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  1. Article ; Online: An Effective Method for Quantifying RNA Expression of IbsC-SibC, a Type I Toxin-Antitoxin System in Escherichia coli.

    Jahanshahi, Shahrzad / Li, Yingfu

    Chembiochem : a European journal of chemical biology

    2020  Volume 21, Issue 21, Page(s) 3120–3130

    Abstract: Toxin and antitoxin (TA) systems are small genetic modules consisting of a toxin protein and an RNA or protein antitoxin. It is difficult to study their functions in a large part due to the lack of effective methods to study toxin RNAs, which usually ... ...

    Abstract Toxin and antitoxin (TA) systems are small genetic modules consisting of a toxin protein and an RNA or protein antitoxin. It is difficult to study their functions in a large part due to the lack of effective methods to study toxin RNAs, which usually exist at exceptionally low levels. Herein, we describe a sensitive reverse transcription quantitative PCR (RT-qPCR) method that is able to quantitate such RNA species. The method was directed at detection of the toxin mRNA of the ibsC-sibC TA pair, and its high specificity was validated by sequencing. The approach was used to determine relative expression of the IbsC and SibC RNAs at different cell-growth phases; this revealed an expression pattern that cannot be explained by the prevailing notion of growth stasis by the toxin and rescue by the antitoxin. The usefulness of the method was further showcased by the determination of average cellular copy numbers of the IbsC-SibC RNAs in wild-type E. coli cells and RNA abundance in E. coli cells engineered with extra copies of the ibsC-sibC genes. With a robust method to quantitate cellular small RNAs at very low concentrations, we are now equipped to study the expression of TA systems under different conditions to gain useful insights about their functions.
    MeSH term(s) Antitoxins/genetics ; Escherichia coli/genetics ; Escherichia coli Proteins/genetics ; Gene Expression Regulation, Bacterial/genetics ; RNA, Bacterial/genetics ; Reverse Transcriptase Polymerase Chain Reaction ; Toxin-Antitoxin Systems/genetics
    Chemical Substances Antitoxins ; Escherichia coli Proteins ; RNA, Bacterial
    Language English
    Publishing date 2020-07-20
    Publishing country Germany
    Document type Journal Article ; Research Support, Non-U.S. Gov't
    ZDB-ID 2020469-3
    ISSN 1439-7633 ; 1439-4227
    ISSN (online) 1439-7633
    ISSN 1439-4227
    DOI 10.1002/cbic.202000280
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  2. Article ; Online: RiboFACSeq: A new method for investigating metabolic and transport pathways in bacterial cells by combining a riboswitch-based sensor, fluorescence-activated cell sorting and next-generation sequencing.

    Ghazi, Zohaib / Jahanshahi, Shahrzad / Li, Yingfu

    PloS one

    2017  Volume 12, Issue 12, Page(s) e0188399

    Abstract: The elucidation of the cellular processes involved in vitamin and cofactor biosynthesis is a challenging task. The conventional approaches to these investigations rely on the discovery and purification of the products (i.e proteins and metabolites) of a ... ...

