A Versatile Method to Design Stem-Loop Primer-Based Quantitative PCR Assays for Detecting Small Regulatory RNA Molecules

Abstract Staphylococcus aureus is an opportunistic human and animal pathogen with an arsenal of virulence factors that are tightly regulated during bacterial infection. The latter is achieved through a sophisticated network of regulatory proteins and regulatory RNAs. Here, we describe the involvement of a novel prophage-carried small regulatory S. aureus RNA, SprY, in the control of virulence genes. An MS2-affinity purification assay reveals that SprY forms a complex in vivo with RNAIII, a major regulator of S. aureus virulence genes. SprY binds to the 13th stem-loop of RNAIII, a key functional region involved in the repression of multiple mRNA targets. mRNAs encoding the repressor of toxins Rot and the extracellular complement binding protein Ecb are among the targets whose expression is increased by SprY binding to RNAIII. Moreover, SprY decreases S. aureus hemolytic activity and virulence. Our results indicate that SprY titrates RNAIII activity by targeting a specific stem loop. Thus, we demonstrate that a prophage-encoded sRNA reduces the pathogenicity of S. aureus through RNA sponge activity.

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A Novel Fur- and Iron-Regulated Small RNA, NrrF, Is Required for Indirect Fur-Mediated Regulation of the sdhA and sdhC Genes in Neisseria meningitidis

Journal of Bacteriology ◽

10.1128/jb.01890-06 ◽

2007 ◽

Vol 189 (10) ◽

pp. 3686-3694 ◽

Cited By ~ 75

Author(s):

J. R. Mellin ◽

Sulip Goswami ◽

Susan Grogan ◽

Brian Tjaden ◽

Caroline A. Genco

Keyword(s):

Neisseria Meningitidis ◽

Gene Transcription ◽

Small Rna ◽

Iron Homeostasis ◽

A Priori ◽

Regulatory Rna ◽

Rna Molecules ◽

Small Regulatory Rna ◽

Regulator Protein ◽

Microarray Studies

ABSTRACT Iron is both essential for bacterial growth and toxic at higher concentrations; thus, iron homeostasis is tightly regulated. In Neisseria meningitidis the majority of iron-responsive gene regulation is mediated by the ferric uptake regulator protein (Fur), a protein classically defined as a transcriptional repressor. Recently, however, microarray studies have identified a number of genes in N. meningitidis that are iron and Fur activated, demonstrating a new role for Fur as a transcriptional activator. Since Fur has been shown to indirectly activate gene transcription through the repression of small regulatory RNA molecules in other organisms, we hypothesized that a similar mechanism could account for Fur-dependent, iron-activated gene transcription in N. meningitidis. In this study, we used a bioinformatics approach to screen for the presence of Fur-regulated small RNA molecules in N. meningitidis MC58. This screen identified one small RNA, herein named NrrF (for neisserial regulatory RNA responsive to iron [Fe]), which was demonstrated to be both iron responsive and Fur regulated and which has a well-conserved orthologue in N. gonorrhoeae. In addition, this screen identified a number of other likely, novel small RNA transcripts. Lastly, we utilized a new bioinformatics approach to predict regulatory targets of the NrrF small RNA. This analysis led to the identification of the sdhA and sdhC genes, which were subsequently demonstrated to be under NrrF regulation in an nrrF mutant. This study is the first report of small RNAs in N. meningitidis and the first to use a bioinformatics approach to identify, a priori, regulatory targets of a small RNA.

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Development of Novel Cardiovascular Therapeutics From Small Regulatory RNA Molecules - An Outline of Key Requirements

Current Pharmaceutical Design ◽

10.2174/138161210791792813 ◽

2010 ◽

Vol 16 (20) ◽

pp. 2252-2268 ◽

Cited By ~ 8

Author(s):

W. Poller ◽

H. Fechner

Keyword(s):

Regulatory Rna ◽

Rna Molecules ◽

Small Regulatory Rna ◽

Cardiovascular Therapeutics

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Small Regulatory Molecules Acting Big in Cancer: Potential Role of Mito-miRs in Cancer

Current Molecular Medicine ◽

10.2174/1566524019666190723165357 ◽

2019 ◽

Vol 19 (9) ◽

pp. 621-631 ◽

Cited By ~ 2

Author(s):

Praveen Sharma ◽

Bharat ◽

Nilambra Dogra ◽

Sandeep Singh

Keyword(s):

Gene Expression ◽

Mitochondrial Genome ◽

Potential Role ◽

Regulatory Rna ◽

Rna Molecules ◽

Small Regulatory Rna ◽

Physiological Processes ◽

Current Review ◽

Posttranscriptional Level

MicroRNAs [miRNAs] are short, non-coding, single stranded RNA molecules regulating gene expression of their targets at the posttranscriptional level by either degrading mRNA or by inhibiting translation. Previously, miRNAs have been reported to be present inside the mitochondria and these miRNAs have been termed as mito-miRs. Origin of these mito-miRs may either be from mitochondrial genome or import from nucleus. The second class of mito-miRs makes it important to unravel the involvement of miRNAs in crosstalk between nucleus and mitochondria. Since miRNAs are involved in various physiological processes, their deregulation is often associated with disease progression, including cancer. The current review focuses on the involvement of miRNAs in different mitochondrial mediated processes. It also highlights the importance of exploring the interaction of miRNAs with mitochondrial genome, which may lead to the development of small regulatory RNA based therapeutic options.

