Base-Pair Opening Dynamics Study of Fluoride Riboswitch in the Bacillus cereus CrcB Gene

Riboswitches are segments of noncoding RNA that bind with metabolites, resulting in a change in gene expression. To understand the molecular mechanism of gene regulation in a fluoride riboswitch, a base-pair opening dynamics study was performed with and without ligands using the Bacillus cereus fluoride riboswitch. We demonstrate that the structural stability of the fluoride riboswitch is caused by two steps depending on ligands. Upon binding of a magnesium ion, significant changes in a conformation of the riboswitch occur, resulting in the greatest increase in their stability and changes in dynamics by a fluoride ion. Examining hydrogen exchange dynamics through NMR spectroscopy, we reveal that the stabilization of the U45·A37 base-pair due to the binding of the fluoride ion, by changing the dynamics while maintaining the structure, results in transcription regulation. Our results demonstrate that the opening dynamics and stabilities of a fluoride riboswitch in different ion states are essential for the genetic switching mechanism.

Download Full-text

AC-motif: a DNA motif containing adenine and cytosine repeat plays a role in gene regulation

Nucleic Acids Research ◽

10.1093/nar/gkab728 ◽

2021 ◽

Vol 49 (17) ◽

pp. 10150-10165

Author(s):

Jeong Hwan Hur ◽

Chan Young Kang ◽

Sungjin Lee ◽

Nazia Parveen ◽

Jihyeon Yu ◽

...

Keyword(s):

Gene Expression ◽

Gene Regulation ◽

Secondary Structure ◽

Base Pair ◽

Mutant Cell ◽

Magnesium Ion ◽

Stable Secondary Structure ◽

Base Pairs ◽

Regulation Effect ◽

Dna Motif

Abstract I-motif or C4 is a four-stranded DNA structure with a protonated cytosine:cytosine base pair (C+:C) found in cytosine-rich sequences. We have found that oligodeoxynucleotides containing adenine and cytosine repeats form a stable secondary structure at a physiological pH with magnesium ion, which is similar to i-motif structure, and have named this structure ‘adenine:cytosine-motif (AC-motif)’. AC-motif contains C+:C base pairs intercalated with putative A+:C base pairs between protonated adenine and cytosine. By investigation of the AC-motif present in the CDKL3 promoter (AC-motifCDKL3), one of AC-motifs found in the genome, we confirmed that AC-motifCDKL3 has a key role in regulating CDKL3 gene expression in response to magnesium. This is further supported by confirming that genome-edited mutant cell lines, lacking the AC-motif formation, lost this regulation effect. Our results verify that adenine-cytosine repeats commonly present in the genome can form a stable non-canonical secondary structure with a non-Watson–Crick base pair and have regulatory roles in cells, which expand non-canonical DNA repertoires.

Download Full-text

Roles of Regulatory RNAs in Nutritional Control

Annual Review of Nutrition ◽

10.1146/annurev-nutr-122319-035633 ◽

2020 ◽

Vol 40 (1) ◽

pp. 77-104

Author(s):

Elizabeth M. McNeill ◽

Kendal D. Hirschi

Keyword(s):

Gene Expression ◽

Gene Regulation ◽

Small Rnas ◽

Noncoding Rna ◽

Small Cell ◽

Altered Expression ◽

Nutritional Interventions ◽

Metabolic Genes ◽

Regulatory Rnas ◽

Dietary Bioactive Compounds

Small RNAs (sRNAs), including microRNAs (miRNAs), are noncoding RNA (ncRNA) molecules involved in gene regulation. sRNAs play important roles in development; however, their significance in nutritional control and as metabolic modulators is still emerging. The mechanisms by which diet impacts metabolic genes through miRNAs remain an important area of inquiry. Recent work has established how miRNAs are transported in body fluids often within exosomes, which are small cell-derived vesicles that function in intercellular communication. The abundance of other recently identified ncRNAs and new insights regarding ncRNAs as dietary bioactive compounds could remodel our understanding about how foods impact gene expression. Although controversial, some groups have shown that dietary RNAs from plants and animals (i.e., milk) are functional in consumers. In the future, regulating sRNAs either directly through dietary delivery or indirectly by altered expression of endogenous sRNA may be part of nutritional interventions for regulating metabolism.

