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Sample records for prevents cytoplasmic contents

  1. Phage shock proteins B and C prevent lethal cytoplasmic membrane permeability in Yersinia enterocolitica.

    Science.gov (United States)

    Horstman, N Kaye; Darwin, Andrew J

    2012-08-01

    The bacterial phage shock protein (Psp) stress response system is activated by events affecting the cytoplasmic membrane. In response, Psp protein levels increase, including PspA, which has been implicated as the master effector of stress tolerance. Yersinia enterocolitica and related bacteria with a defective Psp system are highly sensitive to the mislocalization of pore-forming secretin proteins. However, why secretins are toxic to psp null strains, whereas some other Psp inducers are not, has not been explained. Furthermore, previous work has led to the confounding and disputable suggestion that PspA is not involved in mitigating secretin toxicity. Here we have established a correlation between the amount of secretin toxicity in a psp null strain and the extent of cytoplasmic membrane permeability to large molecules. This leads to a morphological change resembling cells undergoing plasmolysis. Furthermore, using novel strains with dis-regulated Psp proteins has allowed us to obtain unequivocal evidence that PspA is not required for secretin-stress tolerance. Together, our data suggest that the mechanism by which secretin multimers kill psp null cells is by causing a profound defect in the cytoplasmic membrane permeability barrier. This allows lethal molecular exchange with the environment, which the PspB and PspC proteins can prevent. © 2012 Blackwell Publishing Ltd.

  2. Effects of lanthanum on calcium and magnesium contents and cytoplasmic streaming of internodal cells of Chara corallina.

    Science.gov (United States)

    Li, Zijie; Zhang, Zhiyong; Yu, Ming; Zhou, Yunlong; Zhao, Yuliang

    2011-10-01

    Biological and environmental effects of lanthanide series of elements have received much attention recently due to their wide applications. In this study, effects of La(3+) treatments on calcium and magnesium concentrations as well as cytoplasmic streaming of internodal cells of Chara corallina were investigated. At all treatment concentrations (10, 100, and 1,000 μM), La(3+) significantly decreased calcium concentrations in the cell-wall fractions after 5-h treatments. Calcium concentrations in the cell contents and magnesium concentrations in the cell-wall fractions were reduced by 100 and 1,000 μM La(3+) treatments. However, cytoplasmic streaming as an indicator of [Ca(2+)](cyt) was only inhibited at the highest La(3+) concentration (1,000 μM). The results suggest that La(3+) may affect cellular calcium homeostasis by actions other than as a simple Ca(2+) antagonist. La(3+) could partially compensate for calcium deficiency at certain concentrations.

  3. Intraspecific variation in mitochondrial genome sequence, structure, and gene content in Silene vulgaris, an angiosperm with pervasive cytoplasmic male sterility

    Czech Academy of Sciences Publication Activity Database

    Sloan, D.B.; Müller, Karel; McCauley, D.; Taylor, D.R.; Štorchová, Helena

    2012-01-01

    Roč. 196, č. 4 (2012), s. 1228-1239 ISSN 0028-646X R&D Projects: GA ČR GA521/09/0261; GA MŠk(CZ) LC06004; GA MŠk ME09035 Institutional research plan: CEZ:AV0Z50380511 Keywords : cytoplasmic male sterility (CMS) * gynodioecy * intracellular gene transfer Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 6.736, year: 2012

  4. Association of glucose-6-phosphate dehydrogenase activity with oocyte cytoplasmic lipid content, developmental competence, and expression of candidate genes in a sheep model.

    Science.gov (United States)

    Mohammadi-Sangcheshmeh, Abdollah; Veshkini, Arash; Hajarizadeh, Athena; Jamshidi-Adegani, Fatemeh; Zhandi, Mahdi; Abazari-Kia, Amir Hossein; Cinar, Mehmet Ulas; Soleimani, Masoud; Gastal, Eduardo L

    2014-08-01

    To evaluate associations of glucose-6-phosphate dehydrogenase (G6PDH) activity in sheep oocytes with cytoplasmic lipid content, maturational competence, developmental competence to the blastocyst stage, and gene expression of certain molecular markers. Before brilliant cresyl blue (BCB) staining test, oocytes were classified as high, middle, and low cytoplasmic lipid content (HCLC, MCLC, and LCLC) and after the test as having low or high G6PDH-activity (BCB(+) and BCB(-), respectively). After maturation in vitro, a group of oocytes were subjected to IVF followed by in vitro embryo culture and another group was used for evaluation of expression of candidate genes. The cleavage and blastosyst rates were lowest (P BCB(+), and higher (P BCB(+) oocytes than the BCB(-) oocytes. Our gene expression data indicated that mRNA transcript abundance of ITGB2, pZP3, BMP15, and GDF9 genes was similar between BCB oocytes groups. However, the expression of ATP1A1 was higher (P BCB(+) oocytes compared to BCB(-) oocytes. In addition, BAX transcript abundance was similar (P > 0.05) among BCB(+), BCB(-), and control groups, before and after maturation in vitro. Activity of G6PDH in sheep oocytes is highly associated with lipid content, and compared with the morphological parameters might be a more precise and objective predictor for subsequent developmental competence in vitro.

  5. Plant cytoplasm preserved by lightning.

    Science.gov (United States)

    Wang, X

    2004-10-01

    Usually only an organism with hard parts may be preserved in the fossil record. Cytoplasm, which is a physiologically active part of a plant, is rarely seen in the fossil record. Two Cretaceous plant fossils older than 100 million years with exceptional preservation of cytoplasm are reported here. Some cytoplasm is well preserved with subcellular details while other cytoplasm is highly hydrolyzed in the cortex of the same fossil even though both of preservations may be less than 2 microm away. The unique preservation pattern, sharp contrast of preservation in adjacent cells and the exceptional preservation of cytoplasm in the cortex suggest that lightning should play an important role in the preservation of cytoplasm and that cytoplasmic membranes may be more stable than the cell contents. Interpreting the preservation needs knowledge scattering in several formerly unrelated fields of science, including geophysics, botany, biophysics, cytology and microwave fixation technology. This new interpretation of fossilization will shed new light on preservation of cytoplasm and promote cytoplasm fossils from a position of rarity to a position of common research objects available for biological research. The importance of the identification of cytoplasm in fossil lies not in itself but in how much it influences the future research in paleobotany.

  6. Calcium effect on the content of DNA and NYS-stained nuclear, nucleolar and cytoplasmic proteins in cortex cells of pea (Pisum sativum L. roots treated with heavy metals

    Directory of Open Access Journals (Sweden)

    E. Stecka

    2014-01-01

    Full Text Available Using cytophotometric procedures, following Feulgen-NYS staining, the measurements of DNA and nuclear, nucleolar and cytoplasmic protein contents in cortex cells of pea roots growing for 144 h in calcium and/or heavy metals (Cd2+, Cr3+, Pb2+ presence were made. All tested metals treatment reduced the number of nuclei in 4C DNA class and induced appearance of nuclei with DNA amount below 2C, that was expressed in diminished DNA content. The level of NYS proteins in cells underwent also reduction. In lead presence protein content diminished in nucleus. On the other hand, increased amount of nuclear, nucleolar and cytoplasmic proteins was observed in material treated with cadmium while only of nucleolar protein content in chromium presence. In root cells treated with tested metals protein content in nucleus was related with ploidy level, disturbances in this relation appeared in nucleolus and mostly in cytoplasm. Calcium added to chromium and mostly to lead solutions diminished the toxic effect of these metals that was demonstrated by an increase in DNA content, although calcium alone reduced DNA amount in nucleus due to lower number of 4C nuclei accompanied by appearance of 1C and 1-2C DNA classes. Calcium in different ways affected protein content changed by metal treatment. Present in cadmium solution it caused a further reduction in protein content in nucleus, nucleolus and cytoplasm but increased nuclear and cytoplasmic protein when added to lead, and nucleolar proteins - in chromium solution. Moreover, calcium ions presence in metal solutions did not restore the relationship between ploidy level and nucleolar and cytoplasmic NYS stained proteins and it did not disturbe the relation existing in nucleus.

  7. Exploring Black College Females' Perceptions Regarding HIV Prevention Message Content.

    Science.gov (United States)

    Chandler-Coley, Rasheeta; Ross, Henry; Ozoya, Oluwatobi; Lescano, Celia; Flannigan, Timothy

    2017-02-01

    Media messages can facilitate the delivery of accurate information related to HIV and sexually transmitted infection. This study's purpose was to examine preexisting media campaigns from the iMPPACS study to assess age-, gender-, and culturally appropriate components identified by African American females who attend historically Black colleges/universities. In 3 separate focus group sessions, 31 Black female college students (M age = 20) viewed 4 vignettes and heard 3 audio-only clips, then ranked and commented on them based on perceived satisfaction with HIV prevention content and appropriateness of delivery. Conventional qualitative analysis using NVivo software was performed until saturation of content was achieved and themes derived. Six major themes emerged and were designated as (a) social media; (b) mirror image; (c) visually dynamic advertisements; (d) the real world; (e) people, place, things; and (f) HIV knowledge. Visually stimulating content (i.e., graphics) was found to be most appealing in marketing HIV prevention, with brief monologue/dialogue from scenarios that resemble daily life. Socially and culturally relevant HIV prevention messages are important to Black college female students. Participants recommended creating short audiovisual messages that encompass familiar contexts like dorm rooms and appealing graphics for HIV health promotion messages, such as emojis. Future audio-only prevention advertisements for this population should use recognizable voices (e.g., celebrities). Finally, messaging should be promoted on open and closed circuit social media platforms.

  8. Exploring Black College Females’ Perceptions Regarding HIV Prevention Message Content

    Science.gov (United States)

    Chandler, Rasheeta; Ross, Henry; Ozoya, Oluwatobi; Lescano, Celia; Flannigan, Timothy

    2017-01-01

    Media messages can facilitate delivery of accurate HIV/STI-related information. This study’s purpose was to examine pre-existing media campaigns from the iMPPACS study to assess age, gender, and culturally appropriate components identified by African American females who attend HBCUs. In three separate focus group sessions, 31 Black female college students (M age=20) viewed four vignettes and heard three audio-only clips, then ranked and commented on them based on perceived satisfaction of HIV prevention content and appropriateness of delivery. Conventional qualitative analysis, using NVivo software, was employed until saturation of content was achieved and themes derived. Six major themes emerged and were designated as: 1) Social media; 2) Mirror Image; 3) Visually dynamic Advertisements; 4) The Real World; 5) People, Place, Things; and 6) HIV Knowledge. Visually-stimulating content (i.e. graphics) was found to be most appealing in marketing HIV prevention, with brief monologue/dialogue from scenarios that resemble daily life. Socially and culturally relevant HIV prevention messages are important to Black college female students. Participants recommended creating short audio-visual messages that encompass familiar contexts like dorm rooms and appealing graphics for HIV health promotion messages, such as emojis. Future audio-only prevention advertisements for this population should employ recognizable voices (e.g. celebrity). Finally, messaging should be promoted on open and close circuited social media platforms. PMID:28098500

  9. A bandage contact lens prevents extrusion of ocular contents.

    Science.gov (United States)

    Ramjiani, Vipul; Fearnley, Thomas; Tan, Jennifer

    2016-02-01

    A 76 year old male presents with a corneal perforation in a phthisical eye. Definitive treatment in the form of an evisceration was delayed by 38 days. During this period a bandage contact lens prevented extrusion of ocular contents through an enlarging corneal perforation. This case demonstrates that a bandage contact lens can be effective in the immediate management of large corneal perforations whilst awaiting urgent definitive treatment. Copyright © 2015 Elsevier Ltd. All rights reserved.

  10. Interferometric measurements of dry mass content in nuclei and cytoplasm in the life cycle of antheridial filaments cells of Chara vulgaris L. in their successive developmental stages

    Directory of Open Access Journals (Sweden)

    Hanna Kuran

    2015-01-01

    Full Text Available Interferometric measurements of the nucleus and cytoplasm dry mass during interphase in the successive stages of development of antheridial filaments of Chara vulgaris demonstrated that the dry mass and surface area of cell nuclei double in size in each of the successive generations of the filaments, whereas neither the surface nor the dry mass of the cytoplasm increase in such proportion in the same period. In the successive stages of development of the antheridial filaments the dry mass and surface area of the nuclei and cytoplasm gradually diminish.

  11. Content and effects of news stories about uncertain cancer causes and preventive behaviors.

    Science.gov (United States)

    Niederdeppe, Jeff; Lee, Theodore; Robbins, Rebecca; Kim, Hye Kyung; Kresovich, Alex; Kirshenblat, Danielle; Standridge, Kimberly; Clarke, Christopher E; Jensen, Jakob; Fowler, Erika Franklin

    2014-01-01

    This article presents findings from two studies that describe news portrayals of cancer causes and prevention in local TV and test the effects of typical aspects of this coverage on cancer-related fatalism and overload. Study 1 analyzed the content of stories focused on cancer causes and prevention from an October 2002 national sample of local TV and newspaper cancer coverage (n = 122 television stations; n = 60 newspapers). Informed by results from the content analysis, Study 2 describes results from a randomized experiment testing effects of the volume and content of news stories about cancer causes and prevention (n = 601). Study 1 indicates that local TV news stories describe cancer causes and prevention as comparatively more certain than newspapers but include less information about how to reduce cancer risk. Study 2 reveals that the combination of stories conveying an emerging cancer cause and prevention behavior as moderately certain leads to an increased sense of overload, while a short summary of well-established preventive behaviors mitigates these potentially harmful beliefs. We conclude with a series of recommendations for health communication and health journalism practice.

  12. Cytoplasmic bacteriophage display system

    Science.gov (United States)

    Studier, F. William; Rosenberg, Alan H.

    1998-06-16

    Disclosed are display vectors comprising DNA encoding a portion of a structural protein from a cytoplasmic bacteriophage, joined covalently to a protein or peptide of interest. Exemplified are display vectors wherein the structural protein is the T7 bacteriophage capsid protein. More specifically, in the exemplified display vectors the C-terminal amino acid residue of the portion of the capsid protein is joined to the N-terminal residue of the protein or peptide of interest. The portion of the T7 capsid protein exemplified comprises an N-terminal portion corresponding to form 10B of the T7 capsid protein. The display vectors are useful for high copy number display or lower copy number display (with larger fusion). Compositions of the type described herein are useful in connection with methods for producing a virus displaying a protein or peptide of interest.

  13. Televised obesity-prevention advertising across US media markets: exposure and content, 2010-2011.

    Science.gov (United States)

    Kornfield, Rachel; Szczypka, Glen; Powell, Lisa M; Emery, Sherry L

    2015-04-01

    To examine levels of exposure and content characteristics for recent televised obesity-prevention campaigns sponsored by state and community health departments, federal agencies, non-profit organizations and television stations in the USA. Nielsen television ratings for obesity-prevention advertising were collected for the top seventy-five US media markets and were used to calculate household exposure levels for 2010 and 2011. Governmental advertisements were coded for content. United States. Average household exposure to obesity-prevention campaigns was 2·6 advertisements per month. Exposure increased by 31 % between 2010 and 2011, largely driven by increases in federal advertisements. In 2011, the federal government accounted for 62 % of obesity-prevention exposure, non-profit organizations for 9 %, community departments for 8 %, state departments for 3 %, and television station-sponsored public-service announcements for 17 %. The greatest percentage increase between 2010 and 2011 was in community advertising, reflecting efforts funded by the Communities Putting Prevention to Work (CPPW) programme. Among thirty-four state and community campaigns, the majority advocated both healthy eating and physical activity (53 %). Campaigns typically had positive or neutral emotional valence (94 %). Obesity or overweight was mentioned in 47 % of campaigns, but only 9 % specifically advocated weight loss. Exposure to televised obesity-prevention advertising increased from 2010 to 2011 and was higher than previously found in 1999-2003, apart from in 2003 during the federal VERB campaign. Nevertheless, exposure remains low relative to advertising for unhealthy foods. New federal campaigns have increased exposure to obesity-prevention advertising nationally, while CPPW grants have increased exposure for targeted areas.

  14. Content analysis of news coverage on cancer prevention and screening in Japanese newspapers.

    Science.gov (United States)

    Miyawaki, Rina; Ishii, Kaori; Shibata, Ai; Oka, Koichiro

    2017-01-01

    Objectives The present study investigated articles on cancer prevention published in Japanese newspapers in 2011.Methods A content analysis of news coverage on cancer primary prevention and screening was conducted. Articles which mentioned cancer risk, prevention, and screening were extracted. For all articles on prevention, the newspaper's name, month of publication, and information source were checked. Coding variables for articles on primary prevention included causes of human cancer, risk and/or prevention, and recommended screening criteria. Cancer screening articles were classified according to four coding variables: cancer screening site, subjects for screening, examination interval, and whether to promote the screening.Results A total of 272 articles were identified and subsequently coded as either articles on primary prevention or screening. The number of articles on primary prevention was 208. The focus of these articles was mostly on food/nutrition (n=56), cancer-causing infection (n=40), and smoking (n=32). Alcohol drinking (n=12), exercise/physical activity (n=11), and obesity (n=10), which are also major lifestyle factors for cancer, were rarely mentioned. Moreover, cancer risk was more frequently mentioned than prevention. The recommended criteria for major lifestyle factors were mentioned in 13 articles. Screening was mentioned in 92 articles. Breast cancer screening was the most frequently mentioned (n=31). The screening of colon (n=18), cervical (n=18),stomach (n=15), and lung cancer (n=10), which are recommended by the cancer control act, were mentioned in less than 20 articles. Seven articles on screening indicated subjects and interval. Only 39 articles discussed cancer-screening behaviors.Conclusions Articles on cancer prevention were found to be published in Japanese newspapers. However, the number of articles on both primary prevention and screening was not enough, and there were some disparities in the lifestyle factors and

  15. Preventing performance drops of coal mills due to high moisture content

    DEFF Research Database (Denmark)

    Odgaard, Peter Fogh; Stoustrup, Jakob; Mataji, B.

    2007-01-01

    Coal mills pulverize and dry the coal dust before it is blown into the furnace in coal-fired power plants. The coal mills can only deliver the requested coal flow if certain conditions are fulfilled. These are normally considered as constraints on individual variables. However, combinations of more...... than one variable might cause problems even though these individually variables are in an acceptable region. This paper deals with such a problem. The combination of a high load of the power plant, a large load change and high moisture content in the coal, can force the coal mill into a state where...... coal is accumulated instead of being blown into the furnace. This paper suggests a simple method for preventing the accumulation of the coal in the mill, by limiting the requested coal flow considering the coal moisture content and the temperature outside the mill.  ...

  16. Smoking Prevention in China: A Content Analysis of an Anti-Smoking Social Media Campaign.

    Science.gov (United States)

    Jiang, Shaohai; Beaudoin, Christopher E

    2016-07-01

    The China Tobacco Control Media Campaign on Sina Weibo is novel in the context of smoking prevention and cessation in China and has not to date been evaluated. This study draws on health behavior theories and dialogic theory in public relations to analyze microblog campaign postings and their relationships with the outcome of online audience engagement. Microblog postings from May 2011 to January 2015 were content analyzed, showing that the most common persuasive content characteristic was perceived risk, followed by subjective norms and self-efficacy. Perceived risk and self-efficacy postings positively influenced online audience engagement, whereas subjective norm postings was a nonsignificant predictor. Postings were more likely to share information than aim to interact with audience members. However, both information sharing and audience interaction postings were positive predictors of online audience engagement. There was also evidence of main and interactive effects of message originality on online audience engagement. The current study has, to the best of our knowledge, broken new ground in 2 regards: (a) using health behavior theories as a basis for analyzing the content of an anti-smoking social media campaign and (b) examining the content of an anti-smoking media campaign of any type in China.

  17. Cellular Subcompartments through Cytoplasmic Streaming.

    Science.gov (United States)

    Pieuchot, Laurent; Lai, Julian; Loh, Rachel Ann; Leong, Fong Yew; Chiam, Keng-Hwee; Stajich, Jason; Jedd, Gregory

    2015-08-24

    Cytoplasmic streaming occurs in diverse cell types, where it generally serves a transport function. Here, we examine streaming in multicellular fungal hyphae and identify an additional function wherein regimented streaming forms distinct cytoplasmic subcompartments. In the hypha, cytoplasm flows directionally from cell to cell through septal pores. Using live-cell imaging and computer simulations, we identify a flow pattern that produces vortices (eddies) on the upstream side of the septum. Nuclei can be immobilized in these microfluidic eddies, where they form multinucleate aggregates and accumulate foci of the HDA-2 histone deacetylase-associated factor, SPA-19. Pores experiencing flow degenerate in the absence of SPA-19, suggesting that eddy-trapped nuclei function to reinforce the septum. Together, our data show that eddies comprise a subcellular niche favoring nuclear differentiation and that subcompartments can be self-organized as a consequence of regimented cytoplasmic streaming. Copyright © 2015 Elsevier Inc. All rights reserved.

  18. Content

    DEFF Research Database (Denmark)

    Keiding, Tina Bering

    Aim, content and methods are fundamental categories of both theoretical and practical general didactics. A quick glance in recent pedagogical literature on higher education, however, reveals a strong preoccupation with methods, i.e. how teaching should be organized socially (Biggs & Tang, 2007......, is subordinating content to methods as seen in modern didactics, hereby transforming content to a medium for achievement of learning-to-learn skills rather than something valuable in its own right. At the level of general didactics quite few attempts have been made to formulate criteria and categories...... secondary levels. In subject matter didactics, the question of content is more developed, but it is still mostly confined to teaching on lower levels. As for higher education didactics, discussions on selection of content are almost non-existent on the programmatic level. Nevertheless, teachers are forced...

  19. [Fluoride content of bottled natural mineral waters in Spain and prevention of dental caries].

    Science.gov (United States)

    Maraver, Francisco; Vitoria, Isidro; Almerich-Silla, José Manuel; Armijo, Francisco

    2015-01-01

    The aim of the study was to determine the concentration of fluoride in natural mineral waters marketed in Spain in order to prevent tooth decay without the risk of causing dental fluorosis Descriptive and cross-sectional study during 2012. Natural mineral waters marketed in Spain. Three bottles with different bottling dates of 109 natural mineral waters (97 Spanish and 12 imported brands). Determination of fluoride by ion chromatography Median fluoride concentrations of the natural mineral waters bottled in Spain was 0.22 (range 0.00-4.16; interquartile range:0.37). Most samples (61 brands, 62%) contained less than 0.3mg/L. There are 19 Spanish brands with more than 0.6 mg/L. The median level in imported brands was 0.35 (range 0.10-1.21; interquartile range: 0.23). Only 28 of the 109 brands examined (25.6%) specified the fluoride content on the label. Good correlation was observed between the concentrations indicated and those determined. Fluoride concentrations in natural mineral waters showed high variation. Given the growing consumption of natural mineral waters in Spain, this type of information is important to make proper use of fluoride in the primary prevention of dental caries. Copyright © 2014 Elsevier España, S.L.U. All rights reserved.

  20. Family-based childhood obesity prevention interventions: a systematic review and quantitative content analysis.

    Science.gov (United States)

    Ash, Tayla; Agaronov, Alen; Young, Ta'Loria; Aftosmes-Tobio, Alyssa; Davison, Kirsten K

    2017-08-24

    A wide range of interventions has been implemented and tested to prevent obesity in children. Given parents' influence and control over children's energy-balance behaviors, including diet, physical activity, media use, and sleep, family interventions are a key strategy in this effort. The objective of this study was to profile the field of recent family-based childhood obesity prevention interventions by employing systematic review and quantitative content analysis methods to identify gaps in the knowledge base. Using a comprehensive search strategy, we searched the PubMed, PsycIFO, and CINAHL databases to identify eligible interventions aimed at preventing childhood obesity with an active family component published between 2008 and 2015. Characteristics of study design, behavioral domains targeted, and sample demographics were extracted from eligible articles using a comprehensive codebook. More than 90% of the 119 eligible interventions were based in the United States, Europe, or Australia. Most interventions targeted children 2-5 years of age (43%) or 6-10 years of age (35%), with few studies targeting the prenatal period (8%) or children 14-17 years of age (7%). The home (28%), primary health care (27%), and community (33%) were the most common intervention settings. Diet (90%) and physical activity (82%) were more frequently targeted in interventions than media use (55%) and sleep (20%). Only 16% of interventions targeted all four behavioral domains. In addition to studies in developing countries, racial minorities and non-traditional families were also underrepresented. Hispanic/Latino and families of low socioeconomic status were highly represented. The limited number of interventions targeting diverse populations and obesity risk behaviors beyond diet and physical activity inhibit the development of comprehensive, tailored interventions. To ensure a broad evidence base, more interventions implemented in developing countries and targeting racial

  1. Program content of teaching physical education in order to prevent violence in educational institutions

    Directory of Open Access Journals (Sweden)

    Sretenović Zoran

    2013-01-01

    Full Text Available The primary goal of this paper is to show the capacity of the program content of physical education in the implementation of the third specific objective of the Framework Action Plan for the Prevention of Violence in Educational Institutions. Ministry of Education brought this plan in January 2009. The third specific objective of the Framework Action Plan: Provide preventive and protective role of school sport and sporting activities of students. This paper examines the extent to which this goal is achieved, trough regular classes, after-school and extracurricular activities of physical education; indicates the capacity of the teaching field in the creation and development of a safe and supportive environment in the institution; sees level of participation by experts, council and teams in the design and implementation of sports activities in school and the extent to which these activities are implemented in the institution documents, plans and programs of experts, and larger teams; it is the sustainability of these activities achieved in school practices and also to strengthen the competence of teachers in supporting the development of students personality, communication and collaboration. In order to complete document association of council, experts and teams in the institution, there has been an insight into the school curriculum, the development plan, annual plan of the institution, protection program community of students, the curriculum of a school parliament, council of parents, higher vocational and physical education teachers and report on the implementation of the protection program. In order to document by empirical data the capacity of the teaching areas in the function of preventing violence, a sample of 46 primary schools in Sumadija area, evaluation of the implementation of sports and sports activities students in programming classes of physical education, was executed. By these results, it can be concluded that in teaching methods

  2. Analysis of embryo, cytoplasmic and maternal genetic correlations ...

    Indian Academy of Sciences (India)

    maintenance, reproduction and immunity in farm animals. (Wu 2009). An accurate ... Keywords. genetic main correlations; genotype × environment interaction correlations; rapeseed meal; amino acids contents; nutrient quality. Journal of ... cytoplasmic effects, maternal additive and dominance effects for individual amino ...

  3. Contents

    Directory of Open Access Journals (Sweden)

    Editor IJRED

    2012-11-01

    Full Text Available International Journal of Renewable Energy Development www.ijred.com Volume 1             Number 3            October 2012                ISSN 2252- 4940   CONTENTS OF ARTICLES page Design and Economic Analysis of a Photovoltaic System: A Case Study 65-73 C.O.C. Oko , E.O. Diemuodeke, N.F. Omunakwe, and E. Nnamdi     Development of Formaldehyde Adsorption using Modified Activated Carbon – A Review 75-80 W.D.P Rengga , M. Sudibandriyo and M. Nasikin     Process Optimization for Ethyl Ester Production in Fixed Bed Reactor Using Calcium Oxide Impregnated Palm Shell Activated Carbon (CaO/PSAC 81-86 A. Buasri , B. Ksapabutr, M. Panapoy and N. Chaiyut     Wind Resource Assessment in Abadan Airport in Iran 87-97 Mojtaba Nedaei       The Energy Processing by Power Electronics and its Impact on Power Quality 99-105 J. E. Rocha and B. W. D. C. Sanchez       First Aspect of Conventional Power System Assessment for High Wind Power Plants Penetration 107-113 A. Merzic , M. Music, and M. Rascic   Experimental Study on the Production of Karanja Oil Methyl Ester and Its Effect on Diesel Engine 115-122 N. Shrivastava,  , S.N. Varma and M. Pandey  

  4. Regulation of Cytoplasmic and Vacuolar Volumes by Plant Cells in Suspension Culture

    DEFF Research Database (Denmark)

    Owens, Trevor; Poole, Ronald J

    1979-01-01

    Quantitative microscopical measurements have been made of the proportion of cell volume occupied by cytoplasm in a cell suspension culture derived from cotyledons of bush bean (cv. Contender). On a 7-day culture cycle, the content of cytoplasm varies from 25% at the time of transfer to 45...

  5. Regulation of Cytoplasmic and Vacuolar Volumes by Plant Cells in Suspension Culture

    DEFF Research Database (Denmark)

    Owens, Trevor; Poole, Ronald J

    1979-01-01

    by cytoplasm is roughly correlated with protein content, but shows no correlation with cell size or with intracellular concentrations of K or Na. The most striking observation is that the growth of cytoplasmic volume for the culture as a whole appears to be constant throughout the culture cycle, despite...

  6. Diabetes prevention information in Japanese magazines with the largest print runs. Content analysis using clinical guidelines as a standard.

    Science.gov (United States)

    Noda, Emi; Mifune, Taka; Nakayama, Takeo

    2013-01-01

    To characterize information on diabetes prevention appearing in Japanese general health magazines and to examine the agreement of the content with that in clinical practice guidelines for the treatment of diabetes in Japan. We used the Japanese magazines' databases provided by the Media Research Center and selected magazines with large print runs published in 2006. Two medical professionals independently conducted content analysis based on items in the diabetes prevention guidelines. The number of pages for each item and agreement with the information in the guidelines were determined. We found 63 issues of magazines amounting to 8,982 pages; 484 pages included diabetes prevention related content. For 23 items included in the diabetes prevention guidelines, overall agreement of information printed in the magazines with that in the guidelines was 64.5% (471 out of 730). The number of times these items were referred to in the magazines varied widely, from 247 times for food items to 0 times for items on screening for pregnancy-induced diabetes, dyslipidemia, and hypertension. Among the 20 items that were referred to at least once, 18 items showed more than 90% agreement with the guidelines. However, there was poor agreement for information on vegetable oil (2/14, 14%) and for specific foods (5/247, 2%). For the fatty acids category, "fat" was not mentioned in the guidelines; however, the term frequently appeared in magazines. "Uncertainty" was never mentioned in magazines for specific food items. The diabetes prevention related content in the health magazines differed from that defined in clinical practice guidelines. Most information in the magazines agreed with the guidelines, however some items were referred to inappropriately. To disseminate correct information to the public on diabetes prevention, health professionals and the media must collaborate.

  7. Three fire prevention television films varying in "threat" content .... their effectiveness in changing attitudes

    Science.gov (United States)

    Gene C. Bernardi

    1970-01-01

    The campaign to disseminate fire prevention information has for some years relied heavily on a belief in the efficacy of television. Consequently, numerous public service films on fire prevention have been produced and beamed over commercial television channels in the hopes of achieving a reduction in the number of man-caused forest fires. Television continues to be...

  8. School-based suicide prevention: content, process, and the role of trusted adults and peers.

    Science.gov (United States)

    Joshi, Shashank V; Hartley, Samantha N; Kessler, Moira; Barstead, Maura

    2015-04-01

    Suicide is a leading cause of preventable death in youth, and numerous curricula and other prevention and intervention programs have been developed in the last 15 years. Comprehensive suicide prevention planning should include the 4 components of health promotion, prevention/education, intervention, and postvention. School-based suicide prevention and mental health education programs have become more common as an efficient and cost-effective way to reach youth. Process considerations that are based on the principles of therapeutic engagement with patients and families can provide mental health professionals with strategies that can assist education professionals, students, and the larger school community simultaneously. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. A perspective on the mobilization, localization and delivery of molecules in the crowded bacterial cytoplasm.

    Science.gov (United States)

    Trevors, J T

    2012-01-01

    It has been assumed that diffusion of molecules in the bacterial cytoplasm is the mechanism that moves molecules in the absence of cytoplasmic streaming. However, is there an undiscovered mechanism present that mobilizes cytoplasm and its molecular contents, and delivers tRNAs to specific ribosomes at specific bacterial cytoplasmic locations? Mobilization of specific tRNA (and also mRNA transcripts and ribosomes) and cell division proteins to specific intracellular locations may suggest that instructions and/or mechanism(s) are needed. The alternative is that molecular crowding in the cytoplasm is sufficient for gentle contact between mRNA, ribosomes and tRNA. Or is it plausible that the bacterial cytoplasm (and its contents) are mobilized with the outcome being more gentle collisions between molecules than by a diffusion only mechanism? One hypothesis is that cytoplasmic and molecule mobilization and spatial organization are possibly driven by the photons in thermal infrared (IR) radiation and generation of exclusion zone (EZ) water in the cytoplasm.

  10. Bridging the gap between content and context: establishing expert consensus on the content of an exercise training program to prevent lower-limb injuries.

    Science.gov (United States)

    Donaldson, Alex; Cook, Jill; Gabbe, Belinda; Lloyd, David G; Young, Warren; Finch, Caroline F

    2015-05-01

    To achieve expert consensus on the content of an exercise training program (known as FootyFirst) to prevent lower-limb injuries. Three-round online Delphi consultation process. Community Australian Football (AF). Members of the Australian Football Leagues' Medical Officers (n = 94), physiotherapists (n = 50), and Sports Science (n = 19) Associations were invited to participate through e-mail. Five people with more general expertise in sports-related lower-limb injury prevention were also invited to participate. The primary outcome measure was the level of agreement on the appropriateness of the proposed exercises and progressions for inclusion in FootyFirst. Consensus was reached when ≥75% of experts who responded to each item agreed and strongly agreed, or disagreed and strongly disagreed, that an exercise or its progressions were appropriate to include in FootyFirst. Fifty-five experts participated in at least 1 Delphi round. In round 1, consensus was achieved that the proposed warm-up (run through and dynamic stretches) and the exercises and progressions for hamstring strength and for balance, landing, and changing direction were appropriate to include in FootyFirst. There was also consensus in round 1 that progressions for hip/core strength should be included in FootyFirst. Consensus was reached in round 2 that the revised groin strength and hip strength exercises should be included in FootyFirst. Consensus was reached for the progression of the groin strength exercises in round 3. The formal consensus development process has resulted in an evidence-informed, researcher-developed, exercise-based sports injury prevention program that is expert endorsed and specific to the context of AF. Lower-limb injuries are common in running, kicking, and contact sports like AF. These injuries are often costly to treat, and many have high rates of recurrence, making them challenging to treat clinically. Reducing these injuries is a high priority for players, teams, and

  11. Mobile Phone Apps for the Prevention of Unintended Pregnancy: A Systematic Review and Content Analysis.

    Science.gov (United States)

    Mangone, Emily Rose; Lebrun, Victoria; Muessig, Kathryn E

    2016-01-19

    Over 50% of pregnancies in the United States are unintended, meaning that the pregnancy is mistimed, unplanned, or unwanted. Unintended pregnancy increases health risks for mother and child, leads to high economic costs for society, and increases social disparities. Mobile phone ownership is rapidly increasing, providing opportunities to reach at-risk populations with reproductive health information and tailored unintended pregnancy prevention interventions through mobile phone apps. However, apps that offer support for unintended pregnancy prevention remain unevaluated. To identify, describe, and evaluate mobile phone apps that purport to help users prevent unintended pregnancy. We conducted an extensive search of the Apple iTunes and Android Google Play stores for apps that explicitly included or advertised pregnancy prevention or decision-making support in the context of fertility information/tracking, birth control reminders, contraceptive information, pregnancy decision-making, abortion information or counseling, sexual communication/negotiation, and pregnancy tests. We excluded apps that targeted medical professionals or that cost more than US $1.99. Eligible apps were downloaded and categorized by primary purpose. Data extraction was performed on a minimum of 143 attributes in 3 domains: (1) pregnancy prevention best practices, (2) contraceptive methods and clinical services, and (3) user interface. Apps were assigned points for their inclusion of features overall and for pregnancy prevention best practices and contraceptive information. Our search identified 6805 app descriptions in iTunes and Google Play. Of these, 218 unique apps met inclusion criteria and were included in the review. Apps were grouped into 9 categories: fertility trackers (n=72), centers and resources (n=38), birth control reminders (n=35), general sexual and reproductive health (SRH) information (n=17), SRH information targeted specifically to young adults (YA) (n=16), contraceptive

  12. Curious Sex Ratios and Cytoplasmic Genes

    Indian Academy of Sciences (India)

    formation of sperm from mat,!re male germline cells most cytoplasm is lost, including al/ endosym- bionts. In contrast, when eggs are formed from mature female germ line cells, they retain a signi- ficant amount of cytoplasm, and this is likely to contain endosymbionts. As a con- sequence, individuals appearing at each new.

  13. Cytoplasmic Streaming - Skylab Student Experiment ED-63

    Science.gov (United States)

    1973-01-01

    This chart describes the Skylab student experiment (ED-63), Cytoplasmic Streaming, proposed by Cheryl A. Peitz of Arapahoe High School, Littleton, Colorado. Experiment ED-63 was to observe the effect of zero-gravity on cytoplasmic streaming in the aquatic plant named Elodea, commonly called water weed or water thyme. The phenomenon of cytoplasmic streaming is not well understood, but it is recognized as the circulation mechanism of the internal materials or cytoplasm of a cell. Cytoplasm is a gelatinous substance that has the ability to change its viscosity and flow, carrying various cell materials with it. The activity can be stimulated by sunlight or heat. In March 1972, NASA and the National Science Teachers Association selected 25 experiment proposals for flight on Skylab. Science advisors from the Marshall Space Flight Center aided and assisted the students in developing the proposals for flight on Skylab.

  14. Characterization of cytoplasmic male sterility of rice with Lead Rice cytoplasm in comparison with that with Chinsurah Boro II cytoplasm.

    Science.gov (United States)

    Itabashi, Etsuko; Kazama, Tomohiko; Toriyama, Kinya

    2009-02-01

    Rice with LD-type cytoplasmic male sterility (CMS) possesses the cytoplasm of 'Lead Rice' and its fertility is recovered by a nuclear fertility restorer gene Rf1. Rf1 promotes processing of a CMS-associated mitochondrial RNA of atp6-orf79, which consists of atp6 and orf79, in BT-CMS with the cytoplasm of 'Chinsurah Boro II'. In this study, we found that LD-cytoplasm contained a sequence variant of orf79 downstream of atp6. Northern blot analysis showed that atp6-orf79 RNA of LD-cytoplasm was co-transcribed and was processed in the presence of Rf1 in the same manner as in BT-cytoplasm. Western blot analysis showed that the ORF79 peptide did not accumulate in an LD-CMS line, while ORF79 accumulated in a BT-CMS line and was diminished by Rf1. These results suggest that accumulation of ORF79 is not the cause of CMS in LD-cytoplasm and the mechanism of male-sterility induction/fertility restoration in LD-CMS is different from that in BT-CMS.

  15. Mobile Phone Apps for the Prevention of Unintended Pregnancy: A Systematic Review and Content Analysis

    OpenAIRE

    Mangone, Emily Rose; Lebrun, Victoria; Muessig, Kathryn E

    2016-01-01

    Background Over 50% of pregnancies in the United States are unintended, meaning that the pregnancy is mistimed, unplanned, or unwanted. Unintended pregnancy increases health risks for mother and child, leads to high economic costs for society, and increases social disparities. Mobile phone ownership is rapidly increasing, providing opportunities to reach at-risk populations with reproductive health information and tailored unintended pregnancy prevention interventions through mobile phone app...

  16. Comparison of child obesity prevention and control content in mainstream and Spanish-language US parenting magazines.

    Science.gov (United States)

    Kalin, Sari R; Fung, Teresa T

    2013-01-01

    Mass media coverage of child obesity is rising, paralleling the child obesity epidemic's growth, and there is evidence that parents seek parenting advice from media sources. Yet little to no research has examined the coverage of child obesity in parenting magazines or Spanish-language media. The purpose of this study was to use qualitative and quantitative content analysis methods to identify, quantify, and compare strategies for child obesity prevention and control presented in mainstream and Spanish-language US parenting magazines. Child obesity-related editorial content in 68 mainstream and 20 Spanish-language magazine issues published over 32 months was gathered. Magazine content was coded with a manual developed by refining themes from the sample and from an evidence-based child obesity prevention action plan. Seventy-three articles related to child obesity prevention and control were identified. Most focused on parental behavior change rather than environmental change, and only 3 in 10 articles referred to the social context in which parental behavior change takes place. Child obesity-focused articles were not given high prominence; only one in four articles in the entire sample referred to child obesity as a growing problem or epidemic. Key differences between genres reflect culturally important Latino themes, including family focus and changing health beliefs around child weight status. Given mass media's potential influence on parenting practices and public perceptions, nutrition communication professionals and registered dietitians need to work to reframe media coverage of childhood obesity as an environmental problem that requires broad-based policy solutions. Spanish-speaking media can be an ally in helping Latina women change cultural health beliefs around child weight status. Copyright © 2013 Academy of Nutrition and Dietetics. Published by Elsevier Inc. All rights reserved.

  17. Hydrodynamic property of the cytoplasm is sufficient to mediate cytoplasmic streaming in the Caenorhabiditis elegans embryo

    Science.gov (United States)

    Niwayama, Ritsuya; Shinohara, Kyosuke; Kimura, Akatsuki

    2011-01-01

    Cytoplasmic streaming is a type of intracellular transport widely seen in nature. Cytoplasmic streaming in Caenorhabditis elegans at the one-cell stage is bidirectional; the flow near the cortex (“cortical flow”) is oriented toward the anterior, whereas the flow in the central region (“cytoplasmic flow”) is oriented toward the posterior. Both cortical flow and cytoplasmic flow depend on non-muscle-myosin II (NMY-2), which primarily localizes in the cortex. The manner in which NMY-2 proteins drive cytoplasmic flow in the opposite direction from remote locations has not been fully understood. In this study, we demonstrated that the hydrodynamic properties of the cytoplasm are sufficient to mediate the forces generated by the cortical myosin to drive bidirectional streaming throughout the cytoplasm. We quantified the flow velocities of cytoplasmic streaming using particle image velocimetry (PIV) and conducted a three-dimensional hydrodynamic simulation using the moving particle semiimplicit method. Our simulation quantitatively reconstructed the quantified flow velocity distribution resolved through PIV analysis. Furthermore, our PIV analyses detected microtubule-dependent flows during the pronuclear migration stage. These flows were reproduced via hydrodynamic interactions between moving pronuclei and the cytoplasm. The agreement of flow dynamics in vivo and in simulation indicates that the hydrodynamic properties of the cytoplasm are sufficient to mediate cytoplasmic streaming in C. elegans embryos. PMID:21730185

  18. Magnesium Sulfate Prevents Placental Ischemia-Induced Increases in Brain Water Content and Cerebrospinal Fluid Cytokines in Pregnant Rats

    Science.gov (United States)

    Zhang, Linda W.; Warrington, Junie P.

    2016-01-01

    Magnesium sulfate (MgSO4) is the most widely used therapy in the clinic to prevent the progression of preeclampsia, a hypertensive disorder of pregnancy, to eclampsia. Eclampsia, manifested as unexplained seizures and/or coma during pregnancy or postpartum, accounts for ~13% of maternal deaths worldwide. While MgSO4 continues to be used in the clinic, the mechanisms by which it exerts its protective actions are not well understood. In this study, we tested the hypothesis that MgSO4 protects against placental ischemia-induced increases in brain water content and cerebrospinal fluid cytokines. To test this hypothesis, MgSO4 was administered via mini-osmotic pump (60 mg/day, i.p.) to pregnant and placental ischemic rats, induced by mechanical reduction of uterine perfusion pressure, from gestational day 14–19. This treatment regimen of MgSO4 led to therapeutic level of 2.8 ± 0.6 mmol/L Mg in plasma. MgSO4 had no effect on improving placental ischemia-induced changes in mean arterial pressure, number of live fetuses, or fetal and placental weight. Placental ischemia increased, while MgSO4 prevented the increase in water content in the anterior cerebrum. Cytokine and chemokine levels were measured in the cerebrospinal fluid using a multi-plex assay. Results demonstrate that cerebrospinal fluid, obtained via the cisterna magna, had reduced protein, albumin, interleukin (IL)-17A, IL-18, IL-2, eotaxin, fractalkine, interferon gamma, vascular endothelial growth factor (VEGF), and macrophage inflammatory protein (MIP)-2 following MgSO4 treatment. These data support the hypothesis that MgSO4 offers neuroprotection by preventing placental ischemia-induced cerebral edema and reducing levels of cytokines/chemokines in the cerebrospinal fluid. PMID:28008305

  19. Fractal organization of feline oocyte cytoplasm

    Directory of Open Access Journals (Sweden)

    G De Vico

    2009-06-01

    Full Text Available The present study aimed at verifying whether immature cat oocytes with morphologic irregular cytoplasm display selfsimilar features which can be analytically described by fractal analysis. Original images of oocytes collected by ovariectomy were acquired at a final magnification of 400 X with a CCD video camera connected to an optic microscope. After greyscale thresholding segmentation of cytoplasm, image profiles were submitted to fractal analysis using FANAL++, a program which provided an analytical standard procedure for determining the fractal dimension (FD. The presentation of the oocyte influenced the magnitude of the fractal dimension with the highest FD of 1.91 measured on grey-dark cytoplasm characterized by a highly connected network of lipid droplets and intracellular membranes. Fractal analysis provides an effective quantitative descriptor of the real cytoplasm morphology, which can influence the acquirement of in vitro developmental competence, without introducing any bias or shape approximation and thus contributes to an objective and reliable classification of feline oocytes.

  20. Cytoplasmic-anti-neutrophil cytoplasmic antibodies targeting myeloperoxidase in Wegener′s granulomatosis: A rare phenomenon

    Directory of Open Access Journals (Sweden)

    Bhavana M Venkatesh

    2014-01-01

    Full Text Available Wegener′s granulomatosis (WG patients can rarely have antineutrophil cytoplasmic antibodies (ANCAs directed against myeloperoxidase (MPO, producing a cytoplasmic pattern on indirect immunofluorescence (IIF. This has important implications in the diagnosis and pathophysiology of the disease. We present to you a report of three cases of WG, demonstrating a cytoplasmic-ANCA pattern on indirect IIF, but directed against MPO. It is necessary to diagnose a patient taking into account both the autoimmune test results and the clinical features.

  1. Prevention

    Science.gov (United States)

    ... Error processing SSI file About Heart Disease & Stroke Prevention Heart disease and stroke are an epidemic in ... secondhand smoke. Barriers to Effective Heart Disease & Stroke Prevention Many people with key risk factors for heart ...

  2. Intermittent At-Home Suctioning of Esophageal Content for Prevention of Recurrent Aspiration Pneumonia in 4 Dogs with Megaesophagus.

    Science.gov (United States)

    Manning, K; Birkenheuer, A J; Briley, J; Montgomery, S A; Harris, J; Vanone, S L; Gookin, J L

    2016-09-01

    Megaesophagus carries a poor to guarded prognosis due to death from aspiration pneumonia. Options for medical management of regurgitation are limited to strategic oral or gastrostomy tube feeding. To describe the use and efficacy of intermittent esophageal suctioning to prevent regurgitation and associated episodes of aspiration pneumonia in dogs with megaesophagus. Four dogs with acquired idiopathic megaesophagus and recurrent aspiration pneumonia. Retrospective review of medical records of dogs with megaesophagus in which intermittent suctioning of esophageal content was employed for management of recurrent aspiration pneumonia. Intermittent suctioning of the esophagus was initiated in 4 dogs after failure of strict gastrostomy tube feeding failed to prevent regurgitation and repeated episodes of aspiration pneumonia. Suctioning was accomplished by esophagostomy tube in 3 dogs and per os in 1 dog. After initiation of esophageal suctioning, dogs survived for a median of 13.5 additional months (range, 10-30 months) during which time 2 dogs had no additional episodes of aspiration pneumonia and 2 dogs had infrequent episodes of pneumonia, but aspiration was suspected to be a contributing factor in their death. Complications included clogging of the esophagostomy tube, esophagostomy site infections, and esophagitis. Use of intermittent esophageal suctioning in dogs with megaesophagus that continue to regurgitate despite gastrostomy tube feedings can reduce or abolish clinical episodes of aspiration pneumonia. Copyright © 2016 The Authors. Journal of Veterinary Internal Medicine published by Wiley Periodicals, Inc. on behalf of the American College of Veterinary Internal Medicine.

  3. The use of Fagopyrum tataricum gaertn. whole flour to confer preventive contents of rutin to some traditional tuscany biscuits

    Directory of Open Access Journals (Sweden)

    Andrea BRUNORI

    2010-08-01

    Full Text Available To meet the growing interest for new foods that may be regarded as functional aliments, of particular interest appears the utilization of the grain of Fagopyrum tataricum. The high content of rutin available in the grain and whole flour of this species, in fact, offers the opportunity to introduce in the food recipes effective amounts of this bioactive compound. Rutin is credited with a growing multiplicity of health beneficial properties that can be reasonablysecured through the preventive nutrition approach. In this respect, the daily dose of rutin suggested by most dietary supplement preparations is around 50 mg/day. The use of F. tataricum whole flour, where the rutin content usually ranges between 1000 up to 2000 mg/100 g dry weight, allows to reach such an amount with a low percentage introduction of this ingredient in the original recipe. Higher contents of rutin are found in the herb of cultivated species of buckwheat (F. tataricum and F. esculentum. However, the handling of this material may not be as simple as that of the grain made into whole flour. Preliminary results would indicate that an intake of 50 mg of rutin with a single meal (breakfast in this case can be feasible by adding tartary buckwheat whole flour to some traditional Tuscany biscuits without impairing texture, taste and acceptability.In spite of the presence of rutin degrading enzymes, known to be present in tartary buckwheat grain, it would appear that just a negligible degradation of rutin to quercetin occurs during the phase of dough preparation and backing process so that most of the rutin present in the whole flour can be recovered in the biscuits.

  4. Cytoplasmic Streaming in the Drosophila Oocyte.

    Science.gov (United States)

    Quinlan, Margot E

    2016-10-06

    Objects are commonly moved within the cell by either passive diffusion or active directed transport. A third possibility is advection, in which objects within the cytoplasm are moved with the flow of the cytoplasm. Bulk movement of the cytoplasm, or streaming, as required for advection, is more common in large cells than in small cells. For example, streaming is observed in elongated plant cells and the oocytes of several species. In the Drosophila oocyte, two stages of streaming are observed: relatively slow streaming during mid-oogenesis and streaming that is approximately ten times faster during late oogenesis. These flows are implicated in two processes: polarity establishment and mixing. In this review, I discuss the underlying mechanism of streaming, how slow and fast streaming are differentiated, and what we know about the physiological roles of the two types of streaming.

  5. Curious Sex Ratios and Cytoplasmic Genes

    Indian Academy of Sciences (India)

    Home; Journals; Resonance – Journal of Science Education; Volume 2; Issue 6. Curious Sex Ratios and Cytoplasmic Genes Microbes Can Distort the Sex Ratio of Populations. Stephen J Freeland Laurence D Hurst. General Article Volume 2 Issue 6 June 1997 pp 68-78 ...

  6. Detection of antineutrophil cytoplasmic antibodies (ANCAs)

    DEFF Research Database (Denmark)

    Damoiseaux, Jan; Csernok, Elena; Rasmussen, Niels

    2017-01-01

    of diagnosis) from 251 patients with ANCA-associated vasculitis (AAV), including granulomatosis with polyangiitis and microscopic polyangiitis, and from 924 disease controls were tested for the presence of cytoplasmic pattern/perinuclear pattern and atypical ANCA (A-ANCA) by indirect immunofluorescence (IIF...

  7. How crowded is the prokaryotic cytoplasm?

    NARCIS (Netherlands)

    Spitzer, Jan; Poolman, Bert; Ferguson, Stuart

    2013-01-01

    We consider biomacromolecular crowding within the cytoplasm of prokaryotic cells as a two-phase system of 'supercrowded' cytogel and 'dilute' cytosol; we simplify and quantify this model for a coccoid cell over a wide range of biomacromolecular crowding. The key result shows that the supercrowded

  8. Content Validation and Semantic Evaluation of a Check-List Elaborated for the Prevention of Gluten Cross-Contamination in Food Services

    OpenAIRE

    Farage, Priscila; Puppin Zandonadi, Renata; Cortez Ginani, Ver?nica; Gandolfi, Lenora; Pratesi, Riccardo; de Medeiros N?brega, Yanna Karla

    2017-01-01

    Conditions associated to the consumption of gluten have emerged as a major health care concern and the treatment consists on a lifelong gluten-free diet. Providing safe food for these individuals includes adapting to safety procedures within the food chain and preventing gluten cross-contamination in gluten-free food. However, a gluten cross-contamination prevention protocol or check-list has not yet been validated. Therefore, the aim of this study was to perform the content validation and se...

  9. Prevention

    Science.gov (United States)

    ... Contact Aging & Health A to Z Find a Geriatrics Healthcare Professional Medications & Older Adults Making Your Wishes ... Prevention Hearing Loss Heart Attack High Blood Pressure Nutrition Osteoporosis Shingles Skin Cancer Related News Quitting Smoking, ...

  10. Arrest of cytoplasmic streaming induces algal proliferation in green paramecia.

    Directory of Open Access Journals (Sweden)

    Toshiyuki Takahashi

    Full Text Available A green ciliate Paramecium bursaria, bearing several hundreds of endosymbiotic algae, demonstrates rotational microtubule-based cytoplasmic streaming, in which cytoplasmic granules and endosymbiotic algae flow in a constant direction. However, its physiological significance is still unknown. We investigated physiological roles of cytoplasmic streaming in P. bursaria through host cell cycle using video-microscopy. Here, we found that cytoplasmic streaming was arrested in dividing green paramecia and the endosymbiotic algae proliferated only during the arrest of cytoplasmic streaming. Interestingly, arrest of cytoplasmic streaming with pressure or a microtubule drug also induced proliferation of endosymbiotic algae independently of host cell cycle. Thus, cytoplasmic streaming may control the algal proliferation in P. bursaria. Furthermore, confocal microscopic observation revealed that a division septum was formed in the constricted area of a dividing paramecium, producing arrest of cytoplasmic streaming. This is a first report to suggest that cytoplasmic streaming controls proliferation of eukaryotic cells.

  11. Prevention

    DEFF Research Database (Denmark)

    Halken, S; Høst, A

    2001-01-01

    , breastfeeding should be encouraged for 4-6 months. In high-risk infants a documented extensively hydrolysed formula is recommended if exclusive breastfeeding is not possible for the first 4 months of life. There is no evidence for preventive dietary intervention neither during pregnancy nor lactation...... populations. These theories remain to be documented in proper, controlled and prospective studies. Breastfeeding and the late introduction of solid foods (>4 months) is associated with a reduced risk of food allergy, atopic dermatitis, and recurrent wheezing and asthma in early childhood. In all infants....... Preventive dietary restrictions after the age of 4-6 months are not scientifically documented....

  12. Association of Protein Intake with Bone Mineral Density and Bone Mineral Content among Elderly Women: The OSTPRE Fracture Prevention Study.

    Science.gov (United States)

    Isanejad, M; Sirola, J; Mursu, J; Kröger, H; Tuppurainen, M; Erkkilä, A T

    2017-01-01

    It has been hypothesized that high protein intakes are associated with lower bone mineral content (BMC). Previous studies yield conflicting results and thus far no studies have undertaken the interaction of body mass index (BMI) and physical activity with protein intakes in relation to BMC and bone mineral density (BMD). To evaluate the associations of dietary total protein (TP), animal protein (AP) and plant protein (PP) intakes with BMC and BMD and their changes. We tested also the interactions of protein intake with, obesity (BMI ≤30 vs. >30 kg/m2) and physical activity level (passive vs. active). Design/ Setting: Prospective cohort study (Osteoporosis Risk-Factor and Fracture-Prevention Study). Participants/measures: At the baseline, 554 women aged 65-72 years filled out a 3-day food record and a questionnaire covering data on lifestyle, physical activity, diseases, and medications. Intervention group received calcium 1000 mg/d and cholecalciferol 800 IU for 3 years. Control group received neither supplementation nor placebo. Bone density was measured at baseline and year 3, using dual energy x-ray absorptiometry. Multivariable regression analyses were conducted to examine the associations between protein intake and BMD and BMC. In cross-sectional analyses energy-adjusted TP (P≤0·029) and AP (P≤0·045) but not PP (g/d) were negatively associated with femoral neck (FN) BMD and BMC. Women with TP≥1·2 g/kg/body weight (BW) (Ptrend≤0·009) had lower FN, lumbar spine (LS) and total BMD and BMC. In follow-up analysis, TP (g/kg/BW) was inversely associated with LS BMD and LS BMC. The detrimental associations were stronger in women with BMI30 kg/m2 and physical activity.

  13. Cytoplasmic lipid bodies of human neutrophilic leukocytes

    International Nuclear Information System (INIS)

    Weller, P.F.; Ackerman, S.J.; Nicholson-Weller, A.; Dvorak, A.M.

    1989-01-01

    The morphology and function of cytoplasmic lipid bodies in human neutrophils were evaluated. By transmission electron microscopy, neutrophil lipid bodies were cytoplasmic inclusions, usually several microns in diameter, that occasionally coalesced to attain a diameter up to 7 microM. Neutrophil lipid bodies were not enveloped by membrane but were often surrounded by a more electron-dense shell at their periphery. Normal peripheral blood neutrophils contained an average of approximately one lipid body per cell. Lipid bodies appeared in greater numbers in neutrophils from inflammatory lesions. Perturbation of neutrophils during conventional methods of cell isolation and purification modestly increased lipid body numbers in neutrophils, whereas incubation of neutrophils with 1 microM oleic acid rapidly induced lipid body formation over 30 to 60 minutes. After granulocytes were incubated for 2 hours with 3H-fatty acids, including arachidonic, oleic, and palmitic acids, electron microscopic autoradiography demonstrated that lipid bodies represented the predominant intracellular sites of localization of each of the three 3H-fatty acids. There was lesser labeling noted in the perinuclear cisterna, but not in cell membranes. Virtually all of each of the three 3H-fatty acids incorporated by the neutrophils were esterified into chromatographically resolved classes of neutral lipids or phospholipids. These findings indicate that cytoplasmic lipid bodies are more prominent in neutrophils in vivo engaged in inflammatory responses and that these organelles in human neutrophils function as sites of deposition of esterified, incorporated fatty acids

  14. Anti-neutrophil cytoplasm autoantibodies (ANCA) in autoimmune liver diseases

    NARCIS (Netherlands)

    Roozendaal, C.; Kallenberg, Cees

    1999-01-01

    Anti-neutrophil cytoplasm antibodies (ANCA) are autoantibodies directed against cytoplasmic constituents of neutrophil granulocytes and monocytes. ANCA have been detected in serum from patients with inflammatory bowel diseases (mainly ulcerative colitis) and autoimmune mediated liver diseases

  15. Serine phosphorylation of syndecan-2 proteoglycan cytoplasmic domain

    DEFF Research Database (Denmark)

    Oh, E S; Couchman, J R; Woods, A

    1997-01-01

    sequence. We investigated phosphorylation of syndecan-2 cytoplasmic domain by PKC, using purified GST-syndecan-2 fusion proteins and synthetic peptides corresponding to regions of the cytoplasmic domain. A synthetic peptide encompassing the entire cytoplasmic domain of syndecan-2 was phosphorylated by PKC...

  16. Cytoplasm-to-myonucleus ratios following microgravity.

    Science.gov (United States)

    Kasper, C E; Xun, L

    1996-10-01

    The cytoplasmic volume-to-myonucleus ratio in the tibialis anterior and gastrocnemius muscles of juvenile rats after 5.4 days of microgravity was studied. Three groups of rats (n = 8 each) were used. The experimental group (space rats) was flown aboard the space shuttle Discovery (NASA, STS-48), while two ground-based groups, one hindlimb suspended (suspended rats), one non-suspended (control), served as controls. Single fibre analysis revealed a significant decrease in cross-sectional area (microns2) in the gastrocnemius for both the space and the suspended rats; in the tibialis anterior only the suspended rats showed a significant decrease. Myonuclei counts (myonuclei per mm) in both the tibialis anterior and gastrocnemius were significantly increased in the space rats but not in the suspended rats. The mean myonuclear volume (individual nuclei: microns3) in tibialis anterior fibres from the space rats, and in gastrocnemius fibres from both the space and the suspended rats, was significantly lower than that in the respective control group. Estimation of the total myonuclear volume (microns3 per.mm), however, revealed no significant differences between the three groups in either the tibialis anterior or gastrocnemius. The described changes in the cross-sectional area and myonuclei numbers resulted in significant decreases in the cytoplasmic volume-to-myonucleus ratio (microns3 x 10(3)) in both muscles and for both space and suspended rats (tibialis anterior; 15.6 +/- 0.6 (space), 17.2 +/- 1.0 (suspended), 20.8 +/- 0.9 (control): gastrocnemius; 13.4 +/- 0.4 (space) and 14.9 +/- 1.1 (suspended) versus 18.1 +/- 1.1 (control)). These results indicate that even short periods of unweighting due to microgravity or limb suspension result in changes in skeletal muscle fibres which lead to significant decreases in the cytoplasmic volume-to-myonucleus ratio.

  17. Phase separation between nucleoid and cytoplasm in Escherichia coli as defined by immersive refractometry.

    Science.gov (United States)

    Valkenburg, J A; Woldringh, C L

    1984-01-01

    The refractive indices of nucleoid and cytoplasm in Escherichia coli were derived theoretically and experimentally. For the theoretical estimates, we made use of the known macromolecular composition of E. coli B/r (G. Churchward and H. Bremer, J. Theor. Biol. 94:651-670, 1982) and of estimates of cell and nucleoid volumes. These were obtained from micrographs of living bacteria made with a confocal scanning light microscope. The theoretical values were calculated, assuming that all DNA occurred in the nucleoid and that all protein and RNA occurred in the cytoplasm. Comparison with experimental refractive index values directly obtained by immersive refractometry showed that, besides its DNA, the nucleoid must contain an additional amount of solids equivalent to 8.6% (wt/vol) protein. With the nucleoid containing 6.8% (wt/vol) DNA and 8.6% (wt/vol) protein and the cytoplasm containing 21% (wt/vol) protein and 4% (wt/vol) RNA, a mass difference is obtained, which accounts for the phase separation observed between the nucleoid and cytoplasm in living cells by phase-contrast microscopy. The decrease in the refractive index of the nucleoid relative to that of the cytoplasm observed upon, for instance, OsO4 fixation was interpreted as being indicative of the loss of protein content in the nucleoid. Images PMID:6389508

  18. Behavior change in a lifestyle intervention for type 2 diabetes prevention in Dutch primary care: opportunities for intervention content

    NARCIS (Netherlands)

    Vermunt, P.W.A.; Milder, I.E.J.; Wielaard, F.; Baan, C.A.; Schelfhout, J.D.M.; Westert, G.P.; van Oers, J.A.M.

    2013-01-01

    Background Despite the favorable effects of behavior change interventions on diabetes risk, lifestyle modification is a complicated process. In this study we therefore investigated opportunities for refining a lifestyle intervention for type 2 diabetes prevention, based on participant perceptions of

  19. Inborn errors of cytoplasmic triglyceride metabolism.

    Science.gov (United States)

    Wu, Jiang Wei; Yang, Hao; Wang, Shu Pei; Soni, Krishnakant G; Brunel-Guitton, Catherine; Mitchell, Grant A

    2015-01-01

    Triglyceride (TG) synthesis, storage, and degradation together constitute cytoplasmic TG metabolism (CTGM). CTGM is mostly studied in adipocytes, where starting from glycerol-3-phosphate and fatty acyl (FA)-coenzyme A (CoA), TGs are synthesized then stored in cytoplasmic lipid droplets. TG hydrolysis proceeds sequentially, producing FAs and glycerol. Several reactions of CTGM can be catalyzed by more than one enzyme, creating great potential for complex tissue-specific physiology. In adipose tissue, CTGM provides FA as a systemic energy source during fasting and is related to obesity. Inborn errors and mouse models have demonstrated the importance of CTGM for non-adipose tissues, including skeletal muscle, myocardium and liver, because steatosis and dysfunction can occur. We discuss known inborn errors of CTGM, including deficiencies of: AGPAT2 (a form of generalized lipodystrophy), LPIN1 (childhood rhabdomyolysis), LPIN2 (an inflammatory condition, Majeed syndrome, described elsewhere in this issue), DGAT1 (protein loosing enteropathy), perilipin 1 (partial lipodystrophy), CGI-58 (gene ABHD5, neutral lipid storage disease (NLSD) with ichthyosis and "Jordan's anomaly" of vacuolated polymorphonuclear leukocytes), adipose triglyceride lipase (ATGL, gene PNPLA2, NLSD with myopathy, cardiomyopathy and Jordan's anomaly), hormone-sensitive lipase (HSL, gene LIPE, hypertriglyceridemia, and insulin resistance). Two inborn errors of glycerol metabolism are known: glycerol kinase (GK, causing pseudohypertriglyceridemia) and glycerol-3-phosphate dehydrogenase (GPD1, childhood hepatic steatosis). Mouse models often resemble human phenotypes but may diverge markedly. Inborn errors have been described for less than one-third of CTGM enzymes, and new phenotypes may yet be identified.

  20. Online Social Support for the Prevention of Smoking Relapse: A Content Analysis of the WhatsApp and Facebook Social Groups.

    Science.gov (United States)

    Cheung, Yee Tak Derek; Chan, Ching Han Helen; Wang, Man Ping; Li, Ho Cheung William; Lam, Tai-Hing

    2017-06-01

    Online social groups have been increasingly used for smoking cessation intervention. This study aimed to explore the social support components of the online discussion through WhatsApp and Facebook, how these components addressed the need of relapse prevention, and how the participants evaluated this intervention. We coded and analyzed the posts (N = 467) by the 82 recent quitters in WhatsApp and Facebook social groups, who were recruited from the eight smoking cessation clinics in Hong Kong to participate in a pragmatic randomized trial of relapse prevention. Participants' postintervention feedback was collected from the 13 qualitative interviews after the intervention. The WhatsApp social groups had more participants' posts than the Facebook counterparts. The participants' posts in the online social groups could be classified as sharing views and experiences (55.5%), encouragement (28.7%), and knowledge and information (15.8%). About half of the participants' posts (52.9%) addressed the themes listed in the U.S. Clinical Practice Guideline for preventing smoking relapse. The participants perceived the posts as useful reminders for smoking cessation, but avoidance of reporting relapse, inactive discussions, and uninteresting content were barriers to the success of the intervention. Online social groups provided a useful platform for the delivery of cessation support and encouragement of reporting abstinence, which support relapse prevention. The effectiveness of such intervention can be improved by encouraging more self-report of relapse, active discussions, sharing of interesting content, and using an appropriate discussion platform. Quitters who participate in the online social groups can benefit from peer support and information sharing, and hence prevent smoking relapse.

  1. The molecular mechanism and physiological role of cytoplasmic streaming.

    Science.gov (United States)

    Tominaga, Motoki; Ito, Kohji

    2015-10-01

    Cytoplasmic streaming occurs widely in plants ranging from algae to angiosperms. However, the molecular mechanism and physiological role of cytoplasmic streaming have long remained unelucidated. Recent molecular genetic approaches have identified specific myosin members (XI-2 and XI-K as major and XI-1, XI-B, and XI-I as minor motive forces) for the generation of cytoplasmic streaming among 13 myosin XIs in Arabidopsis thaliana. Simultaneous knockout of these myosin XI members led to a reduced velocity of cytoplasmic streaming and marked defects of plant development. Furthermore, the artificial modifications of myosin XI-2 velocity changed plant and cell sizes along with the velocity of cytoplasmic streaming. Therefore, we assume that cytoplasmic streaming is one of the key regulators in determining plant size. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  2. Chiropractic wellness on the web: the content and quality of information related to wellness and primary prevention on the Internet

    Directory of Open Access Journals (Sweden)

    Evans Marion

    2011-02-01

    Full Text Available Abstract Background The Internet has become a common source of information for patients wishing to learn about health information. Previous studies found information related to back pain poor and often contradictory to current guidelines. Wellness has become a common topic in the field of chiropractic and accrediting agencies have standards on delivery of wellness-based content in college curricula as well as directives for clinical applications. The purpose of this study was to evaluate the quality of the information on the Internet using the terms "chiropractic wellness," or "wellness chiropractic". Methods Five commonly used search engines were selected and the first 10 sites found using the strategy above were evaluated by two raters. Demographic assessments of the sites were made along with whether they were Health on the Net Foundation (HON certified, contained standard wellness content, mentioned any Healthy People Focus Areas, and other chiropractic topics. Kappa statistics compared inter-rater agreement. Results Potential patients appeared to be the audience 87% of the time and a private doctor of chiropractic appeared to be the typical site owner. The sites usually promoted the provider. No sites displayed HON certification logo nor did any appear to meet the HON certification criteria. Twenty-six sites (55% promoted regular physical activity in some manner and 18 (38% had information on health risks of tobacco. Four (9% had mental health or stress-reduction content but none had information supportive of vaccination. Some had information contradictory to common public health measures. Conclusions Patients searching the Internet for chiropractic wellness information will often find useless information that will not help them maintain health or become well. Most simply market the chiropractic practice or allow for a patients to provide personal information in exchange for more 'wellness' information. More research should be done on how

  3. Sequencing and annotation of the chloroplast DNAs and identification of polymorphisms distinguishing normal male-fertile and male-sterile cytoplasms of onion.

    Science.gov (United States)

    von Kohn, Christopher; Kiełkowska, Agnieszka; Havey, Michael J

    2013-12-01

    Male-sterile (S) cytoplasm of onion is an alien cytoplasm introgressed into onion in antiquity and is widely used for hybrid seed production. Owing to the biennial generation time of onion, classical crossing takes at least 4 years to classify cytoplasms as S or normal (N) male-fertile. Molecular markers in the organellar DNAs that distinguish N and S cytoplasms are useful to reduce the time required to classify onion cytoplasms. In this research, we completed next-generation sequencing of the chloroplast DNAs of N- and S-cytoplasmic onions; we assembled and annotated the genomes in addition to identifying polymorphisms that distinguish these cytoplasms. The sizes (153 538 and 153 355 base pairs) and GC contents (36.8%) were very similar for the chloroplast DNAs of N and S cytoplasms, respectively, as expected given their close phylogenetic relationship. The size difference was primarily due to small indels in intergenic regions and a deletion in the accD gene of N-cytoplasmic onion. The structures of the onion chloroplast DNAs were similar to those of most land plants with large and small single copy regions separated by inverted repeats. Twenty-eight single nucleotide polymorphisms, two polymorphic restriction-enzyme sites, and one indel distributed across 20 chloroplast genes in the large and small single copy regions were selected and validated using diverse onion populations previously classified as N or S cytoplasmic using restriction fragment length polymorphisms. Although cytoplasmic male sterility is likely associated with the mitochondrial DNA, maternal transmission of the mitochondrial and chloroplast DNAs allows for polymorphisms in either genome to be useful for classifying onion cytoplasms to aid the development of hybrid onion cultivars.

  4. tRNA-dependent cysteine biosynthetic pathway represents a strategy to increase cysteine contents by preventing it from thermal degradation: thermal adaptation of methanogenic archaea ancestor.

    Science.gov (United States)

    Qu, Ge; Wang, Wei; Chen, Ling-Ling; Qian, Shao-Song; Zhang, Hong-Yu

    2009-10-01

    Although cysteine (Cys) is beneficial to stabilize protein structures, it is not prevalent in thermophiles. For instance, the Cys contents in most thermophilic archaea are only around 0.7%. However, methanogenic archaea, no matter thermophilic or not, contain relatively abundant Cys, which remains elusive for a long time. Recently, Klipcan et al. correlated this intriguing property of methanogenic archaea with their unique tRNA-dependent Cys biosynthetic pathway. But, the deep reasons underlying the correlation are ambiguous. Considering the facts that free Cys is thermally labile and the tRNA-dependent Cys biosynthesis avoids the use of free Cys, we speculate that the unique Cys biosynthetic pathway represents a strategy to increase Cys contents by preventing it from thermal degradation, which may be relevant to the thermal adaptation of methanogenic archaea ancestor.

  5. Content Validation and Semantic Evaluation of a Check-List Elaborated for the Prevention of Gluten Cross-Contamination in Food Services.

    Science.gov (United States)

    Farage, Priscila; Puppin Zandonadi, Renata; Cortez Ginani, Verônica; Gandolfi, Lenora; Pratesi, Riccardo; de Medeiros Nóbrega, Yanna Karla

    2017-01-06

    Conditions associated to the consumption of gluten have emerged as a major health care concern and the treatment consists on a lifelong gluten-free diet. Providing safe food for these individuals includes adapting to safety procedures within the food chain and preventing gluten cross-contamination in gluten-free food. However, a gluten cross-contamination prevention protocol or check-list has not yet been validated. Therefore, the aim of this study was to perform the content validation and semantic evaluation of a check-list elaborated for the prevention of gluten cross-contamination in food services. The preliminary version of the check-list was elaborated based on the Brazilian resolution for food safety Collegiate Board Resolution 216 (RDC 216) and Collegiate Board Resolution 275 (RDC 275), the standard 22000 from the International Organization for Standardization (ISO 22000) and the Canadian Celiac Association Gluten-Free Certification Program documents. Seven experts with experience in the area participated in the check-list validation and semantic evaluation. The criteria used for the approval of the items, as to their importance for the prevention of gluten cross-contamination and clarity of the wording, was the achievement of a minimal of 80% of agreement between the experts (W-values ≥ 0.8). Moreover, items should have a mean ≥4 in the evaluation of importance (Likert scale from 1 to 5) and clarity (Likert scale from 0 to 5) in order to be maintained in the instrument. The final version of the check-list was composed of 84 items, divided into 12 sections. After being redesigned and re-evaluated, the items were considered important and comprehensive by the experts (both with W-values ≥ 0.89). The check-list developed was validated with respect to content and approved in the semantic evaluation.

  6. Content Validation and Semantic Evaluation of a Check-List Elaborated for the Prevention of Gluten Cross-Contamination in Food Services

    Directory of Open Access Journals (Sweden)

    Priscila Farage

    2017-01-01

    Full Text Available Conditions associated to the consumption of gluten have emerged as a major health care concern and the treatment consists on a lifelong gluten-free diet. Providing safe food for these individuals includes adapting to safety procedures within the food chain and preventing gluten cross-contamination in gluten-free food. However, a gluten cross-contamination prevention protocol or check-list has not yet been validated. Therefore, the aim of this study was to perform the content validation and semantic evaluation of a check-list elaborated for the prevention of gluten cross-contamination in food services. The preliminary version of the check-list was elaborated based on the Brazilian resolution for food safety Collegiate Board Resolution 216 (RDC 216 and Collegiate Board Resolution 275 (RDC 275, the standard 22000 from the International Organization for Standardization (ISO 22000 and the Canadian Celiac Association Gluten-Free Certification Program documents. Seven experts with experience in the area participated in the check-list validation and semantic evaluation. The criteria used for the approval of the items, as to their importance for the prevention of gluten cross-contamination and clarity of the wording, was the achievement of a minimal of 80% of agreement between the experts (W-values ≥ 0.8. Moreover, items should have a mean ≥4 in the evaluation of importance (Likert scale from 1 to 5 and clarity (Likert scale from 0 to 5 in order to be maintained in the instrument. The final version of the check-list was composed of 84 items, divided into 12 sections. After being redesigned and re-evaluated, the items were considered important and comprehensive by the experts (both with W-values ≥ 0.89. The check-list developed was validated with respect to content and approved in the semantic evaluation.

  7. pH-Responsive Micelle-Based Cytoplasmic Delivery System for Induction of Cellular Immunity

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    Eiji Yuba

    2017-11-01

    Full Text Available (1 Background: Cytoplasmic delivery of antigens is crucial for the induction of cellular immunity, which is an important immune response for the treatment of cancer and infectious diseases. To date, fusogenic protein-incorporated liposomes and pH-responsive polymer-modified liposomes have been used to achieve cytoplasmic delivery of antigen via membrane rupture or fusion with endosomes. However, a more versatile cytoplasmic delivery system is desired for practical use. For this study, we developed pH-responsive micelles composed of dilauroyl phosphatidylcholine (DLPC and deoxycholic acid and investigated their cytoplasmic delivery performance and immunity-inducing capability. (2 Methods: Interaction of micelles with fluorescence dye-loaded liposomes, intracellular distribution of micelles, and antigenic proteins were observed. Finally, antigen-specific cellular immune response was evaluated in vivo using ELIspot assay. (3 Results: Micelles induced leakage of contents from liposomes via lipid mixing at low pH. Micelles were taken up by dendritic cells mainly via macropinocytosis and delivered ovalbumin (OVA into the cytosol. After intradermal injection of micelles and OVA, OVA-specific cellular immunity was induced in the spleen. (4 Conclusions: pH-responsive micelles composed of DLPC and deoxycholic acid are promising as enhancers of cytosol delivery of antigens and the induction capability of cellular immunity for the treatment of cancer immunotherapy and infectious diseases.

  8. Cytoplasmic crystalloids in irradiated rat parotid glands

    International Nuclear Information System (INIS)

    Sholley, M.M.; Pratt, N.E.; Sodicoff, M.

