Introduction: A considerable proportion of mammalian gene expression undergoes circadian oscillations. Post-transcriptional mechanisms likely make important contributions to mRNA abundance rhythms. Aim: We have investigated how microRNAs contribute to core clock and clock-controlled gene expression using mice in which microRNA biogenesis can be inactivated in the liver. Results: While the hepatic core clock was surprisingly resilient to microRNA loss, whole transcriptome sequencing uncovered widespread effects on clock ouput gene expression. Cyclic transcription paired with microRNA-mediated regulation was thus identified as a widespread phenomenon that affected up to 30% of the rhythmic transcriptome and served to post-transcriptionally adjust the phases and amplitudes of rhythmic mRNA accumulation. However, only a few mRNA rhythms were actually generated by microRNAs. Finally, we pinpoint several microRNAs predicted to act as modulators of rhythmic transcripts, and identify rhythmic pathways particularly prone to microRNA regulation. Conclusion: Our study provides a comprehensive analysis of miRNA activity in shaping hepatic circadian gene expression and can serve as a valuable resource for further investigations into the regulatory roles that miRNAs play in liver gene expression and physiology. Overall design: RNA-Seq of rRNA-depleted total RNAs from two independent full time series around-the-clock of Dicer knockout and control mouse livers
MicroRNAs shape circadian hepatic gene expression on a transcriptome-wide scale.
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View SamplesBACKGROUND: The daily gene expression oscillations that underlie mammalian circadian rhythms show striking differences between tissues and involve post-transcriptional regulation. Both aspects remain poorly understood. We have used ribosome profiling to explore the contribution of translation efficiency to temporal gene expression in kidney, and contrasted our findings with liver data available from the same mice. RESULTS: Rhythmic translation of constantly abundant mRNAs affects largely nonoverlapping transcript sets with distinct phase clustering in the two organs. Moreover, tissue differences in translation efficiency modulate the timing and amount of protein biosynthesis from rhythmic mRNAs, consistent with organ-specificity in clock output gene repertoires and rhythmicity parameters. Our comprehensive datasets provided insights into translational control beyond temporal regulation. Between tissues, many transcripts show differences in translation efficiency, which are, however, of markedly smaller scale than mRNA abundance differences. Tissue-specific changes in translation efficiency are associated with specific transcript features and, intriguingly, globally counteracted and compensated transcript abundance variations, leading to higher similarity at the level of protein biosynthesis between both tissues. CONCLUSIONS: We show that tissue-specificity in rhythmic gene expression extends to the translatome and contributes to define the identities, the phases and the expression levels of rhythmic protein biosynthesis. Moreover, translational compensation of transcript abundance divergence leads to overall higher similarity at the level of protein production across organs. The unique resources provided through our study will serve to address fundamental questions of post-transcriptional control and differential gene expression in vivo. Overall design: A total of 48 mice were entrained under 12hours light:dark conditions for 2 weeks and also collected under 12hours light:dark. Mice were sacrificed every two hours during the 24 hours daily cycle. Two replicates per time point, each replicate is a pool of livers or kidneys from 2 animals.
Translational contributions to tissue specificity in rhythmic and constitutive gene expression.
Sex, Cell line, Subject, Time
View SamplesThrough post-transcriptional regulation of gene expression, miRNAs affect numerous regulatory pathways including those crucial for maintaining metabolic balance. Here we demonstrate that a neuronal-specific inhibition of miRNA maturation in adult mice leads to a rapid development of severe obesity, which is equally rapidly reversed. Development of obesity was associated with increased food intake and efficiency, and decreased locomotor activity. The ensuing decrease in body weight resembled a catabolic state with lowered O2-consumption and respiratory-exchange ratio. Brain transcriptome analyses in obese mice identified several obesity-related pathways including leptin, somatostatin, and nemo-like kinase signaling, as well as genes involved in feeding and appetite (e.g. Pmch, Neurotensin). A cluster of genes involved in synaptic plasticity was specifically enriched in post-obese mice that did not appear in obese mice. While other studies have identified a role for miRNAs in obesity our model is unique in that it allows for the study of processes involved in reversing obesity.
A neuron-specific deletion of the microRNA-processing enzyme DICER induces severe but transient obesity in mice.
