bims-spribo Biomed News
on Specialized ribosomes
Issue of 2026–10–04
ten papers selected by
Maxim Bouvet, Università di Torino
  1. Site-specific inhibition of translation initiation via 2'-O-methylation.
  2. Collided ribosomes are rescued by the endonuclease Rae1 and trans- translation.
  3. Fluorescent proteins misreport ribosome abundance under translation inhibition.
  4. PEG precipitation for easy ribosome profiling.
  5. JP-1302 inhibits RNA polymerase I transcription and induces nucleolar stress in cancer cell lines.
  6. NAT10 governs development through the Xrp1-mediated stress response rather than its RNA acetylation activity.
  7. Conformational diversity of collided ribosomes determines stress signaling.
  8. Recoding by N1-methylpseudouridine guides rational mRNA vaccine design.
  9. RBP-Driven RNA sorting and local translation establish the molecular identities of individual sensory axons.
  10. DeCoRE: A computational method to resolve RNA structural heterogeneity from RNA structure probing data by direct RNA sequencing.
  1. Nucleic Acids Res. 2026 Sep 22. pii: gkag916. [Epub ahead of print]54(18):
    Site-specific inhibition of translation initiation via 2'-O-methylation.
    Adam Suh, Stephanie Mou, Emmely A Patrasso, Hannah Serio, Rui Wang, Yue Zhao, Kevin Vasquez, Smriti Sangwan, Yang Yi, Daniel Arango.
      Translation initiation involves a concerted set of intermolecular interactions that efficiently recognize optimal AUG start codons. However, translation initiation complexes often start at upstream non-optimal AUGs or near-cognate codons, leading to the expression of upstream open reading frames (uORFs). Using retrospective analyses of translation preinitiation cryo-EM structures, we identified putative hydrogen bonds between the 2'-OH groups of messenger RNA (mRNA) start codons with 18S ribosomal RNA. Disruption of these interactions using a chemical modification of mRNA, 2'-O-methylation (Nm), repressed translation initiation by preventing the preinitiation complex from recognizing start codons. Notably, 2'-O-methylation in upstream AUG and near-cognate codons inhibits upstream translation initiation while enhancing the expression of canonical ORFs. These findings revealed a transcript- and site-specific inhibitory role for 2'-O-methylation in translation initiation, providing novel insights into the mechanisms of start codon selection in human transcriptomes.
    DOI:  https://doi.org/10.1093/nar/gkag916
  2. bioRxiv. 2026 Sep 24. pii: 2026.09.23.753862. [Epub ahead of print]
    Collided ribosomes are rescued by the endonuclease Rae1 and trans- translation.
    Daniel D Tetreault, Katrina Callan, Cassidy R Prince, Heather A Feaga.
      Ribosome stalling is a major problem in all domains of life. When a ribosome stalls, trailing ribosomes may catch up to and collide with the stalled ribosome, depleting protein synthesis capacity. Here, we describe a novel pathway used by Gram-positive bacteria to rescue ribosome collisions. We used the ATPase defective ABCF protein YdiF(EQ 2 ) to induce ribosome stalling and collisions in Bacillus subtilis . Ribosome profiling (Ribo-seq) of YdiF(EQ 2 )-expressing cells revealed that collided ribosomes are enriched for tmRNA, a functional RNA involved in trans -translation. We confirmed that tmRNA tagging activity is globally increased upon expression of any ATPase defective ABCF as well as in cells treated with the collision-inducing antibiotic erythromycin, suggesting this is a generalizable mechanism to rescue stalled and collided ribosomes. The global increase in tmRNA tagging that occurred in response to both erythromycin and YdiF(EQ2) induced collisions was dependent on the Rae1 endonuclease. Loss of trans -translation in cells experiencing widespread ribosome collisions leads to a severe fitness defect, consistent with the importance of this pathway in rescuing ribosomes stalled on truncated mRNAs that result from ribosome collisions. Altogether, our work supports a model in which Rae1 cleaves mRNA on collided ribosomes, thereby generating a truncated mRNA substrate for trans -translation and leading to rescue and recycling of the collided ribosomes. We term this mechanism C ollision- A ssociated R ae1-induced trans- Translation (CART). CART broadens the repertoire of tools that bacteria use to manage ribosome collisions.
