Project description:MicroRNAs (miRNAs) regulate target mRNAs through a combination of translational repression and mRNA destabilization, with mRNA destabilization dominating at steady state in the few contexts examined globally. Here, we extend the global steady-state measurements to many additional mammalian contexts and find that regardless of the miRNA, cell type, growth condition or translational state, mRNA destabilization explains most (70% to >90%) miRNA-mediated repression. We also determine the relative dynamics of translational repression and mRNA destabilization for endogenous mRNAs as a miRNA is induced. Although translational repression occurs rapidly, its effect on gene expression is relatively weak, such that by the time consequential repression ensues, the effect of mRNA destabilization dominates. These results add to the fundamental understanding of miRNAs, imply that consequential miRNA-mediated repression is largely irreversible and simplify future studies, dramatically extending the known contexts and time points for which monitoring mRNA changes captures most of the direct miRNA effects. 6 samples from a variety of primary cell types
Project description:MicroRNAs (miRNAs) regulate target mRNAs through a combination of translational repression and mRNA destabilization, with mRNA destabilization dominating at steady state in the few contexts examined globally. Here, we extend the global steady-state measurements to additional mammalian contexts and find that regardless of the miRNA, cell type, growth condition, or translational state, mRNA destabilization explains most (66%–>90%) miRNA-mediated repression. We also determine the relative dynamics of translational repression and mRNA destabilization for endogenous mRNAs as a miRNA is induced. Although translational repression occurs rapidly, its effect is relatively weak, such that by the time consequential repression ensues, the effect of mRNA destabilization dominates. These results imply that consequential miRNA-mediated repression is largely irreversible and provide other insights into the nature of miRNA-mediated regulation. They also simplify future studies, dramatically extending the known contexts and time points for which monitoring mRNA changes captures most of the direct miRNA effects.
Project description:MicroRNAs (miRNAs) regulate target mRNAs through a combination of translational repression and mRNA destabilization, with mRNA destabilization dominating at steady state in the few contexts examined globally. Here, we extend the global steady-state measurements to many additional mammalian contexts and find that regardless of the miRNA, cell type, growth condition or translational state, mRNA destabilization explains most (70% to >90%) miRNA-mediated repression. We also determine the relative dynamics of translational repression and mRNA destabilization for endogenous mRNAs as a miRNA is induced. Although translational repression occurs rapidly, its effect on gene expression is relatively weak, such that by the time consequential repression ensues, the effect of mRNA destabilization dominates. These results add to the fundamental understanding of miRNAs, imply that consequential miRNA-mediated repression is largely irreversible and simplify future studies, dramatically extending the known contexts and time points for which monitoring mRNA changes captures most of the direct miRNA effects.
Project description:The cooperation of transcriptional and post-transcriptional controls to shape gene regulation is poorly understood. Here we show that a combination of two simple and non-invasive genomic techniques, coupled with kinetic mathematical modelling, afford insight into the multi-layered regulation of gene expression dynamics in response to oxidative stress in the fission yeast Schizosaccharomyces pombe. This study reveals a dominant role of transcriptional control in response to stress, and it points to the first minutes after stress induction as a critical time when control of mRNA turnover can support transcriptional control for rapid gene regulation. In addition, we uncover specialized gene expression strategies such as simultaneous transcriptional repression and mRNA destabilization for genes encoding ribosomal proteins, delayed mRNA destabilization with varying contribution of transcription for the ribosome biogenesis regulon, dominant roles of mRNA stabilisation for genes participating in protein degradation, and adjustment of mRNA turnover during stress adaptation. We also show that genes regulated independently of the Atf1p transcription factor are mainly controlled by mRNA turnover during oxidative stress.
