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  1. Article ; Online: CRISPR Interference (CRISPRi) Mediated Suppression of OmpR Gene in E. coli: An Alternative Approach to Inhibit Biofilm.

    Zuberi, Azna / Azam, Mohd W / Khan, Asad U

    Current microbiology

    2022  Volume 79, Issue 3, Page(s) 78

    Abstract: Biofilm plays an important role in the community and hospital-acquired infections. Especially E. coli biofilm that contributes towards the significant part of medical devices associated with microbial infections. OmpR/EnvZ, a two-component system, is one ...

    Abstract Biofilm plays an important role in the community and hospital-acquired infections. Especially E. coli biofilm that contributes towards the significant part of medical devices associated with microbial infections. OmpR/EnvZ, a two-component system, is one of the regulatory mechanisms involved in transcription regulation in response to environmental osmolarity changes. The main objective of this study was to elucidate the role of the OmpR/EnvZ two-component system in regulating the biofilm through curli and fimbriae (FimH gene), a contrary approach towards biofilm inhibition. In this study, the CRISPRi technique was used to suppress the expression of the OmpR gene. The RT-PCR assay was performed to quantify mRNA gene expression of curli and biofilm producing genes, and the data were further confirmed by different microscopic, spectroscopic and biofilm quantification assay (Crystal Violet). It is the first time we have shown downregulation of the OmpR gene in biofilm causing clinical isolates of E. coli, which further suppressed the FimH gene, leading to biofilm reduction. The crystal violet assay and microscopic studies also confirmed the biofilm reduction. We conclude that the OmpR gene of the OmpR/EnvZ two-component system could be one of the targets for biofilm mediated infection intervention. Our findings open new vistas to explore the pathways and targets to control biofilm mediated infections.
    MeSH term(s) Bacterial Proteins/genetics ; Biofilms ; Clustered Regularly Interspaced Short Palindromic Repeats ; Escherichia coli/genetics ; Escherichia coli Proteins/genetics
    Chemical Substances Bacterial Proteins ; Escherichia coli Proteins
    Language English
    Publishing date 2022-01-29
    Publishing country United States
    Document type Journal Article
    ZDB-ID 134238-1
    ISSN 1432-0991 ; 0343-8651
    ISSN (online) 1432-0991
    ISSN 0343-8651
    DOI 10.1007/s00284-021-02760-x
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  2. Article: CRISPR Interference (CRISPRi) Mediated Suppression of OmpR Gene in E. coli: An Alternative Approach to Inhibit Biofilm

    Zuberi, Azna / Azam, Mohd W. / Khan, Asad U.

    Current microbiology. 2022 Mar., v. 79, no. 3

    2022  

    Abstract: Biofilm plays an important role in the community and hospital-acquired infections. Especially E. coli biofilm that contributes towards the significant part of medical devices associated with microbial infections. OmpR/EnvZ, a two-component system, is one ...

    Abstract Biofilm plays an important role in the community and hospital-acquired infections. Especially E. coli biofilm that contributes towards the significant part of medical devices associated with microbial infections. OmpR/EnvZ, a two-component system, is one of the regulatory mechanisms involved in transcription regulation in response to environmental osmolarity changes. The main objective of this study was to elucidate the role of the OmpR/EnvZ two-component system in regulating the biofilm through curli and fimbriae (FimH gene), a contrary approach towards biofilm inhibition. In this study, the CRISPRi technique was used to suppress the expression of the OmpR gene. The RT-PCR assay was performed to quantify mRNA gene expression of curli and biofilm producing genes, and the data were further confirmed by different microscopic, spectroscopic and biofilm quantification assay (Crystal Violet). It is the first time we have shown downregulation of the OmpR gene in biofilm causing clinical isolates of E. coli, which further suppressed the FimH gene, leading to biofilm reduction. The crystal violet assay and microscopic studies also confirmed the biofilm reduction. We conclude that the OmpR gene of the OmpR/EnvZ two-component system could be one of the targets for biofilm mediated infection intervention. Our findings open new vistas to explore the pathways and targets to control biofilm mediated infections.
    Keywords Escherichia coli ; biofilm ; fimbriae ; gene expression ; genes ; gentian violet ; microbiology ; osmolarity ; spectroscopy ; transcription (genetics)
    Language English
    Dates of publication 2022-03
    Size p. 78.
    Publishing place Springer US
    Document type Article
    ZDB-ID 134238-1
    ISSN 1432-0991 ; 0343-8651
    ISSN (online) 1432-0991
    ISSN 0343-8651
    DOI 10.1007/s00284-021-02760-x
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  3. Article: bolA

