bims-ribost Biomed News
on Ribostasis and translation stress
Issue of 2026–01–25
fifty-nine papers selected by
Cédric Chaveroux, CNRS
  1. eIF3g binding to GUCG boxes located in mRNA coding regions enhances translation of mild heat shock response genes in the yeast Saccharomyces cerevisiae.
  2. Cockayne syndrome mutation in XPG activate the integrated stress response.
  3. Uridine analogs prevent stress granule formation, not by blocking PKR recognition, but by inhibiting the synthesis of T7 RNA Polymerase byproducts.
  4. A comprehensive view on r-protein binding and rRNA domain structuring during early eukaryotic ribosome formation.
  5. YhfH functions as a tri-function RNA in Bacillus thuringiensis BMB171.
  6. Synthesis of long and functionally active RNAs facilitated by acetal levulinic ester chemistry.
  7. ADAM-tRNA-seq: an optimized approach for demultiplexing and enhanced hierarchal mapping in direct tRNA sequencing.
  8. An elevated level of the mRNA exporter Mex67-Mtr2 in nuclear mRNPs impairs nuclear mRNA export.
  9. The emerging roles of METTL1-mediated tRNA m7G methylation in cancer development and immunotherapy.
  10. Meta-unstable mRNAs in activated CD8+ T cells are defined by interlinked AU-rich elements and m6A mRNA methylation.
  11. Ribosome biogenesis is increased in hepatocellular carcinoma and represents a potential therapeutic target.
  12. The GT1 domain of RNase J ensures RNA quality control through dsRNA binding in Arabidopsis plastids.
  13. Staufen dsRNA-binding domain as modules to design bifunctional antibodies for siRNA delivery.
  14. The two-step purification method ViREn identifies a single NSUN6-mediated 5-methylcytosine modification promoting dengue virus RNA genome turnover.
  15. The Integrated Stress Response in Cancer: Paradox and Therapeutic Promise.
  16. [Expression of Concern] Long non‑coding RNA SNHG20 promotes bladder cancer via activating the Wnt/β‑catenin signalling pathway.
  17. Inhibition of RNase L enhances the expression efficiency of mRNA-LNPs.
  18. Human mitochondrial helicase Twinkle has RNA binding, annealing, and strand-exchange activities.
  19. Nonsense-mediated decay as RNA quality control mechanism: An opportunity for developing viral interference in plants.
  20. Integrative analysis of nanopore direct RNA sequencing data reveals a global impact of pseudouridylation on m6A and m5C modifications.
  21. Degradation determinants are abundant in human noncanonical proteins and minor annotated isoforms.
  22. QuAPPro: an R shiny app for quantification and alignment of polysome profiles.
  23. The RNA binding protein Arid5a is an activator of TNF signaling in rheumatoid arthritis.
  24. Rop-Mediated Suppression of RpoS Production Increases Resistance to Nitric Oxide.
  25. The impact of scaRNA12 deregulation on p53-mediated cellular stress response in B-cell precursor acute lymphoblastic leukaemia.
  26. MeRIP-qPCR Assay for Detecting m6A Modification Levels of Specific RNA in Osteosarcoma Cells.
  27. Transcriptome-wide analysis of Arabidopsis DICER-LIKE1 RNA substrates.
  28. Alternative probe chemistries for single-molecule analysis of long non-coding RNA.
  29. Systems analysis of the kinetics of in vitro transcription from interactions of T7 RNA polymerase and DNA.
  30. A moments-based approach for inferring mechanisms of transcriptional regulation using nascent RNA data.
  31. Structural characterization of the HDV virion and its ribonucleoprotein.
  32. The HigB-like SehA toxin promotes non-replicating Salmonella inside macrophages by inhibiting ribonuclease III-dependent rRNA maturation.
  33. Nuclear MBL-1 modulates mitochondrial morphology through carnitine palmitoyltransferase in Caenorhabditis elegans with toxic trinucleotide repeats.
  34. Biochemical principles of miRNA targeting in flies.
  35. A novel protein encoded by circIMP3 promotes prostate cancer progression by regulating alternative splicing and tumor microenvironment.
  36. A primer on tRNAs, tRNA-m1A58 modification, and tRNA derivatives in T-cell activation.
  37. Concentration-dependent mutational scanning probes the cellular folding landscape of α-synuclein in yeast.
  38. Unconventional monooxygenation by the O2-dependent tRNA wobble uridine hydroxylase TrhO.
  39. Arsenic Trioxide Underpins Delayed Neuroinflammation and Impaired Synaptic Integrity involving Integrative Stress Response Signaling.
  40. Limited effect of short- to mid-term storage conditions on an Australian farmland soil RNA virome.
  41. CircDYRK1A Suppresses Gastric Cancer Progression via Sponging miR-331-5p to Regulate GADD45A.
  42. The RNA-binding protein NOVA-1 regulates circRNA expression, alternative splicing, and aging in C. elegans.
  43. Circadian Regulation of m6A RNA Methylation in Migraine: Mechanisms and Therapeutic Implications.
  44. Mechanisms of astragalus polysaccharide enhancing STM2457 therapeutic efficacy in m6A-mediated OSCC treatment.
  45. Nuclear pore links Fob1-dependent rDNA damage relocation to lifespan control.
  46. SNHG5 enhances colorectal cancer metastasis through RNA-protein interaction with GNB2 and activation of canonical Wnt signaling.
  47. Recent Advances in Chemical Probing Strategies for RNA Structure Determination in Vivo.
  48. Recognition of small Tim chaperones by the mitochondrial Yme1 protease.
  49. RNA-binding protein Epiplakin 1 promotes ovarian cancer immune escape and distal metastasis by enhancing the RNA stability of yes-associated protein.
  50. Mobile RNAs as systemic signaling beyond boundaries for plant stress resistance.
  51. RNA binding motif protein 15 promotes intracerebral hemorrhage progression by regulating mitophagy through E2F transcription factor 1.
  52. A fold switch regulates conformation of an alphavirus RNA-dependent RNA polymerase.
  53. Contributions of selenoproteins to breast cancer etiology and racial disparity.
  54. Structure of the nucleoli in the spermatocytes of three canid species.
  55. Structural plasticity enables broad cAn binding and dual activation of CRISPR-associated ribonuclease Cdn1.
  56. Optimizing Single-Cell Long-Read Sequencing for Enhanced Isoform Detection in Pancreatic Islets.
  57. PIWIL4 regulates gene expression and piRNA levels in RSV-infected airway epithelial cells.
  58. Tuning nuclear rheology through transient chromatin cross-links.
  59. Persulfidation of the zinc finger protein ZRANB2 modulates its RNA binding and alternative splicing function.
  1. Nucleic Acids Res. 2026 Jan 14. pii: gkaf1533. [Epub ahead of print]54(2):
    eIF3g binding to GUCG boxes located in mRNA coding regions enhances translation of mild heat shock response genes in the yeast Saccharomyces cerevisiae.
    Hiroaki Kato, Akihiro Oguro, Yuanhui Mao, Kinsley Ochsner, Hailey Casey, Grace Sakai, Ji Wan, Shingo Usuki, Leiming Tang, Mizuki Asano, Nirmala Bardiya, Kade Kaufman, Susumu Ishiguro, Naoki Tani, Kazuyuki Kumagai, Akira Nakamura, Chingakham Ranjit Singh, Taiichi Sakamoto, Eiji Obayashi, Shu-Bing Qian, Katsura Asano.
      Eukaryotic translation initiation factor 3 (eIF3) is a multi-subunit complex that promotes ribosome recruitment and messenger RNA (mRNA) selection. Here, we show that its eIF3g subunit, along with the binding partner eIF3i, mediates transcript-specific translation under mild heat stress through direct RNA binding. First, SELEX experiments identified a short GUCG-centered motif preferentially recognized by eIF3g, suggesting a sequence-specific binding preference. Next, ribosome profiling of yeast eIF3i mutant revealed that mRNAs containing GUCG motifs in their 5' coding regions exhibit elevated ribosome occupancy in a manner dependent on eIF3g/eIF3i module. A subset of SELEX-identified motifs, collectively termed the GUCG box, was found enriched in the 5'-terminal coding region of the regulated mRNAs. Reporter assays confirmed that these 5'-terminal coding regions are sufficient to drive heat-induced translation. Mutational analyses and biolayer interferometry demonstrated that disruption of the GUCG motif impairs eIF3g binding and diminishes translational induction. Moreover, GUCG motifs are periodically distributed across coding sequences and enriched near start codons, consistent with their role in stabilizing initiating ribosomes. Overall, this study establishes the GUCG box as a bona fide eIF3g-binding motif and validates its functional importance in vivo. These findings provide new insight into how eIF3 mediates stress-adaptive translation through sequence-specific RNA recognition.
