the results of the MS analysis. A flow cytometry assay was designed to observe the influence of various quercetin treatment concentrations on the cell cycle distribution of HepG2 cells. The results indicated that quercetin is able to substantially inhibit proliferation of HepG2 cells and induce an obvious morphological alteration of cells. According to the MS results, the 70 credibly-changed proteins that were identified may play important roles in multiple cellular processes, including protein synthesis, signaling, cytoskeletal processes and metabolism. Among these functional proteins, the expression of cyclin D1 (CCND1) was found to be significantly decreased. RT-PCR and western blot analyses verified the SILAC-MS results of decreased CCND1 expression. In summary, flow cytometry revealed that quercetin is able to induce G1 phase arrest in HepG2 cells. Based on the aforementioned observations, it is suggested that quercetin exerts antitumor activity in HepG2 cells through multiple pathways, including interfering with CCND1 gene expression to disrupt the cell cycle and proliferation of HepG2 cells. In the future, we aim to explore this effect in vivo. PMID:27347174

  12. Triheptanoin for glucose transporter type I deficiency (G1D): Modulation of human ictogenesis, cerebral metabolic rate and cognitive indices by a food supplement

    PubMed Central

    Pascual, Juan M.; Liu, Peiying; Mao, Deng; Kelly, Dorothy; Hernandez, Ana; Sheng, Min; Good, Levi B.; Ma, Qian; Marin-Valencia, Isaac; Zhang, Xuchen; Park, Jason Y.; Hynan, Linda S.; Stavinoha, Peter; Roe, Charles R.; Lu, Hanzhang

    2015-01-01

    Objective G1D is commonly associated with electrographic spike-wave and - less-noticeably – with absence seizures. The G1D syndrome has long been attributed to energy (i.e., ATP-synthetic) failure, as have experimental, toxic-rodent epilepsies to impaired brain metabolism and tricarboxylic acid (TCA) cycle intermediate depletion. Indeed, a (seldom-acknowledged) function of glucose and other substrates is the generation of brain TCAs via carbon-donor reactions collectively named anaplerosis. However, TCAs are preserved in murine G1D. This renders inferences about energy failure premature and suggests a different hypothesis, also grounded on our findings, that consumption of alternate TCA precursors is stimulated, potentially detracting from other functions. Second, common ketogenic diets can ameliorate G1D seizures, but lead to a therapeutically-counterintuitive reduction in blood glucose available to the brain, and they can prove ineffective in 1/3 of cases. While developing G1D treatments, all of this motivated us to: a) uphold (rather than attenuate) the residual brain glucose flux that all G1D patients possess; and b) stimulate the TCA cycle, including anaplerosis. Therefore, we tested the medium-chain triglyceride triheptanoin, a widely-used medical food supplement that can fulfill both of these metabolic roles. The rationale is that ketone bodies derived from ketogenic diets are not anaplerotic, in contrast with triheptanoin metabolites, as we have shown in the G1D mouse brain. Design We supplemented the regular diet of a case series of G1D patients with food-grade triheptanoin. First we confirmed that, despite their frequent electroencephalographic (EEG) presence as spike-waves, most seizures are rarely visible, such that perceptions by patients or others are inadequate for treatment evaluation. Thus, we used EEG, quantitative neuropsychological, blood analytical, and MRI cerebral metabolic rate measurements as main outcomes. Setting Academic and

  13. Production, Characterization, and Biological Evaluation of Well-Defined IgG1 Fc Glycoforms as a Model System for Biosimilarity Analysis.

    PubMed

    Okbazghi, Solomon Z; More, Apurva S; White, Derek R; Duan, Shaofeng; Shah, Ishan S; Joshi, Sangeeta B; Middaugh, C Russell; Volkin, David B; Tolbert, Thomas J

    2016-02-01

    Four different well-defined IgG1 Fc glycoforms are proposed as a model system to examine important biological and physicochemical features for protein drug biosimilar analyses. The IgG1 Fc glycoforms were produced by yeast expression combined with in vitro enzymatic synthesis as a series of sequentially truncated high-mannose IgG1 Fc glycoforms with an anticipated range of biological activity and structural stability. Initial characterization with mass spectrometry, SDS-PAGE, size exclusion HPLC, and capillary isoelectric focusing confirmed that the glycoproteins are overall highly similar with the only major difference being glycosylation state. Binding to the activating Fc receptor, FcγRIIIa was used to evaluate the potential biological activity of the IgG1 Fc glycoproteins. Two complementary methods using biolayer interferometry, 1 with protein G-immobilized IgG1 Fc and the other with streptavidin-immobilized FcγRIIIa, were developed to assess FcγRIIIa affinity in kinetic binding studies. The high-mannose IgG1 Fc and Man5-IgG1 Fc glycoforms were highly similar to one another with high affinity for FcγRIIIa, whereas GlcNAc-Fc had weak affinity, and the nonglycosylated N297Q-Fc had no measurable affinity for FcγRIIIa. These 4 IgG1 Fc glycoforms were also evaluated in terms of physical and chemical stability profiles and then used as a model system to mathematically assess overall biosimilarity, as described in a series of companion articles. PMID:26869419

  14. G1-4A, a polysaccharide from Tinospora cordifolia induces peroxynitrite dependent killer dendritic cell (KDC) activity against tumor cells.

    PubMed

    Pandey, Vipul K; Amin, Prayag J; Shankar, Bhavani S

    2014-12-01

    Dendritic cells (DC) play a central role in the development of an adaptive immune response against tumor. In addition to its role in antigen presentation, DC also possesses cytotoxic activity against tumor cells. We have earlier shown phenotypic and functional maturation of bone marrow derived dendritic cells (BMDC) by G1-4A, an arabinogalactan derived from Tinospora cordifolia. In this study, we have investigated the killer phenotype of BMDC matured in the presence of G1-4A, [mBMDC (G1-4A)] on tumor cells. We have observed several fold increase in killing of tumor cells by mBMDC (G1-4A). The tumoricidal activity was not specific to syngeneic tumors cells but could kill xenogenic tumors also. Nitric oxide released by mBMDC (G1-4A) generates peroxynitrite in tumor cells and is responsible for killing of target cells. This killing was completely abrogated by inducible nitric oxide synthase (iNOS) inhibitor 1400W and NADPH oxidase inhibitor apocyanin. The killed target cells are phagocytosed by BMDC which further activate syngeneic cytotoxic T cells. These results thus show that G1-4A treated mBMDC acquire killer phenotype along with maturation which plays an important role in activation of cytotoxic T cells. PMID:25278461

  15. Validation of a confirmatory analytical method for the determination of aflatoxins B₁, B₂, G₁ and G₂ in foods and feed materials by HPLC with on-line photochemical derivatization and fluorescence detection.

    PubMed

    Muscarella, Marilena; Iammarino, Marco; Nardiello, Donatella; Magro, Sonia Lo; Palermo, Carmen; Centonze, Diego; Palermo, Domenico

    2009-10-01

    A sensitive and selective analytical method for the simultaneous separation and quantitative determination of aflatoxins B1, B2, G1 and G2 in foodstuffs and materials for feed has been validated. The method is based on high performance liquid chromatography with on-line post-column photochemical derivatization and fluorescence detection. The chromatographic separation of aflatoxins was accomplished using a C18 column eluted with an isocratic mobile phase consisting of water, methanol and acetonitrile. The sample preparation required a simple extraction of aflatoxins with MeOH/H2O (80:20, v/v) and a purification step by immunoaffinity column cleanup. The total analysis time, including sample preparation and chromatographic separation, did not exceed 40 min with a run time of 10 min. The on-line photochemical derivatization ensures better results in terms of simplicity, sensitivity and reproducibility with respect to chemical derivatization techniques, and provides an increase of the peak resolution and an extent of automation in comparison with the electrochemical ones. The procedure for the determination of aflatoxins in food samples and cereals for animal consumption was extensively validated following Regulation (EC) No. 882/2004. Detection limits in wheat bran samples of 0.08 μg kg1 for AFB1, 0.02 μg kg1 for AFB2, 0.16 μg kg1 for AFG1 and 0.04 μg kg1 for AFG2 were attained. The method allows high recovery with mean values ranging from 72 to 94% and it satisfies the necessary requirements for sensitivity, linearity, selectivity, precision and ruggedness, demonstrating the conformity of the method with provisions of Regulation (EC) No. 401/2006.

  16. Mycotoxigenic potential of fungi isolated from freshly harvested Argentinean blueberries.

    PubMed

    Munitz, Martin S; Resnik, Silvia L; Pacin, Ana; Salas, Paula M; Gonzalez, Hector H L; Montti, Maria I T; Drunday, Vanesa; Guillin, Eduardo A

    2014-11-01

    Alternaria alternata, A. tenuissima, Fusarium graminearum, F. semitectum, F. verticillioides, Aspergillus flavus, and Aspergillus section Nigri strains obtained from blueberries during the 2009 and 2010 harvest season from Entre Ríos, Argentina were analyzed to determine their mycotoxigenic potential. Taxonomy status at the specific level was determined both on morphological and molecular grounds. Alternariol (AOH), alternariol monomethyl ether (AME), aflatoxins (AFs), zearalenone (ZEA), fumonisins (FBs), and ochratoxin A (OTA) were analyzed by HPLC and the trichotecenes deoxynivalenol (DON), nivalenol (NIV), HT-2 toxin (HT-2), T-2 toxin (T-2), fusarenone X (FUS-X), 3-acetyl-deoxynivalenol (3-AcDON), and 15-acetyl-deoxynivalenol (15-AcDON) by GC. Twenty-five out of forty two strains were able to produce some of the mycotoxins analyzed. Fifteen strains of Aspergillus section Nigri were capable of producing Fumonisin B1 (FB1); two of them also produced Fumonisin B2 (FB2) and one Fumonisin B3 (FB3). One of the F. graminearum isolated produced ZEA, HT-2, and T-2 and the other one was capable of producing ZEA and DON. Two A. alternata isolates produced AOH and AME. Four A. tenuissima were capable of producing AOH and three of them produced AME as well. One Aspergillu flavus strain produced aflatoxin B1 (AFB1), aflatoxin B2 (AFB2), and aflatoxin G1 (AFG1). To our knowledge, this is the first report showing mycotoxigenic capacity of fungal species isolated from blueberries that include other fungi than Alternaria spp.

  17. Tideglusib induces apoptosis in human neuroblastoma IMR32 cells, provoking sub-G0/G1 accumulation and ROS generation.

    PubMed

    Mathuram, Theodore Lemuel; Ravikumar, Vilwanathan; Reece, Lisa M; Karthik, Selvaraju; Sasikumar, Changam Sheela; Cherian, Kotturathu Mammen

    2016-09-01

    Neuroblastoma is the most common tumor amongst children amounting to nearly 15% of cancer deaths. This cancer is peculiar in its characteristics, exhibiting differentiation, maturation and metastatic transformation leading to poor prognosis and low survival rates among children. Chemotherapy, though toxic to normal cells, has shown to improve the survival of the patient with emphasis given more towards targeting angiogenesis. Recently, Tideglusib was designed as an 'Orphan Drug' to target the neurodegenerative Alzheimer's disease and gained significant momentum in its function during clinical trials. Duffy et al. recently reported a reduction in cell viability of human IMR32 neuroblastoma cells when treated with Tideglusib at varying concentrations. We investigated the effects of Tideglusib, at various concentrations, compared to Lithium chloride at various concentrations, on IMR32 cells. Lithium, a known GSK-3 inhibitor, was used as a standard to compare the efficiency of Tideglusib in a dose-dependent manner. Cell viability was assessed by MTT assay. The stages of apoptosis were evaluated by AO/EB staining and nuclear damage was determined by Hoechst 33258 staining. Reactive oxygen species (ROS) and mitochondrial membrane potential (ΔΨm) were assessed by DCFDA dye and Rhodamine-123 dye, respectively. Tideglusib reported a significant dose-dependent increase in pro-apoptotic proteins (PARP, Caspase-9, Caspase-7, Caspase-3) and tumor-related genes (FasL, TNF-α, Cox-2, IL-8, Caspase-3). Anti-GSK3 β, pGSK3 β, Bcl-2, Akt-1, p-Akt1 protein levels were observed with cells exposed to Tideglusib and Lithium chloride. No significant dose-dependent changes were observed for the mRNA expression of collagenase MMP-2, the tumor suppressor p53, or the cell cycle protein p21. Our study also reports Tideglusib reducing colony formation and increasing the level of sub-G0/G1 population in IMR32 cells. Our investigations report the significance of Tideglusib as a promising

  18. Aerosol concentration and size distribution measured below, in, and above cloud from the DOE G-1 during VOCALS-REx

    SciTech Connect

    Kleinman L. I.; Daum, P. H.; Lee, Y.-N.; Lewis, E. R.; Sedlacek III, A. J.; Senum, G. I.; Springston, S. R.; Wang, J.; Hubbe, J.; Jayne, J.; Min, Q.; Yum, S. S.; Allen, G.

