Similar Datasets
Project description:Studies of eRF1 in mammalian cells
| PRJNA1098850 | ENA
Project description:The ribosome-associated quality control (RQC) is a surveillance system for aberrant translation, sensing ribosome collisions. Although the molecular mechanism has been extensively studied, the endogenous targets of RQC in human cells were poorly understood. Here, starting from the study of codon specificity of eukaryotic termination factor eRF1, we unexpectedly find that misrecognition of UUA sense codon by eRF1 leads to ribosome collisions and provides the source of RQC substrates in humans. eRF1-selective Monosome-Seq and Disome-Seq reveal that eRF1 recruitment to ribosome was not restricted to the stop codons but also the sub-cognate sense codons, including the UUA codon. The UUA misrecognition by eRF1 causes ribosome collision without termination reaction. Remarkably, Disome-Seq with the depletion of ASCC3 and 4EHP, key factors in RQC, showed that ribosome stalled at UUA codons are the predominant sub-populations rescued by RQC. Failure to resolve ribosome collisions by RQC triggers p38 phosphorylation and upregulation of stress response transcription factor ATF3. This study presents the impact of sense codon misrecognition by the termination factor on translation homeostasis in human cells.
2024-09-18 | GSE275336 | GEO
Project description:Upon the environmental changes, cells flexibly and rapidly alter the gene expression by the translation control. In plants, the translation of NIP5;1, a boric acid diffusion facilitator, is upregulated through the upstream open reading frame (uORF), which comprises only AUG and stop codons, in response to the shortage of boric acid. However, the molecular details of how the minimum uORF controls the translation of downstream main ORF depending on the boric acid concentration remained unclear. Here we showed that the 80S ribosome (80S) assembled at AUG-stop migrates into the subsequent RNA segment for downstream translation initiation and that the presence of boric acid impedes the process by stable confinement of eukaryotic release factor 1 (eRF1) on 80S on AUG-stop. Ribosome profiling along the in vitro-translated reporter mRNA revealed that the number of the ribosomes on AUG-stop and that along the downstream are anti-correlated in response to boric acid, reflecting the 80S scanning leaving AUG-stop for translation initiation from downstream ORF. Moreover, cryo-electron microscopy (cryo-EM) analysis revealed that, in the presence of boric acid, the initiator methionyl-tRNA (Met-tRNAi) in the P site stably contacts eRF1 in the A site on AUG-stop. In contrast, the absence of boric acid led to weaker eRF1 density, suggesting that eRF1 cannot be stably positioned without boric acid. Consistent with the structural features, the hydrolysis of Met-tRNAi on AUG-stop was accelerated by boric acid, which is likely to be the checkpoint for the subsequent 80S scanning of the downstream. Our results provide a molecular insight into the translation regulation by a minimum and environment-responsive uORF.
2023-09-20 | GSE189222 | GEO
Project description:Limitation for amino acids is thought to regulate translation in mammalian cells primarily by signaling through the kinases mTORC1 and GCN2. We find that limitation for the amino acid arginine causes a selective loss of tRNA charging, which regulates translation through ribosome pausing at two of six arginine codons. Interestingly, limitation for leucine, an essential and abundant amino acid in protein, results in little or no ribosome pausing. Chemical and genetic perturbation of mTORC1 and GCN2 signaling revealed that their robust response to leucine limitation prevents ribosome pausing, while an insufficient response to arginine limitation led to loss of arginine tRNA charging and ribosome pausing. Codon-specific ribosome pausing decreased protein production and triggered premature ribosome termination without significantly reducing mRNA levels. Together, our results suggest that amino acids which are not optimally sensed by the mTORC1 and GCN2 pathways still regulate translation through an evolutionarily conserved mechanism based on synonymous codon usage.
2018-07-19 | GSE113751 | GEO
Project description:Sense codon-misassociated eRF1 elicits widespread ribosome stalling subjected to quality control
| PRJNA1150324 | ENA
Project description:Transfer RNA (tRNA) modifications are crucial for protein synthesis, but their physiological roles remain poorly understood. Here we investigate the impact of N4-acetylcytidine (ac4C), a highly conserved tRNA modification, using a Thumpd1 knockout mouse model. We find that loss of Thumpd1-dependent tRNA acetylation leads to reduced levels of tRNALeu , increased ribosome stalling and collisions, and activation of eIF2α phosphorylation. Thumpd1 knockout mice exhibit growth defects and sterility. Remarkably, concurrent knockout of Thumpd1 and the stress-sensing kinase Gcn2 causes penetrant postnatal lethality, revealing a critical genetic interaction. Our findings demonstrate that a modification restricted to a single site within type II cytosolic tRNAs can regulate ribosome-mediated signaling in mammalian organisms. By providing insight into how tRNA modifications shape signaling and cell fate in response to stress, this work opens up novel strategies for therapeutic intervention and translational control.