    DOI:  https://doi.org/10.1093/nar/gkaf1533
  2. Hum Genet. 2026 Jan 21. 145(1): 15
    Cockayne syndrome mutation in XPG activate the integrated stress response.
    Danhui Zhang, Max Hartmann, Zhouli Cao, Gaojie Zhu, Gregoire Najjar, Cagatay Günes, Steffen Emmert, Karin Scharffetter-Kochanek, Sebastian Iben.
      Xeroderma pigmentosum (XP) and Cockayne syndrome (CS) are diseases provoked by mutations in multifunctional proteins that are involved in DNA repair. DNA-repair deficiency explains the high cancer incidence of XP, whereas cancer-free CS, characterized by growth retardation, neurological degeneration, and premature aging does not present as a classical DNA-repair deficiency disorder. Here, we compared a severe combined XP/CS case provoked by XPG-mutation with an XP "only" patient cell line caused by mutation in the same XPG gene to carve out the pathogenic cellular disturbances that provoke CS. We identified RNA polymerase I transcription and rRNA maturation defects, a highly phosphorylated eukaryotic initiation factor 2 alpha (eIF2alpha), and a shift from cap- to internal ribosomal entry site (IRES) translation, indicating an activated integrated stress response in CS. Disturbances in ribosomal biogenesis and translational control might thus contribute to the development of CS.
    DOI:  https://doi.org/10.1007/s00439-025-02804-3
  3. RNA. 2026 Jan 22. pii: rna.080481.125. [Epub ahead of print]
    Uridine analogs prevent stress granule formation, not by blocking PKR recognition, but by inhibiting the synthesis of T7 RNA Polymerase byproducts.
    Sean J Ihn, Emily Jiang, Nevraj S KeJiou, Yifan E Wang, Laura Farlam-Williams, Alexander F Palazzo, Hyun O Lee.
      mRNA-based therapeutics are commonly produced through T7 RNA Polymerase-mediated in vitro transcription. Introducing these exogenous RNAs into human cells activates an RNA sensor Protein Kinase R (PKR), which suppresses translation initiation and reduces their therapeutic effectiveness. Incorporating uridine analogs into these transcripts prevents PKR activation and translation shutdown, but the underlying mechanism remains unclear. Here, we demonstrate that treating T7 RNA Polymerase-produced transcripts with RNase III, which selectively degrades double-stranded RNA (dsRNA), blocks PKR activation and downstream translation-inhibition events, including eIF2α phosphorylation and stress granule formation in human cells. Interestingly, dsRNAs generated with uridine analogs robustly induce eIF2α phosphorylation and stress granules to the same extent as dsRNA containing uridine. These findings indicate that uridine analogs do not prevent PKR from detecting dsRNA. Instead, we show that uridine analogs decrease the production of T7 RNA Polymerase byproducts, including antisense RNA and dsRNA, which activate PKR and downstream stress responses. Finally, we demonstrate that higher amounts of exogenous RNA, lacking T7 RNA Polymerase byproducts, can induce stress granules independently of PKR and phospho-eIF2α, but dependent on stress granule scaffold proteins G3BP1 and G3BP2. Together, our findings show that uridine analogs mitigate PKR signaling not by blocking mRNA-PKR interactions, but by minimizing dsRNA byproducts from T7 Polymerase transcription. Furthermore, stress granule formation in response to high levels of exogenous RNA can occur through a mechanism that does not depend on PKR but relies on G3BP1 and G3BP2. These insights clarify the role of uridine analogs in PKR activation and may inform future therapeutic RNA design.
    Keywords:  PKR; T7 RNA Polymerase; biomolecular condensates; stress granules; uridine analogs
    DOI:  https://doi.org/10.1261/rna.080481.125
  4. Nucleic Acids Res. 2026 Jan 22. pii: gkag036. [Epub ahead of print]54(3):
    A comprehensive view on r-protein binding and rRNA domain structuring during early eukaryotic ribosome formation.
    Magdalena Gerhalter, Michael Prattes, Lorenz Emanuel Grundmann, Irina Grishkovskaya, Enrico F Semeraro, Gertrude Zisser, Harald Kotisch, Juliane Merl-Pham, Stefanie M Hauck, David Haselbach, Helmut Bergler.
      Formation of the eukaryotic ribosomal subunits follows a strict regime to assemble ribosomal proteins (r-protein) with ribosomal RNAs (rRNA) while removing internal (ITS) and external (ETS) transcribed rRNA spacers. During the early stages of large subunit (LSU) formation, ITS2, together with six assembly factors, forms the characteristic foot structure of early nuclear pre-LSU particles. Here, we address the function of this foot structure during the early stages of ribosome assembly. We present cryo-EM structures from wild-type cells and cells depleted for the foot structure factor Rlp7. We show that compaction of domain I of the 25S rRNA is strictly dependent on the presence of foot factors, while domain II folds independently. Furthermore, Rlp7-depletion accumulated small subunit (SSU) processome intermediates prior to A1 cleavage and compaction of the individual domains of the 18S rRNA, providing also novel insights into the SSU-assembly process. SILAC labeling and affinity purification of co-transcriptionally assembled pre-ribosomes enabled us to resolve the assembly line of most early binding r-proteins step by step. This showed that incorporation of r-proteins in eukaryotes neither follows the bacterial regime nor a strict linear co-transcriptional mode. Instead, seed r-proteins might structurally define the individual rRNA domains before their compaction and fixation in the context of early pre-ribosomes.
    DOI:  https://doi.org/10.1093/nar/gkag036
  5. Nucleic Acids Res. 2026 Jan 14. pii: gkaf1528. [Epub ahead of print]54(2):
    YhfH functions as a tri-function RNA in Bacillus thuringiensis BMB171.
    Jiaxin Qin, Xia Cai, Yizhuo Zhang, Ziqi Wang, Feiyu Pan, Bing Yan, Jun Cai.
      Regulatory RNAs are crucial for transcriptional and posttranscriptional regulation in bacteria and hold significant potential as tools for gene expression in synthetic biology. In our previous study, YhfH in the Bacillus thuringiensis BMB171 strain was identified as a regulatory RNA that functions as an antisense RNA to modulate the expression of LipR, a transcriptional regulator involved in intracellular pH regulation in glucose-rich environments. This study further reveals that YhfH, predominantly expressed during the stationary phase, acts as a small RNA (sRNA) to regulate the expression of the pyrimidine biosynthetic operon, thereby influencing the bacterial growth in pyrimidine-limited media. Additionally, YhfH RNA serves as a messenger RNA (mRNA) encoding a peptide, YhfH-P, which inhibits its own transcription. Moreover, YhfH-P represses the expression of the ilv-leu operon, which is primarily responsible for branched-chain amino acid synthesis, by binding to its promoter. Collectively, YhfH RNA exhibits versatile functions as an sRNA, antisense RNA, and mRNA, thus acting as a "tri-function" RNA. The discovery of YhfH significantly expands our understanding of RNA regulatory potential, and its mechanism of action could provide valuable insights for designing streamlined genetic circuits with diverse regulatory functions in synthetic biology.
    DOI:  https://doi.org/10.1093/nar/gkaf1528
  6. Nucleic Acids Res. 2026 Jan 14. pii: gkaf1525. [Epub ahead of print]54(2):
    Synthesis of long and functionally active RNAs facilitated by acetal levulinic ester chemistry.
    Zidi Lyu, Adam Katolik, Iqra Yaseen, Adrain A Pater, Francis Robert, Sidong Huang, Keith T Gagnon, Peter J Unrau, Masad J Damha.
      Recent advances in RNA-based therapeutics have created a demand for synthetic RNAs that are 100 nucleotides (nts) or longer. In this study, we present the use of 2'-acetal levulinic ester (2'-ALE) phosphoramidites for the synthesis of long RNAs that are at least 215 nts in length. We have developed protocols for rapid (2-4 min) and efficient coupling (>99%) of 2'-ALE monomers and established a rapid, on-column deprotection of RNA strands requiring short alkylamine treatments at room temperature. The results of these studies enabled the successful syntheses of sgRNAs (99 nt), sgRNA tagged with fluorogenic Mango II and Broccoli aptamers (130-170 nt), and 5'-capped minimal mRNAs (200-215 nt), each exhibiting robust functional activity in both cell-free and cellular systems. We also found that the incorporation of 2'-O-methyl-adenosine in the poly(A) tail of synthetic mRNAs markedly enhanced protein expression, highlighting the ALE platform's compatibility for systematic exploration of RNA chemical diversity. Collectively, these results establish 2'-ALE chemistry as a promising platform for the synthesis of long and functionally active RNAs.