Download Full-text

Epigenetics and Noncoding RNA – Principles and Clinical Impact

Osteologie/Osteology ◽

10.1055/a-1527-4585 ◽

2021 ◽

Vol 30 (03) ◽

pp. 201-210

Author(s):

Uwe Kornak ◽

Oliver Bischof ◽

Eric Hesse ◽

Franz Jakob ◽

Regina Ebert ◽

...

Keyword(s):

Gene Expression ◽

Transcriptional Regulation ◽

Dna Sequence ◽

Base Pair ◽

Noncoding Rna ◽

Chromatin Condensation ◽

Regulatory Elements ◽

Diagnostic Tools ◽

Chromatin Looping ◽

Pair Sequence

AbstractGenetics studies the inheritance of genetic information encoded by the base pair sequence and its variants. Sequence variants can have severe consequences as seen in genetically inherited diseases (e. g. osteogenesis Imperfecta, hypophosphatasia). On the other hand, epigenetics deals with inherited and dynamically reversible modifications of chromatin without changing the base pair sequence, resulting in a change in phenotype without a change in genotype. These modifications primarily exert their effects by influencing gene expression. Initially, the definition of epigenetics exclusively comprised inherited changes that persist across several generations without changes in the DNA sequence. This definition has been extended to include also dynamic and partially reversible changes that occur more short-term. These gene modulatory effects introduce new levels of complexity and are crucial determinants of cell fate and organismal development. With its length of approximately two meters, human DNA has to be compacted to fit into the nuclei and fulfill its functions. DNA is wrapped around histone octamers into so-called nucleosomes. DNA, histones, and other DNA-associated proteins together form what is called chromatin. DNA packaging is achieved by variable degrees of chromatin condensation depending on cell type and context. Epigenetic transcriptional regulation modifies the affinity and accessibility of cis-regulatory elements (CREs) for transcription factors and the basic transcriptional machinery and governs interaction between CREs. CREs include promoters, enhancers, silencers, and insulators and are potent modulators of gene expression impacting core cell biological processes such as proliferation and differentiation. Chromatin looping and remodeling by differential covalent modifications of DNA (e. g., methylation or hydroxylation) and histone tails (e. g., acetylation or methylation) elicit fundamental changes in CRE accessibility, thus impacting gene expression. Chromatin looping depends on a specialized machinery including cohesins. Chromatin modifications are mediated by specific enzymes like DNA methylases (DNMTs), histone-modifying enzymes, like histone methyl- and acetyltransferases (KMTs, HATs/KATs), and histone demethylases and deacetylases (KDMs, HDACs). It becomes increasingly evident that epigenetic (dys)regulation plays a decisive role in physiology and pathophysiology, impacting many age-related diseases like cancer and degenerative pathologies (e. g., osteoporosis, Alzheimer’s, or Parkinson’s) in a significant fashion. Recently, small-molecule inhibitors of chromatin-modifying enzymes (e. g., vorinostat) have been identified and successfully introduced in therapy. Significant progress in high-throughput sequencing technologies and big data analysis has broadened our understanding of noncoding (nc) RNAs and DNA sequence regions in (post-)transcriptional regulation and disease development. Among ncRNAs that play vital roles in gene expression are micro- (miRs) and long noncoding RNAs (lncRNAs; e. g., XIST or HOTAIR). By interacting with the coding genome, these RNAs modulate important genetic programs. Interfering RNAs can, for example, enhance the post-transcriptional degradation of transcripts, altering their translation, or assist in the recruitment of chromatin-modifying enzymes to regulate transcription. They can also be packaged into extracellular vesicles as cargo and thus deliver critical information to the microenvironment or even systemically to distant tissues. Therefore, ncRNAs represent a novel playground for therapeutical investigations and supplement epigenetic mechanisms of gene regulation while being subject to epigenetic regulation themselves. Last but not least, dysregulated ncRNAs can also propagate disease. Until recently, the detection of epigenetic phenomena necessitated invasive diagnostic interventions. However, with the arrival of so-called “liquid biopsies” an analysis of circulating cell-free DNA fragments (cfDNA) and RNAs as well as vesicle-packed RNAs through minimal invasively drawn blood samples can be obtained. Such “fragmentomics” and RNAomics approaches on peripheral blood will ultimately serve as diagnostic tools for personalized clinical interventions.