2025-11-27 | GSE311268 | GEO
Project description:Transfer RNA (tRNA) modifications are crucial for protein synthesis, but their physiological roles remain poorly understood. Here we investigate the impact of N4-acetylcytidine (ac4C), a highly conserved tRNA modification, using a Thumpd1 knockout mouse model. We find that loss of Thumpd1-dependent tRNA acetylation leads to reduced levels of tRNALeu, increased ribosome stalling, and activation of eIF2-alpha phosphorylation. Thumpd1 knockout mice exhibit growth defects and sterility. Remarkably, concurrent knockout of Thumpd1 and the stress-sensing kinase Gcn2 causes penetrant postnatal lethality, revealing a critical genetic interaction. Our findings demonstrate that a modification restricted to a single position within type II cytosolic tRNAs can regulate ribosome-mediated signaling in mammalian organisms. Insights into how tRNA modifications shape phenotype and signaling in response to stress provide a foundation for novel strategies for therapeutic intervention and translational control.
2024-08-08 | MSV000095557 | MassIVE
Project description:Transfer RNA (tRNA) modifications are crucial for protein synthesis, but their physiological roles remain poorly understood. Here we investigate the impact of N4-acetylcytidine (ac4C), a highly conserved tRNA modification, using a Thumpd1 knockout mouse model. We find that loss of Thumpd1-dependent tRNA acetylation leads to reduced levels of tRNALeu, increased ribosome stalling and collisions, and activation of eIF2α phosphorylation. Thumpd1 knockout mice exhibit growth defects and sterility. Remarkably, concurrent knockout of Thumpd1 and the stress-sensing kinase Gcn2 causes penetrant postnatal lethality, revealing a critical genetic interaction. Our findings demonstrate that a modification restricted to a single site within type II cytosolic tRNAs can regulate ribosome-mediated signaling in mammalian organisms. By providing insight into how tRNA modifications shape signaling and cell fate in response to stress, this work opens up novel strategies for therapeutic intervention and translational control.
2025-03-25 | GSE272400 | GEO
Project description:Transfer RNA (tRNA) modifications are crucial for protein synthesis, but their physiological roles remain poorly understood. Here we investigate the impact of N4-acetylcytidine (ac4C), a highly conserved tRNA modification, using a Thumpd1 knockout mouse model. We find that loss of Thumpd1-dependent tRNA acetylation leads to reduced levels of tRNALeu, increased ribosome stalling and collisions, and activation of eIF2α phosphorylation. Thumpd1 knockout mice exhibit growth defects and sterility. Remarkably, concurrent knockout of Thumpd1 and the stress-sensing kinase Gcn2 causes penetrant postnatal lethality, revealing a critical genetic interaction. Our findings demonstrate that a modification restricted to a single site within type II cytosolic tRNAs can regulate ribosome-mediated signaling in mammalian organisms. By providing insight into how tRNA modifications shape signaling and cell fate in response to stress, this work opens up novel strategies for therapeutic intervention and translational control.
2025-03-25 | GSE272399 | GEO
Project description:In response to environmental changes, cells flexibly and rapidly alter gene expression, through translational controls. In plants, the translation of NIP5;1, a boric acid diffusion facilitator, is upregulated through upstream open reading frames (uORFs) that comprise only AUG and stop codons, in response to a shortage of boric acid in the environment. However, the molecular details of how the minimum uORF controls the translation of the downstream main ORF, in a boric acid concentration-dependent manner, have remained unclear. Here, combining ribosome profiling, TCP-Seq, structural analysis with cryo-electron microscopy, and biochemical assays, we showed that the 80S ribosome (80S) assembled at AUG-stop migrates into the subsequent RNA segment, followed by downstream translation initiation, and that boric acid impedes this process by the stable confinement of eukaryotic release factor 1 (eRF1) on the 80S on AUG-stop. Our results provide molecular insight into translation regulation by a minimum and environment-responsive uORF.
2023-09-19 | GSE223755 | GEO