    Abstract The elucidation of the cellular processes involved in vitamin and cofactor biosynthesis is a challenging task. The conventional approaches to these investigations rely on the discovery and purification of the products (i.e proteins and metabolites) of a particular transport or biosynthetic pathway, prior to their subsequent analysis. However, the purification of low-abundance proteins or metabolites is a formidable undertaking that presents considerable technical challenges. As a solution, we present an alternative approach to such studies that circumvents the purification step. The proposed approach takes advantage of: (1) the molecular detection capabilities of a riboswitch-based sensor to detect the cellular levels of its cognate molecule, as a means to probe the integrity of the transport and biosynthetic pathways of the target molecule in cells, (2) the high-throughput screening ability of fluorescence-activated cell sorters to isolate cells in which only these specific pathways are disrupted, and (3) the ability of next-generation sequencing to quickly identify the genes of the FACS-sorted populations. This approach was named "RiboFACSeq". Following their identification by RiboFACSeq, the role of these genes in the presumed pathway needs to be verified through appropriate functional assays. To demonstrate the utility of our approach, an adenosylcobalamin (AdoCbl)-responsive riboswitch-based sensor was used in this study to demonstrate that RiboFACSeq can be used to track and sort cells carrying genetic mutations in known AdoCbl transport and biosynthesis genes with desirable sensitivity and specificity. This method could potentially be used to elucidate any pathway of interest, as long as a suitable riboswitch-based sensor can be created. We believe that RiboFACSeq would be especially useful for the elucidation of biological pathways in which the proteins and/or their metabolites are present at very low physiological concentrations in cells, as is the case with vitamin and cofactor biosynthesis.
    MeSH term(s) Cell Separation/methods ; Escherichia coli/metabolism ; Flow Cytometry/methods ; High-Throughput Nucleotide Sequencing/methods ; Riboswitch
    Chemical Substances Riboswitch
    Language English
    Publishing date 2017
    Publishing country United States
    Document type Journal Article
    ISSN 1932-6203
    ISSN (online) 1932-6203
    DOI 10.1371/journal.pone.0188399
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  3. Article ; Online: On a path toward a broad-spectrum anti-viral: inhibition of HIV-1 and coronavirus replication by SR kinase inhibitor harmine.

    Dahal, Subha / Clayton, Kiera / Cabral, Tyler / Cheng, Ran / Jahanshahi, Shahrzad / Ahmed, Choudhary / Koirala, Amrit / Villasmil Ocando, Alonso / Malty, Ramy / Been, Terek / Hernandez, Javier / Mangos, Maria / Shen, David / Babu, Mohan / Calarco, John / Chabot, Benoit / Attisano, Liliana / Houry, Walid A / Cochrane, Alan

    Journal of virology

    2023  Volume 97, Issue 10, Page(s) e0039623

    Abstract: Importance: This study highlights the crucial role RNA processing plays in regulating viral gene expression and replication. By targeting SR kinases, we identified harmine as a potent inhibitor of HIV-1 as well as coronavirus (HCoV-229E and multiple ... ...

    Abstract Importance: This study highlights the crucial role RNA processing plays in regulating viral gene expression and replication. By targeting SR kinases, we identified harmine as a potent inhibitor of HIV-1 as well as coronavirus (HCoV-229E and multiple SARS-CoV-2 variants) replication. Harmine inhibits HIV-1 protein expression and reduces accumulation of HIV-1 RNAs in both cell lines and primary CD4
    MeSH term(s) Humans ; Antiviral Agents/pharmacology ; Antiviral Agents/therapeutic use ; Coronavirus/drug effects ; Coronavirus/physiology ; Coronavirus Infections/drug therapy ; Harmine/pharmacology ; Harmine/therapeutic use ; HIV-1/drug effects ; HIV-1/physiology ; Virus Replication/drug effects
    Chemical Substances Antiviral Agents ; Harmine (4FHH5G48T7)
    Language English
    Publishing date 2023-09-14
    Publishing country United States
    Document type Journal Article ; Research Support, Non-U.S. Gov't
    ZDB-ID 80174-4
    ISSN 1098-5514 ; 0022-538X
    ISSN (online) 1098-5514
    ISSN 0022-538X
    DOI 10.1128/jvi.00396-23
    Database MEDical Literature Analysis and Retrieval System OnLINE

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  4. Article ; Online: Broad spectrum post-entry inhibitors of coronavirus replication: Cardiotonic steroids and monensin.

    Jahanshahi, Shahrzad / Ouyang, Hong / Ahmed, Choudhary / Zahedi Amiri, Ali / Dahal, Subha / Mao, Yu-Qian / Van Ommen, David A J / Malty, Ramy / Duan, Wenming / Been, Terek / Hernandez, Javier / Mangos, Maria / Nurtanto, Jocelyn / Babu, Mohan / Attisano, Liliana / Houry, Walid A / Moraes, Theo J / Cochrane, Alan

    Virology

    2023  Volume 589, Page(s) 109915

    Abstract: A small molecule screen identified several cardiotonic steroids (digitoxin and ouabain) and the ionophore monensin as potent inhibitors of HCoV-229E, HCoV-OC43, and SARS-CoV-2 replication with ... ...