Project description:The cooperation of transcriptional and post-transcriptional controls to shape gene regulation is poorly understood. Here we show that a combination of two simple and non-invasive genomic techniques, coupled with kinetic mathematical modelling, afford insight into the multi-layered regulation of gene expression dynamics in response to oxidative stress in the fission yeast Schizosaccharomyces pombe. This study reveals a dominant role of transcriptional control in response to stress, and it points to the first minutes after stress induction as a critical time when control of mRNA turnover can support transcriptional control for rapid gene regulation. In addition, we uncover specialized gene expression strategies such as simultaneous transcriptional repression and mRNA destabilization for genes encoding ribosomal proteins, delayed mRNA destabilization with varying contribution of transcription for the ribosome biogenesis regulon, dominant roles of mRNA stabilisation for genes participating in protein degradation, and adjustment of mRNA turnover during stress adaptation. We also show that genes regulated independently of the Atf1p transcription factor are mainly controlled by mRNA turnover during oxidative stress. <br><br>An additional file containing normalized data is available on the FTP site for this experiment.
Project description:MicroRNAs (miRNAs) are endogenous ~22-nucleotide RNAs that mediate important gene-regulatory events by pairing to the mRNAs of protein-coding genes to direct their repression. Repression of these regulatory targets leads to decreased translational efficiency and/or decreased mRNA levels, but the relative contributions of these two outcomes have been largely unknown, particularly for endogenous targets expressed at low-to-moderate levels. Here, we use ribosome profiling to measure the overall effects on protein production and compare these to simultaneously measured effects on mRNA levels. For both ectopic and endogenous miRNA regulatory interactions, lowered mRNA levels account for most (≥84%) of the decreased protein production. These results show that changes in mRNA levels closely reflect the impact of miRNAs on gene expression and indicate that destabilization of target mRNAs is the predominant reason for reduced protein output. Examine mRNA expression levels in HeLa cells transfected with miR-1 or miR-155, versus mock-transfected cells, at two different time points post-transfection.
Project description:MicroRNAs (miRNAs) are endogenous ~22-nucleotide RNAs that mediate important gene-regulatory events by pairing to the mRNAs of protein-coding genes to direct their repression. Repression of these regulatory targets leads to decreased translational efficiency and/or decreased mRNA levels, but the relative contributions of these two outcomes have been largely unknown, particularly for endogenous targets expressed at low-to-moderate levels. Here, we use ribosome profiling to measure the overall effects on protein production and compare these to simultaneously measured effects on mRNA levels. For both ectopic and endogenous miRNA regulatory interactions, lowered mRNA levels account for most (≥84%) of the decreased protein production. These results show that changes in mRNA levels closely reflect the impact of miRNAs on gene expression and indicate that destabilization of target mRNAs is the predominant reason for reduced protein output. Examine ribosome footprints and mRNA abundance of HeLa cells transfected with miR-1 or miR-155, versus mock-transfected cells, at two different time points post-transfection. Supplementary processed data files linked below. mir155_summaryTable.txt: log2 fold changes (miR-155-transfected versus mock-transfected HeLa cells, 32hr). mir1_summaryTable.txt: log2 fold changes (miR-1-transfected versus mock-transfected HeLa cells, 32hr).
Project description:Translation and mRNA degradation are intimately connected, yet the mechanisms that regulate them are not fully understood. Here we examine the regulation of translation and mRNA stability in mouse embryonic stem cells (ESCs) and during differentiation. In contrast to previous reports, we found that transcriptional changes account for most of the molecular changes during ESC differentiation. Within ESCs translation level and mRNA stability are positively correlated. The RNA-binding protein DDX6 has been implicated in processes involving both translational repression and mRNA destabilization; in yeast DDX6 connects codon optimality and mRNA stability and in mammals DDX6 is involved in microRNA-mediated repression. We generated DDX6 KO ESCs and found that while there was minimal connection between codon usage and stability changes, the loss of DDX6 leads to the translational depression of microRNA targets. Surprisingly, the translational derepression of microRNA targets occurs without affecting mRNA stability. Furthermore, DDX6 KO ESCs share overlapping phenotypes and global molecular changes with ESCs that completely lack all microRNAs. Together our results demonstrate that the loss of DDX6 decouples the two forms of microRNA induced repression and emphasize that translational repression by microRNAs is underappreciated.