Nascent polypeptide within the exit tunnel stabilizes the ribosome to counteract risky translation
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ABSTRACT:
PROVIDER: PRJNA764085 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR15929424.fastq.gz | Fastqsanger.gz | |||
| SRR15929425.fastq.gz | Fastqsanger.gz | |||
| SRR15929426.fastq.gz | Fastqsanger.gz | |||
| SRR15929427.fastq.gz | Fastqsanger.gz |
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Project description:Continuous translation elongation, irrespective of amino acid sequences, is a prerequisite for living organisms to produce their proteomes. However, the risk of elongation abortion is concealed within nascent polypeptide products. For example, negatively charged sequences with occasional intermittent prolines, termed intrinsic ribosome destabilization (IRD) sequences, destabilizes the translating ribosomal complex. Thus, some nascent chain sequences lead to premature translation cessation. Here, we show that most potential IRD sequences in the middle of open reading frames remain cryptic by two mechanisms: the nascent polypeptide itself that spans the exit tunnel and its bulky amino acid residues that occupy the tunnel entrance region. Thus, nascent polypeptide products have a built-in ability to ensure elongation continuity by serving as a bridge and thus by protecting the large and small ribosomal subunits from dissociation.
2021-12-08 | GSE184331 | GEO
Project description:Continuous translation elongation, irrespective of amino acid sequences, is a prerequisite for living organisms to produce their proteomes. However, nascent polypeptide products bear an inherent risk of elongation abortion. For example, negatively charged sequences with occasional intermittent prolines, termed intrinsic ribosome destabilization (IRD) sequences, weaken the translating ribosomal complex, causing certain nascent chain sequences to prematurely terminate translation. Here, we show that most potential IRD sequences in the middle of open reading frames remain cryptic and do not interrupt translation, due to two features of the nascent polypeptide. Firstly, the nascent polypeptide itself spans the exit tunnel, and secondly, its bulky amino acid residues occupy the tunnel entrance region, thereby serving as a bridge and protecting the large and small ribosomal subunits from dissociation. Thus, nascent polypeptide products have an inbuilt ability to ensure elongation continuity.
| S-EPMC8634131 | biostudies-literature
Project description:Mitochondrial gene expression needs to be balanced with cytosolic translation to produce oxidative phosphorylation complexes. In yeast, translational feedback loops involving lowly expressed proteins called translational activators help to achieve this balance. Synthesis of cytochrome b (Cytb or COB), a core subunit of complex III in the respiratory chain, is controlled by three translational activators and the assembly factor Cbp3-Cbp6. However, the molecular interface between the COB translational feedback loop and complex III assembly are yet unknown. Here, using protein-proximity mapping combined with selective mitoribosome profiling, we reveal the components and dynamic interaction of the molecular switch controlling COB translation. Specifically, we demonstrate that Mrx4, a previously uncharacterized ligand of the mitoribosomal polypeptide tunnel exit, interacts with either the assembly factor Cbp3-Cbp6 or with the translational activator Cbs2. These reciprocal interactions determine whether the translational activator complex with bound COB mRNA can interact with the mRNA channel exit on the small ribosomal subunit for translation initiation. Organization of the feedback loop at the tunnel exit therefore orchestrates mitochondrial translation regulation with respiratory chain biogenesis.
2025-07-16 | GSE283176 | GEO
Project description:Mitochondrial gene expression needs to be balanced with cytosolic translation to produce oxidative phosphorylation complexes. In yeast, translational feedback loops involving lowly expressed proteins called translational activators help to achieve this balance. Synthesis of cytochrome b (Cytb or COB), a core subunit of complex III in the respiratory chain, is controlled by three translational activators and the assembly factor Cbp3-Cbp6. However, the molecular interface between the COB translational feedback loop and complex III assembly is yet unknown. Here, using protein-proximity mapping combined with selective mitoribosome profiling, we reveal the components and dynamics of the molecular switch controlling COB translation. Specifically, we demonstrate that Mrx4, a previously uncharacterized ligand of the mitoribosomal polypeptide tunnel exit, interacts with either the assembly factor Cbp3-Cbp6 or with the translational activator Cbs2. These reciprocal interactions determine whether the translational activator complex with bound COB mRNA can interact with the mRNA channel exit on the small ribosomal subunit for translation initiation. Organization of the feedback loop at the tunnel exit therefore orchestrates mitochondrial translation with respiratory chain biogenesis
2025-07-11 | PXD061279 | Pride
Project description:The Ribosome Assembly Factor Reh1 is Released from the Polypeptide Exit Tunnel in the Pioneer Round of Translation
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Project description:More than 200 assembly factors (AFs) are required for the production of ribosomes in yeast. The stepwise association and dissociation of these AFs with the pre-ribosomal subunits occurs in a strictly hierarchical manner to ensure correct maturation of the pre-rRNAs and assembly of the ribosomal proteins. Although decades of research have provided a wealth of insights into the functions of many AFs, others remain poorly characterized. Pol5 was initially classified with B-type DNA polymerases, however, several lines of evidence indicate the involvement of this protein in ribosome assembly. Here, we show that depletion of Pol5 affects the processing of pre-rRNAs destined for the both the large and small subunits. Furthermore, we identify binding sites for Pol5 in the 5’ external transcribed spacer and within domain III of the 25S rRNA sequence. Consistent with this, we reveal that Pol5 is required for recruitment of ribosomal proteins that form the polypeptide exit tunnel in the LSU and that depletion of Pol5 impairs the release of 5’ ETS fragments from early pre-40S particles. The dual functions of Pol5 in 60S assembly and recycling of pre-40S AFs suggest that this factor could contribute to ensuring the stoichiometric production of ribosomal subunits.
2019-11-05 | GSE132973 | GEO
Project description:Understanding the conformational sampling of translation-arrested ribosome nascent chain complexes is key to understand co-translational folding. Up to now, coupling of cysteine oxidation, disulfide bond formation and structure formation in nascent chains has remained elusive. Here, we investigate the eye-lens protein γB-crystallin in the ribosomal exit tunnel. Using mass spectrometry, theoretical simulations, dynamic nuclear polarization-enhanced solid-state nuclear magnetic resonance and cryo-electron microscopy, we show that thiol groups of cysteine residues undergo S-glutathionylation and S-nitrosylation and form non-native disulfide bonds. Thus, covalent modification chemistry occurs already prior to nascent chain release as the ribosome exit tunnel provides sufficient space even for disulfide bond formation which can guide protein folding.
2020-11-09 | PXD021574 | Pride
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:Pol5 is required for recycling of small subunit biogenesis factors and for formation of the polypeptide exit tunnel of the large ribosomal subunit
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