    DOI:  https://doi.org/10.1093/nar/gkaf1525
  7. Nucleic Acids Res. 2026 Jan 22. pii: gkag022. [Epub ahead of print]54(3):
    ADAM-tRNA-seq: an optimized approach for demultiplexing and enhanced hierarchal mapping in direct tRNA sequencing.
    Rodrigo Alarcon, Daniel Köster, Stine Behrmann, Zoya Ignatova.
      Transfer RNAs (tRNAs) play an essential role in protein synthesis and cellular homeostasis, and their dysregulation is associated with various human pathologies. Recent advances in direct RNA sequencing by the Nanopore platform have enabled simultaneous profiling of tRNA abundance, modifications, and aminoacylation status. However, the high sequence similarity among tRNAs and the lack of robust demultiplexing strategies reduce the accuracy and limit the scalability of current approaches. Here, we developed ADAM-tRNA-seq, a framework that addresses two key limitations of the Nanopore-based direct tRNA sequencing. First, we develop an RNA-based barcode demultiplexing method, that employs a barcode embedded in the sequencing adapter, recognized by the Dorado basecaller. Second, we designed a hierarchy-based mapping strategy that mitigates read loss due to multimapping by classifying reads at the isodecoder, isoacceptor, or isotype levels, thereby enhancing quantification accuracy. We validated ADAM-tRNA-seq using both synthetic tRNAs and a complex human tRNA pool, and systematically optimized it to achieve up to 99% classification precision. Together, these developments enable more accurate, scalable, and comprehensive characterization of tRNA pools across diverse sample types.
    DOI:  https://doi.org/10.1093/nar/gkag022
  8. Nucleic Acids Res. 2026 Jan 22. pii: gkag025. [Epub ahead of print]54(3):
    An elevated level of the mRNA exporter Mex67-Mtr2 in nuclear mRNPs impairs nuclear mRNA export.
    Nataliia Stefanyshena, Katja Sträßer.
      In eukaryotes, nuclear messenger RNA (mRNA) export is a crucial step in gene expression, mediated by the conserved mRNA exporter Mex67-Mtr2 in Saccharomyces cerevisiae and NXF1-NXT1 in humans. Mex67-Mtr2 is recruited to the mRNA by the adaptors Hpr1, Nab2, Yra1, and Npl3, which play important yet incompletely understood roles in this process. Here, we uncover that, counterintuitively, an excess of Mex67 in nuclear messenger ribonucleoprotein particles (mRNPs) impairs nuclear mRNA export. Cells lacking Hpr1, which exhibit a nuclear mRNA export defect, show elevated levels of Nab2, Yra1, and Mex67 in nuclear mRNPs. Remarkably, overexpression of either Nab2 or Yra1 in Δhpr1 cells suppresses this export defect and simultaneously decreases the Mex67 level in nuclear mRNPs to those of wild-type cells. Importantly, a nuclear mRNA export defect is not inherently associated with an elevated Mex67 level in nuclear mRNPs, indicating that the increased Mex67 level in nuclear mRNPs of Δhpr1 cells is likely the cause rather than the consequence of the nuclear mRNA export defect. Thus, the precise regulation of the Mex67-Mtr2 level in nuclear mRNPs is essential for efficient nuclear mRNA export.
    DOI:  https://doi.org/10.1093/nar/gkag025
  9. Front Immunol. 2025 ;16 1706984
    The emerging roles of METTL1-mediated tRNA m7G methylation in cancer development and immunotherapy.
    Qiang Wang, Xiulin Jiang, Yixiao Yuan, Chunhong Li.
      RNA modifications, particularly N7-methylguanosine (m7G), have emerged as critical epigenetic regulators in cancer biology. METTL1, a conserved S-adenosylmethionine-dependent methyltransferase, catalyzes m7G modification primarily on tRNA, often in complex with its cofactor WDR4. This modification stabilizes tRNA structure, protects it from degradation, and enhances the translation efficiency of specific codons, thereby enabling selective protein synthesis. Aberrant METTL1 expression has been observed across multiple cancer types-including lung, liver, colorectal, gastric, breast, cholangiocarcinoma, esophageal, glioma, head and neck, and thyroid cancers-where it promotes tumor proliferation, metastasis, therapy resistance, and metabolic reprogramming. Mechanistically, METTL1-mediated tRNA m7G modification influences downstream mRNA stability and translation, affecting oncogenes, drug resistance genes, and key metabolic regulators. Moreover, METTL1 shapes the tumor immune microenvironment by modulating immune cell infiltration, promoting immunosuppressive populations, and contributing to immune evasion, which has implications for immunotherapy. Collectively, METTL1 functions as a pivotal driver of cancer progression and represents a promising biomarker and therapeutic target, highlighting the potential of targeting tRNA m7G modification in precision oncology.
    Keywords:  METTL1; biomarker; cancer progression; epigenetic regulation; immunotherapy; tRNA modifications; tumor immunity; tumor microenvironment
    DOI:  https://doi.org/10.3389/fimmu.2025.1706984
  10. Nat Commun. 2026 Jan 22. 17(1): 160
    Meta-unstable mRNAs in activated CD8+ T cells are defined by interlinked AU-rich elements and m6A mRNA methylation.
    Paulo A Gameiro, Iosifina P Foskolou, Yumna A Butt, Aniek A M Martens, Klara Kuret Hodnik, Igor Ruiz de Los Mozos, Nordin D Zandhuis, Žan Hozjan, Veronica M Kot, Rupert Faraway, Michiel Vermeulen, Monika C Wolkers, Randall S Johnson, Jernej Ule.
      CD8+ T cells can rapidly produce effector molecules following activation. This activation triggers rapid changes in gene expression that rely on the control of mRNA levels via multiple mechanisms, including RNA modifications. N6-methyladenosine (m6A) is an abundant post-transcriptional modification that promotes the decay of messenger RNAs in the cytosol. However, how recognition of m6A sites is integrated with other regulatory mechanisms that alter the fate of immunoregulatory mRNAs in CD8+ T cells remains unexplored. Here, we apply the m6A-iCLIP and GLORI methods to identify the importance of m6A sites flanked by AU-rich elements (AREs) within the 3'UTRs of CD8+ T cell mRNAs. Presence of such ARE-flanking m6A motifs predicts meta-unstable mRNAs that rapidly decay upon CD8+ T cell activation. We demonstrate interdependent effects of mutations in the identified AREs and RRACHs on TNF mRNA stability. The ARE-flanking m6A sites in these mRNAs show particularly high iCLIP crosslinking of YTHDF proteins, which are also identified by proteomic interactome analyses along with additional novel RNA-binding proteins. Our study reveals a crosstalk between m6A and ARE-dependent mechanisms in CD8+ T cells, providing new approaches for modulating mRNA decay in T cell activation.
    DOI:  https://doi.org/10.1038/s41467-025-67762-w
  11. NAR Cancer. 2026 Mar;8(1): zcaf058
    Ribosome biogenesis is increased in hepatocellular carcinoma and represents a potential therapeutic target.
    Sille Blangstrup Geisler, Kezia Catharina Oxe, Stavroula Boukoura, Ekaterina Dulina, Dorthe Helena Larsen.
      Globally, liver cancer is the sixth most prevalent cancer type and the third leading cause of cancer-related deaths, making the need for improved treatment evident. We conducted a pan-cancer tissue microarray analysis to identify cancer types with upregulated ribosome biogenesis, potentially suitable for treatment with nucleolar-targeting compounds. Our screening identified liver cancer as a potential candidate. Gene expression analysis confirmed upregulation of nucleolar factors facilitating ribosome biogenesis that correlated with poor prognosis. In hepatocellular carcinoma (HCC) cell lines, constituting around 80% of liver cancer cases, we confirmed the upregulation of the nucleolar proteins Treacle, UBF, and Fibrillarin, involved in transcription and processing of ribosomal RNA (rRNA). Measurements of rRNA also confirmed increased nucleolar activity. We treated the HCC cell lines with nucleolar-targeting compounds and observed increased sensitivity in the HCC cell lines. Notably, nucleolar targeting compounds demonstrated a broader therapeutic window than that observed for Sorafenib, a clinically approved targeted therapy. Furthermore, we investigated how nucleolar factors change during HCC stages and found a progressive increase in Treacle and Fibrillarin in advanced stages of HCC. Our results demonstrate aberrant nucleolar activity in HCC and propose targeting ribosome biogenesis as a therapeutic strategy to improve HCC patient outcomes.
    DOI:  https://doi.org/10.1093/narcan/zcaf058
  12. Nucleic Acids Res. 2026 Jan 22. pii: gkag033. [Epub ahead of print]54(3):
    The GT1 domain of RNase J ensures RNA quality control through dsRNA binding in Arabidopsis plastids.