    1981-01-01

    Cytoplasmic crystalloids were found in paroid acinar cells of rats given a large (6400 R) single exposure of X-rays to the head and neck. The crystalloids were first observed 1 day after irradiation and became numerous at 3-4 days. They were associated with autophagic vacuoles, which were seen in acinar cells as early as 3-6 h. Crystalloids sometimes appeared to be forming within autophagic vacuoles, which also contained membranous residues and apparently degenerating secretory material. They were bounded by a single, smooth membrane and had a substructure consisting of dense, parallel longitudinal striations. The crystalloids were also seen in macrophages associated with the basal surfaces of acinar cells. At 3-4 days macrophages were numerous and many contained crystalloids, degenerated secretory droplets, and other cellular debris, which they apparently had phagocytosed. By 6-8 days crystalloids and macrophages were seen infrequently. Regarding mode of formation, removal by macrophages, and ultrastructure, the crystalloids resembled those described by others after ethionine intoxication. Ethionine-induced crystalloids have cytochemical characteristics consistent with a lysosomal identity. The crystalloids in irradiated parotid glands probably reflect a variant type of lysosome, which is a nonspecific manifestation of severe cellular injury and can be elicited by a variety of injurious agents. (author)

  9. A physical perspective on cytoplasmic streaming.

    Science.gov (United States)

    Goldstein, Raymond E; van de Meent, Jan-Willem

    2015-08-06

    Organisms show a remarkable range of sizes, yet the dimensions of a single cell rarely exceed 100 µm. While the physical and biological origins of this constraint remain poorly understood, exceptions to this rule give valuable insights. A well-known counterexample is the aquatic plant Chara, whose cells can exceed 10 cm in length and 1 mm in diameter. Two spiralling bands of molecular motors at the cell periphery drive the cellular fluid up and down at speeds up to 100 µm s(-1), motion that has been hypothesized to mitigate the slowness of metabolite transport on these scales and to aid in homeostasis. This is the most organized instance of a broad class of continuous motions known as 'cytoplasmic streaming', found in a wide range of eukaryotic organisms-algae, plants, amoebae, nematodes and flies-often in unusually large cells. In this overview of the physics of this phenomenon, we examine the interplay between streaming, transport and cell size and discuss the possible role of self-organization phenomena in establishing the observed patterns of streaming.

  10. Cytoplasmic Dynein Promotes HIV-1 Uncoating

    Directory of Open Access Journals (Sweden)

    Paulina Pawlica

    2014-11-01

    Full Text Available Retroviral capsid (CA cores undergo uncoating during their retrograde transport (toward the nucleus, and/or after reaching the nuclear membrane. However, whether HIV-1 CA core uncoating is dependent upon its transport is not understood. There is some evidence that HIV-1 cores retrograde transport involves cytoplasmic dynein complexes translocating on microtubules. Here we investigate the role of dynein-dependent transport in HIV-1 uncoating. To interfere with dynein function, we depleted dynein heavy chain (DHC using RNA interference, and we over-expressed p50/dynamitin. In immunofluorescence microscopy experiments, DHC depletion caused an accumulation of CA foci in HIV-1 infected cells. Using a biochemical assay to monitor HIV-1 CA core disassembly in infected cells, we observed an increase in amounts of intact (pelletable CA cores upon DHC depletion or p50 over-expression. Results from these two complementary assays suggest that inhibiting dynein-mediated transport interferes with HIV-1 uncoating in infected cells, indicating the existence of a functional link between HIV-1 transport and uncoating.

  11. Brain Metabolic DNA in Rat Cytoplasm.

    Science.gov (United States)

    Giuditta, Antonio; Rutigliano, Bruno

    2018-02-09

    Brain metabolic DNA (BMD) is not involved in cell division or DNA repair but is modulated by memory acquisition, sleep processing, and circadian oscillations. Using routine methods of subcellular fractionation, newly synthesized BMD from male rats is shown to be localized in crude nuclear, mitochondrial, and microsomal fractions and in two fractions of purified nuclei. Sub-fractionation of the mitochondrial fraction indicates the prevalent localization of BMD in free mitochondria and to a lesser degree in synaptosomes and myelin. Cesium density profiles of homogenate, subcellular fractions, and purified nuclei obtained after incorporation periods from 30 min to 4 h indicate that BMD synthesis takes place by reverse transcription in cytoplasmic organelles. Following the acquisition of the double-stranded structure, BMD is transferred to nuclei. Kinetic analyses lasting several weeks highlight the massive BMD turnover in subcellular fractions and purified nuclei and its dependence on age. Data are in agreement with the role of BMD as a temporary information store of cell responses of potential use in comparable forthcoming experiences.

  12. Antineutrophil cytoplasm antibody: positivity and clinical correlation.

    Science.gov (United States)

    Martínez Téllez, Goitybell; Torres Rives, Bárbara; Rangel Velázquez, Suchiquil; Sánchez Rodríguez, Vicky; Ramos Ríos, María Antonia; Fuentes Smith, Lisset Evelyn

    2015-01-01

    To determine positivity and clinical correlation of anti-neutrophil cytoplasmic antibodies (ANCA), taking into account the interference of antinuclear antibodies (ANA). A prospective study was conducted in the Laboratory of Immunology of the National Cuban Center of Medical Genetic during one year. Two hounded sixty-seven patients with indication for ANCA determination were included. ANCA and ANA determinations with different cut off points and assays were determined by indirect immunofluorescense. Anti proteinase 3 and antimyeloperoxidase antibodies were determined by ELISA. Most positivity for ANCA was seen in patients with ANCA associated, primary small-vessel vasculitides, rheumatoid arthritis and systemic lupus erythematosus. Presence of ANCA without positivity for proteinase 3 and myeloperoxidase was higher in patients with ANA and little relation was observed between the perinuclear pattern confirmed in formalin and specificity by myeloperoxidase. Highest sensibility and specificity values for vasculitides diagnostic were achieved by ANCA determination using indirect immunofluorescense with a cut off 1/80 and confirming antigenic specificities with ELISA. ANCA can be present in a great number of chronic inflammatory or autoimmune disorders in the population studied. This determination using indirect immunofluorescence and following by ELISA had a great value for vasculitis diagnosis. Anti mieloperoxidasa assay has a higher utility than the formalin assay when ANA is present. Copyright © 2013 Elsevier España, S.L.U. All rights reserved.

  13. Taurine prevention of calcium paradox-related damage in cardiac muscle. Its regulatory action on intracellular cation contents.

    Science.gov (United States)

    Yamauchi-Takihara, K; Azuma, J; Kishimoto, S; Onishi, S; Sperelakis, N

    1988-07-01

    The present study was designed to investigate in chick heart whether oral pretreatment with taurine or taurine added directly to the perfusate has any effect upon calcium paradox-induced heart failure. In both protocols, taurine significantly reduced the mechanical dysfunction resulting from the calcium paradox. Taurine pretreatment partially inhibited the excess accumulation of calcium in the myocardium that occurs upon calcium repletion, and microscopy revealed almost normal structure. This protective effect of taurine was accompanied by (a) reduction of the gain of sodium content that occurs during calcium depletion, and (b) reduction of the late gain in calcium that occurs during calcium repletion. It is proposed that taurine plays a role in the regulation of calcium homeostasis and membrane stabilization.

  14. State of oral cavity hygiene, sIgA level, mineral content of oral fluid, and optimization of preventive intervention in children with enamel hypoplasia and underlying comorbidity.

    Science.gov (United States)

    Liubarets, S F; Kopylova, O V; Belingio, T O; Kolbasynska, V M; Sechina, I M; Liubarets, T F

    2015-12-01

    To advance the efficiency of tooth caries prevention in children with enamel hypoplasia through a com plex intervention focused at amelioration of oral hygiene, secretory immunoglobulin A (sIgA) level, and mineral con tent of oral fluid. Hypoplasia of permanent teeth enamel was the target of research. State of oral cavity hygiene, sIgA, total calcium and orthophosphate content in oral fluid were the research subject. Enamel vital stain ing, transillumination, immunoenzymometric and photometric techniques were applied. Indices of oral cavity hygiene: simplified oral hygiene index (OHI S) by Green and Vermillion (1964) and Silness Loe (1964, 1967) plaque index were the assayed. Assessment of oral cavity hygiene and assay of sIgA, calcium and orthophosphate content in oral fluid were held in 43 children 6-14 years old (n=21, Group 1; n=22, Group 2) with a systemic enamel hypoplasia and underlying diffuse non toxic (euthyroid) goiter. All children were born from the ChNPP accident clean up work ers and evacuees from territories of mandatory resettlement. Children in the Group 1 were prescribed a set of preven tive interventions including recommendations on choice of the oral care tools and teaching the standard dental cleaning skills. Children in the Group 2 additionally used the remineralizing gel with calcium, phosphorus, magnesium and xylitol (Sanction of the State Sanitary and Epidemiological Service of Ukraine # 05.03.02 03/9351 from 07.02.2014). State of oral cavity hygiene in children of the both study groups was assayed by a "dissatisfactory" criteri on according to the OHI S by Green and Vermillion and by a "poor" criterion according to the Silness Loe plaque index. Upon application of preventive intervrntions the state of oral cavity hygiene improved in children of Group 1 and Group 2: there was a significant decrease of OHI S by Green and Vermillion (p=0.0001) and Silness Loe plaque index (p=0.0003). Administration of remineralizing gel resulted in a

  15. Does a parthenogenesis-inducing Wolbachia induce vestigial cytoplasmic incompatibility?

    Science.gov (United States)

    Kraaijeveld, Ken; Reumer, Barbara M.; Mouton, Laurence; Kremer, Natacha; Vavre, Fabrice; van Alphen, Jacques J. M.

    2011-03-01

    Wolbachia is a maternally inherited bacterium that manipulates the reproduction of its host. Recent studies have shown that male-killing strains can induce cytoplasmic incompatibility (CI) when introgressed into a resistant host. Phylogenetic studies suggest that transitions between CI and other Wolbachia phenotypes have also occurred frequently, raising the possibility that latent CI may be widespread among Wolbachia. Here, we investigate whether a parthenogenesis-inducing Wolbachia strain can also induce CI. Parthenogenetic females of the parasitoid wasp Asobara japonica regularly produce a small number of males that may be either infected or not. Uninfected males were further obtained through removal of the Wolbachia using antibiotics and from a naturally uninfected strain. Uninfected females that had mated with infected males produced a slightly, but significantly more male-biased sex ratio than uninfected females that had mated with uninfected males. This effect was strongest in females that mated with males that had a relatively high Wolbachia titer. Quantitative PCR indicated that infected males did not show higher ratios of nuclear versus mitochondrial DNA content. Wolbachia therefore does not cause diploidization of cells in infected males. While these results are consistent with CI, other alternatives such as production of abnormal sperm by infected males cannot be completely ruled out. Overall, the effect was very small (9%), suggesting that if CI is involved it may have degenerated through the accumulation of mutations.

  16. Antineutrophil Cytoplasmic Antibodies, Autoimmune Neutropenia, and Vasculitis

    Science.gov (United States)

    Grayson, Peter C.; Sloan, J. Mark; Niles, John L.; Monach, Paul A.; Merkel, Peter A.

    2011-01-01

    Objectives Reports of an association between antineutrophil cytoplasmic antibodies (ANCA) and autoimmune neutropenia have rarely included cases of proven vasculitis. A case of ANCA-associated vasculitis (AAV) with recurrent neutropenia is described and relevant literature on the association between ANCA, neutropenia, and vasculitis is reviewed. Methods Longitudinal clinical assessments and laboratory findings are described in a patient with AAV and recurrent episodes of profound neutropenia from December 2008 – October 2010. A PubMed database search of the medical literature was performed for papers published from 1960 through October 2010 to identify all reported cases of ANCA and neutropenia. Results A 49 year-old man developed recurrent neutropenia, periodic fevers, arthritis, biopsy-proven cutaneous vasculitis, sensorineural hearing loss, epididymitis, and positive tests for ANCA with specificity for antibodies to both proteinase 3 and myeloperoxidase. Antineutrophil membrane antibodies were detected during an acute neutropenic phase and were not detectable in a post-recovery sample, whereas ANCA titers did not seem to correlate with neutropenia. An association between ANCA and neutropenia has been reported in 74 cases from 24 studies in the context of drug/toxin exposure, underlying autoimmune disease, or chronic neutropenia without underlying autoimmune disease. In these cases, the presence of atypical ANCA patterns and other antibodies were common; however, vasculitis was uncommon and when it occurred was usually limited to the skin and in cases of underlying toxin exposure. Conclusions ANCA is associated with autoimmune neutropenia, but systemic vasculitis rarely occurs in association with ANCA and neutropenia. The interaction between neutrophils and ANCA may provide insight into understanding both autoimmune neutropenia and AAV. PMID:21507463

  17. Aligning HIV/AIDS communication with the oral tradition of Africans: a theory-based content analysis of songs' potential in prevention efforts.

    Science.gov (United States)

    Bekalu, Mesfin Awoke; Eggermont, Steven

    2015-01-01

    Despite a growing recognition of songs as a useful HIV/AIDS campaign strategy, little research has investigated their potential and/or actual impact. In this study, through a theory-based content analysis, we have assessed the prevention domains covered and the health-relevant constructs promoted by 23 AIDS songs widely used to aid prevention efforts in Ethiopia. To identify the health-relevant constructs and reveal their potential to facilitate or inhibit positive changes, the Extended Parallel Process Model (EPPM) has been used. The findings revealed that the songs cover most of the prevention domains that constitute the current agenda of behavior change communication in Sub-Saharan Africa. However, although all the EPPM variables have been found in almost every song, there were significantly more efficacy messages than threat messages. This suggests that although the songs may lead to positive changes in HIV/AIDS-related outcomes among audiences who have already perceived the threat posed by HIV/AIDS, they are less likely to motivate and thereby generate responses from audiences who have less or no threat perceptions. It is argued that given their potential as a culturally appropriate strategy in Sub-Saharan Africa where oral channels of communication play significant roles, songs could be harnessed for better outcomes through a theory-based design.

  18. Pediatric Inflammatory Bowel Disease with Cytoplasmic Staining of Antineutrophil Cytoplasmic Antibodies

    Directory of Open Access Journals (Sweden)

    Omar I. Saadah

    2013-01-01

    Full Text Available Background. It is unusual for the antineutrophil cytoplasmic antibody with cytoplasmic pattern (cANCA to present in patients with inflammatory bowel disease (IBD without vasculitis. The purpose of this study was to describe the occurrence and characteristics of pediatrics IBD with cANCA. Methods. A retrospective review of pediatric IBD associated with cANCA serology in patients from King Abdulaziz University Hospital, Saudi Arabia, between September 2002 and February 2012. Results. Out of 131 patients with IBD screened for cANCAs, cANCA was positive in 7 (5.3% patients of whom 4 had ulcerative colitis and 3 had Crohn's disease. The median age was 8.8 years (2–14.8 years. Six (86% were males. Of the 7 patients, 5 (71% were Saudi Arabians and 2 were of Indian ethnicity. The most common symptoms were diarrhea, abdominal pain, weight loss, and rectal bleeding. None had family history or clinical features suggestive of vasculitis involving renal and respiratory systems. No difference in the disease location or severity was observed between cANCA positive and cANCA negative patients apart from male preponderance in cANCA positive patients. Conclusion. The occurrence of cANCA in pediatric IBD is rare. Apart from male preponderance, there were no peculiar characteristics for the cANCA positive patients.

  19. CTP synthase forms cytoophidia in the cytoplasm and nucleus

    Energy Technology Data Exchange (ETDEWEB)

    Gou, Ke-Mian [MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford OX1 3PT (United Kingdom); State Key Laboratory for Agrobiotechnology, College of Biological Sciences, China Agricultural University, Beijing 100193 (China); Chang, Chia-Chun [Institute of Biotechnology, National Taiwan University, Taipei, Taiwan, ROC (China); Shen, Qing-Ji [MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford OX1 3PT (United Kingdom); Sung, Li-Ying, E-mail: liyingsung@ntu.edu.tw [Institute of Biotechnology, National Taiwan University, Taipei, Taiwan, ROC (China); Agricultural Biotechnology Research Center, Academia Sinica, Taipei 115, Taiwan, ROC (China); Liu, Ji-Long, E-mail: jilong.liu@dpag.ox.ac.uk [MRC Functional Genomics Unit, Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford OX1 3PT (United Kingdom)

    2014-04-15

    CTP synthase is an essential metabolic enzyme responsible for the de novo synthesis of CTP. Multiple studies have recently showed that CTP synthase protein molecules form filamentous structures termed cytoophidia or CTP synthase filaments in the cytoplasm of eukaryotic cells, as well as in bacteria. Here we report that CTP synthase can form cytoophidia not only in the cytoplasm, but also in the nucleus of eukaryotic cells. Both glutamine deprivation and glutamine analog treatment promote formation of cytoplasmic cytoophidia (C-cytoophidia) and nuclear cytoophidia (N-cytoophidia). N-cytoophidia are generally shorter and thinner than their cytoplasmic counterparts. In mammalian cells, both CTP synthase 1 and CTP synthase 2 can form cytoophidia. Using live imaging, we have observed that both C-cytoophidia and N-cytoophidia undergo multiple rounds of fusion upon glutamine analog treatment. Our study reveals the coexistence of cytoophidia in the cytoplasm and nucleus, therefore providing a good opportunity to investigate the intracellular compartmentation of CTP synthase. - Highlights: • CTP synthase forms cytoophidia not only in the cytoplasm but also in the nucleus. • Glutamine deprivation and Glutamine analogs promotes cytoophidium formation. • N-cytoophidia exhibit distinct morphology when compared to C-cytoophidia. • Both CTP synthase 1 and CTP synthase 2 form cytoophidia in mammalian cells. • Fusions of cytoophidia occur in the cytoplasm and nucleus.

  20. Cytoplasmic streaming emerges naturally from hydrodynamic self-organisation of a microfilament suspension

    Science.gov (United States)

    Woodhouse, Francis; Goldstein, Raymond

    2013-03-01

    Cytoplasmic streaming is the ubiquitous phenomenon of deliberate, active circulation of the entire liquid contents of a plant or animal cell by the walking of motor proteins on polymer filament tracks. Its manifestation in the plant kingdom is particularly striking, where many cells exhibit highly organised patterns of flow. How these regimented flow templates develop is biologically unclear, but there is growing experimental evidence to support hydrodynamically-mediated self-organisation of the underlying microfilament tracks. Using the spirally-streaming giant internodal cells of the characean algae Chara and Nitella as our prototype, we model the developing sub-cortical streaming cytoplasm as a continuum microfilament suspension subject to hydrodynamic and geometric forcing. We show that our model successfully reproduces emergent streaming behaviour by evolving from a totally disordered initial state into a steady characean ``conveyor belt'' configuration as a consequence of the cell geometry, and discuss applicability to other classes of steadily streaming plant cells.

  1. On the evolution of cytoplasmic incompatibility in haplodiploid species

    NARCIS (Netherlands)

    Egas, C.J.M.; de Freitas Vala Salvador, F.; Breeuwer, J.A.J.

    2002-01-01

    The most enigmatic sexual manipulation by Wolbachia endosymbionts is cytoplasmic incompatibility (CI): infected mates are reproductively incompatible with uninfected females. In this paper, we extend the theory on population dynamics and evolution of CI, with emphasis on haplodiploid species. First,

  2. Cytoplasmic ATR Activation Promotes Vaccinia Virus Genome Replication

    Directory of Open Access Journals (Sweden)

    Antonio Postigo

    2017-05-01

    Full Text Available In contrast to most DNA viruses, poxviruses replicate their genomes in the cytoplasm without host involvement. We find that vaccinia virus induces cytoplasmic activation of ATR early during infection, before genome uncoating, which is unexpected because ATR plays a fundamental nuclear role in maintaining host genome integrity. ATR, RPA, INTS7, and Chk1 are recruited to cytoplasmic DNA viral factories, suggesting canonical ATR pathway activation. Consistent with this, pharmacological and RNAi-mediated inhibition of canonical ATR signaling suppresses genome replication. RPA and the sliding clamp PCNA interact with the viral polymerase E9 and are required for DNA replication. Moreover, the ATR activator TOPBP1 promotes genome replication and associates with the viral replisome component H5. Our study suggests that, in contrast to long-held beliefs, vaccinia recruits conserved components of the eukaryote DNA replication and repair machinery to amplify its genome in the host cytoplasm.

  3. Refractory disease in antineutrophil cytoplasmic antibodies associated vasculitis

    NARCIS (Netherlands)

    Rutgers, Abraham; Kallenberg, Cornelis

    Purpose of review Induction treatment of antineutrophil cytoplasmic antibodies (ANCA) associated vasculitis (AAV) is not always successful and nonresponding patients are considered refractory. Recent findings Refractory disease should be subdefined to the treatment that was received.

  4. ( Atp9) gene between cytoplasmic male sterile line and its ...

    African Journals Online (AJOL)

    Administrator

    2011-09-07

    , the atp9 gene in soybeans was cloned from a soybean cytoplasmic male sterile line NJCMS2A and its maintainer line NJCMS2B. Sequence alignment was performed, and protein structures were analyzed and compared ...

  5. Cytoplasmic chromatin triggers inflammation in senescence and cancer.

    Science.gov (United States)

    Dou, Zhixun; Ghosh, Kanad; Vizioli, Maria Grazia; Zhu, Jiajun; Sen, Payel; Wangensteen, Kirk J; Simithy, Johayra; Lan, Yemin; Lin, Yanping; Zhou, Zhuo; Capell, Brian C; Xu, Caiyue; Xu, Mingang; Kieckhaefer, Julia E; Jiang, Tianying; Shoshkes-Carmel, Michal; Tanim, K M Ahasan Al; Barber, Glen N; Seykora, John T; Millar, Sarah E; Kaestner, Klaus H; Garcia, Benjamin A; Adams, Peter D; Berger, Shelley L

    2017-10-19

    Chromatin is traditionally viewed as a nuclear entity that regulates gene expression and silencing. However, we recently discovered the presence of cytoplasmic chromatin fragments that pinch off from intact nuclei of primary cells during senescence, a form of terminal cell-cycle arrest associated with pro-inflammatory responses. The functional significance of chromatin in the cytoplasm is unclear. Here we show that cytoplasmic chromatin activates the innate immunity cytosolic DNA-sensing cGAS-STING (cyclic GMP-AMP synthase linked to stimulator of interferon genes) pathway, leading both to short-term inflammation to restrain activated oncogenes and to chronic inflammation that associates with tissue destruction and cancer. The cytoplasmic chromatin-cGAS-STING pathway promotes the senescence-associated secretory phenotype in primary human cells and in mice. Mice deficient in STING show impaired immuno-surveillance of oncogenic RAS and reduced tissue inflammation upon ionizing radiation. Furthermore, this pathway is activated in cancer cells, and correlates with pro-inflammatory gene expression in human cancers. Overall, our findings indicate that genomic DNA serves as a reservoir to initiate a pro-inflammatory pathway in the cytoplasm in senescence and cancer. Targeting the cytoplasmic chromatin-mediated pathway may hold promise in treating inflammation-related disorders.

  6. Antibody-mediated enzyme replacement therapy targeting both lysosomal and cytoplasmic glycogen in Pompe disease.

    Science.gov (United States)

    Yi, Haiqing; Sun, Tao; Armstrong, Dustin; Borneman, Scott; Yang, Chunyu; Austin, Stephanie; Kishnani, Priya S; Sun, Baodong

    2017-05-01

    Pompe disease is characterized by accumulation of both lysosomal and cytoplasmic glycogen primarily in skeletal and cardiac muscles. Mannose-6-phosphate receptor-mediated enzyme replacement therapy (ERT) with recombinant human acid α-glucosidase (rhGAA) targets the enzyme to lysosomes and thus is unable to digest cytoplasmic glycogen. Studies have shown that anti-DNA antibody 3E10 penetrates living cells and delivers "cargo" proteins to the cytosol or nucleus via equilibrative nucleoside transporter ENT2. We speculate that 3E10-mediated ERT with GAA will target both lysosomal and cytoplasmic glycogen in Pompe disease. A fusion protein (FabGAA) containing a humanized Fab fragment derived from the murine 3E10 antibody and the 110 kDa human GAA precursor was constructed and produced in CHO cells. Immunostaining with an anti-Fab antibody revealed that the Fab signals did not co-localize with the lysosomal marker LAMP2 in cultured L6 myoblasts or Pompe patient fibroblasts after incubation with FabGAA. Western blot with an anti-GAA antibody showed presence of the 150 kDa full-length FabGAA in the cell lysates, in addition to the 95- and 76 kDa processed forms of GAA that were also seen in the rhGAA-treated cells. Blocking of mannose-6-phosphate receptor with mannose-6-phosphate markedly reduced the 95- and the 76 kDa forms but not the 150 kDa form. In GAA-KO mice, FabGAA achieved similar treatment efficacy as rhGAA at an equal molar dose in reducing tissue glycogen contents. Our data suggest that FabGAA retains the ability of rhGAA to treat lysosomal glycogen accumulation and has the beneficial potential over rhGAA to reduce cytoplasmic glycogen storage in Pompe disease. FabGAA can be delivered to both the cytoplasm and lysosomes in cultured cells. FabGAA equally reduced lysosomal glycogen accumulation as rhGAA in GAA-KO mice. FabGAA has the beneficial potential over rhGAA to clear cytoplasmic glycogen. This study suggests a novel antibody-enzyme fusion protein therapy

  7. Self-cytoplasmic DNA upregulates the mutator enzyme APOBEC3A leading to chromosomal DNA damage.

    Science.gov (United States)

    Suspène, Rodolphe; Mussil, Bianka; Laude, Hélène; Caval, Vincent; Berry, Noémie; Bouzidi, Mohamed S; Thiers, Valérie; Wain-Hobson, Simon; Vartanian, Jean-Pierre

    2017-04-07

    Foreign and self-cytoplasmic DNA are recognized by numerous DNA sensor molecules leading to the production of type I interferons. Such DNA agonists should be degraded otherwise cells would be chronically stressed. Most human APOBEC3 cytidine deaminases can initiate catabolism of cytoplasmic mitochondrial DNA. Using the human myeloid cell line THP-1 with an interferon inducible APOBEC3A gene, we show that cytoplasmic DNA triggers interferon α and β production through the RNA polymerase III transcription/RIG-I pathway leading to massive upregulation of APOBEC3A. By catalyzing C→U editing in single stranded DNA fragments, the enzyme prevents them from re-annealing so attenuating the danger signal. The price to pay is chromosomal DNA damage in the form of CG→TA mutations and double stranded DNA breaks which, in the context of chronic inflammation, could drive cells down the path toward cancer. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  8. Dynamics of Galectin-3 in the Nucleus and Cytoplasm

    Science.gov (United States)

    Haudek, Kevin C.; Spronk, Kimberly J.; Voss, Patricia G.; Patterson, Ronald J.; Wang, John L.; Arnoys, Eric J.

    2009-01-01

    This review summarizes selected studies on galectin-3 (Gal3) as an example of the dynamic behavior of a carbohydrate-binding protein in the cytoplasm and nucleus of cells. Within the 15-member galectin family of proteins, Gal3 (Mr ~30,000) is the sole representative of the chimera subclass in which a proline- and glycine-rich NH2-terminal domain is fused onto a COOH-terminal carbohydrate recognition domain responsible for binding galactose-containing glycoconjugates. The protein shuttles between the cytoplasm and nucleus on the basis of targeting signals that are recognized by importin(s) for nuclear localization and exportin-1 (CRM1) for nuclear export. Depending on the cell type, specific experimental conditions in vitro, or tissue location, Gal3 has been reported to be exclusively cytoplasmic, predominantly nuclear, or distributed between the two compartments. The nuclear versus cytoplasmic distribution of the protein must reflect, then, some balance between nuclear import and export, as well as mechanisms of cytoplasmic anchorage or binding to a nuclear component. Indeed, a number of ligands have been reported for Gal3 in the cytoplasm and in the nucleus. Most of the ligands appear to bind Gal3, however, through protein-protein interactions rather than through protein-carbohydrate recognition. In the cytoplasm, for example, Gal3 interacts with the apoptosis repressor Bcl-2 and this interaction may be involved in Gal3’s anti-apoptotic activity. In the nucleus, Gal3 is a required pre-mRNA splicing factor; the protein is incorporated into spliceosomes via its association with the U1 small nuclear ribonucleoprotein (snRNP) complex. Although the majority of these interactions occur via the carbohydrate recognition domain of Gal3 and saccharide ligands such as lactose can perturb some of these interactions, the significance of the protein’s carbohydrate-binding activity, per se, remains a challenge for future investigations. PMID:19616076

  9. Generation of micronuclei during interphase by coupling between cytoplasmic membrane blebbing and nuclear budding.

    Directory of Open Access Journals (Sweden)

    Koh-ichi Utani

    Full Text Available Micronucleation, mediated by interphase nuclear budding, has been repeatedly suggested, but the process is still enigmatic. In the present study, we confirmed the previous observation that there are lamin B1-negative micronuclei in addition to the positive ones. A large cytoplasmic bleb was found to frequently entrap lamin B1-negative micronuclei, which were connected to the nucleus by a thin chromatin stalk. At the bottom of the stalk, the nuclear lamin B1 structure appeared broken. Chromatin extrusion through lamina breaks has been referred to as herniation or a blister of the nucleus, and has been observed after the expression of viral proteins. A cell line in which extrachromosomal double minutes and lamin B1 protein were simultaneously visualized in different colors in live cells was established. By using these cells, time-lapse microscopy revealed that cytoplasmic membrane blebbing occurred simultaneously with the extrusion of nuclear content, which generated lamin B1-negative micronuclei during interphase. Furthermore, activation of cytoplasmic membrane blebbing by the addition of fresh serum or camptothecin induced nuclear budding within 1 to 10 minutes, which suggested that blebbing might be the cause of the budding. After the induction of blebbing, the frequency of lamin-negative micronuclei increased. The budding was most frequent during S phase and more efficiently entrapped small extrachromosomal chromatin than the large chromosome arm. Based on these results, we suggest a novel mechanism in which cytoplasmic membrane dynamics pulls the chromatin out of the nucleus through the lamina break. Evidence for such a mechanism was obtained in certain cancer cell lines including human COLO 320 and HeLa. The mechanism could significantly perturb the genome and influence cancer cell phenotypes.

  10. Ofloxacin induces cytoplasmic respiration-deficient mutants in yeast Saccharomyces cerevisiae.

    Science.gov (United States)

    Obernauerová, M; Subík, J; Ebringer, L

    1992-05-01

    Ofloxacin, a new quinolone with potent antibacterial activity, was also found to be effective against yeast. At relatively high concentrations, and at mild alkaline pH, ofloxacin inhibited the growth of yeast cells in medium containing glucose, and prevented growth on glycerol, as carbon and energy source. The cells growing in the presence of ofloxacin exhibited abberrantly budded forms, lost their viability and many of them converted to cytoplasmic respiration-deficient mutants. Induction of mutants was also observed under non-growing conditions. The petite clones analysed exhibited suppressiveness and contained different fragments of the wild-type mitochondrial genome.

  11. Cytoplasmic streaming velocity as a plant size determinant.

    Science.gov (United States)

    Tominaga, Motoki; Kimura, Atsushi; Yokota, Etsuo; Haraguchi, Takeshi; Shimmen, Teruo; Yamamoto, Keiichi; Nakano, Akihiko; Ito, Kohji

    2013-11-11

    Cytoplasmic streaming is active transport widely occurring in plant cells ranging from algae to angiosperms. Although it has been revealed that cytoplasmic streaming is generated by organelle-associated myosin XI moving along actin bundles, the fundamental function in plants remains unclear. We generated high- and low-speed chimeric myosin XI by replacing the motor domains of Arabidopsis thaliana myosin XI-2 with those of Chara corallina myosin XI and Homo sapiens myosin Vb, respectively. Surprisingly, the plant sizes of the transgenic Arabidopsis expressing high- and low-speed chimeric myosin XI-2 were larger and smaller, respectively, than that of the wild-type plant. This size change correlated with acceleration and deceleration, respectively, of cytoplasmic streaming. Our results strongly suggest that cytoplasmic streaming is a key determinant of plant size. Furthermore, because cytoplasmic streaming is a common system for intracellular transport in plants, our system could have applications in artificial size control in plants. Copyright © 2013 Elsevier Inc. All rights reserved.

  12. Assessment of cytoplasm conductivity by nanosecond pulsed electric fields.

    Science.gov (United States)

    Denzi, Agnese; Merla, Caterina; Palego, Cristiano; Paffi, Alessandra; Ning, Yaqing; Multari, Caroline R; Cheng, Xuanhong; Apollonio, Francesca; Hwang, James C M; Liberti, Micaela

    2015-06-01

    The aim of this paper is to propose a new method for the better assessment of cytoplasm conductivity, which is critical to the development of electroporation protocols as well as insight into fundamental mechanisms underlying electroporation. For this goal, we propose to use nanosecond electrical pulses to bypass the complication of membrane polarization and a single cell to avoid the complication of the application of the "mixing formulas." Further, by suspending the cell in a low-conductivity medium, it is possible to force most of the sensing current through the cytoplasm for a more direct assessment of its conductivity. For proof of principle, the proposed technique was successfully demonstrated on a Jurkat cell by comparing the measured and modeled currents. The cytoplasm conductivity was best assessed at 0.32 S/m and it is in line with the literature. The cytoplasm conductivity plays a key role in the understanding of the basis mechanism of the electroporation phenomenon, and in particular, a large error in the cytoplasm conductivity determination could result in a correspondingly large error in predicting electroporation. Methods for a good estimation of such parameter become fundamental.

  13. Connectosomes for Direct Molecular Delivery to the Cellular Cytoplasm.

    Science.gov (United States)

    Gadok, Avinash K; Busch, David J; Ferrati, Silvia; Li, Brian; Smyth, Hugh D C; Stachowiak, Jeanne C

    2016-10-05

    Transport of biomolecules, drugs, and other reagents across the cell's plasma membrane barrier is an inefficient and poorly controlled process, despite its fundamental importance to biotechnology, cell biology, and pharmaceutics. In particular, insufficient membrane permeability frequently limits the accumulation of drugs and reagents in the cytoplasm, undermining their efficacy. While encapsulating drugs in particles increases uptake by cells, inefficient release of drugs from these particles into the cytoplasm ultimately limits drug efficacy. In contrast, gap junctions provide a direct route to the cytoplasm that bypasses the plasma membrane. As transmembrane channels that physically connect the cytoplasm of adjacent cells, gap junctions permit transport of a diverse range of molecules, from ions and metabolites to siRNA, peptides, and chemotherapeutics. To utilize gap junctions for molecular delivery we have developed Connectosomes, cell-derived lipid vesicles that contain functional gap junction channels and encapsulate molecular cargos. Here we show that these vesicles form gap junction channels with cells, opening a direct and efficient route for the delivery of molecular cargo to the cellular cytoplasm. Specifically, we demonstrate that using gap junctions to deliver the chemotherapeutic doxorubicin reduces the therapeutically effective dose of the drug by more than an order of magnitude. Delivering drugs through gap junctions has the potential to boost the effectiveness of existing drugs such as chemotherapeutics, while simultaneously enabling the delivery of membrane-impermeable drugs and reagents.

  14. Aconitum pseudo-laeve var. erectum Inhibits Receptor Activator of Nuclear Factor Kappa-B Ligand-Induced Osteoclastogenesis via the c-Fos/nuclear Factor of Activated T-Cells, Cytoplasmic 1 Signaling Pathway and Prevents Lipopolysaccharide-Induced Bone Loss in Mice

    Directory of Open Access Journals (Sweden)

    Jong Min Baek

    2014-08-01

    Full Text Available Aconitum pseudo-laeve var. erectum (APE has been widely shown in herbal medicine to have a therapeutic effect on inflammatory conditions. However, there has been no evidence on whether the extract of APE is involved in the biological bone metabolism process, particularly osteoclast-mediated bone resorption. In this study, we confirmed that the administration of APE could restore normal skeletal conditions in a murine model of lipopolysaccharide (LPS-induced bone loss via a decrease in the receptor activator of nuclear factor kappa-B ligand (RANKL/osteoprotegerin (OPG ratio and osteoclast number. We then investigated the effect of APE on the RANKL-induced formation and function of osteoclasts to elucidate its underlying molecular mechanisms. APE suppressed the formation of tartrate-resistant acid phosphatase (TRAP-positive cells, as well as the bone-resorbing activity of mature osteoclasts. Furthermore, APE attenuated nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1 and c-Fos without affecting any early signal pathway of osteoclastogenesis. Subsequently, APE significantly downregulated the expression of various genes exclusively expressed in osteoclasts. These results demonstrate that APE restores LPS-induced bone loss through a decrease of the serum RANKL/OPG ratio, and inhibits osteoclast differentiation and function, suggesting the promise of APE as a potential cure for various osteoclast-associated bone diseases.

  15. Solution structure of the dimeric cytoplasmic domain of syndecan-4

    DEFF Research Database (Denmark)

    Shin, J; Lee, W; Lee, D

    2001-01-01

    cytoplasmic domain is dependent on its oligomeric status, the conformation of the syndecan-4 cytoplasmic domain itself is important in the understanding of its biological roles. Gel filtration results show that the syndecan-4 cytoplasmic domain (4L) itself forms a dimer stabilized by ionic interactions......The syndecans, transmembrane proteoglycans which are involved in the organization of cytoskeleton and/or actin microfilaments, have important roles as cell surface receptors during cell-cell and/or cell-matrix interactions. Since previous studies indicate that the function of the syndecan-4...... in the center of the dimeric twist similar to our previously reported 4V structure. The overall topology of the central variable region within the 4L structure is very similar to that of 4V complexed with the phosphatidylinositol 4,5-bisphosphate; however, the intersubunit interaction mode is affected...

  16. Diffusion within the Cytoplasm: A Mesoscale Model of Interacting Macromolecules

    Science.gov (United States)

    Trovato, Fabio; Tozzini, Valentina

    2014-01-01

    Recent experiments carried out in the dense cytoplasm of living cells have highlighted the importance of proteome composition and nonspecific intermolecular interactions in regulating macromolecule diffusion and organization. Despite this, the dependence of diffusion-interaction on physicochemical properties such as the degree of poly-dispersity and the balance between steric repulsion and nonspecific attraction among macromolecules was not systematically addressed. In this work, we study the problem of diffusion-interaction in the bacterial cytoplasm, combining theory and experimental data to build a minimal coarse-grained representation of the cytoplasm, which also includes, for the first time to our knowledge, the nucleoid. With stochastic molecular-dynamics simulations of a virtual cytoplasm we are able to track the single biomolecule motion, sizing from 3 to 80 nm, on submillisecond-long trajectories. We demonstrate that the size dependence of diffusion coefficients, anomalous exponents, and the effective viscosity experienced by biomolecules in the cytoplasm is fine-tuned by the intermolecular interactions. Accounting only for excluded volume in these potentials gives a weaker size-dependence than that expected from experimental data. On the contrary, adding nonspecific attraction in the range of 1–10 thermal energy units produces a stronger variation of the transport properties at growing biopolymer sizes. Normal and anomalous diffusive regimes emerge straightforwardly from the combination of high macromolecular concentration, poly-dispersity, stochasticity, and weak nonspecific interactions. As a result, small biopolymers experience a viscous cytoplasm, while the motion of big ones is jammed because the entanglements produced by the network of interactions and the entropic effects caused by poly-dispersity are stronger. PMID:25468337

  17. Genetic expression of induced rice sterility under alien-cytoplasm

    International Nuclear Information System (INIS)

    Wang Naiyuan; Cai Zhijun; Liang Kangjing; Li Yu

    2005-01-01

    Rice restorer lines were treated with 60 Co γ-ray and 4 male sterile mutants obtained with the fertility of controlled by 4 non-allelic recessive genes, respectively. Sixty combinations were made by using male sterile plants/fertile plants as male parents, and 15 different cytoplasmic substitution lines of the same cell nucleus as female parents. The result showed that F 1 spikelets were normal and fertile, and different numbers of male sterile plants were segregated in F 2 . Complete fertility genotype was not found among interactions between induced male sterile genes and alien-cytoplasms. (authors)

  18. Colicin S8 export: extracellular and cytoplasmic colicin are different.

    Science.gov (United States)

    Garcia Diaz, Maria-Elena; Concepción Curbelo, Juan Luis

    2003-12-01

    The properties of colicin S8 are different for the cytoplasmic, periplasmic and extracellular protein. Interactions with its specific receptors reflect this. Active cell extracts separate into a non-anionic along with an anionic fraction by DEAE-Sephacell chromatography. Previously, we have purified cell-associated colicin S8 as an aggregation of highly related polypeptides; cytoplasmic colicin S8 seems to be post-translationally processed into an aggregation of polypeptides of molecular mass ranging from 45,000 Da to 60,000 Da. We suggest that a conformational change to colicin S8 may occur related to the export process.

  19. Intracellular Fluid Mechanics: Coupling Cytoplasmic Flow with Active Cytoskeletal Gel

    Science.gov (United States)

    Mogilner, Alex; Manhart, Angelika

    2018-01-01

    The cell is a mechanical machine, and continuum mechanics of the fluid cytoplasm and the viscoelastic deforming cytoskeleton play key roles in cell physiology. We review mathematical models of intracellular fluid mechanics, from cytoplasmic fluid flows, to the flow of a viscous active cytoskeletal gel, to models of two-phase poroviscous flows, to poroelastic models. We discuss application of these models to cell biological phenomena, such as organelle positioning, blebbing, and cell motility. We also discuss challenges of understanding fluid mechanics on the cellular scale.

  20. Cytoplasmic transduction peptide (CTP): New approach for the delivery of biomolecules into cytoplasm in vitro and in vivo

    International Nuclear Information System (INIS)

    Kim, Daeyou; Jeon, Choonju; Kim, Jeong-Hwan; Kim, Mi-Seon; Yoon, Cheol-Hee; Choi, In-Soo; Kim, Sung-Hoon; Bae, Yong-Soo

    2006-01-01

    The protein transduction domain (PTD) of HIV-1 TAT has been extensively documented with regard to its membrane transduction potential, as well as its efficient delivery of biomolecules in vivo. However, the majority of PTD and PTD-conjugated molecules translocate to the nucleus rather than to the cytoplasm after transduction, due to the functional nuclear localization sequence (NLS). Here, we report a cytoplasmic transduction peptide (CTP), which was deliberately designed to ensure the efficient cytoplasmic delivery of the CTP-fused biomolecules. In comparison with PTD, CTP and its fusion partners exhibited a clear preference for cytoplasmic localization, and also markedly enhanced membrane transduction potential. Unlike the mechanism underlying PTD-mediated transduction, CTP-mediated transduction occurs independently of the lipid raft-dependent macropinocytosis pathway. The CTP-conjugated Smac/DIABLO peptide (Smac-CTP) was also shown to be much more efficient than Smac-PTD in the blockage of the antiapoptotic properties of XIAP, suggesting that cytoplasmic functional molecules can be more efficiently targeted by CTP-mediated delivery. In in vivo trafficking studies, CTP-fused β-gal exhibited unique organ tropisms to the liver and lymph nodes when systemically injected into mice, whereas PTD-β-gal exhibited no such tropisms. Taken together, our findings implicate CTP as a novel delivery peptide appropriate for (i) molecular targeting to cytoplasmic compartments in vitro, (ii) the development of class I-associated CTL vaccines, and (iii) special drug delivery in vivo, without causing any untoward effects on nuclear genetic material

  1. Experimental Analysis of Cell Function Using Cytoplasmic Streaming

    Science.gov (United States)

    Janssens, Peter; Waldhuber, Megan

    2012-01-01

    This laboratory exercise investigates the phenomenon of cytoplasmic streaming in the fresh water alga "Nitella". Students use the fungal toxin cytochalasin D, an inhibitor of actin polymerization, to investigate the mechanism of streaming. Students use simple statistical methods to analyze their data. Typical student data are provided. (Contains 3…

  2. Endoplasmic-reticulum-mediated microtubule alignment governs cytoplasmic streaming.

    Science.gov (United States)

    Kimura, Kenji; Mamane, Alexandre; Sasaki, Tohru; Sato, Kohta; Takagi, Jun; Niwayama, Ritsuya; Hufnagel, Lars; Shimamoto, Yuta; Joanny, Jean-François; Uchida, Seiichi; Kimura, Akatsuki

    2017-04-01

    Cytoplasmic streaming refers to a collective movement of cytoplasm observed in many cell types. The mechanism of meiotic cytoplasmic streaming (MeiCS) in Caenorhabditis elegans zygotes is puzzling as the direction of the flow is not predefined by cell polarity and occasionally reverses. Here, we demonstrate that the endoplasmic reticulum (ER) network structure is required for the collective flow. Using a combination of RNAi, microscopy and image processing of C. elegans zygotes, we devise a theoretical model, which reproduces and predicts the emergence and reversal of the flow. We propose a positive-feedback mechanism, where a local flow generated along a microtubule is transmitted to neighbouring regions through the ER. This, in turn, aligns microtubules over a broader area to self-organize the collective flow. The proposed model could be applicable to various cytoplasmic streaming phenomena in the absence of predefined polarity. The increased mobility of cortical granules by MeiCS correlates with the efficient exocytosis of the granules to protect the zygotes from osmotic and mechanical stresses.

  3. Evaluation of cytoplasmic genetic effects for production and ...

    African Journals Online (AJOL)

    The model used for WW include direct additive, maternal additive, the covariance between the animal-, permanent maternal environmental-, herd-year-season x sire- and damline effects, while the simplest model which include only direct- and damline effects were used for AFC. The contribution of the cytoplasmic effects to ...

  4. Nuclear proteins hijacked by mammalian cytoplasmic plus strand RNA viruses

    Energy Technology Data Exchange (ETDEWEB)

    Lloyd, Richard E., E-mail: rlloyd@bcm.edu

    2015-05-15

    Plus strand RNA viruses that replicate in the cytoplasm face challenges in supporting the numerous biosynthetic functions required for replication and propagation. Most of these viruses are genetically simple and rely heavily on co-opting cellular proteins, particularly cellular RNA-binding proteins, into new roles for support of virus infection at the level of virus-specific translation, and building RNA replication complexes. In the course of infectious cycles many nuclear-cytoplasmic shuttling proteins of mostly nuclear distribution are detained in the cytoplasm by viruses and re-purposed for their own gain. Many mammalian viruses hijack a common group of the same factors. This review summarizes recent gains in our knowledge of how cytoplasmic RNA viruses use these co-opted host nuclear factors in new functional roles supporting virus translation and virus RNA replication and common themes employed between different virus groups. - Highlights: • Nuclear shuttling host proteins are commonly hijacked by RNA viruses to support replication. • A limited group of ubiquitous RNA binding proteins are commonly hijacked by a broad range of viruses. • Key virus proteins alter roles of RNA binding proteins in different stages of virus replication.

  5. Cytoplasmic nucleophosmin (cNPM) in acute myeloid leukaemia ...

    African Journals Online (AJOL)

    Amani H. Kazem

    2011-08-26

    Aug 26, 2011 ... cytoplasmic positivity for NPM was significantly correlated with increased survival and better out- come after cycles of chemotherapy. .... Statistical Package for Social Science (SPSS) version 17 format and data explore was carried out. ..... ACT cytogenetics laboratory manual. The Association of. Cytogenetic ...

  6. Magnetite nanoparticles as reporters for microcarrier processing in cytoplasm

    Energy Technology Data Exchange (ETDEWEB)

    Reibetanz, Uta, E-mail: uta.reibetanz@medizin.uni-leipzig.de [Translational Centre for Regenerative Medicine (TRM) Leipzig, Universitaet Leipzig, Philipp-Rosenthal-Strasse 55, 04103 Leipzig (Germany); Institute for Medical Physics and Biophysics, Medical Faculty, Universitaet Leipzig, Haertelstrasse 16-18, 04107 Leipzig (Germany); Jankuhn, Steffen, E-mail: jankuhn@uni-leipzig.de [Division of Nuclear Solid State Physics, Faculty of Physics and Geosciences, Universitaet Leipzig, Linnestrasse 5, 04103 Leipzig (Germany); Office for Environmental Protection and Occupational Safety, Universitaet Leipzig, Ritterstrasse 24, 04109 Leipzig (Germany)

    2011-10-15

    The development and therapeutic application of drug delivery systems based on colloidal microcarriers layer-by-layer coated with biopolyelectrolytes requires the investigation of their processing inside the cell for the successful and efficient transport and release of the active agents. The present study is focused on the time-dependent multilayer decomposition and the subsequent release of active agents to the cytoplasm. Magnetite nanoparticles (MNP) were used as reporter agents integrated into the protamine sulfate/dextran sulfate basis multilayer on colloidal SiO{sub 2} cores. This functionalization allows the monitoring of the multilayer decomposition due to the detection of the MNP release, visualized by means of proton-induced X-ray emission (PIXE) by elemental distribution of Si and Fe. The direct correlation between the microcarrier localization in endolysosomes and cytoplasm of HEK293T/17 cells via confocal laser scanning microscopy (CLSM) and the elemental distribution (PIXE) allows tracing the fate of the MNP-coated microcarriers in cytoplasm, and thus the processing of the multilayer. Microcarrier/cell co-incubation experiments of 6 h, 24 h, 48 h, and 72 h show that a MNP release and a slight expansion into the cytoplasm occurs after a longer co-incubation of 72 h.

  7. Actin-mediated cytoplasmic organization of plant cells

    NARCIS (Netherlands)

    Honing, van der H.S.

    2011-01-01

    In this thesis, I present results that give insight in the role of the actin cytoskeleton in the production of an organized cytoplasm in plant cells, which is, for instance, required for proper cell morphogenesis.

    Chapter 1 is a review in which we discuss the possible role of actin-based

  8. Animal models of antineutrophil cytoplasm antibody-associated vasculitis.

    LENUS (Irish Health Repository)

    Salama, Alan D

    2012-01-01

    To provide an update on the experimental models that have been developed recapitulating clinical antineutrophil cytoplasm antibody (ANCA) associated vasculitis. The application of the models in the study of pathogenesis, and the therapeutic implications of this, are covered in the article by van Timmeren and Heeringa in this issue.

  9. Analysis of embryo, cytoplasmic and maternal correlations for quality ...

    Indian Academy of Sciences (India)

    Home; Journals; Journal of Genetics; Volume 85; Issue 2. Analysis of embryo, cytoplasmic and maternal correlations for quality traits of rapeseed (Brassica napus L.) across environments. C. H. Shi H. Z. Zhang J. G. Wu. Research Note Volume 85 Issue 2 August 2006 pp 147-151 ...

  10. The transmission of cytoplasmic genes in Aspergillus nidulans

    NARCIS (Netherlands)

    Coenen, A.

    1997-01-01


    Introduction

    This manuscript concerns the spread of selfish cytoplasmic genes in the fungus Aspergillus nidulans. A.nidulans is a common soil fungus that grows vegetatively by forming a network (mycelium) of hyphae and reproduces

  11. Evaluation of cytoplasmic genetic effects for production and ...

    African Journals Online (AJOL)

    uvp

    2014-12-03

    Dec 3, 2014 ... year-season x sire- and damline effects, while the simplest model which include only direct- and damline effects were ... estimates of additive genetic effects, functional genetic material in the cytoplasm of oocytes must be present .... inheritance in three synthetic lines of beef cattle differing in mature size.

  12. Poison Prevention

    Science.gov (United States)

    ... Prevention Listen Español Text Size Email Print Share Poison Prevention Page Content Article Body Post the Poison Help number 1-800-222-1222 on the ... or empty container of a toxic substance, call Poison Help immediately. More than a million American children ...

  13. Calmodulin antagonists effect on Ca(2+ level in the mitochondria and cytoplasm of myometrium cells

    Directory of Open Access Journals (Sweden)

    S. G. Shlykov

    2015-10-01

    Full Text Available It is known that Са2+-dependent regulation of this cation exchange in mitochondria is carried out with participation of calmodulin. We had shown in a previous work using two experimental models: isolated mitochondria and intact myometrium cells, that calmodulin antagonists reduce the level of mitochondrial membrane polarization. The aim of this work was to investigate the influence of calmodulin antagonists on the level of ionized Са in mitochondria and cytoplasm of uterine smooth muscle cells using spectrofluorometry and confocal microscopy. It was shown that myometrium mitochondria, in the presence of АТР and MgCl2 in the incubation medium, accumulate Са ions in the matrix. Incubation of mitochondria in the presence of СССР inhibited cation accumulation, but did not cease it. Calmodulin antagonist such as trifluoperazine (100 µМ considerably increased the level of ionized Са in the mitochondrial matrix. Preliminary incubation of mitochondria with 100 µМ Са2+, before adding trifluoperazine to the incubation medium, partly prevented influence of the latter on the cation level in the matrix. Incubation of myometrium cells (primary culture with another calmodulin antagonist calmidazolium (10 µМ was accompanied by depolarization of mitochondrial membrane and an increase in the concentration of ionized Са in cytoplasm. Thus, using two models, namely, isolated mitochondria and intact myometrium cells, it has been shown that calmodulin antagonists cause depolarization of mitochondrial membranes and an increase of the ionized Са concentration in both the mitochondrial matrix and the cell cytoplasm.

  14. Fluoride enhances transfection activity of carbonate apatite by increasing cytoplasmic stability of plasmid DNA

    International Nuclear Information System (INIS)

    Chowdhury, E.H.

    2011-01-01

    Highlights: → Cytoplasmic stability of plasmid DNA is enhanced by fluoride incorporation into carbonate apatite carrier. → Fluoridated carbonate apatite promotes a robust increase in transgene expression. → Controlled dissolution of fluoridated carbonate apatite in endosomal acidic environment might buffer the endosomes and prevent degradation of the released DNA. -- Abstract: Intracellular delivery of a functional gene or a nucleic acid sequence to specifically knockdown a harmful gene is a potential approach to precisely treat a critical human disease. The intensive efforts in the last few decades led to the development of a number of viral and non-viral synthetic vectors. However, an ideal delivery tool in terms of the safety and efficacy has yet to be established. Recently, we have developed pH-sensing inorganic nanocrystals of carbonate apatite for efficient and cell-targeted delivery of gene and gene-silencing RNA. Here we show that addition of very low level of fluoride to the particle-forming medium facilitates a robust increase in transgene expression following post-incubation of the particles with HeLa cells. Confocal microscopic observation and Southern blotting prove the cytoplasmic existence of plasmid DNA delivered by likely formed fluoridated carbonate apatite particles while degradation of plasmid DNA presumably by cytoplasmic nucleases was noticed following delivery with apatite particles alone. The beneficial role of fluoride in enhancing carbonate apatite-mediated gene expression might be due to the buffering potential of generated fluoridated apatite in endosomal acidic environment, thereby increasing the half-life of delivered plasmid DNA.

  15. Fluoride enhances transfection activity of carbonate apatite by increasing cytoplasmic stability of plasmid DNA

    Energy Technology Data Exchange (ETDEWEB)

    Chowdhury, E.H., E-mail: md.ezharul.hoque@med.monash.edu.my [Jeffrey Cheah School of Medicine and Health Sciences, Monash University Sunway Campus, Jalan Lagoon Selatan, Bandar Sunway, Selangor Darul Ehsan (Malaysia)

    2011-06-17

    Highlights: {yields} Cytoplasmic stability of plasmid DNA is enhanced by fluoride incorporation into carbonate apatite carrier. {yields} Fluoridated carbonate apatite promotes a robust increase in transgene expression. {yields} Controlled dissolution of fluoridated carbonate apatite in endosomal acidic environment might buffer the endosomes and prevent degradation of the released DNA. -- Abstract: Intracellular delivery of a functional gene or a nucleic acid sequence to specifically knockdown a harmful gene is a potential approach to precisely treat a critical human disease. The intensive efforts in the last few decades led to the development of a number of viral and non-viral synthetic vectors. However, an ideal delivery tool in terms of the safety and efficacy has yet to be established. Recently, we have developed pH-sensing inorganic nanocrystals of carbonate apatite for efficient and cell-targeted delivery of gene and gene-silencing RNA. Here we show that addition of very low level of fluoride to the particle-forming medium facilitates a robust increase in transgene expression following post-incubation of the particles with HeLa cells. Confocal microscopic observation and Southern blotting prove the cytoplasmic existence of plasmid DNA delivered by likely formed fluoridated carbonate apatite particles while degradation of plasmid DNA presumably by cytoplasmic nucleases was noticed following delivery with apatite particles alone. The beneficial role of fluoride in enhancing carbonate apatite-mediated gene expression might be due to the buffering potential of generated fluoridated apatite in endosomal acidic environment, thereby increasing the half-life of delivered plasmid DNA.

  16. Detecting relationships between amylose content and amino acid ...

    Indian Academy of Sciences (India)

    The relationship between the genetic effects of endosperm, cytoplasm and maternal plant on amylose content (AC) and amino acid contents of indica rice was studied using unconditional and conditional analysis methods. The results indicated that the protein content (PC) and brown rice weight (WBR) could significantly ...

  17. Selective export of HLA-F by its cytoplasmic tail.

    Science.gov (United States)

    Boyle, Louise H; Gillingham, Alison K; Munro, Sean; Trowsdale, John

    2006-06-01

    MHC class I molecules exit the endoplasmic reticulum (ER) by an unknown mechanism. Although a selective export mechanism has been proposed for the anterograde transport of class I, a motif responsible for export has never been identified. Although classical class I molecules lacking their cytoplasmic tail are expressed on the cell surface, we found that HLA-F was entirely dependent on its cytoplasmic tail for export from the ER. Two known export motifs were recognizable in HLA-F. A C-terminal valine residue functioned in ER export and interacted with coat complex (COP)II, while an RxR motif also played an important role in anterograde transport and bound to 14-3-3 proteins. This divergent trafficking of HLA-F implicates an alternative function for HLA-F, independent of loading with peptides in the ER.

  18. No single way to explain cytoplasmic maturation of oocytes from prepubertal and cyclic gilts.

    Science.gov (United States)

    Pawlak, P; Cieslak, A; Warzych, E; Zejden, Z; Szumacher-Strabel, M; Molinska-Glura, M; Lechniak, D

    2012-12-01

    The objective of this study was to evaluate selected aspects of cytoplasmic maturation in oocytes from prepubertal and cyclic crossbred gilts before and after in vitro maturation. For this purpose, cortical granule redistribution, mitochondrial DNA content and mitochondria translocation were analyzed. Moreover, for the first time the fatty acid profiles in follicular fluid (FF) of both gilt categories was evaluated. The nuclear maturation (the percentage of metaphase II oocytes was 83% in prepubertal gilts compared with 87% in cyclic gilts), cortical granule relocation from the cortex to peripheral ooplasm (98.7% vs. 98.8% of oocytes, respectively) and mitochondrial DNA content (227 543 vs. 206 660, respectively) was not affected by sexual maturity of the donor gilt. However, the redistribution of active mitochondria during in vitro maturation was observed only in the oocytes of cyclic gilts. With regard to FF analysis, saturated, unsaturated, and monounsaturated fatty acids were significantly more abundant in the FF of prepubertal females. In particular, stearic (C18:0) and palmitic (C16:0) fatty acids had significantly higher concentrations in the FF of prepubertal gilts. In conclusion, although the oocytes of prepubertal gilts matured in vitro at a rate similar to those of cyclic gilts, they differed with respect to the selected factors attributed to cytoplasmic maturation. We suggest that the higher content of particular fatty acids, which is known to have a negative influence on oocyte maturation, as well as impaired mitochondria redistribution are factors limiting the maturation potential of oocytes from prepubertal gilts. Copyright © 2012 Elsevier Inc. All rights reserved.

  19. Influenza a virus assembly intermediates fuse in the cytoplasm.

    Directory of Open Access Journals (Sweden)

    Seema S Lakdawala

    2014-03-01

    Full Text Available Reassortment of influenza viral RNA (vRNA segments in co-infected cells can lead to the emergence of viruses with pandemic potential. Replication of influenza vRNA occurs in the nucleus of infected cells, while progeny virions bud from the plasma membrane. However, the intracellular mechanics of vRNA assembly into progeny virions is not well understood. Here we used recent advances in microscopy to explore vRNA assembly and transport during a productive infection. We visualized four distinct vRNA segments within a single cell using fluorescent in situ hybridization (FISH and observed that foci containing more than one vRNA segment were found at the external nuclear periphery, suggesting that vRNA segments are not exported to the cytoplasm individually. Although many cytoplasmic foci contain multiple vRNA segments, not all vRNA species are present in every focus, indicating that assembly of all eight vRNA segments does not occur prior to export from the nucleus. To extend the observations made in fixed cells, we used a virus that encodes GFP fused to the viral polymerase acidic (PA protein (WSN PA-GFP to explore the dynamics of vRNA assembly in live cells during a productive infection. Since WSN PA-GFP colocalizes with viral nucleoprotein and influenza vRNA segments, we used it as a surrogate for visualizing vRNA transport in 3D and at high speed by inverted selective-plane illumination microscopy. We observed cytoplasmic PA-GFP foci colocalizing and traveling together en route to the plasma membrane. Our data strongly support a model in which vRNA segments are exported from the nucleus as complexes that assemble en route to the plasma membrane through dynamic colocalization events in the cytoplasm.

  20. Nucleoporin Nup98 mediates galectin-3 nuclear-cytoplasmic trafficking

    Energy Technology Data Exchange (ETDEWEB)

    Funasaka, Tatsuyoshi, E-mail: funasaka@staff.kanazawa-u.ac.jp [Laboratory of Molecular and Cellular Biology, Department of Biology, Faculty of Natural Systems, Institute of Science and Engineering, Kanazawa University, Ishikawa (Japan); Balan, Vitaly; Raz, Avraham [Department of Oncology and Pathology, Karmanos Cancer Institute, Wayne State University, School of Medicine, Detroit, MI (United States); Wong, Richard W., E-mail: rwong@staff.kanazawa-u.ac.jp [Laboratory of Molecular and Cellular Biology, Department of Biology, Faculty of Natural Systems, Institute of Science and Engineering, Kanazawa University, Ishikawa (Japan); Bio-AFM Frontier Research Center, Kanazawa Kanazawa University, Ishikawa (Japan)

    2013-04-26

    Highlights: •Nuclear pore protein Nup98 is a novel binding partner of galectin-3. •Nup98 transports galectin-3 into cytoplasm. •Nup98 depletion leads to galectin-3 nuclear transport and induces growth retardation. •Nup98 may involve in ß-catenin pathway through interaction with galectin-3. -- Abstract: Nucleoporin Nup98 is a component of the nuclear pore complex, and is important in transport across the nuclear pore. Many studies implicate nucleoporin in cancer progression, but no direct mechanistic studies of its effect in cancer have been reported. We show here that Nup98 specifically regulates nucleus–cytoplasm transport of galectin-3, which is a ß-galactoside-binding protein that affects adhesion, migration, and cancer progression, and controls cell growth through the ß-catenin signaling pathway in cancer cells. Nup98 interacted with galectin-3 on the nuclear membrane, and promoted galectin-3 cytoplasmic translocation whereas other nucleoporins did not show these functions. Inversely, silencing of Nup98 expression by siRNA technique localized galectin-3 to the nucleus and retarded cell growth, which was rescued by Nup98 transfection. In addition, Nup98 RNA interference significantly suppressed downstream mRNA expression in the ß-catenin pathway, such as cyclin D1 and FRA-1, while nuclear galectin-3 binds to ß-catenin to inhibit transcriptional activity. Reduced expression of ß-catenin target genes is consistent with the Nup98 reduction and the galectin-3–nucleus translocation rate. Overall, the results show Nup98’s involvement in nuclear–cytoplasm translocation of galectin-3 and ß-catenin signaling pathway in regulating cell proliferation, and the results depicted here suggest a novel therapeutic target/modality for cancers.

  1. Conformational changes at cytoplasmic intersubunit interactions control Kir channel gating.

    Science.gov (United States)

    Wang, Shizhen; Borschel, William F; Heyman, Sarah; Hsu, Phillip; Nichols, Colin G

    2017-06-16

    The defining structural feature of inward-rectifier potassium (Kir) channels is the unique Kir cytoplasmic domain. Recently we showed that salt bridges located at the cytoplasmic domain subunit interfaces (CD-Is) of eukaryotic Kir channels control channel gating via stability of a novel inactivated closed state. The cytoplasmic domains of prokaryotic and eukaryotic Kir channels show similar conformational rearrangements to the common gating ligand, phosphatidylinositol bisphosphate (PIP 2 ), although these exhibit opposite coupling to opening and closing transitions. In Kir2.1, mutation of one of these CD-I salt bridge residues (R204A) reduces apparent PIP 2 sensitivity of channel activity, and here we show that Ala or Cys substitutions of the functionally equivalent residue (Arg-165) in the prokaryotic Kir channel KirBac1.1 also significantly decrease sensitivity of the channel to PIP 2 (by 5-30-fold). To further understand the structural basis of CD-I control of Kir channel gating, we examined the effect of the R165A mutation on PIP 2 -induced changes in channel function and conformation. Single-channel analyses indicated that the R165A mutation disrupts the characteristic long interburst closed state of reconstituted KirBac1.1 in giant liposomes, resulting in a higher open probability due to more frequent opening bursts. Intramolecular FRET measurements indicate that, relative to wild-type channels, the R165A mutation results in splaying of the cytoplasmic domains away from the central axis and that PIP 2 essentially induces opposite motions of the major β-sheet in this channel mutant. We conclude that the removal of stabilizing CD-I salt bridges results in a collapsed state of the Kir domain. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  2. CD248 facilitates tumor growth via its cytoplasmic domain

    Directory of Open Access Journals (Sweden)

    Janssens Tom

    2011-05-01

    Full Text Available Abstract Background Stromal fibroblasts participate in the development of a permissive environment for tumor growth, yet molecular pathways to therapeutically target fibroblasts are poorly defined. CD248, also known as endosialin or tumor endothelial marker 1 (TEM1, is a transmembrane glycoprotein expressed on activated fibroblasts. We recently showed that the cytoplasmic domain of CD248 is important in facilitating an inflammatory response in a mouse model of arthritis. Others have reported that CD248 gene inactivation in mice results in dampened tumor growth. We hypothesized that the conserved cytoplasmic domain of CD248 is important in regulating tumor growth. Methods Mice lacking the cytoplasmic domain of CD248 (CD248CyD/CyD were generated and evaluated in tumor models, comparing the findings with wild-type mice (CD248WT/WT. Results As compared to the response in CD248WT/WT mice, growth of T241 fibrosarcomas and Lewis lung carcinomas was significantly reduced in CD248CyD/CyD mice. Tumor size was similar to that seen with CD248-deficient mice. Conditioned media from CD248CyD/CyD fibroblasts were less effective at supporting T241 fibrosarcoma cell survival. In addition to our previous observation of reduced release of activated matrix metalloproteinase (MMP-9, CD248CyD/CyD fibroblasts also had impaired PDGF-BB-induced migration and expressed higher transcripts of tumor suppressor factors, transgelin (SM22α, Hes and Hey1. Conclusions The multiple pathways regulated by the cytoplasmic domain of CD248 highlight its potential as a therapeutic target to treat cancer.

  3. Genetic studies on cytoplasmic male sterility in maize

    Energy Technology Data Exchange (ETDEWEB)

    Laughnan, J.R.

    1992-01-01

    Our research concerns the basic mechanisms of cytoplasmic male sterility (CMS) and fertility restoration in maize. The molecular determination of CMS is in the DNA of the mitochondria (mtDNA) but specific nuclear restorer-of-fertility (Rf) genes can overrule the male-sterile effect of the cytoplasm. Our approach to the study of the Rf genes is threefold. We are attempting to tag the cms-S Rf genes and the cms-T Rf2 gene with controlling elements (CEs). Since we have identified a number of spontaneous Rf genes for cms-S and have demonstrated that they are themselves transposable, we are also searching for cases in which an Rf gene is inserted into a wild-type gene. The other aspect of our research involves the nuclear control over the organization of the mitochondrial genome. We found that the changes in mtDNA organization upon cytoplasmic reversion to fertility were characteristic of the nuclear background in which the reversion event occurred. We have investigated whether these differences are a reflection of differences in the organization of the mtDNA genome before reversion.

  4. Nucleoporin Nup98 mediates galectin-3 nuclear-cytoplasmic trafficking.

    Science.gov (United States)

    Funasaka, Tatsuyoshi; Balan, Vitaly; Raz, Avraham; Wong, Richard W

    2013-04-26

    Nucleoporin Nup98 is a component of the nuclear pore complex, and is important in transport across the nuclear pore. Many studies implicate nucleoporin in cancer progression, but no direct mechanistic studies of its effect in cancer have been reported. We show here that Nup98 specifically regulates nucleus-cytoplasm transport of galectin-3, which is a ß-galactoside-binding protein that affects adhesion, migration, and cancer progression, and controls cell growth through the ß-catenin signaling pathway in cancer cells. Nup98 interacted with galectin-3 on the nuclear membrane, and promoted galectin-3 cytoplasmic translocation whereas other nucleoporins did not show these functions. Inversely, silencing of Nup98 expression by siRNA technique localized galectin-3 to the nucleus and retarded cell growth, which was rescued by Nup98 transfection. In addition, Nup98 RNA interference significantly suppressed downstream mRNA expression in the ß-catenin pathway, such as cyclin D1 and FRA-1, while nuclear galectin-3 binds to ß-catenin to inhibit transcriptional activity. Reduced expression of ß-catenin target genes is consistent with the Nup98 reduction and the galectin-3-nucleus translocation rate. Overall, the results show Nup98's involvement in nuclear-cytoplasm translocation of galectin-3 and ß-catenin signaling pathway in regulating cell proliferation, and the results depicted here suggest a novel therapeutic target/modality for cancers. Copyright © 2013 Elsevier Inc. All rights reserved.

  5. Cytoplasmic streaming in plant cells emerges naturally by microfilament self-organization.

    Science.gov (United States)

    Woodhouse, Francis G; Goldstein, Raymond E

    2013-08-27

    Many cells exhibit large-scale active circulation of their entire fluid contents, a process termed cytoplasmic streaming. This phenomenon is particularly prevalent in plant cells, often presenting strikingly regimented flow patterns. The driving mechanism in such cells is known: myosin-coated organelles entrain cytoplasm as they process along actin filament bundles fixed at the periphery. Still unknown, however, is the developmental process that constructs the well-ordered actin configurations required for coherent cell-scale flow. Previous experimental works on streaming regeneration in cells of Characean algae, whose longitudinal flow is perhaps the most regimented of all, hint at an autonomous process of microfilament self-organization driving the formation of streaming patterns during morphogenesis. Working from first principles, we propose a robust model of streaming emergence that combines motor dynamics with both microscopic and macroscopic hydrodynamics to explain how several independent processes, each ineffectual on its own, can reinforce to ultimately develop the patterns of streaming observed in the Characeae and other streaming species.

  6. Cytoplasmic streaming in Drosophila oocytes varies with kinesin activity and correlates with the microtubule cytoskeleton architecture.

    Science.gov (United States)

    Ganguly, Sujoy; Williams, Lucy S; Palacios, Isabel M; Goldstein, Raymond E

    2012-09-18

    Cells can localize molecules asymmetrically through the combined action of cytoplasmic streaming, which circulates their fluid contents, and specific anchoring mechanisms. Streaming also contributes to the distribution of nutrients and organelles such as chloroplasts in plants, the asymmetric position of the meiotic spindle in mammalian embryos, and the developmental potential of the zygote, yet little is known quantitatively about the relationship between streaming and the motor activity which drives it. Here we use Particle Image Velocimetry to quantify the statistical properties of Kinesin-dependent streaming during mid-oogenesis in Drosophila. We find that streaming can be used to detect subtle changes in Kinesin activity and that the flows reflect the architecture of the microtubule cytoskeleton. Furthermore, based on characterization of the rheology of the cytoplasm in vivo, we establish estimates of the number of Kinesins required to drive the observed streaming. Using this in vivo data as the basis of a model for transport, we suggest that the disordered character of transport at mid-oogenesis, as revealed by streaming, is an important component of the localization dynamics of the body plan determinant oskar mRNA.

  7. Impact-Free Measurement of Microtubule Rotations on Kinesin and Cytoplasmic-Dynein Coated Surfaces.

    Directory of Open Access Journals (Sweden)

    Aniruddha Mitra

    Full Text Available Knowledge about the three-dimensional stepping of motor proteins on the surface of microtubules (MTs as well as the torsional components in their power strokes can be inferred from longitudinal MT rotations in gliding motility assays. In previous studies, optical detection of these rotations relied on the tracking of rather large optical probes present on the outer MT surface. However, these probes may act as obstacles for motor stepping and may prevent the unhindered rotation of the gliding MTs. To overcome these limitations, we devised a novel, impact-free method to detect MT rotations based on fluorescent speckles within the MT structure in combination with fluorescence-interference contrast microscopy. We (i confirmed the rotational pitches of MTs gliding on surfaces coated by kinesin-1 and kinesin-8 motors, (ii demonstrated the superiority of our method over previous approaches on kinesin-8 coated surfaces at low ATP concentration, and (iii identified MT rotations driven by mammalian cytoplasmic dynein, indicating that during collective motion cytoplasmic dynein side-steps with a bias in one direction. Our novel method is easy to implement on any state-of-the-art fluorescence microscope and allows for high-throughput experiments.

  8. Chloroplast and cytoplasmic ribosomes of Euglena: selective binding of dihydrostreptomycin to chloroplast ribosomes.

    Science.gov (United States)

    Schwartzbach, S D; Schiff, J A

    1974-10-01

    Dihydrostreptomycin binds preferentially to chloroplast ribosomes of wild-type Euglena gracilis Klebs var. bacillaris Pringsheim. The K(diss) for the wild-type chloroplast ribosome-dihydrostreptomycin complex is 2 x 10(-7) M, a value comparable with that found for the Escherichia coli ribosome-dihydrostreptomycin complex. Chloroplast ribosomes isolated from the streptomycin-resistant mutant Sm(1) (r)BNgL and cytoplasmic ribosomes from wild-type have a much lower affinity for the antibiotic. The K(diss) for the chloroplast ribosome-dihydrostreptomycin complex of Sm(1) (r) is 387 x 10(-7) M, and the value for the cytoplasmic ribosome-dihydrostreptomycin complex of the wild type is 1,400 x 10(-7) M. Streptomycin competes with dihydrostreptomycin for the chloroplast ribosome binding site, and preincubation of streptomycin with hydroxylamine prevents the binding of streptomycin to the chloroplast ribosome. These results indicate that the inhibition of chloroplast development and replication in Euglena by streptomycin and dihydrostreptomycin is related to the specific inhibition of protein synthesis on the chloroplast ribosomes of Euglena.

  9. Chloroplast and Cytoplasmic Ribosomes of Euglena: Selective Binding of Dihydrostreptomycin to Chloroplast Ribosomes1

    Science.gov (United States)

    Schwartzbach, Steven D.; Schiff, Jerome A.

    1974-01-01

    Dihydrostreptomycin binds preferentially to chloroplast ribosomes of wild-type Euglena gracilis Klebs var. bacillaris Pringsheim. The Kdiss for the wild-type chloroplast ribosome-dihydrostreptomycin complex is 2 × 10−7 M, a value comparable with that found for the Escherichia coli ribosome-dihydrostreptomycin complex. Chloroplast ribosomes isolated from the streptomycin-resistant mutant Sm1rBNgL and cytoplasmic ribosomes from wild-type have a much lower affinity for the antibiotic. The Kdiss for the chloroplast ribosome-dihydrostreptomycin complex of Sm1r is 387 × 10−7 M, and the value for the cytoplasmic ribosome-dihydrostreptomycin complex of the wild type is 1,400 × 10−7 M. Streptomycin competes with dihydrostreptomycin for the chloroplast ribosome binding site, and preincubation of streptomycin with hydroxylamine prevents the binding of streptomycin to the chloroplast ribosome. These results indicate that the inhibition of chloroplast development and replication in Euglena by streptomycin and dihydrostreptomycin is related to the specific inhibition of protein synthesis on the chloroplast ribosomes of Euglena. PMID:4138802

  10. The effect of ionizing irradiation on motion of cytoplasm in cells of Elodea canadensis

    International Nuclear Information System (INIS)

    Tordyiya, N.V.; Grodzyins'kij, D.M.; Danil'chenko, O.O.

    1999-01-01

    The effect of acute irradiation on the velocity of cytoplasm is investigated. It is shown that, for small doses, there is a strong nonlinearity between the velocity of cytoplasm and dose. The nonlinear behavior disappears with increasing a dose

  11. Changes in electrostatic surface potential of Na+/K+-ATPase cytoplasmic headpiece induced by cytoplasmic ligand(s) binding.

    Science.gov (United States)

    Kubala, Martin; Grycova, Lenka; Lansky, Zdenek; Sklenovsky, Petr; Janovska, Marika; Otyepka, Michal; Teisinger, Jan

    2009-09-16

    A set of single-tryptophan mutants of the Na(+)/K(+)-ATPase isolated, large cytoplasmic loop connecting transmembrane helices M4 and M5 (C45) was prepared to monitor effects of the natural cytoplasmic ligands (i.e., Mg(2+) and/or ATP) binding. We introduced a novel method for the monitoring of the changes in the electrostatic surface potential (ESP) induced by ligand binding, using the quenching of the intrinsic tryptophan fluorescence by acrylamide or iodide. This approach opens a new way to understanding the interactions within the proteins. Our experiments revealed that the C45 conformation in the presence of the ATP (without magnesium) substantially differed from the conformation in the presence of Mg(2+) or MgATP or in the absence of any ligand not only in the sense of geometry but also in the sense of the ESP. Notably, the set of ESP-sensitive residues was different from the set of geometry-sensitive residues. Moreover, our data indicate that the effect of the ligand binding is not restricted only to the close environment of the binding site and that the information is in fact transmitted also to the distal parts of the molecule. This property could be important for the communication between the cytoplasmic headpiece and the cation binding sites located within the transmembrane domain.