Specimen part, Time
View SamplesRNA-Seq analysis of Treg cell subsets isolated from lungs of Il10GFPFoxp3Thy1.1 mice. Thy1.1+ Treg cells were FACS-sorted into IL-10–IL-18R–, IL-10+IL-18R– and IL10–IL-18R+ populations on day 5 following intranasal infection with 0.5 LD50 PR8-OTI influenza virus. Overall design: mRNA profiles of each Thy1.1+ Treg cell population (IL-10–IL-18R–, IL-10+IL-18R– and IL10–IL-18R+) from lungs on day 5 following influenza infection from 5 infected mice, sorted into TRIzol LS reagent.
A Distinct Function of Regulatory T Cells in Tissue Protection.
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View SamplesButyrate induces Treg via HDACi activity
Metabolites produced by commensal bacteria promote peripheral regulatory T-cell generation.
Specimen part, Treatment
View SamplesThe basic helix-loop-helix (bHLH) transcription factor hairy and enhancer of split (Hes3) is a member of the Hes/Hey gene family that regulates developmental processes in progenitor cells from various tissues. We demonstrated the Hes3 expression in mouse pancreatic tissue, suggesting it may have a role in modulating beta-cell function. We employed a transfection approach to address specific functions of Hes3. Hes3 RNA interference opposed the growth of the mouse insulinoma cell line Min6. Western blotting and PCR approaches specifically showed that Hes3 RNA interference opposes the expression of Pdx1 and insulin. Likewise, Hes3 knock down reduced evoked insulin release from Min6 cells.
Hes3 is expressed in the adult pancreatic islet and regulates gene expression, cell growth, and insulin release.
Specimen part
View SamplesWe have developed a method to generate muscle stem cells from pluripotent stem cells via teratoma formation. The goal of this study is to compare the transcriptome of a7+ VCAM+ myogenic cells derived from pluripotent stem cells versus satellite cells Overall design: RNA from a7+ VCAM+ myogenic cells derived from teratoma, transplanted muscles, E14.5 mouse embryos, and hindlimbs of 8-week-old mice. In 3 biological replicates
Skeletal Muscle Stem Cells from PSC-Derived Teratomas Have Functional Regenerative Capacity.
Cell line, Subject
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Gene expression profiling reveals aryl hydrocarbon receptor as a possible target for photobiomodulation when using blue light.
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View SamplesPhotobiomodulation (PBM) with blue light induces a biphasic dose response curve in proliferation of immortalized human keratinocytes (HaCaT), with a maximum anti-proliferative effect reached with 30min (41.4J/cm). The aim of this study was to test the photobiomodulatory effect of 41.4J/cm2 blue light irradiation on ROS production, apoptosis and gene expression at different time points after irradiation of HaCaT cells in vitro. ROS concentration was increased 30min after irradiation. However, already 1h after irradiation, cells were able to reduce ROS and balance the concentration to a normal level. The sudden increase in ROS did not damage the cells, which was demonstrated with FACS analysis where HaCaT cells did not show any sign of apoptosis after blue light irradiation. Furthermore, a time course could be seen in gene expression analysis after blue light, with an early response of stimulated genes already 1h after blue light irradiation, leading to the discovery of the aryl hydrocarbon receptor as possible target for blue light irradiation.
Gene expression profiling reveals aryl hydrocarbon receptor as a possible target for photobiomodulation when using blue light.
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View SamplesPhotobiomodulation (PBM) with blue light induces a biphasic dose response curve in proliferation of immortalized human keratinocytes (HaCaT), with a maximum anti-proliferative effect reached with 30min (41.4J/cm). The aim of this study was to test the photobiomodulatory effect of 41.4J/cm2 blue light irradiation on ROS production, apoptosis and gene expression at different time points after irradiation of HaCaT cells in vitro. ROS concentration was increased 30min after irradiation. However, already 1h after irradiation, cells were able to reduce ROS and balance the concentration to a normal level. The sudden increase in ROS did not damage the cells, which was demonstrated with FACS analysis where HaCaT cells did not show any sign of apoptosis after blue light irradiation. Furthermore, a time course could be seen in gene expression analysis after blue light, with an early response of stimulated genes already 1h after blue light irradiation, leading to the discovery of the aryl hydrocarbon receptor as possible target for blue light irradiation.
Gene expression profiling reveals aryl hydrocarbon receptor as a possible target for photobiomodulation when using blue light.
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