    Kevin Baudry, Sébastien Skiada, Carmit Burstein, Varda Liveanu, Arnaud Liehrmann, Maddie Ceminsky, Oded Kleifeld, Etienne Delannoy, Alexandra Launay-Avon, Benoît Castandet, Gadi Schuster, David B Stern.
      RNase J (RNJ) is a ribonuclease found in bacteria, archaea, and plant chloroplasts, and plays diverse roles in RNA maturation and stability. Chloroplast RNJ is encoded by the nuclear RNJ locus and is essential for embryo maturation. Arabidopsis or tobacco plants depleted for RNJ accumulate massive amounts of double-stranded RNA (dsRNA), which interferes with translation and causes chlorosis. Land plant RNJ uniquely contains a C-terminal GT1 domain, a DNA-binding motif found in transcription factors. Here, we have used complementation of an Arabidopsis rnj mutant with versions of RNJ with a mutated or deleted GT1 domain to investigate its role in RNJ function. We show that in vitro, the recombinant GT1 domain binds both dsRNA and DNA, but not single-stranded nucleic acids, with no sequence specificity. Furthermore, while RNJ lacking GT1 binding complements the rnj mutant, these plants accumulate high levels of dsRNA as detected by immunolocalization and RNA-Seq. GT1 mutations also change RNJ solubility in vivo, suggesting that the GT1 domain is involved in localization within the plastid. Taken together, our results suggest that the GT1 domain plays a key role in dsRNA removal through localizing the enzyme and/or selectively binding the dsRNA substrate.
    DOI:  https://doi.org/10.1093/nar/gkag033
  13. Nucleic Acids Res. 2026 Jan 22. pii: gkaf1539. [Epub ahead of print]54(3):
    Staufen dsRNA-binding domain as modules to design bifunctional antibodies for siRNA delivery.
    Yahui Liu, Yan Zheng, Ruolin Xu, Yanan Quan, Wanyi Tai.
      Antibody-small interfering RNA (siRNA) conjugates present an opportunity to expand the siRNA therapy to extrahepatic tissues. However, their investigation is now only confined to a limited number of targets, partially owing to some flaws in structures. Here, we described a modular design of bifunctional antibody that tethers siRNA without conjugation, yielding a diligent one-to-one antibody-siRNA pairing structure feasible for target expansion, charge masking, and further functionalization. Focusing on a noncationic siRNA-recruiting module, Staufen1 dsRBD34, we demonstrated that bifunctional antibodies recruit siRNA independent of base modification and enable target gene silencing on multiple cell types at a stoichiometry (1/1). Notably, by functionalizing siRNA terminus with small-molecule enhancers, the silencing potency of this pairing system can be augmented by seven times (IC50 from 200 to 28 nM) through the endosome-to-cytosol import. Affinity maturation by arginine scanning yields the 32 times higher affinity of dsRBD34 to siRNA, but the augment led to neither stronger silencing nor higher stability in mouse plasma as compared to p19 protein. The competition from sulfated GAGs in circulations can alter the pharmacokinetics of pairs and prevent a practical assessment of their potential in vivo. Altogether, bifunctional antibodies here possess notable properties, but ultrahigh-affinity dsRNA-binding domain is necessary to realize applications.
    DOI:  https://doi.org/10.1093/nar/gkaf1539
  14. Nucleic Acids Res. 2026 Jan 14. pii: gkag003. [Epub ahead of print]54(2):
    The two-step purification method ViREn identifies a single NSUN6-mediated 5-methylcytosine modification promoting dengue virus RNA genome turnover.
    Chia Ching Wu, Isabel S Naarmann-de Vries, Jonas Hartmann, Zarina Nidoieva, Kevin Kopietz, Virginie Marchand, Zeynep Özrendeci, Doris Lindner, Sophia Schelchshorn, Sophia Flad, Michaela Frye, F Nina Papavasiliou, Tanja Schirmeister, Georg Stoecklin, Johanna Schott, Yuri Motorin, Francesca Tuorto, Christoph Dieterich, Mark Helm, Alessia Ruggieri.
      Chemical modifications on cellular and viral RNAs are new layers of post-transcriptional regulation of cellular processes, including RNA stability and translation. Although advances in analytical methods have improved the detection of RNA modifications, precise mapping at single-base resolution remains challenging. Requirements for sensitivity and purity limit accuracy and reproducibility, especially for low abundant viral RNAs extracted from infected cells. Here, we report a two-step method, ViREn, for the enrichment of the genomic RNA (gRNA) of dengue virus (DENV), a positive-sense single-stranded RNA virus. This approach enabled the preparation of gRNA with significantly increased purity and led to the identification of a single high-confidence 5-methylcytosine (m5C) site in DENV gRNA at position 1218. This finding was orthogonally validated by Illumina-based bisulfite sequencing and by Nanopore Oxford Technologies direct RNA sequencing. Strikingly, m5C1218 was detected exclusively in gRNA extracted from infected cells but not in gRNA extracted from viral particles. We identified NSUN6 as the host methyltransferase catalyzing this modification and demonstrated a role for m5C in regulating DENV gRNA turnover. ViREn thus enables the mapping of m5C on low-abundance viral gRNA with unprecedented precision and sensitivity and facilitates future mechanistic studies into the role of RNA modifications in virus replication.
    DOI:  https://doi.org/10.1093/nar/gkag003
  15. Compr Physiol. 2026 Feb;16(1): e70105
    The Integrated Stress Response in Cancer: Paradox and Therapeutic Promise.
    Chenliang Zhang, Ting Zhang, Qiulin Tang, Yu Zeng, Dan Cao.
      The integrated stress response (ISR) is an evolutionarily conserved signaling pathway that converges diverse cellular stresses onto the eIF2α-ATF4 axis, thereby orchestrating a fundamental decision between adaptive survival and cell death. In cancer, malignant cells exploit the ISR to cope with microenvironmental pressure, yet strong or persistent ISR activation can also trigger apoptosis, highlighting its therapeutic potential. However, this duality complicates the targeting of the ISR for cancer therapy. In this review, we systematically outline the upstream regulators and downstream effector networks of the ISR, analyze its context-dependent functions, encompassing both tumor-promoting and tumor-suppressing activities, in tumorigenesis, immune modulation, and therapy resistance, and evaluate the rationale for targeting the ISR under defined conditions. We also comprehensively summarize and discuss recent advances in ISR-targeting agents, including both inhibitors and activators, under preclinical and clinical development, assessing their potential and current constraints. Although numerous challenges remain in therapeutically harnessing the ISR, we conclude that a deeper mechanistic understanding of how the ISR governs cell fate will further establish the ISR as a promising and actionable target for future cancer therapeutics.
    Keywords:  ATF4; apoptosis; drug resistance; eIF2α; integrated stress response; tumor immunotherapy; tumor therapy
    DOI:  https://doi.org/10.1002/cph4.70105
  16. Int J Mol Med. 2026 Mar;pii: 66. [Epub ahead of print]57(3):
    [Expression of Concern] Long non‑coding RNA SNHG20 promotes bladder cancer via activating the Wnt/β‑catenin signalling pathway.
    Qingsong Zhao, Saiyue Gao, Qingyan Du, Ye Liu.
      Following the publication of this paper, it was drawn to the Editor's attention by a concerned reader that the control GAPDH western blots featured in Fig. 5A and B on p. 2845 were apparently the same (even though it is possible that the experiments portrayed in these figure parts were performed under the same experimental conditions). The authors have been contacted by the Editorial Office to offer an explanation for the matter described above, although up to this time no response from them has been forthcoming. Owing to the fact that the Editorial Office has been made aware of potential issues surrounding the scientific integrity of this paper, we are issuing an Expression of Concern to notify readers of this potential problem while the Editorial Office continues to investigate this matter further. [International Journal of Molecular Medicine 42: 2839‑2848, 2018; DOI: 10.3892/ijmm.2018.3819].
    Keywords:  SNHG20; Wnt/β‑catenin signalling pathway; bladder cancer
    DOI:  https://doi.org/10.3892/ijmm.2026.5737
  17. Mol Immunol. 2026 Jan 20. pii: S0161-5890(26)00009-X. [Epub ahead of print]190 101-109
    Inhibition of RNase L enhances the expression efficiency of mRNA-LNPs.