    2012-01-04

    During the VOCALS Regional Experiment, the DOE G-1 aircraft was used to sample a varying aerosol environment pertinent to properties of stratocumulus clouds over a longitude band extending 800 km west from the Chilean coast at Arica. Trace gas and aerosol measurements are presented as a function of longitude, altitude, and dew point in this study. Spatial distributions are consistent with an upper atmospheric source for O{sub 3} and South American coastal sources for marine boundary layer (MBL) CO and aerosol, most of which is acidic sulfate. Pollutant layers in the free troposphere (FT) can be a result of emissions to the north in Peru or long range transport from the west. At a given altitude in the FT (up to 3 km), dew point varies by 40 C with dry air descending from the upper atmospheric and moist air having a boundary layer (BL) contribution. Ascent of BL air to a cold high altitude results in the condensation and precipitation removal of all but a few percent of BL water along with aerosol that served as CCN. Thus, aerosol volume decreases with dew point in the FT. Aerosol size spectra have a bimodal structure in the MBL and an intermediate diameter unimodal distribution in the FT. Comparing cloud droplet number concentration (CDNC) and pre-cloud aerosol (D{sub p} > 100 nm) gives a linear relation up to a number concentration of {approx}150 cm{sup -3}, followed by a less than proportional increase in CDNC at higher aerosol number concentration. A number balance between below cloud aerosol and cloud droplets indicates that {approx}25 % of aerosol with D{sub p} > 100 nm are interstitial (not activated). A direct comparison of pre-cloud and in-cloud aerosol yields a higher estimate. Artifacts in the measurement of interstitial aerosol due to droplet shatter and evaporation are discussed. Within each of 102 constant altitude cloud transects, CDNC and interstitial aerosol were anti-correlated. An examination of one cloud as a case study shows that the

  19. Aerosol concentration and size distribution measured below, in, and above cloud from the DOE G-1 during VOCALS-REx

    SciTech Connect

    Kleinman, L.I.; Daum, P. H.; Lee, Y.-N.; Lewis, E. R.; Sedlacek III, A. J.; Senum, G. I.; Springston, S. R.; Wang, J.; Hubbe, J.; Jayne, J.; Min, Q.; Yum, S. S.; Allen, G.

    2011-06-21

    During the VOCALS Regional Experiment, the DOE G-1 aircraft was used to sample a varying aerosol environment pertinent to properties of stratocumulus clouds over a longitude band extending 800 km west from the Chilean coast at Arica. Trace gas and aerosol measurements are presented as a function of longitude, altitude, and dew point in this study. Spatial distributions are consistent with an upper atmospheric source for O{sub 3} and South American coastal sources for marine boundary layer (MBL) CO and aerosol, most of which is acidic sulfate in agreement with the dominant pollution source being SO{sub 2} from Cu smelters and power plants. Pollutant layers in the free troposphere (FT) can be a result of emissions to the north in Peru or long range transport from the west. At a given altitude in the FT (up to 3 km), dew point varies by 40 C with dry air descending from the upper atmospheric and moist air having a BL contribution. Ascent of BL air to a cold high altitude results in the condensation and precipitation removal of all but a few percent of BL water along with aerosol that served as CCN. Thus, aerosol volume decreases with dew point in the FT. Aerosol size spectra have a bimodal structure in the MBL and an intermediate diameter unimodal distribution in the FT. Comparing cloud droplet number concentration (CDNC) and pre-cloud aerosol (Dp > 100 nm) gives a linear relation up to a number concentration of {approx}150 cm{sup -3}, followed by a less than proportional increase in CDNC at higher aerosol number concentration. A number balance between below cloud aerosol and cloud droplets indicates that {approx}25% of aerosol in the PCASP size range are interstitial (not activated). One hundred and two constant altitude cloud transects were identified and used to determine properties of interstitial aerosol. One transect is examined in detail as a case study. Approximately 25 to 50% of aerosol with D{sub p} > 110 nm were not activated, the difference between the two

  20. Aerosol concentration and size distribution measured below, in, and above cloud from the DOE G-1 during VOCALS-REx

    NASA Astrophysics Data System (ADS)

    Kleinman, L. I.; Daum, P. H.; Lee, Y.-N.; Lewis, E. R.; Sedlacek, A. J., III; Senum, G. I.; Springston, S. R.; Wang, J.; Hubbe, J.; Jayne, J.; Min, Q.; Yum, S. S.; Allen, G.

    2011-06-01

    During the VOCALS Regional Experiment, the DOE G-1 aircraft was used to sample a varying aerosol environment pertinent to properties of stratocumulus clouds over a longitude band extending 800 km west from the Chilean coast at Arica. Trace gas and aerosol measurements are presented as a function of longitude, altitude, and dew point in this study. Spatial distributions are consistent with an upper atmospheric source for O3 and South American coastal sources for marine boundary layer (MBL) CO and aerosol, most of which is acidic sulfate in agreement with the dominant pollution source being SO2 from Cu smelters and power plants. Pollutant layers in the free troposphere (FT) can be a result of emissions to the north in Peru or long range transport from the west. At a given altitude in the FT (up to 3 km), dew point varies by 40 °C with dry air descending from the upper atmospheric and moist air having a BL contribution. Ascent of BL air to a cold high altitude results in the condensation and precipitation removal of all but a few percent of BL water along with aerosol that served as CCN. Thus, aerosol volume decreases with dew point in the FT. Aerosol size spectra have a bimodal structure in the MBL and an intermediate diameter unimodal distribution in the FT. Comparing cloud droplet number concentration (CDNC) and pre-cloud aerosol (Dp > 100 nm) gives a linear relation up to a number concentration of ~150 cm-3, followed by a less than proportional increase in CDNC at higher aerosol number concentration. A number balance between below cloud aerosol and cloud droplets indicates that ~25 % of aerosol in the PCASP size range are interstitial (not activated). One hundred and two constant altitude cloud transects were identified and used to determine properties of interstitial aerosol. One transect is examined in detail as a case study. Approximately 25 to 50 % of aerosol with Dp > 110 nm were not activated, the difference between the two approaches possibly representing

  1. Aerosol concentration and size distribution measured below, in, and above cloud from the DOE G-1 during VOCALS-REx

    NASA Astrophysics Data System (ADS)

    Kleinman, L. I.; Daum, P. H.; Lee, Y.-N.; Lewis, E. R.; Sedlacek, A. J., III; Senum, G. I.; Springston, S. R.; Wang, J.; Hubbe, J.; Jayne, J.; Min, Q.; Yum, S. S.; Allen, G.

    2012-01-01

    During the VOCALS Regional Experiment, the DOE G-1 aircraft was used to sample a varying aerosol environment pertinent to properties of stratocumulus clouds over a longitude band extending 800 km west from the Chilean coast at Arica. Trace gas and aerosol measurements are presented as a function of longitude, altitude, and dew point in this study. Spatial distributions are consistent with an upper atmospheric source for O3 and South American coastal sources for marine boundary layer (MBL) CO and aerosol, most of which is acidic sulfate. Pollutant layers in the free troposphere (FT) can be a result of emissions to the north in Peru or long range transport from the west. At a given altitude in the FT (up to 3 km), dew point varies by 40 °C with dry air descending from the upper atmospheric and moist air having a boundary layer (BL) contribution. Ascent of BL air to a cold high altitude results in the condensation and precipitation removal of all but a few percent of BL water along with aerosol that served as CCN. Thus, aerosol volume decreases with dew point in the FT. Aerosol size spectra have a bimodal structure in the MBL and an intermediate diameter unimodal distribution in the FT. Comparing cloud droplet number concentration (CDNC) and pre-cloud aerosol (Dp>100 nm) gives a linear relation up to a number concentration of ~150 cm-3, followed by a less than proportional increase in CDNC at higher aerosol number concentration. A number balance between below cloud aerosol and cloud droplets indicates that ~25 % of aerosol with Dp>100 nm are interstitial (not activated). A direct comparison of pre-cloud and in-cloud aerosol yields a higher estimate. Artifacts in the measurement of interstitial aerosol due to droplet shatter and evaporation are discussed. Within each of 102 constant altitude cloud transects, CDNC and interstitial aerosol were anti-correlated. An examination of one cloud as a case study shows that the interstitial aerosol appears to have a background

  2. THE HARD X-RAY VIEW OF THE YOUNG SUPERNOVA REMNANT G1.9+0.3

    SciTech Connect

    Zoglauer, Andreas; Boggs, Steven E.; Craig, William W.; Krivonos, Roman A.; Reynolds, Stephen P.; An, Hongjun; Christensen, Finn E.; Fryer, Chris L.; Grefenstette, Brian W.; Harrison, Fiona A.; Madsen, Kristin K.; Miyasaka, Hiromasa; Hailey, Charles J.; Stern, Daniel; Zhang, William W.

    2015-01-10

    NuSTAR observed G1.9+0.3, the youngest known supernova remnant in the Milky Way, for 350 ks and detected emission up to ∼30 keV. The remnant's X-ray morphology does not change significantly across the energy range from 3 to 20 keV. A combined fit between NuSTAR and Chandra shows that the spectrum steepens with energy. The spectral shape can be well fitted with synchrotron emission from a power-law electron energy distribution with an exponential cutoff with no additional features. It can also be described by a purely phenomenological model such as a broken power law or a power law with an exponential cutoff, though these descriptions lack physical motivation. Using a fixed radio flux at 1 GHz of 1.17 Jy for the synchrotron model, we get a column density of N {sub H} = (7.23 ± 0.07) × 10{sup 22} cm{sup –2}, a spectral index of α = 0.633 ± 0.003, and a roll-off frequency of ν{sub rolloff} = (3.07 ± 0.18) × 10{sup 17} Hz. This can be explained by particle acceleration, to a maximum energy set by the finite remnant age, in a magnetic field of about 10 μG, for which our roll-off implies a maximum energy of about 100 TeV for both electrons and ions. Much higher magnetic-field strengths would produce an electron spectrum that was cut off by radiative losses, giving a much higher roll-off frequency that is independent of magnetic-field strength. In this case, ions could be accelerated to much higher energies. A search for {sup 44}Ti emission in the 67.9 keV line results in an upper limit of 1.5 × 10{sup –5} photons cm{sup –2} s{sup –1} assuming a line width of 4.0 keV (1 sigma)

  3. Tideglusib induces apoptosis in human neuroblastoma IMR32 cells, provoking sub-G0/G1 accumulation and ROS generation.

    PubMed

    Mathuram, Theodore Lemuel; Ravikumar, Vilwanathan; Reece, Lisa M; Karthik, Selvaraju; Sasikumar, Changam Sheela; Cherian, Kotturathu Mammen

    2016-09-01

    Neuroblastoma is the most common tumor amongst children amounting to nearly 15% of cancer deaths. This cancer is peculiar in its characteristics, exhibiting differentiation, maturation and metastatic transformation leading to poor prognosis and low survival rates among children. Chemotherapy, though toxic to normal cells, has shown to improve the survival of the patient with emphasis given more towards targeting angiogenesis. Recently, Tideglusib was designed as an 'Orphan Drug' to target the neurodegenerative Alzheimer's disease and gained significant momentum in its function during clinical trials. Duffy et al. recently reported a reduction in cell viability of human IMR32 neuroblastoma cells when treated with Tideglusib at varying concentrations. We investigated the effects of Tideglusib, at various concentrations, compared to Lithium chloride at various concentrations, on IMR32 cells. Lithium, a known GSK-3 inhibitor, was used as a standard to compare the efficiency of Tideglusib in a dose-dependent manner. Cell viability was assessed by MTT assay. The stages of apoptosis were evaluated by AO/EB staining and nuclear damage was determined by Hoechst 33258 staining. Reactive oxygen species (ROS) and mitochondrial membrane potential (ΔΨm) were assessed by DCFDA dye and Rhodamine-123 dye, respectively. Tideglusib reported a significant dose-dependent increase in pro-apoptotic proteins (PARP, Caspase-9, Caspase-7, Caspase-3) and tumor-related genes (FasL, TNF-α, Cox-2, IL-8, Caspase-3). Anti-GSK3 β, pGSK3 β, Bcl-2, Akt-1, p-Akt1 protein levels were observed with cells exposed to Tideglusib and Lithium chloride. No significant dose-dependent changes were observed for the mRNA expression of collagenase MMP-2, the tumor suppressor p53, or the cell cycle protein p21. Our study also reports Tideglusib reducing colony formation and increasing the level of sub-G0/G1 population in IMR32 cells. Our investigations report the significance of Tideglusib as a promising

  4. Effect of altered CH2-associated carbohydrate structure on the functional properties and in vivo fate of chimeric mouse-human immunoglobulin G1

    PubMed Central

    1994-01-01

    Immunoglobulin G (IgG) molecules are glycosylated in CH2 at Asn297; the N-linked carbohydrates attached there have been shown to contribute to antibody (Ab) stability and various effector functions. The carbohydrate attached to the IgG constant region is a complex biantennary structure. Alterations in the structure of oligosaccharide have been associated with human diseases such as rheumatoid arthritis and osteoarthritis. To study the effects of altered carbohydrate structure on Ab effector function, we have used gene transfection techniques to produce mouse-human chimeric IgG1 Abs in the Chinese hamster ovary (CHO) cell line Lec 1, which is incapable of processing the high-mannose intermediate through the terminal glycosylation steps. We also produced IgG1 Abs in Pro-5, the wild-type CHO cell line that is the parent of Lec 1. The Pro-5-produced Ab (IgG1-Pro-5) was similar to IgG1-My 1, a myeloma-produced IgG1 Ab of the same specificity, in its biologic properties such as serum half-life, ability to effect complement-mediated cytolysis, and affinity for Fc gamma RI. Although the Lec 1-produced Ab, IgG1-Lec 1, was properly assembled and retained antigen specificity, it was incapable of complement-mediated hemolysis and was substantially deficient in complement consumption, C1q binding, and C1 activation. IgG1-Lec 1 also showed reduced but significant affinity for Fc gamma R1 receptors. The in vivo half-life of IgG1-Lec 1 was shorter than that of either the myeloma- or Pro-5-produced counterpart, with more being cleared during the alpha-phase and with more rapid clearance during the beta-phase. Clearance of IgG1-Lec 1 could be inhibited by the administration of yeast-derived mannan. Thus the uptake of IgG1-Lec 1 appears to be accelerated by the presence of terminally mannosylated oligosaccharide. Therefore, certain Ab functions as well as the in vivo fate of the protein are dramatically affected by altered carbohydrate structure. Expression of Igs in cell lines with

  5. Whole genome analyses of G1P[8] rotavirus strains from vaccinated and non-vaccinated South African children presenting with diarrhea.