  12. DMPD: Negative regulation of cytoplasmic RNA-mediated antiviral signaling. [Dynamic Macrophage Pathway CSML Database

    Lifescience Database Archive (English)

    Full Text Available 18703349 Negative regulation of cytoplasmic RNA-mediated antiviral signaling. Komur...Show Negative regulation of cytoplasmic RNA-mediated antiviral signaling. PubmedID 18703349 Title Negative r...egulation of cytoplasmic RNA-mediated antiviral signaling. Authors Komuro A, Bamm

  13. Navigating HIV prevention policy and Islam in Malaysia: contention, compatibility or reconciliation? Findings from in-depth interviews among key stakeholders

    Directory of Open Access Journals (Sweden)

    Sima Barmania

    2016-07-01

    Full Text Available Abstract Background Malaysia is a multicultural society, predominantly composed of a Muslim majority population, where Islam is influential. Malaysia has a concentrated HIV epidemic amongst high risk groups, such as, Intravenous Drug Users (IVDU, sex workers, transgender women and Men who have sex with Men (MSM. The objective of this study is to understand how Islam shapes HIV prevention strategies in Malaysia by interviewing the three key stakeholder groups identified as being influential, namely the Ministry of Health, Religious leaders and People living with HIV. Methods Thirty-Five in depth semi structured interviews were undertaken with religious leaders, Ministry of Health and People living with HIV in the last half of 2013 using purposive sampling. Interviews adhered to a topic guide, were audiotaped, and transcribed verbatim and analyzed using a framework analysis. Results Themes including the importance of Islam to health, stakeholder relationships and opinions on HIV prevention emerged. Islam was seen to play a pivotal role in shaping strategies relating to HIV prevention in Malaysia both directly and indirectly. Stakeholders often held different approaches to HIV prevention, which had to be sensitively considered, with some favouring promotion of Islamic principles, whilst others steering towards a more public health centred approach. Conclusions The study suggests that Islam indeed plays an important role in shaping health policies and strategies related to HIV prevention in Malaysia. Certainly, stakeholders do hold differing viewpoints, such as stances of what constitutes the right approach to HIV prevention. However there are also areas of broad consensus, such as the importance in Islamic tradition to prevent harm and disease, which can be crafted into existing and future HIV prevention strategies in Malaysia, as well as the wider Muslim world.

  14. Navigating HIV prevention policy and Islam in Malaysia: contention, compatibility or reconciliation? Findings from in-depth interviews among key stakeholders.

    Science.gov (United States)

    Barmania, Sima; Aljunid, Syed Mohamed

    2016-07-07

    Malaysia is a multicultural society, predominantly composed of a Muslim majority population, where Islam is influential. Malaysia has a concentrated HIV epidemic amongst high risk groups, such as, Intravenous Drug Users (IVDU), sex workers, transgender women and Men who have sex with Men (MSM). The objective of this study is to understand how Islam shapes HIV prevention strategies in Malaysia by interviewing the three key stakeholder groups identified as being influential, namely the Ministry of Health, Religious leaders and People living with HIV. Thirty-Five in depth semi structured interviews were undertaken with religious leaders, Ministry of Health and People living with HIV in the last half of 2013 using purposive sampling. Interviews adhered to a topic guide, were audiotaped, and transcribed verbatim and analyzed using a framework analysis. Themes including the importance of Islam to health, stakeholder relationships and opinions on HIV prevention emerged. Islam was seen to play a pivotal role in shaping strategies relating to HIV prevention in Malaysia both directly and indirectly. Stakeholders often held different approaches to HIV prevention, which had to be sensitively considered, with some favouring promotion of Islamic principles, whilst others steering towards a more public health centred approach. The study suggests that Islam indeed plays an important role in shaping health policies and strategies related to HIV prevention in Malaysia. Certainly, stakeholders do hold differing viewpoints, such as stances of what constitutes the right approach to HIV prevention. However there are also areas of broad consensus, such as the importance in Islamic tradition to prevent harm and disease, which can be crafted into existing and future HIV prevention strategies in Malaysia, as well as the wider Muslim world.

  15. Endogenous Mouse Dicer Is an Exclusively Cytoplasmic Protein.

    Directory of Open Access Journals (Sweden)

    Christian Much

    2016-06-01

    Full Text Available Dicer is a large multi-domain protein responsible for the ultimate step of microRNA and short-interfering RNA biogenesis. In human and mouse cell lines, Dicer has been shown to be important in the nuclear clearance of dsRNA as well as the establishment of chromatin modifications. Here we set out to unambiguously define the cellular localization of Dicer in mice to understand if this is a conserved feature of mammalian Dicer in vivo. To this end, we utilized an endogenously epitope tagged Dicer knock-in mouse allele. From primary mouse cell lines and adult tissues, we determined with certainty by biochemical fractionation and confocal immunofluorescence microscopy that endogenous Dicer is exclusively cytoplasmic. We ruled out the possibility that a fraction of Dicer shuttles to and from the nucleus as well as that FGF or DNA damage signaling induce Dicer nuclear translocation. We also explored Dicer localization during the dynamic and developmental context of embryogenesis, where Dicer is ubiquitously expressed and strictly cytoplasmic in all three germ layers as well as extraembryonic tissues. Our data exclude a direct role for Dicer in the nuclear RNA processing in the mouse.

  16. Endogenous Mouse Dicer Is an Exclusively Cytoplasmic Protein.

    Science.gov (United States)

    Much, Christian; Auchynnikava, Tania; Pavlinic, Dinko; Buness, Andreas; Rappsilber, Juri; Benes, Vladimir; Allshire, Robin; O'Carroll, Dónal

    2016-06-01

    Dicer is a large multi-domain protein responsible for the ultimate step of microRNA and short-interfering RNA biogenesis. In human and mouse cell lines, Dicer has been shown to be important in the nuclear clearance of dsRNA as well as the establishment of chromatin modifications. Here we set out to unambiguously define the cellular localization of Dicer in mice to understand if this is a conserved feature of mammalian Dicer in vivo. To this end, we utilized an endogenously epitope tagged Dicer knock-in mouse allele. From primary mouse cell lines and adult tissues, we determined with certainty by biochemical fractionation and confocal immunofluorescence microscopy that endogenous Dicer is exclusively cytoplasmic. We ruled out the possibility that a fraction of Dicer shuttles to and from the nucleus as well as that FGF or DNA damage signaling induce Dicer nuclear translocation. We also explored Dicer localization during the dynamic and developmental context of embryogenesis, where Dicer is ubiquitously expressed and strictly cytoplasmic in all three germ layers as well as extraembryonic tissues. Our data exclude a direct role for Dicer in the nuclear RNA processing in the mouse.

  17. Effects of cytoplasmic inheritance on preweaning traits of Hereford cattle

    Directory of Open Access Journals (Sweden)

    Mezzadra Carlos Alberto

    2005-01-01

    Full Text Available The influence of cytoplasmic inheritance on birth and weaning weight was evaluated in an experimental Hereford herd. Data on 1,720 records for birth and weaning weights from calves born between 1963 and 2002 were studied. Variance components were estimated using MTDFREML procedures and an animal model was fitted for each trait. Direct and maternal additive effects and permanent environment and maternal lineage effects were treated as random, while year and month of birth, age of dam and sex of the calf were treated as fixed. Identification of maternal lineages was based on pedigree information. The contribution to phenotypic variance of cytoplasmic lineages defined by pedigree information was negligible for both traits. Mitochondrial genotypes of cows present in the herd in 2002 were analyzed by single strand conformation polymorphism (SSCP analysis. Only five different genotypes were identified among 23 maternal lineages. All the animals with records were assigned to maternal genotypes based on pedigree information. The statistical analysis was repeated, removing maternal lineage from the model and including mitochondrial genotype as a fixed effect. No evidence of genotype effects was detected. These results suggest a negligible effect of the mitochondrial genome on the preweaning traits of this Hereford herd.

  18. Response Mechanisms of Bacterial Degraders to Environmental Contaminants on the Level of Cell Walls and Cytoplasmic Membrane

    Directory of Open Access Journals (Sweden)

    Slavomíra Murínová

    2014-01-01

    Full Text Available Bacterial strains living in the environment must cope with the toxic compounds originating from humans production. Surface bacterial structures, cell wall and cytoplasmic membrane, surround each bacterial cell and create selective barriers between the cell interior and the outside world. They are a first site of contact between the cell and toxic compounds. Organic pollutants are able to penetrate into cytoplasmic membrane and affect membrane physiological functions. Bacteria had to evolve adaptation mechanisms to counteract the damage originated from toxic contaminants and to prevent their accumulation in cell. This review deals with various adaptation mechanisms of bacterial cell concerning primarily the changes in cytoplasmic membrane and cell wall. Cell adaptation maintains the membrane fluidity status and ratio between bilayer/nonbilayer phospholipids as well as the efflux of toxic compounds, protein repair mechanisms, and degradation of contaminants. Low energy consumption of cell adaptation is required to provide other physiological functions. Bacteria able to survive in toxic environment could help us to clean contaminated areas when they are used in bioremediation technologies.

  19. Response mechanisms of bacterial degraders to environmental contaminants on the level of cell walls and cytoplasmic membrane.

    Science.gov (United States)

    Murínová, Slavomíra; Dercová, Katarína

    2014-01-01

    Bacterial strains living in the environment must cope with the toxic compounds originating from humans production. Surface bacterial structures, cell wall and cytoplasmic membrane, surround each bacterial cell and create selective barriers between the cell interior and the outside world. They are a first site of contact between the cell and toxic compounds. Organic pollutants are able to penetrate into cytoplasmic membrane and affect membrane physiological functions. Bacteria had to evolve adaptation mechanisms to counteract the damage originated from toxic contaminants and to prevent their accumulation in cell. This review deals with various adaptation mechanisms of bacterial cell concerning primarily the changes in cytoplasmic membrane and cell wall. Cell adaptation maintains the membrane fluidity status and ratio between bilayer/nonbilayer phospholipids as well as the efflux of toxic compounds, protein repair mechanisms, and degradation of contaminants. Low energy consumption of cell adaptation is required to provide other physiological functions. Bacteria able to survive in toxic environment could help us to clean contaminated areas when they are used in bioremediation technologies.

  20. Dietary Intake of Structured Lipids with Different Contents of Medium-Chain Fatty Acids on Obesity Prevention in C57BL/6J Mice.

    Science.gov (United States)

    Zhou, Shengmin; Wang, Yueqiang; Jiang, Yuanrong; Zhang, Zhongfei; Sun, Xiangjun; Yu, Liangli Lucy

    2017-08-01

    Three medium- and long-chain triacylglycerols (MLCT) with different contents of medium-chain fatty acids (MCFA) (10% to 30%, w/w) were prepared and evaluated for their anti-obesity potential in C57BL/6J mice. The group fed with a high fat diet of MLCT containing 30% (w/w) MCFA showed significantly decreased body weight and fat mass (P obesity-inducing high fat rapeseed oil diet. In addition, serum parameters including triacylglycerols, total cholesterol, glucose, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, apolipoprotein A1 and apolipoprotein B in the treatment group fed with 30% (w/w) MCFA were close to those of mice fed with a low fat rapeseed oil diet, but significantly different (P obesity control group. Moreover, the intake of MLCT with high content of MCFA reduced the size of adipocytes. In addition, the visceral fat and liver weights, as well as the liver triacylglycerol for 3 treatment groups were lower than those of the obesity control group. These results demonstrate the great potential of MLCT with high content of MCFA in weight loss. © 2017 Institute of Food Technologists®.

  1. Regular moderate or intense exercise prevents depression-like behavior without change of hippocampal tryptophan content in chronically tryptophan-deficient and stressed mice.

    Directory of Open Access Journals (Sweden)

    Hosung Lee

    Full Text Available Regular exercise has an antidepressant effect in human subjects. Studies using animals have suggested that the antidepressant effect of exercise is attributable to an increase of brain 5-hydroxytryptamine (5-HT; however, the precise mechanism underlying the antidepressant action via exercise is unclear. In contrast, the effect of 5-HT on antidepressant activity has not been clarified, in part because the therapeutic response to antidepressant drugs has a time lag in spite of the rapid increase of brain 5-HT upon administration of these drugs. This study was designed to investigate the contribution of brain 5-HT to the antidepressant effect of exercise. Mice were fed a tryptophan-deficient diet and stressed using chronic unpredictable stress (CUS for 4 weeks with or without the performance of either moderate or intense exercise on a treadmill 3 days per week. The findings demonstrated that the onset of depression-like behavior is attributable not to chronic reduction of 5-HT but to chronic stress. Regular exercise, whether moderate or intense, prevents depression-like behavior with an improvement of adult hippocampal cell proliferation and survival and without the recovery of 5-HT. Concomitantly, the mice that exercised showed increased hippocampal noradrenaline. Regular exercise prevents the impairment of not long-term memory but short-term memory in a 5-HT-reduced state. Together, these findings suggest that: (1 chronic reduction of brain 5-HT may not contribute to the onset of depression-like behavior; (2 regular exercise, whether moderate or intense, prevents the onset of chronic stress-induced depression-like behavior independent of brain 5-HT and dependent on brain adrenaline; and (3 regular exercise prevents chronic tryptophan reduction-induced impairment of not long-term but short-term memory.

  2. A case of community-based fall prevention: Survey of organization and content of minor home help services in Swedish municipalities.

    Science.gov (United States)

    Bernfort, Lars; Eckard, Nathalie; Husberg, Magnus; Alwin, Jenny

    2014-11-01

    The aim of this study was to survey minor home help services provided by Swedish municipalities with the main purpose to prevent fall injuries. If minor home help services were presented on the homepage of a municipality, an initial telephone contact was taken. Thereafter a questionnaire was administered, including questions about target groups, aim with the services, tasks included, costs and restrictions for users, budget, and experienced gains with the services. Municipalities not providing minor home help services were asked about the reason therefore and if the municipality had previously provided the services Results: The questionnaire response rate was 92%. In 191 of Sweden's 290 municipalities services were provided by, or in cooperation with, the municipality. Reasons for not providing the services were mainly financial and lack of demand. Services were more often provided in larger cities and in municipalities located in populous regions. In some municipalities services were performed by persons with functional disabilities or unemployed persons. Both providers and users expressed satisfaction with the services aspects expressed were that services lead to greater sense of safety and social gains the effect of the services in terms of fall prevention is yet to be proved with only a small fall-preventive effect services are probably cost-effective improved quality of life, sense of safety, and being able to offer meaningful work to otherwise unemployed persons are important aspects that might in themselves motivate the provision of minor home help services. © 2014 the Nordic Societies of Public Health.

  3. Nuclear translocation of the cytoplasmic domain of HB-EGF induces gastric cancer invasion

    International Nuclear Information System (INIS)

    Shimura, Takaya; Higashiyama, Shigeki; Joh, Takashi; Yoshida, Michihiro; Fukuda, Shinji; Ebi, Masahide; Hirata, Yoshikazu; Mizoshita, Tsutomu; Tanida, Satoshi; Kataoka, Hiromi; Kamiya, Takeshi

    2012-01-01

    Membrane-anchored heparin-binding epidermal growth factor-like growth factor (proHB-EGF) yields soluble HB-EGF, which is an epidermal growth factor receptor (EGFR) ligand, and a carboxy-terminal fragment of HB-EGF (HB-EGF-CTF) after ectodomain shedding. We previously reported that HB-EGF-CTF and unshed proHB-EGF which has the cytoplasmic domain of proHB-EGF (HB-EGF-C), translocate from the plasma membrane to the nucleus and regulate cell cycle after shedding stimuli. However, the significance of nuclear exported HB-EGF-C in human gastric cancer is unclear. We investigated the relationship between intracellular localization of HB-EGF-C and clinical outcome in 96 gastric cancer patients treated with gastrectomy. Moreover, we established stable gastric cancer cell lines overexpressing wild-type HB-EGF (wt-HB-EGF) and mutated HB-EGF (HB-EGF-mC), which prevented HB-EGF-C nuclear translocation after shedding. Cell motility between these 2 gastric cancer cell lines was investigated using a transwell invasion assay and a wound healing assay. Of the 96 gastric cancer cases, HB-EGF-C immunoreactivity was detected in both the nucleus and cytoplasm in 19 cases (19.8 %) and in the cytoplasm only in 25 cases (26.0 %). The nuclear immunoreactivity of HB-EGF-C was significantly increased in stage pT3/4 tumors compared with pT1/2 tumors (T1/2 vs. T3/4: 11.1 % vs. 36.4 %, P < 0.01). The growth of wt-HB-EGF- and HB-EGF-mC-expressing cells significantly increased compared with control cells, but the growth of HB-EGF-mC-expressing cells was significantly decreased compared with wt-HB-EGF-expressing cells. Gastric cancer cell invasion obviously increased in wt-HB-EGF-expressing cells, but invasion in HB-EGF-mC-expressing cells showed a slight increase compared with control cells. Moreover, wt-HB-EGF overexpression increased the effectiveness of wound healing, but had no significant effect in HB-EGF-mC-expressing cells. Both the function of HB-EGF as an EGFR ligand and a novel signal for

  4. Nuclear translocation of the cytoplasmic domain of HB-EGF induces gastric cancer invasion

    Directory of Open Access Journals (Sweden)

    Shimura Takaya

    2012-05-01

    Full Text Available Abstract Background Membrane-anchored heparin-binding epidermal growth factor-like growth factor (proHB-EGF yields soluble HB-EGF, which is an epidermal growth factor receptor (EGFR ligand, and a carboxy-terminal fragment of HB-EGF (HB-EGF-CTF after ectodomain shedding. We previously reported that HB-EGF-CTF and unshed proHB-EGF which has the cytoplasmic domain of proHB-EGF (HB-EGF-C, translocate from the plasma membrane to the nucleus and regulate cell cycle after shedding stimuli. However, the significance of nuclear exported HB-EGF-C in human gastric cancer is unclear. Methods We investigated the relationship between intracellular localization of HB-EGF-C and clinical outcome in 96 gastric cancer patients treated with gastrectomy. Moreover, we established stable gastric cancer cell lines overexpressing wild-type HB-EGF (wt-HB-EGF and mutated HB-EGF (HB-EGF-mC, which prevented HB-EGF-C nuclear translocation after shedding. Cell motility between these 2 gastric cancer cell lines was investigated using a transwell invasion assay and a wound healing assay. Results Of the 96 gastric cancer cases, HB-EGF-C immunoreactivity was detected in both the nucleus and cytoplasm in 19 cases (19.8 % and in the cytoplasm only in 25 cases (26.0 %. The nuclear immunoreactivity of HB-EGF-C was significantly increased in stage pT3/4 tumors compared with pT1/2 tumors (T1/2 vs. T3/4: 11.1 % vs. 36.4 %, P  Conclusions Both the function of HB-EGF as an EGFR ligand and a novel signal for HB-EGF-C nuclear translocation induce gastric cancer growth, whereas HB-EGF-C nuclear translocation independently plays a critical role in gastric cancer invasion. The present study demonstrated that HB-EGF-C nuclear translocation might be crucial in gastric cancer invasion. HB-EGF-C nuclear translocation may offer a prognostic marker and a new molecular target for gastric cancer therapy.

  5. The Cytoplasm-to-Vacuole Targeting Pathway: A Historical Perspective

    Directory of Open Access Journals (Sweden)

    Midori Umekawa

    2012-01-01

    Full Text Available From today's perspective, it is obvious that macroautophagy (hereafter autophagy is an important pathway that is connected to a range of developmental and physiological processes. This viewpoint, however, is relatively recent, coinciding with the molecular identification of autophagy-related (Atg components that function as the protein machinery that drives the dynamic membrane events of autophagy. It may be difficult, especially for scientists new to this area of research, to appreciate that the field of autophagy long existed as a “backwater” topic that attracted little interest or attention. Paralleling the development of the autophagy field was the identification and analysis of the cytoplasm-to-vacuole targeting (Cvt pathway, the only characterized biosynthetic route that utilizes the Atg proteins. Here, we relate some of the initial history, including some never-before-revealed facts, of the analysis of the Cvt pathway and the convergence of those studies with autophagy.

  6. Propylthiouracil-Induced Vasculitis With Antineutrophil Cytoplasmic Antibody.

    Science.gov (United States)

    Criado, Paulo Ricardo; Grizzo Peres Martins, Ana Claudia; Gaviolli, Camila Fatima; Alavi, Afsaneh

    2015-06-01

    Propylthiouracil (PTU)-associated vasculitis is a potentially life-threatening disease with a recent increase in the reported cases in the medical literature. This increase may suggest that some earlier cases have been unrecognized or assigned to an alternative nosology category. Although the skin can be the only organ affected by PTU-associated vasculitis, there are many reports with multiple-system involvement. Classically, the symptoms appear under a tetrad of fever, sore throat, arthralgia, and skin lesions. Cutaneous lesions in reported cases of PTU vasculitis have most commonly consisted of retiform acral, purpuric plaques, or nodules. We report a case of perinuclear antineutrophil cytoplasmic antibody-associated vasculitis developed during treatment with PTU for Grave's disease. © The Author(s) 2014.

  7. Protein disulfide bond formation in the cytoplasm during oxidative stress.

    Science.gov (United States)

    Cumming, Robert C; Andon, Nancy L; Haynes, Paul A; Park, Minkyu; Fischer, Wolfgang H; Schubert, David

    2004-05-21

    The majority of disulfide-linked cytosolic proteins are thought to be enzymes that transiently form disulfide bonds while catalyzing oxidation-reduction (redox) processes. Recent evidence indicates that reactive oxygen species can act as signaling molecules by promoting the formation of disulfide bonds within or between select redox-sensitive proteins. However, few studies have attempted to examine global changes in disulfide bond formation following reactive oxygen species exposure. Here we isolate and identify disulfide-bonded proteins (DSBP) in a mammalian neuronal cell line (HT22) exposed to various oxidative insults by sequential nonreducing/reducing two-dimensional SDS-PAGE combined with mass spectrometry. By using this strategy, several known cytosolic DSBP, such as peroxiredoxins, thioredoxin reductase, nucleoside-diphosphate kinase, and ribonucleotide-diphosphate reductase, were identified. Unexpectedly, a large number of previously unknown DSBP were also found, including those involved in molecular chaperoning, translation, glycolysis, cytoskeletal structure, cell growth, and signal transduction. Treatment of cells with a wide range of hydrogen peroxide concentrations either promoted or inhibited disulfide bonding of select DSBP in a concentration-dependent manner. Decreasing the ratio of reduced to oxidized glutathione also promoted select disulfide bond formation within proteins from cytoplasmic extracts. In addition, an epitope-tagged version of the molecular chaperone HSP70 forms mixed disulfides with both beta4-spectrin and adenomatous polyposis coli protein in the cytosol. Our findings indicate that disulfide bond formation within families of cytoplasmic proteins is dependent on the nature of the oxidative insult and may provide a common mechanism used to control multiple physiological processes.

  8. Amelioration of neurodegenerative diseases by cell death-induced cytoplasmic delivery of humanin.

    Science.gov (United States)

    Park, Tae-Yoon; Kim, Seung-Hyung; Shin, Yoon-Chul; Lee, Nae-Hyun; Lee, Rae-Kyung Christina; Shim, Jae-Hyuck; Glimcher, Laurie H; Mook-Jung, Inhee; Cheong, Eunji; Kim, Won-Ki; Honda, Fumiko; Morio, Tomohiro; Lim, Jong-Soon; Lee, Sang-Kyou

    2013-03-28

    Inhibition of the early intracellular event that triggers neurodegenerative cascades and reversal of neuronal cell death are essential for effective treatment of Alzheimer's disease (AD). In this study, a novel therapeutic for AD, a transducible humanin with an extended caspase-3 cleavage sequence (tHN-C3), was developed and showed multiple mechanisms of therapeutic action. These included targeted delivery of anti-apoptotic protein humanin through the blood-brain barrier (BBB) to neuronal cells, specific inhibition of caspase-3 activation to inhibit the early triggering of AD progression, and delivery of humanin into the cytoplasm of neuronal cells undergoing apoptosis where it exerts its anti-apoptotic functions effectively. The tHN-C3 prevented neuronal cell death induced by H2O2, or soluble Aβ42, via Bax binding. In animal models of AD induced by amyloid beta, in Tg2576 mice, and in the rat middle cerebral artery occlusion model of stroke, tHN-C3 effectively prevented neuronal cell death, inflammatory cell infiltration into the brain, and improved cognitive memory. The therapeutic effectiveness of tHN-C3 was comparable to that of Aricept, a clinically approved drug for AD treatment. Therefore, tHN-C3 may be a new remedy with multiple therapeutic functions targeting the early and late stages of neurodegeneration in AD and other brain injuries. Copyright © 2013 Elsevier B.V. All rights reserved.

  9. Water diffusion in cytoplasmic streaming in Elodea internodal cells under the effect of antimitotic agents.

    Science.gov (United States)

    Vorob'ev, Vladimir N; Anisimov, Alexander V; Dautova, Nailya R

    2008-07-01

    The translational displacement of the cytoplasmic water in Elodea stem cells resulting from protein motor activity was measured using the NMR method. A 24-h treatment with vincristine results in a reduction of the translational displacement of the cytoplasmic water. With a constant cytoplasmic streaming velocity, the dynamics of the translational displacement of the cytoplasmic water under the effect of taxol are characterized by a continuous increase at a concentration of 0.05 mM, and reaching a plateau at a concentration of 0.5 mM.

  10. The cytoplasmic cage domain of the mechanosensitive channel MscS is a sensor of macromolecular crowding.

    Science.gov (United States)

    Rowe, Ian; Anishkin, Andriy; Kamaraju, Kishore; Yoshimura, Kenjiro; Sukharev, Sergei

    2014-05-01

    and crowding pressure. The cage, therefore, provides feedback on the increasing crowding that disengages the gate and prevents excessive draining and condensation of the cytoplasm. We discuss the structural mechanics of cells surrounded by an elastic cell wall where this MscS-specific feedback mechanism may be necessary.

  11. Cytoplasmic regionalization in starfish oocyte occurrence and localization of cytoplasmic determinants responsible for the formation of archenteron and primary mesenchyme in starfish ( asterias amurensis) oocytes

    Science.gov (United States)

    Zhang, Shicui; Wu, Xianhan; Zhou, Jing; Wang, Renxue; Wu, Shangqin

    1990-09-01

    Starfish oocytes with intact germinal vesicles (GVs) were cut along desired planes with glass needles or ligated using silk thread loops into two parts and allowed to mature in vitro, and inseminated. The experimental results showed that (1) only the parts with GVs or partial GV contents (PGVCs) cleaved, those without any GV materials did not; but nucleated and non-nucleated fragments cut from mature eggs were able to divide; (2) the development of animal parts of oocytes containing GVs or PGVCs was like that of animal fragments of matured oocytes with female pronuclei; most of them gave rise to permanent blastulae, and just a few formed ectodermal vesicles with a little primary mesenchyme; (3) a large part of vegetal fragments with GVs or PGVCs, and the vegetal parts of mature eggs without female pronuclei developed into small but normal embryos; (4) the fragments containing GVs or PGVCs obtained from the oocytes along a plane parallel to the animal-vegetal (A-V) axis developed as normally as the halves (with or without female pronuclei) severed from mature eggs along the same axis. Based on the data above, it was concluded that (1) the non-chromatin materials in the oocyte GVs are indispensable for successful fertilization and cleavage of starfish eggs; (2) some factor (s) located asymmetrically in the vegetal hemispheres of starfish oocytes is (are) responsible for formation of the archenteron and primary mesenchyme. It is evident from the above findings that the oocyte cytoplasm of the starfish had already regionalized before the GV break-down.

  12. Anti-neutrophil cytoplasmic antibodies stimulate release of neutrophil microparticles.

    LENUS (Irish Health Repository)

    Hong, Ying

    2012-01-01

    The mechanisms by which anti-neutrophil cytoplasmic antibodies (ANCAs) may contribute to the pathogenesis of ANCA-associated vasculitis are not well understood. In this study, both polyclonal ANCAs isolated from patients and chimeric proteinase 3-ANCA induced the release of neutrophil microparticles from primed neutrophils. These microparticles expressed a variety of markers, including the ANCA autoantigens proteinase 3 and myeloperoxidase. They bound endothelial cells via a CD18-mediated mechanism and induced an increase in endothelial intercellular adhesion molecule-1 expression, production of endothelial reactive oxygen species, and release of endothelial IL-6 and IL-8. Removal of the neutrophil microparticles by filtration or inhibition of reactive oxygen species production with antioxidants abolished microparticle-mediated endothelial activation. In addition, these microparticles promoted the generation of thrombin. In vivo, we detected more neutrophil microparticles in the plasma of children with ANCA-associated vasculitis compared with that in healthy controls or those with inactive vasculitis. Taken together, these results support a role for neutrophil microparticles in the pathogenesis of ANCA-associated vasculitis, potentially providing a target for future therapeutics.

  13. Cytoplasmic lipid bodies in eosinophils: Central roles in eicosanoid generation

    Directory of Open Access Journals (Sweden)

    Peter F Weller

    1997-01-01

    Full Text Available Lipid bodies are non-membrane bound, lipid-rich cytoplasmic inclusions that form in diverse cell types. Characteristically, lipid body numbers increase when eosinophils and other leukocytes are participating in inflammatory processes. Moreover, lipid bodies are numerous in other sites active in eicosanoid formation, such as the amnion epithelium at parturition. Our interests in lipid bodies relate to the roles that these structures play in arachidonate metabolism by eosinophils and other leukocytes involved in inflammation. Findings indicate that lipid bodies in these leukocytes can function as intracellular domains that are both depots of esterified arachidonate and sites at which regulated enzymatic events relevant to arachidonate metabolism can occur. Lipid bodies are discrete intracellular structures whose formation is specifically inducible early, whose increased numbers correlate with the 'priming' responses of leukocytes to form enhanced amounts of both cyclooxygenase- and lipoxygenase-derived eicosanoids and whose inhibition of formation correlates with reduced synthesis of these eicosanoids. Inhibition of lipid body formation represents a novel pharmacologic target to block the formation of eicosanoid mediators of inflammation.

  14. Gram's Stain Does Not Cross the Bacterial Cytoplasmic Membrane.

    Science.gov (United States)

    Wilhelm, Michael J; Sheffield, Joel B; Sharifian Gh, Mohammad; Wu, Yajing; Spahr, Christian; Gonella, Grazia; Xu, Bolei; Dai, Hai-Lung

    2015-07-17

    For well over a century, Hans Christian Gram's famous staining protocol has been the standard go-to diagnostic for characterizing unknown bacteria. Despite continuous and ubiquitous use, we now demonstrate that the current understanding of the molecular mechanism for this differential stain is largely incorrect. Using the fully complementary time-resolved methods: second-harmonic light-scattering and bright-field transmission microscopy, we present a real-time and membrane specific quantitative characterization of the bacterial uptake of crystal-violet (CV), the dye used in Gram's protocol. Our observations contradict the currently accepted mechanism which depicts that, for both Gram-negative and Gram-positive bacteria, CV readily traverses the peptidoglycan mesh (PM) and cytoplasmic membrane (CM) before equilibrating within the cytosol. We find that not only is CV unable to traverse the CM but, on the time-scale of the Gram-stain procedure, CV is kinetically trapped within the PM. Our results indicate that CV, rather than dyes which rapidly traverse the PM, is uniquely suited as the Gram stain.

  15. Novel clinical and diagnostic aspects of antineutrophil cytoplasmic antibodies.

    Science.gov (United States)

    Schulte-Pelkum, Johannes; Radice, Antonella; Norman, Gary L; Lόpez Hoyos, Marcos; Lakos, Gabriella; Buchner, Carol; Musset, Lucile; Miyara, Makoto; Stinton, Laura; Mahler, Michael

    2014-01-01

    Antineutrophil cytoplasmic antibodies (ANCA) are the serological hallmark of some idiopathic systemic vasculitides. Besides the investigation of ANCA-associated vasculitis (AAV) and constant effort for a standardized nomenclature and classification of the AAV, a main focus of research during the last few years has been to constantly improve the performance of enzyme immunoassays. With the latest so called third generation ELISA, this goal seemed to be fulfilled. The International Consensus Statement on Testing and Reporting of ANCA gave recommendations for standardized strategies for the serological diagnosis of ANCA. New developments now target the system immanent drawbacks of the respective diagnostic methods, be it the need for batching and the long time to result for ELISA, or the high likelihood of error and subjectivity of indirect immunofluorescence (IIF). Random access technology and multiplexing for solid phase assays as well as digital imaging for IIF are tools which may help to expedite and simplify routine diagnostics in the lab and in emergency settings. Recent findings indicate that PR3-ANCA have clinical utility beyond the diagnosis of AAV. PR3-ANCA can also serve as an aid for the differentiation between ulcerative colitis (UC) and Crohn's disease (CrD) and the stratification of UC patients. This review provides a detailed review of what is known about ANCA and highlights the latest research and state-of-the-art developments in this area.

  16. Cytoplasmic sphingosine-1-phosphate pathway modulates neuronal autophagy.

    Science.gov (United States)

    Moruno Manchon, Jose Felix; Uzor, Ndidi-Ese; Dabaghian, Yuri; Furr-Stimming, Erin E; Finkbeiner, Steven; Tsvetkov, Andrey S

    2015-10-19

    Autophagy is an important homeostatic mechanism that eliminates long-lived proteins, protein aggregates and damaged organelles. Its dysregulation is involved in many neurodegenerative disorders. Autophagy is therefore a promising target for blunting neurodegeneration. We searched for novel autophagic pathways in primary neurons and identified the cytosolic sphingosine-1-phosphate (S1P) pathway as a regulator of neuronal autophagy. S1P, a bioactive lipid generated by sphingosine kinase 1 (SK1) in the cytoplasm, is implicated in cell survival. We found that SK1 enhances flux through autophagy and that S1P-metabolizing enzymes decrease this flux. When autophagy is stimulated, SK1 relocalizes to endosomes/autophagosomes in neurons. Expression of a dominant-negative form of SK1 inhibits autophagosome synthesis. In a neuron model of Huntington's disease, pharmacologically inhibiting S1P-lyase protected neurons from mutant huntingtin-induced neurotoxicity. These results identify the S1P pathway as a novel regulator of neuronal autophagy and provide a new target for developing therapies for neurodegenerative disorders.

  17. In silico classification of proteins from acidic and neutral cytoplasms.

    Directory of Open Access Journals (Sweden)

    Yaping Fang

    Full Text Available Protein acidostability is a common problem in biopharmaceutical and other industries. However, it remains a great challenge to engineer proteins for enhanced acidostability because our knowledge of protein acidostabilization is still very limited. In this paper, we present a comparative study of proteins from bacteria with acidic (AP and neutral cytoplasms (NP using an integrated statistical and machine learning approach. We construct a set of 393 non-redundant AP-NP ortholog pairs and calculate a total of 889 sequence based features for these proteins. The pairwise alignments of these ortholog pairs are used to build a residue substitution propensity matrix between APs and NPs. We use Gini importance provided by the Random Forest algorithm to rank the relative importance of these features. A scoring function using the 10 most significant features is developed and optimized using a hill climbing algorithm. The accuracy of the score function is 86.01% in predicting AP-NP ortholog pairs and is 76.65% in predicting non-ortholog AP-NP pairs, suggesting that there are significant differences between APs and NPs which can be used to predict relative acidostability of proteins. The overall trends uncovered in the study can be used as general guidelines for designing acidostable proteins. To best of our knowledge, this work represents the first systematic comparative study of the acidostable proteins and their non-acidostable orthologs.

  18. Nematode development after removal of egg cytoplasm: absence of localized unbound determinants.

    Science.gov (United States)

    Laufer, J S; von Ehrenstein, G

    1981-01-23

    Embryos of Caenorhabditis elegans develop into fertile adults after cell fragments, containing presumptive cytoplasm of somatic and germ line precursors, are extruded from uncleaved eggs or early blastomeres through laser-induced holes in the eggshells. This suggests that the determinate development of this worm is not dependent on the prelocalization of determinants in specific regions of the egg cytoplasm.

  19. High viscosity and anisotropy characterize the cytoplasm of fungal dormant stress resistant spores

    NARCIS (Netherlands)

    Dijksterhuis, J.; Nijsse, J.; Hoekstra, F.A.; Golovina, E.A.

    2007-01-01

    Ascospores of the fungus Talaromyces macrosporus are dormant and extremely stress resistant, whereas fungal conidia¿the main airborne vehicles of distribution¿are not. Here, physical parameters of the cytoplasm of these types of spores were compared. Cytoplasmic viscosity and level of anisotropy as

  20. TRIM5α association with cytoplasmic bodies is not required for antiretroviral activity

    International Nuclear Information System (INIS)

    Song, Byeongwoon; Diaz-Griffero, Felipe; Park, Do Hyun; Rogers, Thomas; Stremlau, Matthew; Sodroski, Joseph

    2005-01-01

    The tripartite motif (TRIM) protein, TRIM5α, restricts infection by particular retroviruses. Many TRIM proteins form cytoplasmic bodies of unknown function. We investigated the relationship between cytoplasmic body formation and the structure and antiretroviral activity of TRIM5α. In addition to diffuse cytoplasmic staining, the TRIM5α proteins from several primate species were located in cytoplasmic bodies of different sizes; by contrast, TRIM5α from spider monkeys did not form cytoplasmic bodies. Despite these differences, all of the TRIM5α proteins exhibited the ability to restrict infection by particular retroviruses. Treatment of cells with geldanamycin, an Hsp90 inhibitor, resulted in disappearance or reduction of the TRIM5α-associated cytoplasmic bodies, yet exerted little effect on the restriction of retroviral infection. Studies of green fluorescent protein-TRIM5α fusion proteins indicated that no TRIM5α domain is specifically required for association with cytoplasmic bodies. Apparently, the formation of cytoplasmic bodies is not required for the antiretroviral activity of TRIM5α

  1. Effect of wild Helianthus cytoplasms on agronomic and oil characteristics of cultivated sunflower (H. annuus L.)

    Science.gov (United States)

    Sunflower (Helianthus annuus L.) productions reliance on a single source of cytoplasmic male-sterility, PET1, derived from H. petiolaris Nutt., makes the crop genetically vulnerable. Twenty diverse cytoplasmic substitution lines from annual and perennial wild species were compared with the inbred li...

  2. [Wegener's granulomatosis with anti-neutrophil cytoplasmic antibodies against anti-cathepsin G antigen].

    Science.gov (United States)

    Ocaña Pérez, E; Peña Casas, A M; del Campo Muñoz, T; Avila Casas, A; Luque Barona, R

    2013-12-01

    Wegener's granulomatosis belongs to the group of small vessel vasculitis associated with anti-neutrophil cytoplasmic antibodies characterized by granulomatous inflammation and necrotising vasculitis in various organs with particular involvement of the upper and lower respiratory tracts and kidneys. Wegener's granulomatosis is a rare disorder in childhood and early diagnosis of this disease is critical to the long-term prognosis of the disease. The presence of positive cytoplasmic antineutrophil cytoplasmic antibody staining or a high titre of proteinase 3 antibodies were added as new criteria of vasculitis in childhood. This article presents a case of Wegener's granulomatosis, with the presence of anti-neutrophil cytoplasm antibodies with cytoplasmic pattern with absence of anti-proteinase 3 antibodies and presence of high levels of anti-cathepsin G antibodies, rarely described in Wegener's granulomatosis. Copyright © 2012 Asociación Española de Pediatría. Published by Elsevier Espana. All rights reserved.

  3. ROS-activated ATM-dependent phosphorylation of cytoplasmic substrates identified by large scale phosphoproteomics screen

    DEFF Research Database (Denmark)

    Kozlov, Sergei V; Waardenberg, Ashley J; Engholm-Keller, Kasper

    2016-01-01

    checkpoints, initiating DNA repair and regulating gene expression. ATM kinase can be activated by a variety of stimuli, including oxidative stress. Here we confirmed activation of cytoplasmic ATM by autophosphorylation at multiple sites. Then we employed a global quantitative phosphoproteomics approach...... to identify cytoplasmic proteins altered in their phosphorylation state in control and A-T (ataxia-telangiectasia) cells in response to oxidative damage. We demonstrated that ATM was activated by oxidative damage in the cytoplasm as well as in the nucleus and identified a total of 9,833 phosphorylation sites......-dependent after H2O2 exposure and another protein (S100A11) demonstrated ATM-dependence for translocation from the cytoplasm to the nucleus. These data provide new insights into the activation of ATM by oxidative stress through identification of novel substrates for ATM in the cytoplasm. 2....

  4. Tubulin dynamics during the cytoplasmic cohesiveness cycle in artificially activated sea urchin eggs.

    Science.gov (United States)

    Coffe, G; Foucault, G; Raymond, M N; Pudles, J

    1983-12-01

    Sedimentation studies and [3H]colchicine-binding assays have demonstrated a relationship between the cytoplasmic cohesiveness cycles and the changes in tubulin organization in Paracentrotus lividus eggs activated by 2.5 mM procaine. The same amount of tubulin (20-25% of the total egg tubulin) is involved in these cyclic process and appears to undergo polymerization and depolymerization cycles. Electron microscopy studies reveal that the microtubules formed during these cytoplasmic cohesiveness cycles are under a particulate form which is sedimentable at low speed. Activation experiments carried out in the presence of cytochalasin B (CB) show that the increase in the cytoplasmic cohesiveness is highly reduced while tubulin polymerization and depolymerization cycles and pronuclear centration are not affected. Although tubulin or actin polymerization can be independently triggered in procaine-activated eggs, the increase in cytoplasmic cohesiveness requires the polymerization of both proteins. However, the cytoplasmic cohesiveness cycles appear to be regulated by tubulin polymerization and depolymerization cycles.

  5. Effects of modification of the HIV-1 Env cytoplasmic tail on immunogenicity of VLP vaccines.

    Science.gov (United States)

    Vzorov, Andrei N; Wang, Li; Chen, Jianjun; Wang, Bao-Zhong; Compans, Richard W

    2016-02-01

    We investigated the effects on assembly and antigenic properties of specific modifications of the transmembrane spanning (TMS) and cytoplasmic tail (CT) domains of HIV-1 Env from a transmitted/founder (T/F) ZM53 Env glycoprotein. A construct containing a short version of the TMS domain derived from the mouse mammary tumor virus (MMTV) Env with or without a GCN4 trimerization sequence in the CT exhibited the highest levels of incorporation into VLPs and induced the highest titers of anti-Env IgG immune responses in a VLP context. Sera from guinea pigs immunized by VLPs with high Env content, and containing the CT trimerization sequence, had increased neutralization activity and antibody avidity. A cross-clade prime-boost regimen with clade B SF162 or clade C ZM53 Env DNA priming and boosting with VLPs containing modified ZM53 Env further enhanced these immune responses. The modified VLPs demonstrate improved potential as HIV-1 vaccine antigens. Copyright © 2016. Published by Elsevier Inc.

  6. Cytoplasmic p21 is a potential predictor for cisplatin sensitivity in ovarian cancer

    International Nuclear Information System (INIS)

    Xia, Xi; Weng, Yanjie; Liao, Shujie; Han, Zhiqiang; Liu, Ronghua; Zhu, Tao; Wang, Shixuan; Xu, Gang; Meng, Li; Zhou, Jianfeng; Ma, Ding; Ma, Quanfu; Li, Xiao; Ji, Teng; Chen, Pingbo; Xu, Hongbin; Li, Kezhen; Fang, Yong; Weng, Danhui

    2011-01-01

    P21 (WAF1/Cip1) binds to cyclin-dependent kinase complexes and inhibits their activities. It was originally described as an inhibitor of cancer cell proliferation. However, many recent studies have shown that p21 promotes tumor progression when accumulated in the cell cytoplasm. So far, little is known about the correlation between cytoplasmic p21 and drug resistance. This study was aimed to investigate the role of p21 in the cisplatin resistance of ovarian cancer. RT-PCR, western blot and immunofluorescence were used to detect p21 expression and location in cisplatin-resistant ovarian cancer cell line C13* and its parental line OV2008. Regulation of cytoplasmic p21 was performed through transfection of p21 siRNA, Akt2 shRNA and Akt2 constitutively active vector in the two cell lines; their effects on cisplatin-induced apoptosis were evaluated by flow cytometry. Tumor tissue sections of clinical samples were analyzed by immunohistochemistry. p21 predominantly localizes to the cytoplasm in C13* compared to OV2008. Persistent exposure to low dose cisplatin in OV2008 leads to p21 translocation from nuclear to cytoplasm, while it had not impact on p21 localization in C13*. Knockdown of cytoplasmic p21 by p21 siRNA transfection in C13* notably increased cisplatin-induced apoptosis through activation of caspase 3. Inhibition of p21 translocation into the cytoplasm by transfection of Akt2 shRNA into C13* cells significantly increased cisplatin-induced apoptosis, while induction of p21 translocation into the cytoplasm by transfection of constitutively active Akt2 in OV2008 enhanced the resistance to cisplatin. Immunohistochemical analysis of clinical ovarian tumor tissues demonstrated that cytoplasmic p21 was negatively correlated with the response to cisplatin based treatment. Cytoplasmic p21 is a novel biomarker of cisplatin resistance and it may represent a potential therapeutic target for ovarian tumors that are refractory to conventional treatment

  7. Agronomic Performance of Flue-Cured Tobacco F1 Hybrids Obtained with Different Sources of Male Sterile Cytoplasm

    Directory of Open Access Journals (Sweden)

    Berbec A

    2014-12-01

    Full Text Available Four cytoplasmic male sterile (cms F1flue-cured hybrids cv. Wiaelica × cv. Virginia Golta (VG, the male fertile analogue and the parental varieties were tested at two locations in Poland in a replicated field trial. The cms sources in the hybrids wereN. suaveolens,N. amplexicaulis,N. bigeloviiand aN. tabacumcms mutant. Under the slight to moderate pressure from black root rot present at the trial sites the hybrids showed a moderate tolerance of the disease characteristic of VG as opposed to medium strong susceptibility of Wislica. Apart from the effect of black root rot tolerance the vegetative vigor of the hybrids (plant height, leaf size, earliness was affected by cytoplasm source. The F1hybrid withN. suaveolens cytoplasm flowered approximately three days later than the remaining hybrids. Of the cms hybrids tested cmsN. bigelovii produced the tallest plants with largest mid-position leaves. Yields of cured leaves were largely influenced by black root rot and were generally higher in VG and in the hybrids than in Wislica. Leaf yields and curability were generally little affected by cms source under low pressure from black root rot. At the site with a relatively high level of black root rot infestation the yields of cmsN. suaveolens were slightly lower but the percentage of light grades slightly higher compared to those of other cms hybrids. CmsN. suaveolens was the best hybrid in terms of money returns at the low black root rot field but it was the poorest hybrid performer under high pressure from the disease. Contents of nitrogen, sugars, nicotine and ash was little affected by source of cms. There was an increased incidence of potato virus Y (PVY and white spots in cmsN. suaveolens and, to a lesser extent, in cmsN. bigelovii as compared to the remaining disease-free entries.

  8. Urinary Biomarkers in Relapsing Antineutrophil Cytoplasmic Antibody-associated Vasculitis

    Science.gov (United States)

    Lieberthal, Jason G.; Cuthbertson, David; Carette, Simon; Hoffman, Gary S.; Khalidi, Nader A.; Koening, Curry L.; Langford, Carol A.; Maksimowicz-McKinnon, Kathleen; Seo, Philip; Specks, Ulrich; Ytterberg, Steven R.; Merkel, Peter A.; Monach, Paul A.

    2015-01-01

    Objective Glomerulonephritis (GN) is common in antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV), but tools for early detection of renal involvement are imperfect. We investigated 4 urinary proteins as markers of active renal AAV: alpha-1 acid glycoprotein (AGP), kidney injury molecule-1 (KIM-1), monocyte chemoattractant protein-1 (MCP-1), and neutrophil gelatinase-associated lipocalin (NGAL). Methods Patients with active renal AAV (n = 20), active nonrenal AAV (n = 16), and AAV in longterm remission (n = 14) were identified within a longitudinal cohort. Urinary biomarker concentrations (by ELISA) were normalized for urine creatinine. Marker levels during active AAV were compared to baseline remission levels (from 1–4 visits) for each patient. Areas under receiver-operating characteristic curves (AUC), sensitivities, specificities, and likelihood ratios (LR) comparing disease states were calculated. Results Baseline biomarker levels varied among patients. All 4 markers increased during renal flares (p < 0.05). MCP-1 discriminated best between active renal disease and remission: a 1.3-fold increase in MCP-1 had 94% sensitivity and 89% specificity for active renal disease (AUC = 0.93, positive LR 8.5, negative LR 0.07). Increased MCP-1 also characterized 50% of apparently nonrenal flares. Change in AGP, KIM-1, or NGAL showed more modest ability to distinguish active renal disease from remission (AUC 0.71–0.75). Hematuria was noted in 83% of active renal episodes, but also 43% of nonrenal flares and 25% of remission samples. Conclusion Either urinary MCP-1 is not specific for GN in AAV, or it identifies early GN not detected by standard assessment and thus has potential to improve care. A followup study with kidney biopsy as the gold standard is needed. PMID:23547217

  9. Multi-Scale Characean Experimental System: From Electrophysiology of Membrane Transporters to Cell-to-Cell Connectivity, Cytoplasmic Streaming and Auxin Metabolism

    Science.gov (United States)

    Beilby, Mary J.

    2016-01-01

    The morphology of characean algae could be mistaken for a higher plant: stem-like axes with leaf-like branchlets anchored in the soil by root-like rhizoids. However, all of these structures are made up of giant multinucleate cells separated by multicellular nodal complexes. The excised internodal cells survive long enough for the nodes to give rise to new thallus. The size of the internodes and their thick cytoplasmic layer minimize impalement injury and allow specific micro-electrode placement. The cell structure can be manipulated by centrifugation, perfusion of cell contents or creation of cytoplasmic droplets, allowing access to both vacuolar and cytoplasmic compartments and both sides of the cell membranes. Thousands of electrical measurements on intact or altered cells and cytoplasmic droplets laid down basis to modern plant electrophysiology. Furthermore, the giant internodal cells and whole thalli facilitate research into many other plant properties. As nutrients have to be transported from rhizoids to growing parts of the thallus and hormonal signals need to pass from cell to cell, Characeae possess very fast cytoplasmic streaming. The mechanism was resolved in the characean model. Plasmodesmata between the internodal cells and nodal complexes facilitate transport of ions, nutrients and photosynthates across the nodes. The internal structure was found to be similar to those of higher plants. Recent experiments suggest a strong circadian influence on metabolic pathways producing indole-3-acetic acid (IAA) and serotonin/melatonin. The review will discuss the impact of the characean models arising from fragments of cells, single cells, cell-to-cell transport or whole thalli on understanding of plant evolution and physiology. PMID:27504112

  10. Skeletal Phenotype of Transgenic Mice Expressing the Beta1 Integrin Cytoplasmic Tail In Osteoblasts

    Science.gov (United States)

    Globus, R. K.; vanderMeulen, M. C. H.; Damsky, D.; Kim, J.-B.; Amblard, D.; Amblard, D.; Nishimura, Y.; Almeida, E.; Iwaniec, U. T.; Wronski, T. J.; hide

    2002-01-01

    To define the physiologic role of beta1 integrin in bone formation and mechanical loading, transgenic mice were generated by expressing the cytoplasmic tall and transmembrane domain of Beta1 integrin under the control of the osteocalcin promoter. In cultured cells, this truncated fragment of Beta1 can act as a dominant negative. Previously, the matrix of calvariae was shown to be abnormal in transgenic (TG) compared to wildtype (WT) mice. In this study, we analyzed appendicular bone in TG and WT, male and female mice at 14, 35, 63, 90 and 365 days old (n=8-12/gp). To assess beta1 integrin function in mechanical loading, a pilot study using hindlimb unloading by tail suspension was performed. 35d old TG and WT females were hindlimb unloaded for 4 wks (n=3-5). Body mass, bone mineral content, histomorphometric (distal femur) and biomechanical parameters were analyzed. Statistical significance (P less than.05) was defined by ANOVA using the Tukey-Kramer post-hoc test. We confirmed transgene expression by immunoprecipitating then immunoblotting bone lysates using an antibody against the beta1 tail. Body masses of TG mice at 63, 90 and 365d old were greater (16-25%) than WT. Some TG female mice at 365d appeared obese; mean abdominal fat mass was 415% greater in TG than WT mice. Tibiae were longer (5-7%) in TG than WT mice at 63 and 90d. Tibial mineral mass of 35d males was 7% lower in TG than WT mice, but at 63d was 21% higher. The % osteoblast surface in 35d TG mice was 20% higher than WT, and at 63d was 17% lower, while % osteoclast surface did not differ. In 365d mice, cancellous bone volume (125%) and endocortical mineral apposition rate (40%) were greater in TG than WT males but not females. In WT mice, hindlimb unloading caused a reduction in mineral mass of tibiae (-20%) and lumbar vertebrae (-22%) relative to normally loaded controls. Surprisingly, hindlimb unloading also caused a relative reduction (-13%) in humerus mass. The effects of hindlimb unloading on

  11. Cytoplasmic vacuoles: clue in the fine-needle aspiration diagnosis of melanoma.

    Science.gov (United States)

    Chen, Liansong; Raza, Anwar S; Simental, Raul G; Iverson, Kiva T; Cobb, Camilla J

    2014-02-01

    For the fine-needle aspiration (FNA) diagnosis of melanoma recognition of characteristic morphologic features is key. In our practice we noted that cytoplasmic vacuoles in Romanowsky-stained FNA smears of melanoma appeared to be a frequent finding. To investigate this premise, we examined 36 consecutive melanoma FNA cases that had both adequate Papanicolaou-stained and Romanowsky-stained smears in an effort to determine the prevalence, and thereby, the potential diagnostic utility of cytoplasmic vacuoles in the diagnosis of melanoma compared to established cytomorphologic parameters of melanoma. We found pigment in 21 cases (58%) and plasmacytoid cells in 28 cases (78%), and in all 36 cases (100%) we found bi/multinucleation, intranuclear cytoplasmic inclusions and prominent nucleoli. Cytoplasmic vacuoles were present in 30 cases (83%) and were found in air-dried Romanowsky-stained smears only. These findings suggest that in FNA smears of melanoma cytoplasmic vacuoles in Romanowsky-stained smears have prevalence and potential diagnostic utility that are comparable to widely recognized cytomorphologic features of melanoma. Numerous articles have focused on the cytomorphologic criteria for the FNA diagnosis of melanoma, but few have noted the presence of cytoplasmic vacuoles in FNA smears of melanoma, and rare reports suggest this finding to be a useful clue to the FNA diagnosis of melanoma. This report appears to be the first to focus on the prevalence and potential diagnostic utility of cytoplasmic vacuoles in FNA samples of melanoma. Copyright © 2013 Society of Plastics Engineers.

  12. When is a carrier not a membrane carrier? The cytoplasmic transport of amphipathic molecules.

    Science.gov (United States)

    Weisiger, R A

    1996-11-01

    After entering the cell, small molecules must penetrate the cytoplasm before they are metabolized, excreted, or can convey information to the cell nucleus. Without efficient cytoplasmic transport, most such molecules would efflux back out of the cell before they could reach their targets. Cytoplasmic movement of amphipathic molecules (e.g., long-chain fatty acids, bilirubin, bile acids) is greatly slowed by their tendency to bind intracellular structures. Soluble cytoplasmic binding proteins reduce this binding by increasing the aqueous solubility of their ligands. These soluble carriers catalyze the transport of hydrophobic molecules across hydrophilic water layers, just as membrane carriers catalyze the transport of hydrophilic molecules across the hydrophobic membrane core. They even display the kinetic features of carrier-mediated transport, including saturation, mutual competition between similar molecules, and countertransport. Recent data suggest that amphipathic molecules cross the cytoplasm very slowly, with apparent diffusion constants 10(2) to 10(4) times smaller than in water. By modulating the rate of cytoplasmic transport, cytosolic binding proteins may regulate transport and metabolism of amphipathic molecules. Storage diseases may cause hepatocellular dysfunction by disrupting normal cytoplasmic transport.

  13. Single-molecule study of molecular mobility in the cytoplasm of Escherichia coli

    Science.gov (United States)

    Lill, Yoriko; Kaserer, Wallace A.; Newton, Salete M.; Lill, Markus; Klebba, Phillip E.; Ritchie, Ken

    2012-08-01

    The cytoplasm of bacterial cells is filled with individual molecules and molecular complexes that rely on diffusion to bring them together for interaction. The mobility of molecules in the cytoplasm has been characterized by several techniques mainly using fluorescent probes and ensemble methods. In order to probe the microenvrionment inside the cytoplasm as viewed by an individual molecule, we have studied single green fluorescent proteins (GFPs) diffusing in the cytoplasm of Escherichia coli cells at observation at rates ranging from 60 to 1000 Hz. Over long times the diffusion shows confinement due to the geometry of the cells themselves. A simulation in model cells using the actual distribution of cell sizes found in the experiments describes accurately the experimental results as well as reveals a short time diffusion coefficient that agrees well with that determined by ensemble methods. Higher short time diffusion coefficients can be obtained by filling the simulated cell with small spheres modeling cytoplasmic molecules and, depending on the density of particles included in the modeled cytoplasm, can approach the diffusion coefficient of GFPs found in water. Thus, single-molecule tracking combined with analysis using simple simulation of Brownian motion is able to reveal the main contributors to the GFP mobility in the cytoplasm of E. coli.

  14. Nup98 Localizes to Both Nuclear and Cytoplasmic Sides of the Nuclear Pore and Binds to Two Distinct Nucleoporin Subcomplexes

    Science.gov (United States)

    Griffis, Eric R.; Xu, Songli; Powers, Maureen A.

    2003-01-01

    The vertebrate nuclear pore is an enormous structure that spans the double membrane of the nuclear envelope. In yeast, most nucleoporins are found symmetrically on both the nuclear and cytoplasmic sides of the structure. However, in vertebrates most nucleoporins have been localized exclusively to one side of the nuclear pore. Herein, we show, by immunofluorescence and immunoelectron microscopy, that Nup98 is found on both sides of the pore complex. Additionally, we find that the pore-targeting domain of Nup98 interacts directly with the cytoplasmic nucleoporin Nup88, a component of the Nup214, Nup88, Nup62 subcomplex. Nup98 was previously described to interact with the nuclear-oriented Nup160, 133, 107, 96 complex through direct binding to Nup96. Interestingly, the same site within Nup98 is involved in binding to both Nup88 and Nup96. Autoproteolytic cleavage of the Nup98 C terminus is required for both of these binding interactions. When cleavage is blocked by a point mutation, a minimal eight amino acids downstream of the cleavage site is sufficient to prevent most binding to either Nup96 or Nup88. Thus, Nup98 interacts with both faces of the nuclear pore, a localization in keeping with its previously described nucleocytoplasmic shuttling activity. PMID:12589057

  15. Cytoplasmic membrane is the target organelle for transition metal mediated damage induced by paraquat in Escherichia coli

    International Nuclear Information System (INIS)

    Kohen, R.; Chevion, M.

    1988-01-01

    Bacterial survival indicates that copper or iron is an essential mediator in paraquat toxicity in Escherichia coli. In this study the authors have identified the cytoplasmic membrane as a target organelle in metal-mediated paraquat toxicity and have demonstrated the complete correlation of the membrane damage with the levels of adventitious copper (or iron). The extent of membrane damage was related by use of four parameters: (a) the level of cellular ATP, (b) the level of cellular potassium, (c) the cellular capacity to accumulate and retain radiolabeled leucine, and (d) the cellular integrity as reflected by transmission electron microscopy (TEM). Exposure of bacterial cells to a combination of paraquat and copper caused a marked decline in parameters a, b, and c. This decline was found to occur in parallel with, or even to precede, the sharp loss of survival of E. coli under the same conditions. Likewise, TEM micrographs clearly indicated alternations in cellular structure that possibly reflect sites of detachment of the cytoplasmic membrane from the bacterial capsule. In contradistinction, copper alone or paraquat alone could not bring about similar changes in cellular structure. These findings are in accord with the suggested site-specific metal-mediated Haber-Weiss mechanism for paraquat toxicity and support our notion that specific chelators of transition metals could reduce or prevent the biological deleterious effects of this herbicide

  16. Analysis of embryo, cytoplasmic and maternal genetic correlations ...

    Indian Academy of Sciences (India)

    2School of Agriculture and Food Science, Zhejiang A & F University, Linan, 311300, People's Republic of China. Abstract. Genetic correlations of nutrient quality traits including lysine, ...... spectroscopy (NIRS) enables the fast and accurate prediction of essential amino acid contents. 2. Results for wheat, barley, corn, triticale ...

  17. Analysis of embryo, cytoplasmic and maternal genetic correlations ...

    Indian Academy of Sciences (India)

    Genetic correlations of nutrient quality traits including lysine, methionine, leucine, isoleucine, phenylalanine, valine and threonine contents in rapeseed meal were analysed by the genetic model for quantitative traits of diploid plants using a diallel design with nine parents of Brassica napus L. These results indicated that the ...

  18. Nuclear translocation of the cytoplasmic domain of HB-EGF induces gastric cancer invasion

    Science.gov (United States)

    2012-01-01

    Background Membrane-anchored heparin-binding epidermal growth factor-like growth factor (proHB-EGF) yields soluble HB-EGF, which is an epidermal growth factor receptor (EGFR) ligand, and a carboxy-terminal fragment of HB-EGF (HB-EGF-CTF) after ectodomain shedding. We previously reported that HB-EGF-CTF and unshed proHB-EGF which has the cytoplasmic domain of proHB-EGF (HB-EGF-C), translocate from the plasma membrane to the nucleus and regulate cell cycle after shedding stimuli. However, the significance of nuclear exported HB-EGF-C in human gastric cancer is unclear. Methods We investigated the relationship between intracellular localization of HB-EGF-C and clinical outcome in 96 gastric cancer patients treated with gastrectomy. Moreover, we established stable gastric cancer cell lines overexpressing wild-type HB-EGF (wt-HB-EGF) and mutated HB-EGF (HB-EGF-mC), which prevented HB-EGF-C nuclear translocation after shedding. Cell motility between these 2 gastric cancer cell lines was investigated using a transwell invasion assay and a wound healing assay. Results Of the 96 gastric cancer cases, HB-EGF-C immunoreactivity was detected in both the nucleus and cytoplasm in 19 cases (19.8 %) and in the cytoplasm only in 25 cases (26.0 %). The nuclear immunoreactivity of HB-EGF-C was significantly increased in stage pT3/4 tumors compared with pT1/2 tumors (T1/2 vs. T3/4: 11.1 % vs. 36.4 %, P HB-EGF- and HB-EGF-mC-expressing cells significantly increased compared with control cells, but the growth of HB-EGF-mC-expressing cells was significantly decreased compared with wt-HB-EGF-expressing cells. Gastric cancer cell invasion obviously increased in wt-HB-EGF-expressing cells, but invasion in HB-EGF-mC-expressing cells showed a slight increase compared with control cells. Moreover, wt-HB-EGF overexpression increased the effectiveness of wound healing, but had no significant effect in HB-EGF-mC-expressing cells. Conclusions Both the function of HB-EGF as an EGFR ligand

  19. Inhibitiory properties of cytoplasmic extract of Lactobacilli isolated from common carp intestine on human chronic myelocytic leukemia K562 cell line: an in vitro study

    Directory of Open Access Journals (Sweden)

    Kabiri F

    2011-03-01

    Full Text Available "n Normal 0 false false false EN-US X-NONE AR-SA MicrosoftInternetExplorer4 st1":*{behavior:url(#ieooui } /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-qformat:yes; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:"Times New Roman"; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:Arial; mso-bidi-theme-font:minor-bidi;} Background: Lactobacillus species are genetically diverse groups of Lactic Acid Bacteria (LAB that have been introduced as probiotics, because of some characteristics such as their anti-tumor properties, helping the intestinal flora balance, production of antibiotics, stimulation of host immune response, etc. The aim of this study was to investigate the effects of cytoplasmic extraction and cell wall of Lactobacillus species isolated from the intestine of common carp on human chronic myelocytic leukemia or K562 cancer cell lines."n"nMethods: The intestinal contents of 115 common carp captured from the natural resources of West Azerbaijan province in Iran were examined for LAB. After isolation, the identification of Lactobacilli was done according to traditional and molecular bacteriological tests. Subsequently, a suspension of each bacterium was prepared and the protein content of the cytoplasm was extracted. Cell wall disintegration was done by cell lysis buffer and sonication. The effects of cytoplasmic extraction and cell wall on K562 cell line proliferation were investigated by MTT assays."n"nResults: The cytoplasmic extraction of the isolated Lactobacilli had significant (p<0.05 anti

  20. Genetic and genotype environment interaction effects for the content ...

    Indian Academy of Sciences (India)

    In view of the paucity of genetic information available on essential amino acids in indica rice, we estimated the genetic main effects and genotype × environment (G × E) interaction effects on the content of essential amino acids. Nine cytoplasmic male sterile lines as females and five restorer lines as males were introduced ...

  1. Anticorpos contra o citoplasma de neutrófilos Antineutrophil cytoplasmic antibodies

    Directory of Open Access Journals (Sweden)

    Ari Stiel Radu

    2005-07-01

    Full Text Available A descoberta do marcador sorológico denominado anticorpo anticitoplasma de neutrófilos revolucionou o diagnóstico e o seguimento das vasculites pulmonares, especialmente da granulomatose de Wegener. Seu padrão pode ser citoplasmático e perinuclear. Sua titulação auxilia no diagnóstico e no seguimento das vasculites pulmonares.The discovery of the serological markers known as antineutrophil cytoplasmic antibodies revolutionized the diagnosis and follow-up treatment of the various forms of pulmonary vasculitis, especially that of Wegener's granulomatosis. The antineutrophil cytoplasmic antibodies pattern can be cytoplasmic or perinuclear. Determination of antineutrophil cytoplasmic antibodies titers aids the diagnosis and follow-up treatment of pulmonary vasculitis.

  2. Nonlinearity in cytoplasm viscosity can generate an essential symmetry breaking in cellular behaviors.

    Science.gov (United States)

    Tachikawa, Masashi; Mochizuki, Atsushi

    2015-01-07

    The cytoplasms of ameboid cells are nonlinearly viscous. The cell controls this viscosity by modulating the amount, localization and interactions of bio-polymers. Here we investigated how the nonlinearity infers the cellular behaviors and whether nonlinearity-specific behaviors exist. We modeled the developed plasmodium of the slime mold Physarum polycephalum as a network of branching tubes and examined the linear and nonlinear viscous cytoplasm flows in the tubes. We found that the nonlinearity in the cytoplasm׳s viscosity induces a novel type of symmetry breaking in the protoplasmic flow. We also show that symmetry breaking can play an important role in adaptive behaviors, namely, connection of behavioral modes implemented on different time scales and transportation of molecular signals from the front to the rear of the cell during cellular locomotion. Copyright © 2014 Elsevier Ltd. All rights reserved.

  3. Novel nuclear-cytoplasmic interaction in wheat (Triticum aestivum) induces vigorous plants

    Science.gov (United States)

    Interspecific hybridization can be considered an accelerator of evolution, otherwise a slow process, solely dependent on mutation and recombination. Upon interspecific hybridization, several novel interactions between nuclear and cytoplasmic genomes emerge which provide additional sources of diversi...

  4. Anti-neutrophil cytoplasm autoantibodies (ANCA). The need for specific and sensitive assays

    DEFF Research Database (Denmark)

    Baslund, B; Petersen, J

    1998-01-01

    Anti-neutrophil cytoplasm antibodies (ANCA) are a group of autoantibodies primarily associated with systemic vasculitis. Hitherto, the method of choice for ANCA detection has been indirect immunofluorescence (IIF). By this method two major patterns can be seen: a cytoplasmic pattern (cANCA) or a ......Anti-neutrophil cytoplasm antibodies (ANCA) are a group of autoantibodies primarily associated with systemic vasculitis. Hitherto, the method of choice for ANCA detection has been indirect immunofluorescence (IIF). By this method two major patterns can be seen: a cytoplasmic pattern (c......ANCA) or a perinuclear pattern (pANCA). The cANCA pattern is most often caused by antibodies directed against proteinase-3 (PR3) and in rare cases it is caused by anti-myeloperoixdase (MPO) antibodies. The pANCA pattern can de caused by antibodies directed against a large group of proteins i.e. MPO, lactofenin...

  5. Characterization of Novel Cytoplasmic PARP in the Brain of Octopus vulgaris

    Science.gov (United States)

    DE LISA, EMILIA; DE MAIO, ANNA; MOROZ, LEONID L.; MOCCIA, FRANCESCO; MENNELLA, MARIA ROSARIA FARAONE; DI COSMO, ANNA

    2014-01-01

    Recent investigation has focused on the participation of the poly (ADP-ribose) polymerase (PARP) reaction in the invertebrate central nervous system (CNS) during the process of long-term memory (LTM). In this paper, we characterize, localize, and assign a possible role to a cytoplasmic PARP in the brain of Octopus vulgaris. PARP activity was assayed in optic lobes, supraesophageal mass, and optic nerves. The highest levels of enzyme were found in the cytoplasmic fraction. Hyper-activation of the enzyme was detected in Octopus brain after visual discrimination training. Finally, cytoplasmic PARP was found to inhibit Octopus vulgaris actin polymerization. We propose that the cytoplasmic PARP plays a role in vivo to induce the cytoskeletonal reorganization that occurs during learning-induced neuronal plasticity. PMID:22815366

  6. Organization of the cytoplasmic reticulum in the central vacuole of parenchyma cells in Allium cepa L.

    Directory of Open Access Journals (Sweden)

    Tomasz J. Wodzicki

    2015-01-01

    Full Text Available An elaborate and complex cytoplasmic reticulum composed of fine filaments and lamellae ranging from 0.1 to 4 microns in size is revealed by viewing the central vacuole of onion bulb parenchyma cells with the scanning election microscope. The larger cytoplasmic strands, visible with the light microscope, are composed of numerous smaller filaments (some tubular which might explain the observed bidirectional movement of particles in these larger strands. The finely divided cytoplasmic network of filaments is continuous with the parietal cytoplasm inclosing the vacuolar sap. In these highly vacuolated cells the mass of the protoplast is in the form of an intravacuolar reticulum immersed in the cell sap. The probable significance of the vacuolar sap in relation to physiological processes of the cell is discussed.

  7. Content Marketing

    OpenAIRE

    Le, Duc

    2013-01-01

    The purpose of this thesis was to introduce the new trend in today’s marketing world: content marketing. It has been employed by many companies and organizations in the world and has been proven success even when it is still a fairly new topic. Five carefully selected theories of content marketing proposed by experts in the field has been collected, compared and displayed as originally and scientifically as possible in this thesis. The chosen theories provide a diversified perspectives of...

  8. Developing content for a mHealth intervention to promote postpartum retention in prevention of mother-to-child HIV transmission programs and early infant diagnosis of HIV: a qualitative study.

    Directory of Open Access Journals (Sweden)

    Thomas A Odeny

    Full Text Available Maternal attendance at postnatal clinic visits and timely diagnosis of infant HIV infection are important steps for prevention of mother-to-child transmission (PMTCT of HIV. We aimed to use theory-informed methods to develop text messages targeted at facilitating these steps.We conducted five focus group discussions with health workers and women attending antenatal, postnatal, and PMTCT clinics to explore aspects of women's engagement in postnatal HIV care and infant testing. Discussion topics were informed by constructs of the Health Belief Model (HBM and prior empirical research. Qualitative data were coded and analyzed according to the construct of the HBM to which they related. Themes were extracted and used to draft intervention messages. We carried out two stages of further messaging development: messages were presented in a follow-up focus group in order to develop optimal phrasing in local languages. We then further refined the messages, pretested them in individual cognitive interviews with selected health workers, and finalized the messages for the intervention.Findings indicated that brief, personalized, caring, polite, encouraging, and educational text messages would facilitate women bringing their children to clinic after delivery, suggesting that text messages may serve as an important "cue to action." Participants emphasized that messages should not mention HIV due to fear of HIV testing and disclosure. Participants also noted that text messages could capitalize on women's motivation to attend clinic for childhood immunizations.Applying a multi-stage content development approach to crafting text messages--informed by behavioral theory--resulted in message content that was consistent across different focus groups. This approach could help answer "why" and "how" text messaging may be a useful tool to support maternal and child health. We are evaluating the effect of these messages on improving postpartum PMTCT retention and infant

  9. Raman microspectroscopy of nucleus and cytoplasm for human colon cancer diagnosis.

    Science.gov (United States)

    Liu, Wenjing; Wang, Hongbo; Du, Jingjing; Jing, Chuanyong

    2017-11-15

    Subcellular Raman analysis is a promising clinic tool for cancer diagnosis, but constrained by the difficulty of deciphering subcellular spectra in actual human tissues. We report a label-free subcellular Raman analysis for use in cancer diagnosis that integrates subcellular signature spectra by subtracting cytoplasm from nucleus spectra (Nuc.-Cyt.) with a partial least squares-discriminant analysis (PLS-DA) model. Raman mapping with the classical least-squares (CLS) model allowed direct visualization of the distribution of the cytoplasm and nucleus. The PLS-DA model was employed to evaluate the diagnostic performance of five types of spectral datasets, including non-selective, nucleus, cytoplasm, ratio of nucleus to cytoplasm (Nuc./Cyt.), and nucleus minus cytoplasm (Nuc.-Cyt.), resulting in diagnostic sensitivity of 88.3%, 84.0%, 98.4%, 84.5%, and 98.9%, respectively. Discriminating between normal and cancerous cells of actual human tissues through subcellular Raman markers is feasible, especially when using the nucleus-cytoplasm difference spectra. The subcellular Raman approach had good stability, and had excellent diagnostic performance for rectal as well as colon tissues. The insights gained from this study shed new light on the general applicability of subcellular Raman analysis in clinical trials. Copyright © 2017 Elsevier B.V. All rights reserved.

  10. Microtubule–microtubule sliding by kinesin-1 is essential for normal cytoplasmic streaming in Drosophila oocytes

    Science.gov (United States)

    Lu, Wen; Winding, Michael; Lakonishok, Margot; Wildonger, Jill

    2016-01-01

    Cytoplasmic streaming in Drosophila oocytes is a microtubule-based bulk cytoplasmic movement. Streaming efficiently circulates and localizes mRNAs and proteins deposited by the nurse cells across the oocyte. This movement is driven by kinesin-1, a major microtubule motor. Recently, we have shown that kinesin-1 heavy chain (KHC) can transport one microtubule on another microtubule, thus driving microtubule–microtubule sliding in multiple cell types. To study the role of microtubule sliding in oocyte cytoplasmic streaming, we used a Khc mutant that is deficient in microtubule sliding but able to transport a majority of cargoes. We demonstrated that streaming is reduced by genomic replacement of wild-type Khc with this sliding-deficient mutant. Streaming can be fully rescued by wild-type KHC and partially rescued by a chimeric motor that cannot move organelles but is active in microtubule sliding. Consistent with these data, we identified two populations of microtubules in fast-streaming oocytes: a network of stable microtubules anchored to the actin cortex and free cytoplasmic microtubules that moved in the ooplasm. We further demonstrated that the reduced streaming in sliding-deficient oocytes resulted in posterior determination defects. Together, we propose that kinesin-1 slides free cytoplasmic microtubules against cortically immobilized microtubules, generating forces that contribute to cytoplasmic streaming and are essential for the refinement of posterior determinants. PMID:27512034

  11. Localization and function of KLF4 in cytoplasm of vascular smooth muscle cell

    International Nuclear Information System (INIS)

    Liu, Yan; Zheng, Bin; Zhang, Xin-hua; Nie, Chan-juan; Li, Yong-hui; Wen, Jin-kun

    2013-01-01

    Highlights: •PDGF-BB prompts the translocation of KLF4 to the cytoplasm. •PDGF-BB promotes interaction between KLF4 and actin in the cytoplasm. •Phosphorylation and SUMOylation of KLF4 participates in regulation of cytoskeletal organization. •KLF4 regulates cytoskeleton by promoting the expression of contraction-associated genes. -- Abstract: The Krüppel-like factor 4 is a DNA-binding transcriptional regulator that regulates a diverse array of cellular processes, including development, differentiation, proliferation, and apoptosis. The previous studies about KLF4 functions mainly focused on its role as a transcription factor, its functions in the cytoplasm are still unknown. In this study, we found that PDGF-BB could prompt the translocation of KLF4 to the cytoplasm through CRM1-mediated nuclear export pathway in vascular smooth muscle cells (VSMCs) and increased the interaction of KLF4 with actin in the cytoplasm. Further study showed that both KLF4 phosphorylation and SUMOylation induced by PDGF-BB participates in regulation of cytoskeletal organization by stabilizing the actin cytoskeleton in VSMCs. In conclusion, these results identify that KLF4 participates in the cytoskeletal organization by stabilizing cytoskeleton in the cytoplasm of VSMCs

  12. Anti-neutrophil cytoplasmic antibodies in rheumatoid arthritis: two case reports and review of literature

    Directory of Open Access Journals (Sweden)

    Spoerl David

    2012-12-01

    Full Text Available Abstract Background Anti-neutrophil cytoplasmic antibodies are typically detected in anti-neutrophil cytoplasmic antibody associated vasculitis, but are also present in a number of chronic inflammatory non-vasculitic conditions like rheumatoid arthritis. Rare cases of granulomatosis with polyangiitis (formerly known as Wegener’s granulomatosis, a vasculitic disorder frequently associated with the presence of anti-neutrophil cytoplasmic antibodies in patients with rheumatoid arthritis have been described in literature. Case presentation We report two middle-aged female patients with rheumatoid arthritis who developed anti-neutrophil cytoplasmic antibodies and symptoms reminiscent of granulomatosis with polyangiitis. Despite the lack of antibodies specific for proteinase 3 and the absence of a classical histology, we report a probable case of granulomatosis with polyangiitis in the first patient, and consider rheumatoid vasculitis in the second patient. Conclusion Taken together with previous reports, these cases highlight that anti-neutrophil cytoplasmic antibodies have to be evaluated very carefully in patients with rheumatoid arthritis. In this context, anti-neutrophil cytoplasmic antibodies detected by indirect immunofluorescence appear to have a low diagnostic value for granulomatosis with polyangiitis. Instead they may have prognostic value for assessing the course of rheumatoid arthritis.