    Hong-My Nguyen, Britteny Cassaidy, Mark Collinge, James C Hickey, Jin Li, Amir Arellano-Saab, Steven W Kumpf, Mitchell Thorn.
      mRNA-LNPs offer a promising platform for therapeutic protein expression, however, achieving efficient and sustained translation remains a significant challenge. One of the major barriers to mRNA-LNP efficacy is the activation of innate immune responses that recognize foreign RNA and suppress subsequent protein synthesis. Among these, the OAS-RNase L pathway, involved in degradation of cytoplasmic mRNA, plays a key role. This study examined the impact of RNase L and RNase L blockade on mRNA-LNP expression efficiency. In THP-1 cells, which express high endogenous levels of RNase L, both genetic ablation and pharmacological inhibition of RNase L led to a marked increase in protein expression. In contrast, HeLa cells, which exhibit low RNase L expression, showed minimal response to RNase L inhibition. In human peripheral blood mononuclear cells (PBMCs), RNase L inhibition also enhanced mRNA expression, while blocking other RNA sensors such as TLR7/8, RIG-I, TLR3, or MAVS, did not. Activation of the OAS-RNase L pathway may be driven by double-stranded secondary structure formed by therapeutic mRNA, resulting in mRNA recognition and degradation. RNase L acts as a key post-transcriptional regulator of mRNA stability and translation. Targeting this pathway offers a strategy to improve the performance of mRNA-based therapeutics.
    Keywords:  2-5 A; DsRNA; Innate immunity; Lipid nanoparticles (LNPs); MRNA-LNPs; OAS-RNase L pathway; Protein translation; RNA degradation; RNA medicine; RNase L
    DOI:  https://doi.org/10.1016/j.molimm.2026.01.003
  18. Nucleic Acids Res. 2026 Jan 14. pii: gkag008. [Epub ahead of print]54(2):
    Human mitochondrial helicase Twinkle has RNA binding, annealing, and strand-exchange activities.
    Anupam Singh, Laura C Johnson, Noe Baruch-Torres, Y Whitney Yin, Smita S Patel.
      Twinkle is the sole replicative helicase in human mitochondria, essential for mitochondrial DNA replication. Beyond its canonical unwinding activity, Twinkle has non-canonical activities, including DNA annealing and strand-exchange. Here, we show that these non-canonical activities extend to RNA. Twinkle binds RNA and catalyzes RNA:DNA hybrid formation through annealing, strand-exchange, and toehold-mediated strand displacement. Twinkle can unwind RNA:DNA forks when loaded onto the DNA tail but not the RNA tail. Although the physiological role of these RNA-related activities remains unclear, we show that Twinkle can strand-exchange an RNA downstream of a stalled replication fork to restart replication. The annealing/strand-exchange activity can be involved in DNA replication initiation and repair, but RNA:DNA hybrids can compromise genome integrity, emphasizing the need to balance unwinding and annealing activities. Interestingly, mitochondrial SSB inhibits the RNA:DNA annealing activity of Twinkle, thus regulating the non-canonical functions of Twinkle. A disease-associated W315L variant, which is defective in DNA replication, retains annealing and strand-exchange functions with both RNA and DNA, resulting in an imbalance between replication and annealing functions that may underlie its pathogenicity. Our findings of Twinkle's RNA-binding and strand-exchange activities may have a connection to its localization within mitochondrial RNA granules.
    DOI:  https://doi.org/10.1093/nar/gkag008
  19. Plant Biol (Stuttg). 2026 Jan 18.
    Nonsense-mediated decay as RNA quality control mechanism: An opportunity for developing viral interference in plants.
    V Rana, R Luha, V Kumar.
      Nonsense-mediated decay (NMD) is a crucial RNA surveillance mechanism that not only prevents synthesis of harmful truncated proteins but also keeps an eye on the quality and quantity check of mRNA in an efficient manner for effective gene expression. Viruses are in a continuous arm race with their host organisms to establish infection. Hence, hosts also constantly evolve new means of defence to exploit and suppress infection by the virus. In maximizing coding capacity in a small genome, several families of plant RNA viruses were documented to have NMD substrate features as a by-product. Although these features aid in viral infection, they are also susceptible to the host NMD pathway. In response, some viruses have evolved to bypass or disrupt the host NMD mechanism via cis-acting or trans-acting viral factors. By targeting the NMD pathway, it is possible to reduce the stability of pathogenic RNAs, leading to decreased pathogen viability and resistance. This review explores the mechanisms of NMD, its role in RNA quality control and the potential applications of manipulating NMD to combat pathogen resistance in agricultural and natural ecosystems. This review discusses the viral system circumventing the host NMD for their successful replication in plants. Moreover, an increased number of studies on the evasion mechanism of RNA viruses from host NMD provide insight into the molecular means of the NMD mechanism and may be linked to device defence strategies in agro-economic plants, which is an interesting future research possibility.
    Keywords:  RNA quality control; mRNA surveillance; nonsense‐mediated decay; pathogen resistance
    DOI:  https://doi.org/10.1111/plb.70176
  20. NPJ Precis Oncol. 2026 Jan 22.
    Integrative analysis of nanopore direct RNA sequencing data reveals a global impact of pseudouridylation on m6A and m5C modifications.
    Mohit Bansal, Anamika Gupta, Jane Ding, Anirban Kundu, Katherine Marlow, Andrew Gibson, Zhangli Su, Sunil Sudarshan, Han-Fei Ding.
      RNA modifications play a crucial role in regulating cellular functions. Among the most abundant modifications in the human transcriptome are pseudouridine (Ψ), N6-methyladenosine (m6A), and 5-methylcytosine (m5C). However, the interplay between these modifications remains poorly understood due to limited integrative studies. To address the gap, we utilized nanopore direct RNA sequencing to quantify the stoichiometry of Ψ, m6A, and m5C after depleting the pseudouridine synthases PUS7 or DKC1. We used the custom tool NanoPsiPy to quantify pseudouridine by analyzing differential U-to-C base-calling errors in nanopore sequencing data. For m6A and m5C, we applied the established tool CHEUI to conduct stoichiometry differential analysis. Our investigation identified both known and novel pseudouridylation sites in tRNA, rRNA, and mRNA targeted by PUS7 or DKC1. Integrative analysis revealed that depletion of PUS7 or DKC1 reduced pseudouridylation levels while simultaneously increasing global m6A and m5C levels, with functional implications for mRNA translation regulation. These findings suggest that pseudouridylation may play an active role in repressing m6A and m5C modifications. This study demonstrates the analytical power of nanopore direct RNA sequencing for investigating co-regulation of RNA modifications.
    DOI:  https://doi.org/10.1038/s41698-026-01278-4
  21. Genome Biol Evol. 2026 Jan 19. pii: evag009. [Epub ahead of print]
    Degradation determinants are abundant in human noncanonical proteins and minor annotated isoforms.
    Claudio Casola, Adekola Owoyemi, Nikolaos Vakirlis.
      The comprehensive characterization of human proteins, a key objective in contemporary biology, has been revolutionized by the identification of thousands of potential novel proteins through ribosome profiling and proteomics. Determining the physiological activity of these noncanonical proteins has proven difficult, because they are encoded by different types of coding regions and tend to share no sequence similarity with canonical polypeptides. Evidence from immunopeptidomic assays combined with a better understanding of the quality control of protein synthesis suggest that many noncanonical proteins may possess low stability in the cellular environment. Here, we tested this hypothesis by analyzing the frequency of multiple sequence features eliciting either proteasomal degradation or autophagy across 91,003 canonical (annotated) protein isoforms and 11,499 noncanonical proteins. Overall, noncanonical proteins were enriched for degradation-related features compared to all canonical proteins. Notably, degradation determinants were also enriched in canonical protein isoforms starting with a non-methionine amino acid. Analyses of original and shuffled sequences showed evidence of selective pressure either against or towards the accumulation of specific degradation signatures only in major isoforms of canonical proteins. However, stability was significantly higher in noncanonical proteins with evidence of phenotypic effects upon knock-out in cell lines. Notably, we found that the C-terminal tail hydrophobicity represents a reliable proxy for degradation propensity with potential applications in identifying functional noncanonical proteins. These findings underscore the critical role of degradation processes in regulating the half-life of noncanonical proteins and demonstrate the power of degradation-associated signatures in discriminating noncanonical genes likely to encode biologically functional molecules.
    Keywords:  cryptic proteins; microproteins; noncanonical ORFs; proteasome; protein degradation
    DOI:  https://doi.org/10.1093/gbe/evag009
  22. BMC Bioinformatics. 2026 Jan 22.
    QuAPPro: an R shiny app for quantification and alignment of polysome profiles.
    Chiara Schiller, Matthias Lemmer, Sonja Reitter, Janina A Lehmann, Kai Fenzl, Johanna Schott.
       BACKGROUND: Polysome profiling is a widespread technique to study mRNA translation. After separation of cellular particles by ultracentrifugation on a sucrose-density gradient, a UV absorbance profile is recorded during elution, which mostly reflects RNA content and shows distinct peaks for ribosomal subunits, monosomes and polysomes with increasing number of ribosomes. This profile can be used to assess global translational activity, or to reveal changes in ribosome biogenesis and translation elongation. In addition, it is also possible to measure the association of fluorescently tagged proteins with ribosomal subunits or polysomes. Alignment and quantification of polysome profiles usually relies on spreadsheet programs, custom R/Python scripts or commercial software.