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Measuring Expression Levels of Small Regulatory RNA Molecules from Body Fluids and Formalin-Fixed, Paraffin-Embedded Samples

RNA Mapping - Methods in Molecular Biology ◽

10.1007/978-1-4939-1062-5_10 ◽

2014 ◽

pp. 105-119 ◽

Cited By ~ 5

Author(s):

Adrienn Gyongyosi ◽

Otto Docs ◽

Zsolt Czimmerer ◽

Laszlo Orosz ◽

Attila Horvath ◽

...

Keyword(s):

Body Fluids ◽

Regulatory Rna ◽

Expression Levels ◽

Rna Molecules ◽

Small Regulatory Rna ◽

Formalin Fixed Paraffin ◽

Formalin Fixed Paraffin Embedded ◽

Formalin Fixed

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Application of Quantitative-PCR to Monitor Netpen Sites in British Columbia (Canada) for Tenacibaculum Species

Pathogens ◽

10.3390/pathogens10040414 ◽

2021 ◽

Vol 10 (4) ◽

pp. 414

Author(s):

Joseph P. Nowlan ◽

Scott R. Britney ◽

John S. Lumsden ◽

Spencer Russell

Keyword(s):

Atlantic Salmon ◽

Quantitative Pcr ◽

Salmo Salar ◽

Bacterial Load ◽

Temporal Distribution ◽

Quality Parameters ◽

Antimicrobial Use ◽

Live Fish ◽

Dead Fish ◽

Pcr Assays

Tenacibaculum are frequently detected from fish with tenacibaculosis at aquaculture sites; however, information on the ecology of these bacteria is sparse. Quantitative-PCR assays were used to detect T. maritimum and T. dicentrarchi at commercial Atlantic salmon (Salmo salar) netpen sites throughout several tenacibaculosis outbreaks. T. dicentrarchi and T. maritimum were identified in live fish, dead fish, other organisms associated with netpens, water samples and on inanimate substrates, which indicates a ubiquitous distribution around stocked netpen sites. Before an outbreak, T. dicentrarchi was found throughout the environment and from fish, and T. maritimum was infrequently identified. During an outbreak, increases in the bacterial load in were recorded and no differences were recorded after an outbreak supporting the observed recrudescence of mouthrot. More bacteria were recorded in the summer months, with more mortality events and antibiotic treatments, indicating that seasonality may influence tenacibaculosis; however, outbreaks occurred in both seasons. Relationships were identified between fish mortalities and antimicrobial use to water quality parameters (temperature, salinity, dissolved oxygen) (p < 0.05), but with low R2 values (<0.25), other variables are also involved. Furthermore, Tenacibaculum species appear to have a ubiquitous spatial and temporal distribution around stocked netpen sites, and with the potential to induce disease in Atlantic salmon, continued research is needed.

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MicroRNAs in central nervous system disorders: current advances in pathogenesis and treatment

The Egyptian Journal of Neurology Psychiatry and Neurosurgery ◽

10.1186/s41983-021-00289-1 ◽

2021 ◽

Vol 57 (1) ◽

Author(s):

Mona Hussein ◽

Rehab Magdy

Keyword(s):

Neurological Disorders ◽

Target Genes ◽

Transcriptional Regulators ◽

Therapeutic Strategies ◽

Regulatory Rna ◽

Altered Expression ◽

Rna Molecules ◽

Central Nervous System Disorders ◽

Novel Targets ◽

Lateral Sclerosis

AbstractMicroRNAs (miRNAs) are a class of short, non-coding, regulatory RNA molecules that function as post transcriptional regulators of gene expression. Altered expression of multiple miRNAs was found to be extensively involved in the pathogenesis of different neurological disorders including Alzheimer’s disease, Parkinson’s disease, stroke, epilepsy, multiple sclerosis, amyotrophic lateral sclerosis, and Huntington’s disease. miRNAs are implicated in the pathogenesis of excitotoxicity, apoptosis, oxidative stress, inflammation, neurogenesis, angiogenesis, and blood–brain barrier protection. Consequently, miRNAs can serve as biomarkers for different neurological disorders. In recent years, advances in the miRNA field led to identification of potentially novel prospects in the development of new therapies for incurable CNS disorders. MiRNA-based therapeutics include miRNA mimics and inhibitors that can decrease or increase the expression of target genes. Better understanding of the mechanisms by which miRNAs are implicated in the pathogenesis of neurological disorders may provide novel targets to researchers for innovative therapeutic strategies.

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1H, 13C and 15N assignment of stem-loop SL1 from the 5'-UTR of SARS-CoV-2

Biomolecular NMR Assignments ◽

10.1007/s12104-021-10047-2 ◽

2021 ◽

Author(s):

Christian Richter ◽

Katharina F. Hohmann ◽

Sabrina Toews ◽

Daniel Mathieu ◽

Nadide Altincekic ◽

...

Keyword(s):

Life Cycle ◽

Chemical Shift ◽

Base Pair ◽

Chemical Shift Assignment ◽

Potential Functions ◽

Regulatory Rna ◽

Structural Element ◽

Stem Loop ◽

Internal Loop ◽

Shift Assignment

AbstractThe stem-loop (SL1) is the 5'-terminal structural element within the single-stranded SARS-CoV-2 RNA genome. It is formed by nucleotides 7–33 and consists of two short helical segments interrupted by an asymmetric internal loop. This architecture is conserved among Betacoronaviruses. SL1 is present in genomic SARS-CoV-2 RNA as well as in all subgenomic mRNA species produced by the virus during replication, thus representing a ubiquitous cis-regulatory RNA with potential functions at all stages of the viral life cycle. We present here the 1H, 13C and 15N chemical shift assignment of the 29 nucleotides-RNA construct 5_SL1, which denotes the native 27mer SL1 stabilized by an additional terminal G-C base-pair.

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