Cycloheximide resistant ribosomes reveal adaptive translation dynamics in C. elegans [Ribo-Seq]
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1282121 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:Cycloheximide resistant ribosomes reveal adaptive translation dynamics in C. elegans
| PRJNA1281666 | ENA
Project description:Protein translation regulation is critical for cellular responses and development, yet how disruptions during the elongation stage shape these processes remains incompletely understood. Here, we identify and validate a single amino acid substitution (P55Q) in the ribosomal protein RPL-36A of Caenorhabditis elegans that confers complete resistance to high concentrations of the elongation inhibitor cycloheximide (CHX). Heterozygous animals carrying both wild-type RPL-36A and RPL-36A(P55Q) exhibit normal development but intermediate CHX resistance, indicating a partial dominant effect. Leveraging RPL-36A(P55Q) as a single-copy positive selection marker for CRISPR-based genome editing, we introduced targeted modifications into multiple ribosomal protein genes, confirming its broad utility for altering essential loci. In L4-stage heterozygotes, where CHX-sensitive and CHX-resistant ribosomes coexist, ribosome profiling revealed increased start-codon occupancy, suggesting early stalling of CHX sensitive ribosomes. Chronic CHX reduced ribosome collisions, evidenced by fewer disomes and unchanged codon distributions in monosomes. Surprisingly, prolonged elongation inhibition did not activate well characterized stress pathways–including ribosome quality control (RQC), the ribotoxic stress response (RSR), or the integrated stress response (ISR)–as indicated by absence of changes in RPS-10 ubiquitination, eIF2α phosphorylation, PMK-1 phosphorylation, or the transcriptional upregulation of ATF-4 target genes. Instead, RNA-normalized ribosome footprints revealed gene-specific changes in translation efficiency, with nucleolar and P granule components significantly decreased while oocyte development genes were increased. Consistent with these observations, we detected premature oogenesis in L4 animals, suggesting that partial translation elongation inhibition reshapes translation efficiency, to fine-tune developmental timing.
2025-06-29 | GSE300790 | GEO
Project description:Cycloheximide resistant ribosomes reveal adaptive translation dynamics in C. elegans [RNA-Seq]
| PRJNA1281580 | ENA
Project description:Protein translation regulation is critical for cellular responses and development, yet how disruptions during the elongation stage shape these processes remains incompletely understood. Here, we identify and validate a single amino acid substitution (P55Q) in the ribosomal protein RPL-36A of Caenorhabditis elegans that confers complete resistance to high concentrations of the elongation inhibitor cycloheximide (CHX). Heterozygous animals carrying both wild-type RPL-36A and RPL-36A(P55Q) exhibit normal development but intermediate CHX resistance, indicating a partial dominant effect. Leveraging RPL-36A(P55Q) as a single-copy positive selection marker for CRISPR-based genome editing, we introduced targeted modifications into multiple ribosomal protein genes, confirming its broad utility for altering essential loci. In L4-stage heterozygotes, where CHX-sensitive and CHX-resistant ribosomes coexist, ribosome profiling revealed increased start-codon occupancy, suggesting early stalling of CHX sensitive ribosomes. Chronic CHX reduced ribosome collisions, evidenced by fewer disomes and unchanged codon distributions in monosomes. Surprisingly, prolonged elongation inhibition did not activate well characterized stress pathways–including ribosome quality control (RQC), the ribotoxic stress response (RSR), or the integrated stress response (ISR)–as indicated by absence of changes in RPS-10 ubiquitination, eIF2α phosphorylation, PMK-1 phosphorylation, or the transcriptional upregulation of ATF-4 target genes. Instead, RNA-normalized ribosome footprints revealed gene-specific changes in translation efficiency, with nucleolar and P granule components significantly decreased while oocyte development genes were increased. Consistent with these observations, we detected premature oogenesis in L4 animals, suggesting that partial translation elongation inhibition reshapes translation efficiency, to fine-tune developmental timing.