    Abstract A small molecule screen identified several cardiotonic steroids (digitoxin and ouabain) and the ionophore monensin as potent inhibitors of HCoV-229E, HCoV-OC43, and SARS-CoV-2 replication with EC
    MeSH term(s) Humans ; Cardiac Glycosides/pharmacology ; Monensin/pharmacology ; Ouabain/pharmacology ; Coronavirus 229E, Human ; Digitoxin/pharmacology ; Antiviral Agents/pharmacology
    Chemical Substances Cardiac Glycosides ; Monensin (906O0YJ6ZP) ; Ouabain (5ACL011P69) ; Digitoxin (E90NZP2L9U) ; Antiviral Agents
    Language English
    Publishing date 2023-10-31
    Publishing country United States
    Document type Journal Article
    ZDB-ID 200425-2
    ISSN 1096-0341 ; 0042-6822
    ISSN (online) 1096-0341
    ISSN 0042-6822
    DOI 10.1016/j.virol.2023.109915
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    In stock of ZB MED Cologne/Königswinter

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    bis Jg. 2021: Bestellungen von Artikeln über das Online-Bestellformular
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  5. Article ; Online: A dual reporter system for detecting RNA interactions in bacterial cells.

    Kuryllo, Kacper / Jahanshahi, Shahrzad / Zhu, Weijia / Brown, Eric D / Li, Yingfu

    Chembiochem : a European journal of chemical biology

    2014  Volume 15, Issue 18, Page(s) 2703–2709

    Abstract: Detecting RNA-partner interactions in cells is often difficult due to a lack of suitable tools. Here we describe a dual reporter system capable of detecting intracellular interactions in which one of the partners is an RNA. The system utilizes two ... ...

    Abstract Detecting RNA-partner interactions in cells is often difficult due to a lack of suitable tools. Here we describe a dual reporter system capable of detecting intracellular interactions in which one of the partners is an RNA. The system utilizes two fluorescent proteins with similar maturation rates but distinct spectral properties, specifically cyan fluorescent protein (CFP) and yellow fluorescent protein (YFP). By placing the CFP gene upstream and the YFP gene downstream of an RNA gene of interest, the production of YFP becomes sensitive to RNA-partner interaction, whereas the synthesis of CFP is not disturbed. Therefore, the RNA-partner interaction can be simply measured by the change in the ratio of fluorescence of YFP over CFP. The utility of our approach is demonstrated through verification of three known RNA-partner interactions in the model bacterium Escherichia coli. Our two-reporter strategy should be broadly useful to the study of RNA-targeted interactions in bacteria.
    MeSH term(s) Bacterial Proteins/analysis ; Bacterial Proteins/genetics ; Bacterial Proteins/metabolism ; Base Sequence ; Cloning, Molecular ; Escherichia coli/genetics ; Escherichia coli/metabolism ; Escherichia coli Proteins/metabolism ; Fluorescent Dyes/analysis ; Fluorescent Dyes/metabolism ; Green Fluorescent Proteins/analysis ; Green Fluorescent Proteins/genetics ; Green Fluorescent Proteins/metabolism ; Luminescent Proteins/analysis ; Luminescent Proteins/genetics ; Luminescent Proteins/metabolism ; RNA, Bacterial/genetics ; RNA, Bacterial/metabolism ; Spectrometry, Fluorescence
    Chemical Substances Bacterial Proteins ; Cyan Fluorescent Protein ; Escherichia coli Proteins ; Fluorescent Dyes ; Luminescent Proteins ; RNA, Bacterial ; yellow fluorescent protein, Bacteria ; Green Fluorescent Proteins (147336-22-9)
    Language English
    Publishing date 2014-12-15
    Publishing country Germany
    Document type Journal Article ; Research Support, Non-U.S. Gov't
    ZDB-ID 2020469-3
    ISSN 1439-7633 ; 1439-4227
    ISSN (online) 1439-7633
    ISSN 1439-4227
    DOI 10.1002/cbic.201402322
    Database MEDical Literature Analysis and Retrieval System OnLINE

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