Alphaherpesvirus-mediated remodeling of the cellular transcriptome results in depletion of m6A-containing transcripts.

The mechanisms by which viruses regulate host mRNAs during infection are still poorly understood. Several host transcripts that encode proteins that contribute to the anti-viral response contain the N6-methyladenosine nucleotide (m6A). In this study, we investigated if and how viruses from different (sub) families specifically affect m6A-containing host transcripts.

Alphaherpesvirus US3 protein-mediated inhibition of the m6A mRNA methyltransferase complex.

Chemical modifications of mRNA, the so-called epitranscriptome, represent an additional layer of post-transcriptional regulation of gene expression. The most common epitranscriptomic modification, N6-methyladenosine (m6A), is generated by a multi-subunit methyltransferase complex. We show that alphaherpesvirus kinases trigger phosphorylation of several components of the m6A methyltransferase complex, including METTL3, METTL14, and WTAP, which correlates with inhibition of the complex and a near complete loss of m6A levels in mRNA of virus-infected cells.

Repurposing an adenine riboswitch into a fluorogenic imaging and sensing tag.

Fluorogenic RNA aptamers are used to genetically encode fluorescent RNA and to construct RNA-based metabolite sensors. Unlike naturally occurring aptamers that efficiently fold and undergo metabolite-induced conformational changes, fluorogenic aptamers can exhibit poor folding, which limits their cellular fluorescence. To overcome this, we evolved a naturally occurring well-folded adenine riboswitch into a fluorogenic aptamer.

Antibody cross-reactivity accounts for widespread appearance of mA in 5'UTRs.

N-methyladenosine (mA) was proposed to be a highly prevalent modification in mRNA 5'UTRs based on mapping studies using an mA-binding antibody. We developed a bioinformatic approach to discover mA and other modifications in mRNA throughout the transcriptome by analyzing preexisting ultra-deep RNA-Seq data for modification-induced misincorporations. Using this approach, we detected appreciable levels of mA only in one mRNA: the mitochondrial MT-ND5 transcript.

mA enhances the phase separation potential of mRNA.

N-methyladenosine (mA) is the most prevalent modified nucleotide in mRNA, with around 25% of mRNAs containing at least one mA. Methylation of mRNA to form mA is required for diverse cellular and physiological processes. Although the presence of mA in an mRNA can affect its fate in different ways, it is unclear how mA directs this process and why the effects of mA can vary in different cellular contexts.

HIF-1α Stabilization Increases miR-210 Eliciting First Trimester Extravillous Trophoblast Mitochondrial Dysfunction.

Preeclampsia is associated with first trimester placental dysfunction. miR-210, a small non-coding RNA, is increased in the preeclamptic placenta. The effects of elevated miR-210 on placental function remain unclear.

Clock Regulation of Metabolites Reveals Coupling between Transcription and Metabolism.

The intricate connection between the circadian clock and metabolism remains poorly understood. We used high temporal resolution metabolite profiling to explore clock regulation of mouse liver and cell-autonomous metabolism. In liver, ∼50% of metabolites were circadian, with enrichment of nucleotide, amino acid, and methylation pathways.

Mapping mA at Individual-Nucleotide Resolution Using Crosslinking and Immunoprecipitation (miCLIP).

N -methyladenosine (m6A) is the most abundant modified base in eukaryotic mRNA and has been linked to diverse effects on mRNA fate. Current m6A mapping approaches localize m6A residues to 100-200 nt-long regions of transcripts. The precise position of m6A in mRNAs cannot be identified on a transcriptome-wide level because there are no chemical methods to distinguish between m6A and adenosine.

MetaPlotR: a Perl/R pipeline for plotting metagenes of nucleotide modifications and other transcriptomic sites.

Summary: An increasing number of studies are mapping protein binding and nucleotide modifications sites throughout the transcriptome. Often, these sites cluster in certain regions of the transcript, giving clues to their function. Hence, it is informative to summarize where in the transcript these sites occur.