    RESULTS: With QuAPPro, we present the first interactive web app that allows quantification and alignment of polysome profiles, independently of the device or software that was used to generate the profiles. QuAPPro was written in R, with a graphical user interface implemented in R shiny. It supports interactive visualization and analysis of polysome profiles, including profile smoothing, baseline selection, alignment along a defined point on the x-axis, quantification of profile subsections and deconvolution for resolving individual peaks. Fluorescence profiles can be aligned and quantified in parallel. Finally, quantification results can be summarized and visualized as bar plots. Every interactive plot can be exported directly in a publication-ready format.
    CONCLUSIONS: This user-friendly tool does not only speed up the analysis of polysome profiles but also facilitates reproducibility and documentation of the process, without the need for programming abilities or commercial software.
    Keywords:  Polysome profiling; Polysomes; Protein biosynthesis; R shiny; Translation
    DOI:  https://doi.org/10.1186/s12859-026-06379-2
  23. JCI Insight. 2026 Jan 23. pii: e196411. [Epub ahead of print]11(2):
    The RNA binding protein Arid5a is an activator of TNF signaling in rheumatoid arthritis.
    Yang Li, Ipsita Dey, Shachi P Vyas, Alzbeta Synackova, Decheng Li, Erik Lubberts, Dana P Ascherman, Peter Draber, Sarah L Gaffen.
      Rheumatoid arthritis (RA) is characterized by joint inflammation and bone erosion. Understanding cytokine pathways, particularly those targeting TNF, is crucial for understanding pathology and advancing treatment development. Arid5a is a noncanonical RNA binding protein (RBP) that augments inflammation through stabilizing proinflammatory mRNAs and enhancing protein translation. We examined published datasets for ARID5A in human RA blood, T cells, and synovial tissues. A stromal cell line, epithelial cells, and primary synovial fibroblasts were used to assess the effect of TNF on Arid5a expression, localization, and function. To determine how TNF induces Arid5a, WT or Traf2-/- stromal cells were treated with NIK or IKK inhibitors. To evaluate the necessity of Arid5a in arthritis progression, Arid5a-/- mice were subjected to collagen-induced arthritis. ARID5A was elevated in patients with RA and reduced by anti-TNF therapy. TNF upregulated Arid5a through the NF-κB1/TRAF2 pathway, causing cytoplasmic relocalization. Arid5a stabilized proinflammatory transcripts and enhanced expression of chemokines that drive RA. Arid5a-/- mice were resistant to collagen-induced arthritis correlating with reduced Th17 cells in synovial tissue. Thus, Arid5a serves as a newly recognized signaling intermediate downstream of TNF that is elevated in human RA and drives pathology in murine CIA, potentially positioning this RBP as a possible therapeutic target.
    Keywords:  Arthritis; Autoimmunity; Chemokines; Cytokines; Immunology
    DOI:  https://doi.org/10.1172/jci.insight.196411
  24. Mol Microbiol. 2026 Jan 20.
    Rop-Mediated Suppression of RpoS Production Increases Resistance to Nitric Oxide.
    Takeshi Shimizu, Shin Suzuki, Ichiyou Fukumoto, Takashi Hamabata.
      We identified the RNA-binding protein Rop, encoded on the pOSAK1 plasmid of enterohaemorrhagic Escherichia coli (EHEC), as a novel factor that enhances nitric oxide (NO) resistance, although it has previously been reported to regulate plasmid copy number. The Rop-induced increase in NO resistance was significantly reduced in several small noncoding RNA (sRNA) gene-deficient EHEC mutants. Among these sRNAs, DsrA, ArcZ, and RprA were directly involved in the translational regulation of rpoS expression, suggesting that Rop modulates rpoS expression through sRNAs. To examine this mechanism, we generated sRNA gene-deficient mutants with an additional deletion of the 5' untranslated region (5' UTR) of rpoS, which is required for translational regulation. The increase in NO resistance by Rop was restored in the double mutant, suggesting that this phenotype is mediated by Rop-dependent interactions between sRNAs and the 5' UTR of rpoS mRNA. Furthermore, Rop promoted rpoS mRNA degradation, an effect that likely suppresses RpoS production and may thereby enhance NO resistance. Finally, an hfq-deficient EHEC mutant exhibited no increase in NO resistance in the presence of Rop, indicating that Hfq is essential for Rop-mediated NO resistance.
    Keywords:  Rop; RpoS; enterohemorrhagic Escherichia coli; general stress response; small RNA
    DOI:  https://doi.org/10.1111/mmi.70051
  25. Br J Haematol. 2026 Jan 19.
    The impact of scaRNA12 deregulation on p53-mediated cellular stress response in B-cell precursor acute lymphoblastic leukaemia.
    Martijn W C Verbeek, Mathijs A Sanders, Dwin G B Grashof, Remco M Hoogenboezem, Iris van Zuijen, Annemarieke Bos, Trui van Gulck, Rosan Olsman, Rishov Mukhopadhyay, Eric M J Bindels, Peter J M Valk, Harmen J G van de Werken, Vincent H J van der Velden, Stefan J Erkeland.
      Small nucleolar ribonucleic acids (snoRNAs) are a class of small non-coding RNAs involved in the post-transcriptional modification of ribosomal RNAs (rRNA) and small nuclear RNAs (snRNA). Mounting evidence indicates that specific snoRNAs are drivers of oncogenesis, but their role in B-cell precursor acute lymphoblastic leukaemia (BCP-ALL) is unknown. We found that a small subset of 30 snoRNAs is commonly deregulated in BCP-ALL. Small Cajal body-specific RNA 12 (scaRNA12) was the strongest downregulated snoRNAs in BCP-ALL. Forced expression of scaRNA12 in BCP-ALL cells largely recovered the level of uridine-46 pseudouridylation of U5 snRNA (U5:Ψ46), an important RNA modification for proper splicing and gene expression regulation. We found that scaRNA12 controls a set of genes that belong to pathways that are frequently affected in BCP-ALL samples. We show that scaRNA12 activates p53, which is commonly affected in BCP-ALL even in the absence of p53 mutations. We show that scaRNA12 expression decreased the expression of upstream p53 regulators and provide novel evidence for a role of scaRNA12 in p53 regulation. We found that forced expression of scaRNA12 in BCP-ALL increased p53 activity and significantly enhanced the sensitivity of BCP-ALL to chemotherapeutic reagents. Together, our results suggest a tumour-suppressing role for scaRNA12 in BCP-ALL.
    Keywords:  BCP‐ALL; doxorubicin; p53; scaRNA12; snoRNA
    DOI:  https://doi.org/10.1111/bjh.70336
  26. J Vis Exp. 2025 Dec 30.
    MeRIP-qPCR Assay for Detecting m6A Modification Levels of Specific RNA in Osteosarcoma Cells.
    Binghan Yan, Junli Chang, Suxia Guo, Peng Zhao, Fulai Zhao, Chujie Zhou, Junjie Tong, Xinyu Zhang, Yuping Hong, Mengchen Yin, Xingyuan Sun, Yanping Yang.
      RNA epitranscriptomic modification plays a critical role in the initiation and development of various cancers, among which N6-methyladenosine (m6A) is the most prevalent and functionally diverse modification. Here, we present a streamlined and reproducible protocol using MeRIP-qPCR to assess m6A modification levels of specific transcripts in osteosarcoma cells. In this assay, total RNA was extracted, followed by incubation with a specific anti-m6A antibody to immunocapture RNA of m6A modification. Then, RNA was fragmented, and enrichment of m6A-modified RNA fragments was achieved using protein G magnetic beads. The enriched RNA was released by proteinase K digestion and purified by RNA-binding beads. Lastly, RNA was subjected to reverse transcription and quantitative polymerase chain reaction (PCR) to assess m6A levels in candidate genes. The results have demonstrated that this method effectively distinguishes methylation differences and provides a reliable tool for exploring the functional mechanisms of m6A modification in osteosarcoma cells. In conclusion, MeRIP-qPCR offers a simple and highly specific approach for studying m6A modification in osteosarcoma cells. This protocol holds great potential for elucidating m6A regulatory networks and identifying novel therapeutic targets.
    DOI:  https://doi.org/10.3791/69880
  27. Nucleic Acids Res. 2026 Jan 22. pii: gkaf1434. [Epub ahead of print]54(3):
    Transcriptome-wide analysis of Arabidopsis DICER-LIKE1 RNA substrates.
    Nicolas G Bologna, Alexis Sarazin, Gregory Schott, Antoine Bouet, Natalia P Achkar, Belen Moro, Florence Jay, Uciel Chorostecki, Emanuel A Devers, Olivier Voinnet.