    Significance: Prolonged ribosome stalling leads to ribosome collisions, which are rescued by specialized factors. While ribosome collisions have been extensively studied in eukaryotes, our understanding of collision rescue in bacteria is in its infancy. Data described here are the first to directly show that tmRNA mediates rescue of collided ribosomes in a Gram-positive bacterium, Bacillus subtilis . This pathway is analogous to what occurs in model organisms such as Escherichia coli and Saccharomyces cerevisiae , but relies on an unrelated nuclease, Rae1. Since B. subtilis and E. coli are on opposite ends of the bacterial phylogenetic tree, and since Rae1 is broadly conserved in bacteria, our findings suggest that mRNA cleavage arose convergently in distantly related bacteria as a strategy to rescue ribosome collisions. Convergent evolution of these pathways highlights the importance of rescuing collided ribosomes in all organisms. Moreover, insights into ribosome rescue in E. coli and B. subtilis can guide studies of ribosome rescue in bacteria with intermediary phylogenetic relatedness to these two model organisms.
    DOI:  https://doi.org/10.64898/2026.09.23.753862
  3. bioRxiv. 2026 Sep 22. pii: 2026.09.17.752513. [Epub ahead of print]
    Fluorescent proteins misreport ribosome abundance under translation inhibition.
    Aniket Zodage, Justin Chen, Nabigha Mogharbel, Hidde de Jong, Suckjoon Jun.
      Fluorescent proteins are widely used as quantitative reporters of protein abundance in living cells. Here we show that this relationship can break down under translation inhibition: the cellular ribosomal abundance can increase while fluorescence intensity remains unchanged or even decreases. Using fluorescent reporters of ribosome abundance in Escherichia coli and Bacillus subtilis , we find that cellular fluorescence intensity quantitatively tracks ribosome abundance, inferred by RNA-to-protein ratio, under nutrient-limited growth but that is no longer true during chloramphenicol treatment. In E. coli , the discrepancy occurs with transcriptional and translational reporters and with both ribosomal protein and ribosomal RNA promoters. To distinguish changes in fluorescence output per reporter from changes in reporter abundance, we constructed a fluorescent protein-LacZ dual reporter in which fluorescence and an independent enzymatic estimate of reporter abundance are obtained from the same protein. Under translation inhibition, LacZ-derived reporter abundance increases whereas fluorescence intensity remains approximately constant, showing that fluorescence output per unit reporter abundance decreases. In a companion study, Bakshi and colleagues demonstrate that the effect extends to other fluorescent proteins and other translation inhibitors, indicating that it is not specific to a particular fluorescent protein or translation inhibitor. Our results show that fluorescent-protein calibration can be condition-dependent and cannot be assumed to transfer across physiological perturbations.
    DOI:  https://doi.org/10.64898/2026.09.17.752513
  4. Methods. 2026 Oct 01. pii: S1046-2023(26)00204-5. [Epub ahead of print]256 109-116
    PEG precipitation for easy ribosome profiling.
    Alberto Agnolin, Yaozu Han, Christopher Sauer, Paul Igor Costea, Max Fabian Felle, Mathis Appelbaum, Leendert W Hamoen.
      Ribosome profiling is a powerful technique to discover translation events, and to reveal translational dynamics at gene and codon level. The procedure includes laborious ribosome isolation steps using density gradient ultracentrifugation. However, ultracentrifuges are not available in every laboratory, and ultracentrifugation is not suitable for high-throughput experimentation. PEG precipitation has been used to purify ribosomes and in this study we have tested whether PEG precipitation can be used as an alternative method to isolate ribosomes for ribosome profiling. We have benchmarked PEG precipitation against spin column-based size-exclusion and standard sucrose density ultracentrifugation methods for ribosome isolation. The methods were tested using the bacterial model system Bacillus subtilis treated with mupirocin, which stalls ribosomes at isoleucine codons, providing a testable reference for ribosome profile comparison. We found that the use of PEG precipitation, size-exclusion spin columns, and a combination of both, yielded ribosome profiles that were comparable to the standard procedure. With these methods, ultracentrifugation can be omitted, facilitating ribosome profiling and enabling high-throughput analyses.
    Keywords:  Bacillus subtilis; PEG precipitation; Ribo-seq; Ribosome isolation; Ribosome profiling
    DOI:  https://doi.org/10.1016/j.ymeth.2026.09.006
  5. Cell Death Discov. 2026 Sep 30. pii: 389. [Epub ahead of print]12(1):
    JP-1302 inhibits RNA polymerase I transcription and induces nucleolar stress in cancer cell lines.
    Sheetanshu Saproo, Styliani Papadaki, Johana Fernandez-Martinez, Pinky Sultana, Dimitris C Kanellis, Jiri Bartek, Mikael S Lindström.