    PubMed

    Magagula, Nonkululeko B; Esona, Mathew D; Nyaga, Martin M; Stucker, Karla M; Halpin, Rebecca A; Stockwell, Timothy B; Seheri, Mapaseka L; Steele, A Duncan; Wentworth, David E; Mphahlele, M Jeffrey

    2015-01-01

    Group A rotaviruses (RVAs) are the leading cause of severe gastroenteritis and eventually death among infants and young children worldwide, and disease prevention and management through vaccination is a public health priority. In August 2009, Rotarix™ was introduced in the South African Expanded Programme on Immunisation. As a result, substantial reductions in RVA disease burden have been reported among children younger than 5 years old. Rotavirus strain surveillance post-vaccination is crucial to, inter alia, monitor and study the evolution of vaccine escape strains. Here, full-genome sequence data for the 11 gene segments from 11 South African G1P[8] rotavirus strains were generated, including 5 strains collected from non-vaccinated children during the 2004-2009 rotavirus seasons and 6 strains collected from vaccinated children during the 2010 rotavirus season. These data were analyzed to gain insights into the overall genetic makeup and evolution of South African G1P[8] rotavirus strains and to compare their genetic backbones with those of common human Wa-like RVAs from other countries, as well as with the Rotarix™ and RotaTeq™ G1P[8] vaccine components. All 11 South African G1P[8] strains revealed a complete Wa-like genotype constellation of G1-P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1. On the basis of sequence similarities, the South African G1P[8] strains (with the exception of strain RVA/Human-wt/ZAF/1262/2004/G1P[8]) were closely related to each other (96-100% identity in all gene segments). Comparison to the Rotarix™ and RotaTeq™ G1P[8] vaccine components revealed a moderate nucleotide identity of 89-96% and 93-95%, respectively. The results indicated that none of the gene segments of these 11 South African G1P[8] strains were vaccine-derived. This study illustrates that large-scale next generation sequencing will provide crucial information on the influence of the vaccination program on evolution of rotavirus strains. This is the first report to describe

  6. Whole genome analyses of G1P[8] rotavirus strains from vaccinated and non-vaccinated South African children presenting with diarrhea.

    PubMed

    Magagula, Nonkululeko B; Esona, Mathew D; Nyaga, Martin M; Stucker, Karla M; Halpin, Rebecca A; Stockwell, Timothy B; Seheri, Mapaseka L; Steele, A Duncan; Wentworth, David E; Mphahlele, M Jeffrey

    2015-01-01

    Group A rotaviruses (RVAs) are the leading cause of severe gastroenteritis and eventually death among infants and young children worldwide, and disease prevention and management through vaccination is a public health priority. In August 2009, Rotarix™ was introduced in the South African Expanded Programme on Immunisation. As a result, substantial reductions in RVA disease burden have been reported among children younger than 5 years old. Rotavirus strain surveillance post-vaccination is crucial to, inter alia, monitor and study the evolution of vaccine escape strains. Here, full-genome sequence data for the 11 gene segments from 11 South African G1P[8] rotavirus strains were generated, including 5 strains collected from non-vaccinated children during the 2004-2009 rotavirus seasons and 6 strains collected from vaccinated children during the 2010 rotavirus season. These data were analyzed to gain insights into the overall genetic makeup and evolution of South African G1P[8] rotavirus strains and to compare their genetic backbones with those of common human Wa-like RVAs from other countries, as well as with the Rotarix™ and RotaTeq™ G1P[8] vaccine components. All 11 South African G1P[8] strains revealed a complete Wa-like genotype constellation of G1-P[8]-I1-R1-C1-M1-A1-N1-T1-E1-H1. On the basis of sequence similarities, the South African G1P[8] strains (with the exception of strain RVA/Human-wt/ZAF/1262/2004/G1P[8]) were closely related to each other (96-100% identity in all gene segments). Comparison to the Rotarix™ and RotaTeq™ G1P[8] vaccine components revealed a moderate nucleotide identity of 89-96% and 93-95%, respectively. The results indicated that none of the gene segments of these 11 South African G1P[8] strains were vaccine-derived. This study illustrates that large-scale next generation sequencing will provide crucial information on the influence of the vaccination program on evolution of rotavirus strains. This is the first report to describe

  7. G1-4A, a Polysaccharide from Tinospora cordifolia Inhibits the Survival of Mycobacterium tuberculosis by Modulating Host Immune Responses in TLR4 Dependent Manner

    PubMed Central

    Gupta, Pramod Kumar; Chakraborty, Pampi; Kumar, Santosh; Singh, Prafull Kumar; Rajan, M. G. R.; Sainis, Krishna B.; Kulkarni, Savita

    2016-01-01

    Rapid emergence of drug resistance in Mycobacterium tuberculosis (MTB) is a major health concern and demands the development of novel adjunct immunotherapeutic agents capable of modulating the host immune responses in order to control the pathogen. In the present study, we sought to investigate the immunomodulatory effects of G1-4A, a polysaccharide derived from the Indian medicinal plant Tinospora cordifolia, in in-vitro and aerosol mouse models of MTB infection. G1-4A treatment of MTB infected RAW264.7 macrophages significantly induced the surface expression of MHC-II and CD-86 molecules, secretion of proinflammatory cytokines (TNF-α, IL-β, IL-6, IL-12, IFN-γ) and nitric oxide leading to reduced intracellular survival of both drug sensitive (H37Rv) as well as multi drug resistant strains (Beijing and LAM) of MTB, which was partially attributed to G1-4A induced NO production in TLR4-MyD88 dependent manner. Similarly, bacillary burden was significantly reduced in the lungs of MTB infected BALB/c mice treated with G1-4A, with simultaneous up-regulation of the expression of TNF-α, INF-γ and NOS2 in the mouse lung along with increased levels of Th1 cytokines like IFN-γ, IL-12 and decreased levels of Th2 cytokine like IL-4 in the serum. Furthermore, combination of G1-4A with Isoniazid (INH) exhibited better protection against MTB compared to that due to INH or G1-4A alone, suggesting its potential as adjunct therapy. Our results demonstrate that modulation of host immune responses by G1-4A might improve the therapeutic efficacy of existing anti-tubercular drugs and provide an attractive strategy for the development of alternative therapies to control tuberculosis. PMID:27148868

  8. G1-4A, a Polysaccharide from Tinospora cordifolia Inhibits the Survival of Mycobacterium tuberculosis by Modulating Host Immune Responses in TLR4 Dependent Manner.

    PubMed

    Gupta, Pramod Kumar; Chakraborty, Pampi; Kumar, Santosh; Singh, Prafull Kumar; Rajan, M G R; Sainis, Krishna B; Kulkarni, Savita

    2016-01-01

    Rapid emergence of drug resistance in Mycobacterium tuberculosis (MTB) is a major health concern and demands the development of novel adjunct immunotherapeutic agents capable of modulating the host immune responses in order to control the pathogen. In the present study, we sought to investigate the immunomodulatory effects of G1-4A, a polysaccharide derived from the Indian medicinal plant Tinospora cordifolia, in in-vitro and aerosol mouse models of MTB infection. G1-4A treatment of MTB infected RAW264.7 macrophages significantly induced the surface expression of MHC-II and CD-86 molecules, secretion of proinflammatory cytokines (TNF-α, IL-β, IL-6, IL-12, IFN-γ) and nitric oxide leading to reduced intracellular survival of both drug sensitive (H37Rv) as well as multi drug resistant strains (Beijing and LAM) of MTB, which was partially attributed to G1-4A induced NO production in TLR4-MyD88 dependent manner. Similarly, bacillary burden was significantly reduced in the lungs of MTB infected BALB/c mice treated with G1-4A, with simultaneous up-regulation of the expression of TNF-α, INF-γ and NOS2 in the mouse lung along with increased levels of Th1 cytokines like IFN-γ, IL-12 and decreased levels of Th2 cytokine like IL-4 in the serum. Furthermore, combination of G1-4A with Isoniazid (INH) exhibited better protection against MTB compared to that due to INH or G1-4A alone, suggesting its potential as adjunct therapy. Our results demonstrate that modulation of host immune responses by G1-4A might improve the therapeutic efficacy of existing anti-tubercular drugs and provide an attractive strategy for the development of alternative therapies to control tuberculosis. PMID:27148868

  9. Deep Phylogenetic Analysis of Haplogroup G1 Provides Estimates of SNP and STR Mutation Rates on the Human Y-Chromosome and Reveals Migrations of Iranic Speakers

    PubMed Central

    Balanovsky, Oleg; Zhabagin, Maxat; Agdzhoyan, Anastasiya; Chukhryaeva, Marina; Zaporozhchenko, Valery; Utevska, Olga; Highnam, Gareth; Sabitov, Zhaxylyk; Greenspan, Elliott; Dibirova, Khadizhat; Skhalyakho, Roza; Kuznetsova, Marina; Koshel, Sergey; Yusupov, Yuldash; Nymadawa, Pagbajabyn; Zhumadilov, Zhaxybay; Pocheshkhova, Elvira; Haber, Marc; A. Zalloua, Pierre; Yepiskoposyan, Levon; Dybo, Anna; Tyler-Smith, Chris; Balanovska, Elena

    2015-01-01

    Y-chromosomal haplogroup G1 is a minor component of the overall gene pool of South-West and Central Asia but reaches up to 80% frequency in some populations scattered within this area. We have genotyped the G1-defining marker M285 in 27 Eurasian populations (n= 5,346), analyzed 367 M285-positive samples using 17 Y-STRs, and sequenced ~11 Mb of the Y-chromosome in 20 of these samples to an average coverage of 67X. This allowed detailed phylogenetic reconstruction. We identified five branches, all with high geographical specificity: G1-L1323 in Kazakhs, the closely related G1-GG1 in Mongols, G1-GG265 in Armenians and its distant brother clade G1-GG162 in Bashkirs, and G1-GG362 in West Indians. The haplotype diversity, which decreased from West Iran to Central Asia, allows us to hypothesize that this rare haplogroup could have been carried by the expansion of Iranic speakers northwards to the Eurasian steppe and via founder effects became a predominant genetic component of some populations, including the Argyn tribe of the Kazakhs. The remarkable agreement between genetic and genealogical trees of Argyns allowed us to calibrate the molecular clock using a historical date (1405 AD) of the most recent common genealogical ancestor. The mutation rate for Y-chromosomal sequence data obtained was 0.78×10-9 per bp per year, falling within the range of published rates. The mutation rate for Y-chromosomal STRs was 0.0022 per locus per generation, very close to the so-called genealogical rate. The “clan-based” approach to estimating the mutation rate provides a third, middle way between direct farther-to-son comparisons and using archeologically known migrations, whose dates are subject to revision and of uncertain relationship to genetic events. PMID:25849548

  10. G1-4A, a Polysaccharide from Tinospora cordifolia Inhibits the Survival of Mycobacterium tuberculosis by Modulating Host Immune Responses in TLR4 Dependent Manner.