      In plants, DICER-LIKE1 (DCL1) orchestrates microRNA (miRNA) biogenesis by cleaving imperfect stem-loop precursors within primary transcripts (pri-miRNAs). However, the full spectrum of DCL1 RNA substrates remains unexplored. Here, we report transcriptome-wide RNA immunoprecipitation and deep-sequencing (RIP-Seq) analyses of the Arabidopsis catalytically inactive DCL1 (DCL1ci), designed to bind but not cleave its targets. In inflorescences, DCL1ci-RIP retrieved nearly all evolutionarily conserved MIRNA loci and uncovered many hitherto unknown young MIRNA loci. Extensive interactions with both pre-miRNA stem-loops and flanking single-stranded regions were detected, suggesting that DCL1 scans pri-miRNAs prior to stem-loop cleavage. Quantitative binding profiles resolved the specific contribution of paralogous MIRNA family members in inflorescences, enabling tissue-level discrimination of pri-miRNA engagement. The analysis also identified hundreds of DCL1ci-interacting non-MIRNA loci, including protein-coding genes, transposons, and intergenic regions, with many lacking canonical stem-loop structures. We show that DCL1 promotes 24-nt small RNA biogenesis mostly from helitron-derived transcripts via a pathway genetically distinct from RNA-directed DNA methylation. Moreover, we identify a conserved stem-loop in the DCL1 5'-UTR suggesting autoregulatory feedback control. Collectively, our study establishes DCLci-RIP as a robust noninvasive approach for profiling DCL substrates, broadens DCL1's functional landscape, and provides a foundation for dissecting dynamic DCL-RNA interactions across developmental and stress contexts.
    DOI:  https://doi.org/10.1093/nar/gkaf1434
  28. bioRxiv. 2025 Dec 06. pii: 2025.12.04.691911. [Epub ahead of print]
    Alternative probe chemistries for single-molecule analysis of long non-coding RNA.
    Kalika R Pai, Aimee M Martin, Julia R Widom.
      Single-molecule microscopy has been widely used to study the structure and dynamics of RNA, but extension to larger systems such as long noncoding RNA (lncRNA) has proven challenging. The length and complex nature of lncRNA limits the techniques that can be readily used to study these aspects, but methods such as single-molecule kinetic analysis of RNA transient structure (SiM-KARTS), where the binding of a short, complementary oligonucleotide probe is used to determine accessibility of a specific region of the RNA, are promising. However, adapting SiM-KARTS to systems as complex as lncRNA requires careful optimization of experimental variables that have not been thoroughly explored. In this work, SiM-KARTS, thermal denaturation experiments, and circular dichroism spectroscopy were used to analyze the binding behaviors of probes with alternative backbone chemistries, specifically DNA with locked nucleic acid residues incorporated and morpholinos. A segment of lncRNA that enabled control over the accessibility of the target sequence was used as a model. We show that optimizing probe backbone chemistry can allow for more precise distinction between different structures of the target RNA, and for fine-tuning of probe binding stability without the structural impacts that other variables such as ionic concentration may have. These results provide design principles for the application of SiM-KARTS to target RNAs of increased complexity such as lncRNA.
    DOI:  https://doi.org/10.64898/2025.12.04.691911
  29. Arch Biochem Biophys. 2026 Jan 16. pii: S0003-9861(26)00008-1. [Epub ahead of print] 110737
    Systems analysis of the kinetics of in vitro transcription from interactions of T7 RNA polymerase and DNA.
    Nathan M Stover, Marieke De Bock, Julie Chen, Jacob Rosenfeld, Maria Del Carme Pons Royo, Allan S Myerson, Richard D Braatz.
      The in vitro transcription reaction (IVT) is of growing importance for the manufacture of RNA vaccines and therapeutics. While the kinetics of the microscopic steps of this reaction (promoter binding, initiation, and elongation) are well studied, the rate law of overall RNA synthesis that emerges from this system is unclear. In this work, we show that a model that incorporates both initiation and elongation steps is essential for describing trends in IVT kinetics in conditions relevant to RNA manufacturing. In contrast to previous reports, we find that the IVT reaction can be either initiation- or elongation-limited depending on solution conditions. This initiation-elongation model is also essential for describing the effect of salts, which disrupt polymerase-promoter binding, on transcription rates. Polymerase-polymerase interactions during elongation are incorporated into our modeling framework and found to have nonzero but unidentifiable effects on macroscopic transcription rates. Finally, we develop an extension of our modeling approach to quantitatively describe and experimentally evaluate RNA- and DNA-templated mechanisms for the formation of double-stranded RNA (dsRNA) impurities.
    Keywords:  RNA polymerase; double‐stranded RNA (dsRNA); enzyme kinetics; mathematical modeling; transcription
    DOI:  https://doi.org/10.1016/j.abb.2026.110737
  30. Biophys J. 2026 Jan 21. pii: S0006-3495(26)00045-7. [Epub ahead of print]
    A moments-based approach for inferring mechanisms of transcriptional regulation using nascent RNA data.
    Kunwen Wen, Yu Liao, Jiandong Wang, Sandeep Choubey, Feng Jiao.
      Unraveling the kinetics of transcriptional regulation is a central challenge in molecular biology. Traditional inference methods based on cell-to-cell mRNA variability are limited because mRNA levels reflect not only transcriptional activity but also post-transcriptional processes. Recent advances in quantifying nascent RNAs across isogenic cells provide a more direct readout of transcriptional dynamics. Here, we present a moments-based theoretical framework that leverages nascent RNA data to infer transcriptional kinetics. We derive closed-form expressions for the first two time-dependent moments of nascent RNA across three canonical transcription models and demonstrate that nascent RNA exhibits distinct transient overshoots and higher Fano factors compared to mature mRNA. Through analysis of synthetic datasets, we show that these features enable more accurate estimation of transcriptional kinetics and allow reliable model selection, distinguishing single from multi-pathway promoter activation with over 75% accuracy even from small sample sizes. We further validated our model selection framework using experimental transcription datasets from yeast and E. coli cells, where nascent RNA data exhibited superior discriminatory power for identifying cross-talk pathway regulation. Our framework thus offers a computationally efficient and mechanistically informative approach for decoding transcriptional dynamics, emphasizing the advantages of time-dependent nascent RNA measurements over mature mRNA in capturing the true kinetics of transcriptional regulation.
    DOI:  https://doi.org/10.1016/j.bpj.2026.01.030
  31. Proc Natl Acad Sci U S A. 2026 Jan 27. 123(4): e2519809123
    Structural characterization of the HDV virion and its ribonucleoprotein.
    Samuel Itskanov, Beatrice Ary, Upasana Mehra, Irene Lew, Nikolai Novikov, Uli Schmitz, Meghan M Holdorf, Rudolf K Beran, Eric B Lansdon.
      Hepatitis D virus (HDV) is a small RNA satellite virus of hepatitis B virus (HBV) which encodes a single protein, HDV delta antigen (HDAg), that is required for replication. Viral replication occurs independently from HBV and relies primarily on host RNA polymerase(s). Bulevirtide, a viral entry inhibitor, is the only approved treatment for chronic HDV but has a low cure rate as a monotherapy, and most patients rebound following cessation of therapy. It is likely that an inhibitor targeting HDV replication is necessary to achieve HDV cure, but the paucity of HDV-derived elements and limited understanding of HDV replication presents a significant therapeutic challenge. Understanding the precise mechanism of interactions between HDAg and viral RNA, and how it is packaged within the virion can inspire structure-guided drug design targeting replication. Using cryoelectron tomography and single particle cryoelectron microscopy, we present reconstructions of the virion and viral RNPs. We observed multiple binding configurations in vitro that suggest a propensity to arrange four RNA segments around repeating units of HDAg in a ladder-like formation. The oligomerization domains of a homo-octameric HDAg complex are directly involved in RNA binding by utilizing the vertices and sides of its square-shaped architecture to bind RNA in a sequence-promiscuous fashion. Structure-function analysis reveals that these RNA contact sites are important for viral replication and their disruption may be a potential avenue for next-generation antivirals to treat HDV.
    Keywords:  HDV; cryo-EM; cryo-ET; ribonucleoprotein
    DOI:  https://doi.org/10.1073/pnas.2519809123
  32. Nucleic Acids Res. 2026 Jan 14. pii: gkaf1534. [Epub ahead of print]54(2):
    The HigB-like SehA toxin promotes non-replicating Salmonella inside macrophages by inhibiting ribonuclease III-dependent rRNA maturation.
    Soomin Choi, Yong-Joon Cho, Seungwoo Baek, Eunna Choi, Yoon Ki Kim, Tae Su Choi, Eun-Jin Lee.