      Cancer cells upregulate ribosomal RNA (rRNA) synthesis to sustain the elevated ribosome production that fuels rapid proliferation, creating a dependency that renders RNA polymerase I (Pol I) transcription a pharmacological vulnerability. JP-1302 is a blood-brain barrier-penetrant 9-anilinoacridine derivative developed as an α2C-adrenergic receptor antagonist, but also recently identified as an RNA Pol II inhibitor at micromolar concentrations (~5-10 μM). Here we show that JP-1302 also suppresses Pol I-driven 47S pre-rRNA synthesis at submicromolar-to-low-micromolar concentrations, evidenced by loss of nascent EU-labelled rRNA and reduced activity of an rDNA promoter-driven luciferase reporter. This was accompanied by preferential proteasome-dependent degradation of the Pol I catalytic subunit POLR1A, independent of α2C-adrenergic receptor signalling, displacement of POLR1A from the rDNA promoter and progressive disruption of nucleolar morphology. Growth inhibitory IC50 values across a panel of normal and cancer cell lines, including glioblastoma, ranged from submicromolar to low micromolar. Nucleolar stress stabilised p53 and induced p21 expression at low-to-intermediate concentrations, without evidence of DNA damage at these doses (no γH2AX increase at concentrations sufficient to abrogate Pol I transcription). Interestingly, p53-null and p53-wild type cells showed comparable S-phase depletion in response to JP-1302. At higher concentrations, JP-1302 additionally caused covalent trapping of TOP2A/TOP2B, accompanied by delayed γH2AX induction, and enrichment of the FACT subunits SSRP1 and SPT16 on chromatin. Together, these findings identify JP-1302 as a pharmacologically distinct inhibitor of ribosome biogenesis with broader, dose-dependent chromatin-disruptive activity.
    DOI:  https://doi.org/10.1038/s41420-026-03381-8
  6. Sci Adv. 2026 Oct 02. 12(40): eaeg0399
    NAT10 governs development through the Xrp1-mediated stress response rather than its RNA acetylation activity.
    Lin Wang, Zhao Xue, Shuyang Gao, Yanhua Wang, Houguang Lu, Bo Peng, Tianyi Fan, Ying Cao, Jian-Quan Ni, Guoliang Lu, Jinzhong Lin, Dong Yan.
      N4-acetylcytidine (ac4C) is installed by N-acetyltransferase 10 (NAT10) and represents the only known acetylation mark on messenger RNA. Since NAT10 also acetylates transfer RNA, ribosomal RNA (rRNA), and proteins, its in vivo molecular mechanisms remain elusive. Here, we demonstrate that knockdown of Drosophila NAT10 induces an eye-to-antenna transformation, c-Jun amino-terminal kinase activation, and cell apoptosis. NAT10 facilitates ac4C modification in rRNA, and its loss impairs rRNA processing and ribosomal assembly. Depletion of NAT10 activates the integrated stress response, ultimately leading to reduced global protein synthesis. Crucially, Xrp1 plays a key role in the stress response and its ablation rescues most NAT10 loss-of-function defects and transcriptomic alterations. We also performed acetylated RNA immunoprecipitation and sequencing (acRIP-seq) on control and NAT10 knockdown flies. Furthermore, a catalytically deficient form of NAT10, which is unable to mediate ac4C acetylation, completely rescues the lethality of NAT10 mutants. Collectively, these findings establish that NAT10's primary developmental function stems from its roles in ribosome biogenesis and Xrp1 activation, which are independent of its RNA acetylation activity.
    DOI:  https://doi.org/10.1126/sciadv.aeg0399
  7. bioRxiv. 2026 Sep 10. pii: 2026.06.16.730951. [Epub ahead of print]
    Conformational diversity of collided ribosomes determines stress signaling.
    Jeffrey J Li, Timo Denk, Matthias Thoms, Allen R Buskirk, Roland Beckmann, Rachel Green.
      Ribosome collisions trigger pathways that clear stalled ribosomes, and when sufficiently abundant, the integrated stress response (ISR) through GCN2 and the ribotoxic stress response (RSR) through ZAK. The inhibitors anisomycin (ANS), emetine (EME), and didemnin B (DDB) are commonly used to induce collisions in studying these responses. Here, we demonstrate that these drugs induce distinct signatures: ANS and DDB potently activate ZAK whereas EME does not. We define transcriptional programs induced by these inhibitors, where collisions induce the RSR and general inhibition of translation leads to an RSR-independent response. Surprisingly, we find that collisions induced by EME, unlike ANS, are not cleared by ASCC3. The cryo-EM structure of human disomes stalled by EME reveals its mechanism of inhibition and a conformation distinct from ANS-stalled disomes. These differences in collision geometry explain the different outcomes in quality control and signaling activation, showing how ribosome stalling events can yield distinct cellular responses.