    PubMed

    Gupta, Pramod Kumar; Chakraborty, Pampi; Kumar, Santosh; Singh, Prafull Kumar; Rajan, M G R; Sainis, Krishna B; Kulkarni, Savita

    2016-01-01

    Rapid emergence of drug resistance in Mycobacterium tuberculosis (MTB) is a major health concern and demands the development of novel adjunct immunotherapeutic agents capable of modulating the host immune responses in order to control the pathogen. In the present study, we sought to investigate the immunomodulatory effects of G1-4A, a polysaccharide derived from the Indian medicinal plant Tinospora cordifolia, in in-vitro and aerosol mouse models of MTB infection. G1-4A treatment of MTB infected RAW264.7 macrophages significantly induced the surface expression of MHC-II and CD-86 molecules, secretion of proinflammatory cytokines (TNF-α, IL-β, IL-6, IL-12, IFN-γ) and nitric oxide leading to reduced intracellular survival of both drug sensitive (H37Rv) as well as multi drug resistant strains (Beijing and LAM) of MTB, which was partially attributed to G1-4A induced NO production in TLR4-MyD88 dependent manner. Similarly, bacillary burden was significantly reduced in the lungs of MTB infected BALB/c mice treated with G1-4A, with simultaneous up-regulation of the expression of TNF-α, INF-γ and NOS2 in the mouse lung along with increased levels of Th1 cytokines like IFN-γ, IL-12 and decreased levels of Th2 cytokine like IL-4 in the serum. Furthermore, combination of G1-4A with Isoniazid (INH) exhibited better protection against MTB compared to that due to INH or G1-4A alone, suggesting its potential as adjunct therapy. Our results demonstrate that modulation of host immune responses by G1-4A might improve the therapeutic efficacy of existing anti-tubercular drugs and provide an attractive strategy for the development of alternative therapies to control tuberculosis.

  11. Persistent decrease in viability as a function of X irradiation of human bladder carcinoma cells in G1 or S phase.

    PubMed

    Leonhardt, E A; Trinh, M; Forrester, H B; Dewey, W C

    1998-04-01

    A persistent decrease in viability after treatment with a variety of mutagenic agents has been observed previously, but the dependence of the decrease on the phase of the cell cycle in which the cells are treated has not been fully explored. Synchronous human bladder carcinoma cells (EJ30-15) were obtained by mitotic selection (88-96% in or near mitosis). As monitored by microscopy and pulse labeling with [3H]dThd, approximately 98% of the cells were in G1 phase when they were irradiated after 3 h of incubation, and approximately 80% were in S phase when they were irradiated after 14 h of incubation. The initial plating efficiencies demonstrated no difference in cell survival when cells were irradiated in G1 or S phase, with normalized clonogenic survival and standard error of 60+/-6% for 3 Gy and 13+/-2% for 6 Gy. However, when the cell populations were allowed to incubate and were replated 5 to 33 days later (5.5 to 36 doublings), a difference between the populations irradiated in G1 and S phase became clear. Cells that were irradiated with 6 Gy regained and maintained the high plating efficiencies (67.9+/-3.6%) of the unirradiated populations much sooner when they were irradiated in S phase compared with irradiation in G1 phase, i.e. 11 days (12 cell doublings) for S phase compared to approximately 20 days (22 cell doublings) for G1 phase. During these periods when the plating efficiencies were increasing, the populations irradiated in G1 phase were multiplying at rates lower than those for the populations irradiated in S phase. Furthermore, after 6 Gy, more giant cells and multinucleated cells were seen in the populations irradiated in G1 phase than in the populations irradiated in S phase. These results indicate that, although the clonogenic survival was the same for cells irradiated in G1 or S phase, the residual damage in progeny of the irradiated cells persisted longer (approximately 20 days compared to 11 days) when cells were irradiated in G1 phase than

  12. Involvement of miR-15a in G0/G1 Phase Cell Cycle Arrest Induced by Porcine Circovirus Type 2 Replication.

    PubMed

    Quan, Rong; Wei, Li; Zhu, Shanshan; Wang, Jing; Cao, Yongchang; Xue, Chunyi; Yan, Xu; Liu, Jue

    2016-01-01

    Many viruses exploit the host cell division cycle to favour their own growth. Here we demonstrated that porcine circovirus type 2 (PCV2), which is a major causative agent of an emerging and important swine disease complex, PCV2-associated diseases, caused G0/G1 cell cycle arrest through degradation of cyclin D1 and E followed by reduction of retinoblastoma phosphorylation in synchronized PCV2-infected cells dependent upon virus replication. This induction of G0/G1 cell cycle arrest promoted PCV2 replication as evidenced by increased viral protein expression and progeny virus production in the synchronized PCV2-infected cells. To delineate a mechanism of miRNAs in regulating PCV2-induced G0/G1 cell cycle arrest, we determined expression levels of some relevant miRNAs and found that only miR-15a but not miR-16, miR-21, and miR-34a was significantly changed in the PCV2-infected cells. We further demonstrated that upregulation of miR-15a promoted PCV2-induced G0/G1 cell cycle arrest via mediating cyclins D1 and E degradation, in which involves PCV2 growth. These results reveal that G0/G1 cell cycle arrest induced by PCV2 may provide favourable conditions for viral protein expression and progeny production and that miR-15a is implicated in PCV2-induced cell cycle control, thereby contributing to efficient viral replication. PMID:27302568

  13. Murine germinal center B cells require functional Fms-like tyrosine kinase 3 signaling for IgG1 class-switch recombination

    PubMed Central

    Svensson, Mattias N. D.; Andersson, Karin M. E.; Wasén, Caroline; Nurkkala-Karlsson, Merja; Jonsson, Ing-Marie; Brisslert, Mikael; Bemark, Mats; Bokarewa, Maria I.

    2015-01-01

    Switched antibody classes are important for efficient immune responses. Aberrant antibody production to otherwise harmless antigens may result in autoimmunity. The protein kinase fms-like tyrosine kinase 3 receptor (Flt3) has an important role during early B-cell development, but the role of Flt3 in peripheral B cells has not been assessed before. Herein we describe a previously unappreciated role for Flt3 in IgG1 class-switch recombination (CSR) and production. We show that Flt3 is reexpressed on B-cell lymphoma 6+ germinal center B cells in vivo and following LPS activation of peripheral B cells in vitro. Absence of Flt3 signaling in Flt3 ligand-deficient mice results in impaired IgG1 CSR and accumulation of IgM-secreting plasma cells. On activated B cells, Flt3 is coexpressed and functions in synergy with the common-gamma chain receptor family. B cells from Flt3 ligand-deficient mice have impaired IL-4R signaling, with reduced phosphorylation of signal transducer and activator of transcription (Stat) 6, and demonstrate a failure to initiate CSR to IgG1 with low expression of γ1 germ-line transcripts, resulting in impaired IgG1 production. Thus, functional synergy between Flt3 and IL-4R signaling is critical for Stat-mediated regulation of sterile γ1 germ-line transcripts and CSR to IgG1. PMID:26627255

  14. Involvement of miR-15a in G0/G1 Phase Cell Cycle Arrest Induced by Porcine Circovirus Type 2 Replication

    PubMed Central

    Quan, Rong; Wei, Li; Zhu, Shanshan; Wang, Jing; Cao, Yongchang; Xue, Chunyi; Yan, Xu; Liu, Jue

    2016-01-01

    Many viruses exploit the host cell division cycle to favour their own growth. Here we demonstrated that porcine circovirus type 2 (PCV2), which is a major causative agent of an emerging and important swine disease complex, PCV2-associated diseases, caused G0/G1 cell cycle arrest through degradation of cyclin D1 and E followed by reduction of retinoblastoma phosphorylation in synchronized PCV2-infected cells dependent upon virus replication. This induction of G0/G1 cell cycle arrest promoted PCV2 replication as evidenced by increased viral protein expression and progeny virus production in the synchronized PCV2-infected cells. To delineate a mechanism of miRNAs in regulating PCV2-induced G0/G1 cell cycle arrest, we determined expression levels of some relevant miRNAs and found that only miR-15a but not miR-16, miR-21, and miR-34a was significantly changed in the PCV2-infected cells. We further demonstrated that upregulation of miR-15a promoted PCV2-induced G0/G1 cell cycle arrest via mediating cyclins D1 and E degradation, in which involves PCV2 growth. These results reveal that G0/G1 cell cycle arrest induced by PCV2 may provide favourable conditions for viral protein expression and progeny production and that miR-15a is implicated in PCV2-induced cell cycle control, thereby contributing to efficient viral replication. PMID:27302568

  15. Overexpression of a glycosyltransferase gene SrUGT74G1 from Stevia improved growth and yield of transgenic Arabidopsis by catechin accumulation.

    PubMed

    Guleria, Praveen; Yadav, Sudesh Kumar

    2014-03-01

    Steviol glycoside and gibberellin biosynthetic routes are known as divergent branches of a common origin in Stevia. A UDP-glycosyltransferase encoded by SrUGT74G1 catalyses the conversion of steviolbioside into stevioside in Stevia rebaudiana leaves. In the present study, transgenic Arabidopsis thaliana overexpressing SrUGT74G1 cDNA from Stevia were developed to check the probability of stevioside biosynthesis in them. However, stevioside accumulation was not evident in transgenics. Also, the transgenic Arabidopsis showed no change in GA3 content on SrUGT74G1 overexpression. Surprisingly, significant accumulation of catechin was noticed in transgenics. The transgenics showed a considerable increase in shoot length, root length and rosette area. An increase in free radical scavenging activity of transgenics was noticed. Moreover, the seed yield of transgenics was also increased by 6-15% than control. Additionally, variation in trichome branching pattern on leaf surface of transgenics was observed. The trichome branching pattern was also validated by exogenous catechin exposure (10, 50, 100 ng ml(-1)) to control plants. Hence, present study reports the probable role of SrUGT74G1 from Stevia in catechin accumulation of transgenic Arabidopsis thaliana. Thus, detailed study in present perspective has revealed the role of Stevia SrUGT74G1 gene in trichome branching pattern, improved vegetative growth, scavenging potential and seed yield by catechin accumulation in transgenic Arabidopsis.

  16. TM4SF5 accelerates G1/S phase progression via cytosolic p27Kip1 expression and RhoA activity.

    PubMed

    Kim, Hyeonjung; Kang, Minkyung; Lee, Sin-Ae; Kwak, Tae Kyoung; Jung, Oisun; Lee, Hyo Jeong; Kim, Sung-Hoon; Lee, Jung Weon

    2010-08-01

    Transmembrane 4 L six family member 5 (TM4SF5) causes epithelial-mesenchymal transition (EMT) for aberrant cell proliferation. However, the effects of TM4SF5 expression on cell cycle are unknown so far. In this study, using hepatocytes that either ectopically or endogenously express TM4SF5 and human hepatocarcinoma tissues, the role of TM4SF5 in G1/S phase progression was examined. We found that TM4SF5 expression accelerated G1/S phase progression with facilitated cyclin D1 and E expression and Rb phosphorylation. Furthermore, TM4SF5 enhanced trafficking of CDK4 and cyclin D1 into the nucleus and induced complex formation between them. However, TM4SF5-facilitated G1/S phase progression was blocked by silencing of p27Kip1 using siRNA or by infection of active RhoA. Pharmacological inhibition of ROCK accelerated the G1/S phase progression of control TM4SF5-unexpressing cells. Altogether, these observations suggest that TM4SF5 accelerates G1/S phase progression with facilitated CDK4/cyclin D1 entry into the nucleus, which might be supported by TM4SF5-mediated actin reorganization through cytosolic p27Kip1 expression and Rho GTPase activity.

  17. Post-transcriptional regulation of cyclins D1, D3 and G1 and proliferation of human cancer cells depend on IMP-3 nuclear localization.

    PubMed

    Rivera Vargas, T; Boudoukha, S; Simon, A; Souidi, M; Cuvellier, S; Pinna, G; Polesskaya, A

    2014-05-29

    RNA-binding proteins of the IMP family (insulin-like growth factor 2 (IGF2) mRNA-binding proteins 1-3) are important post-transcriptional regulators of gene expression. Multiple studies have linked high expression of IMP proteins, and especially of IMP-3, to an unfavorable prognosis in numerous types of cancer. The specific importance of IMP-3 for cancer transformation remains poorly understood. We here show that all three IMPs can directly bind the mRNAs of cyclins D1, D3 and G1 (CCND1, D3 and G1) in vivo and in vitro, and yet only IMP-3 regulates the expression of these cyclins in a significant manner in six human cancer cell lines of different origins. In the absence of IMP-3, the levels of CCND1, D3 and G1 proteins fall dramatically, and the cells accumulate in the G1 phase of the cell cycle, leading to almost complete proliferation arrest. Our results show that, compared with IMP-1 and IMP-2, IMP-3 is enriched in the nucleus, where it binds the transcripts of CCND1, D3 and G1. The nuclear localization of IMP-3 depends on its protein partner HNRNPM and is indispensable for the post-transcriptional regulation of expression of the cyclins. Cytoplasmic retention of IMP-3 and HNRNPM in human cancer cells leads to significant drop in proliferation. In conclusion, a nuclear IMP-3-HNRNPM complex is important for the efficient synthesis of CCND1, D3 and G1 and for the proliferation of human cancer cells.