Deciphering the Duality of Clock and Growth Metabolism in a Cell Autonomous System Using NMR Profiling of the Secretome.

Oscillations in circadian metabolism are crucial to the well being of organism. Our understanding of metabolic rhythms has been greatly enhanced by recent advances in high-throughput systems biology experimental techniques and data analysis. In an in vitro setting, metabolite rhythms can be measured by time-dependent sampling over an experimental period spanning one or more days at sufficent resolution to elucidate rhythms.

Single-nucleotide-resolution mapping of m6A and m6Am throughout the transcriptome.

N(6)-methyladenosine (m6A) is the most abundant modified base in eukaryotic mRNA and has been linked to diverse effects on mRNA fate. Current mapping approaches localize m6A residues to transcript regions 100-200 nt long but cannot identify precise m6A positions on a transcriptome-wide level. Here we developed m6A individual-nucleotide-resolution cross-linking and immunoprecipitation (miCLIP) and used it to demonstrate that antibodies to m6A can induce specific mutational signatures at m6A residues after ultraviolet light-induced antibody-RNA cross-linking and reverse transcription.

Placental expression of miR-517a/b and miR-517c contributes to trophoblast dysfunction and preeclampsia.

Preeclampsia is a pregnancy specific hypertensive disease that confers significant maternal and fetal risks. While the exact pathophysiology of preeclampsia is unknown, it is widely accepted that placental dysfunction is mechanistically involved. Recent studies reported aberrant expression of placenta-specific microRNAs (miRNAs) in preeclampsia including miR-517a/b and miR-517c.

Assessing the prevalence of mycoplasma contamination in cell culture via a survey of NCBI's RNA-seq archive.

Mycoplasmas are notorious contaminants of cell culture and can have profound effects on host cell biology by depriving cells of nutrients and inducing global changes in gene expression. Over the last two decades, sentinel testing has revealed wide-ranging contamination rates in mammalian culture. To obtain an unbiased assessment from hundreds of labs, we analyzed sequence data from 9395 rodent and primate samples from 884 series in the NCBI Sequence Read Archive.

Extensive variation in chromatin states across humans.

The majority of disease-associated variants lie outside protein-coding regions, suggesting a link between variation in regulatory regions and disease predisposition. We studied differences in chromatin states using five histone modifications, cohesin, and CTCF in lymphoblastoid lines from 19 individuals of diverse ancestry. We found extensive signal variation in regulatory regions, which often switch between active and repressed states across individuals.

miR-210 inhibits trophoblast invasion and is a serum biomarker for preeclampsia.

Preeclampsia is characterized by hypertension and proteinuria in pregnant women. Its exact cause is unknown. Preeclampsia increases the risk of maternal and fetal morbidity and mortality.

A functional genomics screen for microRNA regulators of NF-kappaB signaling.

Background: The nuclear factor-KappaB (NF-κB) pathway is conserved from fruit flies to humans and is a key mediator of inflammatory signaling. Aberrant regulation of NF-κB is associated with several disorders including autoimmune disease, chronic inflammation, and cancer, making the NF-κB pathway an attractive therapeutic target. Many regulatory components of the NF-κB pathway have been identified, including microRNAs (miRNAs).

Adhesion regulates MAP kinase/ternary complex factor exchange to control a proliferative transcriptional switch.

Background: The ternary complex factors (TCFs; Elk1, Net, and Sap-1) are growth factor-responsive transcription cofactors of serum response factor (SRF) and are activated by MAP kinase (MAPK) phosphorylation to regulate immediate early gene transcription. Although cell adhesion also can regulate immediate early genes and proliferation, the mechanism for this effect has remained unexplored.

Results: Restricting adhesion and spreading of G(0)-synchronized cells on substrates with decreasing size of micropatterned islands of fibronectin suppressed serum-induced immediate early gene expression and S phase entry.