scholarly journals A novel G-quadruplex motif in the Human MET promoter region

2017 ◽  
Vol 37 (6) ◽  
Author(s):  
Jing Yan ◽  
Deming Zhao ◽  
Liping Dong ◽  
Shuang Pan ◽  
Fengjin Hao ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gene Expression Regulation ◽  
Luciferase Assay ◽  
Klenow Fragment ◽  
Regulate Gene Expression ◽  
Physiological Conditions ◽  
Quadruplex Structure ◽  
G Quadruplex ◽  
Quadruplex Formation

It is known that the guanine-rich strands in proto-oncogene promoters can fold into G-quadruplex structures to regulate gene expression. An intramolecular parallel G-quadruplex has been identified in MET promoter. It acts as a repressor in regulating MET expression. However, the full guanine-rich region in MET promoter forms a hybrid parallel/antiparallel G-quadruplex structure under physiological conditions, which means there are some antiparallel and hybrid parallel/antiparallel G-quadruplex structures in this region. In the present study, our data indicate that g3-5 truncation adopts an intramolecular hybrid parallel/antiparallel G-quadruplex under physiological conditions in vitro. The g3-5 G-quadruplex structure significantly stops polymerization by Klenow fragment in K+ buffer. Furthermore, the results of circular dichroism (CD) spectra and polymerase stop assay directly demonstrate that the G-quadruplex structure in g3-5 fragment can be stabilized by the G-quadruplex ligand TMPyP4 (5,10,15,20-tetra-(N-methyl-4-pyridyl) porphine). But the dual luciferase assay indicates TMPyP4 has no effect on the formation of g3-5 G-quadruplex in HepG2 cells. The findings in the present study will enrich our understanding of the G-quadruplex formation in proto-oncogene promoters and the mechanisms of gene expression regulation.

A multi-functional guanine derivative for studying the DNA G-quadruplex structure

The Analyst ◽  
2017 ◽  
Vol 142 (21) ◽  
pp. 4083-4088 ◽  
Author(s):  
Takumi Ishizuka ◽  
Pei-Yan Zhao ◽  
Hong-Liang Bao ◽  
Yan Xu
Keyword(s):  
Fluorescent Probe ◽  
Living Cells ◽  
Quadruplex Structure ◽  
G Quadruplex ◽  
Quadruplex Formation

A multi-functional guanine derivative, 8FG, as a G-quadruplex stabilizer, a fluorescent probe for the detection of G-quadruplex formation, and a 19F sensor for the observation of the G-quadruplex in vitro and in living cells.

scholarly journals PCIF1 catalyzes m6Am mRNA methylation to regulate gene expression

2018 ◽  
Author(s):  
Erdem Sendinc ◽  
David Valle-Garcia ◽  
Abhinav Dhall ◽  
Hao Chen ◽  
Telmo Henriques ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gene Expression Regulation ◽  
List Type ◽  
Methylation Analysis ◽  
Rna Pol Ii ◽  
Regulate Gene Expression ◽  
Evolutionarily Conserved ◽  
Mrna Methylation

SummarymRNA modifications play an important role in regulating gene expression. One of the most abundant mRNA modifications is N6,2-O-dimethyladenosine (m6Am). Here, we demonstrate that m6Am is an evolutionarily conserved mRNA modification mediated by the Phosphorylated CTD Interacting Factor 1 (PCIF1), which catalyzes m6A methylation on 2-O-methylated adenine located at the 5’ ends of mRNAs. Furthermore, PCIF1 catalyzes only 5’ m6Am methylation of capped mRNAs, but not internal m6A methylation in vitro and in vivo. Our global mRNA methylation analysis revealed that there is no crosstalk between m6Am and m6A mRNA methylation events, suggesting that m6Am is functionally distinct from m6A. Importantly, our data indicate that m6Am negatively impacts translation of methylated mRNAs by antagonizing cap binding protein eIF4E. Together, we identify the first and only human mRNA m6Am methyltransferase and demonstrate a novel mechanism of gene expression regulation through PCIF1-mediated m6Am mRNA methylation in eukaryotes.HighlightsPCIF1 is an evolutionarily conserved mRNA m6Am methyltransferaseLoss of PCIF1 leads to a complete loss of m6Am, whereas m6A level and distribution are not affectedPCIF1 mediated m6Am does not affect RNA Pol II transcription or mRNA stabilitym6Am-Exo-Seq is a robust methodology that enables global m6Am mappingm6Am suppresses cap dependent translation