  18. The possible use of HLA-G1 and G3 in the inhibition of NK cell-mediated swine endothelial cell lysis.

    PubMed

    Matsunami, K; Miyagawa, S; Nakai, R; Murase, A; Shirakura, R

    2001-10-01

    The splicing isoform of HLA-G that is expressed in xenogeneic cells, and its effect on NK-mediated direct cytotoxicity was examined, using stable Chinese hamster ovary (CHO) cell or swine endothelial cell (SEC) transfectants. cDNAs of HLA-G (G1 and G3) and human beta2-microglobulin were prepared and subcloned into the expression vector, pCXN. The transfected HLA-G1 was easily expressed on SEC, and co-transfection with human beta2-microglobulin led to an enhanced level of HLA-G1 expression, as evidenced by flow cytometry. The expressed HLA-G1 significantly suppressed NK-mediated SEC cell lysis, which is an in vitro delayed-type rejection model of a xenograft. On the other hand, the swine leucocyte antigen (SLA) class I molecules could be up-regulated as the result of the transfection of human beta2-microglobulin, but did not down-regulate human NK-mediated SEC lysis. The HLA-G3 was not expressed on CHO and SEC in contrast to HLA-G1, as the result of the transfection. The gene introduction of HLA-G3 in SEC showed no protective effect from human NK cells. However, indirect evidence demonstrated that HLA-G3 transfection resulted in HLA-E expression, but not itself, when transfected to the human cell line, 721.221, thus providing some insight into its natural function in human cells. The present findings suggest that the expression of HLA-G1 on the cell surface could serve as a new approach to overcoming NK-mediated immunity to xenografts.

  19. The transcription factor Swi4 is target for PKA regulation of cell size at the G1 to S transition in Saccharomyces cerevisiae

    PubMed Central

    Amigoni, Loredana; Colombo, Sonia; Belotti, Fiorella; Alberghina, Lilia; Martegani, Enzo

    2015-01-01

    To investigate the specific target of PKA in the regulation of cell cycle progression and cell size we developed a new approach using the yeast strain GG104 bearing a deletion in adenylate cyclase gene and permeable to cAMP ( cyr1Δ, pde2Δ, msn2Δ, msn4Δ). In this strain the PKA activity is absent and can be activated by addition of cAMP in the medium, without any other change of the growth conditions. In the present work we show that the activation of PKA by exogenous cAMP in the GG104 strain exponentially growing in glucose medium caused a marked increase of cell size and perturbation of cell cycle with a transient arrest of cells in G1, followed by an accumulation of cells in G2/M phase with a minimal change in the growth rate. Deletion of CLN1 gene, but not of CLN2, abolished the transient G1 phase arrest. Consistently we found that PKA activation caused a transcriptional repression of CLN1 gene. Transcription of CLN1 is controlled by SBF and MBF dual-regulated promoter. We found that also the deletion of SWI4 gene abolished the transient G1 arrest suggesting that Swi4 is a target responsible for PKA modulation of G1/S phase transition. We generated a SWI4 allele mutated in the consensus site for PKA (Swi4S159A) and we found that expression of Swi4S159A protein in the GG104-Swi4Δ strain did not restore the transient G1 arrest induced by PKA activation, suggesting that Swi4 phosphorylation by PKA regulates CLN1 gene expression and G1/S phase transition. PMID:26046481

  20. Detection of IgG1 antibodies against Mycobacterium tuberculosis DosR and Rpf antigens in tuberculosis patients before and after chemotherapy.

    PubMed

    Mattos, Ana Márcia Menezes; Chaves, Alexandre Silva; Franken, Kees L M C; Figueiredo, Bárbara Bruna Muniz; Ferreira, Ana Paula; Ottenhoff, Tom H M; Teixeira, Henrique Couto

    2016-01-01

    Diagnosis of tuberculosis (TB) remains challenging. Serum IgG1 antibodies against Mycobacterium tuberculosis active growth phase antigens (ESAT-6/CFP-10, Rv0717 and Rv3353), DosR regulon-encoded proteins (Rv1733, Rv1737, Rv2628 and Rv2029), and resuscitation-promoting factors (Rv0867 and Rv2389) were evaluated in TB patients using ELISA. Active TB patients showed elevated levels of IgG1 antibodies against ESAT-6/CFP-10, Rv0717, Rv3353, Rv1733, Rv2628, Rv2029 and Rv0867 in comparison to healthy controls (p < 0.001). These levels remained high after the initiation of treatment, while responses to Rv0717 and Rv1733 peaked early during treatment. IgG1 responses to ESAT-6/CFP-10, Rv3353, Rv2628, Rv2029 and Rv0867 declined to control levels after the completion of 6 months chemotherapy. ROC analysis confirmed the good diagnostic performance of Rv0717, Rv1733, Rv3353, Rv2628, Rv2029 and Rv0867antigens. These data suggest that detecting IgG1 antibodies against M. tuberculosis antigens, including DosR and Rpf proteins, may represent an additional tool in the diagnosis of tuberculosis.

  1. Physical stability comparisons of IgG1-Fc variants: effects of N-glycosylation site occupancy and Asp/Gln residues at site Asn 297.

    PubMed

    Alsenaidy, Mohammad A; Okbazghi, Solomon Z; Kim, Jae Hyun; Joshi, Sangeeta B; Middaugh, C Russell; Tolbert, Thomas J; Volkin, David B

    2014-06-01

    The structural integrity and conformational stability of various IgG1-Fc proteins produced from the yeast Pichia pastoris with different glycosylation site occupancy (di-, mono-, and nonglycosylated) were determined. In addition, the physical stability profiles of three different forms of nonglycosylated Fc molecules (varying amino-acid residues at site 297 in the CH 2 domain due to the point mutations and enzymatic digestion of the Fc glycoforms) were also examined. The physical stability of these IgG1-Fc glycoproteins was examined as a function of pH and temperature by high-throughput biophysical analysis using multiple techniques combined with data visualization tools (three index empirical phase diagrams and radar charts). Across the pH range of 4.0-6.0, the di- and monoglycosylated forms of the IgG1-Fc showed the highest and lowest levels of physical stability, respectively, with the nonglycosylated forms showing intermediate stability depending on solution pH. In the aglycosylated Fc proteins, the introduction of Asp (D) residues at site 297 (QQ vs. DN vs. DD forms) resulted in more subtle changes in structural integrity and physical stability depending on solution pH. The utility of evaluating the conformational stability profile differences between the various IgG1-Fc glycoproteins is discussed in the context of analytical comparability studies. PMID:24740840

  2. Refrigeration of the 18.3 GHz C_3H_2 Transition in Dark Clouds G1.6-0.25

    NASA Technical Reports Server (NTRS)

    Kuiper, T. B. H.; Whiteoak, J. B.; Peng, R. -S.; Peters, W. L., III; Reynolds, J. E.

    1993-01-01

    We have observed the 1_(10)-1_(01) (18.3 GHz) transition of orthocyclopropenylidene, C_(-3)H_(-2), at 24 positions in the unusual dense cloud G1.6- 0.025. Except for one position, the transition is refrigerated, a phenomenon which has not been seen in this transition before.

  3. Recruitment of trimeric proliferating cell nuclear antigen by G1-phase cyclin-dependent kinases following DNA damage with platinum-based antitumour agents

    PubMed Central

    He, G; Kuang, J; Koomen, J; Kobayashi, R; Khokhar, A R; Siddik, Z H

    2013-01-01

    Background: In cycling tumour cells, the binary cyclin-dependent kinase Cdk4/cyclin D or Cdk2/cyclin E complex is inhibited by p21 following DNA damage to induce G1 cell-cycle arrest. However, it is not known whether other proteins are also recruited within Cdk complexes, or their role, and this was investigated. Methods: Ovarian A2780 tumour cells were exposed to the platinum-based antitumour agent 1R,2R-diaminocyclohexane(trans-diacetato)(dichloro)platinum(IV) (DAP), which preferentially induces G1 arrest in a p21-dependent manner. The Cdk complexes were analysed by gel filtration chromatography, immunoblot and mass spectrometry. Results: The active forms of Cdk4 and Cdk2 complexes in control tumour cells have a molecular size of ∼140 kDa, which increased to ∼290 kDa when inhibited following G1 checkpoint activation by DAP. Proteomic analysis identified Cdk, cyclin, p21 and proliferating cell nuclear antigen (PCNA) in the inhibited complex, and biochemical studies provided unequivocal evidence that the increase in ∼150 kDa of the inhibited complex is consistent with p21-dependent recruitment of PCNA as a trimer, likely bound to three molecules of p21. Although p21 alone was sufficient to inhibit the Cdk complex, PCNA was critical for stabilising p21. Conclusion: G1 Cdk complexes inhibited by p21 also recruit PCNA, which inhibits degradation and, thereby, prolongs activity of p21 within the complex. PMID:24104967

  4. Characterization of a G1P[8] rotavirus causing an outbreak of gastroenteritis in the Northern Territory, Australia, in the vaccine era

    PubMed Central

    Donato, Celeste M; Cowley, Daniel; Snelling, Thomas L; Akopov, Asmik; Kirkness, Ewen F; Kirkwood, Carl D

    2014-01-01

    In 2010, a large outbreak of rotavirus gastroenteritis occurred in the Alice Springs region of the Northern Territory, Australia. The outbreak occurred 43 months after the introduction of the G1P[8] rotavirus vaccine Rotarix®. Forty-three infants were hospitalized during the outbreak and analysis of fecal samples from each infant revealed a G1P[8] rotavirus strain. The outbreak strain was adapted to cell culture and neutralization assays were performed using VP7 and VP4 neutralizing monoclonal antibodies. The outbreak strain exhibited a distinct neutralization resistance pattern compared to the Rotarix® vaccine strain. Whole genome sequencing of the 2010 outbreak virus strain demonstrated numerous amino acid differences compared to the Rotarix® vaccine strain in the characterized neutralization epitopes of the VP7 and VP4 proteins. Phylogenetic analysis of the outbreak strain revealed a close genetic relationship to global strains, in particular RVA/Human-wt/BEL/BE0098/2009/G1P[8] and RVA/Human-wt/BEL/BE00038/2008/G1P[8] for numerous genes. The 2010 outbreak strain was likely introduced from a globally circulating population of strains rather than evolving from an endemic Australian strain. The outbreak strain possessed antigenic differences in the VP7 and VP4 proteins compared to the Rotarix® vaccine strain. The outbreak was associated with moderate vaccine coverage and possibly low vaccine take in the population. PMID:26038746

  5. Immunophenotyping and characterization of BNP colostra revealed pathogenic alloantibodies of IgG1 subclass with specifity to platelets, granulocytes and monocytes of all maturation stages.

    PubMed

    Assad, Aryan; Amann, Barbara; Friedrich, Annette; Deeg, Cornelia Annette

    2012-06-15

    Bovine neonatal pancytopenia (BNP) is mainly characterized by multiple haemorrhages, thrombocytopenia and leukocytopenia as a result of bone marrow depletion. BNP can be induced in healthy calves through application of colostrum from BNP donors, proofing that BNP is mediated to maternal alloantibodies. Alloantibody binding to bovine blood cells is present in sera and colostra of BNP donors and is probably initialized by vaccination with a certain BVD vaccine. To understand etiology and pathomechanisms of BNP, we closely characterized disease inducing antibodies regarding immunoglobulin subclass and binding specificities to peripheral blood derived leukocytes and platelets. By exact phenotyping the targeted blood cell subsets, including platelets for the first time, we investigated that BNP alloantibodies are exclusively of IgG1 subclass. Interestingly, IgG1 of BNP colostra bound to 70% leukocytes and 100% platelets irrespective of different bovine breeds and cellular maturity of all specimens tested. Furthermore, staining pattern on platelets as well as leukocyte subsets by BNP-IgG1 alloantibody exposed 100% reactivity to platelets, granulocytes and monocytes. Interestingly, the main part of T-helper cells was not bound by colostral alloantibodies. Our results point to a crucial role of IgG1 antibodies in BNP and to a target antigen that is expressed by all cells of myeloid lineage, but only partially by the lymphoid lineage. PMID:22554492

  6. Knockdown of Sec8 promotes cell-cycle arrest at G1/S phase by inducing p21 via control of FOXO proteins.

    PubMed

    Tanaka, Toshiaki; Iino, Mituyoshi

    2014-02-01

    p21(Cip1) protein inhibits the activity of cyclins at the G(1) checkpoint and influences transition of cells from the G(1) to the S phase of the cell cycle. Moreover, expression of members of the FOXO family (active form of forkhead transcription factors of the O class) in dividing cells promotes cell-cycle arrest at the G(1)/S boundary via regulation of p21(Cip1). Recently, the exocyst complex, including Sec8, has been implicated in various roles independent of its role in secretion, such as cell migration, invadopodia formation, cytokinesis, glucose uptake and neural development. Given the essential roles of the exocyst complex in cellular and developmental processes, disruption of its function may be involved in various diseases such as cancer, diabetes and neuronal disorders. However, the relationship between Sec8 and the cell cycle remains to be elucidated. In this study, knockdown of Sec8 inhibited cell growth and promoted cell-cycle arrest at the G(1)/S phase by control of p21 expression and retinoblastoma protein phosphorylation. Furthermore, Sec8 regulated FOXO family proteins via ubiquitin-proteasome degradation by regulating the expression of the murine double minute 2 (Mdm2) protein but not S-phase kinase-associated protein 2 (Skp2).