  13. Localization and function of KLF4 in cytoplasm of vascular smooth muscle cell

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Yan [Department of Biochemistry and Molecular Biology, The Key Laboratory of Neurobiology and Vascular Biology (China); The Third Hospital of Hebei Medical University, Shijazhuang (China); Zheng, Bin; Zhang, Xin-hua; Nie, Chan-juan; Li, Yong-hui [Department of Biochemistry and Molecular Biology, The Key Laboratory of Neurobiology and Vascular Biology (China); Wen, Jin-kun, E-mail: wjk@hebmu.edu.cn [Department of Biochemistry and Molecular Biology, The Key Laboratory of Neurobiology and Vascular Biology (China)

    2013-06-28

    Highlights: •PDGF-BB prompts the translocation of KLF4 to the cytoplasm. •PDGF-BB promotes interaction between KLF4 and actin in the cytoplasm. •Phosphorylation and SUMOylation of KLF4 participates in regulation of cytoskeletal organization. •KLF4 regulates cytoskeleton by promoting the expression of contraction-associated genes. -- Abstract: The Krüppel-like factor 4 is a DNA-binding transcriptional regulator that regulates a diverse array of cellular processes, including development, differentiation, proliferation, and apoptosis. The previous studies about KLF4 functions mainly focused on its role as a transcription factor, its functions in the cytoplasm are still unknown. In this study, we found that PDGF-BB could prompt the translocation of KLF4 to the cytoplasm through CRM1-mediated nuclear export pathway in vascular smooth muscle cells (VSMCs) and increased the interaction of KLF4 with actin in the cytoplasm. Further study showed that both KLF4 phosphorylation and SUMOylation induced by PDGF-BB participates in regulation of cytoskeletal organization by stabilizing the actin cytoskeleton in VSMCs. In conclusion, these results identify that KLF4 participates in the cytoskeletal organization by stabilizing cytoskeleton in the cytoplasm of VSMCs.

  14. Microtubule-microtubule sliding by kinesin-1 is essential for normal cytoplasmic streaming in Drosophila oocytes.

    Science.gov (United States)

    Lu, Wen; Winding, Michael; Lakonishok, Margot; Wildonger, Jill; Gelfand, Vladimir I

    2016-08-23

    Cytoplasmic streaming in Drosophila oocytes is a microtubule-based bulk cytoplasmic movement. Streaming efficiently circulates and localizes mRNAs and proteins deposited by the nurse cells across the oocyte. This movement is driven by kinesin-1, a major microtubule motor. Recently, we have shown that kinesin-1 heavy chain (KHC) can transport one microtubule on another microtubule, thus driving microtubule-microtubule sliding in multiple cell types. To study the role of microtubule sliding in oocyte cytoplasmic streaming, we used a Khc mutant that is deficient in microtubule sliding but able to transport a majority of cargoes. We demonstrated that streaming is reduced by genomic replacement of wild-type Khc with this sliding-deficient mutant. Streaming can be fully rescued by wild-type KHC and partially rescued by a chimeric motor that cannot move organelles but is active in microtubule sliding. Consistent with these data, we identified two populations of microtubules in fast-streaming oocytes: a network of stable microtubules anchored to the actin cortex and free cytoplasmic microtubules that moved in the ooplasm. We further demonstrated that the reduced streaming in sliding-deficient oocytes resulted in posterior determination defects. Together, we propose that kinesin-1 slides free cytoplasmic microtubules against cortically immobilized microtubules, generating forces that contribute to cytoplasmic streaming and are essential for the refinement of posterior determinants.

  15. Lecithin Prevents Cortical Cytoskeleton Reorganization in Rat Soleus Muscle Fibers under Short-Term Gravitational Disuse.

    Directory of Open Access Journals (Sweden)

    Irina V Ogneva

    , but the levels of Tmod1 and Svil decreased by 2- and 5-fold, respectively. However, gravitational disuse did not result in changes in the mRNA content of Arpc3, Tmod1, Svil or Lcp1. Anti-orthostatic suspension for 6 hours resulted in a decrease in the mRNA content of alpha-actinin-4 (Actn4 and alpha-actinin-1 (Actn1 in group "HS" compared with that of group "C" by 25% and 30%, respectively, as well as a decrease and increase in the ACTN4 protein content in the membrane and cytoplasmic fractions, respectively. Lecithin injection resulted in an increase in the Actn1 and Actn4 mRNA content in group "C+L" by 1.5-fold and more than 2-fold, respectively, compared with the levels in group "C". Moreover, in group "HS+L", the mRNA content did not change in these genes compared with the levels in group "C+L", and the ACTN4 protein content in the membrane and cytoplasmic fractions also remained unchanged. Thus, lecithin prevented the reduction of Actn1 and Actn4 mRNA and the migration of ACTN4 from the cortical cytoskeleton to the cytoplasm.

  16. Prolonged endoplasmic reticulum stress promotes mislocalization of immunoglobulins to the cytoplasm.

    Science.gov (United States)

    Drori, Adi; Misaghi, Shahram; Haimovich, Joseph; Messerle, Martin; Tirosh, Boaz

    2010-05-01

    Signal peptide-dependent insertion of newly synthesized proteins into the endoplasmic reticulum (ER) is a multi-step process, whose fidelity varies with the identity of the protein and the cell type. ER translocation of prions is sensitive to conditions of acute ER stress in a manner that pre-emptively prevents their aggregation and proteo-toxicity. While this has been documented for extreme ER stress conditions and for a special type of proteins, the impact of chronic ER stress on protein translocation in general has not been well characterized. The unfolded protein response (UPR) is a cytoprotective signaling pathway activated by ER stress. The transcription factor X-box-binding protein 1 (XBP-1) is a key element of the mammalian UPR, which is activated in response to ER stress. Deletion of XBP-1 generates constitutive chronic ER stress conditions. Chronic ER stress can also be produced pharmacologically, for example by prolonged treatment with proteasome inhibitors, which abrogates XBP-1 activation. We tested the impact of chronic ER stress on protein insertion into the ER with special emphasis on antibody secreting cells (ASCs), as these cells cope physiologically with prolonged stress conditions. We show that XBP-1 in plasmablasts and fibroblasts controls the ER translocation of US2, a viral-encoded protein with a priori poor insertion efficiency. Using monoclonal antibodies that preferentially recognize ER-mis-inserted micro Ig chains we demonstrate that prolonged treatment of plasmablasts with proteasome inhibitors, as well as deletion of XBP-1, impaired the translocation of mu chains to the ER. Our data suggest that ASCs under prolonged ER stress conditions endure cytoplasmic mislocalization of Ig proteins. This mislocalization may further explain the exquisite sensitivity of multiple myeloma to proteasome inhibitors. (c) 2010 Elsevier Ltd. All rights reserved.

  17. Integration of irradiation with cytoplasmic incompatibility to facilitate a lymphatic filariasis vector elimination approach

    Directory of Open Access Journals (Sweden)

    Dobson Stephen L

    2009-08-01

    Full Text Available Abstract Background Mass drug administration (MDA is the emphasis of an ongoing global lymphatic filariasis (LF elimination program by the World Health Organization, in which the entire 'at risk' human population is treated annually with anti-filarial drugs. However, there is evidence that the MDA strategy may not be equally appropriate in all areas of LF transmission, leading to calls for the augmentation of MDA with anti-vector interventions. One potential augmentative intervention is the elimination of vectors via repeated inundative releases of male mosquitoes made cytoplasmically incompatible via an infection with Wolbachia bacteria. However, with a reduction in the vector population size, there is the risk that an accidental female release would permit the establishment of the incompatible Wolbachia infection type, resulting in population replacement instead of population elimination. To avoid the release of fertile females, we propose the exposure of release individuals to low doses of radiation to sterilize any accidentally released females, reducing the risk of population replacement. Results Aedes polynesiensis pupae of differing ages were irradiated to determine a radiation dose that results in sterility but that does not affect the survival and competitiveness of males. Laboratory assays demonstrate that males irradiated at a female sterilizing dosage of 40 Gy are equally competitive with un-irradiated males. No effect of irradiation on the ability of Wolbachia to affect egg hatch was observed. Conclusion An irradiation dose of 40 Gy is sufficient to cause female sterility, but has no observed negative effect on male fitness. The results support further development of this approach as a preventative measure against accidental population replacement.

  18. Cytoplasmic Kaiso is associated with poor prognosis in non-small cell lung cancer

    International Nuclear Information System (INIS)

    Dai, Shun-Dong; Wang, Yan; Miao, Yuan; Zhao, Yue; Zhang, Yong; Jiang, Gui-Yang; Zhang, Peng-Xin; Yang, Zhi-Qiang; Wang, En-Hua

    2009-01-01

    Kaiso has been identified as a new member of the POZ-zinc finger family of transcription factors that are implicated in development and cancer. Although controversy still exists, Kaiso is supposed to be involved in human cancer. However, there is limited information regarding the clinical significance of cytoplasmic/nuclear Kaiso in human lung cancer. In this study, immunohistochemical studies were performed on 20 cases of normal lung tissues and 294 cases of non-small cell lung cancer (NSCLC), including 50 cases of paired lymph node metastases and 88 cases with complete follow-up records. Three lung cancer cell lines showing primarily nuclear localization of Kaiso were selected to examine whether roles of Kaiso in cytoplasm and in nucleus are identical. Nuclear Kaiso was down-regulated by shRNA technology or addition a specific Kaiso antibody in these cell lines. The proliferative and invasive abilities were evaluated by MTT and Matrigel invasive assay, transcription of Kaiso's target gene matrilysin was detected by RT-PCR. Kaiso was primarily expressed in the cytoplasm of lung cancer tissues. Overall positive cytoplasmic expression rate was 63.61% (187/294). The positive cytoplasmic expression of Kaiso was higher in advanced TNM stages (III+IV) of NSCLC, compared to lower stages (I+II) (p = 0.019). A correlation between cytoplasmic Kaiso expression and lymph node metastasis was found (p = 0.003). In 50 paired cases, cytoplasmic expression of Kaiso was 78.0% (41/50) in primary sites and 90.0% (45/50) in lymph node metastases (p = 0.001). The lung cancer-related 5-year survival rate was significantly lower in patients who were cytoplasmic Kaiso-positive (22.22%), compared to those with cytoplasmic Kaiso-negative tumors (64.00%) (p = 0.005). Nuclear Kaiso staining was seen in occasional cases with only a 5.10% (15/294) positive rate and was not associated with any clinicopathological features of NSCLC. Furthermore, after the down-regulation of the nuclear

  19. Epidermal Growth Factor Cytoplasmic Domain Affects ErbB Protein Degradation by the Lysosomal and Ubiquitin-Proteasome Pathway in Human Cancer Cells

    Directory of Open Access Journals (Sweden)

    Aleksandra Glogowska

    2012-05-01

    Full Text Available The cytoplasmic domains of EGF-like ligands, including EGF cytoplasmic domain (EGFcyt, have important biological functions. Using specific constructs and peptides of human EGF cytoplasmic domain, we demonstrate that EGFcyt facilitates lysosomal and proteasomal protein degradation, and this coincided with growth inhibition of human thyroid and glioma carcinoma cells. EGFcyt and exon 22–23-encoded peptide (EGF22.23 enhanced procathepsin B (procathB expression and procathB-mediated lysosomal degradation of EGFR/ErbB1 as determined by inhibitors for procathB and the lysosomal ATPase inhibitor BafA1. Presence of mbEGFctF, EGFcyt, EGF22.23, and exon 23-encoded peptides suppressed the expression of the deubiqitinating enzyme ubiquitin C-terminal hydrolase-L1 (UCH-L1. This coincided with hyperubiquitination of total cellular proteins and ErbB1/2 and reduced proteasome activity. Upon small interfering RNA-mediated silencing of endogenously expressed UCH-L1, a similar hyperubiquitinylation phenotype, reduced ErbB1/2 content, and attenuated growth was observed. The exon 23-encoded peptide region of EGFcyt was important for these biologic actions. Structural homology modeling of human EGFcyt showed that this molecular region formed an exposed surface loop. Peptides derived from this EGFcyt loop structure may aid in the design of novel peptide therapeutics aimed at inhibiting growth of cancer cells.

  20. Infection of human and non-human cells by a highly fusogenic primary CD4-independent HIV-1 isolate with a truncated envelope cytoplasmic tail

    International Nuclear Information System (INIS)

    Saha, Kunal; Yan Hui; Nelson, Julie A.E.; Zerhouni-Layachi, Bouchra

    2005-01-01

    Truncation of the envelope cytoplasmic tail has enabled FIV, SIV, and some laboratory HIV-1 strains to acquire broader cellular tropism and enhanced fusogenicity. Here we have characterized a primary CD4-independent HIV-1 isolate (92UG046-T8) with a truncated cytoplasmic tail that was able to infect and induce syncytia in primary lymphocytes from human, chimpanzee, and monkey, as well as CD4-negative cell lines from human and monkey. Increased syncytia were also noticeable with 293 cells expressing the cloned envelope from the 92UG046-T8 isolate suggesting envelope-mediated cellular fusion. Except pooled serum from HIV-1-infected individuals, monoclonal anti-envelope antibodies or antibodies/antagonists against CD4, CXCR4, and CCR5 were not able to prevent infection by the 92UG046-T8 isolate. This is the first report showing a primary HIV-1 variant with truncated cytoplasmic tail which is highly fusogenic and can infect a broad range of cells from human and non-human origins. In vivo evolution of similar HIV-1 mutants may have important implications in AIDS pathogenesis

  1. HIV Risk and Prevention

    Science.gov (United States)

    ... Prevention VIH En Español Get Tested Find an HIV testing site near you. Enter ZIP code or city Follow HIV/AIDS CDC HIV CDC HIV/AIDS See RSS | ... Email Updates on HIV Syndicated Content Website Feedback HIV Risk and Prevention Recommend on Facebook Tweet Share ...

  2. Local Content

    CSIR Research Space (South Africa)

    Gibberd, Jeremy

    2016-10-01

    Full Text Available , 1997; Qiu & Tao, 2001; Corkin, 2012; Warner, 2011; Stephenson, 2013). Local content is also seen a way of improving national sustainability performance and developing greener buildings (Olivier et al, 2016; van Reneen, 2014; Gibberd, 2002). As a result...

  3. P granules phase transition induced by cytoplasmic streaming in Caenorhabditis elegans embryo

    Science.gov (United States)

    Wang, Hang; Hu, GuoHui

    2017-01-01

    P granules are germ granules contained in Caenorhabditis elegans germ cells. The first germ cell is specified by the one-cell embryo in which P granules localize to the posterior. Previous studies suggested that the mechanism of the localization phenomena is induced by liquid-liquid phase transition (LLPT), in which the polarity proteins control the saturation point of P granules. In the present study, we propose that the P granules phase transition can be triggered by the cytoplasmic streaming. A two-phase flow model is employed to simulate the localization of P granules, i.e., the cytoplasm is considered as a liquid phase, and the droplet-like P granules are another liquid phase. With the presence of the cytoplasmic streaming, P granules, initially distributing uniformly in the entire one-cell embryo, eventually condense/dissolve in the cytoplasm phase, regulated by difference between the saturation pressure and the hydrodynamic pressure. The numerical results reveal that the cytoplasmic streaming has significant effects on the localization of P granules, as well as the embryo division.

  4. Export of Precursor tRNAIle from the Nucleus to the Cytoplasm in Human Cells.

    Directory of Open Access Journals (Sweden)

    Min Wei

    Full Text Available In the current concept, tRNA maturation in vertebrate cells, including splicing of introns, trimming of 5' leader and 3' trailer, and adding of CCA, is thought to occur exclusively in the nucleus. Here we provide evidence to challenge this concept. Unspliced intron-containing precursor tRNAIle was identified in Human Immunodeficiency Virus type 1 (HIV-1 virions, which are synthesized in the cytoplasm. Northern blot, confocal microscopy and quantitative RT-PCR further verified enrichment of this unspliced tRNAIle within the cytoplasm in human cells. In addition to containing an intron, the cytoplasmic precursor tRNAIle also contains a short incompletely processed 5´ leader and a 3´ trailer, which abundance is around 1000 fold higher than the nuclear precursor tRNAIle with long 5' leader and long 3' trailer. In vitro data also suggest that the cytoplasmic unspliced end-immature precursor tRNAIle could be processed by short isoform of RNase Z, but not long isoform of RNase Z. These data suggest that precursor tRNAs could export from the nucleus to the cytoplasm in human cells, instead of be processed only in the nucleus.

  5. Bayesian Inference of Forces Causing Cytoplasmic Streaming in Caenorhabditis elegans Embryos and Mouse Oocytes.

    Science.gov (United States)

    Niwayama, Ritsuya; Nagao, Hiromichi; Kitajima, Tomoya S; Hufnagel, Lars; Shinohara, Kyosuke; Higuchi, Tomoyuki; Ishikawa, Takuji; Kimura, Akatsuki

    2016-01-01

    Cellular structures are hydrodynamically interconnected, such that force generation in one location can move distal structures. One example of this phenomenon is cytoplasmic streaming, whereby active forces at the cell cortex induce streaming of the entire cytoplasm. However, it is not known how the spatial distribution and magnitude of these forces move distant objects within the cell. To address this issue, we developed a computational method that used cytoplasm hydrodynamics to infer the spatial distribution of shear stress at the cell cortex induced by active force generators from experimentally obtained flow field of cytoplasmic streaming. By applying this method, we determined the shear-stress distribution that quantitatively reproduces in vivo flow fields in Caenorhabditis elegans embryos and mouse oocytes during meiosis II. Shear stress in mouse oocytes were predicted to localize to a narrower cortical region than that with a high cortical flow velocity and corresponded with the localization of the cortical actin cap. The predicted patterns of pressure gradient in both species were consistent with species-specific cytoplasmic streaming functions. The shear-stress distribution inferred by our method can contribute to the characterization of active force generation driving biological streaming.

  6. Immunomodulatory and antitumor effects in vivo by the cytoplasmic fraction of Lactobacillus casei and Bifidobacterium longum.

    Science.gov (United States)

    Lee, Jung-Woo; Shin, Jung-Gul; Kim, Eun Hee; Kang, Hae Eun; Yim, In Been; Kim, Ji Yeon; Joo, Hong-Gu; Woo, Hee Jong

    2004-03-01

    The immunomodulatory and antitumor effects of lactic acid bacteria (LABs) were investigated. Cytoplasmic fraction of Lactobacillus acidophilus, Lactobacillus casei and Bifidobacterium longum were tested for the antiproliferative activity in vitro to SNUC2A, SNU1, NIH/3T3 and Jurkat cell lines by crystal violet assay. All cytoplasmic fraction suppressed proliferation of tumor cells, though L. casei and B. longum were more effective. From these results, cytoplasmic fraction of L. casei and B. longum with Y400 as a control were administered as dietary supplements to Balb/c mice for 2, and 4 consecutive wks. Administration for 4 wks enhanced the number of total T cells, NK cells and MHC class II+ cells, and CD4-CD8+ T cells in flow cytometry analysis. To determine of antitumor activity of LABs preparation in vivo, F9 teratocarcinoma cells were inoculated on mice at 14th day. Body weight was decreased with increased survival rate in all groups with the cytoplasm of LABs. Our results showed that cytoplasmic fraction of LABs had direct antiproliferative effects on tumor cell lines in vitro, effects on immune cells in vivo, and antitumor effects on tumor-bearing mice with prolonged survival periods.

  7. Improved cytoplasmic pH measurements in SNARF-1 stained plant cells by image processing

    Science.gov (United States)

    Kutsuna, Natsumaro; Hasezawa, Seiichiro

    2010-01-01

    Cytoplasmic pH has long been considered to act as a secondary messenger of various cellular responses by affecting the ionization state of proteins.1 In plant biology, cytoplasmic pH has traditionally been measured, especially in guard cells and as a response to plant microorganism interactions, with pH-sensitive microelectrodes.2 More recently, the development of fluorescent pH markers, such as BCECF and SNARF-1, has allowed us to monitor cytoplasmic pH without the need for electrophysiological equipment. However, because of vacuolar structures that occupy a large volume of plant cells, simple measurements of fluorescent intensities are insufficient to provide precise cytoplasmic pH values. In this addendum, we describe our improved method to monitor cytoplasmic pH in plant cells stained by SNARF-1 by image processing using a noise-reducing filter after determination of an optimal ROI size. In addition, further developments for automated region extraction are proposed. PMID:20404517

  8. Opposing prognostic roles of nuclear and cytoplasmic RACGAP1 expression in colorectal cancer patients.

    Science.gov (United States)

    Yeh, Chung-Min; Sung, Wen-Wei; Lai, Hung-Wen; Hsieh, Ming-Ju; Yen, Hsu-Heng; Su, Tzu-Cheng; Chang, Wei-Hsiang; Chen, Chia-Yu; Ko, Jiunn-Liang; Chen, Chih-Jung

    2016-01-01

    Rac GTPase activating protein 1 (RACGAP1) plays a regulatory role in initiation of cytokinesis, control of cell growth and differentiation, and tumor malignancy, making it a potential prognostic biomarker. RACGAP1 is present in the nucleus, but a diffuse distribution in the cytoplasm also occurs. The aim of this study was to determine the impact of nuclear and cytoplasmic expression of RACGAP1 on clinical outcome to provide further evidence of a role in colorectal cancer. RACGAP1 expression was analyzed by immunohistochemistry in 166 cancer specimens from primary colorectal cancer patients. The mean follow-up time after surgery was 5.4 years (range, 0.01-13.10 years). The prognostic value of RACGAP1 on overall survival was validated by Kaplan-Meier analysis and Cox regression models. RACGAP1 is expressed in colorectal specimen and is present in both the nucleus and cytoplasm in different amounts. Colorectal cancer patients had opposite prognoses depending on the site of RACGAP1 expression. Patients with high nuclear RACGAP1 expression had poor outcomes, whereas those with high cytoplasmic RACGAP1 expression had favorable prognosis (P = .003 and P = .001, respectively). Patients with low nuclear but high cytoplasmic RACGAP1 expression had better survival compared with those with other combinations (P colorectal cancer. Copyright © 2015 Elsevier Inc. All rights reserved.

  9. Elucidating the role of select cytoplasmic proteins in altering diffusion of integrin receptors.

    Science.gov (United States)

    Sander, Suzanne; Arora, Neha; Smith, Emily A

    2012-06-01

    Cytoplasmic proteins that affect integrin diffusion in the cell membrane are identified using a combination of fluorescence recovery after photobleaching (FRAP) and RNA interference. Integrin receptors are essential for many cellular events, and alterations in lateral diffusion are one mechanism for modulating their function. In cells expressing native cytoplasmic protein concentrations and spread on a slide containing integrin extracellular ligand, 45 ± 2% of the integrin is mobile with a time-dependent 5.2 ± 0.9 × 10(-9) cm(2)/s diffusion coefficient at 1 s. The time exponent is 0.90 ± 0.07, indicating integrin diffusion moderately slows at longer times. The role of a specific cytoplasmic protein in altering integrin diffusion is revealed through changes in the FRAP curve after reducing the cytoplasmic protein's expression. Decreased expression of cytoplasmic proteins rhea, focal adhesion kinase (FAK), or steamer duck decreases the integrin mobile fraction. For rhea and FAK, there is a concomitant shift to Brownian (i.e., time-independent) diffusion at reduced concentrations of these proteins. In contrast, when the expression of actin 42A, dreadlocks, paxillin, integrin-linked kinase (ILK), or vinculin is reduced, integrin diffusion generally becomes more constrained with an increase in the integrin mobile fraction. This same change in integrin diffusion is measured in the absence of integrin extracellular ligand. The results indicate breaking the extracellular ligand-integrin-cytoskeletal linkage alters integrin diffusion properties, and, in most cases, there is no correlation between integrin and lipid diffusion properties.

  10. Expression of Anion Exchanger 1 Sequestrates p16 in the Cytoplasm in Gastric, Colonic Adenocarcinoma

    Directory of Open Access Journals (Sweden)

    Wei-Wei Shen

    2007-10-01

    Full Text Available p16INK4A (p16 binds to cyclin-dependent kinase 4/6, negatively regulates cell growth. Recent studies have led to an understanding of additional biologic functions for p16; however, the detailed mechanisms involved are still elusive. In this article, we show an unexpected expression of anion exchanger 1 (AEi in the cytoplasm in poorly, moderately differentiated gastric, colonic adenocarcinoma cells, in its interaction with p16, thereby sequestrating the protein in the cytoplasm. Genetic alterations of p16, AEi were not detectable. Forced expression of AEi in these cells sequestrated more p16 in the cytoplasm, whereas small interfering RNA-mediated silencing of AEi in the cells induced the release of p16 from the cytoplasm to the nucleus, leading to cell death, growth inhibition of tumor cells. By analyzing tissue samples obtained from patients with gastric, colonic cancers, we found that 83.33% of gastric cancers, 56.52% of colonic cancers coexpressed AEi, p16 in the cytoplasm. We conclude that AEi plays a crucial role in the pathogenesis of gastric, colonic adenocarcinoma, that p16 dysfunction is a novel pathway of carcinogenesis.

  11. Hydroxytyrosol prevents reduction in liver activity of Δ-5 and Δ-6 desaturases, oxidative stress, and depletion in long chain polyunsaturated fatty acid content in different tissues of high-fat diet fed mice.

    Science.gov (United States)

    Valenzuela, Rodrigo; Echeverria, Francisca; Ortiz, Macarena; Rincón-Cervera, Miguel Ángel; Espinosa, Alejandra; Hernandez-Rodas, María Catalina; Illesca, Paola; Valenzuela, Alfonso; Videla, Luis A

    2017-04-11

    , oxidative stress-related parameters, and tissue n-3 LCPUFA content was observed in HT-treated rats over control animals. HT supplementation prevents metabolic alterations in desaturase activities, oxidative stress status, and n-3 LCPUFA content in the liver and extrahepatic tissues of mice fed HFD.

  12. Cytoplasmic Skp2 expression is associated with p-Akt1 and predicts poor prognosis in human breast carcinomas.

    Directory of Open Access Journals (Sweden)

    Jing Liu

    Full Text Available BACKGROUND: S-phase kinase protein 2 (Skp2, an oncogenic protein, is a key regulator in different cellular and molecular processes, through ubiquitin-proteasome degradation pathway. Increased levels of Skp2 are observed in various types of cancer and associated with poor prognosis. However, in human breast carcinomas, the underlying mechanism and prognostic significance of cytoplasmic Skp2 is still undefined. METHODS: To investigate the role of cytoplasmic Skp2 expression in human breast carcinomas, we immnohistochemically assessed cytoplasmic Skp2, p-Akt1, and p27 expression in 251 patients with invasive ductal carcinomas of the breast. Association of cytoplasmic Skp2 expression with p-Akt1 and p27 was analyzed as well as correspondence with other clinicopathological parameters. Disease-free survival and overall survival were determined based on the Kaplan-Meier method and Cox regression models. RESULTS: Cytoplasmic of Skp2 was detected in 165 out of 251 (65.7% patients. Cytoplasmic Skp2 expression was associated with larger tumor size, more advanced histological grade, and positive HER2 expression. Increased cytoplasmic Skp2 expression correlated with p-Akt1 expression, with 54.2% (51/94 of low p-Akt1-expressing breast carcinomas, but 72.6% (114/157 of high p-Akt1-expressing breast carcinomas exhibiting cytoplasmic Skp2 expression. Elevated cytoplasmic Skp2 expression with low p-Akt1 expression was associated with poor disease-free and overall survival (DFS and OS, and Cox regression models demonstrated that cytoplasmic Skp2 expression was an independent prognostic marker for invasive breast carcinomas. CONCLUSION: Cytoplasmic Skp2 expression is associated with aggressive prognostic factors, such as larger tumor size, and advanced histological grade of the breast cancers. Results demonstrate that combined cytoplasmic Skp2 and p-Akt1 expression may be prognostic for patients with invasive breast carcinomas, and cytoplasmic Skp2 may serve as a

  13. Transcytosis of Aminopeptidase N in caco-2 cells is mediated by a Non-cytoplasmic Signal

    DEFF Research Database (Denmark)

    Vogel, L K; Norén, Ove; Sjöström, H

    1995-01-01

    of the transmembrane or cytoplasmic domain of aminopeptidase N for transport of aminopeptidase N by the indirect pathway by analysis of mutated forms of aminopeptidase N recombinantly expressed in Caco-2 cells. A tail-less and two secretory forms of aminopeptidase N, all deprived of the cytoplasmic tail, were...... transported to the basolateral plasma membrane in proportions equivalent to the wild type enzyme. This shows that no cytoplasmic basolateral sorting signal is involved in directing aminopeptidase N to the basolateral plasma membrane. Both the wild type and the tail-less aminopeptidase N were transcytosed from...... the basolateral to the apical plasma membrane, whereas no transcytosis of two secretory forms could be detected, showing that the transmembrane domain is important for efficient transcytosis to take place. A significant difference in transcytosis kinetics of the human and the porcine wild type aminopeptidase N...

  14. A Mechanism for Cytoplasmic Streaming: Kinesin-Driven Alignment of Microtubules and Fast Fluid Flows.

    Science.gov (United States)

    Monteith, Corey E; Brunner, Matthew E; Djagaeva, Inna; Bielecki, Anthony M; Deutsch, Joshua M; Saxton, William M

    2016-05-10

    The transport of cytoplasmic components can be profoundly affected by hydrodynamics. Cytoplasmic streaming in Drosophila oocytes offers a striking example. Forces on fluid from kinesin-1 are initially directed by a disordered meshwork of microtubules, generating minor slow cytoplasmic flows. Subsequently, to mix incoming nurse cell cytoplasm with ooplasm, a subcortical layer of microtubules forms parallel arrays that support long-range, fast flows. To analyze the streaming mechanism, we combined observations of microtubule and organelle motions with detailed mathematical modeling. In the fast state, microtubules tethered to the cortex form a thin subcortical layer and undergo correlated sinusoidal bending. Organelles moving in flows along the arrays show velocities that are slow near the cortex and fast on the inward side of the subcortical microtubule layer. Starting with fundamental physical principles suggested by qualitative hypotheses, and with published values for microtubule stiffness, kinesin velocity, and cytoplasmic viscosity, we developed a quantitative coupled hydrodynamic model for streaming. The fully detailed mathematical model and its simulations identify key variables that can shift the system between disordered (slow) and ordered (fast) states. Measurements of array curvature, wave period, and the effects of diminished kinesin velocity on flow rates, as well as prior observations on f-actin perturbation, support the model. This establishes a concrete mechanistic framework for the ooplasmic streaming process. The self-organizing fast phase is a result of viscous drag on kinesin-driven cargoes that mediates equal and opposite forces on cytoplasmic fluid and on microtubules whose minus ends are tethered to the cortex. Fluid moves toward plus ends and microtubules are forced backward toward their minus ends, resulting in buckling. Under certain conditions, the buckling microtubules self-organize into parallel bending arrays, guiding varying directions

  15. Dynamics of highly polydisperse colloidal suspensions as a model system for bacterial cytoplasm.

    Science.gov (United States)

    Hwang, Jiye; Kim, Jeongmin; Sung, Bong June

    2016-08-01

    There are various kinds of macromolecules in bacterial cell cytoplasm. The size polydispersity of the macromolecules is so significant that the crystallization and the phase separation could be suppressed, thus stabilizing the liquid state of bacterial cytoplasm. On the other hand, recent experiments suggested that the macromolecules in bacterial cytoplasm should exhibit glassy dynamics, which should be also affected significantly by the size polydispersity of the macromolecules. In this work, we investigate the anomalous and slow dynamics of highly polydisperse colloidal suspensions, of which size distribution is chosen to mimic Escherichia coli cytoplasm. We find from our Langevin dynamics simulations that the diffusion coefficient (D_{tot}) and the displacement distribution functions (P(r,t)) averaged over all colloids of different sizes do not show anomalous and glassy dynamic behaviors until the system volume fraction ϕ is increased up to 0.82. This indicates that the intrinsic polydispersity of bacterial cytoplasm should suppress the glass transition and help maintain the liquid state of the cytoplasm. On the other hand, colloids of each kind show totally different dynamic behaviors depending on their size. The dynamics of colloids of different size becomes non-Gaussian at a different range of ϕ, which suggests that a multistep glass transition should occur. The largest colloids undergo the glass transition at ϕ=0.65, while the glass transition does not occur for smaller colloids in our simulations even at the highest value of ϕ. We also investigate the distribution (P(θ,t)) of the relative angles of displacement for macromolecules and find that macromolecules undergo directionally correlated motions in a sufficiently dense system.

  16. Distinct cytoplasmic and nuclear functions of the stress induced protein DDIT3/CHOP/GADD153.

    Directory of Open Access Journals (Sweden)

    Alexandra Jauhiainen

    Full Text Available DDIT3, also known as GADD153 or CHOP, encodes a basic leucine zipper transcription factor of the dimer forming C/EBP family. DDIT3 is known as a key regulator of cellular stress response, but its target genes and functions are not well characterized. Here, we applied a genome wide microarray based expression analysis to identify DDIT3 target genes and functions. By analyzing cells carrying tamoxifen inducible DDIT3 expression constructs we show distinct gene expression profiles for cells with cytoplasmic and nuclear localized DDIT3. Of 175 target genes identified only 3 were regulated by DDIT3 in both cellular localizations. More than two thirds of the genes were downregulated, supporting a role for DDIT3 as a dominant negative factor that could act by either cytoplasmic or nuclear sequestration of dimer forming transcription factor partners. Functional annotation of target genes showed cell migration, proliferation and apoptosis/survival as the most affected categories. Cytoplasmic DDIT3 affected more migration associated genes, while nuclear DDIT3 regulated more cell cycle controlling genes. Cell culture experiments confirmed that cytoplasmic DDIT3 inhibited migration, while nuclear DDIT3 caused a G1 cell cycle arrest. Promoters of target genes showed no common sequence motifs, reflecting that DDIT3 forms heterodimers with several alternative transcription factors that bind to different motifs. We conclude that expression of cytoplasmic DDIT3 regulated 94 genes. Nuclear translocation of DDIT3 regulated 81 additional genes linked to functions already affected by cytoplasmic DDIT3. Characterization of DDIT3 regulated functions helps understanding its role in stress response and involvement in cancer and degenerative disorders.

  17. Day-to-night variations of cytoplasmic pH in a crassulacean acid metabolism plant.

    Science.gov (United States)

    Hafke, J B; Neff, R; Hütt, M T; Lüttge, U; Thiel, G

    2001-01-01

    In crassulacean acid metabolism (CAM) large amounts of malic acid are redistributed between vacuole and cytoplasm in the course of night-to-day transitions. The corresponding changes of the cytoplasmic pH (pHcyt) were monitored in mesophyll protoplasts from the CAM plant Kalanchoe daigremontiana Hamet et Perrier by ratiometric fluorimetry with the fluorescent dye 2',7'-bis-(2-carboxyethyl)-5-(and-6-)carboxyfluorescein as a pHcyt indicator. At the beginning of the light phase, pHcyt was slightly alkaline (about 7.5). It dropped during midday by about 0.3 pH units before recovering again in the late-day-to-early-dark phase. In the physiological context the variation in pHcyt may be a component of CAM regulation. Due to its pH sensitivity, phosphoenolpyruvate carboxylase appears as a likely target enzyme. From monitoring delta pHcyt in response to loading the cytoplasm with the weak acid salt K-acetate a cytoplasmic H(+)-buffer capacity in the order of 65 mM H+ per pH unit was estimated at a pHcyt of about 7.5. With this value, an acid load of the cytoplasm by about 10 mM malic acid can be estimated as the cause of the observed drop in pHcyt. A diurnal oscillation in pHcyt and a quantitatively similar cytoplasmic malic acid is predicted from an established mathematical model which allows simulation of the CAM dynamics. The similarity of model predictions and experimental data supports the view put forward in this model that a phase transition of the tonoplast is an essential functional element in CAM dynamics.

  18. Cytoplasmic retention of protein tyrosine kinase 6 promotes growth of prostate tumor cells.

    Science.gov (United States)

    Brauer, Patrick M; Zheng, Yu; Wang, Lin; Tyner, Angela L

    2010-10-15

    Protein tyrosine kinase 6 (PTK6) is an intracellular tyrosine kinase that is nuclear in epithelial cells of the normal prostate, but cytoplasmic in prostate tumors and in the PC3 prostate tumor cell line. The impact of altered PTK6 intracellular localization in prostate tumor cells has not been extensively explored. Knockdown of endogenous cytoplasmic PTK6 resulted in decreased PC3 cell proliferation and colony formation, suggesting that cytoplasmic PTK6 stimulates oncogenic pathways. In contrast, reintroduction of PTK6 into nuclei of PC3 cells had a negative effect on growth. Enhanced tyrosine phosphorylation of the PTK6 substrate Sam68 was detected in cells expressing nuclear-targeted PTK6. We found that mechanisms regulating nuclear localization of PTK6 are intact in PC3 cells. Transiently overexpressed PTK6 readily enters the nucleus. Ectopic expression of ALT-PTK6, a catalytically inactive splice variant of PTK6, did not affect localization of endogenous PTK6 in PC3 cells. Using leptomycin B, we confirmed that cytoplasmic localization of endogenous PTK6 is not due to Crm-1/exportin-1 mediated nuclear export. In addition, overexpression of the PTK6 nuclear substrate Sam68 is not sufficient to bring PTK6 into the nucleus. While exogenous PTK6 was readily detected in the nucleus when transiently expressed at high levels, low-level expression of inducible wild type PTK6 in stable cell lines resulted in its cytoplasmic retention. Our results suggest that retention of PTK6 in the cytoplasm of prostate cancer cells disrupts its ability to regulate nuclear substrates and leads to aberrant growth. In prostate cancer, restoring PTK6 nuclear localization may have therapeutic advantages.

  19. Two Outer Membrane Proteins Contribute to Caulobacter crescentus Cellular Fitness by Preventing Intracellular S-Layer Protein Accumulation

    Energy Technology Data Exchange (ETDEWEB)

    Overton, K. Wesley; Park, Dan M.; Yung, Mimi C.; Dohnalkova, Alice; Smit, John; Jiao, Yongqin

    2016-09-23

    ABSTRACT

    Surface layers, or S-layers, are two-dimensional protein arrays that form the outermost layer of many bacteria and archaea. They serve several functions, including physical protection of the cell from environmental threats. The high abundance of S-layer proteins necessitates a highly efficient export mechanism to transport the S-layer protein from the cytoplasm to the cell exterior.content content-type='genus-species'>Caulobacter crescentuscontent>is unique in that it has two homologous, seemingly redundant outer membrane proteins, RsaFaand RsaFb, which together with other components form a type I protein translocation pathway for S-layer export. These proteins have homology tocontent content-type='genus-species'>Escherichia colicontent>TolC, the outer membrane channel of multidrug efflux pumps. Here we provide evidence that, unlike TolC, RsaFaand RsaFbare not involved in either the maintenance of membrane stability or the active export of antimicrobial compounds. Rather, RsaFaand RsaFbare required to prevent intracellular accumulation and aggregation of the S-layer protein RsaA; deletion of RsaFaand RsaFbled to a general growth defect and lowered cellular fitness. Using Western blotting, transmission electron microscopy, and transcriptome sequencing (RNA-seq), we show that loss of both RsaFaand RsaFbled to accumulation of insoluble RsaA in the cytoplasm, which in turn caused upregulation of a number of genes involved in protein misfolding and degradation pathways. These findings provide new insight into the requirement for RsaFaand RsaFbin cellular fitness and tolerance to antimicrobial agents and further our understanding of the S-layer export mechanism on both the transcriptional and translational levels incontent content

  20. Conservation and divergence between cytoplasmic and muscle-specific actin capping proteins: insights from the crystal structure of cytoplasmic Cap32/34 from Dictyostelium discoideum

    Directory of Open Access Journals (Sweden)

    Eckert Christian

    2012-06-01

    Full Text Available Abstract Background Capping protein (CP, also known as CapZ in muscle cells and Cap32/34 in Dictyostelium discoideum, plays a major role in regulating actin filament dynamics. CP is a ubiquitously expressed heterodimer comprising an α- and β-subunit. It tightly binds to the fast growing end of actin filaments, thereby functioning as a “cap” by blocking the addition and loss of actin subunits. Vertebrates contain two somatic variants of CP, one being primarily found at the cell periphery of non-muscle tissues while the other is mainly localized at the Z-discs of skeletal muscles. Results To elucidate structural and functional differences between cytoplasmic and sarcomercic CP variants, we have solved the atomic structure of Cap32/34 (32 = β- and 34 = α-subunit from the cellular slime mold Dictyostelium at 2.2 Å resolution and compared it to that of chicken muscle CapZ. The two homologs display a similar overall arrangement including the attached α-subunit C-terminus (α-tentacle and the flexible β-tentacle. Nevertheless, the structures exhibit marked differences suggesting considerable structural flexibility within the α-subunit. In the α-subunit we observed a bending motion of the β-sheet region located opposite to the position of the C-terminal β-tentacle towards the antiparallel helices that interconnect the heterodimer. Recently, a two domain twisting attributed mainly to the β-subunit has been reported. At the hinge of these two domains Cap32/34 contains an elongated and highly flexible loop, which has been reported to be important for the interaction of cytoplasmic CP with actin and might contribute to the more dynamic actin-binding of cytoplasmic compared to sarcomeric CP (CapZ. Conclusions The structure of Cap32/34 from Dictyostelium discoideum allowed a detailed analysis and comparison between the cytoplasmic and sarcomeric variants of CP. Significant structural flexibility could particularly be found within the

  1. Choking Prevention

    Science.gov (United States)

    ... Healthy Living Healthy Living Healthy Living Nutrition Fitness Sports Oral Health Emotional Wellness Growing Healthy Sleep Safety & Prevention Safety & Prevention Safety and Prevention Immunizations At Home ...

  2. Comparative structural analysis of cytoplasmic and chloroplastic 5S rRNA from spinach.

    Science.gov (United States)

    Pieler, T; Digweed, M; Bartsch, M; Erdmann, V A

    1983-01-01

    5S rRNAs from Spinacea oleracea cytoplasmic and chloroplastic ribosomes have been subjected to digestion with the single strand specific nuclease S1 and to chemical modification of cytidines by sodium bisulphite in order to probe the RNA structure. According to these data, cytoplasmic 5S rRNA can be folded as proposed in the general eukaryotic 5S rRNA structure (1) and 5S rRNA from chloroplastides is shown to be more related to the general eubacterial structure (2). Images PMID:6340063

  3. Comparative structural analysis of cytoplasmic and chloroplastic 5S rRNA from spinach.

    OpenAIRE

    Pieler, T; Digweed, M; Bartsch, M; Erdmann, V A

    1983-01-01

    5S rRNAs from Spinacea oleracea cytoplasmic and chloroplastic ribosomes have been subjected to digestion with the single strand specific nuclease S1 and to chemical modification of cytidines by sodium bisulphite in order to probe the RNA structure. According to these data, cytoplasmic 5S rRNA can be folded as proposed in the general eukaryotic 5S rRNA structure (1) and 5S rRNA from chloroplastides is shown to be more related to the general eubacterial structure (2).

  4. Titration of a cytoplasmic polyhedrosis virus by a tissue microculture assay: some applications.

    Science.gov (United States)

    Belloncik, S; Chagnon, A

    1980-01-01

    A simple tissue microculture technique was developed for the titration of a cytoplasmic polyhedrosis virus (CPV) from Euxoa scandens. The procedure was similar to the 50% tissue culture infectious dose assay, but a single infected cell, detected by the presence of cytoplasmic polyhedra, was scored rather than the degeneration of cell monolayers. The filtration of CPV suspensions resulted in decreased virus titers under certain conditions. This microculture assay was used to determine the effect of cell disruption methods on virus yields. Sonication of infected cells was more efficient than freeze-thawing for the recovery of nonoccluded virus.

  5. Silicon scaffolds promoting three-dimensional neuronal web of cytoplasmic processes.

    Science.gov (United States)

    Papadopoulou, Evie L; Samara, Athina; Barberoglou, Marios; Manousaki, Aleka; Pagakis, Stamatis N; Anastasiadou, Ema; Fotakis, Costas; Stratakis, Emmanuel

    2010-06-01

    Primary neurons were grown on structured silicon (Si) substrates, in the absence of chemotropic factors or synthetic extracellular matrix. The Si substrates used for the study comprise hierarchical structures in the micro- and nanolength scales. The substrates were structured via femtosecond laser irradiation of the Si wafer, in a reactive SF(6) environment. Electron microscopy revealed that the neurons formed an elaborate web of cytoplasmic processes in the absence of glial elements. The neuronal cytoplasm autografted the depth of the spikes, and the neurite sprouting took place over the spikes surface. Here we demonstrate how microfabrication of a Si surface provides an excellent platform for multifaceted studies of neuronal specimens.

  6. Wild Nicotiana Species as a Source of Cytoplasmic Male Sterility in Nicotianatabacum

    Directory of Open Access Journals (Sweden)

    Nikova V

    2014-12-01

    Full Text Available The results of our experiments executed to obtain tobacco male sterile lines through interspecific hybridization are summarized. Ten wild species from the genus Nicotiana: N. excelsior (exc, N. amplexicaulis (amp, N. rustica (rus, Nicotianaglauca (gla, N. velutina (vel, N. benthamiana (ben, N. maritima (mar, N. paniculata (pan, N. longiflora (lon and N. africana (afr were used as cytoplasmic donors and N. tabacum, cv. HarmanliiskaBasma (HB as a donor of the nucleus. Genetic effects of cytoplasmic-nuclear interaction of the studied species are discussed. Our results suggested that cytoplasmic male sterility (CMS was expressed when the cytoplasms of the above mentioned wild Nicotiana species were combined with the nucleus of N. tabacum. The 10 sources of CMS obtained in tobacco were characterized by altered flower phenotypes. Flowers are classified into types according the stamen, pistil and corolla modification. All these CMS sources were backcrossed to Oriental tobaccos, cvs. Tekne, Nevrokop B-12, Kroumovgrad 90 and Djebel 576, to develop corresponding CMS lines. The investigated cytoplasms produced compete male sterility in all those cultivars. The CMS lines preserved flower types, specific for every “sterile” cytoplasm. The extent of male organ modifications varied from apparently normal (but pollenless stamens in CMS (pan, (afr, some plants of (vel (mar through different degrees of malformations (shriveled anther on shortened filaments (lon, pinnate-like anthers on filaments of normal length (amp, petal - (ben, pistil- or stigma-like structures (rus, (gla to lack of male reproductive organs in (exc and in some plants of (vel, (mar, (rus and (gla. Most of the above mentioned cytoplasms had normal female gametophyte and good seed productivity. Alterations of the pistils were observed in CMS (rus, (exc and (ben causing reduction of the seed set. Electrophoresis of seed proteins of the tobacco cultivars and their CMS lines also suggested that

  7. The early noncoding region of human papillomavirus type 16 is regulated by cytoplasmic polyadenylation factors

    DEFF Research Database (Denmark)

    Glahder, Jacob-Andreas Harald; Kristiansen, Karen; Durand, Marjorie

    2010-01-01

    (CPEs). We show here that a fragment of the early 3'end comprising four of the five CPE-like regions when inserted downstream of a reporter gene confers regulation of the gene expression. A key protein involved in cytoplasmic polyadenylation is CPEB. We show that the human CPEB1 can repress the activity...... of the reporter construct containing the HPV-16 early sequences. This repression can be counteracted by a human cytoplasmic poly(A) polymerase, hGLD-2 fused to CPEB1. The hGLD-2/CPEB1 fusion protein facilitates furthermore poly(A) elongation of early HPV transcripts....

  8. Content Analysis

    Directory of Open Access Journals (Sweden)

    George Bedinelli Rossi

    2014-09-01

    Full Text Available This study introduces the various definitions and types of content analysis. This type of analysis historically presents itself as a quantitative approach to data analysis and currently shows up as a qualitative approach. The most common types are the conceptual and relational analysis. The latter receives influences of linguistic, cognitive and mental models and it is subdivided in affective extraction, analysis of proximity and cognitive mapping. Regarding the importance of this type of analysis, we have quantitative and qualitative character and the latter approach can be used to identify hypotheses, theoretical constructs or even models that can be tested by multivariate statistical techniques or even by experiments. 

  9. Cytological study of radiation induced alterations in cytoplasmic factors controlling male sterility in corn. Progress report, February 28, 1975--December 1, 1975

    International Nuclear Information System (INIS)

    Edwardson, J.R.

    1975-01-01

    Progress is reported on the following research projects: cytoplasmic constituents of the embryo of various gymnosperms and angiosperms; cytoplasmic male sterility in corn; modification of cytoplasmic sterility factors using gamma radiation, EMS, and ethidium bromide; selection for sterile, blight-resistant corn plants; electron microscopy study of abnormal mitochondria in cytoplasm of corn; cytoplasmic male sterility in Petunia; non-Mendelian variegation in Petunia and Nicotiana; graft transmission of cytoplasmic male sterility; cytoplasmic male sterility in Vicia faba; and studies on Blakeslee's I virus in Datura

  10. CONTINUOUS MEASUREMENT OF THE CYTOPLASMIC PH IN LACTOCOCCUS-LACTIS WITH A FLUORESCENT PH INDICATOR

    NARCIS (Netherlands)

    MOLENAAR, D; ABEE, T; KONINGS, WN

    1991-01-01

    The cytoplasmic pH of Lactococcus lactis was studied with the fluorescent pH indicator 2',7'-bis-(2-carboxyethyl)-5 (and-6)-carboxyfluorescein (BCECF). A novel method was applied for loading bacterial cells with BCECF, which consists of briefly treating a dense cell suspension with acid in the

  11. Non-coding RNA may be associated with cytoplasmic male sterility in Silene vulgaris

    Czech Academy of Sciences Publication Activity Database

    Stone, James D.; Koloušková, Pavla; Sloan, D.B.; Štorchová, Helena

    2017-01-01

    Roč. 68, č. 7 (2017), s. 1599-1612 ISSN 0022-0957 R&D Projects: GA ČR(CZ) GA16-09220S Institutional support: RVO:61389030 Keywords : Cytoplasmic male sterility * Editing * Mitochondrion * Non-coding RNA * Silene vulgaris * Splicing * Transcriptome Subject RIV: EF - Botanics OBOR OECD: Plant sciences, botany Impact factor: 5.830, year: 2016

  12. Clinical significance of anti-neutrophil cytoplasmic antibodies (ANCA) in autoimmune liver diseases

    NARCIS (Netherlands)

    Roozendaal, C; de Jong, MA; van den Berg, AP; Limburg, PC; Kallenberg, CGM; van Wijk, R.

    Background/Aims: The clinical relevance of anti-neutrophil cytoplasmic antibodies (ANCA) in autoimmune liver disease is unclear. Defining the antigenic specificities of ANCA in these diseases may improve their clinical significance. Methods: We studied the target antigens of ANCA in 88 patients with

  13. A duplicated coxI gene is associated with cytoplasmic male sterility ...

    Indian Academy of Sciences (India)

    Home; Journals; Journal of Genetics; Volume 86; Issue 2. A duplicated coxI gene is associated with cytoplasmic male sterility in an alloplasmic Brassica juncea line derived from somatic hybridization with Diplotaxis catholica. Aruna Pathania Rajesh Kumar V. Dinesh Kumar Ashutosh K. K. Dwivedi P. B. Kirti P. Prakash V. L. ...

  14. Cytoplasmic male sterility in Petunia hybrida : a structural and histochemical analysis

    NARCIS (Netherlands)

    Bino, R.J.

    1986-01-01

    This thesis presents an analysis of the structural and histochemical aspects of cytoplasmic male sterility (cms) in Petuniahybrida . In petunia and in other crops, cms is the most commonly used tool for hybrid seed production. Application of the trait

  15. Molecular sieving properties of the cytoplasm of Escherichia coli and consequences of osmotic stress

    NARCIS (Netherlands)

    Mika, Jacek T.; van den Bogaart, Geert; Veenhoff, Liesbeth; Krasnikov, Victor; Poolman, Bert

    We determined the diffusion coefficients (D) of (macro)molecules of different sizes (from ~0.5 to 600 kDa) in the cytoplasm of live Escherichia coli cells under normal osmotic conditions and osmotic upshift. D values decreased with increasing molecular weight of the molecules. Upon osmotic upshift,

  16. Pathogenesis of antineutrophil cytoplasmic autoantibody-associated vasculitis and potential targets for biologic treatment

    NARCIS (Netherlands)

    Sanders, J.S.F.; Abdulahad, W H; Stegeman, C A; Kallenberg, C G M

    2014-01-01

    Antineutrophil cytoplasmic autoantibody (ANCA)-associated vasculitides (AAV) are autoimmune diseases in which the small vessels are inflamed. Clinical observations suggest a pathogenic role for ANCA. Such a role is supported by in vitro experimental data and animal models, particularly for

  17. Identifying cytoplasmic input to the cell wall of growing Chara corallina.

    Science.gov (United States)

    Proseus, Timothy E; Boyer, John S

    2006-01-01

    Plants enlarge mostly because the walls of certain cells enlarge, with accompanying input of wall constituents and other factors from the cytoplasm. However, the enlargement can occur without input, suggesting an uncertain relationship between cytoplasmic input and plant growth. Therefore, the role of the input was investigated by quantitatively comparing growth in isolated walls (no input) with that in living cells (input occurring). Cell walls were isolated from growing internodes of Chara corallina and filled with pressurized oil to control turgor pressure while elongation was monitored. Turgor pressure in living cells was similarly controlled and monitored by adding/removing cell solution. Temperature was varied in some experiments. At all pressures and temperatures, isolated walls displayed turgor-driven growth indistinguishable in every respect from that in living cells, except the rate decelerated in the isolated walls while the living cells grew rapidly. The growth in the isolated walls was highly responsive to temperature, in contrast to the elastic extension that has been shown to be insensitive to similar temperatures. Consequently, strong intermolecular bonds were responsible for growth and weak bonds for elastic extension. Boiling the walls gave the same results, indicating that enzyme activities were not controlling these bonds. However, pectin added to isolated walls reversed their growth deceleration and returned the rate to that in the living cells. The pectin was similar to that normally produced by the cytoplasm and deposited in the wall, suggesting that continued cytoplasmic input of pectin may play a role in sustaining turgor-driven growth in Chara.

  18. Ion permeability of the cytoplasmic membrane limits the maximum growth temperature of bacteria and archaea

    NARCIS (Netherlands)

    van de Vossenberg, J.L C M; Ubbink-Kok, T.; Elferink, M.G.L.; Driessen, A.J.M.; Konings, W.N

    1995-01-01

    Protons and sodium ions are the most commonly used coupling ions in energy transduction in bacteria and archaea. At their growth temperature, the permeability of the cytoplasmic membrane of thermophilic bacteria to protons is high compared with that of sodium ions. In some thermophiles, sodium is

  19. Cytoplasmic tethering of a RING protein RBCK1 by its splice variant lacking the RING domain

    International Nuclear Information System (INIS)

    Yoshimoto, Nobuo; Tatematsu, Kenji; Koyanagi, Tomoyoshi; Okajima, Toshihide; Tanizawa, Katsuyuki; Kuroda, Shun'ichi

    2005-01-01

    RBCC protein interacting with PKC 1 (RBCK1) is a transcription factor belonging to the RING-IBR protein family and has been shown to shuttle between the nucleus and cytoplasm, possessing both the nuclear export and localization signals within its amino acid sequence. RBCK2, lacking the C-terminal half of RBCK1 including the RING-IBR domain, has also been identified as an alternative splice variant of RBCK1. RBCK2 shows no transcriptional activity and instead it represses the transcriptional activity of RBCK1. Here, we show that RBCK2 is present usually in the cytoplasm containing two Leu-rich regions that presumably serve as a nuclear export signal (NES). Moreover, an NES-disrupted RBCK1 that is mostly localized within the nucleus is translocated to the cytoplasm when coexpressed with RBCK2, suggesting that RBCK2 serves as a cytoplasmic tethering protein for RBCK1. We propose a novel and general function of RING-lacking splice variants of RING proteins to control the intracellular localization and functions of the parental RING proteins by forming a hetero-oligomeric complex

  20. Plasma exchange in antineutrophil cytoplasmic antibody-associated vasculitis--a 25-year perspective

    DEFF Research Database (Denmark)

    Szpirt, Wladimir M

    2015-01-01

    Demonstration of a pathogenic role for antineutrophil cytoplasmic antibodies (ANCA) underlies the scientific rationale for plasma exchange (PLEX) in the treatment of ANCA-associated vasculitis (AAV). Most clinical evidence of efficacy concerns the use of PLEX for the recovery of renal function...

  1. Effects of ozone on apoplast/cytoplasm partitioning of ascorbic acid in snap bean

    Energy Technology Data Exchange (ETDEWEB)

    Burkey, K.O. [North Carolina State Univ., Dept. of Crop Science and Botany, Raleigh, NC (United States)

    1999-07-01

    Apoplast/cytoplasm partitioning of ascorbic acid (AA) was examined in four genotypes of snap bean (Phaseolus vulgaris L.) known to differ in ozone sensitivity. Plants were grown in pots under field conditions using open-top chambers to establish charcoal-filtered (CF) air (36 nmol mol{sup -1} ozone) or elevated ozone (77 nmol mol{sup -1} ozone) treatments, AA in fully expanded leaves of 36-day-old plants was separated into apoplast and cytoplasm fractions by vacuum infiltration methods using glucose 6-phosphate as a marker for cytoplasm contamination, Apoplast ascorbate levels ranged from 30 to 150 nmol g{sup -1} fresh weight. Ozone-sensitive genotypes partitioned 1-2% of total AA into the apoplast under CF conditions and up to 7% following a 7-day ozone exposure. In contrast, an ozone-tolerant genotype partitioned 3-4% of total leaf AA into the leaf apoplast in both CF and ozone-treated plants. The results suggest that genetic background and ozone stress are factors that affect AA levels in the extracellular space. For all genotypes, the fraction of AA in the oxidized form was higher in the apoplast compared to the cytoplasm, indicative of a more oxidizing environment within the cell wall. (au)

  2. Cytoplasmic Transport Machinery of the SPF27 Homologue Num1 in Ustilago maydis.

    Science.gov (United States)

    Zhou, Lu; Obhof, Theresa; Schneider, Karina; Feldbrügge, Michael; Nienhaus, G Ulrich; Kämper, Jörg

    2018-02-26

    In the phytopathogenic basidiomycete Ustilago maydis, the Num1 protein has a pivotal function in hyphal morphogenesis. Num1 functions as a core component of the spliceosome-associated Prp19/CDC5 complex (NTC). The interaction of Num1 with the kinesin motor Kin1 suggests a connection between a component of the splicing machinery and cytoplasmic trafficking processes. Previously it was shown that Num1 localizes predominantly in the nucleus; however, due to the diffraction-limited spatial resolution of conventional optical microscopy, it was not possible to attribute the localization to specific structures within the cytoplasm. We have now employed super-resolution localization microscopy to visualize Num1 in the cytoplasm by fusing it to a tandem dimeric Eos fluorescent protein (tdEosFP). The Num1 protein is localized within the cytoplasm with an enhanced density in the vicinity of microtubules. Num1 movement is found predominantly close to the nucleus. Movement is dependent on its interaction partner Kin1, but independent of Kin3. Our results provide strong evidence that, in addition to its involvement in splicing in the nucleus, Num1 has an additional functional role in the cytosol connected to the Kin1 motor protein.

  3. Molecular analysis of a new cytoplasmic male sterile genotype in sunflower

    NARCIS (Netherlands)

    Spassova, Mariana; Christov, Michail; Bohorova, Natasha; Petrov, Peter; Dudov, Kalin; Atanassov, Atanas; Nijkamp, H. John J.; Hille, Jaques

    1992-01-01

    Mitochondrial DNA from 1 fertile and 6 cytoplasmic male sterile (CMS) sunflower genotypes was studied. The CMS genotypes had been obtained either by specific crosses between different Helianthus species or by mutagenesis. CMS-associated restriction fragment length polymorphisms (RFLPs) were found in

  4. Dynamics of vegetative cytoplasm during generative cell formation and pollen maturation in Arabidopsis thaliana

    Science.gov (United States)

    Kuang, A.; Musgrave, M. E.

    1996-01-01

    Ultrastructural changes of pollen cytoplasm during generative cell formation and pollen maturation in Arabidopsis thaliana were studied. The pollen cytoplasm develops a complicated ultrastructure and changes dramatically during these stages. Lipid droplets increase after generative cell formation and their organization and distribution change with the developmental stage. Starch grains in amyloplasts increase in number and size during generative and sperm cell formation and decrease at pollen maturity. The shape and membrane system of mitochondria change only slightly. Dictyosomes become very prominent, and numerous associated vesicles are observed during and after sperm cell formation. Endoplasmic reticulum appears extensively as stacks during sperm cell formation. Free and polyribosomes are abundant in the cytoplasm at all developmental stages although they appear denser at certain stages and in some areas. In mature pollen, all organelles are randomly distributed throughout the vegetative cytoplasm and numerous small particles appear. Organization and distribution of storage substances and appearance of these small particles during generative and sperm cell formation and pollen maturation are discussed.

  5. Urinary matrix metalloproteinases reflect renal damage in anti-neutrophil cytoplasm autoantibody-associated vasculitis

    NARCIS (Netherlands)

    Sanders, J.S.F.; Huitema, M.G.; Hanemaaijer, R.; Goor, H. van; Kallenberg, C.G.M.; Stegeman, C.A.

    2007-01-01

    Renal expression of MMP-2, -9, and tissue inhibitor of MMP-1 (TIMP-1) correlates with histological disease activity in anti-neutrophil cytoplasm autoantibody (ANCA)-associated vasculitis (AAV). We studied whether urinary and plasma levels of MMP-2, -9, and TIMP-1 reflect renal expression of these

  6. Mitochondrial DNA variation within P-type cytoplasmic male sterility of Plantago lanceolata L

    NARCIS (Netherlands)

    Groenendijk, C.F.M.; Sandbrink, J.M.; Van Brederode, J.; Van Damme, J.M.M.

    1997-01-01

    MtDNA restriction fragment polymorphisms were found between cytoplasmic male-sterility types P and R of Plantago lanceolata with the homologous probe pPl311 and maize mtDNA fragments derived from the regions of atp1, cox1 and cox2. No mtDNA differences were observed between male-sterile and restored

  7. A duplicated coxI gene is associated with cytoplasmic male sterility ...

    Indian Academy of Sciences (India)

    which have been implicated in male sterility (reviewed in. Keywords. Brassica juncea; coxI gene; cytoplasmic male sterility; somatic hybrid; transcript variation. ... drial recombination has led to duplication of coxI gene which appears to be associated with the CMS. Materials and methods. Plant material. B. juncea cv.

  8. Probing the biology of cell boundary conditions through confinement of Xenopus cell-free cytoplasmic extracts.

    Science.gov (United States)

    Bermudez, Jessica G; Chen, Hui; Einstein, Lily C; Good, Matthew C

    2017-01-01

    Cell-free cytoplasmic extracts prepared from Xenopus eggs and embryos have for decades provided a biochemical system with which to interrogate complex cell biological processes in vitro. Recently, the application of microfabrication and microfluidic strategies in biology has narrowed the gap between in vitro and in vivo studies by enabling formation of cell-size compartments containing functional cytoplasm. These approaches provide numerous advantages over traditional biochemical experiments performed in a test tube. Most notably, the cell-free cytoplasm is confined using a two- or three-dimensional boundary, which mimics the natural configuration of a cell. This strategy enables characterization of the spatial organization of a cell, and the role that boundaries play in regulating intracellular assembly and function. In this review, we describe the marriage of Xenopus cell-free cytoplasm and confinement technologies to generate synthetic cell-like systems, the recent biological insights they have enabled, and the promise they hold for future scientific discovery. © 2017 Wiley Periodicals, Inc.

  9. Evaluating cytoplasmic and nuclear levels of inflammatory cytokines in cancer cells by western blotting.

    Science.gov (United States)

    Gatla, Himavanth R; Singha, Bipradeb; Persaud, Valerie; Vancurova, Ivana

    2014-01-01

    Increased expression and cellular release of inflammatory cytokines, interleukin-8 (IL-8; CXCL8), and high mobility group box-1 (HMGB1) are associated with increased cell proliferation, angiogenesis, and metastasis during cancer progression. In prostate and ovarian cancer cells, increased levels of IL-8 and HMGB1 correlate with poor prognosis. We have recently shown that proteasome inhibition by bortezomib (BZ) specifically increases IL-8 release from metastatic prostate and ovarian cancer cells. In this chapter, we describe a protocol to analyze the cytoplasmic and nuclear levels of IL-8 and HMGB1 in prostate and ovarian cancer cells by western blotting. IL-8 is localized in the cytoplasm in both cell types, and its protein levels are significantly increased by BZ. In contrast, HMGB1 is localized in the nucleus, and BZ increases its nuclear levels only in ovarian cancer cells. The protocol includes isolation of cytoplasmic and nuclear extracts, followed by SDS electrophoresis and western blotting, and can be easily modified to analyze the cytoplasmic and nuclear cytokine levels in other cell types.

  10. Secretion of Cpn0796 from Chlamydia pneumoniae into the host cell cytoplasm by an autotransporter mechanism

    DEFF Research Database (Denmark)

    Vandahl, Brian B S; Stensballe, Allan; Roepstorff, Peter

    2005-01-01

    in infected cells, whereas only the 65 kDa full-length Cpn0796 could be detected in purified Chlamydia. Confocal immunofluorescence microscopy demonstrated that Cpn0796 was localized in the Chlamydia membrane in young inclusions. However, at 36 h post infection and later Cpn0796 was detected in the cytoplasm...

  11. Probing cytoplasmic organization and the actin cytoskeleton of plant cells with optical tweezers

    NARCIS (Netherlands)

    Ketelaar, T.; Honing, van der H.S.; Emons, A.M.C.

    2010-01-01

    In interphase plant cells, the actin cytoskeleton is essential for intracellular transport and organization. To fully understand how the actin cytoskeleton functions as the structural basis for cytoplasmic organization, both molecular and physical aspects of the actin organization have to be

  12. Clinical Outcomes of Remission Induction Therapy for Severe Antineutrophil Cytoplasmic Antibody-Associated Vasculitis

    NARCIS (Netherlands)

    Miloslavsky, E. M.; Specks, U.; Merkel, P. A.; Seo, P.; Spiera, R.; Langford, C. A.; Hoffman, G. S.; Kallenberg, C. G. M.; St Clair, E. W.; Tchao, N. K.; Viviano, L.; Ding, L.; Sejismundo, L. P.; Mieras, K.; Ikle, D.; Jepson, B.; Mueller, M.; Brunetta, P.; Allen, N. B.; Fervenza, F. C.; Geetha, D.; Keogh, K.; Kissin, E. Y.; Monach, P. A.; Peikert, T.; Stegeman, C.; Ytterberg, S. R.; Stone, J. H.

    Objective. To evaluate the reasons that complete remission is not achieved or maintained with original treatment in some patients with antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) treated with rituximab (RTX) or with cyclophosphamide/azathioprine (CYC/AZA). Methods. The

  13. Rituximab for the Treatment of Relapses in Antineutrophil Cytoplasmic Antibody-Associated Vasculitis

    NARCIS (Netherlands)

    Miloslavsky, E. M.; Specks, U.; Merkel, P. A.; Seo, P.; Spiera, R.; Langford, C. A.; Hoffman, G. S.; Kallenberg, C. G. M.; Clair, E. W. St.; Tchao, N. K.; Viviano, L.; Ding, L.; Ikle, D.; Villarreal, M.; Jepson, B.; Brunetta, P.; Allen, N. B.; Fervenza, F. C.; Geetha, D.; Keogh, K.; Kissin, E. Y.; Monach, P. A.; Peikert, T.; Stegeman, C.; Ytterberg, S. R.; Stone, J. H.

    2014-01-01

    Objective. Disease relapses are frequent in anti-neutrophil cytoplasmic antibody-associated vasculitis (AAV). This study was undertaken to evaluate outcomes in patients with AAV who are re-treated with rituximab (RTX) and prednisone for severe disease relapses. Methods. The Rituximab in AAV trial

  14. Pleckstrin Homology Domain Diffusion in Dictyostelium Cytoplasm Studied Using Fluorescence Correlation Spectroscopy

    NARCIS (Netherlands)

    Engel, Ruchira; Hink, Mark A.; Bosgraaf, Leonard; Haastert, Peter J.M. van; Visser, Antonie J.W.G.

    2004-01-01

    The translocation of pleckstrin homology (PH) domain-containing proteins from the cytoplasm to the plasma membrane plays an important role in the chemotaxis mechanism of Dictyostelium cells. The diffusion of three PH domain-green fluorescent protein (GFP) fusions (PH2-GFP, PH10-GFP, and PH-CRAC

  15. Pleckstrin homology domain diffusion in Dictyostelium cytoplasm studied using fluorescence correlation spectroscopy

    NARCIS (Netherlands)

    Ruchira, A.; Hink, M.A.; Bosgraaf, L.; Haastert, van P.J.M.; Visser, A.J.W.G.

    2004-01-01

    The translocation of pleckstrin homology (PH) domain-containing proteins from the cytoplasm to the plasma membrane plays an important role in the chemotaxis mechanism of Dictyostelium cells. The diffusion of three PH domain-green fluorescent protein (GFP) fusions (PH2-GFP, PH10-GFP, and PH-CRAC

  16. Cytoplasmic CUG RNA foci are insufficient to elicit key DM1 features.

    Directory of Open Access Journals (Sweden)

    Warunee Dansithong

    Full Text Available The genetic basis of myotonic dystrophy type I (DM1 is the expansion of a CTG tract located in the 3' untranslated region of DMPK. Expression of mutant RNAs encoding expanded CUG repeats plays a central role in the development of cardiac disease in DM1. Expanded CUG tracts form both nuclear and cytoplasmic aggregates, yet the relative significance of such aggregates in eliciting DM1 pathology is unclear. To test the pathophysiology of CUG repeat encoding RNAs, we developed and analyzed mice with cardiac-specific expression of a beta-galactosidase cassette in which a (CTG(400 repeat tract was positioned 3' of the termination codon and 5' of the bovine growth hormone polyadenylation signal. In these animals CUG aggregates form exclusively in the cytoplasm of cardiac cells. A key pathological consequence of expanded CUG repeat RNA expression in DM1 is aberrant RNA splicing. Abnormal splicing results from the functional inactivation of MBNL1, which is hypothesized to occur due to MBNL1 sequestration in CUG foci or from elevated levels of CUG-BP1. We therefore tested the ability of cytoplasmic CUG foci to elicit these changes. Aggregation of CUG RNAs within the cytoplasm results both in Mbnl1 sequestration and in approximately a two fold increase in both nuclear and cytoplasmic Cug-bp1 levels. Significantly, despite these changes RNA splice defects were not observed and functional analysis revealed only subtle cardiac dysfunction, characterized by conduction defects that primarily manifest under anesthesia. Using a human myoblast culture system we show that this transgene, when expressed at similar levels to a second transgene, which encodes expanded CTG tracts and facilitates both nuclear focus formation and aberrant splicing, does not elicit aberrant splicing. Thus the lack of toxicity of cytoplasmic CUG foci does not appear to be a consequence of low expression levels. Our results therefore demonstrate that the cellular location of CUG RNA

  17. Cytoplasmic effects on DNA methylation between male sterile lines and the maintainer in wheat (Triticum aestivum L.).

    Science.gov (United States)

    Ba, Qingsong; Zhang, Gaisheng; Niu, Na; Ma, Shoucai; Wang, Junwei

    2014-10-01

    Male sterile cytoplasm plays an important role in hybrid wheat, and three-line system including male sterile (A line), its maintainer (B line) and restoring (R line) has played a major role in wheat hybrid production. It is well known that DNA methylation plays an important role in gene expression regulation during biological development in wheat. However, no reports are available on DNA methylation affected by different male sterile cytoplasms in hybrid wheat. We employed a methylation-sensitive amplified polymorphism technique to characterize nuclear DNA methylation in three male sterile cytoplasms. A and B lines share the same nucleus, but have different cytoplasms which is male sterile for the A and fertile for the B. The results revealed a relationship of DNA methylation at these sites specifically with male sterile cytoplasms, as well as male sterility, since the only difference between the A lines and B line was the cytoplasm. The DNA methylation was markedly affected by male sterile cytoplasms. K-type cytoplasm affected the methylation to a much greater degree than T-type and S-type cytoplasms, as indicated by the ratio of methylated sites, ratio of fully methylated sites, and polymorphism between A lines and B line for these cytoplasms. The genetic distance between the cytoplasm and nucleus for the K-type is much greater than for the T- and S-types because the former is between Aegilops genus and Triticum genus and the latter is within Triticum genus between Triticum spelta and Triticum timopheevii species. Thus, this difference in genetic distance may be responsible for the variation in methylation that we observed. Copyright © 2014. Published by Elsevier B.V.

  18. Nucleus and cytoplasm segmentation in microscopic images using K-means clustering and region growing.

    Science.gov (United States)

    Sarrafzadeh, Omid; Dehnavi, Alireza Mehri

    2015-01-01

    Segmentation of leukocytes acts as the foundation for all automated image-based hematological disease recognition systems. Most of the time, hematologists are interested in evaluation of white blood cells only. Digital image processing techniques can help them in their analysis and diagnosis. The main objective of this paper is to detect leukocytes from a blood smear microscopic image and segment them into their two dominant elements, nucleus and cytoplasm. The segmentation is conducted using two stages of applying K-means clustering. First, the nuclei are segmented using K-means clustering. Then, a proposed method based on region growing is applied to separate the connected nuclei. Next, the nuclei are subtracted from the original image. Finally, the cytoplasm is segmented using the second stage of K-means clustering. The results indicate that the proposed method is able to extract the nucleus and cytoplasm regions accurately and works well even though there is no significant contrast between the components in the image. In this paper, a method based on K-means clustering and region growing is proposed in order to detect leukocytes from a blood smear microscopic image and segment its components, the nucleus and the cytoplasm. As region growing step of the algorithm relies on the information of edges, it will not able to separate the connected nuclei more accurately in poor edges and it requires at least a weak edge to exist between the nuclei. The nucleus and cytoplasm segments of a leukocyte can be used for feature extraction and classification which leads to automated leukemia detection.

  19. Myosin-Powered Membrane Compartment Drives Cytoplasmic Streaming, Cell Expansion and Plant Development.

    Science.gov (United States)

    Peremyslov, Valera V; Cole, Rex A; Fowler, John E; Dolja, Valerian V

    2015-01-01

    Using genetic approaches, particle image velocimetry and an inert tracer of cytoplasmic streaming, we have made a mechanistic connection between the motor proteins (myosins XI), cargo transported by these motors (distinct endomembrane compartment defined by membrane-anchored MyoB receptors) and the process of cytoplasmic streaming in plant cells. It is shown that the MyoB compartment in Nicotiana benthamiana is highly dynamic moving with the mean velocity of ~3 μm/sec. In contrast, Golgi, mitochondria, peroxisomes, carrier vesicles and a cytosol flow tracer share distinct velocity profile with mean velocities of 0.6-1.5 μm/sec. Dominant negative inhibition of the myosins XI or MyoB receptors using overexpression of the N. benthamiana myosin cargo-binding domain or MyoB myosin-binding domain, respectively, resulted in velocity reduction for not only the MyoB compartment, but also each of the tested organelles, vesicles and cytoplasmic streaming. Furthermore, the extents of this reduction were similar for each of these compartments suggesting that MyoB compartment plays primary role in cytosol dynamics. Using gene knockout analysis in Arabidopsis thaliana, it is demonstrated that inactivation of MyoB1-4 results in reduced velocity of mitochondria implying slower cytoplasmic streaming. It is also shown that myosins XI and MyoB receptors genetically interact to contribute to cell expansion, plant growth, morphogenesis and proper onset of flowering. These results support a model according to which myosin-dependent, MyoB receptor-mediated transport of a specialized membrane compartment that is conserved in all land plants drives cytoplasmic streaming that carries organelles and vesicles and facilitates cell growth and plant development.