Comparison of gene expression and activation of transcription factors in organoid-derived monolayer intestinal epithelial cells and organoids

2021 ◽  
Author(s):  
Yu Takahashi ◽  
Yu Inoue ◽  
Keitaro Kuze ◽  
Shintaro Sato ◽  
Makoto Shimizu ◽  
...  
Keyword(s):  
Gene Expression ◽  
Epithelial Cells ◽  
Gene Expression Regulation ◽  
Intestinal Epithelial Cells ◽  
Three Dimensional ◽  
Culture Conditions ◽  
Dependent Manner ◽  
Intestinal Epithelial

Abstract Intestinal organoids better represent in vivo intestinal properties than conventionally used established cell lines in vitro. However, they are maintained in three-dimensional culture conditions that may be accompanied by handling complexities. We characterized the properties of human organoid-derived two-dimensionally cultured intestinal epithelial cells (IECs) compared with those of their parental organoids. We found that the expression of several intestinal markers and functional genes were indistinguishable between monolayer IECs and organoids. We further confirmed that their specific ligands equally activate intestinal ligand-activated transcriptional regulators in a dose-dependent manner. The results suggest that culture conditions do not significantly influence the fundamental properties of monolayer IECs originating from organoids, at least from the perspective of gene expression regulation. This will enable their use as novel biological tools to investigate the physiological functions of the human intestine.

scholarly journals Analysis of G-quadruplexes upstream of herpesvirus miRNAs: evidence of G-quadruplex mediated regulation of KSHV miR-K12–1-9,11 cluster and HCMV miR-US33

2020 ◽  
Vol 21 (1) ◽  
Author(s):  
Shivani Kumar ◽  
Divya Choudhary ◽  
Anupam Patra ◽  
Neel Sarovar Bhavesh ◽  
Perumal Vivekanandan
Keyword(s):  
Promoter Activity ◽  
Wild Type ◽  
Regulatory Regions ◽  
Biophysical Characterization ◽  
Regulate Gene Expression ◽  
G Quadruplex ◽  
Wild Type Promoter ◽  
Shed Light

Abstract Background G-quadruplexes regulate gene expression, recombination, packaging and latency in herpesviruses. Herpesvirus-encoded miRNAs have been linked to important biological functions. The presence and the biological role of G-quadruplexes have not been studied in the regulatory regions of virus miRNA. We hypothesized that herpesvirus-encoded miRNAs are regulated by G-quadruplexes in their promoters. Results We analyzed the 1 kb regulatory regions of all herpesvirus-encoded miRNAs for the presence of putative quadruplex-forming sequences (PQS). Over two-third (67%) of the regulatory regions of herpesvirus miRNAs had atleast 1 PQS. The 200 bp region of the promoter proximal to herpesvirus miRNA is particularly enriched for PQS. We chose to study the G-quadruplex motifs in the promoters of miR-K12 cluster in Kaposi's sarcoma-associated Herpesvirus (KSHV miR-K12–1-9,11) and the miR-US33 encoded by Human Cytomegalovirus (HCMV miR-US33). Biophysical characterization indicates that the G-quadruplex motifs in the promoters of the KSHV miR-K12 cluster and the HCMV miR-US33 form stable intramolecular G-quadruplexes in vitro. Mutations disrupting the G-quadruplex motif in the promoter of the KSHV miR-K12 cluster significantly inhibits promoter activity, while those disrupting the motif in the promoter of HCMV miR-US33 significantly enhance the promoter activity as compared to that of the respective wild-type promoter. Similarly, the addition of G-quadruplex binding ligands resulted in the modulation of promoter activity of the wild-type promoters (with intact G-quadruplex) but not the mutant promoters (containing quadruplex-disrupting mutations). Conclusion Our findings highlight previously unknown mechanisms of regulation of virus-encoded miRNA and also shed light on new roles for G-quadruplexes in herpesvirus biology.