  7. 26 CFR 1.1033(g)-1 - Condemnation of real property held for productive use in trade or business or for investment.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 26 Internal Revenue 11 2011-04-01 2011-04-01 false Condemnation of real property held for... (CONTINUED) Common Nontaxable Exchanges § 1.1033(g)-1 Condemnation of real property held for productive use..., of real property held either for productive use in trade or business or for investment (not...

  8. Peptide-Mediated Interference of PB2-eIF4G1 Interaction Inhibits Influenza A Viruses' Replication in Vitro and in Vivo.

    PubMed

    Yuan, Shuofeng; Chu, Hin; Ye, Jiahui; Hu, Meng; Singh, Kailash; Chow, Billy K C; Zhou, Jie; Zheng, Bo-Jian

    2016-07-01

    Influenza viruses are obligate parasites that hijack the host cellular system. Previous results have shown that the influenza virus PB2 subunit confers a dependence of host eukaryotic translation initiation factor 4-γ 1 (eIF4G1) for viral mRNA translation. Here, we demonstrated that peptide-mediated interference of the PB2-eIF4G1 interaction inhibited virus replication in vitro and in vivo. Remarkably, intranasal administration of the peptide provided 100% protection against lethal challenges of influenza A viruses in BALB/c mice, including H1N1, H5N1, and H7N9 influenza virus subtypes. Mapping of the PB2 protein indicated that the eIF4G1 binding sites resided within the PB2 cap-binding domain. Virtual docking analysis suggested that the inhibitory peptide associated with the conserved amino acid residues that were essential to PB2 cap-binding activity. Overall, our results identified the PB2-eIF4G1 interactive site as a druggable target for influenza therapeutics. PMID:27626099

  9. The Pseudomonas syringae Type III Effector HopG1 Induces Actin Remodeling to Promote Symptom Development and Susceptibility during Infection1[OPEN

    PubMed Central

    Shimono, Masaki; Porter, Katie; Kvitko, Brian H.; Henty-Ridilla, Jessica; Creason, Allison; Chang, Jeff H.; Staiger, Christopher J.

    2016-01-01

    The plant cytoskeleton underpins the function of a multitude of cellular mechanisms, including those associated with developmental- and stress-associated signaling processes. In recent years, the actin cytoskeleton has been demonstrated to play a key role in plant immune signaling, including a recent demonstration that pathogens target actin filaments to block plant defense and immunity. Herein, we quantified spatial changes in host actin filament organization after infection with Pseudomonas syringae pv. tomato DC3000 (Pst DC3000), demonstrating that the type-III effector HopG1 is required for pathogen-induced changes to actin filament architecture and host disease symptom development during infection. Using a suite of pathogen effector deletion constructs, coupled with high-resolution microscopy, we found that deletion of hopG1 from Pst DC3000 resulted in a reduction in actin bundling and a concomitant increase in the density of filament arrays in Arabidopsis, both of which correlate with host disease symptom development. As a mechanism underpinning this activity, we further show that the HopG1 effector interacts with an Arabidopsis mitochondrial-localized kinesin motor protein. Kinesin mutant plants show reduced disease symptoms after pathogen infection, which can be complemented by actin-modifying agents. In total, our results support a model in which HopG1 induces changes in the organization of the actin cytoskeleton as part of its virulence function in promoting disease symptom development. PMID:27217495

  10. A method for the low-level (ng g(-1)) determination of perfluorooctanoate in paper and textile by liquid chromatography with tandem mass spectrometry.

    PubMed

    Stadalius, Marilyn; Connolly, Paul; L'Empereur, Karen; Flaherty, John M; Isemura, Tsuguhide; Kaiser, Mary A; Knaup, Wolfgang; Noguchi, Masahiro

    2006-08-01

    The determination of perfluorooctanoate (PFO) in articles of commerce has become increasingly important to understand if treated products are a possible source of PFO. An LC-MS/MS method for the determination of PFO in paper and textile using a dual labeled 13C-PFOA internal standard was successfully developed and validated. Residues of PFO were determined using an isocratic, reversed-phase high-performance liquid chromatography (HPLC) method with an ammonium acetate/methanol buffer. Ions monitored were 413 (parent) and 369 (daughter) for PFO and 415 (parent) and 370 (daughter) for dual labeled 13C-PFOA internal standard. As a precaution against ubiquitous PFO that occasionally occurs in mobile phase or instrument components, two Hypercarb cartridges (4 mm) were placed before the HPLC injector. Any PFO that was captured by the cartridges was removed before each injection by flushing the system with 100% methanol prior to equilibration with the isocratic mobile phase. Overall recovery and standard deviation over a 3 day validation regimen for samples (n=54-55) fortified with PFOA at 5, 50, and 200 ng g(-1) were 114+/-4.9% for textile and 110+/-7.6% for paper. The results also established a limit of detection (LOD) of 1 ng g(-1) in textile and 2 ng g(-1) in paper based upon S/N of the 5.0 ng g(-1) fortification versus the untreated paper and textile.

  11. 26 CFR 1.1033(g)-1 - Condemnation of real property held for productive use in trade or business or for investment.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 26 Internal Revenue 11 2013-04-01 2013-04-01 false Condemnation of real property held for... (CONTINUED) Common Nontaxable Exchanges § 1.1033(g)-1 Condemnation of real property held for productive use..., of real property held either for productive use in trade or business or for investment (not...

  12. Intrinsic Properties of immunoglobulin IgG1 Isotype-Switched B Cell Receptors Promote Microclustering and the Initiation of Signaling

    PubMed Central

    Liu, Wanli; Meckel, Tobias; Tolar, Pavel; Sohn, Hae Won; Pierce, Susan K.

    2010-01-01

    Summary Memory B cells express high affinity, immunoglobulin B cell receptors (IgG-BCRs) that enhance B cell responses giving rise to the rapid production of high affinity, IgG antibodies. Despite the central role of IgG-BCRs in memory responses, the mechanisms by which the IgG-BCRs function to enhance B cell responses are not fully understood. Using high-resolution live-cell imaging we showed that independent of affinity, IgG1-BCRs dramatically enhanced the earliest BCR-intrinsic events that followed within seconds of B cells’ encounter with membrane bound antigen including BCR oligomerization and BCR microcluster growth, leading to Syk kinase recruitment and calcium responses. The enhancement of these early events was dependent on a membrane proximal region of the IgG1 cytoplasmic tail not previously appreciated to play a role in IgG1-BCR signaling. Thus, intrinsic properties of the IgG1-BCR enhance early antigen-driven events that ultimately translate into heightened signaling. PMID:20620943

  13. Characterization of a G1P[8] rotavirus causing an outbreak of gastroenteritis in the Northern Territory, Australia, in the vaccine era.

    PubMed

    Donato, Celeste M; Cowley, Daniel; Snelling, Thomas L; Akopov, Asmik; Kirkness, Ewen F; Kirkwood, Carl D

    2014-07-01

    In 2010, a large outbreak of rotavirus gastroenteritis occurred in the Alice Springs region of the Northern Territory, Australia. The outbreak occurred 43 months after the introduction of the G1P[8] rotavirus vaccine Rotarix(®). Forty-three infants were hospitalized during the outbreak and analysis of fecal samples from each infant revealed a G1P[8] rotavirus strain. The outbreak strain was adapted to cell culture and neutralization assays were performed using VP7 and VP4 neutralizing monoclonal antibodies. The outbreak strain exhibited a distinct neutralization resistance pattern compared to the Rotarix(®) vaccine strain. Whole genome sequencing of the 2010 outbreak virus strain demonstrated numerous amino acid differences compared to the Rotarix(®) vaccine strain in the characterized neutralization epitopes of the VP7 and VP4 proteins. Phylogenetic analysis of the outbreak strain revealed a close genetic relationship to global strains, in particular RVA/Human-wt/BEL/BE0098/2009/G1P[8] and RVA/Human-wt/BEL/BE00038/2008/G1P[8] for numerous genes. The 2010 outbreak strain was likely introduced from a globally circulating population of strains rather than evolving from an endemic Australian strain. The outbreak strain possessed antigenic differences in the VP7 and VP4 proteins compared to the Rotarix(®) vaccine strain. The outbreak was associated with moderate vaccine coverage and possibly low vaccine take in the population. PMID:26038746

  14. Susceptibility and intra-species transmission of the H9N2 G1 prototype lineage virus in Japanese quail and turkeys.

    PubMed

    Bonfante, Francesco; Patrono, Livia Victoria; Aiello, Roberta; Beato, Maria Serena; Terregino, Calogero; Capua, Ilaria

    2013-07-26

    Avian influenza viruses of the H9N2 subtype have circulated in the poultry population in Asia, Far and Middle East since the mid-1990 s. One of the most widespread lineages established in poultry is the G1 lineage. This lineage has undergone further evolution and reassortment since its first detection in 1997 and G1-like H9N2 viruses still circulate. In this study we have investigated the susceptibility of quail and turkeys to the H9N2 G1-lineage prototype strain (A/quail/Hong Kong/G1/97). Contact transmission experiments were carried out in both avian species. Animals were infected oro-nasally with increasing doses of the virus (10(3)-10(6) EID 50/0.1 ml) and sentinel birds were introduced 4 days post infection (pi) in each experimental group. Quail were more susceptible than turkeys, as they were readily infected with lower challenge doses. Interestingly, infection of turkeys was associated with worse clinical condition. Transmission was detected in both species. Quail infected with a dose less than or equal to 10(4) EID50 transmitted the virus to the sentinels without showing any signs of disease. These findings reinforce the hypothesis that quail may ensure the perpetuation of H9N2 viruses in poultry, acting as a silent reservoir. PMID:23597652

  15. Soluble and membrane-bound Drosophila melanogaster CYP6G1 expressed in Escherichia coli: purification, activity, and binding properties toward multiple pesticides.

    PubMed

    Cheesman, Matthew J; Traylor, Matthew J; Hilton, Margaret E; Richards, Katelyn E; Taylor, Matthew C; Daborn, Phillip J; Russell, Robyn J; Gillam, Elizabeth M J; Oakeshott, John G

    2013-05-01

    Cytochrome P450 CYP6G1 has been implicated in the resistance of Drosophila melanogaster to numerous pesticides. While in vivo and in vitro studies have provided insight to the diverse functions of this enzyme, direct studies on the isolated CYP6G1 enzyme have not been possible due to the need for a source of recombinant enzyme. In the current study, the Cyp6g1 gene was isolated from D. melanogaster and re-engineered for heterologous expression in Escherichia coli. Approximately 460 nmol L⁻¹ of P450 holoenzyme were obtained in 500 mL cultures. The recombinant enzyme was located predominantly within the bacterial cytosol. A two-step purification protocol using Ni-chelate affinity chromatography followed by removal of detergent on a hydroxyapatite column produced essentially homogenous enzyme from both soluble and membrane fractions. Recombinant CYP6G1 exhibited p-nitroanisole O-dealkylation activity but was not active against eleven other typical P450 marker substrates. Substrate-induced binding spectra and IC₅₀ values for inhibition of p-nitroanisole O-dealkylation were obtained for a wide selection of pesticides, namely DDT, imidacloprid, chlorfenvinphos, malathion, endosulfan, dieldrin, dicyclanil, lufenuron and carbaryl, supporting previous in vivo and in vitro studies on Drosophila that have suggested that the enzyme is involved in multi-pesticide resistance in insects. PMID:23470655

  16. Characterization of a G1P[8] rotavirus causing an outbreak of gastroenteritis in the Northern Territory, Australia, in the vaccine era.

    PubMed

    Donato, Celeste M; Cowley, Daniel; Snelling, Thomas L; Akopov, Asmik; Kirkness, Ewen F; Kirkwood, Carl D

    2014-07-01

    In 2010, a large outbreak of rotavirus gastroenteritis occurred in the Alice Springs region of the Northern Territory, Australia. The outbreak occurred 43 months after the introduction of the G1P[8] rotavirus vaccine Rotarix(®). Forty-three infants were hospitalized during the outbreak and analysis of fecal samples from each infant revealed a G1P[8] rotavirus strain. The outbreak strain was adapted to cell culture and neutralization assays were performed using VP7 and VP4 neutralizing monoclonal antibodies. The outbreak strain exhibited a distinct neutralization resistance pattern compared to the Rotarix(®) vaccine strain. Whole genome sequencing of the 2010 outbreak virus strain demonstrated numerous amino acid differences compared to the Rotarix(®) vaccine strain in the characterized neutralization epitopes of the VP7 and VP4 proteins. Phylogenetic analysis of the outbreak strain revealed a close genetic relationship to global strains, in particular RVA/Human-wt/BEL/BE0098/2009/G1P[8] and RVA/Human-wt/BEL/BE00038/2008/G1P[8] for numerous genes. The 2010 outbreak strain was likely introduced from a globally circulating population of strains rather than evolving from an endemic Australian strain. The outbreak strain possessed antigenic differences in the VP7 and VP4 proteins compared to the Rotarix(®) vaccine strain. The outbreak was associated with moderate vaccine coverage and possibly low vaccine take in the population.