  20. Human B cells fail to secrete type I interferons upon cytoplasmic DNA exposure.

    Science.gov (United States)

    Gram, Anna M; Sun, Chenglong; Landman, Sanne L; Oosenbrug, Timo; Koppejan, Hester J; Kwakkenbos, Mark J; Hoeben, Rob C; Paludan, Søren R; Ressing, Maaike E

    2017-11-01

    Most cells are believed to be capable of producing type I interferons (IFN I) as part of an innate immune response against, for instance, viral infections. In macrophages, IFN I is potently induced upon cytoplasmic exposure to foreign nucleic acids. Infection of these cells with herpesviruses leads to triggering of the DNA sensors interferon-inducible protein 16 (IFI16) and cyclic GMP-AMP (cGAMP) synthase (cGAS). Thereby, the stimulator of interferon genes (STING) and the downstream molecules TANK-binding kinase 1 (TBK1) and interferon regulatory factor 3 (IRF3) are sequentially activated culminating in IFN I secretion. Human gamma-herpesviruses, such as Epstein-Barr virus (EBV), exploit B cells as a reservoir for persistent infection. In this study, we investigated whether human B cells, similar to macrophages, engage the cytoplasmic DNA sensing pathway to induce an innate immune response. We found that the B cells fail to secrete IFN I upon cytoplasmic DNA exposure, although they express the DNA sensors cGAS and IFI16 and the signaling components TBK1 and IRF3. In primary human B lymphocytes and EBV-negative B cell lines, this deficiency is explained by a lack of detectable levels of the central adaptor protein STING. In contrast, EBV-transformed B cell lines did express STING, yet both these lines as well as STING-reconstituted EBV-negative B cells did not produce IFN I upon dsDNA or cGAMP stimulation. Our combined data show that the cytoplasmic DNA sensing pathway is dysfunctional in human B cells. This exemplifies that certain cell types cannot induce IFN I in response to cytoplasmic DNA exposure providing a potential niche for viral persistence. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  1. Myosin-Powered Membrane Compartment Drives Cytoplasmic Streaming, Cell Expansion and Plant Development.

    Directory of Open Access Journals (Sweden)

    Valera V Peremyslov

    Full Text Available Using genetic approaches, particle image velocimetry and an inert tracer of cytoplasmic streaming, we have made a mechanistic connection between the motor proteins (myosins XI, cargo transported by these motors (distinct endomembrane compartment defined by membrane-anchored MyoB receptors and the process of cytoplasmic streaming in plant cells. It is shown that the MyoB compartment in Nicotiana benthamiana is highly dynamic moving with the mean velocity of ~3 μm/sec. In contrast, Golgi, mitochondria, peroxisomes, carrier vesicles and a cytosol flow tracer share distinct velocity profile with mean velocities of 0.6-1.5 μm/sec. Dominant negative inhibition of the myosins XI or MyoB receptors using overexpression of the N. benthamiana myosin cargo-binding domain or MyoB myosin-binding domain, respectively, resulted in velocity reduction for not only the MyoB compartment, but also each of the tested organelles, vesicles and cytoplasmic streaming. Furthermore, the extents of this reduction were similar for each of these compartments suggesting that MyoB compartment plays primary role in cytosol dynamics. Using gene knockout analysis in Arabidopsis thaliana, it is demonstrated that inactivation of MyoB1-4 results in reduced velocity of mitochondria implying slower cytoplasmic streaming. It is also shown that myosins XI and MyoB receptors genetically interact to contribute to cell expansion, plant growth, morphogenesis and proper onset of flowering. These results support a model according to which myosin-dependent, MyoB receptor-mediated transport of a specialized membrane compartment that is conserved in all land plants drives cytoplasmic streaming that carries organelles and vesicles and facilitates cell growth and plant development.

  2. Streptococcus faecalis mutants defective in regulation of cytoplasmic pH.

    Science.gov (United States)

    Kobayashi, H; Unemoto, T

    1980-09-01

    We have isolated two acid-sensitive mutants of Streptococcus faecalis (ATCC 9790), designated AS13 and AS25, which grew at pH 7.5 but not at pH below 6.0. The ionophore gramicidin D, which collapsed the pH gradient between the cytoplasm and the medium, had little effect on the growth of these mutants, indicating that growing cells maintain only a small pH gradient. In the presence of gramicidin D the growth rates of the parent and mutant strains were identical over a range of pH values. When glucose was added to a cell suspension at pH 6.4, the parent strain generated a pH gradient of 1.0 unit, interior alkaline; AS13 generated a pH gradient of only 0.5 units, and AS25 generated no measurable pH gradient. The proton permeability of the mutant strains was the same as that of the parent strain. These results suggest that a cytoplasmic pH of around 7.5 is required for the growth of the cells and that the mutant strains are unable to establish a neutral cytoplasmic pH in acidic medium because of damage to the regulatory system of the cytoplasmic pH. Mutant strains also have a reduced capacity to extrude protons and take up potassium. Therefore, it is likely that these cation transport systems are involved in the regulation of cytoplasmic pH.

  3. The dynamics of gynodioecy in Plantago lanceolata L. .1. Frequencies of male-steriles and their cytoplasmic male sterility types

    NARCIS (Netherlands)

    De Haan, A.A.; Luyten, R.M.J.M.; Bakx-Schotman, Tanja; Van Damme, J.M.M.

    1997-01-01

    The maintenance of a gynodioecious breeding system (hermaphrodites and male-steriles) was studied in Plantago lanceolata. Cytoplasmic-nuclear inheritance is important in the maintenance of male-steriles. The male-sterile trait is cytoplasmically based (CMS), and male fertility can be restored by

  4. Characterisation and expression of the mitochondrial genome of a new type of cytoplasmic male-sterile sunflower

    NARCIS (Netherlands)

    Spassova, Mariana; Moneger, Françoise; Leaver, Christopher J.; Petrov, Peter; Atanassov, Atanas; Nijkamp, H. John J.; Hille, Jacques

    1994-01-01

    A new cytoplasmic male sterile sunflower, CMS3, was characterised in relation to the Petiolaris (PET1) cytoplasmic male-sterile sunflower, CMS89. Southern blot analysis showed that the mitochondrial genome of CMS3 contains unique rearrangements in at least five loci (atp6, atp9, atpA, nad1 + 5 and

  5. Cytoplasmic continuity revisited: closure of septa of the filamentous fungus Schizophyllum commune in response to environmental conditions

    NARCIS (Netherlands)

    van Peer, A.F.; Muller, W.H.; Boekhout, T.; Lugones, L.; Wosten, H.A.

    2009-01-01

    BACKGROUND: Mycelia of higher fungi consist of interconnected hyphae that are compartmentalized by septa. These septa contain large pores that allow streaming of cytoplasm and even organelles. The cytoplasm of such mycelia is therefore considered to be continuous. METHODOLOGY/PRINCIPAL FINDINGS:

  6. Characterization of phosphorylation sites in the cytoplasmic domain of the 300 kDa mannose-6-phosphate receptor

    DEFF Research Database (Denmark)

    Rosorius, O; Mieskes, G; Issinger, O G

    1993-01-01

    kinases phosphorylate the cytoplasmic tail exclusively on serine residues. Inhibition studies using synthetic peptides, partial sequencing of isolated tryptic phosphopeptides and co-migration with tryptic phosphopeptides from MPR 300 labelled in vivo showed that (i) PKA phosphorylates the cytoplasmic MPR...

  7. Homologous Transcription Factors DUX4 and DUX4c Associate with Cytoplasmic Proteins during Muscle Differentiation.

    Directory of Open Access Journals (Sweden)

    Eugénie Ansseau

    Full Text Available Hundreds of double homeobox (DUX genes map within 3.3-kb repeated elements dispersed in the human genome and encode DNA-binding proteins. Among these, we identified DUX4, a potent transcription factor that causes facioscapulohumeral muscular dystrophy (FSHD. In the present study, we performed yeast two-hybrid screens and protein co-purifications with HaloTag-DUX fusions or GST-DUX4 pull-down to identify protein partners of DUX4, DUX4c (which is identical to DUX4 except for the end of the carboxyl terminal domain and DUX1 (which is limited to the double homeodomain. Unexpectedly, we identified and validated (by co-immunoprecipitation, GST pull-down, co-immunofluorescence and in situ Proximal Ligation Assay the interaction of DUX4, DUX4c and DUX1 with type III intermediate filament protein desmin in the cytoplasm and at the nuclear periphery. Desmin filaments link adjacent sarcomere at the Z-discs, connect them to sarcolemma proteins and interact with mitochondria. These intermediate filament also contact the nuclear lamina and contribute to positioning of the nuclei. Another Z-disc protein, LMCD1 that contains a LIM domain was also validated as a DUX4 partner. The functionality of DUX4 or DUX4c interactions with cytoplasmic proteins is underscored by the cytoplasmic detection of DUX4/DUX4c upon myoblast fusion. In addition, we identified and validated (by co-immunoprecipitation, co-immunofluorescence and in situ Proximal Ligation Assay as DUX4/4c partners several RNA-binding proteins such as C1QBP, SRSF9, RBM3, FUS/TLS and SFPQ that are involved in mRNA splicing and translation. FUS and SFPQ are nuclear proteins, however their cytoplasmic translocation was reported in neuronal cells where they associated with ribonucleoparticles (RNPs. Several other validated or identified DUX4/DUX4c partners are also contained in mRNP granules, and the co-localizations with cytoplasmic DAPI-positive spots is in keeping with such an association. Large muscle RNPs

  8. Nuclear and cytoplasmic accumulation of Ep-ICD is frequently detected in human epithelial cancers.

    Directory of Open Access Journals (Sweden)

    Ranju Ralhan

    Full Text Available BACKGROUND: We previously demonstrated that nuclear and cytoplasmic accumulation of the intracellular domain (Ep-ICD of epithelial cell adhesion molecule (EpCAM accompanied by a reciprocal reduction of its extracellular domain (EpEx, occurs in aggressive thyroid cancers. This study was designed to determine whether similar accumulation of Ep-ICD is a common event in other epithelial cancers. METHODOLOGY AND RESULTS: Ten epithelial cancers were immunohistochemically analyzed using Ep-ICD and EpEx domain-specific antibodies. The subcellular localization of EpEx and Ep-ICD in the human colon adenocarcinoma cell line CX-1 was observed using immunofluorescence. Nuclear and cytoplasmic Ep-ICD expression was increased in cancers of the breast (31 of 38 tissues, 82%, prostate (40 of 49 tissues, 82%, head and neck (37 of 57 tissues, 65% and esophagus (17 of 46 tissues, 37% compared to their corresponding normal tissues that showed membrane localization of the protein. Importantly, Ep-ICD was not detected in the nuclei of epithelial cells in most normal tissues. High nuclear and cytoplasmic Ep-ICD accumulation also occurred in the other six epithelial cancer types analyzed - lung, colon, liver, bladder, pancreatic, and ovarian. A concomitant reduction in membrane EpEx expression was observed in a subset of all cancer types. Receiver operating characteristic curve analysis revealed nuclear Ep-ICD distinguished breast cancers with 82% sensitivity and 100% specificity and prostate cancers with 82% sensitivity and 78% specificity. Similar findings were observed for cytoplasmic accumulation of Ep-ICD in these cancers. We provide clinical evidence of increased nuclear and cytoplasmic Ep-ICD accumulation and a reduction in membranous EpEx in these cancers. CONCLUSIONS: Increased nuclear and cytoplasmic Ep-ICD was observed in all epithelial cancers analyzed and distinguished them from normal tissues with high-sensitivity, specificity, and AUC. Development of a

  9. Cytoplasmic expression of C-MYC protein is associated with risk stratification of mantle cell lymphoma

    Directory of Open Access Journals (Sweden)

    Yi Gong

    2017-06-01

    Full Text Available Aim To investigate the association of C-MYC protein expression and risk stratification in mantle cell lymphoma (MCL, and to evaluate the utility of C-MYC protein as a prognostic biomarker in clinical practice. Methods We conducted immunohistochemical staining of C-MYC, Programmed cell death ligand 1 (PD-L1, CD8, Ki-67, p53 and SRY (sex determining region Y -11 (SOX11 to investigate their expression in 64 patients with MCL. The staining results and other clinical data were evaluated for their roles in risk stratification of MCL cases using ANOVA, Chi-square, and Spearman’s Rank correlation coefficient analysis. Results Immunohistochemical staining in our study indicated that SOX11, Ki-67 and p53 presented nuclear positivity of tumor cells, CD8 showed membrane positivity in infiltrating T lymphocytes while PD-L1 showed membrane and cytoplasmic positivity mainly in macrophage cells and little in tumor cells. We observed positive staining of C-MYC either in the nucleus or cytoplasm or in both subcellular locations. There were significant differences in cytoplasmic C-MYC expression, Ki-67 proliferative index of tumor cells, and CD8 positive tumor infiltrating lymphocytes (CD8+TIL among three risk groups (P = 0.000, P = 0.037 and P=0.020, respectively. However, no significant differences existed in the expression of nuclear C-MYC, SOX11, p53, and PD-L1 in MCL patients with low-, intermediate-, and high risks. In addition, patient age and serum LDH level were also significantly different among 3 groups of patients (P = 0.006 and P = 0.000, respectively. Spearman’s rank correlation coefficient analysis indicated that cytoplasmic C-MYC expression, Ki-67 index, age, WBC, as well as LDH level had significantly positive correlations with risk stratification (P = 0.000, 0.015, 0.000, 0.029 and 0.000, respectively, while CD8+TIL in tumor microenvironment negatively correlated with risk stratification of patients (P = 0.006. Patients with

  10. Cytoplasmic expression of C-MYC protein is associated with risk stratification of mantle cell lymphoma.

    Science.gov (United States)

    Gong, Yi; Zhang, Xi; Chen, Rui; Wei, Yan; Zou, Zhongmin; Chen, Xinghua

    2017-01-01

    To investigate the association of C-MYC protein expression and risk stratification in mantle cell lymphoma (MCL), and to evaluate the utility of C-MYC protein as a prognostic biomarker in clinical practice. We conducted immunohistochemical staining of C-MYC, Programmed cell death ligand 1 (PD-L1), CD8, Ki-67, p53 and SRY (sex determining region Y) -11 (SOX11) to investigate their expression in 64 patients with MCL. The staining results and other clinical data were evaluated for their roles in risk stratification of MCL cases using ANOVA, Chi-square, and Spearman's Rank correlation coefficient analysis. Immunohistochemical staining in our study indicated that SOX11, Ki-67 and p53 presented nuclear positivity of tumor cells, CD8 showed membrane positivity in infiltrating T lymphocytes while PD-L1 showed membrane and cytoplasmic positivity mainly in macrophage cells and little in tumor cells. We observed positive staining of C-MYC either in the nucleus or cytoplasm or in both subcellular locations. There were significant differences in cytoplasmic C-MYC expression, Ki-67 proliferative index of tumor cells, and CD8 positive tumor infiltrating lymphocytes (CD8+TIL) among three risk groups ( P  = 0.000, P  = 0.037 and P =0.020, respectively). However, no significant differences existed in the expression of nuclear C-MYC, SOX11, p53, and PD-L1 in MCL patients with low-, intermediate-, and high risks. In addition, patient age and serum LDH level were also significantly different among 3 groups of patients ( P  = 0.006 and P  = 0.000, respectively). Spearman's rank correlation coefficient analysis indicated that cytoplasmic C-MYC expression, Ki-67 index, age, WBC, as well as LDH level had significantly positive correlations with risk stratification ( P  = 0.000, 0.015, 0.000, 0.029 and 0.000, respectively), while CD8+TIL in tumor microenvironment negatively correlated with risk stratification of patients ( P  = 0.006). Patients with increased positive

  11. Enhancement of antiviral capacity of transgenic silkworms against cytoplasmic polyhedrosis virus via knockdown of multiple viral genes.

    Science.gov (United States)

    Jiang, Liang; Peng, Zhengwen; Guo, Huizhen; Sun, Jingchen; Sun, Qiang; Xia, Fei; Huang, Chunlin; Xu, Guowen; Xia, Qingyou

    2017-12-01

    Bombyx mori cytoplasmic polyhedrosis virus (BmCPV), a major pathogen of silkworms, causes serious economic losses in sericulture. The BmCPV genome contains 10 discrete dsRNA segments; among these, S1, S2, S3, S4, S6, and S7 encode virus structural proteins, whereas S5, S8, S9, and S10 encode nonstructural proteins. In an attempt to create an anti-BmCPV silkworm strain, we constructed transgenic RNAi vector pb-CNS for knockdown of S5, S8, S9, and S10, and pb-SNS targeting S1, S2, S4, S5, and S8. Transgenic silkworm line CNS and SNS were generated via microinjection of the practical diapause silkworm strain Furong. Following infection via the oral administration of a high dose of BmCPV, the mortality rates of the nontransgenic control, CNS, and SNS were 91%, 37%, and 41%, respectively. qPCR showed that the viral mRNA content in CNS and SNS was significantly lower than that in the nontransgenic line. The economic traits of CNS and SNS were not affected. These results suggest that the knockdown of multiple BmCPV genes significantly enhances the antiviral capacity of the silkworm. Copyright © 2017. Published by Elsevier Ltd.

  12. Citrus leprosis virus cytoplasmic type 2 (CiLV-C2 detection in Meta and Casanare States

    Directory of Open Access Journals (Sweden)

    Guillermo León

    2014-07-01

    Full Text Available This study discusses the diagnosis and detection of the new citrus leprosis virus cytoplasmic type 2 (CiLV-C2 in samples collected in Meta and Casanare States, using molecular techniques performed RT-PCR on plant tissue and mite vectors Brevipalpus phoenicis (Geijskes. RT-PCR tests were carry out using specific primers MP and CPG. The use of CPG primers allows the detection of CiLV-C2, in tissue plant samples with leprosis symptoms on fruits and leaves of Valencia orange (Citrus sinensis L. and Swinglea gluinosa Merr. leaves, which confirms the presence of this virus in Colombia. The MP primers did not detect the CiLV-C2 in any of performed tests. The RT-PCR technique allows detection of CiLV-C2 in samples of B. phoenicis mite vectors, with the specific CPG primers. Molecular detection of CiLV-C2 by RT-PCR technique is a fundamental tool for the virus diagnosis in Colombia, and should be used in diagnosis and prevention programs of citrus leprosis in the country.

  13. IP3 Receptor-Dependent Cytoplasmic Ca2+ Signals Are Tightly Controlled by Cavβ3

    Directory of Open Access Journals (Sweden)

    Anouar Belkacemi

    2018-01-01

    Full Text Available Voltage-gated calcium channels (Cavs are major Ca2+ entry pathways in excitable cells. Their β subunits facilitate membrane trafficking of the channel’s ion-conducting α1 pore and modulate its gating properties. We report that one β subunit, β3, reduces Ca2+ release following stimulation of phospholipase C-coupled receptors and inositol 1,4,5-trisphosphate (IP3 formation. This effect requires the SH3-HOOK domain of Cavβ3, includes physical β3/IP3 receptor interaction, and prevails when agonist-induced IP3 formation is bypassed by photolysis of caged IP3. In agreement with β3 acting as a brake on Ca2+ release, fibroblast migration is enhanced in vitro, and in vivo, closure of skin wounds is accelerated in the absence of β3. To mediate specific physiological responses and to prevent Ca2+ toxicity, cytoplasmic Ca2+ signals must be tightly controlled. The described function of β3, unrelated to its function as a Cav subunit, adds to this tight control.

  14. Monoacylated Cellular Prion Proteins Reduce Amyloid-β-Induced Activation of Cytoplasmic Phospholipase A2 and Synapse Damage

    Directory of Open Access Journals (Sweden)

    Ewan West

    2015-06-01

    Full Text Available Alzheimer’s disease (AD is a progressive neurodegenerative disease characterized by the accumulation of amyloid-β (Aβ and the loss of synapses. Aggregation of the cellular prion protein (PrPC by Aβ oligomers induced synapse damage in cultured neurons. PrPC is attached to membranes via a glycosylphosphatidylinositol (GPI anchor, the composition of which affects protein targeting and cell signaling. Monoacylated PrPC incorporated into neurons bound “natural Aβ”, sequestering Aβ outside lipid rafts and preventing its accumulation at synapses. The presence of monoacylated PrPC reduced the Aβ-induced activation of cytoplasmic phospholipase A2 (cPLA2 and Aβ-induced synapse damage. This protective effect was stimulus specific, as treated neurons remained sensitive to α-synuclein, a protein associated with synapse damage in Parkinson’s disease. In synaptosomes, the aggregation of PrPC by Aβ oligomers triggered the formation of a signaling complex containing the cPLA2.a process, disrupted by monoacylated PrPC. We propose that monoacylated PrPC acts as a molecular sponge, binding Aβ oligomers at the neuronal perikarya without activating cPLA2 or triggering synapse damage.

  15. Rape prevention

    Science.gov (United States)

    Date rape - prevention; Sexual assault - prevention ... Centers for Disease Control and Prevention website. Sexual assault and abuse and STDs. In: 2015 sexually transmitted diseases treatment guidelines 2015. www.cdc.gov/std/tg2015/sexual- ...

  16. Dengue Prevention

    Science.gov (United States)

    ... Address What's this? Submit What's this? Submit Button Prevention Recommend on Facebook Tweet Share Compartir This photograph ... medications to treat a dengue infection. This makes prevention the most important step, and prevention means avoiding ...

  17. The Use of Living Cancer Cells Expressing Green Fluorescent Protein in the Nucleus and Red Fluorescence Protein in the Cytoplasm for Real-time Confocal Imaging of Chromosome and Cytoplasmic Dynamics During Mitosis.

    Science.gov (United States)

    Suetsugu, Atsushi; Jiang, Ping; Yang, Meng; Yamamoto, Norio; Moriwaki, Hisataka; Saji, Shigetoyo; Hoffman, Robert M

    2015-05-01

    A library of dual-color fluorescent cancer cells with green fluorescent protein (GFP), linked to histone H2B, expressed in the nucleus and red fluorescent protein (RFP) expressed in the cytoplasm was previously genetically engineered. The aim of the current study was to use the dual-color cancer cells to visualize chromosome and cytoplasmic dynamics during mitosis. Using an Olympus FV1000 confocal microscope, a library of dual-color cells from the major cancer types was cultured on plastic. The cells were imaged by confocal microscopy to demonstrate chromosome and cytoplasmic dynamics during mitosis. Nuclear GFP expression enabled visualization of chromosomes behavior, whereas simultaneous cytoplasmic RFP expression enabled visualization of cytoplasmic behavior during mitosis. Thus, total cellular dynamics can be visualized at high resolution, including individual chromosomes in some cases, in living dual-color cells in real time. Dual-color cancer cells expressing H2B-GFP in the nucleus and RFP in the cytoplasm provide unique tools for visualizing subcellular nuclear and cytoplasm dynamics, including the behavior of individual chromosomes during mitosis. The dual-color cells can be used to evaluate chromosomal loss or gain in real time during treatment with a variety of agents or as the cells are selected for increased or decreased malignancy in culture or in vivo. The dual color cells will be a useful tool to discover and evaluate novel strategies for killing cancer cells. Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  18. Plague Prevention

    Science.gov (United States)

    ... Healthcare Professionals Clinicians Public Health Officials Veterinarians Prevention History of Plague Resources FAQ Prevention Recommend on Facebook Tweet Share Compartir Reduce rodent habitat around your ...

  19. Antineutrophil cytoplasmic antibody–negative pauci-immune glomerulonephritis with massive intestinal bleeding

    Directory of Open Access Journals (Sweden)

    Miyeon Kim

    2015-09-01

    Full Text Available A 61-year-old woman was admitted to hospital because of generalized edema and proteinuria. Her renal function deteriorated rapidly. Serum immunoglobulin and complement levels were within normal ranges. An autoantibody examination showed negative for antinuclear antibody and antineutrophil cytoplasmic antibody. Histologic examination of a renal biopsy specimen revealed that all of the glomeruli had severe crescent formations with no immune deposits. The patient was treated with steroid pulse therapy with cyclophosphamide followed by oral prednisolone. Fifteen days later, she experienced massive recurrent hematochezia. Angiography revealed an active contrast extravasation in a branch of the distal ileal artery. We selectively embolized with a permanent embolic agent. On the 45th hospital day, the patient suddenly lost consciousness. Brain computed tomography showed intracerebral hemorrhage. We report a case of antineutrophil cytoplasmic antibody–negative pauci-immune glomerulonephritis with massive intestinal bleeding and cerebral hemorrhage.

  20. Development of new semidwarf sources for rice with different cytoplasms (CV Basmati 370 and Gloria)

    International Nuclear Information System (INIS)

    Deus, J.E.; Suarez, E.; Fuentes, J.L.; Alvarez, A.; Padron, E.

    2001-01-01

    In Cuba semidwarf rice varieties grow on 98% of the area. Virtually all carry the same Dee-geo-won-gen dwarfing gene. Also, most if not all the cultivars have the same cytoplasm. The induced mutations approach was undertaken in order to generate alternative genetic sources of dwarfing with different cytoplasms and to improve the grain quality of Cuban rice. The seeds of two varieties, Basmati 370 and Gloria, were irradiated with 200 and 300 Gy of 60 Co gamma rays. In several generations of selection, progeny testing and preliminary yield tests 10 mutants (6 from Basmati 370 and 4 from Gloria), whose yields exceeded the source cultivars, have been advanced to replicated yield trials. Some of the mutant lines are also resistant to lodging and to blast (Pyricularis grisea). It is concluded the induced mutations can be used successfully to improve plant type and other agronomic traits in rice. The induced mutants will be used also in hybridization programs. (author)

  1. Molecular characterization of cytoplasmic male sterility (CMS) in perennial ryegrass (Lolium perenne L.)

    DEFF Research Database (Denmark)

    Islam, Md Shofiqul

    2011-01-01

    to increase biomass yield, improve nutritional value and tolerance towards abiotic and biotic stress. Cytoplasmic male sterility (CMS) is an efficient tool to control pollination for hybrid seed production. In order to identify the causative polymorphism of the CMS phenotype, a cytoplasmic male sterile plant......Perennial ryegrass (Lolium perenne L.) is a key grassland species in temperate climates. As an out breeding crop, ryegrass is currently being bred as population and synthetic families, thereby not fully exploiting the genetically available heterosis. Thus, hybrid breeding has the potential...... using 454 next-generation sequencing technology, resulting in approximately 800,000 high quality single reads. Here we report on the sequencing and the assembly of the mitochondrial genome from perennial ryegrass. Moreover, the assembly and annotation of the male-sterile and fertile mitochondrial...

  2. Parvovirus particles and movement in the cellular cytoplasm and effects of the cytoskeleton

    Energy Technology Data Exchange (ETDEWEB)

    Lyi, Sangbom Michael; Tan, Min Jie Alvin, E-mail: tanmja@gis.a-star.edu.sg; Parrish, Colin R., E-mail: crp3@cornell.edu

    2014-05-15

    Cell infection by parvoviruses requires that capsids be delivered from outside the cell to the cytoplasm, followed by genome trafficking to the nucleus. Here we microinject capsids into cells that lack receptors and followed their movements within the cell over time. In general the capsids remained close to the positions where they were injected, and most particles did not move to the vicinity of or enter the nucleus. When 70 kDa-dextran was injected along with the capsids that did not enter the nucleus in significant amounts. Capsids conjugated to peptides containing the SV40 large T-antigen nuclear localization signal remained in the cytoplasm, although bovine serum albumen conjugated to the same peptide entered the nucleus rapidly. No effects of disruption of microfilaments, intermediate filaments, or microtubules on the distribution of the capsids were observed. These results suggest that movement of intact capsids within cells is primarily associated with passive processes.

  3. The influence of Romanowsky-Giemsa type stains on nuclear and cytoplasmic features of cytological specimens.

    Science.gov (United States)

    Schulte, E; Wittekind, D

    1989-04-01

    The aim of the present study was to compare the staining pattern of the standard azure B-eosin Y stain with commercial May-Grünwald-Giemsa (MGG) stains on cytological specimens by means of high resolution image analysis. Several cytological specimens (blood smears, abdominal serous effusions, bronchial scrape material) were air dried, methanol fixed and stained with the standard azure B-eosin Y stain and with commercial May-Grünwald-Giemsa stains. Integrated optical density (IOD) and colour intensities of cell nuclei and cytoplasm were measured with the IBAS 2000 image analyser. Commercial MGG stains gave much higher coefficients of variation for all parameters than the standard stain. Reproducibility of cell nuclei segmentation versus cytoplasm was significantly better for the standard stain. Contamination of the standard stain with methylene blue partly copied the staining pattern of commercial stains. The standard azure B-eosin Y stain is recommended for high resolution image analysis (HRIA) of cytological samples.

  4. Attractant Signaling by an Aspartate Chemoreceptor Dimer with a Single Cytoplasmic Domain

    Science.gov (United States)

    Gardina, Paul J.; Manson, Michael D.

    1996-10-01

    Signal transduction across cell membranes often involves interactions among identical receptor subunits, but the contribution of individual subunits is not well understood. The chemoreceptors of enteric bacteria mediate attractant responses by interrupting a phosphotransfer circuit initiated at receptor complexes with the protein kinase CheA. The aspartate receptor (Tar) is a homodimer, and oligomerized cytoplasmic domains stimulate CheA activity much more than monomers do in vitro. Intragenic complementation was used to show in Escherichia coli that heterodimers containing one full-length and one truncated Tar subunit mediated responses to aspartate in the presence of full-length Tar homodimers that could not bind aspartate. Thus, a Tar dimer containing only one cytoplasmic domain can initiate an attractant (inhibitory) signal, although it may not be able to stimulate kinase activity of CheA.

  5. Dendritic Homeostasis Disruption in a Novel Frontotemporal Dementia Mouse Model Expressing Cytoplasmic Fused in Sarcoma

    Directory of Open Access Journals (Sweden)

    Gen Shiihashi

    2017-10-01

    Full Text Available Cytoplasmic aggregation of fused in sarcoma (FUS is detected in brain regions affected by amyotrophic lateral sclerosis (ALS and frontotemporal dementia (FTD, which compose the disease spectrum, FUS proteinopathy. To understand the pathomechanism of ALS-FTD-associated FUS, we examined the behavior and cellular properties of an ALS mouse model overexpressing FUS with nuclear localization signal deletion. Mutant FUS transgenic mice showed hyperactivity, social interactional deficits, and impaired fear memory retrieval, all of which are compatible with FTD phenotypes. Histological analyses showed decreased dendritic spine and synaptic density in the frontal cortex before neuronal loss. Examination of cultured cells confirmed that mutant but not wild-type FUS was associated with decreased dendritic growth, mRNA levels, and protein synthesis in dendrites. These data suggest that cytoplasmic FUS aggregates impair dendritic mRNA trafficking and translation, in turn leading to dendritic homeostasis disruption and the development of FTD phenotypes.

  6. Evidence for a cytoplasmic pathway of oxalate biosynthesis in Aspergillus niger

    Energy Technology Data Exchange (ETDEWEB)

    Kubicek, C.P.; Schreferl-Kunar, G.; Woehrer, W.; Roehr, M.

    1988-03-01

    Oxalate accumulation of up to 8 g/liter was induced in Aspergillus niger by shifting the pH from 6 to 8. This required the presence of P/sub i/ and a nitrogen source and was inhibited by the protein synthesis inhibitor cycloheximide. Exogenously added /sup 14/CO/sub 2/ was not incorporated into oxalate, but was incorporated into acetate and malate, thus indicating the biosynthesis of oxalate by hydrolytic cleavage of oxaloacetate. Inhibition of mitochondrial citrate metabolism by fluorocitrate did not significantly decrease the oxalate yield. The putative enzyme that was responsible for this oxaloacetate hydrolase (EC 3.7.1.1), which was induced de novo during the pH shift. Subcellular fractionation of oxalic acid-forming mycelia of A. niger showed that this enzyme is located in the cytoplasm of A. niger. The results are consistent with a cytoplasmic pathway of oxalate formation which does not involve the tricarboxylic acid cycle.

  7. Proteomic Analyses Reveal a Role of Cytoplasmic Droplets as an Energy Source during Epididymal Sperm Maturation

    Science.gov (United States)

    Yuan, Shuiqiao; Zheng, Huili; Zheng, Zhihong; Yan, Wei

    2013-01-01

    A small portion of cytoplasm is generally retained as the cytoplasmic droplet (CD) on the flagellum of spermatozoa after spermiation in mice. CDs are believed to play a role in osmoadaptation by allowing water entrance or exit. However, many lines of evidence suggest that CDs may have roles beyond osmoregulation. To gain more insights, we purified CDs from murine epididymal spermatozoa and conducted proteomic analyses on proteins highly enriched in CDs. Among 105 proteins identified, 71 (68%) were enzymes involved in energy metabolism. We also found that sperm mitochondria underwent a reactivation process and glycolytic enzymes were further distributed and incorporated into different regions of the flagellum during epididymal sperm maturation. Both processes appeared to require CDs. Our data suggest that the CD represents a transient organelle that serves as an energy source essential for epididymal sperm maturation. PMID:24155961

  8. Evolution of the cytoplasmic and mitochondrial phosphagen kinases unique to annelid groups.

    Science.gov (United States)

    Tanaka, Kumiko; Uda, Kouji; Shimada, Mayumi; Takahashi, Ken-Ichi; Gamou, Shinobu; Ellington, W Ross; Suzuki, Tomohiko

    2007-11-01

    Creatine kinase (CK) is a member of a group of phosphoryl transfer enzymes called phosphagen kinases that play a key role in cellular energy transactions in animals. Three CK isoform gene families are known-cytoplasmic CK (CK), flagellar CK (fCK), and mitochondrial CK (MiCK). Each of the isoforms has a unique gene structure (intron/exon organization). A broad array of other phosphagen kinases is present in animals. Some of these enzymes are found only in annelids and closely related groups including glyocyamine kinase (GK), lombricine kinase (LK), taurocyamine kinase (TK), and a unique arginine kinase (AK) restricted to annelids. Phylogenetic analyses of these annelid phosphagen kinases indicate that they appear to have evolved from a CK-like ancestor. To gain a greater understanding of the relationship of the CK isoforms to the annelid enzymes, we have determined the intron/exon organization of the genes for the following phosphagen kinases: Eisenia LK, Sabellastarte AK, and Arenicola mitochondrial TK (MiTK). Analysis of genomic database for the polychaete Capitella sp. yielded two putative LK genes [cytoplasmic LK and mitochondrial LK (MiLK)]. The intron/exon organization of these genes was compared with available data for cytoplasmic and mitochondrial CKs, and an annelid GK. Surprisingly, these annelid genes, irrespective of whether they are cytoplasmic (LK, AK, and GK) or mitochondrial (MiTK and MiLK), had the same 8-intron/9-exon organization and were strikingly similar to MiCK genes sharing seven of eight splice junctions. These results support the view that the MiCK gene is basal and ancestral to the phosphagen kinases unique to annelids.

  9. Interactions of Radiofrequency Radiation with NITELLA: Electrical Excitation and Perturbation of the Control of Cytoplasmic Streaming.

    Science.gov (United States)

    1982-03-01

    electrical state of the membrane in response to microwave exposure. Liu, et. al. (1982) exposed Chara corallina to 2.5-3.0 GHz microwaves, 36 w/kg for up to...of the effects of continuous wave, pulse- and amplitude modulated microwaves on single excitable cells of Chara corallina . Bioelectromagnetics (In...cells. J. of Ultrastructure Res. 71:321-330. Williamson, R.E. 1974. Actin in the alga Chara corallina . Nature. 241:801-802. -. 1975. Cytoplasmic

  10. Plant cytoplasmic GAPDH: redox post-translational modifications and moonlighting properties

    Directory of Open Access Journals (Sweden)

    Mirko eZaffagnini

    2013-11-01

    Full Text Available Glyceraldehyde-3-phosphate dehydrogenase (GAPDH is a ubiquitous enzyme involved in glycolysis and shown, particularly in animal cells, to play additional roles in several unrelated non-metabolic processes such as control of gene expression and apoptosis. This functional versatility is regulated, in part at least, by redox post-translational modifications that alter GAPDH catalytic activity and influence the subcellular localization of the enzyme. In spite of the well established moonlighting (multifunctional properties of animal GAPDH, little is known about non-metabolic roles of GAPDH in plants. Plant cells contain several GAPDH isoforms with different catalytic and regulatory properties, located both in the cytoplasm and in plastids, and participating in glycolysis and the Calvin-Benson cycle. A general feature of all GAPDH proteins is the presence of an acidic catalytic cysteine in the active site that is overly sensitive to oxidative modifications, including glutathionylation and S-nitrosylation. In Arabidopsis, oxidatively-modified cytoplasmic GAPDH has been successfully used as a tool to investigate the role of reduced glutathione, thioredoxins and glutaredoxins in the control of different types of redox post-translational modifications. Oxidative modifications inhibit GAPDH activity, but might enable additional functions in plant cells. Mounting evidence support the concept that plant cytoplasmic GAPDH may fulfill alternative, non-metabolic functions that are triggered by redox post-translational modifications of the protein under stress conditions. The aim of this review is to detail the molecular mechanisms underlying the redox regulation of plant cytoplasmic GAPDH in the light of its crystal structure, and to provide a brief inventory of the well known redox-dependent multi-facetted properties of animal GAPDH, together with the emerging roles of oxidatively-modified GAPDH in stress signaling pathways in plants.

  11. Reprogramming of round spermatids by the germinal vesicle cytoplasm in mice.

    Directory of Open Access Journals (Sweden)

    Peng-Cheng Kong

    Full Text Available The birthrate following round spermatid injection (ROSI remains low in current and evidence suggests that factors in the germinal vesicle (GV cytoplasm and certain substances in the GV such as the nucleolus might be responsible for genomic reprogramming and embryonic development. However, little is known whether the reprogramming factors in GV oocyte cytoplasm and/or nucleolus in GV are beneficial to the reprogramming of round spermatids and development of ROSI embryos. Here, round spermatids were treated with GV cytolysates and injected this round spermatid alone or co-injected with GV oocyte nucleolus into mature metaphase II oocytes. Subsequent embryonic development was assessed morphologically and by Oct4 expression in blastocysts. There was no significant difference between experimental groups at the zygote to four-cell development stages. Blastocysts derived from oocytes which were injected with cytolysate treated-round spermatid alone or co-injected with nucleoli injection yielded 63.6% and 70.3% high quality embryos, respectively; comparable to blastocysts derived by intracytoplasmic sperm injection (ICSI, but higher than these oocytes which were co-injected with lysis buffer-treated round spermatids and nucleoli or injected with the lysis buffer-treated round spermatids alone. Furthermore, the proportion of live offspring resulting from oocytes which were co-injected with cytolysate treated-round spermatids and nucleoli or injected with cytolysate treated-round spermatids alone was higher than those were injected with lysis buffer treated-round spermaids, but comparable with the ICSI group. Our results demonstrate that factors from the GV cytoplasm improve round spermatid reprogramming, and while injection of the extra nucleolus does not obviously improve reprogramming its potential contribution, although which cannot be definitively excluded. Thus, some reprogramming factors are evidently present in GV oocyte cytoplasm and could

  12. High ASMA+ Fibroblasts and Low Cytoplasmic HMGB1+ Breast Cancer Cells Predict Poor Prognosis.

    Science.gov (United States)

    Amornsupak, Kamolporn; Jamjuntra, Pranisa; Warnnissorn, Malee; O-Charoenrat, Pornchai; Sa-Nguanraksa, Doonyapat; Thuwajit, Peti; Eccles, Suzanne A; Thuwajit, Chanitra

    2017-10-01

    The influence of cancer-associated fibroblasts (CAFs) and high mobility group box 1 (HMGB1) has been recognized in several cancers, although their roles in breast cancer are unclear. The present study aimed to determine the levels and prognostic significance of α-smooth muscle actin-positive (ASMA + ) CAFs, plus HMGB1 and receptor for advanced glycation end products (RAGE) in cancer cells. A total of 127 breast samples, including 96 malignant and 31 benign, were examined for ASMA, HMGB1, and RAGE by immunohistochemistry. The χ 2 test and Fisher's exact test were used to test the association of each protein with clinicopathologic parameters. The Kaplan-Meier method or log-rank test and Cox regression were used for survival analysis. ASMA + fibroblast infiltration was significantly increased in the tumor stroma compared with that in benign breast tissue. The levels of cytoplasmic HMGB1 and RAGE were significantly greater in the breast cancer tissue than in the benign breast tissues. High ASMA expression correlated significantly with large tumor size, clinical stage III-IV, and angiolymphatic and perinodal invasion. In contrast, increased cytoplasmic HMGB1 correlated significantly with small tumor size, pT stage, early clinical stage, luminal subtype (but not triple-negative subtype), and estrogen receptor and progesterone receptor expression. The levels of ASMA (hazard ratio, 14.162; P = .010) and tumor cytoplasmic HMGB1 (hazard ratio, 0.221; P = .005) could serve as independent prognostic markers for metastatic relapse in breast cancer patients. The ASMA-high/HMGB1-low profile provided the most reliable prediction of metastatic relapse. We present for the first time, to the best of our knowledge, the potential clinical implications of the combined assessment of ASMA + fibroblasts and cytoplasmic HMGB1 in breast cancer. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  13. Cytoplasmic BRMS1 expression in malignant melanoma is associated with increased disease-free survival

    Directory of Open Access Journals (Sweden)

    Slipicevic Ana

    2012-02-01

    Full Text Available Abstract Background/aims Breast cancer metastasis suppressor 1 (BRMS1 blocks metastasis in melanoma xenografts; however, its usefulness as a biomarker in human melanomas has not been widely studied. The goal was to measure BRMS1 expression in benign nevi, primary and metastatic melanomas and evaluate its impact on disease progression and prognosis. Methods Paraffin-embedded tissue from 155 primary melanomas, 69 metastases and 15 nevi was examined for BRMS1 expression using immunohistochemistry. siRNA mediated BRMS1 down-regulation was used to study impact on invasion and migration in melanoma cell lines. Results A significantly higher percentage of nevi (87%, compared to primary melanomas (20% and metastases (48%, expressed BRMS1 in the nucelus (p Waf1/Cip1 (p = 0.009. Cytoplasmic score index was inversely associated with nuclear p-Akt (p = 0.013 and positively associated with cytoplasmic p-ERK1/2 expression (p = 0.033. Nuclear BRMS1 expression in ≥ 10% of primary melanoma cells was associated with thicker tumors (p = 0.016 and decreased relapse-free period (p = 0.043. Nuclear BRMS1 was associated with expression of fatty acid binding protein 7 (FABP7; p = 0.011, a marker of invasion in melanomas. In line with this, repression of BRMS1 expression reduced the ability of melanoma cells to migrate and invade in vitro. Conclusion Our data suggest that BRMS1 is localized in cytoplasm and nucleus of melanocytic cells and that cellular localization determines its in vivo effect. We hypothesize that cytoplasmic BRMS1 restricts melanoma progression while nuclear BRMS1 possibly promotes melanoma cell invasion. Please see related article: http://www.biomedcentral.com/1741-7015/10/19

  14. Plant cytoplasmic GAPDH: redox post-translational modifications and moonlighting properties

    Science.gov (United States)

    Zaffagnini, Mirko; Fermani, Simona; Costa, Alex; Lemaire, Stéphane D.; Trost, Paolo

    2013-01-01

    Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a ubiquitous enzyme involved in glycolysis and shown, particularly in animal cells, to play additional roles in several unrelated non-metabolic processes such as control of gene expression and apoptosis. This functional versatility is regulated, in part at least, by redox post-translational modifications that alter GAPDH catalytic activity and influence the subcellular localization of the enzyme. In spite of the well established moonlighting (multifunctional) properties of animal GAPDH, little is known about non-metabolic roles of GAPDH in plants. Plant cells contain several GAPDH isoforms with different catalytic and regulatory properties, located both in the cytoplasm and in plastids, and participating in glycolysis and the Calvin-Benson cycle. A general feature of all GAPDH proteins is the presence of an acidic catalytic cysteine in the active site that is overly sensitive to oxidative modifications, including glutathionylation and S-nitrosylation. In Arabidopsis, oxidatively modified cytoplasmic GAPDH has been successfully used as a tool to investigate the role of reduced glutathione, thioredoxins and glutaredoxins in the control of different types of redox post-translational modifications. Oxidative modifications inhibit GAPDH activity, but might enable additional functions in plant cells. Mounting evidence support the concept that plant cytoplasmic GAPDH may fulfill alternative, non-metabolic functions that are triggered by redox post-translational modifications of the protein under stress conditions. The aim of this review is to detail the molecular mechanisms underlying the redox regulation of plant cytoplasmic GAPDH in the light of its crystal structure, and to provide a brief inventory of the well known redox-dependent multi-facetted properties of animal GAPDH, together with the emerging roles of oxidatively modified GAPDH in stress signaling pathways in plants. PMID:24282406

  15. Occurrence, pathology, and ultrastructure of iridovirus and cytoplasmic polyhedrosis viruses in daphnids from the Czech Republic

    Czech Academy of Sciences Publication Activity Database

    Vávra, Jiří; Bílý, Tomáš; Nebesářová, Jana; Federici, B. A.

    2016-01-01

    Roč. 140, Jul 19 (2016), s. 35-38 ISSN 0022-2011 R&D Projects: GA MŠk LM2015062; GA TA ČR(CZ) TE01020118 Institutional support: RVO:60077344 Keywords : iridovirus * cytoplasmic polhedrosis virus * Crustacean * Daphnid * geographical occurrence * Czech Republic * AF microscopy * electron tomography Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 2.379, year: 2016

  16. Differential effect of HDAC3 on cytoplasmic and nuclear huntingtin aggregates.

    Directory of Open Access Journals (Sweden)

    Tatsuo Mano

    Full Text Available Histone deacetylases (HDACs are potential therapeutic targets of polyglutamine (pQ diseases including Huntington's disease (HD that may function to correct aberrant transcriptional deactivation caused by mutant pQ proteins. HDAC3 is a unique class 1 HDAC found in both the cytoplasm and in the nucleus. However, the precise functions of HDAC3 in the two cellular compartments are only vaguely known. HDAC3 directly binds to huntingtin (Htt with short pQ and this interaction is important for suppressing neurotoxicity induced by HDAC3. With long pQ Htt, the interaction with HDAC3 is inhibited, and this supposedly promotes neuronal death, indicating that HDAC3 would be a good therapeutic target for HD. However, the knockout of one HDAC3 allele did not show any efficacy in reducing neurodegenerative symptoms in a mouse model of HD. Therefore, the role of HDAC3 in the pathogenesis of HD has yet to be fully elucidated. We attempted to resolve this issue by focusing on the different roles of HDAC3 on cytoplasmic and nuclear Htt aggregates. In addition to supporting the previous findings, we found that HDAC3 preferentially binds to nuclear Htt over cytoplasmic ones. Specific HDAC3 inhibitors increased the total amount of Htt aggregates by increasing the amount of nuclear aggregates. Both cytoplasmic and nuclear Htt aggregates were able to suppress endogenous HDAC3 activity, which led to decreased nuclear proteasome activity. Therefore, we concluded that Htt aggregates impair nuclear proteasome activity through the inhibition of HDAC3. Our findings provide new insights regarding cross-compartment proteasome regulation.

  17. The role of non-coding RNAs in cytoplasmic male sterility in flowering plants

    Czech Academy of Sciences Publication Activity Database

    Štorchová, Helena

    2017-01-01

    Roč. 18, č. 11 (2017), č. článku 2429. E-ISSN 1422-0067 R&D Projects: GA ČR GA16-09220S Institutional support: RVO:61389030 Keywords : Cytoplasmic male sterility * Gene expression * Global transcriptome * Non-coding RNA * Pollen development Subject RIV: EF - Botanics OBOR OECD: Plant sciences, botany Impact factor: 3.226, year: 2016

  18. Regulation and Impact of Cytoplasmic ARID1A in Ovarian Cancer

    Science.gov (United States)

    2016-03-01

    broadly accepted to be a tumor suppressor in an increasing number of cancers, including ovarian . Silencing ARID1A in ovarian surface epithelium...consequence of its tumor suppressor role in chromatin remodeling and transcriptional regulation. We recently used a well annotated ovarian cancer tissue...and advanced stage disease. In ovarian cancer patients whose tumors have loss of nuclear ARID1A with prevalent cytoplasmic ARID1A had a very grave

  19. Ubiquitin ligase ITCH recruitment suppresses the aggregation and cellular toxicity of cytoplasmic misfolded proteins

    OpenAIRE

    Chhangani, Deepak; Upadhyay, Arun; Amanullah, Ayeman; Joshi, Vibhuti; Mishra, Amit

    2014-01-01

    The protein quality control (QC) system protects cells against cellular toxicity induced by misfolded proteins and maintains overall cellular fitness. Inefficient clearance of or failure to degrade damaged proteins causes several diseases, especially age-linked neurodegenerative disorders. Attenuation of misfolded protein degradation under severe stress conditions leads to the rapid over-accumulation of toxic proteinaceous aggregates in the cytoplasmic compartment. However, the precise cytopl...

  20. Development of Novel Cytoplasmic Male Sterile Source from Dongxiang Wild Rice (Oryza rufipogon

    Directory of Open Access Journals (Sweden)

    Xian-hua SHEN

    2013-09-01

    Full Text Available This study was conducted to develop and characterize a novel cytoplasmic male sterile (CMS source which was identified from Dongxiang wild rice (Oryza rufipogon by crossing Dongxiang wild rice as female with Zhongzao 35, an indica inbred variety, as male and continuous backcrossing with Zhongzao 35. Observation under optical microscope manifested that this novel CMS belonged to typical abortion type with less pollen compared with wild abortive type cytoplasm (CMS-WA. Sequential planting showed that this novel CMS has complete and stable male sterility. Testcross experiment showed that all the 24 tested materials including maintainer and restorer lines of CMS-WA and Honglian type cytoplasm (CMS-HL and other indica inbred varieties are the maintainers with complete maintaining ability, suggesting that this novel CMS has fertility restoration totally different from CMS-WA and CMS-HL and belongs to a novel type of CMS. So far, we only discovered a unique fertility restoration source for this novel CMS. Inheritance analysis showed that the fertility restoration of this CMS was governed by three pairs of independent dominant genes. Prospect for application of this novel CMS system in hybrid rice breeding was also discussed.

  1. Cytoplasmic proteasomes are not indispensable for cell growth in Saccharomyces cerevisiae

    Energy Technology Data Exchange (ETDEWEB)

    Tsuchiya, Hikaru; Arai, Naoko; Tanaka, Keiji, E-mail: tanaka-kj@igakuken.or.jp; Saeki, Yasushi, E-mail: saeki-ys@igakuken.or.jp

    2013-07-05

    Highlights: •We succeeded to control the proteasome localization by the anchor-away technique. •Nuclear proteasome-depleted cells showed a lethal phenotype. •Cytoplasmic proteasomes are not indispensable for cell growth in dividing cells. -- Abstract: The 26S proteasome is an essential protease complex responsible for the degradation of ubiquitinated proteins in eukaryotic cells. In rapidly proliferating yeast cells, proteasomes are mainly localized in the nucleus, but the biological significance of the proteasome localization is still unclear. In this study, we investigated the relationship between the proteasome localization and the functions by the anchor-away technique, a ligand-dependent sequestration of a target protein into specific compartment(s). Anchoring of the proteasome to the plasma membrane or the ribosome resulted in conditional depletion of the nuclear proteasomes, whereas anchoring to histone resulted in the proteasome sequestration into the nucleus. We observed that the accumulation of ubiquitinated proteins in all the proteasome-targeted cells, suggesting that both the nuclear and cytoplasmic proteasomes have proteolytic functions and that the ubiquitinated proteins are produced and degraded in each compartment. Consistent with previous studies, the nuclear proteasome-depleted cells exhibited a lethal phenotype. In contrast, the nuclear sequestration of the proteasome resulted only in a mild growth defect, suggesting that the cytoplasmic proteasomes are not basically indispensable for cell growth in rapidly growing yeast cells.

  2. A pentatricopeptide repeat protein restores nap cytoplasmic male sterility in Brassica napus.

    Science.gov (United States)

    Liu, Zhi; Dong, Faming; Wang, Xiang; Wang, Tao; Su, Rui; Hong, Dengfeng; Yang, Guangsheng

    2017-07-10

    Two forms of male-sterile cytoplasm, designated nap and pol, are found in the oilseed rape species, Brassica napus. The nap cytoplasm is observed in most B. napus varieties, and it confers male sterility on a limited number of cultivars that lack the corresponding restorer gene, Rfn. In the present study, using linkage analysis in combination with 5652 BC1 progeny derived from a cross between a nap cytoplasmic male sterility (CMS) line 181A and a restorer line H5, we delimited the Rfn gene to a 10.5 kb region on chromosome A09, which contained three putative ORFs. Complementation by transformation rescue revealed that the introduction of ORF2, which encodes a pentatricopeptide repeat (PPR) protein, resulted in the recovery of fertility of nap CMS plants. Expression analysis suggested that the Rfn was highly expressed in flower buds and it was preferentially expressed in the tapetum and meiocytes during anther development. Further RNA gel blots and immunodetection suggested that the Rfn gene may play a complicated role in restoring the nap CMS. Our work laid the foundation for dissecting the molecular basis of CMS fertility restoration and the nuclear-mitochondrial interactions in CMS/Rf systems. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  3. Genetic studies on cytoplasmic male sterility in maize. Progress report, April 15, 1990--April 14, 1992

    Energy Technology Data Exchange (ETDEWEB)

    Laughnan, J.R.

    1992-05-01

    Our research concerns the basic mechanisms of cytoplasmic male sterility (CMS) and fertility restoration in maize. The molecular determination of CMS is in the DNA of the mitochondria (mtDNA) but specific nuclear restorer-of-fertility (Rf) genes can overrule the male-sterile effect of the cytoplasm. Our approach to the study of the Rf genes is threefold. We are attempting to tag the cms-S Rf genes and the cms-T Rf2 gene with controlling elements (CEs). Since we have identified a number of spontaneous Rf genes for cms-S and have demonstrated that they are themselves transposable, we are also searching for cases in which an Rf gene is inserted into a wild-type gene. The other aspect of our research involves the nuclear control over the organization of the mitochondrial genome. We found that the changes in mtDNA organization upon cytoplasmic reversion to fertility were characteristic of the nuclear background in which the reversion event occurred. We have investigated whether these differences are a reflection of differences in the organization of the mtDNA genome before reversion.

  4. Cytoplasmic tail domain of glycoprotein B is essential for HHV-6 infection

    Energy Technology Data Exchange (ETDEWEB)

    Mahmoud, Nora F. [Division of Clinical Virology, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe (Japan); Faculty of Pharmacy, Suez Canal University, Ismailia (Egypt); Jasirwan, Chyntia [Division of Clinical Virology, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe (Japan); Division of Hepatobiliary, Department of Internal Medicine, Faculty of Medicine, University of Indonesia (Indonesia); Kanemoto, Satoshi; Wakata, Aika; Wang, Bochao; Hata, Yuuki [Division of Clinical Virology, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe (Japan); Nagamata, Satoshi [Division of Clinical Virology, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe (Japan); Department of Obstetrics and Gynecology, Kobe University Graduate School of Medicine, Kobe (Japan); Kawabata, Akiko [Division of Clinical Virology, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe (Japan); Tang, Huamin [Division of Clinical Virology, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe (Japan); Department of Immunology, Nanjing Medical University, Nanjing (China); Mori, Yasuko, E-mail: ymori@med.kobe-u.ac.jp [Division of Clinical Virology, Center for Infectious Diseases, Kobe University Graduate School of Medicine, Kobe (Japan)

    2016-03-15

    Human herpesvirus 6 (HHV-6) glycoprotein B (gB) is an abundantly expressed viral glycoprotein required for viral entry and cell fusion, and is highly conserved among herpesviruses. The present study examined the function of HHV-6 gB cytoplasmic tail domain (CTD). A gB CTD deletion mutant was constructed which, in contrast to its revertant, could not be reconstituted. Moreover, deletion of gB cytoplasmic tail impaired the intracellular transport of gB protein to the trans-Golgi network (TGN). Taken together, these results suggest that gB CTD is critical for HHV-6 propagation and important for intracellular transportation. - Highlights: • Glycoprotein B (gB) is highly conserved among herpesviruses. • HHV-6 gB is also abundantly expressed in virions. • In the present study, we showed the function of HHV-6 gB cytoplasmic tail domain (CTD). • We found that deletion of gB CTD impairs the intracellular transport of gB protein to the trans-Golgi network (TGN), and CTD of gB is critical for HHV-6 propagation.

  5. Isotachophoresis for fractionation and recovery of cytoplasmic RNA and nucleus from single cells.

    Science.gov (United States)

    Kuriyama, Kentaro; Shintaku, Hirofumi; Santiago, Juan G

    2015-07-01

    There is a substantial need for simultaneous analyses of RNA and DNA from individual single cells. Such analysis provides unique evidence of cell-to-cell differences and the correlation between gene expression and genomic mutation in highly heterogeneous cell populations. We present a novel microfluidic system that leverages isotachophoresis to fractionate and isolate cytoplasmic RNA and genomic DNA (gDNA) from single cells. The system uniquely enables independent, sequence-specific analyses of these critical markers. Our system uses a microfluidic chip with a simple geometry and four end-channel electrodes, and completes the entire process in RNA output reservoirs, each containing high quality and purity aliquots with no measurable cross-contamination of cytoplasmic RNA versus gDNA. We demonstrate our system with simultaneous, sequence-specific quantitation using off-chip RT-qPCR and qPCR for simultaneous cytoplasmic RNA and gDNA analyses, respectively. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  6. The cytoplasm of living cells: a functional mixture of thousands of components

    International Nuclear Information System (INIS)

    Sear, Richard P

    2005-01-01

    Inside every living cell is the cytoplasm: a fluid mixture of thousands of different macromolecules, predominantly proteins. This mixture is where most of the biochemistry occurs that enables living cells to function, and it is perhaps the most complex liquid on earth. Here we take an inventory of what is actually in this mixture. Recent genome-sequencing work has given us for the first time at least some information on all of these thousands of components. Having done so we consider two physical phenomena in the cytoplasm: diffusion and possible phase separation. Diffusion is slower in the highly crowded cytoplasm than in dilute solution. Reasonable estimates of this slow-down can be obtained and their consequences explored; for example, monomer-dimer equilibria are established approximately 20 times more slowly than in a dilute solution. Phase separation in all except exceptional cells appears not to be a problem, despite the high density and so strong protein-protein interactions present. We suggest that this may be partially a by-product of the evolution of other properties, and partially a result of the huge number of components present

  7. Cytoplasmic tail domain of glycoprotein B is essential for HHV-6 infection

    International Nuclear Information System (INIS)

    Mahmoud, Nora F.; Jasirwan, Chyntia; Kanemoto, Satoshi; Wakata, Aika; Wang, Bochao; Hata, Yuuki; Nagamata, Satoshi; Kawabata, Akiko; Tang, Huamin; Mori, Yasuko

    2016-01-01

    Human herpesvirus 6 (HHV-6) glycoprotein B (gB) is an abundantly expressed viral glycoprotein required for viral entry and cell fusion, and is highly conserved among herpesviruses. The present study examined the function of HHV-6 gB cytoplasmic tail domain (CTD). A gB CTD deletion mutant was constructed which, in contrast to its revertant, could not be reconstituted. Moreover, deletion of gB cytoplasmic tail impaired the intracellular transport of gB protein to the trans-Golgi network (TGN). Taken together, these results suggest that gB CTD is critical for HHV-6 propagation and important for intracellular transportation. - Highlights: • Glycoprotein B (gB) is highly conserved among herpesviruses. • HHV-6 gB is also abundantly expressed in virions. • In the present study, we showed the function of HHV-6 gB cytoplasmic tail domain (CTD). • We found that deletion of gB CTD impairs the intracellular transport of gB protein to the trans-Golgi network (TGN), and CTD of gB is critical for HHV-6 propagation.

  8. HDAC4 reduction: a novel therapeutic strategy to target cytoplasmic huntingtin and ameliorate neurodegeneration.

    Directory of Open Access Journals (Sweden)

    Michal Mielcarek

    2013-11-01

    Full Text Available Histone deacetylase (HDAC 4 is a transcriptional repressor that contains a glutamine-rich domain. We hypothesised that it may be involved in the molecular pathogenesis of Huntington's disease (HD, a protein-folding neurodegenerative disorder caused by an aggregation-prone polyglutamine expansion in the huntingtin protein. We found that HDAC4 associates with huntingtin in a polyglutamine-length-dependent manner and co-localises with cytoplasmic inclusions. We show that HDAC4 reduction delayed cytoplasmic aggregate formation, restored Bdnf transcript levels, and rescued neuronal and cortico-striatal synaptic function in HD mouse models. This was accompanied by an improvement in motor coordination, neurological phenotypes, and increased lifespan. Surprisingly, HDAC4 reduction had no effect on global transcriptional dysfunction and did not modulate nuclear huntingtin aggregation. Our results define a crucial role for the cytoplasmic aggregation process in the molecular pathology of HD. HDAC4 reduction presents a novel strategy for targeting huntingtin aggregation, which may be amenable to small-molecule therapeutics.

  9. Deciphering the role of nuclear and cytoplasmic IKKα in skin cancer

    Science.gov (United States)

    Alameda, Josefa P.; Gaspar, Miriam; Ramírez, Ángel; Navarro, Manuel; Page, Angustias; Suárez-Cabrera, Cristian; Fernández, M. Guadalupe; Mérida, Jose R.; Paramio, Jesús M.; García-Fernández, Rosa A.; Fernández-Aceñero, M. Jesús; Casanova, M. Llanos

    2016-01-01

    Nonmelanoma skin cancers (NMSC) are the most common human malignancies. IKKα is an essential protein for skin development and is also involved in the genesis and progression of NMSC, through mechanisms not fully understood. While different studies show that IKKα protects against skin cancer, others indicate that it promotes NMSC. To resolve this controversy we have generated two models of transgenic mice expressing the IKKα protein in the nucleus (N-IKKα mice) or the cytoplasm (C-IKKα mice) of keratinocytes. Chemical skin carcinogenesis experiments show that tumors developed by both types of transgenic mice exhibit histological and molecular characteristics that make them more prone to progression and invasion than those developed by Control mice. However, the mechanisms through which IKKα promotes skin tumors are different depending on its subcellular localization; while IKKα of cytoplasmic localization increases EGFR, MMP-9 and VEGF-A activities in tumors, nuclear IKKα causes tumor progression through regulation of c-Myc, Maspin and Integrin-α6 expression. Additionally, we have found that N-IKKα skin tumors mimic the characteristics associated to aggressive human skin tumors with high risk to metastasize. Our results show that IKKα has different non-overlapping roles in the nucleus or cytoplasm of keratinocytes, and provide new targets for intervention in human NMSC progression. PMID:27121058

  10. Involvement of hGLD-2 in cytoplasmic polyadenylation of human p53 mRNA

    DEFF Research Database (Denmark)

    Glahder, Jacob-Andreas Harald; Norrild, Bodil

    2011-01-01

    Cytoplasmic polyadenylation is a post-transcriptional mechanism regulating mRNA stability and translation. The human p53 3'-untranslated region (3'-UTR) contains two regions similar to cytoplasmic polyadenylation elements (CPEs) just upstream of the poly(A) hexanucleotide. Evaluation of the p53 CPE......-like elements was performed by luciferase reporter assays, qPCR, and poly(A) assays. Herein, we report the down regulation of a luciferase reporter fused to the p53 3'-UTR, when human CPE-binding protein 1 (hCPEB1) is overexpressed. This inhibition is partially rescued when hCPEB1fused to hGLD-2 [a human...... cytoplasmic poly(A) polymerase] is overexpressed instead. The stability of a luciferase mRNA containing the p53 3'-UTR downstream, is decreased when hCPEB1 is overexpressed as seen by qPCR. Expression of hGLD-2 restores the mRNA stability. This is due to elongation of the poly(A) tail as seen by a PCR...

  11. Ubiquitin ligase ITCH recruitment suppresses the aggregation and cellular toxicity of cytoplasmic misfolded proteins.

    Science.gov (United States)

    Chhangani, Deepak; Upadhyay, Arun; Amanullah, Ayeman; Joshi, Vibhuti; Mishra, Amit

    2014-05-28

    The protein quality control (QC) system protects cells against cellular toxicity induced by misfolded proteins and maintains overall cellular fitness. Inefficient clearance of or failure to degrade damaged proteins causes several diseases, especially age-linked neurodegenerative disorders. Attenuation of misfolded protein degradation under severe stress conditions leads to the rapid over-accumulation of toxic proteinaceous aggregates in the cytoplasmic compartment. However, the precise cytoplasmic quality control degradation mechanism is unknown. In the present study, we demonstrate that the Nedd4-like E3 ubiquitin ligase ITCH specifically interacts with mutant bona fide misfolded proteins and colocalizes with their perinuclear aggregates. In a cell culture model, we demonstrate ITCH recruitment by cytoplasmic inclusions containing polyglutamine-expanded huntingtin or ataxin-3 proteins. Transient overexpression of ITCH dramatically induced the degradation of thermally denatured misfolded luciferase protein. Partial depletion of ITCH increased the rate of aggregate formation and cell death generated by expanded polyglutamine proteins. Finally, we demonstrate that overexpression of ITCH alleviates the cytotoxic potential of expanded polyglutamine proteins and reduces aggregation. These observations indicate that ITCH is involved in the cytosolic quality control pathway and may help to explain how abnormal proteins are targeted by QC ubiquitin-protein ligases.

  12. Particle-Rich Cytoplasmic Structure (PaCS: Identification, Natural History, Role in Cell Biology and Pathology

    Directory of Open Access Journals (Sweden)

    Enrico Solcia

    2014-09-01

    Full Text Available Cytoplasmic structures showing a selective concentration of both polyubiquitinated proteins and proteasome have been described in various epithelial, hematopoietic, mesenchymal and neural cells in vitro or in fetal tissues, as well as in chronically-infected, mutated preneoplastic and neoplastic tissues. These cytoplasmic structures differ from other ubiquitin-reactive cytoplasmic bodies, like sequestosomes, aggresome-like-induced structures in dendritic cells (DALIS/non-dendritic cells (ALIS and aggresomes in showing distinctive ultrastructural organization (particle-rich cytoplasmic structure or PaCS, a cytochemical pattern and a functional profile. Their formation can be induced in vitro in dendritic or natural killer cells by trophic factors and interleukin treatment. They originate in close connection with ribosomes, while, as a result of their growth, the cytoskeleton and other surrounding organelles are usually dislocated outside their core. Interestingly, these particulate cytoplasmic structures are often found to fill cytoplasmic blebs forming proteasome- and polyubiquitinated protein-discharging vesicles, called ectosomes, which are found to detach from the cell and freely float in the extracellular space. To clearly point out the importance of the polyubiquitinated proteins and proteasome containing cytoplasmic structures, their role in cell biology and pathology has been carefully analyzed.

  13. Influence of Genetic Drift of Restoring Gene (Rf on Seed Purity of Yuetai A, a Honglian-Type Cytoplasmic Male Sterile Line in Hybrid Rice

    Directory of Open Access Journals (Sweden)

    Ji-feng Wang

    2008-06-01

    Full Text Available The seed samples of Yuetai A, a Honglian (HL type cytoplasmic male sterile (CMS line in hybrid rice were investigated to assess the seed purity and to analyze the cause of off-type plants by imitating the biological contaminant to Yuetai A in Nanjing, Jiangsu Province and Lingshui, Hainan Province during 2004–2006. The seed impurity of Yuetai A mainly resulted from the genetic drift of restoring fertility gene (Rf after biological contamination but not from its sterility unstability. All of the ten maintainer lines, five restorer lines and three thermo-sensitive genic male sterile lines used in the study could pollinate Yuetai A and Yuetai B to produce F1 plants, directly or indirectly resulting in Rf-gene drifting into Yuetai A and generating ‘iso-cytoplasm restoring-generations’. Furthermore, high outcrossing rate and similar heading date of Yuetai A with many varieties used in rice production might easily result in the biological contamination. After removing all plants with Rf-gene mixed in Yuetai A and preventing Rf-gene drifting into Yuetai A, the seed purity of Yuetai A and Yuetai B had been raised to 100%.

  14. Preventing eating disorders.

    Science.gov (United States)

    Shaw, Heather; Stice, Eric; Becker, Carolyn Black

    2009-01-01

    This article reviews eating disorder (ED) prevention programs, highlighting features that define successful programs and particularly promising interventions, and how they might be further refined. The field of ED prevention has advanced considerably both theoretically and methodologically compared with the earlier ED prevention programs, which were largely psychoeducational and met with limited success. Recent meta-analytic findings show that more than half (51%) of ED prevention interventions reduced ED risk factors and more than a quarter (29%) reduced current or future eating pathology (EP). A couple of brief programs have been shown to reduce the risk for future onset of EP and obesity. Selected interactive, multisession programs offered to participants older than 15 years, delivered by professional interventionists and including body acceptance or dissonance-induction content, produced larger effects. Understanding and applying these results can help inform the design of more effective prevention programs in the future.

  15. Nucleocapsid Protein from Fig Mosaic Virus Forms Cytoplasmic Agglomerates That Are Hauled by Endoplasmic Reticulum Streaming

    Science.gov (United States)

    Ishikawa, Kazuya; Miura, Chihiro; Maejima, Kensaku; Komatsu, Ken; Hashimoto, Masayoshi; Tomomitsu, Tatsuya; Fukuoka, Misato; Yusa, Akira; Yamaji, Yasuyuki

    2014-01-01

    ABSTRACT Although many studies have demonstrated intracellular movement of viral proteins or viral replication complexes, little is known about the mechanisms of their motility. In this study, we analyzed the localization and motility of the nucleocapsid protein (NP) of Fig mosaic virus (FMV), a negative-strand RNA virus belonging to the recently established genus Emaravirus. Electron microscopy of FMV-infected cells using immunogold labeling showed that NPs formed cytoplasmic agglomerates that were predominantly enveloped by the endoplasmic reticulum (ER) membrane, while nonenveloped NP agglomerates also localized along the ER. Likewise, transiently expressed NPs formed agglomerates, designated NP bodies (NBs), in close proximity to the ER, as was the case in FMV-infected cells. Subcellular fractionation and electron microscopic analyses of NP-expressing cells revealed that NBs localized in the cytoplasm. Furthermore, we found that NBs moved rapidly with the streaming of the ER in an actomyosin-dependent manner. Brefeldin A treatment at a high concentration to disturb the ER network configuration induced aberrant accumulation of NBs in the perinuclear region, indicating that the ER network configuration is related to NB localization. Dominant negative inhibition of the class XI myosins, XI-1, XI-2, and XI-K, affected both ER streaming and NB movement in a similar pattern. Taken together, these results showed that NBs localize in the cytoplasm but in close proximity to the ER membrane to form enveloped particles and that this causes passive movements of cytoplasmic NBs by ER streaming. IMPORTANCE Intracellular trafficking is a primary and essential step for the cell-to-cell movement of viruses. To date, many studies have demonstrated the rapid intracellular movement of viral factors but have failed to provide evidence for the mechanism or biological significance of this motility. Here, we observed that agglomerates of nucleocapsid protein (NP) moved rapidly

  16. Coordinated movement of cytoplasmic and transmembrane domains of RyR1 upon gating.

    Directory of Open Access Journals (Sweden)

    Montserrat Samsó

    2009-04-01

    Full Text Available Ryanodine receptor type 1 (RyR1 produces spatially and temporally defined Ca2+ signals in several cell types. How signals received in the cytoplasmic domain are transmitted to the ion gate and how the channel gates are unknown. We used EGTA or neuroactive PCB 95 to stabilize the full closed or open states of RyR1. Single-channel measurements in the presence of FKBP12 indicate that PCB 95 inverts the thermodynamic stability of RyR1 and locks it in a long-lived open state whose unitary current is indistinguishable from the native open state. We analyzed two datasets of 15,625 and 18,527 frozen-hydrated RyR1-FKBP12 particles in the closed and open conformations, respectively, by cryo-electron microscopy. Their corresponding three-dimensional structures at 10.2 A resolution refine the structure surrounding the ion pathway previously identified in the closed conformation: two right-handed bundles emerging from the putative ion gate (the cytoplasmic "inner branches" and the transmembrane "inner helices". Furthermore, six of the identifiable transmembrane segments of RyR1 have similar organization to those of the mammalian Kv1.2 potassium channel. Upon gating, the distal cytoplasmic domains move towards the transmembrane domain while the central cytoplasmic domains move away from it, and also away from the 4-fold axis. Along the ion pathway, precise relocation of the inner helices and inner branches results in an approximately 4 A diameter increase of the ion gate. Whereas the inner helices of the K+ channels and of the RyR1 channel cross-correlate best with their corresponding open/closed states, the cytoplasmic inner branches, which are not observed in the K+ channels, appear to have at least as important a role as the inner helices for RyR1 gating. We propose a theoretical model whereby the inner helices, the inner branches, and the h1 densities together create an efficient novel gating mechanism for channel opening by relaxing two right

  17. A novel nucleo-cytoplasmic hybrid clone formed via androgenesis in polyploid gibel carp

    Directory of Open Access Journals (Sweden)

    Zhou Li

    2011-03-01

    Full Text Available Abstract Background Unisexual vertebrates have been demonstrated to reproduce by gynogenesis, hybridogenesis, parthenogenesis, or kleptogenesis, however, it is uncertain how the reproduction mode contributes to the clonal diversity. Recently, polyploid gibel carp has been revealed to possess coexisting dual modes of unisexual gynogenesis and sexual reproduction and to have numerous various clones. Using sexual reproduction mating between clone D female and clone A male and subsequent 7 generation multiplying of unisexual gynogenesis, we have created a novel clone strain with more than several hundred millions of individuals. Here, we attempt to identify genetic background of the novel clone and to explore the significant implication for clonal diversity contribution. Methods Several nuclear genome markers and one cytoplasmic marker, the mitochondrial genome sequence, were used to identify the genetic organization of the randomly sampled individuals from different generations of the novel clone. Results Chromosome number, Cot-1 repetitive DNA banded karyotype, microsatellite patterns, AFLP profiles and transferrin alleles uniformly indicated that nuclear genome of the novel clone is identical to that of clone A, and significantly different from that of clone D. However, the cytoplasmic marker, its complete mtDNA genome sequence, is same to that of clone D, and different from that of clone A. Conclusions The present data indicate that the novel clone is a nucleo-cytoplasmic hybrid between the known clones A and D, because it originates from the offspring of gonochoristic sexual reproduction mating between clone D female and clone A male, and contains an entire nuclear genome from the paternal clone A and a mtDNA genome (cytoplasm from the maternal clone D. It is suggested to arise via androgenesis by a mechanism of ploidy doubling of clone A sperm in clone D ooplasm through inhibiting the first mitotic division. Significantly, the selected nucleo-cytoplasmic

  18. A novel nucleo-cytoplasmic hybrid clone formed via androgenesis in polyploid gibel carp

    Science.gov (United States)

    2011-01-01

    Background Unisexual vertebrates have been demonstrated to reproduce by gynogenesis, hybridogenesis, parthenogenesis, or kleptogenesis, however, it is uncertain how the reproduction mode contributes to the clonal diversity. Recently, polyploid gibel carp has been revealed to possess coexisting dual modes of unisexual gynogenesis and sexual reproduction and to have numerous various clones. Using sexual reproduction mating between clone D female and clone A male and subsequent 7 generation multiplying of unisexual gynogenesis, we have created a novel clone strain with more than several hundred millions of individuals. Here, we attempt to identify genetic background of the novel clone and to explore the significant implication for clonal diversity contribution. Methods Several nuclear genome markers and one cytoplasmic marker, the mitochondrial genome sequence, were used to identify the genetic organization of the randomly sampled individuals from different generations of the novel clone. Results Chromosome number, Cot-1 repetitive DNA banded karyotype, microsatellite patterns, AFLP profiles and transferrin alleles uniformly indicated that nuclear genome of the novel clone is identical to that of clone A, and significantly different from that of clone D. However, the cytoplasmic marker, its complete mtDNA genome sequence, is same to that of clone D, and different from that of clone A. Conclusions The present data indicate that the novel clone is a nucleo-cytoplasmic hybrid between the known clones A and D, because it originates from the offspring of gonochoristic sexual reproduction mating between clone D female and clone A male, and contains an entire nuclear genome from the paternal clone A and a mtDNA genome (cytoplasm) from the maternal clone D. It is suggested to arise via androgenesis by a mechanism of ploidy doubling of clone A sperm in clone D ooplasm through inhibiting the first mitotic division. Significantly, the selected nucleo-cytoplasmic hybrid female

  19. Preventing Breast Cancer: Making Progress

    Science.gov (United States)

    ... Home Current Issue Past Issues Preventing Breast Cancer: Making Progress Past Issues / Fall 2006 Table of Contents ... the time a woman is taking the pills, notes Leslie Ford, M.D., associate director for NCI's ...

  20. Genetics, Disease Prevention and Treatment

    Science.gov (United States)

    Skip to main content FAQ About Genetics, Disease Prevention and Treatment Enter Search Term(s): Español Research Funding An Overview Bioinformatics Current Grants Education and Training Funding Extramural Research News ...