Download Full-text

Noncoding RNA participation in gene expression regulation in yeast Saccharomyces cerevisiae

Ecological genetics ◽

10.17816/ecogen913-14 ◽

2011 ◽

Vol 9 (1) ◽

pp. 3-14

Author(s):

Olga V Kochenova

Keyword(s):

Gene Expression ◽

Saccharomyces Cerevisiae ◽

Gene Regulation ◽

Antisense Rna ◽

Noncoding Rna ◽

Gene Expression Regulation ◽

Regulation Of Gene Expression ◽

Yeast Genome ◽

Yeast Saccharomyces Cerevisiae ◽

Main Components

Saccharomyces cerevisiae lacks the main components of RNAi-dependent gene silencing. Nevertheless, regulation of gene expression in S. cerevisiae could be accomplished via some other types of noncoding RNA, particularly via antisense RNA. Although, there is a high percent of untranslated RNA in yeast genome only few evidences of noncoding RNA gene regulation exist in yeast S. cerevisiae, some of them are reviewed in the present paper.

Download Full-text

Faculty Opinions recommendation of A long noncoding RNA maintains active chromatin to coordinate homeotic gene expression.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.9550956.10207054 ◽

2011 ◽

Author(s):

Pamela Geyer

Keyword(s):

Gene Expression ◽

Long Noncoding Rna ◽

Noncoding Rna ◽

Homeotic Gene ◽

Active Chromatin ◽

Homeotic Gene Expression

Download Full-text

The Therapeutic Potential and Role of miRNA, lncRNA, and circRNA in Osteoarthritis

Current Gene Therapy ◽

10.2174/1566523219666190716092203 ◽

2019 ◽

Vol 19 (4) ◽

pp. 255-263 ◽

Cited By ~ 13

Author(s):

Yuangang Wu ◽

Xiaoxi Lu ◽

Bin Shen ◽

Yi Zeng

Keyword(s):

Gene Expression ◽

Gene Therapy ◽

Extracellular Matrix ◽

Inflammatory Reaction ◽

Noncoding Rna ◽

Rna Binding ◽

Rna Binding Proteins ◽

Protein Translation ◽

Cochrane Library

Background: Osteoarthritis (OA) is a disease characterized by progressive degeneration, joint hyperplasia, narrowing of joint spaces, and extracellular matrix metabolism. Recent studies have shown that the pathogenesis of OA may be related to non-coding RNA, and its pathological mechanism may be an effective way to reduce OA. Objective: The purpose of this review was to investigate the recent progress of miRNA, long noncoding RNA (lncRNA) and circular RNA (circRNA) in gene therapy of OA, discussing the effects of this RNA on gene expression, inflammatory reaction, apoptosis and extracellular matrix in OA. Methods: The following electronic databases were searched, including PubMed, EMBASE, Web of Science, and the Cochrane Library, for published studies involving the miRNA, lncRNA, and circRNA in OA. The outcomes included the gene expression, inflammatory reaction, apoptosis, and extracellular matrix. Results and Discussion: With the development of technology, miRNA, lncRNA, and circRNA have been found in many diseases. More importantly, recent studies have found that RNA interacts with RNA-binding proteins to regulate gene transcription and protein translation, and is involved in various pathological processes of OA, thus becoming a potential therapy for OA. Conclusion: In this paper, we briefly introduced the role of miRNA, lncRNA, and circRNA in the occurrence and development of OA and as a new target for gene therapy.

Download Full-text

OP0248 DEVELOPMENT OF A 3D SKIN MICROTISSUE MODEL FOR FIBROTIC DISEASES

Annals of the Rheumatic Diseases ◽

10.1136/annrheumdis-2021-eular.3306 ◽

2021 ◽

Vol 80 (Suppl 1) ◽

pp. 151.2-152

Author(s):

E. Pachera ◽

G. Kania ◽

A. Juengel ◽

M. Calcagni ◽

O. Distler

Keyword(s):