scholarly journals Yy1 regulates Senp1 contributing to AMPA receptor GluR1 expression following neuronal depolarization

2019 ◽  
Vol 26 (1) ◽  
Author(s):  
Tao Wu ◽  
Mary E. Donohoe
Keyword(s):  
Gene Expression ◽  
Ampa Receptor ◽  
Neuronal Activity ◽  
Cortical Neurons ◽  
Expression Patterns ◽  
Luciferase Assay ◽  
Regulation Mechanism ◽  
Phosphatase Inhibitors ◽  
Ampa Receptor Subunit

Abstract Background Neuronal activity-induced changes in gene expression patterns are important mediators of neuronal plasticity. Many neuronal genes can be activated or inactivated in response to neuronal depolarization. Mechanisms that activate gene transcription are well established, but activity-dependent mechanisms that silence transcription are less understood. It is also not clear what is the significance of inhibiting these genes during neuronal activity. Methods Quantitative Real Time-PCR, western blot and immunofluorescence staining were performed to examine the expression of Senp1 and GluR1 in mouse cortical neurons. The alterations of Yy1 phosphorylation upon neuronal depolarization and the interaction of Yy1 with Brd4 were studied by protein co-immunoprecipitation. The regulators of Yy1 phosphorylation were identified by phosphatase inhibitors. Chromatin immunoprecipitation, in vitro DNA binding assay, luciferase assay and gene knockdown experiments were used to validate the roles of Yy1 and its phosphorylation as well as Brd4 in regulating Senp1 expression. Results We report that neuronal depolarization deactivates the transcription of the SUMO protease Senp1, an important component regulating synaptic transmission, scaling, and plasticity, through Yy1. In un-stimulated neurons, Senp1 transcription is activated by a Yy1-Brd4 transcription factor protein complex assembled on the Senp1 promoter. Upon membrane depolarization, however, Yy1 is dephosphorylated and the Yy1-Brd4 complex is evicted from the Senp1 promoter, reducing Senp1 transcription levels. Both Yy1 and Senp1 promote the expression of AMPA receptor subunit GluR1, a pivotal component in learning and memory. Conclusions These results reveal an axis of Yy1/Brd4-Senp1 which regulates the expression of GluR1 during neuronal depolarization. This implicates a regulation mechanism in silencing gene expression upon neuronal activity.

scholarly journals Different Effects of Cisplatin and Transplatin on the Higher-Order Structure of DNA and Gene Expression

2019 ◽  
Vol 21 (1) ◽  
pp. 34 ◽  
Author(s):  
Toshifumi Kishimoto ◽  
Yuko Yoshikawa ◽  
Kenichi Yoshikawa ◽  
Seiji Komeda
Keyword(s):  
Gene Expression ◽  
Nuclear Dna ◽  
Biological Effects ◽  
Inhibitory Effect ◽  
Higher Order ◽  
Luciferase Assay ◽  
Order Structure ◽  
Force Microscopy ◽  
Order Of Magnitude

Despite the effectiveness of cisplatin as an anticancer agent, its trans-isomer, transplatin, is clinically ineffective. Although both isomers target nuclear DNA, there is a large difference in the magnitude of their biological effects. Here, we compared their effects on gene expression in an in vitro luciferase assay and quantified their effects on the higher-order structure of DNA using fluorescence microscopy (FM) and atomic force microscopy (AFM). The inhibitory effect of cisplatin on gene expression was about 7 times that of transplatin. Analysis of the fluctuation autocorrelation function of the intrachain Brownian motion of individual DNA molecules showed that cisplatin increases the spring and damping constants of DNA by one order of magnitude and these visco-elastic characteristics tend to increase gradually over several hours. Transplatin had a weaker effect, which tended to decrease with time. These results agree with a stronger inhibitory effect of cisplatin on gene expression. We discussed the characteristic effects of the two compounds on the higher-order DNA structure and gene expression in terms of the differences in their binding to DNA.

scholarly journals The Effects of Molecular Crowding on the Structure and Stability of G-Quadruplexes with an Abasic Site