  1. Content identification: binary content fingerprinting versus binary content encoding

    Science.gov (United States)

    Ferdowsi, Sohrab; Voloshynovskiy, Svyatoslav; Kostadinov, Dimche

    2014-02-01

    In this work, we address the problem of content identification. We consider content identification as a special case of multiclass classification. The conventional approach towards identification is based on content fingerprinting where a short binary content description known as a fingerprint is extracted from the content. We propose an alternative solution based on elements of machine learning theory and digital communications. Similar to binary content fingerprinting, binary content representation is generated based on a set of trained binary classifiers. We consider several training/encoding strategies and demonstrate that the proposed system can achieve the upper theoretical performance limits of content identification. The experimental results were carried out both on a synthetic dataset with different parameters and the FAMOS dataset of microstructures from consumer packages.

  2. Susceptibility to virus-cell fusion at the plasma membrane is reduced through expression of HIV gp41 cytoplasmic domains

    International Nuclear Information System (INIS)

    Malinowsky, Katharina; Luksza, Julia; Dittmar, Matthias T.

    2008-01-01

    The cytoplasmic tail of the HIV transmembrane protein plays an important role in viral infection. In this study we analyzed the role of retroviral cytoplasmic tails in modulating the cytoskeleton and interfering with virus-cell fusion. HeLaP4 cells expressing different HIV cytoplasmic tail constructs showed reduced acetylated tubulin levels whereas the cytoplasmic tail of MLV did not alter microtubule stability indicating a unique function for the lentiviral cytoplasmic tail. The effect on tubulin is mediated through the membrane proximal region of the HIV cytoplasmic tail and was independent of membrane localization. Site-directed mutagenesis identified three motifs in the HIV-2 cytoplasmic tail required to effect the reduction in acetylated tubulin. Both the YxxΦ domain and amino acids 21 to 45 of the HIV-2 cytoplasmic tail need to be present to change the level of acetylated tubulin in transfected cells. T-cells stably expressing one HIV-2 cytoplasmic tail derived construct showed also a reduction in acetylated tubulin thus confirming the importance of this effect not only for HeLaP4 and 293T cells. Challenge experiments using transiently transfected HeLaP4 cells and T cells stably expressing an HIV cytoplasmic tail construct revealed both reduced virus-cell fusion and replication of HIV-1 NL4.3 compared to control cells. In the virus-cell fusion assay only virions pseudotyped with either HIV or MLV envelopes showed reduced fusion efficiency, whereas VSV-G pseudotyped virions where not affected by the expression of HIV derived cytoplasmic tail constructs, indicating that fusion at the plasma but not endosomal membrane is affected. Overexpression of human histone-deacetylase 6 (HDAC6) and constitutively active RhoA resulted in a reduction of acetylated tubulin and reduced virus-cell fusion as significant as that observed following expression of HIV cytoplasmic tail constructs. Inhibition of HDAC6 showed a strong increase in acetylated tubulin and increase of

  3. Chemotaxis cluster 1 proteins form cytoplasmic arrays in Vibrio cholerae and are stabilized by a double signaling domain receptor DosM

    DEFF Research Database (Denmark)

    Briegel, Ariane; Ortega, Davi R; Mann, Petra

    2016-01-01

    motile bacteria contain one or more additional, sometimes purely cytoplasmic, chemoreceptor systems. Vibrio cholerae contains three chemotaxis clusters (I, II, and III). Here, using electron cryotomography, we explore V. cholerae's cytoplasmic chemoreceptor array and establish that it is formed...

  4. Plasma exchange and glucocorticoid dosing in the treatment of anti-neutrophil cytoplasm antibody associated vasculitis (PEXIVAS)

    DEFF Research Database (Denmark)

    Walsh, Michael; Merkel, Peter A; Peh, Chen Au

    2013-01-01

    Granulomatosis with polyangiitis (GPA, Wegener's) and microscopic polyangiitis (MPA) are small vessel vasculitides collectively referred to as anti-neutrophil cytoplasm antibody-associated vasculitis (AAV). AAV is associated with high rates of morbidity and mortality due to uncontrolled disease...

  5. Two nuclear export signals of Cdc6 are differentially associated with CDK-mediated phosphorylation residues for cytoplasmic translocation.

    Science.gov (United States)

    Hwang, In Sun; Woo, Sang Uk; Park, Ji-Woong; Lee, Seung Ki; Yim, Hyungshin

    2014-02-01

    Cdc6 is cleaved at residues 442 and 290 by caspase-3 during apoptosis producing p49-tCdc6 and p32-tCdc6, respectively. While p32-tCdc6 is unable to translocate into the cytoplasm, p49-tCdc6 retains cytoplasmic translocation activity, but it has a lower efficiency than wild-type Cdc6. We hypothesized that a novel nuclear export signal (NES) sequence exists between amino acids 290 and 442. Cdc6 contains a novel NES in the region of amino acids 300-315 (NES2) that shares sequence similarity with NES1 at residues 462-476. In mutant versions of Cdc6, we replaced leucine with alanine in NES1 and NES2 and co-expressed the mutant constructs with cyclin A. We observed that the cytoplasmic translocation of these mutants was reduced in comparison to wild-type Cdc6. Moreover, the cytoplasmic translocation of a mutant in which all four leucine residues were mutated to alanine was significantly inhibited in comparison to the translocation of wild-type Cdc6. The Crm1 binding activities of Cdc6 NES mutants were consistent with the efficiency of its cytoplasmic translocation. Further studies have revealed that L468 and L470 of NES1 are required for cytoplasmic translocation of Cdc6 phosphorylated at S74, while L311 and L313 of NES2 accelerate the cytoplasmic translocation of Cdc6 phosphorylated at S54. These results suggest that the two NESs of Cdc6 work cooperatively and distinctly for the cytoplasmic translocation of Cdc6 phosphorylated at S74 and S54 by cyclin A/Cdk2. Copyright © 2013 Elsevier B.V. All rights reserved.

  6. Accessory factors of cytoplasmic viral RNA sensors required for antiviral innate immune response

    Directory of Open Access Journals (Sweden)

    Hiroyuki eOshiumi

    2016-05-01

    Full Text Available Type I interferon (IFN induces many antiviral factors in host cells. RIG-I-like receptors (RLRs are cytoplasmic viral RNA sensors that trigger the signal to induce the innate immune response that includes type I IFN production. RIG-I and MDA5 are RLRs that form nucleoprotein filaments along viral double-stranded RNA, resulting in the activation of MAVS adaptor molecule. The MAVS protein forms a prion-like aggregation structure, leading to type I IFN production. RIG-I and MDA5 undergo post-translational modification. TRIM25 and Riplet ubiquitin ligases deliver a K63-linked polyubiquitin moiety to the RIG-I N-terminal caspase activation and recruitment domains (CARDs and C-terminal region; the polyubiquitin chain then stabilizes the two-CARD tetramer structure required for MAVS assembly. MDA5 activation is regulated by phosphorylation. RIOK3 is a protein kinase that phosphorylates the MDA5 protein in a steady state, and PP1α/γ dephosphorylate this protein, resulting in its activation. RIG-I and MDA5 require cytoplasmic RNA helicases for their efficient activation. LGP2, another RLR, is an RNA helicase involved in RLR signaling. This protein does not possess N-terminal CARDs and thus cannot trigger downstream signaling by itself. Recent studies have revealed that this protein modulates MDA5 filament formation, resulting in enhanced type I IFN production. Several other cytoplasmic RNA helicases are involved in RLR signaling. DDX3, DHX29, DHX36, and DDX60 RNA helicases have been reported to be involved in RLR-mediated type I IFN production after viral infection. However, the underlying mechanism is largely unknown. Future studies are required to reveal the role of RNA helicases in the RLR signaling pathway.

  7. Secretion of Bacterial Lipoproteins: Through the Cytoplasmic Membrane, the Periplasm and Beyond

    Science.gov (United States)

    Zückert, Wolfram R.

    2014-01-01

    Bacterial lipoproteins are peripherally anchored membrane proteins that play a variety of roles in bacterial physiology and virulence in monoderm (single membrane-enveloped, e.g., grampositive) and diderm (double membrane-enveloped, e.g., gram-negative) bacteria. After export of prolipoproteins through the cytoplasmic membrane, which occurs predominantly but not exclusively via the general secretory or Sec pathway, the proteins are lipid-modified at the cytoplasmic membrane in a multistep process that involves sequential modification of a cysteine residue and cleavage of the signal peptide by the signal II peptidase Lsp. In both monoderms and diderms, signal peptide processing is preceded by acylation with a diacylglycerol through preprolipoprotein diacylglycerol transferase (Lgt). In diderms but also some monoderms, lipoproteins are further modified with a third acyl chain through lipoprotein N-acyl transferase (Lnt). Fully modified lipoproteins that are destined to be anchored in the inner leaflet of the outer membrane (OM) are selected, transported and inserted by the Lol (lipoprotein outer membrane localization) pathway machinery, which consists of the inner-membrane (IM) ABC transporterlike LolCDE complex, the periplasmic LolA chaperone and the OM LolB lipoprotein receptor. Retention of lipoproteins in the cytoplasmic membrane results from Lol avoidance signals that were originally described as the “+2 rule”. Surface localization of lipoproteins in diderms is rare in most bacteria, with the exception of several spirochetal species. Type 2 (T2SS) and type 5 (T5SS) secretion systems are involved in secretion of specific surface lipoproteins of γ-proteobacteria. In the model spirochete Borrelia burgdorferi, surface lipoprotein secretion does not follow established sorting rules, but remains dependent on N-terminal peptide sequences. Secretion through the outer membrane requires maintenance of lipoproteins in a translocation-competent unfolded conformation

  8. Occurrence of proteinaceous 10-nm filaments throughout the cytoplasm of algae of the order Dasycladales.

    Science.gov (United States)

    Berger, S; Wittke, W; Traub, P

    1998-05-01

    Previously, whole-mount electron microscopy of nuclei extruded together with residual cytoplasm from the rhizoids of several algal species of the order Dasycladales has revealed the occurrence of an intra- and perinuclear network of 10-nm filaments morphologically indistinguishable from that of mammalian vimentin intermediate filaments. The present investigation demonstrates the existence of a filament system throughout the cytoplasm of the rhizoid, stalk, and apical tip of these giant cells. However, while the perinuclear 10-nm filaments interconnecting the nuclear surface with a perinuclear layer of large, electron-dense bodies filled with nucleoprotein material are of smooth appearance, those continuing within and beyond the perinuclear bodies are densely covered with differently sized, globular structures and, therefore, are of a very rough appearance. The filaments in the very apical tip of the cells are mainly of the smooth type. The transition from smooth to rough filaments seems to occur in the numerous perinuclear dense bodies surrounding the large nucleus. Digestion of the rough filaments with proteinase K removes the globules from the filament surface, revealing that throughout the nonvacuolar, intracellular space the filaments have the same basic 10-nm structure. On the other hand, gold-conjugated RNase A strongly binds to the filament-attached globules but not to the smooth, perinuclear, and the proteinase K-treated, rough filaments. In addition, an antibody raised against Xp54, a highly conserved protein which in Xenopus oocytes is an integral component of stored mRNP particles, decorates the rough but not the smooth 10-nm filaments. These results support the notion that the 10-nm filament system of Dasycladales cells plays a role in the transient storage of ribonucleoprotein particles in the cytoplasm and possibly fulfils a supportive function in the actomyosin-based transport of such material to various cytological destinations.

  9. Divergent evolution in the cytoplasmic domains of PRLR and GHR genes in Artiodactyla

    Directory of Open Access Journals (Sweden)

    Li Meng-Hua

    2009-07-01

    Full Text Available Abstract Background Prolactin receptor (PRLR and growth hormone receptor (GHR belong to the large superfamily of class 1 cytokine receptors. Both of them have been identified as candidate genes affecting key quantitative traits, like growth and reproduction in livestock. We have previously studied the molecular anatomy of the cytoplasmic domain of GHR in different cattle breeds and artiodactyl species. In this study we have analysed the corresponding cytoplasmic signalling region of PRLR. Results We sequenced PRLR gene exon 10, coding for the major part of the cytoplasmic domain, from cattle, American bison, European bison, yak, sheep, pig and wild boar individuals. We found different patterns of variation in the two receptors within and between ruminants and pigs. Pigs and bison species have no variation within GHR exon 10, but show high haplotype diversity for the PRLR exon 10. In cattle, PRLR shows lower diversity than GHR. The Bovinae PRLR haplotype network fits better the known phylogenetic relationships between the species than that of the GHR, where differences within cattle breeds are larger than between the different species in the subfamily. By comparison with the wild boar haplotypes, a high number of subsequent nonsynonymous substitutions seem to have accumulated in the pig PRLR exon 10 after domestication. Conclusion Both genes affect a multitude of traits that have been targets of selection after domestication. The genes seem to have responded differently to different selection pressures imposed by human artificial selection. The results suggest possible effects of selective sweeps in GHR before domestication in the pig lineage or species divergence in the Bison lineage. The PRLR results may be explained by strong directional selection in pigs or functional switching.

  10. Imaging Nuclear-Cytoplasmic Dynamics in Primary and Metastatic Colon Cancer in Nude Mice.

    Science.gov (United States)

    Hasegawa, Kosuke; Suetsugu, Atsushi; Nakamura, Miki; Matsumoto, Takuro; Aoki, Hitomi; Kunisada, Takahiro; Bouvet, Michael; Shimizu, Masahito; Hoffman, Robert M

    2016-05-01

    Colon cancer frequently results in metastasis to the liver, where it becomes the main cause of death. However, the cell cycle in primary tumors and metastases is poorly understood. We developed a mouse model of liver metastasis using the human colon cancer cell line HCT-116, which expresses green fluorescent protein (GFP) in the nucleus and red fluorescent protein (RFP) in the cytoplasm (HCT-116-GFP-RFP). HCT-116 GFP-RFP cells were injected into the spleen of nu/nu nude mice. HCT-116-GFP-RFP cells subsequently formed primary tumors in the spleen, as well as metastatic colonies in the liver and retroperitoneum by 28 days after cell transplantation. Using an Olympus FV1000 confocal microscope, it was possible to clearly image mitosis of the dual-colored colon cancer cells in the primary tumor as well as liver and other metastases. Multi-nucleate cancer cells, in addition to mono-nucleate cancer cells and their mitosis, were observed in the primary tumor and metastasis. Multi-nucleate HCT-116-GFP-RFP cells were also observed after culture of the primary and metastatic tumors. A similar ratio of mono-nucleate, multi-nucleate, and mitotic cells grew from the primary and metastatic tumors in culture, suggesting similarity of the nuclear-cytoplasmic dynamics of primary and metastatic cancer cells, further emphasizing the stochastic nature of metastasis. Our results demonstrate a similar heterogeneity of nuclear-cytoplasmic dynamics within primary tumors and metastases, which may be an important factor in the stochastic nature of metastasis. Copyright© 2016 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  11. Structural and biophysical characterization of the cytoplasmic domains of human BAP29 and BAP31.

    Directory of Open Access Journals (Sweden)

    Esben M Quistgaard

    Full Text Available Two members of the B-cell associated 31 (BAP31 family are found in humans; BAP29 and BAP31. These are ubiquitously expressed receptors residing in the endoplasmic reticulum. BAP31 functions in sorting of membrane proteins and in caspase-8 mediated apoptosis, while BAP29 appears to mainly corroborate with BAP31 in sorting. The N-terminal half of these proteins is membrane-bound while the C-terminal half is cytoplasmic. The latter include the so called variant of death effector domain (vDED, which shares weak sequence homology with DED domains. Here we present two structures of BAP31 vDED determined from a single and a twinned crystal, grown at pH 8.0 and pH 4.2, respectively. These structures show that BAP31 vDED forms a dimeric parallel coiled coil with no structural similarity to DED domains. Solution studies support this conclusion and strongly suggest that an additional α-helical domain is present in the C-terminal cytoplasmic region, probably forming a second coiled coil. The thermal stability of BAP31 vDED is quite modest at neutral pH, suggesting that it may assemble in a dynamic fashion in vivo. Surprisingly, BAP29 vDED is partially unfolded at pH 7, while a coiled coil is formed at pH 4.2 in vitro. It is however likely that folding of the domain is triggered by other factors than low pH in vivo. We found no evidence for direct interaction of the cytoplasmic domains of BAP29 and BAP31.

  12. Cell cycle-dependent microtubule-based dynamic transport of cytoplasmic dynein in mammalian cells.

    Directory of Open Access Journals (Sweden)

    Takuya Kobayashi

    Full Text Available BACKGROUND: Cytoplasmic dynein complex is a large multi-subunit microtubule (MT-associated molecular motor involved in various cellular functions including organelle positioning, vesicle transport and cell division. However, regulatory mechanism of the cell-cycle dependent distribution of dynein has not fully been understood. METHODOLOGY/PRINCIPAL FINDINGS: Here we report live-cell imaging of cytoplasmic dynein in HeLa cells, by expressing multifunctional green fluorescent protein (mfGFP-tagged 74-kDa intermediate chain (IC74. IC74-mfGFP was successfully incorporated into functional dynein complex. In interphase, dynein moved bi-directionally along with MTs, which might carry cargos such as transport vesicles. A substantial fraction of dynein moved toward cell periphery together with EB1, a member of MT plus end-tracking proteins (+TIPs, suggesting +TIPs-mediated transport of dynein. In late-interphase and prophase, dynein was localized at the centrosomes and the radial MT array. In prometaphase and metaphase, dynein was localized at spindle MTs where it frequently moved from spindle poles toward chromosomes or cell cortex. +TIPs may be involved in the transport of spindle dyneins. Possible kinetochore and cortical dyneins were also observed. CONCLUSIONS AND SIGNIFICANCE: These findings suggest that cytoplasmic dynein is transported to the site of action in preparation for the following cellular events, primarily by the MT-based transport. The MT-based transport may have greater advantage than simple diffusion of soluble dynein in rapid and efficient transport of the limited concentration of the protein.

  13. Nuclear, but not cytoplasmic, localization of survivin as a negative prognostic factor for survival in upper urinary tract urothelial carcinoma.

    Science.gov (United States)

    Kitamura, Hiroshi; Torigoe, Toshihiko; Hirohashi, Yoshihiko; Asanuma, Hiroko; Inoue, Ryuta; Nishida, Sachiyo; Tanaka, Toshiaki; Masumori, Naoya; Sato, Noriyuki; Tsukamoto, Taiji

    2013-01-01

    Survivin, a member of the inhibitor of apoptosis protein gene family, inhibits apoptosis and promotes mitosis. We determined whether nuclear or cytoplasmic localization of survivin could predict survival of patients with upper urinary tract urothelial carcinoma (UUTUC). Immunohistochemical staining for survivin was carried out on archival specimens from 125 consecutive patients with UUTUC who underwent radical nephroureterectomy. Nuclear and cytoplasmic staining of survivin was scored and compared with clinicopathologic features and cancer-specific survival (CSS). Nuclear expression of survivin was significantly correlated with tumor grade (p nuclear expression of survivin vs. 73 % for those without nuclear expression of survivin (hazard ratio = 2.19; 95 % confidence interval = 1.02-4.70; p = 0.043). The 5-year cancer-specific survival rates of patients with cytoplasmic survivin-negative and -positive tumors were 66 and 67 %, respectively. There was no difference in survival between patients with cytoplasmic survivin-negative tumors and those with cytoplasmic survivin-positive tumors. Using univariate analysis, nuclear survivin expression, tumor grade, pathological T stage, pathological N stage, and lymphovascular invasion were the predictive variables for CSS. In contrast, cytoplasmic survivin expression had no prognostic relevance. These data suggest that nuclear accumulation of survivin represents biologic aggressiveness and that nuclear survivin is a negative prognostic marker in patients with resected UUTUC.

  14. Characterization of phosphorylation sites in the cytoplasmic domain of the 300 kDa mannose-6-phosphate receptor

    DEFF Research Database (Denmark)

    Rosorius, O; Mieskes, G; Issinger, O G

    1993-01-01

    The human 300 kDa mannose-6-phosphate receptor (MPR 300) is phosphorylated in vivo at serine residues of its cytoplasmic domain. Two-dimensional separation can resolve tryptic phosphopeptides into four major species. To identify the kinases involved in MPR 300 phosphorylation and the phosphorylat......The human 300 kDa mannose-6-phosphate receptor (MPR 300) is phosphorylated in vivo at serine residues of its cytoplasmic domain. Two-dimensional separation can resolve tryptic phosphopeptides into four major species. To identify the kinases involved in MPR 300 phosphorylation...... and the phosphorylation sites the entire coding sequence of the cytoplasmic tail was expressed in Escherichia coli. The isolated cytoplasmic domain was used as a substrate for four purified serine/threonine kinases [casein kinase II (CK II), protein kinase A (PKA), protein kinase C and Ca2+/calmodulin kinase]. All...... kinases phosphorylate the cytoplasmic tail exclusively on serine residues. Inhibition studies using synthetic peptides, partial sequencing of isolated tryptic phosphopeptides and co-migration with tryptic phosphopeptides from MPR 300 labelled in vivo showed that (i) PKA phosphorylates the cytoplasmic MPR...

  15. Central Diabetes Insipidus in Refractory Antineutrophil Cytoplasmic Antibody-associated Vasculitis.

    Science.gov (United States)

    Ohashi, Keiji; Morishita, Michiko; Watanabe, Haruki; Sada, Ken-Ei; Katsuyama, Takayuki; Miyawaki, Yoshia; Katsuyama, Eri; Narazaki, Mariko; Tatebe, Noriko; Watanabe, Katsue; Kawabata, Tomoko; Wada, Jun

    2017-11-01

    We herein describe two cases of refractory antineutrophil cytoplasmic antibody-associated vasculitis (AAV) complicated with diabetes insipidus (DI) possibly related to hypertrophic pachymeningitis (HP). One patient had microscopic polyangiitis and HP, which were refractory to cyclophosphamide, azathioprine, rituximab, mycophenolate mofetil (MMF), and mizoribine. Remission was finally achieved with the use of etanercept, but DI occurred 5 years later. The other patient had granulomatosis with polyangiitis, which that was refractory to cyclophosphamide, methotrexate, MMF, and rituximab. DI subsequently developed, but was successfully treated with etanercept. Dura mater hypertrophy was macroscopically observed in the latter case.

  16. Pattern formation by interaction of three cytoplasmic factors in the egg of the leafhopper Euscelis plebejus.

    Science.gov (United States)

    Vogel, O

    1983-09-01

    Developmental capacities of different parts of the Euscelis plebejus egg were tested by translocating posterior pole material and subsequent transverse constriction of the egg posterior to the translocated material. The results support the hypothesis that at least three cytoplasmic factors of maternal origin are necessary to form a complete germ band. Those factors do not act autonomously. The anterior and posterior factors require interaction with the middle factor in order to cause formation of head and abdomen, respectively. The middle factor, on the other hand, forms a complete thorax only if it is in contact with the posterior factor.

  17. Isolation of cell nuclei using inert macromolecules to mimic the crowded cytoplasm.

    Directory of Open Access Journals (Sweden)

    Ronald Hancock

    Full Text Available Cell nuclei are commonly isolated and studied in media which include millimolar concentrations of cations, which conserve the nuclear volume by screening the negative charges on chromatin and maintaining its compaction. However, two factors question if these ionic conditions correctly reproduce the environment of nuclei in vivo: the small-scale motion and conformation of chromatin in vivo are not reproduced in isolated nuclei, and experiments and theory suggest that small ions in the cytoplasm are not free in the soluble phase but are predominantly bound to macromolecules. We studied the possible role in maintaining the structure and functions of nuclei in vivo of a further but frequently overlooked property of the cytoplasm, the crowding or osmotic effects caused by diffusible macromolecules whose concentration, measured in several studies, is in the range of 130 mg/ml. Nuclei which conserved their volume in the cell and their ultrastructure seen by electron microscopy were released from K562 cells in media containing the inert polymer 70 kDa Ficoll (50% w/v or 70 kDa dextran (35% w/v to replace the diffusible cytoplasmic molecules which were dispersed on cell lysis with digitonin, with 100 microM K-Hepes buffer as the only source of ions. Immunofluorescence labelling and experiments using cells expressing GFP-fusion proteins showed that internal compartments (nucleoli, PML and coiled bodies, foci of RNA polymerase II were conserved in these nuclei, and nascent RNA transcripts could be elongated. Our observations are consistent with the hypothesis that crowding by diffusible cytoplasmic macromolecules is a crucial but overlooked factor which supports the nucleus in vivo by equilibrating the opposing osmotic pressure cause by the high concentration of macromolecules in the nucleus, and suggest that crowded media provide more physiological conditions to study nuclear structure and functions. They may also help to resolve the long-standing paradox

  18. Prospective study of radioimmunoassay for antibodies against neutrophil cytoplasm in diagnosis of systemic vasculitis

    Energy Technology Data Exchange (ETDEWEB)

    Savage, C.O.S.; Winearls, C.G.; Jones, S.; Marshall, P.D.; Lockwood, C.M.

    1987-06-20

    The diagnosis and management of Wegener's granulomatosis and microscopic polyarteritis are complicated by the lack of specific diagnostic tests. The diagnostic performance of a solid-phase radioimmunoassay, which detects the autoantibodies against neutrophil cytoplasm present in these disorders, was assessed in a prospective study of patients with suspected vasculitis and/or rapidly progressive nephritis. The assay had a sensitivity and specificity of 96% when carried out in combination with a specific inhibition stage and indirect immunofluorescence staining of alcohol-fixed normal neutrophils.

  19. Prospective study of radioimmunoassay for antibodies against neutrophil cytoplasm in diagnosis of systemic vasculitis

    International Nuclear Information System (INIS)

    Savage, C.O.S.; Winearls, C.G.; Jones, S.; Marshall, P.D.; Lockwood, C.M.

    1987-01-01

    The diagnosis and management of Wegener's granulomatosis and microscopic polyarteritis are complicated by the lack of specific diagnostic tests. The diagnostic performance of a solid-phase radioimmunoassay, which detects the autoantibodies against neutrophil cytoplasm present in these disorders, was assessed in a prospective study of patients with suspected vasculitis and/or rapidly progressive nephritis. The assay had a sensitivity and specificity of 96% when carried out in combination with a specific inhibition stage and indirect immunofluorescence staining of alcohol-fixed normal neutrophils. (author)

  20. p25alpha relocalizes in oligodendroglia from myelin to cytoplasmic inclusions in multiple system atrophy

    DEFF Research Database (Denmark)

    Song, Yun Ju C; Lundvig, Ditte M S; Huang, Yue

    2007-01-01

    immunohistochemistry revealed a cellular redistribution of p25alpha immunoreactivity from the myelin to the oligodendroglial cell soma, with no overall change in p25alpha protein concentration using immunoblotting. Concomitantly, an approximately 80% reduction in the concentration of full-length MBP protein...... cytoplasmic inclusions. Overall, the data indicate that changes in the cellular interactions between MBP and p25alpha occur early in MSA and contribute to abnormalities in myelin and subsequent alpha-synuclein aggregation and the ensuing neuronal degeneration that characterizes this disease....

  1. Multimedia content classification metrics for content adaptation

    OpenAIRE

    Fernandes, Rui; Andrade, M.T.

    2015-01-01

    Multimedia content consumption is very popular nowadays. However, not every content can be consumed in its original format: the combination of content, transport and access networks, consumption device and usage environment characteristics may all pose restrictions to that purpose. One way to provide the best possible quality to the user is to adapt the content according to these restrictions as well as user preferences. This adaptation stage can be best executed if knowledge about the conten...

  2. Multimedia content classification metrics for content adaptation

    OpenAIRE

    Fernandes, Rui; Andrade, M.T.

    2016-01-01

    Multimedia content consumption is very popular nowadays. However, not every content can be consumed in its original format: the combination of content, transport and access networks, consumption device and usage environment characteristics may all pose restrictions to that purpose. One way to provide the best possible quality to the user is to adapt the content according to these restrictions as well as user preferences. This adaptation stage can be best executed if knowledge about the conten...

  3. Preventing Addiction.

    Science.gov (United States)

    Moore, Susan Fordney

    The purpose of this paper is to provide the beginning counselor with an overview of prevention concepts. Prevention is a relatively new emphasis in community efforts to stem the rising costs of substance abuse and other high-risk behaviors. The paper discusses agent, host, and environmental prevention models and how they relate to causal theories…

  4. The presequence of Euglena LHCPII, a cytoplasmically synthesized chloroplast protein, contains a functional endoplasmic reticulum-targeting domain.

    Science.gov (United States)

    Kishore, R; Muchhal, U S; Schwartzbach, S D

    1993-01-01

    The precursor to the Euglena light-harvesting chlorophyll a/b-binding protein of photosystem II (pLHCPII) is unique; it is a polyprotein, synthesized on membrane-bound ribosomes and transported to the Golgi apparatus prior to chloroplast localization. A cDNA corresponding to the 5' end of LHCPII mRNA has been isolated and sequenced. The deduced amino acid sequence of this cDNA indicates that Euglena pLHCPII contains a 141-amino acid N-terminal extension. The N-terminal extension contains three hydrophobic domains and a potential signal peptidase cleavage site at amino acid 35. Cotranslational processing by canine microsomes removed approximately 35 amino acids from an in vitro synthesized 33-kDa pLHCPII composed of a 141-amino acid N-terminal extension and a 180-amino acid partial LHCPII unit truncated at the beginning of the third membrane-spanning hydrophobic domain. Processed pLHCPII was degraded by exogenous protease, indicating that it had not been translocated to the microsomal lumen. Extraction with 0.1 M Na2CO3, pH 11.5, did not remove the processed pLHCPII from the microsomal membrane. A stop-transfer membrane anchor sequence appears to anchor the nascent protein within the membrane, preventing translocation into the lumen. Taken together, these results provide biochemical evidence for a functional cleaved signal sequence within the N-terminal extension of a Euglena cytoplasmically synthesized chloroplast-localized protein. Images Fig. 2 Fig. 3 PMID:8265635

  5. Whole Mitochondrial Genome Sequencing and Re-Examination of a Cytoplasmic Male Sterility-Associated Gene in Boro-Taichung-Type Cytoplasmic Male Sterile Rice.

    Science.gov (United States)

    Kazama, Tomohiko; Toriyama, Kinya

    2016-01-01

    Nuclear genome substitutions between subspecies can lead to cytoplasmic male sterility (CMS) through incompatibility between nuclear and mitochondrial genomes. Boro-Taichung (BT)-type CMS rice was obtained by substituting the nuclear genome of Oryza sativa subsp. indica cultivar Chinsurah Boro II with that of Oryza sativa subsp. japonica cultivar Taichung 65. In BT-type CMS rice, the mitochondrial gene orf79 is associated with male sterility. A complete sequence of the Boro-type mitochondrial genome responsible for BT-type CMS has not been determined to date. Here, we used pyrosequencing to construct the Boro-type mitochondrial genome. The contiguous sequences were assembled into five circular DNA molecules, four of which could be connected into a single circle. The two resulting subgenomic circles were unable to form a reliable master circle, as recombination between them was scarcely detected. We also found an unequal abundance of DNA molecules for the two loci of atp6. These results indicate the presence of multi-partite DNA molecules in the Boro-type mitochondrial genome. Expression patterns were investigated for Boro-type mitochondria-specific orfs, which were not found in the mitochondria from the standard japonica cultivar Nipponbare. Restorer of fertility 1 (RF1)-dependent RNA processing has been observed in orf79-containing RNA but was not detected in other Boro-type mitochondria-specific orfs, supporting the conclusion that orf79 is a unique CMS-associated gene in Boro-type mitochondria.

  6. Scallop protein with endogenous high taurine and glycine content prevents high-fat, high-sucrose-induced obesity and improves plasma lipid profile in male C57BL/6J mice

    DEFF Research Database (Denmark)

    Tastesen, Hanne Sørup; Keenan, Alison H.; Madsen, Lise

    2014-01-01

    -sucrose diets with protein sources of increasing endogenous taurine content, i.e., chicken, cod, crab and scallop, for 6 weeks. The energy intake was lower in crab and scallop-fed mice than in chicken and cod-fed mice, but only scallop-fed mice gained less body and fat mass. Liver mass was reduced in scallop......-fasted states. Dietary intake of taurine and glycine correlated negatively with body mass gain and total fat mass, while intake of all other amino acids correlated positively. Furthermore taurine and glycine intake correlated positively with improved plasma lipid profile, i.e., lower levels of plasma lipids...

  7. Ultrasound-dependent cytoplasmic internalization of a peptide-sonosensitizer conjugate.

    Science.gov (United States)

    Inaba, Yuki; Watanabe, Kazunori; Kitamatsu, Mizuki; Nakata, Eiji; Harada, Atsushi; Ohtsuki, Takashi

    2017-08-01

    A method to induce cytoplasmic peptide delivery, using ultrasound, was demonstrated using a molecular conjugate of a cell-penetrating peptide (CPP), a functional peptide, and a sonosensitizer. As a model of such molecular conjugates, TatBim-RB, consisting of the Tat CPP, the Bim apoptosis inducing peptide, and the sonosensitizer rose bengal was synthesized. CPPs have been widely used for intracellular delivery of various cargos; however, CPP-fused molecules tend to become entrapped in endosomes, as was observed for TatBim-RB molecules applied to cells. To promote escape of the entrapped TatBim-RB molecules, cells were irradiated with ultrasound, which successfully induced endosomal escape and cytoplasmic dispersion of TatBim-RB, and subsequently apoptosis. Our results suggest that this peptide-sonosensitizer conjugate strategy may facilitate numerous kinds of medicinal chemistry studies, and furthermore, this specific conjugate may exhibit potential as a novel therapeutic agent for the promotion of apoptosis. Copyright © 2017 Elsevier Ltd. All rights reserved.

  8. Cytoplasmic Dynein Transports Axonal Microtubules in a Polarity-Sorting Manner

    Directory of Open Access Journals (Sweden)

    Anand N. Rao

    2017-06-01

    Full Text Available Axonal microtubules are predominantly organized into a plus-end-out pattern. Here, we tested both experimentally and with computational modeling whether a motor-based polarity-sorting mechanism can explain this microtubule pattern. The posited mechanism centers on cytoplasmic dynein transporting plus-end-out and minus-end-out microtubules into and out of the axon, respectively. When cytoplasmic dynein was acutely inhibited, the bi-directional transport of microtubules in the axon was disrupted in both directions, after which minus-end-out microtubules accumulated in the axon over time. Computational modeling revealed that dynein-mediated transport of microtubules can establish and preserve a predominantly plus-end-out microtubule pattern as per the details of the experimental findings, but only if a kinesin motor and a static cross-linker protein are also at play. Consistent with the predictions of the model, partial depletion of TRIM46, a protein that cross-links axonal microtubules in a manner that influences their polarity orientation, leads to an increase in microtubule transport.

  9. Internal epithelia in Drosophila display rudimentary competence to form cytoplasmic networks of transgenic human vimentin.

    Science.gov (United States)

    Gullmets, Josef; Torvaldson, Elin; Lindqvist, Julia; Imanishi, Susumu Y; Taimen, Pekka; Meinander, Annika; Eriksson, John E

    2017-12-01

    Cytoplasmic intermediate filaments (cIFs) are found in all eumetazoans, except arthropods. To investigate the compatibility of cIFs in arthropods, we expressed human vimentin (hVim), a cIF with filament-forming capacity in vertebrate cells and tissues, transgenically in Drosophila Transgenic hVim could be recovered from whole-fly lysates by using a standard procedure for intermediate filament (IF) extraction. When this procedure was used to test for the possible presence of IF-like proteins in flies, only lamins and tropomyosin were observed in IF-enriched extracts, thereby providing biochemical reinforcement to the paradigm that arthropods lack cIFs. In Drosophila , transgenic hVim was unable to form filament networks in S2 cells and mesenchymal tissues; however, cage-like vimentin structures could be observed around the nuclei in internal epithelia, which suggests that Drosophila retains selective competence for filament formation. Taken together, our results imply that although the filament network formation competence is partially lost in Drosophila , a rudimentary filament network formation ability remains in epithelial cells. As a result of the observed selective competence for cIF assembly in Drosophila , we hypothesize that internal epithelial cIFs were the last cIFs to disappear from arthropods.-Gullmets, J., Torvaldson, E., Lindqvist, J., Imanishi, S. Y., Taimen, P., Meinander, A., Eriksson, J. E. Internal epithelia in Drosophila display rudimentary competence to form cytoplasmic networks of transgenic human vimentin. © FASEB.

  10. Association of nad7a Gene with Cytoplasmic Male Sterility in Pigeonpea

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    Pallavi Sinha

    2015-07-01

    Full Text Available Cytoplasmic male sterility (CMS has been exploited in the commercial pigeonpea [ (L. Millsp.] hybrid breeding system; however, the molecular mechanism behind this system is unknown. To understand the underlying molecular mechanism involved in A CMS system derived from (Haines Maesen, 34 mitochondrial genes were analyzed for expression profiling and structural variation analysis between CMS line (ICRISAT Pigeonpea A line, ICPA 2039 and its cognate maintainer (ICPB 2039. Expression profiling of 34 mitochondrial genes revealed nine genes with significant fold differential gene expression at ≤ 0.01, including one gene, , with 1366-fold higher expression in CMS line as compared with the maintainer. Structural variation analysis of these mitochondrial genes identified length variation between ICPA 2039 and ICPB 2039 for (subunit of gene. Sanger sequencing of and genes in the CMS and the maintainer lines identified two single nucleotide polymorphisms (SNPs in upstream region of and a deletion of 10 bp in in the CMS line. Protein structure evaluation showed conformational changes in predicted protein structures for between ICPA 2039 and ICPB 2039 lines. All above analyses indicate association of gene with the CMS for A cytoplasm in pigeonpea. Additionally, one polymerase chain reaction (PCR based Indel marker ( has been developed and validated for testing genetic purity of A derived CMS lines to strengthen the commercial hybrid breeding program in pigeonpea.

  11. PKCepsilon stimulated arginine methylation of RIP140 for its nuclear-cytoplasmic export in adipocyte differentiation.

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    Pawan Gupta

    2008-07-01

    Full Text Available Receptor interacting protein 140 (RIP140 is a versatile transcriptional co-repressor that plays roles in diverse metabolic processes including fat accumulation in adipocytes. Previously we identified three methylated arginine residues in RIP140, which rendered its export to the cytoplasm; but it was unclear what triggered RIP140 arginine methylation.In this study, we determined the activated PKCepsilon as the specific trigger for RIP140 arginine methylation and its subsequent export. We identified two PKCepsilon-phosphorylated residues of RIP140, Ser-102 and Ser-1003, which synergistically stimulated direct binding of RIP140 by 14-3-3 that recruited protein arginine methyl transferase 1 to methylate RIP140. The methylated RIP140 then preferentially recruited exportin 1 for nuclear export. As a result, the nuclear gene-repressive activity of RIP140 was reduced. In RIP140 null adipocyte cultures, the defect in fat accumulation was effectively rescued by the phosphorylation-deficient mutant RIP140 that resided predominantly in the nucleus, but less so by the phospho-mimetic RIP140 that was exported to the cytoplasm.This study uncovers a novel means, via a cascade of protein modifications, to inactivate, or suppress, the nuclear action of an important transcription coregulator RIP140, and delineates the first specific phosphorylation-arginine methylation cascade that could alter protein subcellular distribution and biological activity.

  12. Cucumber Pti1-L is a cytoplasmic protein kinase involved in defense responses and salt tolerance.

    Science.gov (United States)

    Oh, Sang-Keun; Jang, Hyun A; Lee, Sang Sook; Cho, Hye Sun; Lee, Dong-Hee; Choi, Doil; Kwon, Suk-Yoon

    2014-06-15

    Homologs of the cytoplasmic protein kinase Pti1 are found in diverse plant species. A clear role of Pti1 in plant defense response has not been established. We identified a Pti1 homolog in cucumber (CsPti1-L). CsPti1-L expression was induced when cucumber plants were challenged with the fungal pathogen Sphaerotheca fuliginea or with salt treatment. CsPti1-L expression in cucumber leaves also was induced by methyl jasmonate, salicylic acid, and abscisic acid. CsPti1-L exhibited autophosphorylation activity and was targeted to the cytoplasm. Transgenic Nicotiana benthamiana expressing CsPti1-L exhibited greater cell death and increased ion leakage in response to the bacterial pathogen Pseudomonas syringae pv. tomato DC3000, resistance to Botrytis cinerea infection, and higher tolerance to salt stress. RT-PCR analysis of transgenic N. benthamiana overexpressing CsPti1-L revealed constitutive upregulation of multiple genes involved in plant-defense and osmotic-stress responses. Our results suggest a functional role for CsPti1-L as a positive regulator of pathogen-defense and salt-stress responses. Copyright © 2014 Elsevier GmbH. All rights reserved.

  13. Uptake and degradation of cytoplasmic RNA by lysosomes in the perfused rat liver

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    Heydrick, S.J.; Lardeux, B.; Mortimore, G.E.

    1987-05-01

    The release of (/sup 14/C)cytidine has been shown previously to be a valid marker for RNA degradation in rat hepatocytes. The breakdown of RNA measured with this marker in perfused livers prelabeled in vivo with (6-/sup 14/C)orotic acid was found to be regulated acutely by perfusate amino acids over a wide range, from 0.29 to 3.48%/h. This regulation paralleled that of lysosomal proteolysis. Chloroquine inhibited RNA degradation 60-70%. In subsequent cell fractionation studies labelled cytidine was released; the distribution of this release paralleled that of a lysosomal marker enzyme. The release plateaued after two hours, defining a distinct lysosomal pool of RNA. The lysosomal location of the RNA pool was confirmed in experiments where a 22% increase in the apparent pool size was obtained by lowering the homogenate pH from 7.0 to 5.5. The pool size correlated linearly with the rate of RNA degradation measured during perfusion, giving a turnover constant in reasonable agreement with values reported for autophagy. These results indicate that cytoplasmic RNA degradation occurs primarily in the lysosome and is regulated under these conditions by the amino acid control of lysosomal sequestration of cytoplasm.

  14. [Anti-neutrophil cytoplasmic antibodies (ANCA) in patients with symptomatic and asymptomatic HIV infection].

    Science.gov (United States)

    Habegger de Sorrentino, A; Motta, P; Iliovich, E; Sorrentino, A P

    1997-01-01

    The cytopathic effect of HIV on CD4 T cells, as well as the active autoimmune mechanism occurring during infection, have been documented. Of the cytokines involved in the pathogenesis of AIDS, the main one produced by the monocyte-macrophage series is tumor necrosis factor alfa (TNF alpha). This cytokine induces antigens such as proteinase 3 (Pr 3) or mieloperoxidase (MPO). Anti-neutrophil cytoplasmic antibodies (ANCA) are directed against this type of PMN antigens. In the present paper, the role of anti-neutrophil cytoplasmic antibodies (ANCA) in HIV infected patients as responsible for autoimmune phenomena in relation to opportunistic infections, was studied. A total of 88 serum samples belonging to 49 asymptomatic and 39 symptomatic HIV infected patients were tested for ANCA by an indirect immunofluorescence (IIF) test over a neutrophil substrate. ANCA were detected in 53.8% of symptomatic patients as compared to 4.1% in asymptomatic cases (p tuberculosis is a frequent finding in HIV infected patients from Northeastern Argentina. When the presence of ANCA in TB(+) HIV(+) and TB(+) HIV(-) patients was studied, it was seen that positive-ANCA significantly correlated with the first group (p pulmonar TB, could indicate that the virus may not be responsible for the induction of these antibodies.

  15. Cytoplasmic inheritance of parent-offspring cell structure in the clonal diatom Cyclotella meneghiniana.

    Science.gov (United States)

    Shirokawa, Yuka; Shimada, Masakazu

    2016-11-16

    In cytoplasmic inheritance, structural states of a parent cell could be transmitted to offspring cells via two mechanisms. The first is referred to as the hangover of parent structure, where the structure itself remains and faithfully transmits within offspring cells; the second is structural inheritance, wherein the parent structure functions as a template for development of new offspring structure. We estimated to what extent the parent structure affects the development of offspring structure by structural inheritance, using a clone of the diatom Cyclotella meneghiniana The cell has two siliceous valves (a cell wall part at both cell poles): one is inherited from the parent and the other is newly formed. We estimated cytoplasmic heritability by comparing valve traits (central fultoportulae (CTFP), striae, central area, and cell diameter) of parent and new offspring valves, using single-cell isolation and valve labelling. Parent-offspring valve trait regressions showed that all traits, except CTFP, were significantly correlated. We formulated a quantitative genetic model considering the diatom inheritance system and revealed short-term rapid evolution compared with other inheritance systems. Diatom structural inheritance will have evolved to enable clonal populations to rapidly acquire and maintain suitable structures for temporal changes in environments and life-cycle stages. © 2016 The Author(s).

  16. Transcriptional Regulation of Culex pipiens Mosquitoes by Wolbachia Influences Cytoplasmic Incompatibility

    Science.gov (United States)

    Harris, Simon; Kambris, Zakaria; Sutton, Elizabeth R.; Bonsall, Michael B.; Parkhill, Julian; Sinkins, Steven P.

    2013-01-01

    Cytoplasmic incompatibility (CI) induced by the endosymbiont Wolbachia pipientis causes complex patterns of crossing sterility between populations of the Culex pipiens group of mosquitoes. The molecular basis of the phenotype is yet to be defined. In order to investigate what host changes may underlie CI at the molecular level, we examined the transcription of a homolog of the Drosophila melanogaster gene grauzone that encodes a zinc finger protein and acts as a regulator of female meiosis, in which mutations can cause sterility. Upregulation was observed in Wolbachia-infected C. pipiens group individuals relative to Wolbachia-cured lines and the level of upregulation differed between lines that were reproductively incompatible. Knockdown analysis of this gene using RNAi showed an effect on hatch rates in a Wolbachia infected Culex molestus line. Furthermore, in later stages of development an effect on developmental progression in CI embryos occurs in bidirectionally incompatible crosses. The genome of a wPip Wolbachia strain variant from Culex molestus was sequenced and compared with the genome of a wPip variant with which it was incompatible. Three genes in inserted or deleted regions were newly identified in the C. molestus wPip genome, one of which is a transcriptional regulator labelled wtrM. When this gene was transfected into adult Culex mosquitoes, upregulation of the grauzone homolog was observed. These data suggest that Wolbachia-mediated regulation of host gene expression is a component of the mechanism of cytoplasmic incompatibility. PMID:24204251

  17. Short Arginine Motifs Drive Protein Stickiness in the Escherichia coli Cytoplasm.

    Science.gov (United States)

    Kyne, Ciara; Crowley, Peter B

    2017-09-19

    Although essential to numerous biotech applications, knowledge of molecular recognition by arginine-rich motifs in live cells remains limited. 1 H, 15 N HSQC and 19 F NMR spectroscopies were used to investigate the effects of C-terminal -GR n (n = 1-5) motifs on GB1 interactions in Escherichia coli cells and cell extracts. While the "biologically inert" GB1 yields high-quality in-cell spectra, the -GR n fusions with n = 4 or 5 were undetectable. This result suggests that a tetra-arginine motif is sufficient to drive interactions between a test protein and macromolecules in the E. coli cytoplasm. The inclusion of a 12 residue flexible linker between GB1 and the -GR 5 motif did not improve detection of the "inert" domain. In contrast, all of the constructs were detectable in cell lysates and extracts, suggesting that the arginine-mediated complexes were weak. Together these data reveal the significance of weak interactions between short arginine-rich motifs and the E. coli cytoplasm and demonstrate the potential of such motifs to modify protein interactions in living cells. These interactions must be considered in the design of (in vivo) nanoscale assemblies that rely on arginine-rich sequences.

  18. Characterization of the injection funnel during intracytoplasmic sperm injection reflects cytoplasmic maturity of the oocyte.

    Science.gov (United States)

    Krause, Iris; Pohler, Uwe; Grosse, Stefan; Shebl, Omar; Petek, Erwin; Chandra, Arvind; Ebner, Thomas

    2016-10-01

    To quantify cytoplasmic maturity on the basis of intracytoplasmic sperm injection (ICSI) injection funnel manifestation and to evaluate influence factors of the latter. Prospective study. Private fertility center. A total of 31 patients with good ovarian response. Mature and immature oocytes were injected intracytoplasmatically. Formation and persistence of an injection funnel was documented and measured. ICSI funnel size, persistence of injection funnel, rates of degeneration and fertilization, embryo quality. Funnel volume in germinal vesicle stage oocytes (prophase I [PI]) was significantly smaller than that of metaphase I (MI) and MII oocytes. Immature eggs (PI, MI) almost never showed a persistent funnel 2-4 minutes after ICSI, whereas in MII eggs the funnel was still observable in 35% (117/334) of the cases. Uni- and multivariate analysis revealed that pipette type and stimulation protocol significantly influenced appearance of injection funnel. Funnel volume in oocytes that fertilized regularly was significantly higher compared with three-polar body and degenerated oocytes. Oocyte maturation within the follicle is closely associated with a remarkable change in cytoplasm viscosity from an aqueous to a more viscous subtype. Precise evaluation of the injection funnel may help to explain deviations from expected ICSI outcome and could also assist in optimizing controlled ovarian hyperstimulation. Copyright © 2016 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  19. Gas6 downregulation impaired cytoplasmic maturation and pronuclear formation independent to the MPF activity.

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    Kyeoung-Hwa Kim

    Full Text Available Previously, we found that the growth arrest-specific gene 6 (Gas6 is more highly expressed in germinal vesicle (GV oocytes than in metaphase II (MII oocytes using annealing control primer (ACP-PCR technology. The current study was undertaken to investigate the role of Gas6 in oocyte maturation and fertilization using RNA interference (RNAi. Interestingly, despite the specific and marked decrease in Gas6 mRNA and protein expression in GVs after Gas6 RNAi, nuclear maturation including spindle structures and chromosome segregation was not affected. The only discernible effect induced by Gas6 RNAi was a change in maturation promoting factor (MPF activity. After parthenogenetic activation, Gas6 RNAi-treated oocytes at the MII stage had not developed further and arrested at MII (90.0%. After stimulation with Sr(2+, Gas6-silenced MII oocytes had markedly reduced Ca(2+ oscillation and exhibited no exocytosis of cortical granules. In these oocytes, sperm penetration occurred during fertilization but not pronucleus (PN formation. By roscovitine and colcemid treatment, we found that the Gas6 knockdown affected cytoplasmic maturation directly, independent to the changed MPF activity. These results strongly suggest that 1 the Gas6 signaling itself is important to the cytoplasmic maturation, but not nuclear maturation, and 2 the decreased Gas6 expression and decreased MPF activity separately or mutually influence sperm head decondensation and PN formation.

  20. Behavioral and other phenotypes in a cytoplasmic Dynein light intermediate chain 1 mutant mouse.

    Science.gov (United States)

    Banks, Gareth T; Haas, Matilda A; Line, Samantha; Shepherd, Hazel L; Alqatari, Mona; Stewart, Sammy; Rishal, Ida; Philpott, Amelia; Kalmar, Bernadett; Kuta, Anna; Groves, Michael; Parkinson, Nicholas; Acevedo-Arozena, Abraham; Brandner, Sebastian; Bannerman, David; Greensmith, Linda; Hafezparast, Majid; Koltzenburg, Martin; Deacon, Robert; Fainzilber, Mike; Fisher, Elizabeth M C

    2011-04-06

    The cytoplasmic dynein complex is fundamentally important to all eukaryotic cells for transporting a variety of essential cargoes along microtubules within the cell. This complex also plays more specialized roles in neurons. The complex consists of 11 types of protein that interact with each other and with external adaptors, regulators and cargoes. Despite the importance of the cytoplasmic dynein complex, we know comparatively little of the roles of each component protein, and in mammals few mutants exist that allow us to explore the effects of defects in dynein-controlled processes in the context of the whole organism. Here we have taken a genotype-driven approach in mouse (Mus musculus) to analyze the role of one subunit, the dynein light intermediate chain 1 (Dync1li1). We find that, surprisingly, an N235Y point mutation in this protein results in altered neuronal development, as shown from in vivo studies in the developing cortex, and analyses of electrophysiological function. Moreover, mutant mice display increased anxiety, thus linking dynein functions to a behavioral phenotype in mammals for the first time. These results demonstrate the important role that dynein-controlled processes play in the correct development and function of the mammalian nervous system.

  1. Potential role for MATER in cytoplasmic lattice formation in murine oocytes.

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    Boram Kim

    2010-09-01

    Full Text Available Mater and Padi6 are maternal effect genes that are first expressed during oocyte growth and are required for embryonic development beyond the two-cell stage in the mouse. We have recently found that PADI6 localizes to, and is required for the formation of, abundant fibrillar Triton X-100 (Triton insoluble structures termed the oocyte cytoplasmic lattices (CPLs. Given their similar expression profiles and mutant mouse phenotypes, we have been testing the hypothesis that MATER also plays a role in CPL formation and/or function.Herein, we show that PADI6 and MATER co-localize throughout the oocyte cytoplasm following Triton extraction, suggesting that MATER co-localizes with PADI6 at the CPLs. Additionally, the solubility of PADI6 was dramatically increased in Mater(tm/tm oocytes following Triton extraction, suggesting that MATER is involved in CPL nucleation. This prediction is supported by transmission electron microscopic analysis of Mater(+/+ and Mater(tm/tm germinal vesicle stage oocytes which illustrated that volume fraction of CPLs was reduced by 90% in Mater(tm/tm oocytes compared to Mater(+/+ oocytes.Taken together, these results suggest that, similar to PADI6, MATER is also required for CPL formation. Given that PADI6 and MATER are essential for female fertility, these results not only strengthen the hypothesis that the lattices play a critical role in mediating events during the oocyte-to-embryo transition but also increase our understanding of the molecular nature of the CPLs.

  2. Formation, function, and exhaustion of notochordal cytoplasmic vacuoles within intervertebral disc: current understanding and speculation.

    Science.gov (United States)

    Wang, Feng; Gao, Zeng-Xin; Cai, Feng; Sinkemani, Arjun; Xie, Zhi-Yang; Shi, Rui; Wei, Ji-Nan; Wu, Xiao-Tao

    2017-08-22

    Notochord nucleus pulposus cells are characteristic of containing abundant and giant cytoplasmic vacuoles. This review explores the embryonic formation, biological function, and postnatal exhaustion of notochord vacuoles, aiming to characterize the signal network transforming the vacuolated nucleus pulposus cells into the vacuole-less chondrocytic cells. Embryonically, the cytoplasmic vacuoles within vertebrate notochord originate from an evolutionarily conserved vacuolation process during neurulation, which may continue to provide mechanical and signal support in constructing a mammalian intervertebral disc. For full vacuolation, a vacuolating specification from dorsal organizer cells, synchronized convergent extension, well-structured notochord sheath, and sufficient post-Golgi trafficking in notochord cells are required. Postnatally, age-related and species-specific exhaustion of vacuolated nucleus pulposus cells could be potentiated by Fas- and Fas ligand-induced apoptosis, intolerance to mechanical stress and nutrient deficiency, vacuole-mediated proliferation check, and gradual de-vacuolation within the avascular and compression-loaded intervertebral disc. These results suggest that the notochord vacuoles are active and versatile organelles for both embryonic notochord and postnatal nucleus pulposus, and may provide novel information on intervertebral disc degeneration to guide cell-based regeneration.

  3. Positive Selection Driving Cytoplasmic Genome Evolution of the Medicinally Important Ginseng Plant Genus Panax

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    Peng Jiang

    2018-04-01

    Full Text Available Panax L. (the ginseng genus is a shade-demanding group within the family Araliaceae and all of its species are of crucial significance in traditional Chinese medicine. Phylogenetic and biogeographic analyses demonstrated that two rounds of whole genome duplications accompanying with geographic and ecological isolations promoted the diversification of Panax species. However, contributions of the cytoplasmic genomes to the adaptive evolution of Panax species remained largely uninvestigated. In this study, we sequenced the chloroplast and mitochondrial genomes of 11 accessions belonging to seven Panax species. Our results show that heterogeneity in nucleotide substitution rate is abundant in both of the two cytoplasmic genomes, with the mitochondrial genome possessing more variants at the total level but the chloroplast showing higher sequence polymorphisms at the genic regions. Genome-wide scanning of positive selection identified five and 12 genes from the chloroplast and mitochondrial genomes, respectively. Functional analyses further revealed that these selected genes play important roles in plant development, cellular metabolism and adaptation. We therefore conclude that positive selection might be one of the potential evolutionary forces that shaped nucleotide variation pattern of these Panax species. In particular, the mitochondrial genes evolved under stronger selective pressure compared to the chloroplast genes.

  4. Role of cytoplasm-specific introgression in the evolution of the polyploid wheats

    Science.gov (United States)

    Gill, Bikram S.; Chen, P. D.

    1987-01-01

    Studies of N-banded mitotic and meiotic karyotypes of Triticum turgidum L. (2n = 28; AABB) and Triticum timopheevii Zhuk. (2n = 28; AAGG) and hybrids between them, along with observations of meiotic pairing between telocentrics of the AB-genome chromosomes and their respective homologues and homeologues in T. timopheevii, showed that chromosome 4 (m4) of Triticum monococcum L. is present (as 4At) in T. timopheevii but is lacking in T. turgidum. Neither 4A nor 4B pairs with 4At, but 4A pairs with 4G and, for this reason and because of its banding pattern, must be considered a B-genome chromosome. T. timopheevii chromosomes 4At and 3At are involved in a reciprocal translocation, and 2At, 1G, 2G, and 5G are also involved in translocations. Chromosome arm 4BL occasionally pairs with 7G. The satellites are on the short arms of chromosomes 6At and 6G of T. timopheevii and 1B and 6B of T. turgidum. It is suggested that (i) T. timopheevii orginated as an allotetraploid of Aegilops speltoides Tausch/T. monococcum and (ii) T. turgidum was derived from T. timopheevii by introgressive hybridization with an unknown diploid species, which contributed its distinctive cytoplasm, chromosome 4B or a substantial portion of it, and additional chromosome segments. Rapid fixation of 4B in T. turgidum was ensured by cytoplasm-specific transmission. Images PMID:16578821

  5. Holophytochrome-interacting proteins in Physcomitrella: putative actors in phytochrome cytoplasmic signaling

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    Anna Lena eErmert

    2016-05-01

    Full Text Available Phytochromes are the principle photoreceptors in light-regulated plant development, primarily acting via translocation of the light-activated photoreceptor into the nucleus and subsequent gene regulation. However, several independent lines of evidence indicate unambiguously that an additional cytoplasmic signaling mechanism must exist. Directional responses in filament tip cells of the moss Physcomitrella patens are steered by phy4 which has been shown to interact physically with the blue light receptor phototropin at the plasma membrane. This complex might perceive and transduce vectorial information leading to cytoskeleton reorganization and finally a directional growth response. We developed yeast two-hybrid procedures using photochemically-functional, full-length phy4 as bait in Physcomitrella cDNA library screens and growth assays under different light conditions, revealing Pfr-dependent interactions possibly associated with phytochrome cytoplasmic signaling. Candidate proteins were then expressed in planta with fluorescent protein tags to determine their intracellular localization in darkness and red light. Of 14 candidates, 12 were confirmed to interact with phy4 in planta using bimolecular fluorescence complementation. We discuss the roles these putative holophytochrome-interacting proteins (HIP's might have in signaling.

  6. Host cytoplasmic processing bodies assembled by Trypanosoma cruzi during infection exert anti-parasitic activity.

    Science.gov (United States)

    Seto, Eri; Onizuka, Yoko; Nakajima-Shimada, Junko

    2015-12-01

    Processing bodies (PBs) are cytoplasmic granules containing mRNAs and proteins involved in translation and degradation of mRNAs. PBs are constitutively present in cells and are induced to accumulate when external stressors including microbial infection are applied to cells, followed by a rapid translational arrest. We have examined the impact of Trypanosoma cruzi (T. cruzi, Tc) infection on host cytoplasmic PB assembly. Within 24h post-infection, we found the average number of PB foci per cell increased by more than 2-fold. Protein levels of PB components were unaltered during infection. These results indicated that Tc infection caused accumulation of PBs by changing the localization pattern of PB protein components. To elucidate the role of the accumulated PBs on Tc infection, we knocked down PBs using a siRNA specific for PB components EDC4 and Lsm14A, which are involved in mRNA decapping and translational repression, respectively. We observed that the inhibition of PB accumulation significantly enhanced the infectivity and growth of intracellular amastigotes. Depletion of PBs did not affect nitric oxide (NO) production during Tc infection, indicating that the growth promotion was not caused by modulation of NO-mediated killing of Tc. Our results suggest that the accumulated PBs partially contribute to anti-parasitic responses by manipulating the host's mRNA metabolism. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  7. The Mechanism by Which Luteolin Disrupts the Cytoplasmic Membrane of Methicillin-Resistant Staphylococcus aureus.

    Science.gov (United States)

    Zhang, Tao; Qiu, Yunguang; Luo, Qichao; Zhao, Lifen; Yan, Xin; Ding, Qiaoce; Jiang, Hualiang; Yang, Huaiyu

    2018-02-01

    Methicillin-resistant Staphylococcus aureus (MRSA) is one of the most versatile human pathogens. Luteolin (LUT) has anti-MRSA activity by disrupting the MRSA cytoplasmic membrane. However, the mechanism by which luteolin disrupts the membrane remains unclear. Here, we performed differential scanning calorimetry (DSC) and all-atomic molecular dynamics (AA-MD) simulations to investigate the interactions and effects of LUT on model membranes composed of phosphatidylcholine (PC) and phosphatidylglycerol (PG). We detected the transition thermodynamic parameters of dipalmitoylphosphatidylcholine (DPPC) liposomes, dipalmitoylphosphatidylglycerol (DPPG) liposomes, and liposomes composed of both DPPC and DPPG at different LUT concentrations and showed that LUT molecules were located between polar heads and the hydrophobic region of DPPC/DPPG. In the MD trajectories, LUT molecules ranging from 5 to 50 had different effects on the membranes thickness, fluidity and ordered property of lipids, and lateral pressure of lipid bilayers composed of dioleoylphosphatidylcholine (DOPC) and dioleoylphosphatidylglycerol (DOPG). Also, most LUT molecules were distributed in the region between the phosphorus atoms and C9 atoms of DOPC and DOPG. On the basis of the combination of these results, we conclude that the distinct effects of LUT on lipid bilayers composed of PCs and PGs may elucidate the mechanism by which LUT disrupts the cytoplasmic membrane of MRSA.

  8. Differential Proteomic Analysis of Anthers between Cytoplasmic Male Sterile and Maintainer Lines in Capsicum annuum L.

    Directory of Open Access Journals (Sweden)

    Zhiming Wu

    2013-11-01

    Full Text Available Cytoplasmic male sterility (CMS, widely used in the production of hybrid seeds, is a maternally inherited trait resulting in a failure to produce functional pollen. In order to identify some specific proteins associated with CMS in pepper, two-dimensional gel electrophoresis (2-DE was applied to proteomic analysis of anthers/buds between a CMS line (designated NA3 and its maintainer (designated NB3 in Capsicum annuum L. Thirty-three spots showed more than 1.5-fold in either CMS or its maintainer. Based on mass spectrometry, 27 spots representing 23 distinct proteins in these 33 spots were identified. Proteins down-regulated in CMS anthers/buds includes ATP synthase D chain, formate dehydrogenase, alpha-mannosidas, RuBisCO large subunit-binding protein subunit beta, chloroplast manganese stabilizing protein-II, glutathione S-transferase, adenosine kinase isoform 1T-like protein, putative DNA repair protein RAD23-4, putative caffeoyl-CoA 3-O-methyltransferase, glutamine synthetase (GS, annexin Cap32, glutelin, allene oxide cyclase, etc. In CMS anthers/buds, polyphenol oxidase, ATP synthase subunit beta, and actin are up-regulated. It was predicted that male sterility in NA3 might be related to energy metabolism turbulence, excessive ethylene synthesis, and suffocation of starch synthesis. The present study lays a foundation for future investigations of gene functions associated with pollen development and cytoplasmic male sterility, and explores the molecular mechanism of CMS in pepper.

  9. Structural changes in human cytomegalovirus cytoplasmic assembly sites in the absence of UL97 kinase activity

    International Nuclear Information System (INIS)

    Azzeh, Maysa; Honigman, Alik; Taraboulos, Albert; Rouvinski, Alexander; Wolf, Dana G.

    2006-01-01

    Studies of human cytomegalovirus (HCMV) UL97 kinase deletion mutant (ΔUL97) indicated a multi-step role for this kinase in early and late phases of the viral life cycle, namely, in DNA replication, capsid maturation and nuclear egress. Here, we addressed its possible involvement in cytoplasmic steps of HCMV assembly. Using the ΔUL97 and the UL97 kinase inhibitor NGIC-I, we demonstrate that the absence of UL97 kinase activity results in a modified subcellular distribution of the viral structural protein assembly sites, from compact structures impacting upon the nucleus to diffuse perinuclear structures punctuated by large vacuoles. Infection by either wild type or ΔUL97 viruses induced a profound reorganization of wheat germ agglutinin (WGA)-positive Golgi-related structures. Importantly, the viral-induced Golgi remodeling along with the reorganization of the nuclear architecture was substantially altered in the absence of UL97 kinase activity. These findings suggest that UL97 kinase activity might contribute to organization of the viral cytoplasmic assembly sites

  10. No influence of Indy on lifespan in Drosophila after correction for genetic and cytoplasmic background effects.

    Directory of Open Access Journals (Sweden)

    Janne M Toivonen

    2007-06-01

    Full Text Available To investigate whether alterations in mitochondrial metabolism affect longevity in Drosophila melanogaster, we studied lifespan in various single gene mutants, using inbred and outbred genetic backgrounds. As positive controls we included the two most intensively studied mutants of Indy, which encodes a Drosophila Krebs cycle intermediate transporter. It has been reported that flies heterozygous for these Indy mutations, which lie outside the coding region, show almost a doubling of lifespan. We report that only one of the two mutants lowers mRNA levels, implying that the lifespan extension observed is not attributable to the Indy mutations themselves. Moreover, neither Indy mutation extended lifespan in female flies in any genetic background tested. In the original genetic background, only the Indy mutation associated with altered RNA expression extended lifespan in male flies. However, this effect was abolished by backcrossing into standard outbred genetic backgrounds, and was associated with an unidentified locus on the X chromosome. The original Indy line with long-lived males is infected by the cytoplasmic symbiont Wolbachia, and the longevity of Indy males disappeared after tetracycline clearance of this endosymbiont. These findings underscore the critical importance of standardisation of genetic background and of cytoplasm in genetic studies of lifespan, and show that the lifespan extension previously claimed for Indy mutants was entirely attributable to confounding variation from these two sources. In addition, we saw no effects on lifespan of expression knockdown of the Indy orthologues nac-2 and nac-3 in the nematode Caenorhabditis elegans.

  11. Analysis of trafficking of Rev and transdominant Rev proteins in living cells using green fluorescent protein fusions: transdominant Rev blocks the export of Rev from the nucleus to the cytoplasm.

    Science.gov (United States)

    Stauber, R; Gaitanaris, G A; Pavlakis, G N

    1995-11-10

    Expression of gag/pol and env genes of human immunodeficiency virus requires the viral Rev protein. Mutant Rev proteins, displaying a transdominant phenotype (TDRev), were shown to inhibit Rev function. To investigate the underlying mechanism of this inhibition, the green fluorescent protein (GFP) of Aequorea victoria was fused to Rev and TDRev, which allowed the study of their trafficking and interactions in living human cells. Both Rev-GFP and TDRev-GFP were shown to retain appropriate nucleolar localization and function. Upon actinomycin D treatment, Rev-GFP was transported to the cytoplasm within 1.5 hr, while TDRev, although partially dissociated from the nucleolus, was retained in the nucleus. Coexpression of Rev-GFP and TDRev in the same cell demonstrated that TDRev inhibited the transport of Rev-GFP from the nucleus to the cytoplasm. This inhibition was specific for Rev, since the export of the functionally analogous Rex protein of the human T-cell leukemia virus type I was not inhibited by TDRev. These results indicate that Rev and TDRev form heteromultimers in the nucleolus and that this interaction prevents Rev's export from the nucleus to the cytoplasm. In addition to providing a model for the function of TDRev, these results also demonstrate the successful application of protein fusions to GFP to study localization and trafficking of proteins in living mammalian cells.

  12. Cytoplasmic CXCR4 expression in breast cancer: induction by nitric oxide and correlation with lymph node metastasis and poor prognosis

    Directory of Open Access Journals (Sweden)

    Kodama Rieko

    2008-11-01

    Full Text Available Abstract Background Lymph nodes constitute the first site of metastasis for most malignancies, and the extent of lymph node involvement is a major criterion for evaluating patient prognosis. The CXC chemokine receptor 4 (CXCR4 has been shown to play an important role in lymph node metastasis. Nitric oxide (NO may also contribute to induction of metastatic ability in human cancers. Methods CXCR4 expression was analyzed in primary human breast carcinoma with long-term follow-up. The relationship between nitrotyrosine levels (a biomarker for peroxynitrate formation from NO in vivo and lymph node status, CXCR4 immunoreactivity, and other established clinico-pathological parameters, as well as prognosis, was analyzed. Nitrite/nitrate levels and CXCR4 expressions were assessed in MDA-MB-231 and SK-BR-3 breast cancer cell lines after induction and/or inhibition of NO synthesis. Results CXCR4 staining was predominantly cytoplasmic; this was observed in 50%(56/113 of the tumors. Cytoplasmic CXCR4 expression was significantly correlated with nitrotyrosine levels and lymph node metastasis. Kaplan-Meier survival curves showed that cytoplasmic CXCR4 expression was associated with reduced disease-free and overall survival. In multivariate analysis, cytoplasmic CXCR4 expression emerged as a significant independent predictor for overall and disease-free survival. Cytoplasmic expression of functional CXCR4 in MDA-MB-231 and SK-BR-3 cells was increased by treatment with the NO donor DETA NONOate. This increase was abolished by L-NAME, an inhibitor of NOS. Conclusion Our data showed a role for NO in stimulating cytoplasmic CXCR4 expression in vitro. Formation of the biomarker nitrotyrosine was also correlated with CXCR4 expression and lymph node metastasis in vivo. In addition, cytoplasmic CXCR4 expression may serve as a significant prognostic factor for long-term survival in breast cancer.

  13. Cytoplasmic CXCR4 expression in breast cancer: induction by nitric oxide and correlation with lymph node metastasis and poor prognosis

    International Nuclear Information System (INIS)

    Yasuoka, Hironao; Tsujimoto, Masahiko; Yoshidome, Katsuhide; Nakahara, Masaaki; Kodama, Rieko; Sanke, Tokio; Nakamura, Yasushi

    2008-01-01

    Lymph nodes constitute the first site of metastasis for most malignancies, and the extent of lymph node involvement is a major criterion for evaluating patient prognosis. The CXC chemokine receptor 4 (CXCR4) has been shown to play an important role in lymph node metastasis. Nitric oxide (NO) may also contribute to induction of metastatic ability in human cancers. CXCR4 expression was analyzed in primary human breast carcinoma with long-term follow-up. The relationship between nitrotyrosine levels (a biomarker for peroxynitrate formation from NO in vivo) and lymph node status, CXCR4 immunoreactivity, and other established clinico-pathological parameters, as well as prognosis, was analyzed. Nitrite/nitrate levels and CXCR4 expressions were assessed in MDA-MB-231 and SK-BR-3 breast cancer cell lines after induction and/or inhibition of NO synthesis. CXCR4 staining was predominantly cytoplasmic; this was observed in 50%(56/113) of the tumors. Cytoplasmic CXCR4 expression was significantly correlated with nitrotyrosine levels and lymph node metastasis. Kaplan-Meier survival curves showed that cytoplasmic CXCR4 expression was associated with reduced disease-free and overall survival. In multivariate analysis, cytoplasmic CXCR4 expression emerged as a significant independent predictor for overall and disease-free survival. Cytoplasmic expression of functional CXCR4 in MDA-MB-231 and SK-BR-3 cells was increased by treatment with the NO donor DETA NONOate. This increase was abolished by L-NAME, an inhibitor of NOS. Our data showed a role for NO in stimulating cytoplasmic CXCR4 expression in vitro. Formation of the biomarker nitrotyrosine was also correlated with CXCR4 expression and lymph node metastasis in vivo. In addition, cytoplasmic CXCR4 expression may serve as a significant prognostic factor for long-term survival in breast cancer

  14. Characterization of Elements Regulating the Nuclear-to-Cytoplasmic Translocation of ICP0 in Late Herpes Simplex Virus 1 Infection.

    Science.gov (United States)

    Samrat, Subodh Kumar; Ha, Binh L; Zheng, Yi; Gu, Haidong

    2018-01-15

    Infected cell protein 0 (ICP0) of herpes simplex virus 1 (HSV-1) is an immediate early protein containing a RING-type E3 ubiquitin ligase. It targets several host factors for proteasomal degradation and subsequently activates viral expression. ICP0 has a nuclear localization sequence and functions in the nucleus early during infection. However, later in infection, ICP0 is found solely in the cytoplasm. The molecular mechanism and biological function of the ICP0 nuclear-to-cytoplasmic translocation are not well understood. In this study, we sought to characterize elements important for this translocation. We found that (i) in human embryonic lung fibroblast (HEL) cells, ICP0 C-terminal residues 741 to 775 were necessary but not sufficient for the nuclear-to-cytoplasmic translocation; (ii) the loss of ICP0 E3 ubiquitin ligase activity, which led to defective viral replication in nonpermissive cells, also caused mutant ICP0 to be retained in the nucleus of HEL cells; (iii) in permissive U2OS cells, however, ICP0 lacking E3 ligase activity was translocated to the cytoplasm at a pace faster than that of wild-type ICP0, suggesting that nuclear retention of ICP0 occurs in an ICP0 E3 ligase-dependent manner; and (iv) the ICP0 C terminus and late viral proteins cooperate in order to overcome nuclear retention and stimulate ICP0 cytoplasmic translocation. Taken together, less ICP0 nuclear retention may contribute to the permissiveness of U2OS cells to HSV-1 in the absence of functional ICP0. IMPORTANCE A distinct characteristic for eukaryotes is the compartmentalization of cell metabolic pathways, which allows greater efficiency and specificity of cellular functions. ICP0 of HSV-1 is a multifunctional viral protein that travels through different compartments as infection progresses. Its main regulatory functions are carried out in the nucleus, but it is translocated to the cytoplasm late during HSV-1 infection. To understand the biological significance of cytoplasmic ICP0 in

  15. In vivo evidence of TonB shuttling between the cytoplasmic and outer membrane in Escherichia coli.

    Science.gov (United States)

    Larsen, Ray A; Letain, Tracy E; Postle, Kathleen

    2003-07-01

    Gram-negative bacteria are able to convert potential energy inherent in the proton gradient of the cytoplasmic membrane into active nutrient transport across the outer membrane. The transduction of energy is mediated by TonB protein. Previous studies suggest a model in which TonB makes sequential and cyclic contact with proteins in each membrane, a process called shuttling. A key feature of shuttling is that the amino-terminal signal anchor must quit its association with the cytoplasmic membrane, and TonB becomes associated solely with the outer membrane. However, the initial studies did not exclude the possibility that TonB was artifactually pulled from the cytoplasmic membrane by the fractionation process. To resolve this ambiguity, we devised a method to test whether the extreme TonB amino-terminus, located in the cytoplasm, ever became accessible to the cys-specific, cytoplasmic membrane-impermeant molecule, Oregon Green(R) 488 maleimide (OGM) in vivo. A full-length TonB and a truncated TonB were modified to carry a sole cysteine at position 3. Both full-length TonB and truncated TonB (consisting of the amino-terminal two-thirds) achieved identical conformations in the cytoplasmic membrane, as determined by their abilities to cross-link to the cytoplasmic membrane protein ExbB and their abilities to respond conformationally to the presence or absence of proton motive force. Full-length TonB could be amino-terminally labelled in vivo, suggesting that it was periplasmically exposed. In contrast, truncated TonB, which did not associate with the outer membrane, was not specifically labelled in vivo. The truncated TonB also acted as a control for leakage of OGM across the cytoplasmic membrane. Further, the extent of labelling for full-length TonB correlated roughly with the proportion of TonB found at the outer membrane. These findings suggest that TonB does indeed disengage from the cytoplasmic membrane during energy transduction and shuttle to the outer membrane.

  16. Prevention: Exercise

    Medline Plus

    Full Text Available ... Steroid Injections Lumbar Zygapophysical (Facet) Joint Injections PREVENTION Lifestyle Choices 10 Tips for a Healthy Back Smoking Weight Patient Safety Exercise Strengthening Strengthen ...

  17. Prevention: Exercise

    Medline Plus

    Full Text Available ... Injections PREVENTION Lifestyle Choices 10 Tips for a Healthy Back Smoking Weight Patient Safety Exercise Strengthening Strengthen Your Core! Stretching/Flexibility Aerobic Exercise ...

  18. Prevention: Exercise

    Medline Plus

    Full Text Available ... Watchful Waiting and Education Injection Treatments for Spinal Pain Epidural Steroid Injections Lumbar Zygapophysical (Facet) Joint Injections PREVENTION Lifestyle Choices 10 ...

