Elucidating the role of Zuo1 protein on the translational control of gene expression in Saccharomyces cerevisiae
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ABSTRACT:
PROVIDER: PRJNA635625 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR11866762_1.fastq.gz | Fastqsanger.gz | |||
| SRR11866762_2.fastq.gz | Fastqsanger.gz | |||
| SRR11866763_1.fastq.gz | Fastqsanger.gz | |||
| SRR11866763_2.fastq.gz | Fastqsanger.gz | |||
| SRR11866764_1.fastq.gz | Fastqsanger.gz |
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Project description:Ribosome dynamics during mRNA translation elongation regulate mRNA stability. Yet, known regulators of ribosome transit, such as codon usage, cannot fully explain transcriptome-wide decay rates. Here, we demonstrate that nascent polypeptide folding modulates elongation rates, with Zuotin (Zuo1) serving as an essential mediator. Using reporter constructs encoding co-translationally unstructured proteins and RNA sequencing under proteotoxic stress, we show that Zuo1 is required for selective destabilization of transcripts whose nascent peptides fail to fold properly. This process relies on the co-translational mRNA decay factor Not5, which detects slowed ribosomes. 35S labeling indicates that nascent peptide folding defects correlate with reduced elongation rates in a Zuo1-dependent manner, and ribosome profiling reveals that global protein misfolding induces Zuo1-dependent ribosome pausing. These findings position Zuo1 as a key mediator linking nascent peptide folding status to ribosome dynamics and mRNA stability. Furthermore, this work suggests an expanded role for Not5 beyond codon optimality sensing.
2023-06-01 | GSE151370 | GEO
Project description:Ribosome dynamics during mRNA translation elongation regulate mRNA stability. Yet, known regulators of ribosome transit, such as codon usage, cannot fully explain transcriptome-wide decay rates. Here, we demonstrate that nascent polypeptide folding status modulates elongation rates, with Zuotin (Zuo1) serving as an essential mediator, and this also influences mRNA stability. Using reporter constructs encoding co-translationally unstructured proteins and RNA sequencing under proteotoxic stress, we show that Zuo1 is required for the selective destabilization of transcripts whose nascent peptides fail to fold properly. This process is independent of codon optimality but relies on the co-translational mRNA decay factor Not5, which detects slowed ribosomes. 35S labeling indicates that nascent peptide folding defects correlate with reduced elongation rates in a Zuo1-dependent manner, and ribosome profiling reveals that global protein misfolding induces Zuo1-dependent ribosome pausing. These findings position Zuo1 as a key mediator linking nascent peptide folding status to ribosome dynamics and mRNA stability. Furthermore, this work suggests an expanded role for Not5 beyond codon optimality sensing.
2026-07-24 | GSE319659 | GEO
Project description:Nascent chain folding status modulates ribosome dynamics and mRNA stability through the ribosome-associated chaperone Zuo1
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Project description:The yeast Hsp70 chaperone Ssb interacts with ribosomes and nascent chains to co-translationally assist protein folding. Here, we present a proteome-wide analysis of Hsp70 function during translation, based on in vivo selective ribosome profiling, that reveals mechanistic principles coordinating translation with chaperone-assisted protein folding. Ssb binds most cytosolic, nuclear, and mitochondrial proteins and a subset of ER proteins, supporting its general chaperone function. Position-resolved analysis of Ssb engagement reveals compartment- and protein-specific nascent chain binding profiles that are coordinated by emergence of positively charged peptide stretches enriched in aromatic amino acids. Ssbs’ function is temporally coordinated by RAC but independent from NAC. Analysis of ribosome footprint densities along orfs reveals that ribosomes translate faster at times of Ssb binding. This is coordinated by biases in mRNA secondary structure, and codon usage as well as the action of Ssb, suggesting chaperones may allow higher protein synthesis rates by actively coordinating protein synthesis with co-translational folding.
2017-07-17 | GSE93830 | GEO
Project description:The nascent polypeptide-associated complex (NAC) is a conserved ribosome-bound factor with essential yet incompletely understood roles in protein biogenesis1. Here, we show that NAC is a multifaceted regulator that coordinates translation elongation, cotranslational folding, and organelle targeting through distinct interactions with nascent polypeptides both inside and outside the ribosome exit tunnel. Using NAC-selective ribosome profiling in C. elegans, we identify thousands of sequence-specific NAC binding events across the nascent proteome, revealing broad cotranslational engagement with hydrophobic and helical motifs in cytosolic, nuclear, ER, and mitochondrial proteins. Unexpectedly, we discover an intra-tunnel sensing mode, where NAC engages ribosomes with extremely short nascent polypeptides inside the exit tunnel in a sequence-specific manner. Moreover, initial NAC interactions induce an early elongation slowdown that tunes ribosome flux and prevent ribosome collisions, linking NAC’s chaperone activity to kinetic control of translation. We propose NAC action protects aggregation-prone intermediates by shielding amphipathic helices, thus promoting cytonuclear folding. NAC also supports mitochondrial membrane protein biogenesis and ER targeting by early recognition of signal sequences and transmembrane domain. Our findings establish NAC as an early-acting, multifaceted orchestrator of cotranslational proteostasis, with distinct mechanisms of action on nascent chains depending on their sequence features and subcellular destinations.