2011 ◽  
Vol 2011 ◽  
pp. 1-9 ◽  
Author(s):  
Takeshi Fujimoto ◽  
Shu-ichi Nakano ◽  
Daisuke Miyoshi ◽  
Naoki Sugimoto
Keyword(s):  
Site Effects ◽  
Enthalpy Change ◽  
Molecular Crowding ◽  
Abasic Site ◽  
Structure And Stability ◽  
Quadruplex Structure ◽  
Abasic Sites ◽  
Crowding Effects ◽  
G Quadruplex ◽  
Quadruplex Formation

Both cellular environmental factors and chemical modifications critically affect the properties of nucleic acids. However, the structure and stability of DNA containing abasic sites under cell-mimicking molecular crowding conditions remain unclear. Here, we investigated the molecular crowding effects on the structure and stability of the G-quadruplexes including a single abasic site. Structural analysis by circular dichroism showed that molecular crowding by PEG200 did not affect the topology of the G-quadruplex structure with or without an abasic site. Thermodynamic analysis further demonstrated that the degree of stabilization of the G-quadruplex by molecular crowding decreased with substitution of an abasic site for a single guanine. Notably, we found that the molecular crowding effects on the enthalpy change for G-quadruplex formation had a linear relationship with the abasic site effects depending on its position. These results are useful for predicting the structure and stability of G-quadruplexes with abasic sites in the cell-mimicking conditions.

scholarly journals MicroRNA-378a-3p is overexpressed in psoriasis and modulates cell cycle arrest in keratinocytes via targeting BMP2 gene

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Wipasiri Soonthornchai ◽  
Pattarin Tangtanatakul ◽  
Kornvalee Meesilpavikkai ◽  
Virgil Dalm ◽  
Patipark Kueanjinda ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Cell Cycle Progression ◽  
Gene Expression Regulation ◽  
Mechanistic Model ◽  
Ultraviolet B ◽  
Bone Morphogenetic Protein 2 ◽  
Luciferase Assay ◽  
Cycle Arrest

AbstractPsoriasis is a chronic autoimmune skin disease driven by dysregulations at the cellular, genomic and genetic levels. MicroRNAs are key mediators of gene expression regulation. However, how microRNAs control the pathogenesis of psoriasis is still unclear. Here, we reported a significant up-regulation of miR-378a-3p (miR-378a) in skin biopsies from active psoriatic lesions while it was down-regulated after treatment with methotrexate or narrow-band ultraviolet B phototherapy. Using the keratinocyte in vitro model, we showed that miR-378a disturbed the cell cycle progression, causing cell cycle arrest at G1 phase. Transcriptomic analysis of keratinocytes with miR-378a overexpression and depletion revealed several important biological mechanisms related to inflammation and tight junction. Target mRNA transcript assessed by luciferase assay identified bone morphogenetic protein 2 as a novel target gene of miR-378a. These findings offer a mechanistic model where miR-378a contributes to the pathogenesis of psoriasis.

scholarly journals The parallel G-quadruplex structure of vertebrate telomeric repeat sequences is not the preferred folding topology under physiological conditions

2011 ◽  
Vol 39 (13) ◽  
pp. 5768-5775 ◽  
Author(s):  
Robert Hänsel ◽  
Frank Löhr ◽  
Silvie Foldynová-Trantírková ◽  
Ernst Bamberg ◽  
Lukáš Trantírek ◽  
...  
Keyword(s):  
Telomeric Repeat ◽  
Physiological Conditions ◽  
Quadruplex Structure ◽  
Repeat Sequences ◽  
G Quadruplex

scholarly journals Effect of ATRX and G-Quadruplex Formation by the VNTR Sequence on α-Globin Gene Expression

ChemBioChem ◽  
2016 ◽  
Vol 17 (10) ◽  
pp. 928-935 ◽  
Author(s):  
Yue Li ◽  
Junetha Syed ◽  
Yuki Suzuki ◽  
Sefan Asamitsu ◽  
Norifumi Shioda ◽  
...  
Keyword(s):  
Gene Expression ◽  
Globin Gene ◽  
G Quadruplex ◽  
Quadruplex Formation ◽  
Globin Gene Expression
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