Helicase Ded1 fine-tunes the reprogramming of gene expression conferred by inhibition of TORC1 by rapamycin.
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ABSTRACT:
ORGANISM(S): Saccharomyces cerevisiae
PROVIDER: GSE260591 | GEO | 2025/01/30
REPOSITORIES: GEO
Similar Datasets
Project description:Helicase Ded1 fine-tunes the reprogramming of gene expression conferred by inhibition of TORC1 by rapamycin.
| PRJNA1082295 | ENA
Project description:DEAD-box RNA helicases are ATP-dependent RNA binding proteins and RNA-dependent ATPases that possess weak, nonprocessive unwinding activity in vitro, but they can form long-lived complexes on RNAs when the ATPase activity is inhibited. Ded1 is a yeast DEAD-box protein, the functional ortholog of mammalian DDX3, that is considered important for the scanning efficiency of the 48S pre-initiation complex ribosomes to the AUG start codon. We used a modified PAR-CLIP technique, which we call quicktime PAR-CLIP (qtPAR-CLIP) to crosslink Ded1 to 4-thiouridine-incorported RNAs in vivo using UV light centered at 365 nm. The irradiation conditions are largely benign to the yeast cells and to Ded1, and we are able to obtain a high efficiency of crosslinking under physiological conditions. We find that Ded1 forms crosslinks on the open reading frames of many different mRNAs, but it forms the most extensive interactions on a relatively few mRNAs, and particularly on mRNAs encoding certain ribosomal proteins and translation factors. Under glucose-depletion conditions the crosslinking pattern shifts to mRNAs encoding metabolic and stress-related proteins, which reflects the altered translation. These data are consistent with Ded1 functioning in the regulation of translation elongation, perhaps by pausing or stabilizing the ribosomes through its ATP-dependent binding.
2023-08-01 | GSE228828 | GEO
Project description:DEAD-box RNA helicases eIF4A and Ded1 are believed to promote translation initiation by resolving mRNA secondary structures that impede ribosome attachment at the mRNA 5’ end or subsequent scanning of the 5’UTR, but whether they perform distinct functions or act redundantly in vivo is poorly understood. We compared the effects of mutations in Ded1 or eIF4A on global translational efficiencies (TEs) in yeast by ribosome footprint profiling. Despite similar reductions in bulk translation, inactivation of a cold-sensitive Ded1 mutant substantially reduced the TEs of >600 mRNAs, whereas inactivation of a temperature-sensitive eIF4A mutant yielded <40 similarly impaired mRNAs. The broader requirement for Ded1 did not reflect more pervasive secondary structures at low temperature, as inactivation of temperature-sensitive and cold-sensitive ded1 mutants gave highly correlated results. Interestingly, Ded1-dependent mRNAs exhibit greater than average 5’UTR length and propensity for secondary structure, implicating Ded1 in scanning though structured 5' UTRs. Reporter assays confirmed that cap- distal stem-loop insertions increase dependence on Ded1 but not eIF4A for efficient translation. While only a small fraction of mRNAs is strongly dependent on eIF4A, this dependence is significantly correlated with requirements for Ded1 and 5’UTR features characteristic of Ded1- dependent mRNAs. Our findings suggest that Ded1 is critically required to promote scanning through secondary structures within 5’UTRs; and while eIF4A cooperates with Ded1 in this function, it also promotes a step of initiation common to virtually all yeast mRNAs. We compared the effects of mutations in Ded1 or eIF4A on global translational efficiencies (TEs) in yeast by ribosome footprint profiling.The study includes 32 samples, comprised of 16 mRNA-Seq samples and 16 ribosome footprint profiling samples, derived from biological replicates of 3 mutant strains, ded1-cs, ded1-ts and tif1-ts, and the corresponding wild-type strains. The tif1-ts mutant and its wild-type counterpart were analyzed at 30°C and 37°C.
2015-03-02 | E-GEOD-66411 | biostudies-arrayexpress
Project description:The AMP-activated protein kinase (AMPK) and the target of rapamycin complex 1 (TORC1) are central kinase modules of two opposing signaling pathways that control eukaryotic cell growth and metabolism in response to the availability of energy and nutrients. Accordingly, energy depletion activates AMPK to inhibit growth, while nutrients and high energy levels activate TORC1 to promote growth. Both in mammals and lower eukaryotes such as yeast, the AMPK and TORC1 pathways are wired to each other at different levels, which ensures homeostatic control of growth and metabolism. In this context, a previous study (Hughes Hallett et al., 2015) reported that AMPK in yeast, that is Snf1, prevents the transient TORC1 reactivation during the early phase following acute glucose starvation, but the underlying mechanism has remained elusive. Using a combination of unbiased mass spectrometry (MS)-based phosphoproteomics, genetic, biochemical, and physiological experiments, we show here that Snf1 temporally maintains TORC1 inactive in glucose-starved cells primarily through the TORC1-regulatory protein Pib2. Our data, therefore, extend the function of Pib2 to a hub that integrates both glucose and, as reported earlier, glutamine signals to control TORC1. We further demonstrate that Snf1 phosphorylates the TORC1 effector kinase Sch9 within its N-terminal region and thereby antagonizes the phosphorylation of a C-terminal TORC1-target residue within Sch9 itself that is critical for its activity. The consequences of Snf1-mediated phosphorylation of Pib2 and Sch9 are physiologically additive and sufficient to explain the role of Snf1 in short-term inhibition of TORC1 in acutely glucose-starved cells.