2025-12-22 | GSE263661 | GEO
Project description:The nascent polypeptide-associated complex (NAC) is a conserved ribosome-bound factor with essential yet incompletely understood roles in protein biogenesis1. Here, we show that NAC is a multifaceted regulator that coordinates translation elongation, cotranslational folding, and organelle targeting through distinct interactions with nascent polypeptides both inside and outside the ribosome exit tunnel. Using NAC-selective ribosome profiling in C. elegans, we identify thousands of sequence-specific NAC binding events across the nascent proteome, revealing broad cotranslational engagement with hydrophobic and helical motifs in cytosolic, nuclear, ER, and mitochondrial proteins. Unexpectedly, we discover an intra-tunnel sensing mode, where NAC engages ribosomes with extremely short nascent polypeptides inside the exit tunnel in a sequence-specific manner. Moreover, initial NAC interactions induce an early elongation slowdown that tunes ribosome flux and prevent ribosome collisions, linking NAC’s chaperone activity to kinetic control of translation. We propose NAC action protects aggregation-prone intermediates by shielding amphipathic helices, thus promoting cytonuclear folding. NAC also supports mitochondrial membrane protein biogenesis and ER targeting by early recognition of signal sequences and transmembrane domain. Our findings establish NAC as an early-acting, multifaceted orchestrator of cotranslational proteostasis, with distinct mechanisms of action on nascent chains depending on their sequence features and subcellular destinations.
2025-12-22 | GSE263659 | GEO
Project description:The cellular environment is critical for efficient protein maturation, but how proteins fold during biogenesis remains poorly understood. To understand how the cellular environment modulates folding, we studied the cotranslational chaperone-assisted folding of Escherichia coli dihydrofolate reductase (DHFR). To sample folding intermediates along the pathway of vectorial synthesis, we prepared a series of stalled ribosome:nascent chain complexes (RNCs) representing snapshots of DHFR folding in vivo. Proteomic analysis of RNCs revealed their composition and allowed us to define chaperone interactions as a function of NC length. These data set the stage for further studies aimed at resolving the conformation of the nascent polypeptide on the ribosome.
2025-05-06 | PXD036784 | Pride
Project description:The cellular environment is critical for efficient protein maturation, but how proteins fold during biogenesis remains poorly understood. We used hydrogen/deuterium exchange (HDX) mass spectrometry (MS) to define, at peptide resolution, the cotranslational chaperone-assisted folding pathway of Escherichia coli dihydrofolate reductase. On the ribosome, the nascent polypeptide folds via structured intermediates not populated during refolding from denaturant. Association with the ribosome allows these intermediates to form, as otherwise destabilizing C-terminal sequences remain confined in the ribosome exit tunnel. We find that partially-folded nascent chains recruit the chaperone Trigger factor, which uses a large composite hydrophobic/hydrophilic interface to engage folding intermediates without disrupting their structure. In addition, we comprehensively mapped dynamic interactions between the nascent chain and ribosomal proteins, tracing the path of the emerging polypeptide during synthesis. Our work provides a high-resolution description of de novo protein folding dynamics, thereby revealing new mechanisms by which cellular factors shape the conformational search for the native state.
2024-07-12 | PXD036945 | Pride
Project description:The folding of most proteins occurs during the course of their translation while their tRNA-bound C-termini are embedded in the ribosome. How the close proximity of nascent proteins to the ribosome influences their folding thermodynamics remains poorly understood. Here, we have developed a mass spectrometry-based approach for determining the stabilities of nascent polypeptide chains using methionine oxidation as a folding probe. This approach enables quantitative measurements sub-global folding stabilities of ribosome nascent chains (RNCs) within complex protein mixtures and extracts. To validate the methodology, we analyzed the folding thermodynamics of three model proteins (DHFR, CheY and DinB) in soluble and ribosome-bound states. The data indicated that the ribosome can significantly alter the stability of nascent polypeptides. Ribosome-induced stability modulations were highly variable among different folding domains and were dependent on localized charge distributions within nascent polypeptides. The results implicated electrostatic interactions between the ribosome surface and nascent polypeptides as the cause of ribosome-induced stability modulations. The study establishes a robust proteomic methodology for analyzing localized stabilities within ribosome-bound nascent polypeptides and sheds light on how the ribosome influences the thermodynamics of protein folding.
2023-08-09 | PXD037995 | Pride
Project description:Protein folding is assisted by molecular chaperones that bind nascent polypeptides during mRNA translation. Several structurally-distinct classes of chaperone promote de novo folding, suggesting that their activities are coordinated at the ribosome. We used biochemical reconstitution and structural proteomics to explore the molecular basis for cotranslational chaperone action in bacteria. We found that chaperone binding is disfavoured close to the ribosome, allowing folding to precede chaperone recruitment. Trigger factor recognises compact folding intermediates exposing extensive unfolded surface and dictates DnaJ access to nascent chains. DnaJ uses a large surface to bind structurally diverse intermediates, and recruits DnaK to sequence-diverse solvent-accessible sites. Neither Trigger factor, DnaJ nor DnaK destabilize cotranslational folding intermediates. Instead, the chaperones collaborate to protect incipient structure in the nascent polypeptide well beyond the ribosome exit tunnel. Our findings show how the chaperone network selects and modulates cotranslational folding intermediates.
2024-07-03 | PXD048623 | Pride