  19. Praline metabolism by germinating Lilium longiflorum pollen. I. Labelling of cytoplasmic, wall and culture medium molecules

    Directory of Open Access Journals (Sweden)

    W. V. Dashek

    2014-01-01

    Full Text Available Radioactivity occurs in trithloroacetic acid (TCA-soluble and precipitable, cytoplasm and salt-washed walls following germination of Lilium longiflorum, cv. 'Ace' pollen in medium containing [14C]-proline (Pro. Sephadex gel filtration on G-25 through G-100 was employed to determine whether radioactivity in cytoplasm, wall and growth medium from pollen fed [14C]-Pro or [3H]=Pro plus [14C]-arafbinose (Ara was contained within molecules possessing molecular weights of 5,000 to 100,000 daltones or greater. G-25 elution profiles of a crude cytoplasmic fraction (15,000 X g supernatant from [14C]-Pro labelled pollen yielded a radioctive void volume peak and a retarded peak. The void volume peak contained hydroxyproline (Hyp, and exhibited a coincidence of [3H]-Pro and [14C] -Ara labelling when pollen was double labelled with the two isotopes. This peak also contained radioactivity when pollen was germinated in 2-[3H]-myo-inositol. Germination in medium supplemented with 100 µM 2,2'-dipyridyl eliminated radioactivity from 2-[3H]-myo-inositol or [14C]-,Pro in the peak. Filtratian on G-25 of aTCA-soluble fraction of a salt-extract of walls from [14C]-Pro labelled pollen resulted in void volume and two retarded peaks. Void volume and two retarded peaks were also obtained upon G-25 filtration of a cellulase-digest of walls from [M]-Pro labeled pollen. The void volume peak contained Hyp, Lys, Gly, Ala, Ser, Glu and Asp acids, Val, Tyr, Leu or lieu and Pro. Sephadex G-90, 75, and 100 elution profiles of cellulasedigests of walls from [3H]-,Pro and [14C]-Ara labelled pollen yielded radioactive retarded and Hyp-containing void volume peaks with a coincidence of [3H] and [14C] labelling. Label in the void volume was obtained when either rhozyme P11- or pepsin-digests of walls from [14C]-Pro labelled pollen were gel filtered on G-50. Paper electrophoresis coupled with paper chromatography of acid hydrolyzates of salt-washed wall fractions demonstrated 15 of the

  20. Secretion of bacterial lipoproteins: through the cytoplasmic membrane, the periplasm and beyond.

    Science.gov (United States)

    Zückert, Wolfram R

    2014-08-01

    Bacterial lipoproteins are peripherally anchored membrane proteins that play a variety of roles in bacterial physiology and virulence in monoderm (single membrane-enveloped, e.g., gram-positive) and diderm (double membrane-enveloped, e.g., gram-negative) bacteria. After export of prolipoproteins through the cytoplasmic membrane, which occurs predominantly but not exclusively via the general secretory or Sec pathway, the proteins are lipid-modified at the cytoplasmic membrane in a multistep process that involves sequential modification of a cysteine residue and cleavage of the signal peptide by the signal II peptidase Lsp. In both monoderms and diderms, signal peptide processing is preceded by acylation with a diacylglycerol through preprolipoprotein diacylglycerol transferase (Lgt). In diderms but also some monoderms, lipoproteins are further modified with a third acyl chain through lipoprotein N-acyl transferase (Lnt). Fully modified lipoproteins that are destined to be anchored in the inner leaflet of the outer membrane (OM) are selected, transported and inserted by the Lol (lipoprotein outer membrane localization) pathway machinery, which consists of the inner-membrane (IM) ABC transporter-like LolCDE complex, the periplasmic LolA chaperone and the OM LolB lipoprotein receptor. Retention of lipoproteins in the cytoplasmic membrane results from Lol avoidance signals that were originally described as the "+2 rule". Surface localization of lipoproteins in diderms is rare in most bacteria, with the exception of several spirochetal species. Type 2 (T2SS) and type 5 (T5SS) secretion systems are involved in secretion of specific surface lipoproteins of γ-proteobacteria. In the model spirochete Borrelia burgdorferi, surface lipoprotein secretion does not follow established sorting rules, but remains dependent on N-terminal peptide sequences. Secretion through the outer membrane requires maintenance of lipoproteins in a translocation-competent unfolded conformation

  1. Membranous and Cytoplasmic Expression of PD-L1 in Ovarian Cancer Cells.

    Science.gov (United States)

    Qu, Qiu-Xia; Xie, Fang; Huang, Qin; Zhang, Xue-Guang

    2017-01-01

    Expression of programmed death-ligand 1 (PD-L1) on tumor cells represents a powerful immune evasion pathway, but the role of intracellular or cytoplasmic PD-L1 has not been investigated in ovarian cancer cells. Flow cytometry (FCM), Real-time PCR (qPCR), immunohistochemistry (IHC) and western blot were used to determine the expression of PD-L1 in ovarian cancer cells. The cytokines detected in the tumor or tumor associated macrophage (TAM) were used to treat cancer cells. PD-L1 blockade and silencing were used to elucidate the functional significance of cancer-related PD-L1 expression. Based on the results presented, PD-L1 was found variably expressed in the cytoplasm and the cell surface of both HO8910 and SKOV3 cells. TAM or IFN-γ, TNF-α, IL-10 and IL-6 released from TAM stimulated the expression of PD-L1 at the surface of the cancer cells. The IHC results were consistent with the data in vitro showing infiltration of TAM correlated with membranous PD-L1. The increases of PD-L1 at the surface were not due to a shift in the proportion of surface versus intracellular protein, but the contribution of extracellular signal-regulated kinase (ERK)1/2 and phosphoinositide 3-kinase (PI3K) pathway activation. As a consequence, inducible membranous PD-L1 expression on SKOV3 inhibited CD8+ T cell function, and cytoplasmic PD-L1 promoted cancer cell growth. Additionally, in mouse models, both PD-L1 and PD-1 mAb resulted in tumor growth inhibition and demonstrated a potential to decrease the number of PD-1+CD8+T cells. We conclude that TAM induced PD-L1 on the cancer cells represents an immune evasion mechanism. The observations confirm the therapeutic potential of PD-L1/PD-1 mAb to reactivate anti-tumor immunity in ovarian cancer. © 2017 The Author(s). Published by S. Karger AG, Basel.

  2. Functional Hierarchy of Herpes Simplex Virus 1 Viral Glycoproteins in Cytoplasmic Virion Envelopment and Egress

    Science.gov (United States)

    Chouljenko, Dmitry V.; Kim, In-Joong; Chouljenko, Vladimir N.; Subramanian, Ramesh; Walker, Jason D.

    2012-01-01

    Herpes simplex virus 1 (HSV-1) viral glycoproteins gD (carboxyl terminus), gE, gK, and gM, the membrane protein UL20, and membrane-associated protein UL11 play important roles in cytoplasmic virion envelopment and egress from infected cells. We showed previously that a recombinant virus carrying a deletion of the carboxyl-terminal 29 amino acids of gD (gDΔct) and the entire gE gene (ΔgE) did not exhibit substantial defects in cytoplasmic virion envelopment and egress (H. C. Lee et al., J. Virol. 83:6115–6124, 2009). The recombinant virus ΔgM2, engineered not to express gM, produced a 3- to 4-fold decrease in viral titers and a 50% reduction in average plaque sizes in comparison to the HSV-1(F) parental virus. The recombinant virus containing all three mutations, gDΔct-ΔgM2-ΔgE, replicated approximately 1 log unit less efficiently than the HSV-1(F) parental virus and produced viral plaques which were on average one-third the size of those of HSV-1(F). The recombinant virus ΔUL11-ΔgM2, engineered not to express either UL11 or gM, replicated more than 1 log unit less efficiently and produced significantly smaller plaques than UL11-null or gM-null viruses alone, in agreement with the results of Leege et al. (T. Leege et al., J. Virol. 83:896-907, 2009). Analyses of particle-to-PFU ratios, relative plaque size, and kinetics of virus growth and ultrastructural visualization of glycoprotein-deficient mutant and wild-type virions indicate that gDΔct, gE, and gM function in a cooperative but not redundant manner in infectious virion morphogenesis. Overall, comparisons of single, double, and triple mutant viruses generated in the same HSV-1(F) genetic background indicated that lack of either UL20 or gK expression caused the most severe defects in cytoplasmic envelopment, egress, and infectious virus production, followed by the double deletion of UL11 and gM. PMID:22318149

  3. Mapping of ESE-1 subdomains required to initiate mammary epithelial cell transformation via a cytoplasmic mechanism

    Directory of Open Access Journals (Sweden)

    Tentler John J

    2011-08-01

    Full Text Available Abstract Background The ETS family transcription factor ESE-1 is often overexpressed in human breast cancer. ESE-1 initiates transformation of MCF-12A cells via a non-transcriptional, cytoplasmic process that is mediated by a unique 40-amino acid serine and aspartic acid rich (SAR subdomain, whereas, ESE-1's nuclear transcriptional property is required to maintain the transformed phenotype of MCF7, ZR-75-1 and T47D breast cancer cells. Results To map the minimal functional nuclear localization (NLS and nuclear export (NES signals, we fused in-frame putative NLS and NES motifs between GFP and the SAR domain. Using these GFP constructs as reporters of subcellular localization, we mapped a single NLS to six basic amino acids (242HGKRRR247 in the AT-hook and two CRM1-dependent NES motifs, one to the pointed domain (NES1: 102LCNCALEELRL112 and another to the DNA binding domain (DBD, (NES2: 275LWEFIRDILI284. Moreover, analysis of a putative NLS located in the DBD (316GQKKKNSN323 by a similar GFP-SAR reporter or by internal deletion of the DBD, revealed this sequence to lack NLS activity. To assess the role of NES2 in regulating ESE-1 subcellular localization and subsequent transformation potency, we site-specifically mutagenized NES2, within full-length GFP-ESE-1 and GFP-NES2-SAR reporter constructs. These studies show that site-specific mutation of NES2 completely abrogates ESE-1 transforming activity. Furthermore, we show that exclusive cytoplasmic targeting of the SAR domain is sufficient to initiate transformation, and we report that an intact SAR domain is required, since block mutagenesis reveals that an intact SAR domain is necessary to maintain its full transforming potency. Finally, using a monoclonal antibody targeting the SAR domain, we demonstrate that the SAR domain contains a region accessible for protein - protein interactions. Conclusions These data highlight that ESE-1 contains NLS and NES signals that play a critical role in

  4. Live confocal imaging of intrashell cytoplasm and calcification process in a symbiotic large benthic foraminifer Amphisorus kudakajimensis

    Science.gov (United States)

    Ohno, Yoshikazu; Fuita, Kazuhiko; Toyofuku, Takashi; Nakamura, Takashi

    2015-04-01

    Algal symbiont-bearing large benthic foraminifers are primary and carbonate producers as well as paleoenvironmental indicators in tropical and subtropical reef environments. Despite their importance, their cellular physiology is not well known. In this study, we have developed methods to observe in vivo images of a living symbiotic porcelaneous large foraminifer, Amphisorus kudakajimensis. Nikon A1 confocal laser scanning microscope with Calcein-AM as a fluorescent indicator was used to visualize morphology and streaming of cytoplasm in living A. kudakajimensis. The observation showed that in the marginal part of intrashell cytoplasm, cytoplasm gradually became highly branched and vacuolated. We also succeeded in observing elevated pH (pH 9.0) and lowered pH (pH 6.0) vesicles in reticulopodia-like cytoplasms, using a cell membrane-impermeable fluorescent pH indicator with low toxicity (HPTS). This study demonstrate the use of confocal microscopy in studying cytoplasmic dynamics and initial calcification process such as seawater endocytosis and alkalization of seawater vacuoles.

  5. An ABC transporter B family protein, ABCB19, is required for cytoplasmic streaming and gravitropism of the inflorescence stems.

    Science.gov (United States)

    Okamoto, Keishi; Ueda, Haruko; Shimada, Tomoo; Tamura, Kentaro; Koumoto, Yasuko; Tasaka, Masao; Morita, Miyo Terao; Hara-Nishimura, Ikuko

    2016-01-01

    A significant feature of plant cells is the extensive motility of organelles and the cytosol, which was originally defined as cytoplasmic streaming. We suggested previously that a three-way interaction between plant-specific motor proteins myosin XIs, actin filaments, and the endoplasmic reticulum (ER) was responsible for cytoplasmic streaming. (1) Currently, however, there are no reports of molecular components for cytoplasmic streaming other than the actin-myosin-cytoskeleton and ER-related proteins. In the present study, we found that elongated cells of inflorescence stems of Arabidopsis thaliana exhibit vigorous cytoplasmic streaming. Statistical analysis showed that the maximal velocity of plastid movements is 7.26 µm/s, which is much faster than the previously reported velocities of organelles. Surprisingly, the maximal velocity of streaming in the inflorescence stem cells was significantly reduced to 1.11 µm/s in an Arabidopsis mutant, abcb19-101, which lacks ATP BINDING CASSETTE SUBFAMILY B19 (ABCB19) that mediates the polar transport of the phytohormone auxin together with PIN-FORMED (PIN) proteins. Polar auxin transport establishes the auxin concentration gradient essential for plant development and tropisms. Deficiency of ABCB19 activity eventually caused enhanced gravitropic responses of the inflorescence stems and abnormally flexed inflorescence stems. These results suggest that ABCB19-mediated auxin transport plays a role not only in tropism regulation, but also in cytoplasmic streaming.

  6. HuR represses Wnt/β-catenin-mediated transcriptional activity by promoting cytoplasmic localization of β-catenin

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Inae; Hur, Jung; Jeong, Sunjoo, E-mail: sjsj@dankook.ac.kr

    2015-01-30

    Highlights: • Wnt signaling as well as β-catenin overexpression enhance HuR cytoplasmic export. • HuR overexpression promotes cytoplasmic localization of β-catenin from the perinuclear fraction. • Wnt/β-catenin-mediated transcriptional activity is repressesed by HuR. - Abstract: β-Catenin is the key transcriptional activator of canonical Wnt signaling in the nucleus; thus, nuclear accumulation of β-catenin is a critical step for expressing target genes. β-Catenin accumulates in the nucleus of cancer cells where it activates oncogenic target genes. Hu antigen R (HuR) is a RNA binding protein that regulates multiple post-transcriptional processes including RNA stability. Thus, cytoplasmic HuR protein may be involved in tumorigenesis by stabilizing oncogenic transcripts, but the molecular mechanism remains unclear. Here, we observed that Wnt/β-catenin signaling induced export of the HuR protein, whereas HuR overexpression promoted accumulation of the β-catenin protein in the cytoplasm. Thus, Wnt/β-catenin-mediated transcriptional activity in the nucleus was reduced by overexpressing HuR. These results suggest novel and uncharacterized cytoplasmic β-catenin functions related to HuR-mediated RNA metabolism in cancer cells.

  7. HuR represses Wnt/β-catenin-mediated transcriptional activity by promoting cytoplasmic localization of β-catenin

    International Nuclear Information System (INIS)

    Kim, Inae; Hur, Jung; Jeong, Sunjoo

    2015-01-01

    Highlights: • Wnt signaling as well as β-catenin overexpression enhance HuR cytoplasmic export. • HuR overexpression promotes cytoplasmic localization of β-catenin from the perinuclear fraction. • Wnt/β-catenin-mediated transcriptional activity is repressesed by HuR. - Abstract: β-Catenin is the key transcriptional activator of canonical Wnt signaling in the nucleus; thus, nuclear accumulation of β-catenin is a critical step for expressing target genes. β-Catenin accumulates in the nucleus of cancer cells where it activates oncogenic target genes. Hu antigen R (HuR) is a RNA binding protein that regulates multiple post-transcriptional processes including RNA stability. Thus, cytoplasmic HuR protein may be involved in tumorigenesis by stabilizing oncogenic transcripts, but the molecular mechanism remains unclear. Here, we observed that Wnt/β-catenin signaling induced export of the HuR protein, whereas HuR overexpression promoted accumulation of the β-catenin protein in the cytoplasm. Thus, Wnt/β-catenin-mediated transcriptional activity in the nucleus was reduced by overexpressing HuR. These results suggest novel and uncharacterized cytoplasmic β-catenin functions related to HuR-mediated RNA metabolism in cancer cells

  8. Balanced nuclear and cytoplasmic activities of EDS1 are required for a complete plant innate immune response.

    Directory of Open Access Journals (Sweden)

    Ana V García

    2010-07-01

    Full Text Available An important layer of plant innate immunity to host-adapted pathogens is conferred by intracellular nucleotide-binding/oligomerization domain-leucine rich repeat (NB-LRR receptors recognizing specific microbial effectors. Signaling from activated receptors of the TIR (Toll/Interleukin-1 Receptor-NB-LRR class converges on the nucleo-cytoplasmic immune regulator EDS1 (Enhanced Disease Susceptibility1. In this report we show that a receptor-stimulated increase in accumulation of nuclear EDS1 precedes or coincides with the EDS1-dependent induction and repression of defense-related genes. EDS1 is capable of nuclear transport receptor-mediated shuttling between the cytoplasm and nucleus. By enhancing EDS1 export from inside nuclei (through attachment of an additional nuclear export sequence (NES or conditionally releasing EDS1 to the nucleus (by fusion to a glucocorticoid receptor (GR in transgenic Arabidopsis we establish that the EDS1 nuclear pool is essential for resistance to biotrophic and hemi-biotrophic pathogens and for transcriptional reprogramming. Evidence points to post-transcriptional processes regulating receptor-triggered accumulation of EDS1 in nuclei. Changes in nuclear EDS1 levels become equilibrated with the cytoplasmic EDS1 pool and cytoplasmic EDS1 is needed for complete resistance and restriction of host cell death at infection sites. We propose that coordinated nuclear and cytoplasmic activities of EDS1 enable the plant to mount an appropriately balanced immune response to pathogen attack.

  9. Post-translational methylation of high mobility group box 1 (HMGB1) causes its cytoplasmic localization in neutrophils.

    Science.gov (United States)

    Ito, Ichiaki; Fukazawa, Jutarou; Yoshida, Michiteru

    2007-06-01

    High mobility group box 1 (HMGB1) protein plays multiple roles in transcription, replication, and cellular differentiation. HMGB1 is also secreted by activated monocytes and macrophages and passively released by necrotic or damaged cells, stimulating inflammation. HMGB1 is a novel antigen of anti-neutrophil cytoplasmic antibodies (ANCA) observed in the sera of patients with ulcerative colitis and autoimmune hepatitis, suggesting that HMGB1 is secreted from neutrophils to the extracellular milieu. However, the actual distribution of HMGB1 in the cytoplasm of neutrophils and the mechanisms responsible for it are obscure. Here we show that HMGB1 in neutrophils is post-translationally mono-methylated at Lys42. The methylation alters the conformation of HMGB1 and weakens its DNA binding activity, causing it to become largely distributed in the cytoplasm by passive diffusion out of the nucleus. Thus, post-translational methylation of HMGB1 causes its cytoplasmic localization in neutrophils. This novel pathway explains the distribution of nuclear HMGB1 to the cytoplasm and is important for understanding how neutrophils release HMGB1 to the extracellular milieu.

  10. The effect of the cytoplasmic tail of influenza C virus CM2 protein on its biochemical properties and intracellular processing.

    Science.gov (United States)

    Shimotai, Yoshitaka; Goto, Takanari; Matsuzaki, Yoko; Muraki, Yasushi; Sugawara, Kanetsu; Hongo, Seiji

    2015-09-01

    CM2 is an integral membrane protein encoded by the influenza C virus M gene. To examine the effects of the cytoplasmic tail of CM2 on its biochemical properties, deletion and substitution mutations were introduced into CM2 cytoplasmic tail at residues 47-115, and the expressed CM2 mutants were investigated. Although the cytoplasmic tail is not essential for the oligomerization of CM2, it may affect the degree of oligomerization. The residues 47-48, 67-69, 73-90 and 113-115 were all required for the proper expression of CM2. Pulse-chase experiments suggest that residues 47-48, 67-69, 73-75 and 79-87 stabilize CM2, thereby affecting CM2 expression. The C-terminal region at residues 61-115 is not essential for CM2 transport to the cell surface, and a 14-amino-acid, but not an 11-amino-acid, cytoplasmic tail is sufficient for the cell surface expression of CM2. These results suggest that either certain amino acid sequences or the length of the CM2 cytoplasmic tail are necessary for the proper conformational maturation, stability, expression level and intracellular transport of CM2.

  11. Glucocorticoid control of rat growth hormone gene expression: Effect on cytoplasmic messenger ribonucleic acid production and degradation

    International Nuclear Information System (INIS)

    Gertz, B.J.; Gardner, D.G.; Baxter, J.D.

    1987-01-01

    The effect of the glucocorticoid dexamethasone on the production and degradation of rat GH (rGH) cytoplasmic mRNA was studied in cultured rat pituitary tumor (GC) cells. The incorporation of [3H]uridine into both rGH cytoplasmic mRNA and the pyrimidine nucleotide precursor pool was determined in hormone-treated and control cells. From these measurements glucocorticoid effects on absolute production rates of rGH cytoplasmic mRNA were determined and compared to effects on rGH mRNA accumulation. Rat GH mRNA half-life was then calculated based on a first-order decay model. Rat GH mRNA half-life was also directly assayed by: (1) pulse-chase studies and (2) measuring the kinetics of decay of rGH mRNA in cells after transfer from serum-containing to hormone-deficient media. From these independent analyses rGH mRNA half-life estimates ranged from 28-55 h in different experiments. Within individual experiments there was little variability of rGH mRNA decay rates; glucocorticoids were found not to alter the stability of rGH cytoplasmic mRNA. Glucocorticoid induction of rGH cytoplasmic mRNA accumulation was accounted for solely on the basis of increased mRNA production

  12. An ABC transporter B family protein, ABCB19, is required for cytoplasmic streaming and gravitropism of the inflorescence stems

    Science.gov (United States)

    Okamoto, Keishi; Ueda, Haruko; Shimada, Tomoo; Tamura, Kentaro; Koumoto, Yasuko; Tasaka, Masao; Morita, Miyo Terao; Hara-Nishimura, Ikuko

    2016-01-01

    Abstract A significant feature of plant cells is the extensive motility of organelles and the cytosol, which was originally defined as cytoplasmic streaming. We suggested previously that a three-way interaction between plant-specific motor proteins myosin XIs, actin filaments, and the endoplasmic reticulum (ER) was responsible for cytoplasmic streaming.1 Currently, however, there are no reports of molecular components for cytoplasmic streaming other than the actin-myosin-cytoskeleton and ER-related proteins. In the present study, we found that elongated cells of inflorescence stems of Arabidopsis thaliana exhibit vigorous cytoplasmic streaming. Statistical analysis showed that the maximal velocity of plastid movements is 7.26 µm/s, which is much faster than the previously reported velocities of organelles. Surprisingly, the maximal velocity of streaming in the inflorescence stem cells was significantly reduced to 1.11 µm/s in an Arabidopsis mutant, abcb19-101, which lacks ATP BINDING CASSETTE SUBFAMILY B19 (ABCB19) that mediates the polar transport of the phytohormone auxin together with PIN-FORMED (PIN) proteins. Polar auxin transport establishes the auxin concentration gradient essential for plant development and tropisms. Deficiency of ABCB19 activity eventually caused enhanced gravitropic responses of the inflorescence stems and abnormally flexed inflorescence stems. These results suggest that ABCB19-mediated auxin transport plays a role not only in tropism regulation, but also in cytoplasmic streaming. PMID:26337543

  13. Differences in cytoplasmic maturation between the BCB+ and control porcine oocytes do not justify application of the BCB test for a standard IVM protocol.

    Science.gov (United States)

    Pawlak, Piotr; Warzych, Ewelina; Chabowska, Agnieszka; Lechniak, Dorota

    2014-03-07

    The Brilliant Cresyl Blue (BCB) test relies on G6PDH activity and a simple protocol for the selection of higher quality oocytes. Although the BCB+ oocytes of all the species that have been investigated are characterized by superior quality when compared to BCB- counterparts, application of the test for embryo production still remains an open issue. The aim of our study was to compare BCB+ and the control oocytes (not subjected to the BCB test) in terms of selected aspects of cytoplasmic maturation (mtDNA copy number, mitochondria distribution, relative transcript abundance of six marker genes). The results of our study revealed more relevant differences within the BCB+ and the control oocytes (before and after IVM) than between the two categories of oocytes. There was no difference in the transcript abundance of the BCB+ and the control oocytes in 5 out of 6 analyzed genes (BMP15, GDF9, ATP5A1, EEF1A, ZAR1) and in mtDNA content (pre-IVM 179609 vs. 176595 and post-IVM 187243 vs. 246984, respectively). With regard to mitochondria distribution in pre- and post-IVM oocytes, there was nonsignificant tendency for a more frequent occurrence of the expected patterns in the BCB+ group. The results of the present study do not support the application of BCB staining in a routine IVM protocol due to relatively high similarity in selected parameters characterizing cytoplasmic maturation of BCB+ and control oocytes. This high similarity may results from the limited amount of less competent BCB- oocytes (10%) still present among nonselected oocytes of proper morphology.

  14. A role for complexes of survival of motor neurons (SMN) protein with gemins and profilin in neurite-like cytoplasmic extensions of cultured nerve cells

    International Nuclear Information System (INIS)

    Sharma, Aarti; Lambrechts, Anja; Le thi Hao; Le, Thanh T.; Sewry, Caroline A.; Ampe, Christophe; Burghes, Arthur H.M.; Morris, Glenn E.

    2005-01-01

    Spinal muscular atrophy (SMA) is caused by reduced levels of SMN (survival of motor neurons protein) and consequent loss of motor neurons. SMN is involved in snRNP transport and nuclear RNA splicing, but axonal transport of SMN has also been shown to occur in motor neurons. SMN also binds to the small actin-binding protein, profilin. We now show that SMN and profilin II co-localise in the cytoplasm of differentiating rat PC12 cells and in neurite-like extensions, especially at their growth cones. Many components of known SMN complexes were also found in these extensions, including gemin2 (SIP-1), gemin6, gemin7 and unrip (unr-interacting protein). Coilin p80 and Sm core protein immunoreactivity, however, were seen only in the nucleus. SMN is known to associate with β-actin mRNA and specific hnRNPs in axons and in neurite extensions of cultured nerve cells, and SMN also stimulates neurite outgrowth in cultures. Our results are therefore consistent with SMN complexes, rather than SMN alone, being involved in the transport of actin mRNPs along the axon as in the transport of snRNPs into the nucleus by similar SMN complexes. Antisense knockdown of profilin I and II isoforms inhibited neurite outgrowth of PC12 cells and caused accumulation of SMN and its associated proteins in cytoplasmic aggregates. BIAcore studies demonstrated a high affinity interaction of SMN with profilin IIa, the isoform present in developing neurons. Pathogenic missense mutations in SMN, or deletion of exons 5 and 7, prevented this interaction. The interaction is functional in that SMN can modulate actin polymerisation in vitro by reducing the inhibitory effect of profilin IIa. This suggests that reduced SMN in SMA might cause axonal pathfinding defects by disturbing the normal regulation of microfilament growth by profilins

  15. Role of the beta1-integrin cytoplasmic tail in mediating invasin-promoted internalization of Yersinia

    DEFF Research Database (Denmark)

    Gustavsson, Anna; Armulik, Annika; Brakebusch, Cord

    2002-01-01

    Invasin of Yersinia pseudotuberculosis binds to beta1-integrins on host cells and triggers internalization of the bacterium. To elucidate the mechanism behind the beta1-integrin-mediated internalization of Yersinia, a beta1-integrin-deficient cell line, GD25, transfected with wild-type beta1A, beta......1B or different mutants of the beta1A subunit was used. Both beta1A and beta1B bound to invasin-expressing bacteria, but only beta1A was able to mediate internalization of the bacteria. The cytoplasmic region of beta1A, differing from beta1B, contains two NPXY motifs surrounding a double threonine...... noted that cells affected in bacterial internalization exhibited reduced spreading capability when seeded onto invasin, suggesting a correlation between the internalization of invasin-expressing bacteria and invasin-induced spreading. Likewise, integrins defective in forming peripheral focal complex...

  16. Brain synaptosomes harbor more than one cytoplasmic system of protein synthesis.

    Science.gov (United States)

    Cefaliello, Carolina; Eyman, Maria; Melck, Dominique; De Stefano, Rosanna; Ferrara, Eugenia; Crispino, Marianna; Giuditta, Antonio

    2014-11-01

    Synaptosomal protein synthesis from rat brain is selectively increased by learning and is massively enhanced during the recovery period from brain ischemia. To lay the groundwork for identification of the involved synaptic elements, we examined the effects induced by varying the concentrations of extracellular cations and endogenous calcium. Most of the recorded rate response curves exhibited biphasic profiles that suggested the presence of more than one translation system. Because comparable profiles were obtained by fully inhibiting mitochondrial translation, the data indicated the involvement of cytoplasmic translation systems present in different synaptosomal classes. Their properties may be individually investigated by exploiting the partially inhibited conditions we have described. The identification of the synaptic elements from which they originated and their newly synthesized proteins will significantly expand our understanding of the synaptic contribution to brain plastic events. Copyright © 2014 Wiley Periodicals, Inc.

  17. Analysis of a solid-phase radioimmunoassay for antibodies to cytoplasmic antigen fractions of Candida albicans

    International Nuclear Information System (INIS)

    Mauch, H.; Bromback, J.

    1981-01-01

    An indirect solid-phase radioimmunoassay (SPRIA) in individual polystyrene microtiter cups has been adapted for measurement of antibody to various cytoplasmic and carbohydrate antigen fractions of Candida albicans. The assay was optimized for sensitivity, precision and linearization of serum dilution curves. The optimized procedure allows computerized measurement of anti-Candida antibodies and can be used for measurement of antibody over a wide concentration range. The procedure obviates variation due to changes in day-to-day counts as a result of isotope decay and end-point antibody dilutions. The assay has been used to demonstrate a Poisson-like distribution of antibody levels in the sera of persons showing no symptoms of candidiasis. The minimum antibody level detectable by the assay is about two orders of magnitude lower than the lowest level found in human serum and 4 orders of magnitude lower than the most sensitive test used hitherto, the hemagglutination test. (Auth.)

  18. Cytoplasmic Dynein Is Required for the Spatial Organization of Protein Aggregates in Filamentous Fungi

    Directory of Open Access Journals (Sweden)

    Martin J. Egan

    2015-04-01

    Full Text Available Eukaryotes have evolved multiple strategies for maintaining cellular protein homeostasis. One such mechanism involves neutralization of deleterious protein aggregates via their defined spatial segregation. Here, using the molecular disaggregase Hsp104 as a marker for protein aggregation, we describe the spatial and temporal dynamics of protein aggregates in the filamentous fungus Aspergillus nidulans. Filamentous fungi, such as A. nidulans, are a diverse group of species of major health and economic importance and also serve as model systems for studying highly polarized eukaryotic cells. We find that microtubules promote the formation of Hsp104-positive aggregates, which coalesce into discrete subcellular structures in a process dependent on the microtubule-based motor cytoplasmic dynein. Finally, we find that impaired clearance of these inclusions negatively impacts retrograde trafficking of endosomes, a conventional dynein cargo, indicating that microtubule-based transport can be overwhelmed by chronic cellular stress.

  19. Early Outcomes in Children With Antineutrophil Cytoplasmic Antibody-Associated Vasculitis

    DEFF Research Database (Denmark)

    Morishita, Kimberly A; Moorthy, Lakshmi N; Lubieniecka, Joanna M

    2017-01-01

    OBJECTIVE: To characterize the early disease course in childhood-onset antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) and the 12-month outcomes in children with AAV. METHODS: Eligible subjects were children entered into the Pediatric Vasculitis Initiative study who were...... (Pediatric Vasculitis Activity Score [PVAS] of 0) at 12 months with a corticosteroid dosage of ..., presence of damage at 12 months (measured by a modified Pediatric Vasculitis Damage Index [PVDI]; score 0 = no damage, score 1 = one damage item present), and relapse rates at 12 months. RESULTS: In total, 105 children with AAV were included in the study. The median age at diagnosis was 13.8 years...

  20. The 75-kilodalton cytoplasmic Chlamydia trachomatis L2 polypeptide is a DnaK-like protein

    DEFF Research Database (Denmark)

    Birkelund, Svend; Lundemose, AG; Christiansen, Gunna

    1990-01-01

    ,980-base-pair open reading frame revealed 94% homology with a 75-kilodalton protein from C. trachomatis serovar D and 57% homology with the DnaK proteins of E. coli and of Bacillus megaterium, while amino acid homology with human heat shock protein 70 (hsp70) was 42%. The promoter region was identified......The gene coding for the 75-kilodalton cytoplasmic Chlamydia trachomatis L2 polypeptide has been cloned in Escherichia coli, and the nucleotide sequence has been determined. The cloned DNA fragment contained the coding region as well as the putative promoter. The deduced amino acid sequence of the 1...... by computer search and by primer extension of mRNA synthesized in recombinant E. coli. The promoter region which differed from the putative promoter region in serovar D was shown to be a mixed promoter type in which the -10 region showed a regular TATA box configuration while the -35 region showed high...

  1. Anti-Neutrophil Cytoplasmic Antibody-associated Vasculitis (AAV) Restricted to the Limbs: A Case Report.

    Science.gov (United States)

    Ojima, Yoshie; Sawada, Kinya; Fujii, Hiroshi; Shirai, Tsuyoshi; Saito, Ayako; Kagaya, Saeko; Aoki, Satoshi; Takeuchi, Yoichi; Ishii, Tomonori; Nagasawa, Tasuku

    2017-12-27

    A previously healthy 58-year-old man was admitted for muscle pain and weakness (manual muscle testing [MMT] of 4/4 for upper and lower limbs). We detected elevated levels of inflammatory makers and PR3-anti-neutrophil cytoplasmic antibody (ANCA). Subsequently, the muscle weakness rapidly progressed to an MMT of 2 for all limbs. Magnetic resonance imaging indicated muscle edema, and the CK level increased to 29,998 U/L. mPSL and cyclophosphamide pulse therapy improved the patient symptoms. MMT recovered to 4 for all limbs. A muscle biopsy showed degenerated muscle fibers surrounded by neutrophil-predominant infiltration. In addition, lamina elastic breakdown and fibrinoid necrosis of arterioles were observed. A final diagnosis of microscopic polyangiitis (MPA) limited to the muscles was made.

  2. Developmental potential of human oocytes reconstructed by transferring somatic cell nuclei into polyspermic zygote cytoplasm

    International Nuclear Information System (INIS)

    Fan, Yong; Chen, Xinjie; Luo, Yumei; Chen, Xiaolin; Li, Shaoying; Huang, Yulin; Sun, Xiaofang

    2009-01-01

    The generation of patient-specific nuclear transfer embryonic stem cells holds huge promise in modern regenerative medicine and cell-based drug discovery. Since human in vivo matured oocytes are not readily available, human therapeutic cloning is developing slowly. Here, we investigated for the first time whether human polyspermic zygotes could support preimplantation development of cloned embryos. Our results showed that polyspermic zygotes could be used as recipients for human somatic cell nuclear transfer (SCNT). The preimplantation developmental potential of SCNT embryos from polyspermic zygotes was limited to the 8-cell stage. Since ES cell lines can be derived from single blastomeres, these results may have important significance for human ES cells derived by SCNT. In addition, confocal images demonstrated that all of the SCNT embryos that failed to cleave showed abnormal microtubule organization. The results of the present study suggest that polyspermic human zygotes could be used as a potential source of recipient cytoplasm for SCNT.

  3. N-tail translocation of mature beta-lactamase across the Escherichia coli cytoplasmic membrane.

    Science.gov (United States)

    Mitsopoulos, C; Hashemzadeh-Bonehi, L; Broome-Smith, J K

    1997-12-08

    Mature beta-lactamase was attached to the N-terminus of human glycophorin C, an N-out membrane protein lacking a cleavable signal peptide (an N-tail membrane protein). When synthesised in Escherichia coli more than 30% of the intact mature beta-lactamase-glycophorin C molecules assembled N-out, C-in into the cytoplasmic membrane. The N-tail translocated beta-lactamase folded into an enzymatically active form, but it was more susceptible to proteolysis than the equivalent portion of beta-lactamase-glycophorin C synthesised with an N-terminal signal peptide. Its translocation was virtually abolished when the N-out domain of glycophorin C was truncated or when the basic residues C-terminally flanking the glycophorin C membrane-spanning segment were replaced with neutral ones.

  4. Developmental potential of human oocytes reconstructed by transferring somatic cell nuclei into polyspermic zygote cytoplasm

    Energy Technology Data Exchange (ETDEWEB)

    Fan, Yong; Chen, Xinjie; Luo, Yumei; Chen, Xiaolin; Li, Shaoying; Huang, Yulin [Institute of Gynecology and Obstetrics, The Third Affiliated Hospital of Guangzhou Medical College, Duobao Road 63, Guangzhou, Guangdong (China); Sun, Xiaofang, E-mail: xiaofangsun@hotmail.com [Institute of Gynecology and Obstetrics, The Third Affiliated Hospital of Guangzhou Medical College, Duobao Road 63, Guangzhou, Guangdong (China)

    2009-04-24

    The generation of patient-specific nuclear transfer embryonic stem cells holds huge promise in modern regenerative medicine and cell-based drug discovery. Since human in vivo matured oocytes are not readily available, human therapeutic cloning is developing slowly. Here, we investigated for the first time whether human polyspermic zygotes could support preimplantation development of cloned embryos. Our results showed that polyspermic zygotes could be used as recipients for human somatic cell nuclear transfer (SCNT). The preimplantation developmental potential of SCNT embryos from polyspermic zygotes was limited to the 8-cell stage. Since ES cell lines can be derived from single blastomeres, these results may have important significance for human ES cells derived by SCNT. In addition, confocal images demonstrated that all of the SCNT embryos that failed to cleave showed abnormal microtubule organization. The results of the present study suggest that polyspermic human zygotes could be used as a potential source of recipient cytoplasm for SCNT.

  5. Response of amphibian egg cytoplasm to novel gravity orientation and centrifugation

    Science.gov (United States)

    Neff, A. W.; Wakahara, M.; Jurand, A.; Malacinski, G. M.

    1983-01-01

    The effects of inversion and centrifugation on the compartmentalization of cytoplasm in Xenopus laevis eggs are investigated experimentally. The rearrangement of yolk-platelet compartments (YPC) characterized by morphology, density, and viscosity differences is studied in fertilized, unfertilized, and unfertilized electrically activated eggs in normal, and inverted positions and with and without centrifugation at 10-183 x g for 5 min. The eggs are fixed and embedded in plastic or paraffin prior to sagittal sectioning (0.5, 4, or 8 microns) and microscopic examination; the results are presented in a diagram and discussed. A density-compartment model combining both animal/vegetal and dorsal/ventral polarities is proposed: YPC determined without gravity orientation during oogenesis respond to both sperm entrance point and gravity after fertilization, and the response involves breaking of the radial symmetry of the egg. It is predicted that Xenopus eggs in a microgravity environment will encounter difficulties in establishing a primary embryonic axis.

  6. Superdiffusion dominates intracellular particle motion in the supercrowded cytoplasm of pathogenic Acanthamoeba castellanii

    Science.gov (United States)

    Reverey, Julia F.; Jeon, Jae-Hyung; Bao, Han; Leippe, Matthias; Metzler, Ralf; Selhuber-Unkel, Christine

    2015-06-01

    Acanthamoebae are free-living protists and human pathogens, whose cellular functions and pathogenicity strongly depend on the transport of intracellular vesicles and granules through the cytosol. Using high-speed live cell imaging in combination with single-particle tracking analysis, we show here that the motion of endogenous intracellular particles in the size range from a few hundred nanometers to several micrometers in Acanthamoeba castellanii is strongly superdiffusive and influenced by cell locomotion, cytoskeletal elements, and myosin II. We demonstrate that cell locomotion significantly contributes to intracellular particle motion, but is clearly not the only origin of superdiffusivity. By analyzing the contribution of microtubules, actin, and myosin II motors we show that myosin II is a major driving force of intracellular motion in A. castellanii. The cytoplasm of A. castellanii is supercrowded with intracellular vesicles and granules, such that significant intracellular motion can only be achieved by actively driven motion, while purely thermally driven diffusion is negligible.

  7. Preventing Mother-to-Child Transmission of HIV

    Science.gov (United States)

    ... a Preventive HIV Vaccine? HIV/AIDS Clinical Trials HIV Prevention The Basics of HIV Prevention Preventing Mother-to-Child Transmission of HIV Post- ... Related Content Protecting Baby from HIV (infographic) AIDSource | HIV Prevention: Mother-to-Child Transmission MedlinePlus | HIV/AIDS and ...

  8. ATP and magnesium drive conformational changes of the Na+/K+-ATPase cytoplasmic headpiece.

    Science.gov (United States)

    Grycova, Lenka; Sklenovsky, Petr; Lansky, Zdenek; Janovska, Marika; Otyepka, Michal; Amler, Evzen; Teisinger, Jan; Kubala, Martin

    2009-05-01

    Conformational changes of the Na(+)/K(+)-ATPase isolated large cytoplasmic segment connecting transmembrane helices M4 and M5 (C45) induced by the interaction with enzyme ligands (i.e. Mg(2+) and/or ATP) were investigated by means of the intrinsic tryptophan fluorescence measurement and molecular dynamic simulations. Our data revealed that this model system consisting of only two domains retained the ability to adopt open or closed conformation, i.e. behavior, which is expected from the crystal structures of relative Ca(2+)-ATPase from sarco(endo)plasmic reticulum for the corresponding part of the entire enzyme. Our data revealed that the C45 is found in the closed conformation in the absence of any ligand, in the presence of Mg(2+) only, or in the simultaneous presence of Mg(2+) and ATP. Binding of the ATP alone (i.e. in the absence of Mg(2+)) induced open conformation of the C45. The fact that the transmembrane part of the enzyme was absent in our experiments suggested that the observed conformational changes are consequences only of the interaction with ATP or Mg(2+) and may not be related to the transported cations binding/release, as generally believed. Our data are consistent with the model, where ATP binding to the low-affinity site induces conformational change of the cytoplasmic part of the enzyme, traditionally attributed to E2-->E1 transition, and subsequent Mg(2+) binding to the enzyme-ATP complex induces in turn conformational change traditionally attributed to E1-->E2 transition.

  9. Anti-neutrophil cytoplasmic antibody-associated vasculitis with renal involvement: Analysis of 89 cases.

    Science.gov (United States)

    Caravaca-Fontán, Fernando; Yerovi, Estefanía; Delgado-Yagu E, María; Galeano, Cristina; Pampa-Saico, Saúl; Tenorio, Maria Teresa; Liaño, Fernando

    2017-01-06

    The anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis with renal involvement are associated with high morbi-mortality. In this study we analyse if the prognosis of these diseases have improved in recent years, and which factors influence the outcomes. Retrospective single-centre observational study, which included all patients diagnosed with microscopic polyangiitis and granulomatosis with polyangiitis with renal involvement in the last 25 years. Demographic, clinical and biochemical parameters of prognostic interest were recorded. The differences between four chronological periods were analysed, along with the determinants of a poor outcome (death or end-stage renal disease). Eighty-nine patients were included (mean age 64±15 years). Sixty-four patients (72%) had microscopic polyangiitis and 25 (28%) granulomatosis with polyangiitis. During the study period, 37 (42%) patients died. Through Cox regression analysis, the best determinants of mortality were the initial glomerular filtration rate (HR 0.911; P=.003), Charlson comorbidity index (HR 1.513; P<.0001) and tobacco smoking (HR 1.816; P=.003). 35% developed end-stage renal disease, and the best determinants (by competing-risk regression) were: initial glomerular filtration rate (sub-hazard ratio [SHR]: 0.791; P<.0001), proteinuria (SHR: 1.313; P<.0001), and smoking status (SHR: 1.848; P=.023). No differences were found in patients' mortality or renal survival between the different study periods. Prognosis of anti-neutrophil cytoplasm antibodies vasculitis with renal involvement treated with conventional immunosuppressive therapy remains unsatisfactory, and continues to have increased long-term complications and mortality. Copyright © 2016 Elsevier España, S.L.U. All rights reserved.

  10. A new cytoplasmic monoheme cytochrome c from Acidithiobacillus ferrooxidans involved in sulfur oxidation.

    Science.gov (United States)

    Liu, Yuandong; Guo, Shuhui; Yu, Runlan; Zou, Kai; Qiu, Guanzhou

    2014-03-01

    Acidithiobacillus ferrooxidans can obtain energy from the oxidation of various reduced inorganic sulfur compounds (RISCs, e.g., sulfur) and ferrous iron in bioleaching so has multiple branched respiratory pathways with a diverse range of electron transporters, especially cytochrome c proteins. A cytochrome c family gene, afe1130, which has never been reported before, was found by screening the whole genome of A. ferrooxidans. Here we report the differential gene transcription, bioinformatics analysis, and molecular modeling of the protein encoded by the afe1130 gene (AFE1130). The differential transcription of the target afe1130 gene versus the reference rrs gene in the A. ferrooxidans, respectively, on the culture conditions of sulfur and ferrous energy sources was performed through quantitative reverse transcription polymerase chain reaction (qRT-PCR) with a SYBR green-based assay according to the standard curves method. The qRT-PCR results showed that the afe1130 gene in sulfur culture condition was obviously more transcribed than that in ferrous culture condition. Bioinformatics analysis indicated that the AFE1130 was affiliated to the subclass ID of class I of cytochrome c and located in cytoplasm. Molecular modeling results exhibited that the AFE1130 protein consisted of 5 alpha-helices harboring one heme c group covalently bonded by Cys13 and Cys16 and ligated by His17 and Met62 and owned a big raised hydrophobic surface responsible for attaching to inner cytomembrane. So the AFE1130 in A. ferrooxidans plays a role in the RISCs oxidation in bioleaching in cytoplasm bound to inner membrane.

  11. Aspartate tightens the anchoring of staphylococcal lipoproteins to the cytoplasmic membrane.

    Science.gov (United States)

    Kumari, Nimerta; Götz, Friedrich; Nguyen, Minh-Thu

    2017-12-01

    In gram-negative bacteria, the ABC transporter LolCDE complex translocates outer membrane-specific lipoproteins (Lpp) from the inner membrane to the outer membrane. Lpp possessing aspartate (Asp) at position +2 are not translocated because it functions as a LolCDE avoidance signal. In gram-positive bacteria, lacking an outer membrane and the Lol system, Lpp are only anchored at the outer leaflet of the cytoplasmic membrane. However, the release of Lpp particularly in pathogenic or commensal species is crucial for immune modulation. Here, we provide evidence that in Staphylococcus aureus Asp at position +2 plays a role in withholding Lpp to the cytoplasmic membrane. Screening of published exoproteomic data of S. aureus revealed that Lpp mainly with Gly or Ser at position +2 were found in exoproteome, but there was no Lpp with Asp+2. The occurrence of Lpp with Asp+2 is infrequent in gram-positive bacteria. In S. aureus USA300 only seven of the 67 Lpp possess Asp+2; among them five Lpp represented Lpl lipoproteins involved in host cell invasion. Our study demonstrated that replacing the Asp+2 present in Lpl8 with a Ser enhances its release into the supernatant. However, there is no different release of Asp+2 and Ser+2 in mprF mutant that lacks the positive charge of lysyl-phosphatidylglycerol (Lys-PG). Moreover, substitution of Ser+2 by Asp in SitC (MntC) did not lead to a decreased release indicating that in staphylococci positions +3 and +4 might also be important for a tighter anchoring of Lpp. Here, we show that Asp in position +2 and adjacent amino acids contribute in tightening the anchoring of Lpp by interaction of the negative charged Asp with the positive charged Lys-PG. © 2017 The Authors. MicrobiologyOpen published by John Wiley & Sons Ltd.

  12. Production of bovine hand-made cloned embryos by zygote-oocyte cytoplasmic hemi-complementation.

    Science.gov (United States)

    Mezzalira, Joana Claudia; Ohlweiler, Lain Uriel; da Costa Gerger, Renato Pereira; Casali, Renata; Vieira, Fabiano Koerich; Ambrósio, Carlos Eduardo; Miglino, Maria Angélica; Rodrigues, José Luiz; Mezzalira, Alceu; Bertolini, Marcelo

    2011-02-01

    The aim of this study was to evaluate the effect of the cytoplast type and activation process on development of cloned embryos. Bovine oocytes (MII) or zygotes at the one-cell stage (IVF) were manually bisected and segregated in MII or IVF hemi-cytoplasts or hemi-karyoplasts. Adult skin cells from a bovine female were used as nucleus donors (SC). Experimental groups were composed of IVF embryos; parthenogenetic embryos; hand-made cloned (HMC) embryos; and reconstructed HMC embryos using IVF hemi-cytoplast + MII hemi-cytoplast + SC (G-I); IVF hemi-cytoplast + IVF hemi-cytoplast + SC (G-II); MII hemi-cytoplast + IVF hemi-karyoplast (G-III); and IVF hemi-cytoplast + IVF hemi-karyoplast (G-IV). Embryos from G-I to G-IV were allocated to subgroups as sperm-activated (SA) or were further chemically activated (SA + CA). Embryos from all groups and subgroups were in vitro cultured in the WOW system. Blastocyst development in subgroup G-I SA (28.2%) was similar to IVF (27.0%) and HMC (31.4%) controls, perhaps due to a to a more suitable activation process and/or better complementation of cytoplasmic reprogramming factors, with the other groups and subgroups having lower levels of development. No blastocyst development was observed when using IVF hemi-karyoplasts (G-III and G-IV), possibly due to the manipulation process during a sensitive biological period. In summary, the presence of cytoplasmic factors from MII hemi-oocytes and the sperm activation process from hemi-zygotes appear to be necessary for adequate in vitro development, as only the zygote-oocyte hemi-complementation was as efficient as controls for the generation of bovine cloned blastocysts.

  13. Cytoplasmic delivery and selective, multicomponent labeling with oligoarginine-linked protein tags.

    Science.gov (United States)

    Zou, Xiaoyan; Rajendran, Megha; Magda, Darren; Miller, Lawrence W

    2015-03-18

    Strategies that leverage bio-orthogonal interactions between small molecule ligands and genetically encoded amino acid sequences can be used to attach high-performance fluorophores to proteins in living cells. However, a major limitation of chemical protein labeling is that cells' plasma membranes are impermeable to many useful probes and biolabels. Here, we show that conjugation to nonaarginine, a cell penetrating peptide (CPP), enables passive cytoplasmic delivery of otherwise membrane-impermeant, small molecule protein labels. Heterodimers consisting of a luminescent Tb(3+) complex, Lumi4, linked to benzyl guanine, benzyl cytosine, and trimethoprim were conjugated to the peptide CysArg9 with a reducible disulfide linker. When added to culture medium, the peptide conjugates rapidly (cells. The benzyl guanine, benzyl cytosine, and trimethoprim derivatives bind selectively to fusion proteins tagged with SNAP-Tag, CLIP-Tag, and Escherichia coli dihydrofolate reductase (eDHFR), respectively. Furthermore, eDHFR and SNAP-Tag fusions can be labeled with Lumi4 analogues in the same cell, and this labeling can be detected using two-color, time-gated Förster resonance energy transfer (FRET) microscopy. Finally, we present quantitative data showing that cytoplasmic uptake of nonaarginine-conjugated probes occurs in multiple cell types (MDCK, HeLa, NIH 3T3), most cells in a culture (>75%) are loaded with probe, and the cellular probe concentration can be controlled by varying incubation conditions. CPP-mediated delivery of Lumi4-linked protein labels will greatly increase the utility of lanthanide-based FRET microscopy. Moreover, our results strongly suggest that this approach can be adapted to deliver a wide variety of protein-targeted fluorophores or other functional probes that were previously unavailable for intracellular imaging studies.

  14. Genetic Diversity and Structure of Lolium Species Surveyed on Nuclear Simple Sequence Repeat and Cytoplasmic Markers

    Directory of Open Access Journals (Sweden)

    Hongwei Cai

    2017-04-01

    Full Text Available To assess the genetic diversity and population structure of Lolium species, we used 32 nuclear simple sequence repeat (SSR markers and 7 cytoplasmic gene markers to analyze a total of 357 individuals from 162 accessions of 9 Lolium species. This survey revealed a high level of polymorphism, with an average number of alleles per locus of 23.59 and 5.29 and an average PIC-value of 0.83 and 0.54 for nuclear SSR markers and cytoplasmic gene markers, respectively. Analysis of molecular variance (AMOVA revealed that 16.27 and 16.53% of the total variation was due to differences among species, with the remaining 56.35 and 83.47% due to differences within species and 27.39 and 0% due to differences within individuals in 32 nuclear SSR markers set and 6 chloroplast gene markers set, respectively. The 32 nuclear SSR markers detected three subpopulations among 357 individuals, whereas the 6 chloroplast gene markers revealed three subpopulations among 160 accessions in the STRUCTURE analysis. In the clustering analysis, the three inbred species clustered into a single group, whereas the outbreeding species were clearly divided, especially according to nuclear SSR markers. In addition, almost all Lolium multiflorum populations were clustered into group C4, which could be further divided into three subgroups, whereas Lolium perenne populations primarily clustered into two groups (C2 and C3, with a few lines that instead grouped with L. multiflorum (C4 or Lolium rigidum (C6. Together, these results will useful for the use of Lolium germplasm for improvement and increase the effectiveness of ryegrass breeding.

  15. Coupled Ca2+/H+ transport by cytoplasmic buffers regulates local Ca2+ and H+ ion signaling.

    Science.gov (United States)

    Swietach, Pawel; Youm, Jae-Boum; Saegusa, Noriko; Leem, Chae-Hun; Spitzer, Kenneth W; Vaughan-Jones, Richard D

    2013-05-28

    Ca(2+) signaling regulates cell function. This is subject to modulation by H(+) ions that are universal end-products of metabolism. Due to slow diffusion and common buffers, changes in cytoplasmic [Ca(2+)] ([Ca(2+)]i) or [H(+)] ([H(+)]i) can become compartmentalized, leading potentially to complex spatial Ca(2+)/H(+) coupling. This was studied by fluorescence imaging of cardiac myocytes. An increase in [H(+)]i, produced by superfusion of acetate (salt of membrane-permeant weak acid), evoked a [Ca(2+)]i rise, independent of sarcolemmal Ca(2+) influx or release from mitochondria, sarcoplasmic reticulum, or acidic stores. Photolytic H(+) uncaging from 2-nitrobenzaldehyde also raised [Ca(2+)]i, and the yield was reduced following inhibition of glycolysis or mitochondrial respiration. H(+) uncaging into buffer mixtures in vitro demonstrated that Ca(2+) unloading from proteins, histidyl dipeptides (HDPs; e.g., carnosine), and ATP can underlie the H(+)-evoked [Ca(2+)]i rise. Raising [H(+)]i tonically at one end of a myocyte evoked a local [Ca(2+)]i rise in the acidic microdomain, which did not dissipate. The result is consistent with uphill Ca(2+) transport into the acidic zone via Ca(2+)/H(+) exchange on diffusible HDPs and ATP molecules, energized by the [H(+)]i gradient. Ca(2+) recruitment to a localized acid microdomain was greatly reduced during intracellular Mg(2+) overload or by ATP depletion, maneuvers that reduce the Ca(2+)-carrying capacity of HDPs. Cytoplasmic HDPs and ATP underlie spatial Ca(2+)/H(+) coupling in the cardiac myocyte by providing ion exchange and transport on common buffer sites. Given the abundance of cellular HDPs and ATP, spatial Ca(2+)/H(+) coupling is likely to be of general importance in cell signaling.

  16. Synthetic anti-endotoxin peptides inhibit cytoplasmic LPS-mediated responses.

    Science.gov (United States)

    Pfalzgraff, Anja; Heinbockel, Lena; Su, Qi; Brandenburg, Klaus; Weindl, Günther

    2017-09-15

    Toll-like receptor (TLR) 4-independent recognition of lipopolysaccharide (LPS) in the cytosol by inflammatory caspases leads to non-canonical inflammasome activation and induction of IL-1 secretion and pyroptosis. The discovery of this novel mechanism has potential implications for the development of effective drugs to treat sepsis since LPS-mediated hyperactivation of caspases is critically involved in endotoxic shock. Previously, we demonstrated that Pep19-2.5, a synthetic anti-endotoxin peptide, efficiently neutralises pathogenicity factors of Gram-negative and Gram-positive bacteria and protects against sepsis in vivo. Here, we report that Pep19-2.5 inhibits the effects of cytoplasmic LPS in human myeloid cells and keratinocytes. In THP-1 monocytes and macrophages, the peptide strongly reduced secretion of IL-1β and LDH induced by intracellular LPS. In contrast, the TLR4 signaling inhibitor TAK-242 abrogates LPS-induced TNF and IL-1β secretion, but not pyroptotic cell death. Furthermore, Pep19-2.5 suppressed LPS-induced HMGB-1 production and caspase-1 activation in THP-1 monocytes. Consistent with this observation, we found impaired IL-1β and IL-1α release in LPS-stimulated primary monocytes in the presence of Pep19-2.5 and reduced LDH release and IL-1B and IL-1A expression in LPS-transfected HaCaT keratinocytes. Additionally, Pep19-2.5 completely abolished IL-1β release induced by LPS/ATP in macrophages via canonical inflammasome activation. In conclusion, we provide evidence that anti-endotoxin peptides inhibit the inflammasome/IL-1 axis induced by cytoplasmic LPS sensing in myeloid cells and keratinocytes and activation of the classical inflammasome by LPS/ATP which may contribute to the protection against bacterial sepsis and skin infections with intracellular Gram-negative bacteria. Copyright © 2017 Elsevier Inc. All rights reserved.

  17. The desmin network is a determinant of the cytoplasmic stiffness of myoblasts.

    Science.gov (United States)

    Charrier, Elisabeth E; Montel, Lorraine; Asnacios, Atef; Delort, Florence; Vicart, Patrick; Gallet, François; Batonnet-Pichon, Sabrina; Hénon, Sylvie

    2018-02-01

    The mechanical properties of cells are essential to maintain their proper functions, and mainly rely on their cytoskeleton. A lot of attention has been paid to actin filaments, demonstrating their central role in the cells mechanical properties, but much less is known about the participation of intermediate filament (IF) networks. Indeed the contribution of IFs, such as vimentin, keratins and lamins, to cell mechanics has only been assessed recently. We study here the involvement of desmin, an IF specifically expressed in muscle cells, in the rheology of immature muscle cells. Desmin can carry mutations responsible for a class of muscle pathologies named desminopathies. In this study, using three types of cell rheometers, we assess the consequences of expressing wild-type (WT) or mutated desmin on the rheological properties of single myoblasts. We find that the mechanical properties of the cell cortex are not correlated to the quantity, nor the quality of desmin expressed. On the contrary, the overall cell stiffness increases when the amount of WT or mutated desmin polymerised in cytoplasmic networks increases. However, myoblasts become softer when the desmin network is partially depleted by the formation of aggregates induced by the expression of a desmin mutant. We demonstrate that desmin plays a negligible role in the mechanical properties of the cell cortex but is a determinant of the overall cell stiffness. More particularly, desmin participates to the cytoplasm viscoelasticity. Desminopathies are associated with muscular weaknesses attributed to a disorganisation of the structure of striated muscle that impairs the active force generation. The present study evidences for the first time the key role of desmin in the rheological properties of myoblasts, raising the hypothesis that desmin mutations could also alter the passive mechanical properties of muscles, thus participating to the lack of force build up in muscle tissue. © 2018 Société Française des

  18. Phase-Separated Liposomes Enhance the Efficiency of Macromolecular Delivery to the Cellular Cytoplasm.

    Science.gov (United States)

    Imam, Zachary I; Kenyon, Laura E; Ashby, Grant; Nagib, Fatema; Mendicino, Morgan; Zhao, Chi; Gadok, Avinash K; Stachowiak, Jeanne C

    2017-10-01

    From viruses to organelles, fusion of biological membranes is used by diverse biological systems to deliver macromolecules across membrane barriers. Membrane fusion is also a potentially efficient mechanism for the delivery of macromolecular therapeutics to the cellular cytoplasm. However, a key shortcoming of existing fusogenic liposomal systems is that they are inefficient, requiring a high concentration of fusion-promoting lipids in order to cross cellular membrane barriers. Toward addressing this limitation, our experiments explore the extent to which membrane fusion can be amplified by using the process of lipid membrane phase separation to concentrate fusion-promoting lipids within distinct regions of the membrane surface. We used confocal fluorescence microscopy to investigate the integration of fusion-promoting lipids into a ternary lipid membrane system that separated into liquid-ordered and liquid-disordered membrane phases. Additionally, we quantified the impact of membrane phase separation on the efficiency with which liposomes transferred lipids and encapsulated macromolecules to cells, using a combination of confocal fluorescence imaging and flow cytometry. Here we report that concentrating fusion-promoting lipids within phase-separated lipid domains on the surfaces of liposomes significantly increases the efficiency of liposome fusion with model membranes and cells. In particular, membrane phase separation enhanced the delivery of lipids and model macromolecules to the cytoplasm of tumor cells by at least 4-fold in comparison to homogenous liposomes. Our findings demonstrate that phase separation can enhance membrane fusion by locally concentrating fusion-promoting lipids on the surface of liposomes. This work represents the first application of lipid membrane phase separation in the design of biomaterials-based delivery systems. Additionally, these results lay the ground work for developing fusogenic liposomes that are triggered by physical and

  19. Frequent Nuclear/Cytoplasmic Localization of β-Catenin without Exon 3 Mutations in Malignant Melanoma

    Science.gov (United States)

    Rimm, David L.; Caca, Karel; Hu, Gang; Harrison, Frank B.; Fearon, Eric R.

    1999-01-01

    β-Catenin has a critical role in E-cadherin-mediated cell-cell adhesion, and it also functions as a downstream signaling molecule in the wnt pathway. Mutations in the putative glycogen synthase kinase 3β phosphorylation sites near the β-catenin amino terminus have been found in some cancers and cancer cell lines. The mutations render β-catenin resistant to regulation by a complex containing the glycogen synthase kinase 3β, adenomatous polyposis coli, and axin proteins. As a result, β-catenin accumulates in the cytosol and nucleus and activates T-cell factor/lymphoid enhancing factor transcription factors. Previously, 6 of 27 melanoma cell lines were found to have β-catenin exon 3 mutations affecting the N-terminal phosphorylation sites (Rubinfeld B, Robbins P, Elgamil M, Albert I, Porfiri E, Polakis P: Stabilization of beta-catenin by genetic defects in melanoma cell lines. Science 1997, 275:1790–1792). To assess the role of β-catenin defects in primary melanomas, we undertook immunohistochemical and DNA sequencing studies in 65 melanoma specimens. Nuclear and/or cytoplasmic localization of β-catenin, a potential indicator of wnt pathway activation, was seen focally within roughly one third of the tumors, though a clonal somatic mutation in β-catenin was found in only one case (codon 45 Ser→Pro). Our findings demonstrate that β-catenin mutations are rare in primary melanoma, in contrast to the situation in melanoma cell lines. Nonetheless, activation of β-catenin, as indicated by its nuclear and/or cytoplasmic localization, appears to be frequent in melanoma, and in some cases, it may reflect focal and transient activation of the wnt pathway within the tumor. PMID:10027390

  20. Organelle Simple Sequence Repeat Markers Help to Distinguish Carpelloid Stamen and Normal Cytoplasmic Male Sterile Sources in Broccoli

    Science.gov (United States)

    Shu, Jinshuai; Liu, Yumei; Li, Zhansheng; Zhang, Lili; Fang, Zhiyuan; Yang, Limei; Zhuang, Mu; Zhang, Yangyong; Lv, Honghao

    2015-01-01

    We previously discovered carpelloid stamens when breeding cytoplasmic male sterile lines in broccoli (Brassica oleracea var. italica). In this study, hybrids and multiple backcrosses were produced from different cytoplasmic male sterile carpelloid stamen sources and maintainer lines. Carpelloid stamens caused dysplasia of the flower structure and led to hooked or coiled siliques with poor seed setting, which were inherited in a maternal fashion. Using four distinct carpelloid stamens and twelve distinct normal stamens from cytoplasmic male sterile sources and one maintainer, we used 21 mitochondrial simple sequence repeat (mtSSR) primers and 32 chloroplast SSR primers to identify a mitochondrial marker, mtSSR2, that can differentiate between the cytoplasm of carpelloid and normal stamens. Thereafter, mtSSR2 was used to identify another 34 broccoli accessions, with an accuracy rate of 100%. Analysis of the polymorphic sequences revealed that the mtSSR2 open reading frame of carpelloid stamen sterile sources had a deletion of 51 bases (encoding 18 amino acids) compared with normal stamen materials. The open reading frame is located in the coding region of orf125 and orf108 of the mitochondrial genomes in Brassica crops and had the highest similarity with Raphanus sativus and Brassica carinata. The current study has not only identified a useful molecular marker to detect the cytoplasm of carpelloid stamens during broccoli breeding, but it also provides evidence that the mitochondrial genome is maternally inherited and provides a basis for studying the effect of the cytoplasm on flower organ development in plants. PMID:26407159

  1. Multimerization of the cytoplasmic domain of syndecan-4 is required for its ability to activate protein kinase C

    DEFF Research Database (Denmark)

    Oh, E S; Woods, A; Couchman, J R

    1997-01-01

    The transmembrane proteoglycan syndecan-4, which is a coreceptor with integrins in cytoskeleton-matrix interactions, appears to be multimerized in vivo. Both purified and recombinant core proteins form sodium dodecyl sulfate-resistant oligomers, and we now report that a synthetic peptide......, potentiated the activity of PKCalpha, and only oligomeric syndecan-4 cytoplasmic peptides were active. Changes in peptide sequence caused parallel loss of stable oligomeric status and ability to regulate a mixture of PKCalphabetagamma activity. A synthetic peptide encompassing the whole cytoplasmic domain...

  2. Preventive analgesia

    DEFF Research Database (Denmark)

    Dahl, Jørgen B; Kehlet, Henrik

    2011-01-01

    This paper will discuss the concepts of pre-emptive and preventive analgesia in acute and persistent postsurgical pain, based on the most recent experimental and clinical literature, with a special focus on injury-induced central sensitization and the development from acute to chronic pain. Recent...... of preventive analgesia for persistent postoperative pain are promising. However, clinicians must be aware of the demands for improved design of their clinical studies in order to get more conclusive answers regarding the different avenues for intervention. Summary: The concept of preventive analgesia is still...

  3. Cytoplasmic fungal lipases release fungicides from ultra-deformable vesicular drug carriers.

    Science.gov (United States)

    Steinberg, Gero

    2012-01-01

    The Transfersome® is a lipid vesicle that contains membrane softeners, such as Tween 80, to make it ultra-deformable. This feature makes the Transfersome® an efficient carrier for delivery of therapeutic drugs across the skin barrier. It was reported that TDT 067 (a topical formulation of 15 mg/ml terbinafine in Transfersome® vesicles) has a much more potent antifungal activity in vitro compared with conventional terbinafine, which is a water-insoluble fungicide. Here we use ultra-structural studies and live imaging in a model fungus to describe the underlying mode of action. We show that terbinafine causes local collapse of the fungal endoplasmic reticulum, which was more efficient when terbinafine was delivered in Transfersome® vesicles (TFVs). When applied in liquid culture, fluorescently labeled TFVs rapidly entered the fungal cells (T(1/2)~2 min). Entry was F-actin- and ATP-independent, indicating that it is a passive process. Ultra-structural studies showed that passage through the cell wall involves significant deformation of the vesicles, and depends on a high concentration of the surfactant Tween 80 in their membrane. Surprisingly, the TFVs collapsed into lipid droplets after entry into the cell and the terbinafine was released from their interior. With time, the lipid bodies were metabolized in an ATP-dependent fashion, suggesting that cytosolic lipases attack and degrade intruding TFVs. Indeed, the specific monoacylglycerol lipase inhibitor URB602 prevented Transfersome® degradation and neutralized the cytotoxic effect of Transfersome®-delivered terbinafine. These data suggest that (a) Transfersomes deliver the lipophilic fungicide Terbinafine to the fungal cell wall, (b) the membrane softener Tween 80 allows the passage of the Transfersomes into the fungal cell, and (c) fungal lipases digest the invading Transfersome® vesicles thereby releasing their cytotoxic content. As this mode of action of Transfersomes is independent of the drug cargo

  4. Cytoplasmic fungal lipases release fungicides from ultra-deformable vesicular drug carriers.

    Directory of Open Access Journals (Sweden)

    Gero Steinberg

    Full Text Available The Transfersome® is a lipid vesicle that contains membrane softeners, such as Tween 80, to make it ultra-deformable. This feature makes the Transfersome® an efficient carrier for delivery of therapeutic drugs across the skin barrier. It was reported that TDT 067 (a topical formulation of 15 mg/ml terbinafine in Transfersome® vesicles has a much more potent antifungal activity in vitro compared with conventional terbinafine, which is a water-insoluble fungicide. Here we use ultra-structural studies and live imaging in a model fungus to describe the underlying mode of action. We show that terbinafine causes local collapse of the fungal endoplasmic reticulum, which was more efficient when terbinafine was delivered in Transfersome® vesicles (TFVs. When applied in liquid culture, fluorescently labeled TFVs rapidly entered the fungal cells (T(1/2~2 min. Entry was F-actin- and ATP-independent, indicating that it is a passive process. Ultra-structural studies showed that passage through the cell wall involves significant deformation of the vesicles, and depends on a high concentration of the surfactant Tween 80 in their membrane. Surprisingly, the TFVs collapsed into lipid droplets after entry into the cell and the terbinafine was released from their interior. With time, the lipid bodies were metabolized in an ATP-dependent fashion, suggesting that cytosolic lipases attack and degrade intruding TFVs. Indeed, the specific monoacylglycerol lipase inhibitor URB602 prevented Transfersome® degradation and neutralized the cytotoxic effect of Transfersome®-delivered terbinafine. These data suggest that (a Transfersomes deliver the lipophilic fungicide Terbinafine to the fungal cell wall, (b the membrane softener Tween 80 allows the passage of the Transfersomes into the fungal cell, and (c fungal lipases digest the invading Transfersome® vesicles thereby releasing their cytotoxic content. As this mode of action of Transfersomes is independent of the

  5. Mutations in the Transmembrane Domain and Cytoplasmic Tail of Hendra Virus Fusion Protein Disrupt Virus-Like-Particle Assembly.

    Science.gov (United States)

    Cifuentes-Muñoz, Nicolás; Sun, Weina; Ray, Greeshma; Schmitt, Phuong Tieu; Webb, Stacy; Gibson, Kathleen; Dutch, Rebecca Ellis; Schmitt, Anthony P

    2017-07-15

    Hendra virus (HeV) is a zoonotic paramyxovirus that causes deadly illness in horses and humans. An intriguing feature of HeV is the utilization of endosomal protease for activation of the viral fusion protein (F). Here we investigated how endosomal F trafficking affects HeV assembly. We found that the HeV matrix (M) and F proteins each induced particle release when they were expressed alone but that their coexpression led to coordinated assembly of virus-like particles (VLPs) that were morphologically and physically distinct from M-only or F-only VLPs. Mutations to the F protein transmembrane domain or cytoplasmic tail that disrupted endocytic trafficking led to failure of F to function with M for VLP assembly. Wild-type F functioned normally for VLP assembly even when its cleavage was prevented with a cathepsin inhibitor, indicating that it is endocytic F trafficking that is important for VLP assembly, not proteolytic F cleavage. Under specific conditions of reduced M expression, we found that M could no longer induce significant VLP release but retained the ability to be incorporated as a passenger into F-driven VLPs, provided that the F protein was competent for endocytic trafficking. The F and M proteins were both found to traffic through Rab11-positive recycling endosomes (REs), suggesting a model in which F and M trafficking pathways converge at REs, enabling these proteins to preassemble before arriving at plasma membrane budding sites. IMPORTANCE Hendra virus and Nipah virus are zoonotic paramyxoviruses that cause lethal infections in humans. Unlike that for most paramyxoviruses, activation of the henipavirus fusion protein occurs in recycling endosomal compartments. In this study, we demonstrate that the unique endocytic trafficking pathway of Hendra virus F protein is required for proper viral assembly and particle release. These results advance our basic understanding of the henipavirus assembly process and provide a novel model for the interplay between

  6. Wolbachia density and cytoplasmic incompatibility in Aedes albopictus: concerns with using artificial Wolbachia infection as a vector suppression tool.

    Directory of Open Access Journals (Sweden)

    Maurizio Calvitti

    Full Text Available The mosquito Aedes albopictusi is a competent vector of harmful human pathogens, including viruses causing dengue and chikungunya. Cytoplasmic incompatibility (CI induced by endosymbiotic Wolbachia can be used to produce functionally sterile males that can be released in the field as a suppression tool against this mosquito. Because the available sexing methods are not efficient enough to avoid unintentional release of a few transinfected females, we assessed the CI pattern in crosses between wPip Wolbachia-transinfected (ARwP females and wild-type males of Ae. albopictus in this study. Quantitative polymerase chain reaction was used to monitor the titer of the Wolbachia strains that naturally infect Ae. albopictus, that is, wAlbA and wAlbB, in age-controlled males and females. Data were coupled with incompatibility level detected when the above-mentioned males were crossed with ARwP females. Wolbachia infection titer was also monitored in samples of wild caught males. Incompatibility level was positively correlated only with wAlbA density. Crosses between wild-type males having very low wAlbA density (<0.001 wAlbA/actin copy numbers and ARwP females were partially fertile (CIcorr = 68.06 ± 6.20. Individuals with low wAlbA titer were frequently found among sampled wild males (30%-50% depending on the site and period. ARwP males can be as considered as a very promising tool for suppressing Ae. albopictus. However, crosses between wild males having low wAlbA density and ARwP females may be partially fertile. In the case of local establishment of the transinfected mosquito line, this occurrence may favor the replacement of the wild-type mosquitoes with the ARwP line, thus reducing the long-term efficacy of incompatible insect technique. Various alternative strategies have been discussed to prevent this risk and to exploit Wolbachia as a tool to control Ae. albopictus.

  7. Prevention: Exercise

    Medline Plus

    Full Text Available ... Exercise Strength Training for the Elderly Other Back Pack Safety Pregnancy and Back Pain Preventing Osteoporosis Back ... in very slightly. Hold a ball directly in front of you. Keep your abdominal muscles tight and ...

  8. Prevention: Exercise

    Medline Plus

    Full Text Available ... Tips for a Healthy Back Smoking Weight Patient Safety Exercise Strengthening Strengthen Your Core! Stretching/Flexibility Aerobic ... Strength Training for the Elderly Other Back Pack Safety Pregnancy and Back Pain Preventing Osteoporosis Back Pain ...

  9. Prevent Shingles

    Science.gov (United States)

    ... Submit What's this? Submit Button Past Emails Prevent Shingles Language: English (US) Español (Spanish) Recommend on Facebook ... that can result in vision loss. Older Adults & Shingles As you get older, you are more likely ...

  10. Prevention: Exercise

    Medline Plus

    Full Text Available ... Epidural Steroid Injections Lumbar Zygapophysical (Facet) Joint Injections PREVENTION Lifestyle Choices 10 Tips for a Healthy Back Smoking Weight Patient Safety Exercise Strengthening Strengthen Your Core! ...

  11. Prevention: Exercise

    Medline Plus

    Full Text Available ... and Education Injection Treatments for Spinal Pain Epidural Steroid Injections Lumbar Zygapophysical (Facet) Joint Injections PREVENTION Lifestyle ... Z Spine Specialists Videos 9 for Spine Epidural Steroid Injections Exercise: The Backbone of Spine Treatment Spondylolisthesis ...

  12. Prevention: Exercise

    Medline Plus

    Full Text Available ... A SPECIALIST Prevention Strengthening Exercise Committee Exercise Committee Core Strengthening Many popular forms of exercise focus on ... acute pain, you should stop doing it. Transverse Core Strengthening This strengthens the muscles that cross from ...

  13. Prevention: Exercise

    Medline Plus

    Full Text Available ... Lumbar Zygapophysical (Facet) Joint Injections PREVENTION Lifestyle Choices 10 Tips for a Healthy Back Smoking Weight Patient ... the floor; rotate from side to side. Repeat 10 times. Check with your physician; if you are ...

  14. Prevention: Exercise

    Medline Plus

    Full Text Available ... Pregnancy and Back Pain Preventing Osteoporosis Back Pain Basics Book RESOURCES Patient ... popular forms of exercise focus on core strengthening, or building the muscles that provide support for your body. Pilates, yoga and martial arts ...

  15. HIV Prevention

    Centers for Disease Control (CDC) Podcasts

    2012-02-01

    Dr. Kevin Fenton, Director of CDC’s National Center for HIV/AIDS, Viral Hepatitis, STD, and TB Prevention, talks about steps people can take to protect their health from HIV.  Created: 2/1/2012 by National Center for HIV/AIDS, Viral Hepatitis, STD, and TB Prevention (NCHHSTP).   Date Released: 2/1/2012.

  16. Successful Pregnancy in a Couple with Severe Male Factor Infertility after Selection of Sperm with Cytoplasmic Droplets

    OpenAIRE

    Bellish, Jenna; McCulloh, David H.; Ahmad, Khaliq; McGovern, Peter G.