| S-EPMC9937656 | biostudies-literature
Project description:Ded1 is a DEAD-box RNA helicase with essential roles in translation initiation. It binds to the eukaryotic initiation factor 4F (eIF4F) complex and promotes 48S preinitiation complex assembly and start-site scanning of 5' untranslated regions of mRNAs. Most prior studies of Ded1 cellular function were conducted in steady-state conditions during nutrient-rich growth. In this work, however, we examine its role in the translational response during target of rapamycin (TOR)C1 inhibition and identify a novel function of Ded1 as a translation repressor. We show that C-terminal mutants of DED1 are defective in down-regulating translation following TORC1 inhibition using rapamycin. Furthermore, following TORC1 inhibition, eIF4G1 normally dissociates from translation complexes and is degraded, and this process is attenuated in mutant cells. Mapping of the functional requirements for Ded1 in this translational response indicates that Ded1 enzymatic activity and interaction with eIF4G1 are required, while homo-oligomerization may be dispensable. Our results are consistent with a model wherein Ded1 stalls translation and specifically removes eIF4G1 from translation preinitiation complexes, thus removing eIF4G1 from the translating mRNA pool and leading to the codegradation of both proteins. Shared features among DED1 orthologues suggest that this role is conserved and may be implicated in pathologies such as oncogenesis.
| S-EPMC6743465 | biostudies-literature
Project description:We replaced the DED1 ORF with DBP1 and inserted this either twice or four times in a diploid cell. We determined that with twice as much DBP1 expressable, the same amount of protein was made as for DED1, and there were few changes in gene expression. The transcripts that were translated better with DED1 were ones with highly structured 5' leaders, consistent with Dbp1 acting as a less effective initiation helicase than Ded1.
2025-01-29 | GSE262934 | GEO
Project description:Target of rapamycin complex 1 (TORC1) is implicated in growth control and aging from yeast to humans. Fission yeast is emerging as a popular model organism to study TOR signaling, although rapamycin has been thought to not affect cell growth in this organism. Here we analyzed the effects of rapamycin and caffeine, singly and combined, on multiple cellular processes in fission yeast. The two drugs led to diverse and specific phenotypes that depended on TORC1 signaling pathway inhibition, including prolonged chronological lifespan, inhibition of global translation, inhibition of cell growth and division, and reprogramming of global gene expression mimicking nitrogen starvation. Rapamycin and caffeine differentially affected these various TORC1-dependent processes. Combined drug treatment augmented most phenotypes and effectively blocked cell growth. Although rapamycin showed a much more subtle effect on global translation than did caffeine, rapamycin was more effective in prolonging chronological lifespan. Rapamycin prolonged the lifespan of non-growing cells only when applied during the growth phase but not when applied after cells had stopped proliferation. The doses of rapamycin and caffeine strongly correlated with growth inhibition and with lifespan extension. This comprehensive analysis will inform future studies into TORC1 function and cellular aging in fission yeast and beyond.
2013-05-05 | E-MTAB-1489 | biostudies-arrayexpress
Project description:To determine how TORC1 and PKA cooperate to regulate cell growth, we performed temporal analysis of gene expression in yeast switched from a non-fermentable substrate, to glucose, in the presence and absence of TORC1 and PKA inhibitors. Quantitative analysis of these data reveals that PKA drives the expression of key cell growth genes during transitions into, and out of, the rapid growth state in glucose, while TORC1 is important for the steady-state expression of the same genes.
2019-07-19 | GSE133591 | GEO
Project description:The goals of this study are to compare NGS-derived transcriptomes (RNA-Seq) derived from ded1-ref mutant and normal maize endosperm tissue. Homozygous ded1-ref mutants exhibit a seed defect. Maize Ded1 encodes a transcription factor. This study characterizes DEGs between ded1-ref and normal endosperm and serves to identify direct target genes together with DAP-seq data.
2022-06-10 | GSE183302 | GEO
Project description:DEAD-box RNA helicases eIF4A and Ded1 are believed to promote translation initiation by resolving mRNA secondary structures that impede ribosome attachment at the mRNA 5’ end or subsequent scanning of the 5’UTR, but whether they perform distinct functions or act redundantly in vivo is poorly understood. We compared the effects of mutations in Ded1 or eIF4A on global translational efficiencies (TEs) in yeast by ribosome footprint profiling. Despite similar reductions in bulk translation, inactivation of a cold-sensitive Ded1 mutant substantially reduced the TEs of >600 mRNAs, whereas inactivation of a temperature-sensitive eIF4A mutant yielded <40 similarly impaired mRNAs. The broader requirement for Ded1 did not reflect more pervasive secondary structures at low temperature, as inactivation of temperature-sensitive and cold-sensitive ded1 mutants gave highly correlated results. Interestingly, Ded1-dependent mRNAs exhibit greater than average 5’UTR length and propensity for secondary structure, implicating Ded1 in scanning though structured 5' UTRs. Reporter assays confirmed that cap- distal stem-loop insertions increase dependence on Ded1 but not eIF4A for efficient translation. While only a small fraction of mRNAs is strongly dependent on eIF4A, this dependence is significantly correlated with requirements for Ded1 and 5’UTR features characteristic of Ded1- dependent mRNAs. Our findings suggest that Ded1 is critically required to promote scanning through secondary structures within 5’UTRs; and while eIF4A cooperates with Ded1 in this function, it also promotes a step of initiation common to virtually all yeast mRNAs.
2015-03-02 | GSE66411 | GEO