  17. Physical stability comparisons of IgG1-Fc variants: effects of N-glycosylation site occupancy and Asp/Gln residues at site Asn 297.

    PubMed

    Alsenaidy, Mohammad A; Okbazghi, Solomon Z; Kim, Jae Hyun; Joshi, Sangeeta B; Middaugh, C Russell; Tolbert, Thomas J; Volkin, David B

    2014-06-01

    The structural integrity and conformational stability of various IgG1-Fc proteins produced from the yeast Pichia pastoris with different glycosylation site occupancy (di-, mono-, and nonglycosylated) were determined. In addition, the physical stability profiles of three different forms of nonglycosylated Fc molecules (varying amino-acid residues at site 297 in the CH 2 domain due to the point mutations and enzymatic digestion of the Fc glycoforms) were also examined. The physical stability of these IgG1-Fc glycoproteins was examined as a function of pH and temperature by high-throughput biophysical analysis using multiple techniques combined with data visualization tools (three index empirical phase diagrams and radar charts). Across the pH range of 4.0-6.0, the di- and monoglycosylated forms of the IgG1-Fc showed the highest and lowest levels of physical stability, respectively, with the nonglycosylated forms showing intermediate stability depending on solution pH. In the aglycosylated Fc proteins, the introduction of Asp (D) residues at site 297 (QQ vs. DN vs. DD forms) resulted in more subtle changes in structural integrity and physical stability depending on solution pH. The utility of evaluating the conformational stability profile differences between the various IgG1-Fc glycoproteins is discussed in the context of analytical comparability studies.

  18. Neisseria meningitidis Group A IgG1 and IgG2 Subclass Immune Response in African Children Aged 12–23 Months Following Meningococcal Vaccination

    PubMed Central

    Holme, Daniel; Findlow, Helen; Sow, Samba O.; Idoko, Olubukola T.; Preziosi, Marie-Pierre; Carlone, George; Plikaytis, Brian D.; Borrow, Ray

    2015-01-01

    Background. A group A meningococcal conjugate vaccine, PsA-TT, was licensed in 2010 and was previously studied in a phase 2 clinical trial to evaluate its safety and immunogenicity in African children 12–23 months of age. Methods. Subjects received either PsA-TT; meningococcal group A, C, W, Y polysaccharide vaccine (PsACWY); or Haemophilus influenzae type b conjugate vaccine (Hib-TT). Forty weeks following primary vaccination, the 3 groups were further randomized to receive either PsA-TT, one-fifth dose of PsACWY, or Hib-TT. Group A–specific immunoglobulin G (IgG) subclass response was characterized using an enzyme-linked immunosorbent assay. Results. The predominant IgG subclass response, regardless of vaccine, was IgG1. One month following primary vaccination, the geometric mean concentrations (GMCs) of IgG1 and IgG2 in the PsA-TT group were 21.73 µg/mL and 6.27 µg/mL, whereas in the PsACWY group the mean GMCs were 2.01 µg/mL and 0.97 µg/mL, respectively (P < .0001). Group A–specific IgG1 and IgG2 GMCs remained greater in the PsA-TT group than in the PsACWY group 40 weeks following primary vaccination (P < .0001). One week following revaccination, those given 2 doses of PsA-TT had the greatest IgG1 and IgG2 GMCs of 125.23 µg/mL and 36.12 µg/mL, respectively (P = .0008), and demonstrated a significant increase in IgG1:IgG2 mean ratio, indicative of the T-cell–dependent response associated with conjugate vaccines. Conclusions. Vaccination of African children aged 12–24 months with either PsA-TT or PsACWY elicited a predominantly IgG1 response. The IgG1:IgG2 mean ratio decreased following successive vaccination with PsACWY, indicating a shift toward IgG2, suggestive of the T-cell–independent immune response commonly associated with polysaccharide antigens. Clinical Trials Registration. SRCTN78147026. PMID:26553689

  19. Mycoflora and Co-Occurrence of Fumonisins and Aflatoxins in Freshly Harvested Corn in Different Regions of Brazil

    PubMed Central

    Rocha, Liliana O.; Nakai, Viviane K.; Braghini, Raquel; Reis, Tatiana A.; Kobashigawa, Estela; Corrêa, Benedito

    2009-01-01

    Natural mycoflora and co-occurrence of fumonisins (FB1, FB2) and aflatoxins (AFB1, AFB2, AFG1 and AFG2) in freshly harvested corn grain samples from four regions of Brazil were investigated. Fusarium verticillioides was predominant in all samples. Analysis of fumonisins showed that 98% of the samples were contaminated with FB1 and 74.5% with FB1 + FB2, with toxin levels ranging from 0.015 to 9.67 μg/g for FB1 and from 0.015 to 3.16 μg/g for FB2. Twenty-one (10.5%) samples were contaminated with AFB1, seven (3.5%) with AFB2 and only one (0.5%) with AFG1 and AFG2 Co-contamination with aflatoxins and fumonisins was observed in 7% of the samples. The highest contamination of fumonisins and aflatoxins was observed in Nova Odessa (SP) and Várzea Grande (MT), respectively. The lowest contamination of these mycotoxins was found in Várzea Grande and Nova Odessa, respectively. PMID:20087478

  20. Simultaneous detection of multiple mycotoxins in broiler feeds using a liquid chromatography tandem-mass spectrometry

    PubMed Central

    KONGKAPAN, Jutamart; POAPOLATHEP, Saranya; ISARIYODOM, Supaporn; KUMAGAI, Susumu; POAPOLATHEP, Amnart

    2015-01-01

    Mycotoxins are secondary fungal metabolites that are typically present in grain and feed ingredients used for animal feeds. An analytical method using LC-ESI-MS/MS was developed to quantify nine mycotoxins, consisting of aflatoxin B1 (AFB1), AFB2, AFG1, AFG2, T-2 toxin, deoxynivalenol (DON), nivalenol (NIV), zearalenone (ZEA) and ochratoxin A (OTA) in broiler feeds. In total, 100 samples of broiler feeds were collected from poultry farms in Central Thailand. The survey found that AFB1 and ZEA were the most prevalent mycotoxins in the feed samples at percentages of 93% and 63%, respectively. The limit of detections (LODs) of investigated mycotoxins was 0.20–0.78 ng/g. AFB2, DON, AFG1, NIV and T-2 toxin were also detectable at low contamination levels with percentages of 20%, 9%, 7%, 5% and 1%, respectively, whereas OTA and AFG2 were not detected in any of the feed samples. These results suggest that there is a very low level of risk of the exposure to mycotoxins in feeds obtained from broiler farms in Central Thailand. PMID:26477362

  1. Glucose transporter type I deficiency (G1D) at 25 (1990–2015): Presumptions, facts and the lives of persons with this rare disease

    PubMed Central

    Pascual, Juan M.; Ronen, Gabriel M.

    2015-01-01

    As is often the case for rare diseases, the number of published reviews and case reports of Glucose transporter type I deficiency (G1D) approaches or exceeds that of original research. This can indicate medical interest, but also scientific stagnation. In assessing this state of affairs here, we focus not on what is peculiar or disparate about G1D, but on the assumptions that have reigned thus far undisputed, and critique them as a potential impediment to progress. To summarize the most common G1D phenotype, we trace the 25-year story of G1D in parallel with the natural history of one of two index patients, identified in 1990 by one of us (G.M.R.) and brought up to date by the other (J.M.P.) while later examining widely-repeated but little-scrutinized statements. Among them are those that pertain to assumptions about brain fuels; energy-failure; cerebrospinal glucose concentration; the purpose of ketogenic diet; the role of the defective blood brain barrier; genotype-phenotype correlations; a bewildering array of phenotypes; ictogenesis, seizures and the electroencephalogram; the use of mice to model the disorder; and what treatments may and may not be expected to accomplish. We reach the forgone conclusion that the proper study of mankind - and of one of its ailments (G1D) - is man itself (rather than mice, isolated cells or extrapolated inferences), and propose a framework for rigorous investigation that we hope will lead to a better understanding and to better treatments for this and for rare disorders in general. These considerations, together with experience drawn from other disorders, lead, as a logical consequence, to the nullification of the view that therapeutic development (i.e., trials) for rare diseases could or should be accelerated without the most vigorous scientific scrutiny: Trial and error constitute an inseparable couple, such that, at the present time, hastening the former is bound to precipitate the latter. PMID:26341673

  2. Chronic treatment with the G protein-coupled receptor 30 agonist G-1 decreases blood pressure in ovariectomized mRen2.Lewis rats.

    PubMed

    Lindsey, Sarah Hoffmann; Cohen, Jonathan A; Brosnihan, K Bridget; Gallagher, Patricia E; Chappell, Mark C

    2009-08-01

    The mRen2.Lewis congenic strain is an estrogen-sensitive model of hypertension whereby estrogen depletion produces a significant and sustained increase in blood pressure. The recent identification of G protein-coupled receptor 30 (GPR30) as a third estrogen receptor isotype prompted us to test the hypothesis that this novel receptor exhibits beneficial cardiovascular actions in the hypertensive female mRen2.Lewis rat. Intact female, ovariectomized female (OVX), and male mRen2.Lewis rats were treated with the selective GPR30 agonist G-1 or vehicle via osmotic minipump for 2 wk. G-1 significantly reduced systolic blood pressure in OVX (178 +/- 7 to 142 +/- 10 mm Hg, P < 0.001, n = 8) but not intact female (144 +/- 3 to 143 +/- 5 mm Hg, P > 0.05, n = 5) or male mRen2.Lewis rats (207 +/- 7 to 192 +/- 5 mm Hg, P > 0.05, n = 7). G-1 did not alter uterine or body weight in OVX, suggesting activation of a receptor distinct from estrogen receptor-alpha and -beta. In isolated aortic rings from OVX, G-1 reduced constriction in response to angiotensin II. Vascular angiotensin-converting enzyme and angiotensin type 1 receptor mRNA were also lower, whereas angiotensin-converting enzyme-2 mRNA was increased. G-1 treatment in OVX was not associated with alterations in either endothelial nitric oxide synthase expression or acetylcholine-induced relaxation. Immunohistochemical staining for GPR30 was evident in both the intima and media of the aorta. We conclude that the novel estrogen receptor GPR30 may contribute to the beneficial cardiovascular actions of estrogen in female mRen2.Lewis rats through regulation of vascular components of the renin-angiotensin system.

  3. Identification of Cofilin-1 Induces G0/G1 Arrest and Autophagy in Angiotensin-(1-7)-treated Human Aortic Endothelial Cells from iTRAQ Quantitative Proteomics

    PubMed Central

    Wang, Huang-Joe; Chen, Sung-Fang; Lo, Wan-Yu

    2016-01-01

    The angiotensin-converting enzyme 2/angiotensin-(1-7)/Mas axis is a pathway that acts against the detrimental effects of the renin-angiotensin system. However, the effects of angiotensin-(1-7) on endothelial protein expression and the related phenotypes are unclear. We performed a duplicate of iTRAQ quantitative proteomic analysis on human aortic endothelial cells (HAECs) treated with angiotensin-(1-7) for 6 hours. Cofilin-1 was identified as a highly abundant candidate with consistent >30% coverage and >1.2-fold overexpression in the angiotensin-(1-7)-treated group. Gene ontology analysis showed that the “regulation_of_mitosis” was significantly altered, and cell cycle analysis indicated that the 6-hour angiotensin-(1-7) treatment significantly induced G0/G1 arrest. Knockdown of the cofilin-1 (CFL1) gene suggested the G0/G1 phase arrest was mediated by the modulation of p27 and the p21/Cyclin/CDK complex by Cofilin-1. Interestingly, quiescent HAECs escaped G0/G1 arrest upon angiotensin-(1-7) treatment for 24 hours, and angiotensin-(1-7) induced autophagy by upregulating Beclin-1 and microtubule-associated protein 1 light chain 3b-II expression, which was also attenuated by A779 pre-treatment and CFL1 knockdown. After pre-treatment with 3-methyladenine (3MA), treatment with angiotensin-(1-7) for 24 h induced significant G0/G1 phase arrest and apoptosis, suggesting a pro-survival role of autophagy in this context. In conclusion, Cofilin-1 plays a dominant role in angiotensin-(1-7)-induced G0/G1 arrest and autophagy to maintain cellular homeostasis in HAECs. PMID:27748441

  4. PAXX and XLF DNA repair factors are functionally redundant in joining DNA breaks in a G1-arrested progenitor B-cell line.