    Azam, Mohd W / Zuberi, Azna / Khan, Asad U

    Journal of biological research (Thessalonike, Greece)

    2020  Volume 27, Page(s) 10

    Abstract: Background: Biofilm formation is a complex phenomenon of bacterial cells, involved in several human infections. Its formation is regulated and controlled by several protein factors. The BolA-like proteins (: Methods: Knockdown mutants of : Results!# ...

    Abstract Background: Biofilm formation is a complex phenomenon of bacterial cells, involved in several human infections. Its formation is regulated and controlled by several protein factors. The BolA-like proteins (
    Methods: Knockdown mutants of
    Results: The
    Conclusions: Curli fibers and fimbriae are found to be involved in biofilm formation leading to the pathogenicity of the bacterial cell. BolA is a conserved protein and is found to play a significant role in curli and fimbriae formation in
    Language English
    Publishing date 2020-06-17
    Publishing country Greece
    Document type Journal Article
    ZDB-ID 2186880-3
    ISSN 2241-5793 ; 1790-045X
    ISSN (online) 2241-5793
    ISSN 1790-045X
    DOI 10.1186/s40709-020-00120-7
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  4. Article ; Online: bolA gene involved in curli amyloids and fimbriae production in E. coli

    Mohd W. Azam / Azna Zuberi / Asad U. Khan

    Journal of Biological Research - Thessaloniki, Vol 27, Iss 1, Pp 1-

    exploring pathways to inhibit biofilm and amyloid formation

    2020  Volume 12

    Abstract: Abstract Background Biofilm formation is a complex phenomenon of bacterial cells, involved in several human infections. Its formation is regulated and controlled by several protein factors. The BolA-like proteins (bolA gene) are conserved in both ... ...

    Abstract Abstract Background Biofilm formation is a complex phenomenon of bacterial cells, involved in several human infections. Its formation is regulated and controlled by several protein factors. The BolA-like proteins (bolA gene) are conserved in both prokaryotes and eukaryotes. The BolA protein is a transcription factor involved in bacterial cell motility and biofilm formation. This study was initiated to elucidate the role of the bolA gene in the curli biogenesis and amyloid production as well as to observe changes in the expression of fimH, a fimbriae gene. Methods Knockdown mutants of Escherichia coli MG1655 bolA gene (bolA-KD) were generated using CRISPR interference. The results obtained, were validated through gene expression using RT-PCR, microscopic analysis and different biofilm and amyloid assays. Results The bolA knockdown mutants showed a decrement in curli amyloid fibers, in fimbriae production and biofilm formation. We have also observed a reduction in EPS formation, eDNA production and extracellular protein content. Gene expression data showed that bolA downregulation caused the suppression of csgA and csgD of curli that led to the reduction in curli fiber and the amyloid formation and also the suppression of fimH, leading to the loss of fimbriae. Conclusions Curli fibers and fimbriae are found to be involved in biofilm formation leading to the pathogenicity of the bacterial cell. BolA is a conserved protein and is found to play a significant role in curli and fimbriae formation in E. coli. This study further proved that CRISPRi mediated suppression of the bolA gene leads to inhibition of biofilm formation through curli and fimbriae inhibition. Hence, it may be proposed as a possible target for intervention of biofilm mediated infections.
    Keywords Escherichia coli ; Biofilm ; bolA ; CRISPRi ; dCas9 ; Extra polymeric substance (EPS) ; Biology (General) ; QH301-705.5
    Subject code 572
    Language English
    Publishing date 2020-06-01T00:00:00Z
    Publisher Aristotle University of Thessaloniki
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  5. Article ; Online: CRISPR Interference (CRISPRi) Inhibition of luxS Gene Expression in