Gene Expression ◽

Noncoding Rna ◽

Clinical Investigation ◽

3D Culture ◽

Human Keratinocytes ◽

Dermal Fibroblasts ◽

Proof Of Concept ◽

Research Support ◽

Preclinical Research ◽

Matrix Gene

Background:Traditional preclinical approaches, such as two-dimensional cell culture and animal models, are often inadequate to mimic the pathophysiological features of complex diseases such as systemic sclerosis (SSc). Human specific targets, such as the recently described pro-fibrotic long non coding RNA (lncRNA) H19X1, are becoming increasingly relevant in preclinical research, creating the need of new strategies and tools in translational medicine. The employment of novel three-dimensional (3D) culture systems, where multiple cell types are included, is filling an important gap left by the traditional preclinical methods.Objectives:To develop an easy to produce 3D fibrotic skin microtissues model for translational proof of concept studies.Methods:Two thousand five hundred dermal fibroblasts isolated from skin of SSc patients were seeded in ultra-low attachment 96-well plates. Fibroblast were let to aggregate into spheres for 48h. Two thousand five hundred primary normal human keratinocytes were added to the culture and let to layer onto the fibroblast spheres for 72h. H19X silencing experiments were used as proof of concept studies. H19X silencing with antisense oligonucleotides or transfections with a scrambled control were performed in fibroblasts prior to the sphere formation for 24h. TGFβ (10 ng/ml) was added to microtissue to exacerbate the fibrotic phenotype. Haematoxylin eosin staining as well as immunohistochemistry staining for vimentin and cytokeratin 10 was performed. Skin microtissues were processed for RNA and protein isolation. Pro-collagen Iα1 and fibronectin were quantified in the supernatants with ELISA.Results:The microtissues presented a core of SSc fibroblast as revealed by vimentin staining and an external layer of keratinocytes as revealed by cytokeratin 10 staining, mimicking the human skin architecture. Gene expression analysis following TGFβ stimulation displayed induced expression of extracellular matrix gene COL1A1 (p=0.044) and the myofibroblast marker ACTA2 (p=0.018), indicating that the microtissues were able to develop a fibrotic response. Microtissues, where H19X was silenced, displayed reduced gene expression of COL1A1 and ACTA2 after TGFβ stimulation (COL1A1 p=0.007, ACTA2 p=0.045). Additionally, H19X silencing led to lower levels of αSMA protein expression (p=0.009) and pro-collagen1α1 secretion (p=0.039) in the supernatant of the microtissue cultures as revealed by Western Blot and ELISA, respectively. FN1 expression and fibronectin protein levels were not significantly reduced in the microtissues after H19X silencing.Conclusion:We were able to produce a 3D microtissue resembling skin architecture that can respond to fibrotic stimuli. Knockdown experiments of pro-fibrotic lncRNA H19X confirmed the potential of the model as screening platform for novel pro-fibrotic effectors. A future aim will be to optimize the model for high-throughput automated screening platforms.References:[1]Pachera, E., et al. (2020). “Long noncoding RNA H19X is a key mediator of TGF-β–driven fibrosis.” The Journal of Clinical Investigation 130(9): 4888-4905.Disclosure of Interests:Elena Pachera: None declared, Gabriela Kania: None declared, Astrid Juengel: None declared, Maurizio Calcagni Speakers bureau: Arthrex, Consultant of: Medartis, Arthrex, SilkBiomaterials, Grant/research support from: Medartis, Oliver Distler Speakers bureau: Actelion, Bayer, Boehringer Ingelheim, Medscape, Novartis, Roche, Consultant of: Abbvie, Actelion, Acceleron Pharma, Amgen, AnaMar, Arxx Therapeutics, Bayer, Baecon Discovery, Blade Therapeutics, Boehringer, CSL Behring, ChemomAb, Corpuspharma, Curzion Pharmaceuticals, Ergonex, Galapagos NV, GSK, Glenmark Pharmaceuticals, Inventiva, Italfarmaco, iQvia, -Kymera, Medac, Medscape, Mitsubishi Tanabe Pharma, MSD, Roche, Sanofi, UCB, Grant/research support from: Abbvie, Actelion, Bayer, Boehringer Ingelheim, Kymera Therapeutics, Mitsubishi Tanabe

Download Full-text

CFTR Cooperative Cis-Regulatory Elements in Intestinal Cells

International Journal of Molecular Sciences ◽

10.3390/ijms22052599 ◽

2021 ◽

Vol 22 (5) ◽

pp. 2599

Author(s):

Mégane Collobert ◽

Ozvan Bocher ◽

Anaïs Le Nabec ◽

Emmanuelle Génin ◽

Claude Férec ◽

...