    DOI:  https://doi.org/10.64898/2026.06.16.730951
  8. Nat Commun. 2026 Sep 28. pii: 10029. [Epub ahead of print]17(1):
    Recoding by N1-methylpseudouridine guides rational mRNA vaccine design.
    Panagiotis Poulis, Giovanni Robecchi, Arina O Kurochkina, Michele Felletti, Marina V Rodnina.
      N1-methylpseudouridine (m1Ψ) is a key modification used in SARS-CoV-2 mRNA vaccines that reduces immunogenicity and increases mRNA stability. Recent studies suggested that m1Ψ can promote ribosomal frameshifting, a translational error generating aberrant peptides that elicit immune responses, raising concerns about unintended antigenicity. Here, we systematically examined the efficiency of frameshifting induced by m1Ψ and the underlying mechanism, aiming to inform future mRNA vaccine design. Using mRNA-based dual-fluorescence reporters in cells, fully in vitro reconstituted translation system, and single-molecule FRET microscopy, we show that m1Ψ increases +1 frameshifting on UUUC motifs in therapeutic mRNAs. Frameshifting occurs when the peptidyl-tRNA pauses in the ribosomal P site, where m1Ψ both weakens codon-anticodon interactions and promotes a frameshifting-prone tRNA conformation. Replacing slippery UUUC motifs with UUCC or UUUU eliminates this effect. Our results reveal how a clinically relevant mRNA modification promotes recoding and show that codon optimization mitigates this risk in therapeutic mRNA design.
    DOI:  https://doi.org/10.1038/s41467-026-77796-3
  9. Nat Commun. 2026 08 31. pii: 10345. [Epub ahead of print]17(1):
    RBP-Driven RNA sorting and local translation establish the molecular identities of individual sensory axons.
    Elizabeth S Silagi, Ezechukwu Nduka, Jesus Zuniga Paiz, Maria F Pazyra-Murphy, Shamsuddin A Bhuiyan, William Renthal, Rosalind A Segal.
      Neurons extend long axons that traverse distinct microenvironments, yet how these subcellular compartments acquire and maintain specialized identities remains unclear. Here, we use spatial translatomics to define the local translatomes of somatosensory dorsal root ganglion (DRG) neurons. Translating Ribosome Affinity Purification and RNA sequencing (TRAP-seq) reveal thousands of mRNAs preferentially translated within central axons, peripheral axons, or DRG somata, establishing compartment-specific translational programs. Many of these transcripts encode ion channels and neurotransmitter receptors that may confer distinct electrophysiological and regenerative properties to each axon. Integration of the TRAP-seq data with published RNA-seq identify locally translated components that change following neuropathic injury and could thereby adjust neuronal activity. We identify RNA regulons coordinated by RNA-binding proteins (RBPs) SFPQ and SRSF10, which preferentially bind and traffic mRNAs to peripheral or central axons, respectively. These findings indicate that RBP-guided RNA sorting and local translation enable the establishment and dynamic local modulation of somatosensory function.
    DOI:  https://doi.org/10.1038/s41467-026-77192-x
  10. Sci Adv. 2026 Oct 02. 12(40): eaef0670
    DeCoRE: A computational method to resolve RNA structural heterogeneity from RNA structure probing data by direct RNA sequencing.
    Yong Huang, Jinsong Zhang, Lei Sun, Qiangfeng Cliff Zhang.
      RNA molecules are inherently dynamic, forming complex structural ensembles of alternative conformations that direct their cellular functions. Current methods for resolving RNA structural heterogeneity and dynamics have been developed for short-read sequencing-based structure probing, limiting their sensitivity and resolution. Here, we introduce DeCoRE (deconvolving complex RNA structural ensembles), a computational method that leverages direct RNA sequencing to resolve RNA structural heterogeneity and dynamics at the individual transcript level. DeCoRE combines enhanced structural signal detection with direct clustering of sequencing reads to sensitively deconvolve RNA structural ensembles at single-nucleotide resolution. We demonstrate DeCoRE's capabilities by resolving the known alternative conformations of the lysine and TPP riboswitches and uncover a previously uncharacterized cotranscriptional folding pathway of bacterial ribosomal RNA, where two conformations coexist during early elongation and progressively converge into a single mature conformation. DeCoRE represents a powerful tool for dissecting RNA structural heterogeneity and dynamics, advancing our understanding of RNA function.
    DOI:  https://doi.org/10.1126/sciadv.aef0670
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