    Zuberi, Azna / Misba, Lama / Khan, Asad U

    Frontiers in cellular and infection microbiology

    2017  Volume 7, Page(s) 214

    Abstract: Biofilm is a sessile bacterial accretion embedded in self-producing matrix. It is the root cause of about 80% microbial infections in human. Among them, ...

    Abstract Biofilm is a sessile bacterial accretion embedded in self-producing matrix. It is the root cause of about 80% microbial infections in human. Among them,
    Language English
    Publishing date 2017
    Publishing country Switzerland
    Document type Journal Article
    ZDB-ID 2619676-1
    ISSN 2235-2988 ; 2235-2988
    ISSN (online) 2235-2988
    ISSN 2235-2988
    DOI 10.3389/fcimb.2017.00214
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  6. Article ; Online: MED12 mutation activates the tryptophan/kynurenine/AHR pathway to promote growth of uterine leiomyomas.

    Zuberi, Azna / Huang, Yongchao / Dotts, Ariel J / Wei, Helen / Coon, John S / Liu, Shimeng / Iizuka, Takashi / Wu, Olivia / Sotos, Olivia / Saini, Priyanka / Chakravarti, Debabrata / Boyer, Thomas G / Dai, Yang / Bulun, Serdar E / Yin, Ping

    JCI insight

    2023  Volume 8, Issue 18

    Abstract: Uterine leiomyomas cause heavy menstrual bleeding, anemia, and pregnancy loss in millions of women worldwide. Driver mutations in the transcriptional mediator complex subunit 12 (MED12) gene in uterine myometrial cells initiate 70% of leiomyomas that ... ...

    Abstract Uterine leiomyomas cause heavy menstrual bleeding, anemia, and pregnancy loss in millions of women worldwide. Driver mutations in the transcriptional mediator complex subunit 12 (MED12) gene in uterine myometrial cells initiate 70% of leiomyomas that grow in a progesterone-dependent manner. We showed a distinct chromatin occupancy landscape of MED12 in mutant MED12 (mut-MED12) versus WT-MED12 leiomyomas. Integration of cistromic and transcriptomics data identified tryptophan 2,3-dioxygenase (TDO2) as the top mut-MED12 target gene that was significantly upregulated in mut-MED12 leiomyomas when compared with adjacent myometrium and WT-MED12 leiomyomas. TDO2 catalyzes the conversion of tryptophan to kynurenine, an aryl hydrocarbon receptor (AHR) ligand that we confirmed to be significantly elevated in mut-MED12 leiomyomas. Treatment of primary mut-MED12 leiomyoma cells with tryptophan or kynurenine stimulated AHR nuclear translocation, increased proliferation, inhibited apoptosis, and induced AHR-target gene expression, whereas blocking the TDO2/kynurenine/AHR pathway by siRNA or pharmacological treatment abolished these effects. Progesterone receptors regulated the expression of AHR and its target genes. In vivo, TDO2 expression positively correlated with the expression of genes crucial for leiomyoma growth. In summary, activation of the TDO2/kynurenine/AHR pathway selectively in mut-MED12 leiomyomas promoted tumor growth and may inform the future development of targeted treatments and precision medicine.
    MeSH term(s) Female ; Humans ; Tryptophan ; Kynurenine/metabolism ; Uterine Neoplasms/genetics ; Uterine Neoplasms/pathology ; Tryptophan Oxygenase/genetics ; Tryptophan Oxygenase/metabolism ; Receptors, Aryl Hydrocarbon/genetics ; Receptors, Aryl Hydrocarbon/metabolism ; Leiomyoma/genetics ; Leiomyoma/metabolism ; Leiomyoma/pathology ; Mutation ; Mediator Complex/genetics ; Mediator Complex/metabolism
    Chemical Substances Tryptophan (8DUH1N11BX) ; Kynurenine (343-65-7) ; Tryptophan Oxygenase (EC 1.13.11.11) ; Receptors, Aryl Hydrocarbon ; MED12 protein, human ; Mediator Complex
    Language English
    Publishing date 2023-09-22
    Publishing country United States
    Document type Journal Article ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
    ISSN 2379-3708
    ISSN (online) 2379-3708
    DOI 10.1172/jci.insight.171305
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  7. Article: CRISPRi Induced Suppression of Fimbriae Gene (