Keyword(s):

Gene Expression ◽

Cystic Fibrosis ◽

Gene Regulation ◽

Genetic Diseases ◽

Regulatory Elements ◽

Cftr Gene ◽

Intestinal Cells ◽

Cooperative Effects ◽

Cis Acting ◽

Study Techniques

About 8% of the human genome is covered with candidate cis-regulatory elements (cCREs). Disruptions of CREs, described as “cis-ruptions” have been identified as being involved in various genetic diseases. Thanks to the development of chromatin conformation study techniques, several long-range cystic fibrosis transmembrane conductance regulator (CFTR) regulatory elements were identified, but the regulatory mechanisms of the CFTR gene have yet to be fully elucidated. The aim of this work is to improve our knowledge of the CFTR gene regulation, and to identity factors that could impact the CFTR gene expression, and potentially account for the variability of the clinical presentation of cystic fibrosis as well as CFTR-related disorders. Here, we apply the robust GWAS3D score to determine which of the CFTR introns could be involved in gene regulation. This approach highlights four particular CFTR introns of interest. Using reporter gene constructs in intestinal cells, we show that two new introns display strong cooperative effects in intestinal cells. Chromatin immunoprecipitation analyses further demonstrate fixation of transcription factors network. These results provide new insights into our understanding of the CFTR gene regulation and allow us to suggest a 3D CFTR locus structure in intestinal cells. A better understand of regulation mechanisms of the CFTR gene could elucidate cases of patients where the phenotype is not yet explained by the genotype. This would thus help in better diagnosis and therefore better management. These cis-acting regions may be a therapeutic challenge that could lead to the development of specific molecules capable of modulating gene expression in the future.

Download Full-text

Interaction between Host MicroRNAs and the Gut Microbiota in Colorectal Cancer

mSystems ◽

10.1128/msystems.00205-17 ◽

2018 ◽

Vol 3 (3) ◽

Cited By ~ 35

Author(s):

Ce Yuan ◽

Michael B. Burns ◽

Subbaya Subramanian ◽

Ran Blekhman

Keyword(s):

Gene Expression ◽

Colorectal Cancer ◽

Gut Microbiota ◽

Mirna Expression ◽

Gut Microbiome ◽

Noncoding Rna ◽

Microbial Community Composition ◽

Host Cells ◽

Microbiome Composition ◽

Small Noncoding Rna

ABSTRACT Although variation in gut microbiome composition has been linked with colorectal cancer (CRC), the factors that mediate the interactions between CRC tumors and the microbiome are poorly understood. MicroRNAs (miRNAs) are known to regulate CRC progression and are associated with patient survival outcomes. In addition, recent studies suggested that host miRNAs can also regulate bacterial growth and influence the composition of the gut microbiome. Here, we investigated the association between miRNA expression and microbiome composition in human CRC tumor and normal tissues. We identified 76 miRNAs as differentially expressed (DE) in tissue from CRC tumors and normal tissue, including the known oncogenic miRNAs miR-182, miR-503, and mir-17~92 cluster. These DE miRNAs were correlated with the relative abundances of several bacterial taxa, including Firmicutes , Bacteroidetes , and Proteobacteria . Bacteria correlated with DE miRNAs were enriched with distinct predicted metabolic categories. Additionally, we found that miRNAs that correlated with CRC-associated bacteria are predicted to regulate targets that are relevant for host-microbiome interactions and highlight a possible role for miRNA-driven glycan production in the recruitment of pathogenic microbial taxa. Our work characterized a global relationship between microbial community composition and miRNA expression in human CRC tissues. IMPORTANCE Recent studies have found an association between colorectal cancer (CRC) and the gut microbiota. One potential mechanism by which the microbiota can influence host physiology is through affecting gene expression in host cells. MicroRNAs (miRNAs) are small noncoding RNA molecules that can regulate gene expression and have important roles in cancer development. Here, we investigated the link between the gut microbiota and the expression of miRNA in CRC. We found that dozens of miRNAs are differentially regulated in CRC tumors and adjacent normal colon and that these miRNAs are correlated with the abundance of microbes in the tumor microenvironment. Moreover, we found that microbes that have been previously associated with CRC are correlated with miRNAs that regulate genes related to interactions with microbes. Notably, these miRNAs likely regulate glycan production, which is important for the recruitment of pathogenic microbial taxa to the tumor. This work provides a first systems-level map of the association between microbes and host miRNAs in the context of CRC and provides targets for further experimental validation and potential interventions.

Download Full-text