2025-06-29 | GSE300687 | GEO
Project description:Protein translation regulation is critical for cellular responses and development, yet how disruptions during the elongation stage shape these processes remains incompletely understood. Here, we identify and validate a single amino acid substitution (P55Q) in the ribosomal protein RPL-36A of Caenorhabditis elegans that confers complete resistance to high concentrations of the elongation inhibitor cycloheximide (CHX). Heterozygous animals carrying both wild-type RPL-36A and RPL-36A(P55Q) exhibit normal development but intermediate CHX resistance, indicating a partial dominant effect. Leveraging RPL-36A(P55Q) as a single-copy positive selection marker for CRISPR-based genome editing, we introduced targeted modifications into multiple ribosomal protein genes, confirming its broad utility for altering essential loci. In L4-stage heterozygotes, where CHX-sensitive and CHX-resistant ribosomes coexist, ribosome profiling revealed increased start-codon occupancy, suggesting early stalling of CHX sensitive ribosomes. Chronic CHX reduced ribosome collisions, evidenced by fewer disomes and unchanged codon distributions in monosomes. Surprisingly, prolonged elongation inhibition did not activate well characterized stress pathways–including ribosome quality control (RQC), the ribotoxic stress response (RSR), or the integrated stress response (ISR)–as indicated by absence of changes in RPS-10 ubiquitination, eIF2α phosphorylation, PMK-1 phosphorylation, or the transcriptional upregulation of ATF-4 target genes. Instead, RNA-normalized ribosome footprints revealed gene-specific changes in translation efficiency, with nucleolar and P granule components significantly decreased while oocyte development genes were increased. Consistent with these observations, we detected premature oogenesis in L4 animals, suggesting that partial translation elongation inhibition reshapes translation efficiency, to fine-tune developmental timing.
2025-06-28 | GSE300626 | GEO
Project description:Regulation of the efficiency with which an mRNA is translated into proteins represents a key mechanism for controlling gene expression. Such regulation impacts the number of actively translating ribosomes per mRNA molecule, referred to as translation efficiency (TE), which can be monitored using ribosome profiling and RNA-seq, or by evaluating the position of an mRNA in a polysome gradient. Here we show that in budding yeast, under nutrient limiting conditions, the commonly used translation inhibitor cycloheximide induces rapid transcriptional upregulation of hundreds of genes involved in ribosome biogenesis. Cycloheximide also prevents translation of these newly transcribed messages, leading to an apparent drop in TE of these genes under conditions that include key transitions of the yeast metabolic cycle, meiosis, and amino acid starvation; an effect which is abolished when cycloheximide pretreatment is omitted. This response requires TORC1 signaling, and is modulated by the genetic background as well as the vehicle used to deliver the drug. The present work highlights an important caveat to the use of translation inhibitors when measuring TE, and will hopefully aid in future experimental design as well as interpretation of prior results
2019-03-08 | GSE125038 | GEO
Project description:Global restriction of protein synthesis is a hallmark of cellular stress. Using hydrogen peroxide, we monitor the transcript level and also the translation status for each RNA using cycloheximide to freeze elongating ribosomes. Polyribosome fractionation of cell extracts was used to separate highly translated and poorly translated mRNAs that were then separately analysed.
2006-06-01 | E-MEXP-526 | biostudies-arrayexpress
Project description:Charcot-Marie-Tooth (CMT) disease can be caused by mutations in Aminoacyl-tRNA-Synthetases, including G240R mutation in Glycyl-tRNA-Synthetase (GARS). Ribo-seq generates snapshots of translating ribosomes on mRNA and therefore allows analysis of ribosome pausing mRNA. Here we performed Ribo-seq on lysates of HEK293T cells overexpressing GARS, WT or G240R, to dissect mechanism of CMT linked with translation. We found that GARS G240R causes pausing of ribosomes with glycine codons in A-site. The effect is specific for 21 nt ribosome-protected fragments, produced by ribosomes with empty A-sites, suggestive of the deficit of charged Glycyl-tRNA in GARS G240R-CMT.
2021-09-14 | E-MTAB-10342 | biostudies-arrayexpress
Project description:Selective translation by alternative bacterial ribosomes
| PRJNA525595 | ENA
Project description:The ribosome has considerably increased in size during metazoan evolution in the form of an RNA shell that could serve as a platform for yet unknown protein interactions. Here, we have comprehensively identified the mammalian ‘ribo-interactome’ by establishing a ribosome affinity purification method. Our findings reveal a multitude of novel ribosome interacting factors, encompassing unanticipated functional categories including energy metabolism, cell redox homeostasis, as well as key protein and RNA modifying enzymes. These findings led us to characterize ufmylation, a novel posttranslational modification on ribosomes, and define its substrates. We further show that pyruvate kinase, a key enzyme for stem cell and cancer metabolism, is an RNA binding protein that unexpectedly interacts with specialized sub-pools of ribosomes at the endoplasmic reticulum (ER) and coordinates the localization and translation of ER destined mRNAs. Collectively, these studies uncover that the ribo-interactome imbues a new layer of regulatory potential in translating the genome.
2017-06-01 | GSE96998 | GEO