    Zuberi, Azna / Ahmad, Nayeem / Khan, Asad U

    Frontiers in immunology

    2017  Volume 8, Page(s) 1552

    Abstract: Urinary tract infection (UTI) is one the common infections caused by the recalcitrant nature of biofilms, developed after the pathogen has adhered to the inner lining of the urinary tract. Although significant research has been made in recent years to ... ...

    Abstract Urinary tract infection (UTI) is one the common infections caused by the recalcitrant nature of biofilms, developed after the pathogen has adhered to the inner lining of the urinary tract. Although significant research has been made in recent years to control these types of infection, but as of yet, no approach has sufficiently been able to reduce the prevalence of UTIs. The main objective of this study was to prevent UTIs through targeting the fimH gene, which is the major virulent factor responsible for biofilm formation. The novelty of this work lies in the use of CRISPRi, a gene specific editing tool to control such types of infections. Accordingly, the system was designed to target
    Language English
    Publishing date 2017
    Publishing country Switzerland
    Document type Journal Article
    ZDB-ID 2606827-8
    ISSN 1664-3224
    ISSN 1664-3224
    DOI 10.3389/fimmu.2017.01552
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  8. Article ; Online: MED12 mutation activates the tryptophan/kynurenine/AHR pathway to promote growth of uterine leiomyomas

    Azna Zuberi / Yongchao Huang / Ariel J. Dotts / Helen Wei / John S. Coon V / Shimeng Liu / Takashi Iizuka / Olivia Wu / Olivia Sotos / Priyanka Saini / Debabrata Chakravarti / Thomas G. Boyer / Yang Dai / Serdar E. Bulun / Ping Yin

    JCI Insight, Vol 8, Iss

    2023  Volume 18

    Abstract: Uterine leiomyomas cause heavy menstrual bleeding, anemia, and pregnancy loss in millions of women worldwide. Driver mutations in the transcriptional mediator complex subunit 12 (MED12) gene in uterine myometrial cells initiate 70% of leiomyomas that ... ...