    2015-01-01

    We present live births resulting from two separate IVF cycles in a couple in which ICSI was performed with sperm specifically selected for presence of small cytoplasmic droplets. These cycles followed previous cycles using standard sperm selection methods in which very poor embryo development and no pregnancies ensued. The male partner was diagnosed with severe male factor infertility including elevated DNA fragmentation.

  17. Empagliflozin decreases myocardial cytoplasmic Na+ through inhibition of the cardiac Na+/H+ exchanger in rats and rabbits

    NARCIS (Netherlands)

    Baartscheer, Antonius; Schumacher, Cees A.; Wüst, Rob C.I.; Fiolet, Jan W.T.; Stienen, Ger J.M.; Coronel, Ruben; Zuurbier, Coert J.

    2017-01-01

    Aims/hypothesis: Empagliflozin (EMPA), an inhibitor of the renal sodium–glucose cotransporter (SGLT) 2, reduces the risk of cardiovascular death in patients with type 2 diabetes. The underlying mechanism of this effect is unknown. Elevated cardiac cytoplasmic Na+ ([Na+]c) and Ca2+ ([Ca2+]c)

  18. Empagliflozin decreases myocardial cytoplasmic Na+ through inhibition of the cardiac Na+/H+ exchanger in rats and rabbits

    NARCIS (Netherlands)

    Baartscheer, Antonius; Schumacher, Cees A.; Wüst, Rob C. I.; Fiolet, Jan W. T.; Stienen, Ger J. M.; Coronel, Ruben; Zuurbier, Coert J.

    2017-01-01

    Empagliflozin (EMPA), an inhibitor of the renal sodium-glucose cotransporter (SGLT) 2, reduces the risk of cardiovascular death in patients with type 2 diabetes. The underlying mechanism of this effect is unknown. Elevated cardiac cytoplasmic Na+ ([Na+](c)) and Ca2+ ([Ca2+](c)) concentrations and

  19. Cyclin D1 localizes in the cytoplasm of keratinocytes during skin differentiation and regulates cell–matrix adhesion

    Science.gov (United States)

    Fernández-Hernández, Rita; Rafel, Marta; Fusté, Noel P; Aguayo, Rafael S; Casanova, Josep M; Egea, Joaquim; Ferrezuelo, Francisco; Garí, Eloi

    2013-01-01

    The function of Cyclin D1 (CycD1) has been widely studied in the cell nucleus as a regulatory subunit of the cyclin-dependent kinases Cdk4/6 involved in the control of proliferation and development in mammals. CycD1 has been also localized in the cytoplasm, where its function nevertheless is poorly characterized. In this work we have observed that in normal skin as well as in primary cultures of human keratinocytes, cytoplasmic localization of CycD1 correlated with the degree of differentiation of the keratinocyte. In these conditions, CycD1 co-localized in cytoplasmic foci with exocyst components (Sec6) and regulators (RalA), and with β1 integrin, suggesting a role for CycD1 in the regulation of keratinocyte adhesion during differentiation. Consistent with this hypothesis, CycD1 overexpression increased β1 integrin recycling and drastically reduced the ability of keratinocytes to adhere to the extracellular matrix. We propose that localization of CycD1 in the cytoplasm during skin differentiation could be related to the changes in detachment ability of keratinocytes committed to differentiation. PMID:23839032

  20. The nectin-1α transmembrane domain, but not the cytoplasmic tail, influences cell fusion induced by HSV-1 glycoproteins

    International Nuclear Information System (INIS)

    Subramanian, Ravi P.; Dunn, Jennifer E.; Geraghty, Robert J.

    2005-01-01

    Nectin-1 is a receptor for herpes simplex virus (HSV), a member of the immunoglobulin superfamily, and a cellular adhesion molecule. To study domains of nectin-1α involved in cell fusion, we measured the ability of nectin-1α/nectin-2α chimeras, nectin-1α/CD4 chimeras, and transmembrane domain and cytoplasmic tail mutants of nectin-1α to promote cell fusion induced by HSV-1 glycoproteins. Our results demonstrate that only chimeras and mutants containing the entire V-like domain and a link to the plasma membrane conferred cell-fusion activity. The transmembrane domain and cytoplasmic tail of nectin-1 were not required for any viral receptor or cell adhesion function tested. Cellular cytoplasmic factors that bind to the nectin-1α cytoplasmic tail, therefore, did not influence virus entry or cell fusion. Interestingly, the efficiency of cell fusion was reduced when membrane-spanning domains of nectin-1α and gD were replaced by glycosylphosphatidylinositol tethers, indicating that transmembrane domains may play a modulatory role in the gD/nectin-1α interaction in fusion

  1. H-Ras activation promotes cytoplasmic accumulation and phosphoinositide 3-OH kinase association of beta-catenin in epidermal keratinocytes

    DEFF Research Database (Denmark)

    Espada, J; Pérez-Moreno, M; Braga, V M

    1999-01-01

    keratinocytes. Microinjection or stable expression of V12Ras into keratinocytes promotes the loss of E-cadherin and alpha-catenin and relocalization of beta-catenin to the cytoplasm and nucleus. Moreover, these effects are dependent on PI3K (phosphoinositide 3-OH kinase) activity. Interestingly, a strong...

  2. Higher cytoplasmic and nuclear poly(ADP-ribose) polymerase expression in familial than in sporadic breast cancer

    NARCIS (Netherlands)

    Klauke, M.L.; Hoogerbrugge-van der Linden, N.; Budczies, J.; Bult, P.; Prinzler, J.; Radke, C.; van Krieken, J.H.; Dietel, M.; Denkert, C.; Muller, B.M.

    2012-01-01

    Poly(ADP-ribose) polymerase 1 (PARP) is a key element of the single-base excision pathway for repair of DNA single-strand breaks. To compare the cytoplasmic and nuclear poly(ADP-ribose) expression between familial (BRCA1, BRCA2, or non BRCA1/2) and sporadic breast cancer, we investigated 39 sporadic

  3. Role of membrane-associated and cytoplasmic fatty acid-binding proteins in cellular fatty acid metabolism

    NARCIS (Netherlands)

    Glatz, J. F.; van Nieuwenhoven, F. A.; Luiken, J. J.; Schaap, F. G.; van der Vusse, G. J.

    1997-01-01

    A number of membrane-associated and cytoplasmic fatty acid-binding proteins (FABPs) are now being implicated in the cellular uptake and intracellular transport of long-chain fatty acids (FA). These proteins each have the capacity of non-covalent binding of FA, are present in tissues actively

  4. Cytoplasmic and Genomic Effects on Meiotic Pairing in Brassica Hybrids and Allotetraploids from Pair Crosses of Three Cultivated Diploids

    Science.gov (United States)

    Cui, Cheng; Ge, Xianhong; Gautam, Mayank; Kang, Lei; Li, Zaiyun

    2012-01-01

    Interspecific hybridization and allopolyploidization contribute to the origin of many important crops. Synthetic Brassica is a widely used model for the study of genetic recombination and “fixed heterosis” in allopolyploids. To investigate the effects of the cytoplasm and genome combinations on meiotic recombination, we produced digenomic diploid and triploid hybrids and trigenomic triploid hybrids from the reciprocal crosses of three Brassica diploids (B. rapa, AA; B. nigra, BB; B. oleracea, CC). The chromosomes in the resultant hybrids were doubled to obtain three allotetraploids (B. juncea, AA.BB; B. napus, AA.CC; B. carinata, BB.CC). Intra- and intergenomic chromosome pairings in these hybrids were quantified using genomic in situ hybridization and BAC-FISH. The level of intra- and intergenomic pairings varied significantly, depending on the genome combinations and the cytoplasmic background and/or their interaction. The extent of intragenomic pairing was less than that of intergenomic pairing within each genome. The extent of pairing variations within the B genome was less than that within the A and C genomes, each of which had a similar extent of pairing. Synthetic allotetraploids exhibited nondiploidized meiotic behavior, and their chromosomal instabilities were correlated with the relationship of the genomes and cytoplasmic background. Our results highlight the specific roles of the cytoplasm and genome to the chromosomal behaviors of hybrids and allopolyploids. PMID:22505621

  5. Conformational alterations resulting from mutations in cytoplasmic domains of the alpha subunit of the Na,K-ATPase

    DEFF Research Database (Denmark)

    Blostein, R; Daly, S E; MacAulay, Nanna

    1998-01-01

    This paper summarizes experiments concerned with the functional consequences of mutations in cytoplasmic regions of the alpha 1 subunit of the Na,K-ATPase, in particular the amino terminus, the first cytoplasmic loop between transmembrane segments M2 and M3, and the major cytoplasmic loop between M......4 and M5. In the first mutation (alpha 1M32), 32 residues were removed from the N-terminus. The second mutation (E233K) was in the putative beta strand of M2-M3 loop and the third, comprised the replacement of the amino terminal half of loop M4-M5 of the Na,K-ATPase with the homologous segment...... suggesting that changes in apparent affinity for ATP are secondary to changes in K+ ligation. The question of cation selectivity, in particular that of Na+ versus protons, has been addressed in structure/function analysis of a cytoplasmic chimera involving the M4-M5 loop. Transport studies performed...

  6. CRISPR/Cas9-mediated mutation revealed cytoplasmic tail is dispensable for IZUMO1 function and male fertility

    DEFF Research Database (Denmark)

    Young, Samantha A M; Miyata, Haruhiko; Satouh, Yuhkoh

    2016-01-01

    manipulation system of CRISPR/Cas9 to generate a point mutation resulting in a premature stop codon, producing mice with truncated IZUMO1. Mice without the cytoplasmic tail of IZUMO1 showed normal fertility but decreased the amount of protein, indicating that whilst this region is important for the expression...

  7. Synthesis, processing and export of cytoplasmic endo-ß-1,4-xylanase from barley aleurone during germination

    NARCIS (Netherlands)

    Caspers, M.P.M.; Lok, F.; Sinjorgo, K.M.C.; Zeijl, M. van; Nielsen, K.A.; Cameron-Mills, V.

    2001-01-01

    We have identified the major endo-β-l,4-xylanase (XYN-1) in the aleurone of germinating barley grain, and show that it is expressed as a precursor of Mr 61 500 with both N- and C-terminal propeptides. XYN-1 is synthesized as an inactive enzyme in the cytoplasm, and only becomes active at a late

  8. Differential detection of cytoplasmic Wilms tumor 1 expression by immunohistochemistry, western blotting and mRNA quantification.

    Science.gov (United States)

    Maki, Takehiro; Ikeda, Hiroaki; Kuroda, Aki; Kyogoku, Noriaki; Yamamura, Yoshiyuki; Tabata, Yukiko; Abiko, Takehiro; Tsuchikawa, Takahiro; Hida, Yasuhiro; Shichinohe, Toshiaki; Tanaka, Eiichi; Kaga, Kichizo; Hatanaka, Kanako; Matsuno, Yoshihiro; Imai, Naoko; Hirano, Satoshi

    2017-01-01

    Wilms tumor 1 (WT1) is considered to be a promising target of cancer treatment because it has been reported to be frequently expressed at high levels in various malignancies. Although WT1-targeted cancer treatment has been initiated, conclusive detection methods for WT1 are not established. The present study aimed to consolidate immunohistochemistry for WT1 with statistical basis. Transfected cells with forced WT1 expression yielded specific western blot bands and nuclear immunostaining; cytoplasmic immunostaining was not specifically recognized. Immunohistochemistry, western blotting, and quantitative reverse transcriptase-polymerase chain reaction were performed in 35 human cell lines using multiple WT1 antibodies and their results were quantified. Relationships among the quantified results were statistically analyzed; the nuclear immunostaining positively correlated with western blot bands and mRNA expression levels, whereas cytoplasmic immunostaining did not. These results indicate that nuclear immunostaining reflects WT1 expression but cytoplasmic immunostaining does not. The nuclear immunostaining was barely (3/541) observed in primary cancer of esophagus, bile duct, pancreas and lung. Although the present study has some limitations, the results indicate that the cytoplasmic immunostaining does not correlate with actual WT1 expression and prompts researchers to carefully evaluate target molecule expression in treatment of cancer.

  9. Effect of HIV-1 envelope cytoplasmic tail on adenovirus primed virus encoded virus-like particle immunizations

    DEFF Research Database (Denmark)

    Andersson, Anne Marie C; Ragonnaud, Emeline; Seaton, Kelly E.

    2016-01-01

    cytoplasmic tail on epitope presentation and subsequent immune responses, which is relevant for the interpretation of current clinical trials that are using truncated Env as an immunogen. The regimens described here provide similar neutralization titers, and thus are useful for investigating the importance...

  10. Syndecan-4 proteoglycan cytoplasmic domain and phosphatidylinositol 4,5-bisphosphate coordinately regulate protein kinase C activity

    DEFF Research Database (Denmark)

    Oh, E S; Woods, A; Lim, S T

    1998-01-01

    adhesions and actin stress fibers. The cytoplasmic domain of syndecan-4 core protein directly interacts with and potentiates PKCalpha activity, and it can directly interact with the phos- phoinositide PIP2. We, therefore, investigated whether the interaction of inositol phosphates and inositol phospholipids...

  11. DGCR8 Localizes to the Nucleus as well as Cytoplasmic Structures in Mammalian Spermatogenic Cells and Epididymal Sperm

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    Akane Nakano

    2013-01-01

    Full Text Available The localization of DGCR8 in spermatogenic cells and sperm from rat and mouse was studied by immunofluorescence and immunoelectron microscopy. Spermatogenic cells from these species yielded similar DGCR8 localization pattern. Immunofluorescence microscopy results showed that DGCR8 localized to both the cytoplasm and nucleus. In the cytoplasm, diffuse cytosolic and discrete granular staining was observed. Dual staining showed that DGCR8 colocalized to the granules with MAEL (a nuage marker. In the nucleus of spermatocytes, both the nucleoli and nucleoplasm were stained, whereas in the nucleus of early spermatids small spots were stained. In late spermatids, DGCR8 localized to the tip of their head and to small granules (neck granules of the neck cytoplasm. The neck granules were also observed in the neck of epididymal sperm. Immunoelectron microscopy results showed that DGCR8 localized to nuage structures. Moreover, DGCR8 localized to nonnuage structures in late spermatids. DGCR8 also localized to the nucleolus and euchromatin in spermatocytes and round spermatids and to small granules in the nucleus of late spermatids. The results suggest that in spermatogenic cells DGCR8 localizes not only to the nuclei but also to the cytoplasmic structures such as nuage and nonnuage structures. Furthermore, DGCR8 seems to be imported into the egg with neck granules in sperm during fertilization.

  12. Cytoplasmic free Ca2+ is essential for multiple steps in malaria parasite egress from infected erythrocytes

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    Glushakova Svetlana

    2013-01-01

    Full Text Available Abstract Background Egress of Plasmodium falciparum, from erythrocytes at the end of its asexual cycle and subsequent parasite invasion into new host cells, is responsible for parasite dissemination in the human body. The egress pathway is emerging as a coordinated multistep programme that extends in time for tens of minutes, ending with rapid parasite extrusion from erythrocytes. While the Ca2+ regulation of the invasion of P. falciparum in erythrocytes is well established, the role of Ca2+ in parasite egress is poorly understood. This study analysed the involvement of cytoplasmic free Ca2+ in infected erythrocytes during the multistep egress programme of malaria parasites. Methods Live-cell fluorescence microscopy was used to image parasite egress from infected erythrocytes, assessing the effect of drugs modulating Ca2+ homeostasis on the egress programme. Results A steady increase in cytoplasmic free Ca2+ is found to precede parasite egress. This increase is independent of extracellular Ca2+ for at least the last two hours of the cycle, but is dependent upon Ca2+ release from internal stores. Intracellular BAPTA chelation of Ca2+ within the last 45 minutes of the cycle inhibits egress prior to parasitophorous vacuole swelling and erythrocyte membrane poration, two characteristic morphological transformations preceding parasite egress. Inhibitors of the parasite endoplasmic reticulum (ER Ca2+-ATPase accelerate parasite egress, indicating that Ca2+ stores within the ER are sufficient in supporting egress. Markedly accelerated egress of apparently viable parasites was achieved in mature schizonts using Ca2+ ionophore A23187. Ionophore treatment overcomes the BAPTA-induced block of parasite egress, confirming that free Ca2+ is essential in egress initiation. Ionophore treatment of immature schizonts had an adverse effect inducing parasitophorous vacuole swelling and killing the parasites within the host cell. Conclusions The parasite egress

  13. Cytoplasmic compartmentalization of the fetal piRNA pathway in mice.

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    Alexei A Aravin

    2009-12-01

    Full Text Available Derepression of transposable elements (TEs in the course of epigenetic reprogramming of the mouse embryonic germline necessitates the existence of a robust defense that is comprised of PIWI/piRNA pathway and de novo DNA methylation machinery. To gain further insight into biogenesis and function of piRNAs, we studied the intracellular localization of piRNA pathway components and used the combination of genetic, molecular, and cell biological approaches to examine the performance of the piRNA pathway in germ cells of mice lacking Maelstrom (MAEL, an evolutionarily conserved protein implicated in transposon silencing in fruit flies and mice. Here we show that principal components of the fetal piRNA pathway, MILI and MIWI2 proteins, localize to two distinct types of germinal cytoplasmic granules and exhibit differential association with components of the mRNA degradation/translational repression machinery. The first type of granules, pi-bodies, contains the MILI-TDRD1 module of the piRNA pathway and is likely equivalent to the enigmatic "cementing material" first described in electron micrographs of rat gonocytes over 35 years ago. The second type of granules, piP-bodies, harbors the MIWI2-TDRD9-MAEL module of the piRNA pathway and signature components of P-bodies, GW182, DCP1a, DDX6/p54, and XRN1 proteins. piP-bodies are found predominantly in the proximity of pi-bodies and the two frequently share mouse VASA homolog (MVH protein, an RNA helicase. In Mael-mutant gonocytes, MIWI2, TDRD9, and MVH are lost from piP-bodies, whereas no effects on pi-body composition are observed. Further analysis revealed that MAEL appears to specifically facilitate MIWI2-dependent aspects of the piRNA pathway including biogenesis of secondary piRNAs, de novo DNA methylation, and efficient downregulation of TEs. Cumulatively, our data reveal elaborate cytoplasmic compartmentalization of the fetal piRNA pathway that relies on MAEL function.

  14. Structural Basis for Redox Regulation of Cytoplasmic and Chloroplastic Triosephosphate Isomerases from Arabidopsis thaliana

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    Laura Margarita López-Castillo

    2016-12-01

    Full Text Available In plants triosephosphate isomerase (TPI interconverts glyceraldehyde 3-phosphate (G3P and dihydroxyacetone phosphate (DHAP during glycolysis, gluconeogenesis, and the Calvin-Benson cycle. The nuclear genome of land plants encodes two tpi genes, one gene product is located in the cytoplasm and the other is imported into the chloroplast. Herein we report the crystal structures of the TPIs from the vascular plant Arabidopsis thaliana (AtTPIs and address their enzymatic modulation by redox agents. Cytoplasmic TPI (cTPI and chloroplast TPI (pdTPI share more than 60% amino acid identity and assemble as (β-α8 dimers with high structural homology. cTPI and pdTPI harbor two and one accessible thiol groups per monomer respectively. cTPI and pdTPI present a cysteine at an equivalent structural position (C13 and C15 respectively and cTPI also contains a specific solvent accessible cysteine at residue 218 (cTPI-C218. Site directed mutagenesis of residues pdTPI-C15, cTPI-C13 and cTPI-C218 to serine substantially decreases enzymatic activity, indicating that the structural integrity of these cysteines is necessary for catalysis. AtTPIs exhibit differential responses to oxidative agents, cTPI is susceptible to oxidative agents such as diamide and H2O2, whereas pdTPI is resistant to inhibition. Incubation of AtTPIs with the sulfhydryl conjugating reagents methylmethane thiosulfonate (MMTS and glutathione inhibits enzymatic activity. However, the concentration necessary to inhibit pdTPI is at least two orders of magnitude higher than the concentration needed to inhibit cTPI. Western-blot analysis indicates that residues cTPI-C13, cTPI-C218, and pdTPI-C15 conjugate with glutathione. In summary, our data indicate that AtTPIs could be redox regulated by the derivatization of specific AtTPI cysteines (cTPI-C13 and pdTPI-C15 and cTPI-C218. Since AtTPIs have evolved by gene duplication, the higher resistance of pdTPI to redox agents may be an adaptive consequence to

  15. Origin-independent plasmid replication occurs in vaccinia virus cytoplasmic factories and requires all five known poxvirus replication factors

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    Moss Bernard

    2005-03-01

    Full Text Available Abstract Background Replication of the vaccinia virus genome occurs in cytoplasmic factory areas and is dependent on the virus-encoded DNA polymerase and at least four additional viral proteins. DNA synthesis appears to start near the ends of the genome, but specific origin sequences have not been defined. Surprisingly, transfected circular DNA lacking specific viral sequences is also replicated in poxvirus-infected cells. Origin-independent plasmid replication depends on the viral DNA polymerase, but neither the number of additional viral proteins nor the site of replication has been determined. Results Using a novel real-time polymerase chain reaction assay, we detected a >400-fold increase in newly replicated plasmid in cells infected with vaccinia virus. Studies with conditional lethal mutants of vaccinia virus indicated that each of the five proteins known to be required for viral genome replication was also required for plasmid replication. The intracellular site of replication was determined using a plasmid containing 256 repeats of the Escherichia coli lac operator and staining with an E. coli lac repressor-maltose binding fusion protein followed by an antibody to the maltose binding protein. The lac operator plasmid was localized in cytoplasmic viral factories delineated by DNA staining and binding of antibody to the viral uracil DNA glycosylase, an essential replication protein. In addition, replication of the lac operator plasmid was visualized continuously in living cells infected with a recombinant vaccinia virus that expresses the lac repressor fused to enhanced green fluorescent protein. Discrete cytoplasmic fluorescence was detected in cytoplasmic juxtanuclear sites at 6 h after infection and the area and intensity of fluorescence increased over the next several hours. Conclusion Replication of a circular plasmid lacking specific poxvirus DNA sequences mimics viral genome replication by occurring in cytoplasmic viral factories

  16. HIV-1 and M-PMV RNA Nuclear Export Elements Program Viral Genomes for Distinct Cytoplasmic Trafficking Behaviors.

    Science.gov (United States)

    Pocock, Ginger M; Becker, Jordan T; Swanson, Chad M; Ahlquist, Paul; Sherer, Nathan M

    2016-04-01

    Retroviruses encode cis-acting RNA nuclear export elements that override nuclear retention of intron-containing viral mRNAs including the full-length, unspliced genomic RNAs (gRNAs) packaged into assembling virions. The HIV-1 Rev-response element (RRE) recruits the cellular nuclear export receptor CRM1 (also known as exportin-1/XPO1) using the viral protein Rev, while simple retroviruses encode constitutive transport elements (CTEs) that directly recruit components of the NXF1(Tap)/NXT1(p15) mRNA nuclear export machinery. How gRNA nuclear export is linked to trafficking machineries in the cytoplasm upstream of virus particle assembly is unknown. Here we used long-term (>24 h), multicolor live cell imaging to directly visualize HIV-1 gRNA nuclear export, translation, cytoplasmic trafficking, and virus particle production in single cells. We show that the HIV-1 RRE regulates unique, en masse, Rev- and CRM1-dependent "burst-like" transitions of mRNAs from the nucleus to flood the cytoplasm in a non-localized fashion. By contrast, the CTE derived from Mason-Pfizer monkey virus (M-PMV) links gRNAs to microtubules in the cytoplasm, driving them to cluster markedly to the centrosome that forms the pericentriolar core of the microtubule-organizing center (MTOC). Adding each export element to selected heterologous mRNAs was sufficient to confer each distinct export behavior, as was directing Rev/CRM1 or NXF1/NXT1 transport modules to mRNAs using a site-specific RNA tethering strategy. Moreover, multiple CTEs per transcript enhanced MTOC targeting, suggesting that a cooperative mechanism links NXF1/NXT1 to microtubules. Combined, these results reveal striking, unexpected features of retroviral gRNA nucleocytoplasmic transport and demonstrate roles for mRNA export elements that extend beyond nuclear pores to impact gRNA distribution in the cytoplasm.

  17. Germline transgenesis in pigs by cytoplasmic microinjection of Sleeping Beauty transposons.

    Science.gov (United States)

    Ivics, Zoltán; Garrels, Wiebke; Mátés, Lajos; Yau, Tien Yin; Bashir, Sanum; Zidek, Vaclav; Landa, Vladimír; Geurts, Aron; Pravenec, Michal; Rülicke, Thomas; Kues, Wilfried A; Izsvák, Zsuzsanna

    2014-04-01

    The pig has emerged as an important large animal model in biomedical and pharmaceutical research. We describe a protocol for high-efficiency germline transgenesis and sustained transgene expression in pigs by using the Sleeping Beauty (SB) transposon system. The protocol is based on co-injection of a plasmid encoding the SB100X hyperactive transposase, together with a second plasmid carrying a transgene flanked by binding sites for the transposase, into the cytoplasm of porcine zygotes. The transposase mediates excision of the transgene cassette from the plasmid vector and its permanent insertion into the genome to produce stable transgenic animals. This method compares favorably in terms of both efficiency and reliable transgene expression to classic pronuclear microinjection or somatic cell nuclear transfer (SCNT), and it offers comparable efficacies to lentiviral approaches, without limitations on vector design, issues of transgene silencing and the toxicity and biosafety concerns of working with viral vectors. Microinjection of the vectors into zygotes and transfer of the embryos to recipient animals can be performed in 1 d; generation of germline-transgenic lines by using this protocol takes ∼1 year.

  18. Functional organization of cytoplasmic inclusion bodies in cells infected by respiratory syncytial virus.

    Science.gov (United States)

    Rincheval, Vincent; Lelek, Mickael; Gault, Elyanne; Bouillier, Camille; Sitterlin, Delphine; Blouquit-Laye, Sabine; Galloux, Marie; Zimmer, Christophe; Eleouet, Jean-François; Rameix-Welti, Marie-Anne

    2017-09-15

    Infection of cells by respiratory syncytial virus induces the formation of cytoplasmic inclusion bodies (IBs) where all the components of the viral RNA polymerase complex are concentrated. However, the exact organization and function of these IBs remain unclear. In this study, we use conventional and super-resolution imaging to dissect the internal structure of IBs. We observe that newly synthetized viral mRNA and the viral transcription anti-terminator M2-1 concentrate in IB sub-compartments, which we term "IB-associated granules" (IBAGs). In contrast, viral genomic RNA, the nucleoprotein, the L polymerase and its cofactor P are excluded from IBAGs. Live imaging reveals that IBAGs are highly dynamic structures. Our data show that IBs are the main site of viral RNA synthesis. They further suggest that shortly after synthesis in IBs, viral mRNAs and M2-1 transiently concentrate in IBAGs before reaching the cytosol and suggest a novel post-transcriptional function for M2-1.Respiratory syncytial virus (RSV) induces formation of inclusion bodies (IBs) sheltering viral RNA synthesis. Here, Rincheval et al. identify highly dynamic IB-associated granules (IBAGs) that accumulate newly synthetized viral mRNA and the viral M2-1 protein but exclude viral genomic RNA and RNA polymerase complexes.

  19. Seborrheic inclusion cyst of the skin positive for cytoplasmic inclusion bodies and HPV antigen.

    Science.gov (United States)

    Terada, Tadashi

    2012-01-01

    Seborrheic inclusion cyst (SIC) is a very rare variant of epidermal cyst of the skin. SIC shows seborrheic keratosis (SK)-like lesion in epidermal cyst. SIC is extremely rare; only 6 case reports have been published in the English literature. However, no immunohistochemical study of SIC has been reported. A 41-year-old Japanese man noticed a subcutaneous tumor in the neck. Physical examination showed slightly mobile tumor in the subcutaneous tissue, and total excision was performed. Grossly, the tumor (1 x 1 x 0.8 cm) was cyst containing atheromatous keratin. Microscopically, the lesion is a cyst containing keratins. About one half of the cyst showed features of epidermal cyst consisting of mature squamous epithelium with granular layers. The other one half showed SK-like epidermal proliferation. The SK-like area showed basaloid cell proliferation with pseudohorn cysts. No significant atypia was noted. Many eosinophilic cytoplasmic inclusion bodies were noted in the SK-like area. Immunohistochemically, the SK-like area was positive for pancytokeratin AE1/3, pancytokeratin CAM5.2, p63, and Ki-67 (labeling=8%) and HPV, but negative for p53. The pathological diagnosis was SIC.

  20. Antineutrophil Cytoplasmic Antibodies Testing in a Large Cohort of Unselected Greek Patients

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    Konstantinos Tsiveriotis

    2011-01-01

    Full Text Available Objective. To retrospectively evaluate ANCA testing in a cohort of unselected Greek in- and outpatients. Methods. In 10803 consecutive serum samples, ANCA were tested by indirect immunofluorescence (IIF and ELISA. ELISA in inpatients was performed only on IIF positive sera. Results. Low prevalence (6.0% of IIF positive samples was observed. Among these samples, 63.5% presented perinuclear (p-ANCA, 9.3% cytoplasmic (c-ANCA and 27.2% atypical (x-ANCA pattern. 16.1% of p-ANCA were antimyeloperoxidase (anti-MPO positive, whereas 68.3% of c-ANCA were antiproteinase-3 (anti-PR3 positive. Only 17 IIF negative outpatients' samples were ELISA positive. ANCA-associated vasculitides (AAV, connective tissue disorders and gastrointestinal disorders represented 20.5%, 23.9%, and 21.2% of positive results, respectively. AAV patients exhibited higher rates of MPO/PR3 specificity compared to non-AAV (93.8% versus 8%. Conclusions. This first paper on Greek patients supports that screening for ANCA by IIF and confirming positive results by ELISA minimize laboratory charges without sacrificing diagnostic accuracy.

  1. Regulation of cytoplasmic calcium: interactions between prostaglandins, prostacyclin, thromboxane A2, zinc, copper and taurine.

    Science.gov (United States)

    Horrobin, D F; Manku, M S; Cunnane, S; Karmazyn, M; Morgan, R O; Ally, A I; Karmall, R A

    1978-02-01

    The regulation of cytoplasmic calcium is a key process in nerve tissue. Using a smooth muscle model we have shown that prostaglandin (PG) E2 probably regulates entry from extracellular fluid, whereas the release from intracellular stores depends on the interplay between thromboxane (TX) A2, PGEI and prostacyclin. Hormones and other agents interact with this system in the following ways: vasopressin, angiotensin and inositol mobilize arachidonic acid from membrane phospholipids and increase synthesis of PGE2 and TXA2, cortisol blocks this action. Prolactin and zinc mobilize dihomo-gamma-linolenic acid and increase synthesis of PGEI. These effects can be blocked by cortisol, lithium and taurine, three agents which on their own have no effect on basal PG production. Epileptogenic agents like penicillin and picrotoxin also stimulate PG synthesis, while diphenylhydantoin is a PG antagonist and diazepam is a TXA2 antagonist. The effects of all these agents occur at concentrations which are physiological in the case of the natural ones, and readily attained in human plasma in the case of the drgus. In view of recent evidence that calcium may be important in demyelination and considering the established role it plays in nerve conduction and synaptic transmission, we suggest that these observations may be of significance in understanding Friedreich's ataxia.

  2. Multiple Wolbachia determinants control the evolution of cytoplasmic incompatibilities in Culex pipiens mosquito populations.

    Science.gov (United States)

    Atyame, Celestine M; Duron, Olivier; Tortosa, Pablo; Pasteur, Nicole; Fort, Philippe; Weill, Mylene

    2011-01-01

    Wolbachia are maternally inherited endosymbionts that can invade arthropod populations through manipulation of their reproduction. In mosquitoes, Wolbachia induce embryonic death, known as cytoplasmic incompatibility (CI), whenever infected males mate with females either uninfected or infected with an incompatible strain. Although genetic determinants of CI are unknown, a functional model involving the so-called mod and resc factors has been proposed. Natural populations of Culex pipiens mosquito display a complex CI relationship pattern associated with the highest Wolbachia (wPip) genetic polymorphism reported so far. We show here that C. pipiens populations from La Réunion, a geographically isolated island in the southwest of the Indian Ocean, are infected with genetically closely related wPip strains. Crossing experiments reveal that these Wolbachia are all mutually compatible. However, crosses with genetically more distant wPip strains indicate that Wolbachia strains from La Réunion belong to at least five distinct incompatibility groups (or crossing types). These incompatibility properties which are strictly independent from the nuclear background, formally establish that in C. pipiens, CI is controlled by several Wolbachia mod/resc factors. © 2010 Blackwell Publishing Ltd.

  3. Chlamydia trachomatis GlgA is secreted into host cell cytoplasm.

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    Chunxue Lu

    Full Text Available Glycogen has been localized both inside and outside Chlamydia trachomatis organisms. We now report that C. trachomatis glycogen synthase (GlgA was detected in both chlamydial organism-associated and -free forms. The organism-free GlgA molecules were localized both in the lumen of chlamydial inclusions and in the cytosol of host cells. The cytosolic GlgA displayed a distribution pattern similar to that of a known C. trachomatis-secreted protease, CPAF. The detection of GlgA was specific since the anti-GlgA antibody labeling was only removed by preabsorption with GlgA but not CPAF fusion proteins. GlgA was detectable at 12h and its localization into host cell cytosol only became apparent at 24h after infection. The cytosolic localization of GlgA was conserved among all C. trachomatis serovars. However, the significance of the GlgA secretion into host cell cytoplasm remains unclear since, while expression of chlamydial GlgA in HeLa cells increased glycogen stores, it did not affect a subsequent infection with C. trachomatis. Similar to several other C. trachomatis-secreted proteins, GlgA is immunogenic in women urogenitally infected with C. trachomatis, suggesting that GlgA is expressed and may be secreted into host cell cytosol during C. trachomatis infection in humans. These findings have provided important information for further understanding C. trachomatis pathogenic mechanisms.

  4. Are children born through Intra-Cytoplasmic Sperm Injection (ICSI having a lower intelligence quotient?

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    Mahbobeh Faramarzi

    2016-03-01

    Full Text Available Background: There is still concern about delayed mental development for children born through Intra-Cytoplasmic Sperm Injection (ICSI. In the present study, the intelligence quotient (IQ of ICSI children at the age of 5–6 years was compared with that of the control group of naturally born (NB children. Outcome measure: Full-scale IQ of the mental development in children born through ICSI in 5–6 years. Design: Cross-sectional study. Setting: Fatemehzahra Infertility and Reproductive Health Research Center of Babol – Iran. Participants: 28 live-birth singleton children 5–6 years of the first generation born after ICSI treatment. Methods: The mental development of 28 born through ICSI children at 5–6 years compared with 32 naturally born children as control using intelligence quotient (IQ test obtained from the Wechsler Preschool and Primary Scale of Intelligence (WPPSI (Iranian version. Results: The mean full-scale IQ was 105.9 ± 14 for ICSI and 107.2 ± 12.9 for NB children which showed no significant difference. The only significant possible predictors of slightly lower full-scale IQ in ICSI children were lower parental education level and longer childcare in day center. Conclusion: Our finding provides some reassurance for parents of children born through ICSI regarding the child’s preschool IQ.

  5. [Bacterial endosymbiosis within the cytoplasm of Acanthamoeba lugdunensis isolated from a contact lens storage case].

    Science.gov (United States)

    Chung, D I; Kong, H H; Kim, T H; Hwang, M Y; Yu, H S; Yun, H C; Seol, S Y

    1997-06-01

    Transmission electron microscopy of an Acanthamoeba isolate (KA/L5) from a contact lens case revealed bacterial endosymbionts within cytoplasm of the amoebae. The Acanthamoeba isolate belonged to the morphological group II. Based on the polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) of 18S ribosomal RNA coding DNA (rDNA), the isolate was identified as A. lugdunensis. Strain typing by isoenzyme analysis using isoelectric focusing (IEF) and mitochondrial (Mt) DNA RFLP revealed that the isolate was closely related with KA/L1, the most predominant type of isolates from contact lens storage cases, KA/E2, a clinical isolate, KA/W4, previously reported to host endosymbionts, and L3a strains of A. lugdunensis. The endosymbionts were similar to those of KA/W4 in aspects that they were randomly distributed in both trophozoites and cysts, and were rod-shaped bacteria measuring approximately 1.38 x 0.50 microns. But the number of endosymbionts per amoeba was significantly lower than that of KA/W4. They were neither limited by phagosomal membranes nor included in lacunaelike structure.

  6. Wolbachia age-sex-specific density in Aedes albopictus: a host evolutionary response to cytoplasmic incompatibility?

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    Pablo Tortosa

    2010-03-01

    Full Text Available Wolbachia bacteria have invaded many arthropod species by inducing Cytoplasmic Incompatibility (CI. These symbionts represent fascinating objects of study for evolutionary biologists, but also powerful potential biocontrol agents. Here, we assess the density dynamics of Wolbachia infections in males and females of the mosquito Aedes albopitcus, an important vector of human pathogens, and interpret the results within an evolutionary framework.Wolbachia densities were measured in natural populations and in age controlled mosquitoes using quantitative PCR. We show that the density dynamics of the wAlbA Wolbachia strain infecting Aedes albopictus drastically differ between males and females, with a very rapid decay of infection in males only.Theory predicts that Wolbachia and its hosts should cooperate to improve the transmission of infection to offspring, because only infected eggs are protected from the effects of CI. However, incompatible matings effectively lower the fertility of infected males, so that selection acting on the host genome should tend to reduce the expression of CI in males, for example, by reducing infection density in males before sexual maturation. The rapid decay of one Wolbachia infection in Aedes albopictus males, but not in females, is consistent with this prediction. We suggest that the commonly observed reduction in CI intensity with male age reflects a similar evolutionary process. Our results also highlight the importance of monitoring infection density dynamics in both males and females to assess the efficiency of Wolbachia-based control strategies.

  7. The Cytoplasmic Adaptor Protein Dok7 Activates the Receptor Tyrosine Kinase MuSK via Dimerization

    Energy Technology Data Exchange (ETDEWEB)

    Bergamin, E.; Hallock, P; Burden, S; Hubbard, S

    2010-01-01

    Formation of the vertebrate neuromuscular junction requires, among others proteins, Agrin, a neuronally derived ligand, and the following muscle proteins: LRP4, the receptor for Agrin; MuSK, a receptor tyrosine kinase (RTK); and Dok7 (or Dok-7), a cytoplasmic adaptor protein. Dok7 comprises a pleckstrin-homology (PH) domain, a phosphotyrosine-binding (PTB) domain, and C-terminal sites of tyrosine phosphorylation. Unique among adaptor proteins recruited to RTKs, Dok7 is not only a substrate of MuSK, but also an activator of MuSK's kinase activity. Here, we present the crystal structure of the Dok7 PH-PTB domains in complex with a phosphopeptide representing the Dok7-binding site on MuSK. The structure and biochemical data reveal a dimeric arrangement of Dok7 PH-PTB that facilitates trans-autophosphorylation of the kinase activation loop. The structure provides the molecular basis for MuSK activation by Dok7 and for rationalizing several Dok7 loss-of-function mutations found in patients with congenital myasthenic syndromes.

  8. Katanin p80, NuMA and cytoplasmic dynein cooperate to control microtubule dynamics.

    Science.gov (United States)

    Jin, Mingyue; Pomp, Oz; Shinoda, Tomoyasu; Toba, Shiori; Torisawa, Takayuki; Furuta, Ken'ya; Oiwa, Kazuhiro; Yasunaga, Takuo; Kitagawa, Daiju; Matsumura, Shigeru; Miyata, Takaki; Tan, Thong Teck; Reversade, Bruno; Hirotsune, Shinji

    2017-01-12

    Human mutations in KATNB1 (p80) cause severe congenital cortical malformations, which encompass the clinical features of both microcephaly and lissencephaly. Although p80 plays critical roles during brain development, the underlying mechanisms remain predominately unknown. Here, we demonstrate that p80 regulates microtubule (MT) remodeling in combination with NuMA (nuclear mitotic apparatus protein) and cytoplasmic dynein. We show that p80 shuttles between the nucleus and spindle pole in synchrony with the cell cycle. Interestingly, this striking feature is shared with NuMA. Importantly, p80 is essential for aster formation and maintenance in vitro. siRNA-mediated depletion of p80 and/or NuMA induced abnormal mitotic phenotypes in cultured mouse embryonic fibroblasts and aberrant neurogenesis and neuronal migration in the mouse embryonic brain. Importantly, these results were confirmed in p80-mutant harboring patient-derived induced pluripotent stem cells and brain organoids. Taken together, our findings provide valuable insights into the pathogenesis of severe microlissencephaly, in which p80 and NuMA delineate a common pathway for neurogenesis and neuronal migration via MT organization at the centrosome/spindle pole.

  9. Tubular lysosome morphology and distribution within macrophages depend on the integrity of cytoplasmic microtubules

    International Nuclear Information System (INIS)

    Swanson, J.; Bushnell, A.; Silverstein, S.C.

    1987-01-01

    Pinocytosis of the fluorescent dye lucifer yellow labels elongated, membrane-bound tubular organelles in several cell types, including cultured human monocytes, thioglycolate-elicited mouse peritoneal macrophages, and the macrophage-like cell line J774.2. These tubular structures can be identified as lysosomes by acid phosphatase histochemistry and immunofluorescence localization of cathepsin L. The abundance of tubular lysosomes is markedly increased by treatment with phorbol 12-myristate 13-acetate. When labeled by pinocytosis of microperoxidase and examined by electron microscopic histochemistry, the tubular lysosomes have an outside diameter of ≅ 75 nm and a length of several micrometers; they radiate from the cell's centrosphere in alignment with cytoplasmic microtubules and intermediate filaments. Incubation of phorbol myristate acetate-treated macrophages at 4 0 C or in medium containing 5 μM colchicine or nocodazole at 37 0 C leads to disassembly of microtubules and fragmentation of the tubular lysosomes. Return of the cultures to 37 0 C or removal of nocodazole from the medium leads to reassembly of microtubules and the reappearance of tubular lysosomes within 10-20 min. The authors conclude that microtubules are essential for the maintenance of tubular lysosome morphology and that, in macrophages, a significant proportion of the lysosomal compartment is contained within these tubular structures

  10. Molecular mechanisms for mediating light-dependent nucleo/cytoplasmic partitioning of phytochrome photoreceptors.

    Science.gov (United States)

    Klose, Cornelia; Viczián, András; Kircher, Stefan; Schäfer, Eberhard; Nagy, Ferenc

    2015-05-01

    The photoreceptors phytochromes monitor the red/far-red part of the spectrum, exist in the biologically active Pfr (far-red absorbing) or inactive Pr (red absorbing) forms, and function as red/far-red light-regulated molecular switches to modulate plant development and growth. Phytochromes are synthesized in the cytoplasm, and light induces translocation of the Pfr conformer into the nucleus. Nuclear import of phytochromes is a highly regulated process and is fine-tuned by the quality and quantity of light. It appears that phytochrome A (phyA) and phytochrome B (phyB) do not possess active endogenous nuclear import signals (NLSs), thus light-induced translocation of these photoreceptors into the nucleus requires direct protein–protein interactions with their NLS-containing signaling partners. Sub-cellular partitioning of the various phytochrome species is mediated by different molecular machineries. Translocation of phyA into the nucleus is promoted by FAR-RED ELONGATED HYPOCOTYL 1 (FHY1) and FHY1-LIKE (FHL), but the identity of nuclear transport facilitators mediating the import of phyB-E into the nucleus remains elusive. Phytochromes localized in the nucleus are associated with specific protein complexes, termed photobodies. The size and distribution of these structures are regulated by the intensity and duration of irradiation, and circumstantial evidence indicates that they are involved in fine-tuning phytochrome signaling.

  11. Circumvention of nuclear factor kappaB-induced chemoresistance by cytoplasmic-targeted anthracyclines.

    Science.gov (United States)

    Bilyeu, Jennifer D; Panta, Ganesh R; Cavin, Lakita G; Barrett, Christina M; Turner, Eddie J; Sweatman, Trevor W; Israel, Mervyn; Lothstein, Leonard; Arsura, Marcello

    2004-04-01

    Nuclear factor kappaB (NF-kappaB) has been implicated in inducible chemoresistance against anthracyclines. In an effort to improve the cytotoxicity of anthracyclines while reducing their cardiotoxic effects, we have developed a novel class of extranuclear-localizing 14-O-acylanthracyclines that bind to the phorbol ester/diacylglycerol-binding C1b domain of conventional and novel protein kinase C (PKC) isoforms, thereby promoting an apoptotic response. Because PKCs have been shown to be involved in NF-kappaB activation, in this report, we determined the mechanism of NF-kappaB activation by N-benzyladriamycin-14-valerate (AD 198) and N-benzyladriamycin-14-pivalate (AD 445), two novel 14-O-acylanthracylines. We show that the induction of NF-kappaB activity in response to drug treatment relies on the activation of PKC-delta and NF-kappaB-activating kinase (NAK), independent of ataxia telengectasia mutated and p53 activities. In turn, NAK activates the IKK complex through phosphorylation of the IKK-2 subunit. We find that neither NF-kappaB activation nor ectopic expression of Bcl-X(L) confers protection from AD 198-induced cell killing. Overall, our data indicate that activation of novel PKC isoforms by cytoplasmic-targeted 14-O-acylanthracyclines promotes an apoptotic response independent of DNA damage, which is unimpeded by inducible activation of NF-kappaB.

  12. Comparison of disease activity measures for anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitis

    Science.gov (United States)

    Merkel, PA; Cuthbertson, DD; Hellmich, B; Hoffman, GS; Jayne, DRW; Kallenberg, CGM; Krischer, JP; Luqmani, R; Mahr, AD; Matteson, EL; Specks, U; Stone, JH

    2011-01-01

    Aim Currently, several different instruments are used to measure disease activity and extent in clinical trials of anti-neutrophil cytoplasmic autoantibody (ANCA)-associated vasculitis, leading to division among investigative groups and difficulty comparing study results. An exercise comparing six different vasculitis instruments was performed. Methods A total of 10 experienced vasculitis investigators from 5 countries scored 20 cases in the literature of Wegener granulomatosis or microscopic polyangiitis using 6 disease assessment tools: the Birmingham Vasculitis Activity Score (BVAS), The BVAS for Wegener granulomatosis (BVAS/WG), BVAS 2003, a Physician Global Assessment (PGA), the Disease Extent Index (DEI) and the Five Factor Score (FFS). Five cases were rescored by all raters. Results Reliability of the measures was extremely high (intraclass correlations for the six measures all=0.98). Within each instrument, there were no significant differences or outliers among the scores from the 10 investigators. Test/retest reliability was high for each measure: range=0.77 to 0.95. The scores of the five acute activity measures correlated extremely well with one another. Conclusions Currently available tools for measuring disease extent and activity in ANCA-associated vasculitis are highly correlated and reliable. These results provide investigators with confidence to compare different clinical trial data and helps form common ground as international research groups develop new, improved and universally accepted vasculitis disease assessment instruments. PMID:18664546

  13. Detection of Antibodies to Brucella Cytoplasmic Proteins in the Cerebrospinal Fluid of Patients with Neurobrucellosis

    Science.gov (United States)

    Baldi, Pablo C.; Araj, George F.; Racaro, Graciela C.; Wallach, Jorge C.; Fossati, Carlos A.

    1999-01-01

    The diagnosis of human neurobrucellosis usually relies on the detection of antibodies to Brucella lipopolysaccharide (LPS) in cerebrospinal fluid (CSF) by agglutination tests or enzyme-linked immunosorbent assay (ELISA). Here we describe the detection of immunoglobulin G (IgG) to cytoplasmic proteins (CP) of Brucella spp. by ELISA and Western blotting in seven CSF samples from five patients with neurobrucellosis. While IgG to CP (titers of 200 to 12,800) and IgG to LPS (800 to 6,400) were found in the CSF of these patients, these antibodies were not detected in CSF samples from two patients who had systemic brucellosis without neurological involvement. The latter, however, had serum IgG and IgM to both LPS and CP. No reactivity to these antigens was found in CSF samples from 14 and 20 patients suffering from nonbrucellar meningitis and noninfectious diseases, respectively. These findings suggest that, in addition to its usefulness in the serological diagnosis of human systemic brucellosis, the ELISA with CP antigen can be used for the specific diagnosis of human neurobrucellosis. PMID:10473531

  14. Phosphorylation of the dimeric cytoplasmic domain of the phytosulfokine receptor, PSKR1

    KAUST Repository

    Muleya, V.

    2016-08-04

    Phytosulfokines (PSKs) are plant peptide hormones that co-regulate plant growth, differentiation and defense responses. PSKs signal through a plasma membrane localized leucine-rich repeat receptor-like kinase (phytosulfokine receptor 1, PSKR1) that also contains a functional cytosolic guanylate cyclase with its cyclase catalytic center embedded within the kinase domain. To functionally characterize this novel type of overlapping dual catalytic function, we investigated the phosphorylation of PSKR1 in vitro Tandem mass spectrometry of the cytoplasmic domain of PSKR1 (PSKR1cd) revealed at least 11 phosphorylation sites (8 serines, 2 threonines and 1 tyrosine) within the PSKR1cd. Phosphomimetic mutations of three serine residues (Ser686, Ser696 and Ser698) in tandem at the juxta-membrane position resulted in enhanced kinase activity in the on-mutant that was suppressed in the off-mutant, but both mutations reduced guanylate cyclase activity. Both the on and off phosphomimetic mutations of the phosphotyrosine (Tyr888) residue in the activation loop suppressed kinase activity, while neither mutation affected guanylate cyclase activity. Size exclusion and analytical ultracentrifugation analysis of the PSKR1cd suggest that it is reversibly dimeric in solution, which was further confirmed by biflourescence complementation. Taken together, these data suggest that in this novel type of receptor domain architecture, specific phosphorylation and dimerization are possibly essential mechanisms for ligand-mediated catalysis and signaling.

  15. Cytoplasmic expression of a thermostable invertase from Thermotoga maritima in Lactococcus lactis.

    Science.gov (United States)

    Pek, Han Bin; Lim, Pei Yu; Liu, Chengcheng; Lee, Dong-Yup; Bi, Xuezhi; Wong, Fong Tian; Ow, Dave Siak-Wei

    2017-05-01

    To evaluate the secretory and cytoplasmic expression of a thermostable Thermogata maritima invertase in Lactococcus lactis. The thermostable invertase from T. maritima was cloned with and without the USP45 secretory peptide into the pNZ8148 vector for nisin-inducible expression in L. lactis. The introduction of an USP45 secretion peptide at the N-terminal of the enzyme led to a loss of protein solubility. Computational homology modeling and hydrophobicity studies indicated that the USP45 peptide exposes a stretch of hydrophobic amino acids on the protein surface resulting in lower solubility. Removal of the USP45 secretion peptide allowed a soluble and functional invertase to be expressed intracellularly in L. lactis. Immobilized metal affinity chromatography purification of the cell lysate with nickel-NTA gave a single protein band on SDS-PAGE, while E. coli-expressed invertase consistently co-purified with an additional band. The yields of the purified invertase from E. coli and L. lactis were 14.1 and 6.3 mg/l respectively. Invertase can be expressed in L. lactis and purified in a functional form. L. lactis is a suitable host for the production of food-grade invertase for use in the food and biotechnology industries.

  16. EB1 and cytoplasmic dynein mediate protrusion dynamics for efficient 3-dimensional cell migration.

    Science.gov (United States)

    Jayatilaka, Hasini; Giri, Anjil; Karl, Michelle; Aifuwa, Ivie; Trenton, Nicholaus J; Phillip, Jude M; Khatau, Shyam; Wirtz, Denis

    2018-03-01

    Microtubules have long been implicated to play an integral role in metastatic disease, for which a critical step is the local invasion of tumor cells into the 3-dimensional (3D) collagen-rich stromal matrix. Here we show that cell migration of human cancer cells uses the dynamic formation of highly branched protrusions that are composed of a microtubule core surrounded by cortical actin, a cytoskeletal organization that is absent in cells on 2-dimensional (2D) substrates. Microtubule plus-end tracking protein End-binding 1 and motor protein dynein subunits light intermediate chain 2 and heavy chain 1, which do not regulate 2D migration, critically modulate 3D migration by affecting RhoA and thus regulate protrusion branching through differential assembly dynamics of microtubules. An important consequence of this observation is that the commonly used cancer drug paclitaxel is 100-fold more effective at blocking migration in a 3D matrix than on a 2D matrix. This work reveals the central role that microtubule dynamics plays in powering cell migration in a more pathologically relevant setting and suggests further testing of therapeutics targeting microtubules to mitigate migration.-Jayatilaka, H., Giri, A., Karl, M., Aifuwa, I., Trenton, N. J., Phillip, J. M., Khatau, S., Wirtz, D. EB1 and cytoplasmic dynein mediate protrusion dynamics for efficient 3-dimensional cell migration.

  17. "Cytoplasmic domain effects on exposure of co-receptor-binding sites of HIV-1 Env".

    Science.gov (United States)

    Vzorov, Andrei N; Compans, Richard W

    2016-11-01

    We defined the effects of the cytoplasmic domain (CT) of the Env glycoprotein on co-receptor usage of HIV-1 by reciprocal exchanges of regions containing V3-V5 loops between CD4-dependent and CD4-independent isolates. Primary HIV-1 isolate Env clones CD8 CXCR4-tropic 92UG046 CT84 with an 84-aa truncated CT domain, CD4 CXCR4-tropic 92UG046, and CD4 CCR5-tropic SF162 with full-length (FL) CT domains were used for comparison. The parental 92UG046 Env with CT84 was not fusogenic, but a chimeric SF162 V3-V5-CT84 with an 84-aa truncated CT domain, which demonstrated a switched co-receptor specificity, exhibited syncytium-formation activity with 3T3T4X4 cells. The wild-type (WT) SF162 Env with CT84 or full-length CT was fusogenic in 3T3T4R5 cells. By exchange of V3-V5 loops, we were able to alter WT SF162 to switch its co-receptor preference, which was not dependent on CT domain length. These results provide evidence that CT domains can induce conformational changes in functional regions of gp120 and determine receptor tropism but do not modulate HIV-1 co-receptor specificity.

  18. Solution Structure and Membrane Interaction of the Cytoplasmic Tail of HIV-1 gp41 Protein.

    Science.gov (United States)

    Murphy, R Elliot; Samal, Alexandra B; Vlach, Jiri; Saad, Jamil S

    2017-11-07

    The cytoplasmic tail of gp41 (gp41CT) remains the last HIV-1 domain with an unknown structure. It plays important roles in HIV-1 replication such as mediating envelope (Env) intracellular trafficking and incorporation into assembling virions, mechanisms of which are poorly understood. Here, we present the solution structure of gp41CT in a micellar environment and characterize its interaction with the membrane. We show that the N-terminal 45 residues are unstructured and not associated with the membrane. However, the C-terminal 105 residues form three membrane-bound amphipathic α helices with distinctive structural features such as variable degree of membrane penetration, hydrophobic and basic surfaces, clusters of aromatic residues, and a network of cation-π interactions. This work fills a major gap by providing the structure of the last segment of HIV-1 Env, which will provide insights into the mechanisms of Gag-mediated Env incorporation as well as the overall Env mobility and conformation on the virion surface. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. The effect of phenobarbitone on cytoplasmic NADP-linked dehydrogenase activities in rat liver.

    Science.gov (United States)

    Shas, S; Pearson, D J

    1978-02-13

    Phenobarbitone administered in drinking water (0.5 g/l) or by daily intraperitoneal injection (100 mg/kg body weight) consistently caused an elevation of hepatic NADP-linked malic enzyme in rats maintained on a pellet diet. Three to four days appeared to be required for maximum response. The effect was also observed in animals maintained on a protein rich diet, in which the basic hepatic malic enzyme activity was low, but not in animals maintained on a sucrose rich diet, in which the basic enzyme activity was almost twice normal. Methyl cholanthrene, administered by daily intraperitoneal injection (40 mg/kg body weight), resulted in elevated hepatic levels not only of malic enzyme but also of the pentose phosphate pathway dehydrogenases. The timing of the "starve-refeed" response of the hepatic NADP-linked dehydrogenases in phenobarbitone-treated rats was similar to that in controls, and similar maximum enzyme activities were reached. The role of cytoplasmic NADP-linked dehydrogenases in the provision of reducing equivalent is discussed, particularly in relation to hepatic microsomal drug metabolism.

  20. Exploring nucleo-cytoplasmic large DNA viruses in Tara Oceans microbial metagenomes.

    Science.gov (United States)

    Hingamp, Pascal; Grimsley, Nigel; Acinas, Silvia G; Clerissi, Camille; Subirana, Lucie; Poulain, Julie; Ferrera, Isabel; Sarmento, Hugo; Villar, Emilie; Lima-Mendez, Gipsi; Faust, Karoline; Sunagawa, Shinichi; Claverie, Jean-Michel; Moreau, Hervé; Desdevises, Yves; Bork, Peer; Raes, Jeroen; de Vargas, Colomban; Karsenti, Eric; Kandels-Lewis, Stefanie; Jaillon, Olivier; Not, Fabrice; Pesant, Stéphane; Wincker, Patrick; Ogata, Hiroyuki

    2013-09-01

    Nucleo-cytoplasmic large DNA viruses (NCLDVs) constitute a group of eukaryotic viruses that can have crucial ecological roles in the sea by accelerating the turnover of their unicellular hosts or by causing diseases in animals. To better characterize the diversity, abundance and biogeography of marine NCLDVs, we analyzed 17 metagenomes derived from microbial samples (0.2-1.6 μm size range) collected during the Tara Oceans Expedition. The sample set includes ecosystems under-represented in previous studies, such as the Arabian Sea oxygen minimum zone (OMZ) and Indian Ocean lagoons. By combining computationally derived relative abundance and direct prokaryote cell counts, the abundance of NCLDVs was found to be in the order of 10(4)-10(5) genomes ml(-1) for the samples from the photic zone and 10(2)-10(3) genomes ml(-1) for the OMZ. The Megaviridae and Phycodnaviridae dominated the NCLDV populations in the metagenomes, although most of the reads classified in these families showed large divergence from known viral genomes. Our taxon co-occurrence analysis revealed a potential association between viruses of the Megaviridae family and eukaryotes related to oomycetes. In support of this predicted association, we identified six cases of lateral gene transfer between Megaviridae and oomycetes. Our results suggest that marine NCLDVs probably outnumber eukaryotic organisms in the photic layer (per given water mass) and that metagenomic sequence analyses promise to shed new light on the biodiversity of marine viruses and their interactions with potential hosts.

  1. C-reactive protein enhances the respiratory burst of neutrophils-induced by antineutrophil cytoplasmic antibody.

    Science.gov (United States)

    Xu, Peng-cheng; Hao, Jian; Yang, Xiao-wei; Chang, Dong-yuan; Chen, Min; Zhao, Ming-hui

    2012-10-01

    Serum C-reactive protein (CRP) was one of the useful biomarkers for evaluating the disease activity in antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV). Cumulating studies proved that CRP was pathogenic in a variety of diseases. In the current study, the in vitro effects of CRP to prime neutrophils for ANCA-induced respiratory burst were investigated with flow cytometry. Without TNF-α in the reactive system, ANCA could only induce a slight level of respiratory burst of neutrophils. CRP could enhance the respiratory burst of neutrophils induced by ANCA against myeloperoxidse [mean fluorescence intensity (MFI, 68.45 ± 16.87 vs. 58.65 ± 15.09, P Heat-treated CRP could not enhance the levels of neutrophils respiratory burst induced by ANCA or increase the expression of membrane proteinase 3 of neutrophils. So CRP can prime neutrophils and enhance the respiratory burst induced by ANCA and might be pathogenic in AAV. Copyright © 2012 Elsevier Ltd. All rights reserved.

  2. Flexibility of the Cytoplasmic Domain of the Phototaxis Transducer II from Natronomonas pharaonis

    Directory of Open Access Journals (Sweden)

    Ivan L. Budyak

    2008-01-01

    Full Text Available Chemo- and phototaxis systems in bacteria and archaea serve as models for more complex signal transduction mechanisms in higher eukaryotes. Previous studies of the cytoplasmic fragment of the phototaxis transducer (pHtrII-cyt from the halophilic archaeon Natronomonas pharaonis showed that it takes the shape of a monomeric or dimeric rod under low or high salt conditions, respectively. CD spectra revealed only approximately 24% helical structure, even in 4 M KCl, leaving it an open question how the rod-like shape is achieved. Here, we conducted CD, FTIR, and NMR spectroscopic studies under different conditions to address this question. We provide evidence that pHtrII-cyt is highly dynamic with strong helical propensity, which allows it to change from monomeric to dimeric helical coiled-coil states without undergoing dramatic shape changes. A statistical analysis of predicted disorder for homologous sequences suggests that structural flexibility is evolutionarily conserved within the methyl-accepting chemotaxis protein family.

  3. Molecular cloning and characterization of Antheraea mylitta cytoplasmic polyhedrosis virus polyhedrin gene and its variant forms

    International Nuclear Information System (INIS)

    Sinha-Datta, Uma; Chavali, Venkata Ramana Murthy; Ghosh, Ananta K.

    2005-01-01

    The segments 10 (S10) of the 11 double stranded RNA genomes from Antheraea mylitta cytoplasmic polyhedrosis virus (AmCPV) encoding a novel polyhedrin polypeptide was converted to cDNA, cloned, and sequenced. Three cDNA clones consisting of 1502 (AmCPV10-1), 1120 (AmCPV10-2), and 1415 (AmCPV10-3) nucleotides encoding polyhedrin of 254, 339, and 319 amino acids with molecular masses of 29, 39, and 37 kDa, respectively, were obtained, and verified by Northern analysis. These clones showed 70-94% sequence identity among them but none with any sequences in databases. The expression of AmCPV10-1 cDNA encoded polyhedrin in Sf-9 cells was detected by immunoblot analysis and formation of polyhedra by electron microscopy, as observed in AmCPV-infected gut cells, but no expression of AmCPV10-2 or AmCPV10-3 cDNA was detected, indicating that during AmCPV replication, along with functional S10 RNA, some defective variant forms of S10 RNAs are packaged in virion particles

  4. Regulation of Hippo pathway transcription factor TEAD by p38 MAPK-induced cytoplasmic translocation.

    Science.gov (United States)

    Lin, Kimberly C; Moroishi, Toshiro; Meng, Zhipeng; Jeong, Han-Sol; Plouffe, Steven W; Sekido, Yoshitaka; Han, Jiahuai; Park, Hyun Woo; Guan, Kun-Liang

    2017-07-28

    The Hippo pathway controls organ size and tissue homeostasis, with deregulation leading to cancer. The core Hippo components in mammals are composed of the upstream serine/threonine kinases Mst1/2, MAPK4Ks and Lats1/2. Inactivation of these upstream kinases leads to dephosphorylation, stabilization, nuclear translocation and thus activation of the major functional transducers of the Hippo pathway, YAP and its paralogue TAZ. YAP/TAZ are transcription co-activators that regulate gene expression primarily through interaction with the TEA domain DNA-binding family of transcription factors (TEAD). The current paradigm for regulation of this pathway centres on phosphorylation-dependent nucleocytoplasmic shuttling of YAP/TAZ through a complex network of upstream components. However, unlike other transcription factors, such as SMAD, NF-κB, NFAT and STAT, the regulation of TEAD nucleocytoplasmic shuttling has been largely overlooked. In the present study, we show that environmental stress promotes TEAD cytoplasmic translocation via p38 MAPK in a Hippo-independent manner. Importantly, stress-induced TEAD inhibition predominates YAP-activating signals and selectively suppresses YAP-driven cancer cell growth. Our data reveal a mechanism governing TEAD nucleocytoplasmic shuttling and show that TEAD localization is a critical determinant of Hippo signalling output.

  5. Naturalising Representational Content

    Science.gov (United States)

    Shea, Nicholas

    2014-01-01

    This paper sets out a view about the explanatory role of representational content and advocates one approach to naturalising content – to giving a naturalistic account of what makes an entity a representation and in virtue of what it has the content it does. It argues for pluralism about the metaphysics of content and suggests that a good strategy is to ask the content question with respect to a variety of predictively successful information processing models in experimental psychology and cognitive neuroscience; and hence that data from psychology and cognitive neuroscience should play a greater role in theorising about the nature of content. Finally, the contours of the view are illustrated by drawing out and defending a surprising consequence: that individuation of vehicles of content is partly externalist. PMID:24563661

  6. Lead content of calcium supplements.

    Science.gov (United States)

    Ross, E A; Szabo, N J; Tebbett, I R

    2000-09-20

    Substantial quantities of lead have been reported in some over-the-counter calcium supplement preparations, including not only bone-meal and dolomite, but also over-the-counter natural and refined calcium carbonate formulations. Examination of this issue is warranted given recent increases in physician recommendations for calcium supplements for prevention and treatment of osteoporosis. To determine the lead content of calcium supplements and to quantify the lead exposure from popular brands of calcium in dosages used for childhood recommended daily allowance, osteoporosis, and phosphate binding in dialysis patients. Analysis of lead content in 21 formulations of nonprescription calcium carbonate (including 7 natural [ie, oyster shell] and 14 refined), 1 brand of prescription-only calcium acetate, and 1 noncalcium synthetic phosphate binder conducted in March 2000. Lead content, assayed using electrothermal atomic absorption, expressed as micrograms of lead per 800 mg/d of elemental calcium, per 1500 mg/d of calcium, and for a range of dosages for patients with renal failure. Six microg/d of lead was considered the absolute dietary limit, with no more than 1 microg/d being the goal for supplements. Four of 7 natural products had measurable lead content, amounting to approximately 1 microg/d for 800 mg/d of calcium, between 1 and 2 microg/d for 1500 mg/d of calcium, and up to 10 microg/d for renal dosages. Four of the 14 refined products had similar lead content, including up to 3 microg/d of lead in osteoporosis calcium dosages and up to 20 microg/d in high renal dosages. No lead was detected in the calcium acetate or polymer products. Lead was present even in some brand name products from major pharmaceutical companies not of natural oyster shell derivation. Despite increasingly stringent limits of lead exposure, many calcium supplement formulations contain lead and thereby may pose an easily avoidable public health concern. JAMA. 2000;284:1425-1429.

  7. Na/K pump inactivation, subsarcolemmal Na measurements, and cytoplasmic ion turnover kinetics contradict restricted Na spaces in murine cardiac myocytes

    OpenAIRE

    Lu, Fang-Min; Hilgemann, Donald W.

    2017-01-01

    The Na/K pump exports cytoplasmic Na ions while importing K ions, and its activity is thought to be affected by restricted intracellular Na diffusion in cardiac myocytes. Lu and Hilgemann find instead that the pump can enter an inactivated state and that inactivation can be relieved by cytoplasmic Na.

  8. Bullying Prevention

    Science.gov (United States)

    Kemp, Patrice

    2016-01-01

    The focus of the milestone project is to focus on bridging the gap of bullying and classroom instruction methods. There has to be a defined expectations and level of accountability that has to be defined when supporting and implementing a plan linked to bullying prevention. All individuals involved in the student's learning have to be aware of…

  9. Prevention: Exercise

    Medline Plus

    Full Text Available ... Strengthen Your Core! Stretching/Flexibility Aerobic Exercise Cervical Exercise Strength Training for the Elderly Other Back Pack Safety Pregnancy and Back Pain Preventing Osteoporosis Back Pain Basics Book RESOURCES ... The Backbone of Spine Treatment Spondylolisthesis BLOG FIND ...

  10. Prevent Pneumonia

    Centers for Disease Control (CDC) Podcasts

    2015-08-06

    CDC’s Matthew Westercamp explains what pneumonia is, its symptoms, and how to prevent it.  Created: 8/6/2015 by National Center for Immunization and Respiratory Diseases (NCIRD), Division of Bacterial Diseases (DBD), Respiratory Diseases Branch (RDB).   Date Released: 8/6/2015.

  11. HIV Prevention

    Science.gov (United States)

    ... Abroad Treatment Basic Statistics Get Tested Find an HIV testing site near you. Enter ZIP code or city Follow HIV/AIDS CDC HIV CDC HIV/AIDS See RSS | ... Collapse All Is abstinence the only 100% effective HIV prevention option? Yes. Abstinence means not having oral, ...

  12. Prevention: Exercise

    Medline Plus

    Full Text Available ... Information Feature Articles Patient Q&A Success Stories Definitions Anatomy of the Spine Definitions A-Z Spine Specialists Videos 9 for Spine Epidural Steroid Injections Exercise: The Backbone of Spine Treatment Spondylolisthesis BLOG FIND A SPECIALIST Prevention ...

  13. Opioid Overdose Prevention: Safety Advice for Patients & Family Members

    Science.gov (United States)

    SAMHSA Opioid Overdose Prevention TOOLKIT: Safety Advice for Patients & Family Members TABLE OF CONTENTS Safety Advice for Patients & Family ... Advice for Patients & Family Members. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 What Are Opioids. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 Preventing Overdose . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1 If You Suspect an ...

  14. Chemotaxis cluster 1 proteins form cytoplasmic arrays in Vibrio cholerae and are stabilized by a double signaling domain receptor DosM.

    Science.gov (United States)

    Briegel, Ariane; Ortega, Davi R; Mann, Petra; Kjær, Andreas; Ringgaard, Simon; Jensen, Grant J

    2016-09-13

    Nearly all motile bacterial cells use a highly sensitive and adaptable sensory system to detect changes in nutrient concentrations in the environment and guide their movements toward attractants and away from repellents. The best-studied bacterial chemoreceptor arrays are membrane-bound. Many motile bacteria contain one or more additional, sometimes purely cytoplasmic, chemoreceptor systems. Vibrio cholerae contains three chemotaxis clusters (I, II, and III). Here, using electron cryotomography, we explore V. cholerae's cytoplasmic chemoreceptor array and establish that it is formed by proteins from cluster I. We further identify a chemoreceptor with an unusual domain architecture, DosM, which is essential for formation of the cytoplasmic arrays. DosM contains two signaling domains and spans the two-layered cytoplasmic arrays. Finally, we present evidence suggesting that this type of receptor is important for the structural stability of the cytoplasmic array.

  15. Drug Abuse Prevention Starts with Parents

    Science.gov (United States)

    ... Stages Listen Español Text Size Email Print Share Drug Abuse Prevention Starts with Parents Page Content Article ... Learn the facts about the harmful effects of drugs. Talk with your child about the negative effects ...

  16. Components of the E. coli envelope are affected by and can react to protein over-production in the cytoplasm

    Directory of Open Access Journals (Sweden)

    Villa Riccardo

    2009-06-01

    Full Text Available Abstract Background Protein over-expression in bacteria is still the easiest, cheapest and therefore preferred way to obtain large amounts of proteins for industrial and laboratory scale preparations. Several studies emphasized the importance of understanding cellular and molecular mechanisms triggered by protein over-production in order to obtain higher yield and better quality of the recombinant product. Almost every step leading to a fully functional polypeptide has been investigated, from mRNA stability to the role of molecular chaperones, from aggregation to bottlenecks in the secretory pathway. In this context, we focused on the still poorly addressed relationship between protein production in the cytoplasm and the bacterial envelope, an active and reactive cell compartment that controls interactions with the environment and several major cellular processes. Results available to date show that the accumulation of foreign proteins in the cytoplasm induces changes in the membrane lipids and in the levels of mRNAs for some membrane proteins. However, a direct connection between membrane protein expression levels and soluble/aggregated protein accumulation in the cytoplasm has never been reported. Results By the use of a combined physiological and proteomic approach, we investigated the effects on the cell membrane of E. coli of the overexpression of two recombinant proteins, the B. cepacia lipase (BCL and the green fluorescent protein (GFP. Both polypeptides are expressed in the cytoplasm at similar levels but GFP is fully soluble whereas inactive BCL accumulates in inclusion bodies. Growth and viability of the transformed cells were tested in the presence of different drugs. We found that chloramphenycol preferentially inhibited the strain over-producing GFP while SDS was more effective when BCL inclusion bodies accumulated in the cytoplasm. In contrast, both proteins induced a similar response in the membrane proteome, i.e. increased levels

  17. Membrane-bound MinDE complex acts as a toggle switch that drives Min oscillation coupled to cytoplasmic depletion of MinD

    Science.gov (United States)

    Vecchiarelli, Anthony G.; Li, Min; Mizuuchi, Michiyo; Hwang, Ling Chin; Seol, Yeonee; Neuman, Keir C.; Mizuuchi, Kiyoshi

    2016-01-01

    The Escherichia coli Min system self-organizes into a cell-pole to cell-pole oscillator on the membrane to prevent divisions at the cell poles. Reconstituting the Min system on a lipid bilayer has contributed to elucidating the oscillatory mechanism. However, previous in vitro patterns were attained with protein densities on the bilayer far in excess of those in vivo and failed to recapitulate the standing wave oscillations observed in vivo. Here we studied Min protein patterning at limiting MinD concentrations reflecting the in vivo conditions. We identified “burst” patterns—radially expanding and imploding binding zones of MinD, accompanied by a peripheral ring of MinE. Bursts share several features with the in vivo dynamics of the Min system including standing wave oscillations. Our data support a patterning mechanism whereby the MinD-to-MinE ratio on the membrane acts as a toggle switch: recruiting and stabilizing MinD on the membrane when the ratio is high and releasing MinD from the membrane when the ratio is low. Coupling this toggle switch behavior with MinD depletion from the cytoplasm drives a self-organized standing wave oscillator. PMID:26884160

  18. Qualitative Content Analysis

    OpenAIRE

    Philipp Mayring

    2000-01-01

    The article describes an approach of systematic, rule guided qualitative text analysis, which tries to preserve some methodological strengths of quantitative content analysis and widen them to a concept of qualitative procedure. First the development of content analysis is delineated and the basic principles are explained (units of analysis, step models, working with categories, validity and reliability). Then the central procedures of qualitative content analysis, inductive development of ca...

  19. Fluorone dyes have binding sites on both cytoplasmic and extracellular domains of Na,K-ATPase.

    Science.gov (United States)

    Havlíková, Marika; Huličiak, Miroslav; Bazgier, Václav; Berka, Karel; Kubala, Martin

    2013-02-01

    Combination of fluorescence techniques and molecular docking was used to monitor interaction of Na,K-ATPase and its large cytoplasmic loop connecting fourth and fifth transmembrane helices (C45) with fluorone dyes (i.e. eosin Y, 5(6)-carboxyeosin, rose bengal, fluorescein, and erythrosine B). Our data suggested that there are at least two binding sites for all used fluorone dyes, except of 5(6)-carboxyeosin. The first binding site is located on C45 loop, and it is sensitive to the presence of nucleotide. The other site is located on the extracellular part of the enzyme, and it is sensitive to the presence of Na(+) or K(+) ions. The molecular docking revealed that in the open conformation of C45 loop (which is obtained in the presence of ATP) all used fluorone dyes occupy position directly inside the ATP-binding pocket, while in the closed conformation (i.e. in the absence of any ligand) they are located only near the ATP-binding site depending on their different sizes. On the extracellular part of the protein, the molecular docking predicts two possible binding sites with similar binding energy near Asp897(α) or Gln69(β). The former was identified as a part of interaction site between α- and β-subunits, the latter is in contact with conserved FXYD sequence of the γ-subunit. Our findings provide structural explanation for numerous older studies, which were performed with fluorone dyes before the high-resolution structures were known. Further, fluorone dyes seem to be good probes for monitoring of intersubunit interactions influenced by Na(+) and K(+) binding. Copyright © 2012 Elsevier B.V. All rights reserved.

  20. Cytoplasmic binding and disposition kinetics of diclofenac in the isolated perfused rat liver

    Science.gov (United States)

    Weiss, Michael; Kuhlmann, Olaf; Hung, Daniel Y; Roberts, Michael S

    2000-01-01

    The binding kinetics of diclofenac to hepatocellular structures were evaluated in the perfused rat liver using the multiple indicator dilution technique and a stochastic model of organ transit time density.The single-pass, in situ rat liver preparation was perfused with buffer solution (containing 2% albumin) at 30 ml min−1. Diclofenac and [14C]-sucrose (extracellular reference) were injected simultaneously as a bolus dose into the portal vein (six experiments in three rats). An analogous series of experiments was performed with [14C]-diclofenac and [3H]-sucrose.The diclofenac outflow data were analysed using three models of intracellular distribution kinetics, assuming (1) instantaneous distribution and binding (well-mixed model), (2) ‘slow' binding at specific intracellular sites after instantaneous distribution throughout the cytosol (slow binding model), and (3) ‘slowing' of cytoplasmic diffusion due to instantaneous binding (slow diffusion model).The slow binding model provided the best description of the data. The rate constants for cellular influx and sequestration were 0.126±0.026 and 0.013±0.009 s−1, respectively. The estimated ratio of cellular initial distribution volume to extracellular volume of 2.82 indicates an almost instantaneous distribution in the cellular water space, while the corresponding ratio of 5.54 estimated for the apparent tissue distribution volume suggests a relatively high hepatocellular binding. The non-instantaneous intracellular equilibration process was characterized by time constants of the binding and unbinding process of 53.8 and 49.5 s, respectively. The single-pass availability of diclofenac was 86%. The results obtained with [14C]-diclofenac and [3H]-sucrose were not statistically different. PMID:10903973