    PubMed

    Kumar, Vipul; Alt, Frederick W; Frock, Richard L

    2016-09-20

    Classical nonhomologous end joining (C-NHEJ) is a major mammalian DNA double-strand break (DSB) repair pathway. Core C-NHEJ factors, such as XRCC4, are required for joining DSB intermediates of the G1 phase-specific V(D)J recombination reaction in progenitor lymphocytes. Core factors also contribute to joining DSBs in cycling mature B-lineage cells, including DSBs generated during antibody class switch recombination (CSR) and DSBs generated by ionizing radiation. The XRCC4-like-factor (XLF) C-NHEJ protein is dispensable for V(D)J recombination in normal cells, but because of functional redundancy, it is absolutely required for this process in cells deficient for the ataxia telangiectasia-mutated (ATM) DSB response factor. The recently identified paralogue of XRCC4 and XLF (PAXX) factor has homology to these two proteins and variably contributes to ionizing radiation-induced DSB repair in human and chicken cells. We now report that PAXX is dispensable for joining V(D)J recombination DSBs in G1-arrested mouse pro-B-cell lines, dispensable for joining CSR-associated DSBs in a cycling mouse B-cell line, and dispensable for normal ionizing radiation resistance in both G1-arrested and cycling pro-B lines. However, we find that combined deficiency for PAXX and XLF in G1-arrested pro-B lines abrogates DSB joining during V(D)J recombination and sensitizes the cells to ionizing radiation exposure. Thus, PAXX provides core C-NHEJ factor-associated functions in the absence of XLF and vice versa in G1-arrested pro-B-cell lines. Finally, we also find that PAXX deficiency has no impact on V(D)J recombination DSB joining in ATM-deficient pro-B lines. We discuss implications of these findings with respect to potential PAXX and XLF functions in C-NHEJ.

  5. Colletotrichum orbiculare Regulates Cell Cycle G1/S Progression via a Two-Component GAP and a GTPase to Establish Plant Infection[OPEN

    PubMed Central

    2015-01-01

    Morphogenesis in filamentous fungi depends on appropriate cell cycle progression. Here, we report that cells of the cucumber anthracnose fungus Colletotrichum orbiculare regulate G1/S progression via a two-component GAP, consisting of Budding-uninhibited-by-benomyl-2 (Bub2) and Byr-four-alike-1 (Bfa1) as well as its GTPase Termination-of-M-phase-1 (Tem1) to establish successful infection. In a random insertional mutagenesis screen of infection-related morphogenesis, we isolated a homolog of Saccharomyces cerevisiae, BUB2, which encodes a two-component Rab GAP protein that forms a GAP complex with Bfa1p and negatively regulates mitotic exit. Interestingly, disruption of either Co BUB2 or Co BFA1 resulted in earlier onset of nuclear division and decreased the time of phase progression from G1 to S during appressorium development. S. cerevisiae GTPase Tem1p is the downstream target of the Bub2p/Bfa1p GAP complex. Introducing the dominant-negative form of Co Tem1 into Co bub2Δ or Co bfa1Δ complemented the defect in G1/S progression, indicating that Co Bub2/Co Bfa1 regulates G1/S progression via Co Tem1. Based on a pathogenicity assay, we found that Co bub2Δ and Co bfa1Δ reduced pathogenesis by attenuating infection-related morphogenesis and enhancing the plant defense response. Thus, during appressorium development, C. orbiculare Bub2/Bfa1 regulates G1/S progression via Co Tem1, and this regulation is essential to establish plant infection. PMID:26320225

  6. Inhibition of Rac1 activity induces G1/S phase arrest through the GSK3/cyclin D1 pathway in human cancer cells.

    PubMed

    Liu, Linna; Zhang, Hongmei; Shi, Lei; Zhang, Wenjuan; Yuan, Juanli; Chen, Xiang; Liu, Juanjuan; Zhang, Yan; Wang, Zhipeng

    2014-10-01

    Rac1 has been shown to regulate the cell cycle in cancer cells. Yet, the related mechanism remains unclear. Thus, the present study aimed to investigate the mechanism involved in the regulation of G1/S phase transition by Rac1 in cancer cells. Inhibition of Rac1 by inhibitor NSC23766 induced G1/S phase arrest and inhibited the proliferation of A431, SW480 and U2-OS cells. Suppression of GSK3 by shRNA partially rescued G1/S phase arrest and inhibition of proliferation. Incubation of cells with NSC23766 reduced p-AKT and inactivated p-GSK3α and p-GSK3β, increased p-cyclin D1 expression and decreased the level of cyclin D1 protein. Consequently, cyclin D1 targeting transcriptional factor E2F1 expression, which promotes G1 to S phase transition, was also reduced. In contrast, constitutive active Rac1 resulted in increased p-AKT and inactivated p-GSK3α and p-GSK3β, decreased p-cyclin D1 expression and enhanced levels of cyclin D1 and E2F1 expression. Moreover, suppression of GSK3 did not alter p-AKT or Rac1 activity, but decreased p-cyclin D1 and increased total cyclin D1 protein. However, neither Rac1 nor GSK3 inhibition altered cyclin D1 at the RNA level. Moreover, after inhibition of Rac1 or GSK3 following proteasome inhibitor MG132 treatment, cyclin D1 expression at the protein level remained constant, indicating that Rac1 and GSK3 may regulate cyclin D1 turnover through phosphorylation and degradation. Therefore, our findings suggest that inhibition of Rac1 induces cell cycle G1/S arrest in cancer cells by regulation of the GSK3/cyclin D1 pathway.

  7. Relative biological effectiveness of 6 MeV neutrons with respect to cell inactivation and disturbances of the G1 phase.

    PubMed

    Zölzer, F; Streffer, C

    2008-02-01

    The relative biological effectiveness (RBE) of neutrons and other types of densely ionizing radiation appears to be close to 1.0 for the induction of strand breaks, but considerably higher RBEs have been found for cellular end points such as colony-forming ability. This may be due to differences in the processing of strand breaks or to the involvement of other lesions whose yields are more dependent on radiation quality. Because cell cycle delays may be of great importance in the processing of DNA damage, we determined the RBE for disturbances of the G1 phase in four different cell types (Be11 melanoma, 4197 squamous cell carcinoma, EA14 glioma, GM6419 fibroblasts) and compared them with the RBE for cell inactivation. The method we used to determine the progress from G1 into S was as follows: Cells were serum-deprived for a number of days and then stimulated to grow with culture medium containing normal amounts of serum. Immediately before the change of medium, cells were exposed to graded doses of either 240 kV X rays or 6 MeV neutrons. At different times afterward, cells were labeled with BrdU and the numbers of active S-phase cells were assessed using two-parameter flow cytometry. For all four cell types, cells started to progress from G1 into S after a few hours. Radiation suppressed this process in all cases, but there were some interesting differences. For Be11 and 4197 cells, the most obvious effect was a delay in G1; the labeling index increased a few hours later in irradiated samples than in controls, and there was no significant effect on the maximum labeling index. For EA14 and GM6419 cells, although smaller doses were used because of greater radiosensitivity, a delay of the entry into S phase was again noticeable, but the most significant effect was a reduction in the maximum percentage of active S-phase cells after stimulation, indicating a permanent or long-term arrest in G1. The RBE for the G1 delay was the same for all four cell types, about 2

  8. Comparative In Vitro and In Vivo Studies of Porcine Rotavirus G9P[13] and Human Rotavirus Wa G1P[8

    PubMed Central

    Shao, Lulu; Fischer, David D.; Kandasamy, Sukumar; Rauf, Abdul; Langel, Stephanie N.; Wentworth, David E.; Stucker, Karla M.; Halpin, Rebecca A.; Lam, Ham Ching; Marthaler, Douglas

    2015-01-01

    ABSTRACT The changing epidemiology of group A rotavirus (RV) strains in humans and swine, including emerging G9 strains, poses new challenges to current vaccines. In this study, we comparatively assessed the pathogenesis of porcine RV (PRV) G9P[13] and evaluated the short-term cross-protection between this strain and human RV (HRV) Wa G1P[8] in gnotobiotic pigs. Complete genome sequencing demonstrated that PRV G9P[13] possessed a human-like G9 VP7 genotype but shared higher overall nucleotide identity with historic PRV strains. PRV G9P[13] induced longer rectal virus shedding and RV RNAemia in pigs than HRV Wa G1P[8] and generated complete short-term cross-protection in pigs challenged with HRV or PRV, whereas HRV Wa G1P[8] induced only partial protection against PRV challenge. Moreover, PRV G9P[13] replicated more extensively in porcine monocyte-derived dendritic cells (MoDCs) than did HRV Wa G1P[8]. Cross-protection was likely not dependent on serum virus-neutralizing (VN) antibodies, as the heterologous VN antibody titers in the sera of G9P[13]-inoculated pigs were low. Thus, our results suggest that heterologous protection by the current monovalent G1P[8] HRV vaccine against emerging G9 strains should be evaluated in clinical and experimental studies to prevent further dissemination of G9 strains. Differences in the pathogenesis of these two strains may be partially attributable to their variable abilities to replicate and persist in porcine immune cells, including dendritic cells (DCs). Additional studies are needed to evaluate the emerging G9 strains as potential vaccine candidates and to test the susceptibility of various immune cells to infection by G9 and other common HRV/PRV genotypes. IMPORTANCE The changing epidemiology of porcine and human group A rotaviruses (RVs), including emerging G9 strains, may compromise the efficacy of current vaccines. An understanding of the pathogenesis and genetic, immunological, and biological features of the new emerging

  9. Association of Human Immunoglobulin G1 Heavy Chain Variants With Neutralization Capacity and Antibody-Dependent Cellular Cytotoxicity Against Human Cytomegalovirus.

    PubMed

    Vietzen, Hannes; Görzer, Irene; Puchhammer-Stöckl, Elisabeth

    2016-10-15

    Human cytomegalovirus (HCMV) infection is limited by HCMV-specific antibody functions. Here the association between the genetic marker (GM) 3/17 variants in the immunoglobulin G1 (IgG1) heavy chain constant region, virus neutralization, and natural killer (NK)-cell activation was investigated. In 100 HCMV-seropositive individuals, the GM3/17 polymorphism, serum 50% HCMV antibody neutralization titer (NT50), and in vitro HCMV-specific antibody NK-cell activation were assessed. The HCMV NT50 was higher in heterozygous GM3/17 persons than in GM3/3 persons (P = .0276). Furthermore, individuals expressing GM3/17 exhibited significantly higher NK-cell activation than persons carrying GM3/3 (P < .0001) or GM17/17 (P = .0095). Thus, persons expressing GM3/17 have potentially a selective advantage in HCMV defense.

  10. Interleukin-1beta partially alleviates cyclosporin A-induced suppression of IgG1 isotype response to thyroglobulin in BALB/c mice in vivo.

    PubMed Central

    Dalai, S K; Miriyala, B; Kar, S K

    1998-01-01

    Cyclosporin A (CsA) at 120 mg/kg body weight when injected subcutaneously into BALB/c mice along with thyroglobulin emulsified in incomplete Freund's adjuvant (IFA) was found to suppress antigen-specific IgG titre by 86%. Isotyping revealed that both IgG1 and IgG2a titres were suppressed by 87% and 57%, respectively. But under identical conditions when complete Freund's adjuvant (CFA) was used, the suppression of antigen-specific IgG, IgG1 and IgG2a titres was 50%, 51% and 55%, respectively. Injection of anti-IL-1beta-neutralizing hamster monoclonal antibodies along with thyroglobulin and CsA emulsified in CFA increased the suppression of antigen-specific IgG titre. Under such conditions the IgG1 titre was suppressed more than the IgG2a titre. Recombinant human interleukin-1 receptor antagonist (rhuIL-1ra) also enhanced the suppression caused by CsA in the presence of CFA but control hamster immunoglobulin had no such effect. Recombinant human IL-1beta, when administered along with thyroglobulin and CsA emulsified in IFA, alleviated the suppression of antigen-specific IgG titre and the IgG1 titre was alleviated more than the IgG2a titre. Under identical conditions, rhuIL-1ra did not alleviate CsA-induced suppression. Lymphocytes from the lymph nodes of thyroglobulin-sensitized BALB/c mice when stimulated in vitro by thyroglobulin in the presence of CsA, secreted very little interferon-gamma (IFN-gamma) and IL-4, but on addition of an optimal dose of rhuIL-1beta, IFN-gamma and IL-4 secretion was partially restored. PMID:9767461

  11. Draft Genome Sequence of Geobacillus icigianus Strain G1w1T Isolated from Hot Springs in the Valley of Geysers, Kamchatka (Russian Federation).

    PubMed

    Bryanskaya, Alla V; Rozanov, Aleksey S; Logacheva, Maria D; Kotenko, Anastasia V; Peltek, Sergey E

    2014-10-23

    The Geobacillus icigianus G1w1(T) strain was isolated from sludge samples of unnamed vaporing hydrotherm