    Abstract Uterine leiomyomas cause heavy menstrual bleeding, anemia, and pregnancy loss in millions of women worldwide. Driver mutations in the transcriptional mediator complex subunit 12 (MED12) gene in uterine myometrial cells initiate 70% of leiomyomas that grow in a progesterone-dependent manner. We showed a distinct chromatin occupancy landscape of MED12 in mutant MED12 (mut-MED12) versus WT-MED12 leiomyomas. Integration of cistromic and transcriptomics data identified tryptophan 2,3-dioxygenase (TDO2) as the top mut-MED12 target gene that was significantly upregulated in mut-MED12 leiomyomas when compared with adjacent myometrium and WT-MED12 leiomyomas. TDO2 catalyzes the conversion of tryptophan to kynurenine, an aryl hydrocarbon receptor (AHR) ligand that we confirmed to be significantly elevated in mut-MED12 leiomyomas. Treatment of primary mut-MED12 leiomyoma cells with tryptophan or kynurenine stimulated AHR nuclear translocation, increased proliferation, inhibited apoptosis, and induced AHR-target gene expression, whereas blocking the TDO2/kynurenine/AHR pathway by siRNA or pharmacological treatment abolished these effects. Progesterone receptors regulated the expression of AHR and its target genes. In vivo, TDO2 expression positively correlated with the expression of genes crucial for leiomyoma growth. In summary, activation of the TDO2/kynurenine/AHR pathway selectively in mut-MED12 leiomyomas promoted tumor growth and may inform the future development of targeted treatments and precision medicine.
    Keywords Metabolism ; Reproductive biology ; Medicine ; R
    Subject code 570
    Language English
    Publishing date 2023-09-01T00:00:00Z
    Publisher American Society for Clinical investigation
    Document type Article ; Online
    Database BASE - Bielefeld Academic Search Engine (life sciences selection)

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  9. Article ; Online: Mono-(2-ethyl-5-hydroxyhexyl) phthalate promotes uterine leiomyoma cell survival through tryptophan-kynurenine-AHR pathway activation.

    Iizuka, Takashi / Yin, Ping / Zuberi, Azna / Kujawa, Stacy / Coon, John S / Björvang, Richelle D / Damdimopoulou, Pauliina / Pacyga, Diana C / Strakovsky, Rita S / Flaws, Jodi A / Bulun, Serdar E

    Proceedings of the National Academy of Sciences of the United States of America

    2022  Volume 119, Issue 47, Page(s) e2208886119

    Abstract: Uterine leiomyoma is the most common tumor in women and causes severe morbidity in 15 to 30% of reproductive-age women. Epidemiological studies consistently indicate a correlation between leiomyoma development and exposure to endocrine-disrupting ... ...

    Abstract Uterine leiomyoma is the most common tumor in women and causes severe morbidity in 15 to 30% of reproductive-age women. Epidemiological studies consistently indicate a correlation between leiomyoma development and exposure to endocrine-disrupting chemical phthalates, especially di-(2-ethylhexyl) phthalate (DEHP); however, the underlying mechanisms are unknown. Here, among the most commonly encountered phthalate metabolites, we found the strongest association between the urine levels of mono(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP), the principal DEHP metabolite, and the risk of uterine leiomyoma diagnosis (
    MeSH term(s) Humans ; Female ; Diethylhexyl Phthalate/toxicity ; Diethylhexyl Phthalate/urine ; Kynurenine ; Tryptophan ; Cell Survival ; Receptors, Aryl Hydrocarbon/genetics ; Receptors, Aryl Hydrocarbon/metabolism ; Large Neutral Amino Acid-Transporter 1 ; Environmental Exposure/adverse effects ; Phthalic Acids ; Leiomyoma/chemically induced ; Leiomyoma/urine ; Environmental Pollutants
    Chemical Substances phthalic acid (6O7F7IX66E) ; Diethylhexyl Phthalate (C42K0PH13C) ; Kynurenine (343-65-7) ; Tryptophan (8DUH1N11BX) ; Receptors, Aryl Hydrocarbon ; Large Neutral Amino Acid-Transporter 1 ; Phthalic Acids ; Environmental Pollutants
    Language English
    Publishing date 2022-11-14
    Publishing country United States
    Document type Journal Article ; Research Support, U.S. Gov't, Non-P.H.S. ; Research Support, N.I.H., Extramural ; Research Support, Non-U.S. Gov't
    ZDB-ID 209104-5
    ISSN 1091-6490 ; 0027-8424
    ISSN (online) 1091-6490
    ISSN 0027-8424
    DOI 10.1073/pnas.2208886119
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    In stock of ZB MED Cologne/Königswinter

    Zs.A 1148: Show issues Location:
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    Jg. 1995 - 2021: Lesesall (1.OG)
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