      Many bacteria are often resistant to antibiotic treatment because they can slow down their growth rate, thereby attenuating the drug's effectiveness. A similar growth slowdown is observed in pathogens that persist inside their hosts. The bacterial toxin-antitoxin systems serve as tunable phenotypic switches by slowing down growth through the expression of a toxin component, though its biological target has been largely unknown. Here, we investigate the biological target of SehA, a HigB-like type II toxin from the intracellular pathogen Salmonella Typhimurium. Contrary to its predicted endoribonuclease function, it does not exhibit endoribonuclease activity. Instead, it inhibits ribonuclease III that mediates the initial cleavage for ribosomal RNA processing. The toxin binds to the dsRNA-binding domain of RNase III, thereby decreasing ribosome assembly and bacterial growth. Given that the SehA toxin induces persister formation within macrophages in an RNase III-dependent manner, inhibition of RNase III represents a newly identified biological target for pathogen persistence within hosts.
    DOI:  https://doi.org/10.1093/nar/gkaf1534
  33. Proc Natl Acad Sci U S A. 2026 Jan 27. 123(4): e2514994123
    Nuclear MBL-1 modulates mitochondrial morphology through carnitine palmitoyltransferase in Caenorhabditis elegans with toxic trinucleotide repeats.
    Joana Teixeira, Mikko J Frilander, Ove Eriksson, Susana M D A Garcia.
      Expansion of nucleotide repeat sequences is linked to a growing number of neuromuscular degenerative disorders. Metabolic changes, including disruptions in mitochondrial function and dynamics, characterize these disorders and are believed to contribute to organismal toxicity. To investigate how toxic RNA repeats affect mitochondria, we used a Caenorhabditis elegans model that expresses expanded CUG repeat RNAs in muscle cells and recapitulates muscle dysfunction. We found that the RNA-binding protein Muscleblind-like 1 (MBL-1) is essential for normal mitochondrial function and regulates organelle morphology. In animals expressing expanded CUG repeats, where MBL-1 function is impaired, we identified two distinct mechanisms of mitochondrial disruption: altered mitochondrial morphology regulated by MBL-1, and oxidative phosphorylation (OxPhos) dysfunction occurring independently of MBL-1. Our data further show that changes in mitochondrial morphology are specifically linked to nuclear MBL-1 dysfunction, which affects cpt-3 expression, a gene encoding carnitine palmitoyltransferase-an enzyme required for fatty acid transport into mitochondria. This mechanism is conserved, with similar disruptions observed in patients with Myotonic Dystrophy type 1. Importantly, our findings indicate that increased organelle fragmentation is not central to cellular pathogenesis. Instead, OxPhos dysfunction appears to be a primary contributor to organismal toxicity.
    Keywords:  Caenorhabditis elegans; RNA repeat toxicity; carnitine palmitoyl transferase; mitochondrial dysfunction; muscleblind-like
    DOI:  https://doi.org/10.1073/pnas.2514994123
  34. Nat Commun. 2026 Jan 20.
    Biochemical principles of miRNA targeting in flies.
    Joel Vega-Badillo, Phillip D Zamore, Karina Jouravleva.
      MicroRNAs direct Argonaute proteins to repress complementary target mRNAs via mRNA degradation or translational inhibition. While mammalian miRNA targeting has been well studied, the principles by which Drosophila miRNAs bind their target RNAs remain to be fully characterized. Here, we use RNA Bind-n-Seq to systematically identify binding sites and measure their affinities for five highly expressed Drosophila miRNAs. Our results reveal a narrower range of binding site diversity in flies compared to mammals, with fly miRNAs favoring canonical seed-matched sites and exhibiting limited tolerance for imperfections within these sites. We also identified non-canonical site types, including nucleation-bulged and 3'-only sites, whose binding affinities are comparable to canonical sites. These findings establish a foundation for future computational models of Drosophila miRNA targeting, enabling predictions of regulatory outcomes in response to cellular signals, and advancing our understanding of miRNA-mediated regulation in flies.
    DOI:  https://doi.org/10.1038/s41467-026-68360-0
  35. Front Cell Dev Biol. 2025 ;13 1722674
    A novel protein encoded by circIMP3 promotes prostate cancer progression by regulating alternative splicing and tumor microenvironment.
    Wenren Zuo, Weizhou Huang, Haojie Chen, Yan Xu, Yang Zhang.
       Introduction: Prostate cancer (PC) is one of the most prevalent malignancies in men, with rising incidence and mortality rates globally. Despite advances in therapeutic options such as androgen deprivation therapy and chemotherapy, effective cures, especially for advanced stages of the disease, remain limited. Recent research has highlighted the significant roles of alternative splicing (AS) and noncoding RNAs in tumor progression and drug resistance. This study aims to investigate the role of circIMP3, derived from the IMP3 gene, in prostate cancer development.
    Methods: In this study, we employed quantitative PCR, RNA sequencing, and immunoblotting to identify and characterize circIMP3 in prostate cancer tissues and patient blood samples. Functional assays, including cell proliferation and in vivo tumorigenicity assays, were conducted to assess the biological role of circIMP3 in PC cells. RNA immunoprecipitation sequencing (RIP-seq) was used to identify alternative splicing events regulated by circIMP3. Additionally, exosome isolation and uptake assays were performed to explore the paracrine signaling function of circIMP3 within the tumor microenvironment (TME).
    Results: We identified circIMP3, which is significantly upregulated in both prostate cancer tissues and peripheral blood of patients. CircIMP3 contains an internal ribosome entry site (IRES) and encodes a previously uncharacterized 288-amino-acid protein, circIMP3_288aa. Functional assays revealed that circIMP3_288aa promotes cell proliferation in vitro and accelerates tumor growth in vivo. Mechanistically, circIMP3_ 288aa regulates the alternative splicing of FBXW7, leading to impaired c-Myc ubiquitination and stabilization, which enhances oncogenic signaling. RIP-seq analysis identified over 2,000 alternative splicing events regulated by IMP3, with a notable enrichment in pathways related to ubiquitin-mediated proteolysis. Furthermore, circIMP3 is secreted into the TME via exosomes, where it is taken up by recipient cells, contributing to their proliferation.
    Discussion: Our findings demonstrate that circIMP3 acts as a key regulator of both intracellular alternative splicing and extracellular paracrine signaling within the TME. The ability of circIMP3 to influence FBXW7 splicing and stabilize c-Myc provides a mechanistic basis for its role in promoting oncogenesis in prostate cancer. Clinically, high expression levels of circIMP3 correlate with poorer event-free survival in prostate cancer patients, suggesting its potential as a prognostic biomarker. Additionally, the detection of circIMP3 in peripheral blood positions it as a promising target for liquid biopsy applications in PC diagnosis and monitoring.
    Keywords:  Fbxw7; IMP3; alternative splicing; c-Myc; circular RNA; prostate cancer
    DOI:  https://doi.org/10.3389/fcell.2025.1722674
  36. Front Cell Dev Biol. 2025 ;13 1683213
    A primer on tRNAs, tRNA-m1A58 modification, and tRNA derivatives in T-cell activation.
    Chuanxiang Zhao, Juxiang Shen.
      Upon antigen recognition, naive CD4+ T cells are activated, exiting quiescence to undergo rapid activation, clonal expansion, and differentiation into effector functions against pathogens. T-cell activation and clonal expansion necessitate the biosynthesis of millions of new protein copies. Recent technological advancements in small RNA sequencing have revealed a highly complex and dynamic repertoire of cellular tRNAs, tRNA-m1A58 modification, and tRNA derivatives during T-cell activation. This review outlines the basic framework of the biogenesis and biological functions of tRNAs, tRNA-m1A58 modification, and tRNA derivatives. Importantly, we elucidate how m1A58 modification regulates translation through multilevel mechanisms involving initiation, elongation, and termination. Furthermore, this review provides a comprehensive overview of the dynamic changes in tRNA expression repertoires and the impacts of tRNA-m1A58 modification and tRNA derivatives on T-cell activation. This review aims to offer novel insights into the molecular mechanisms underlying T-cell activation, facilitating the development of more effective therapeutic strategies for treating T-cell-related diseases.
    Keywords:  T cells; tRFs; tRNA; tRNA derivatives; tRNA-m1A58 modification
    DOI:  https://doi.org/10.3389/fcell.2025.1683213
  37. Protein Sci. 2026 Feb;35(2): e70456
    Concentration-dependent mutational scanning probes the cellular folding landscape of α-synuclein in yeast.
    Daeun Noh, Robert W Newberry.
      The misfolding and aggregation of α-synuclein is a central molecular event in the etiology of Parkinson's disease and related disorders. α-synuclein misfolding and pathology are both concentration-dependent, but it is not clear precisely how changes in concentration alter the folding landscape within cells. Whereas most conventional structural biology approaches offer limited resolution in living systems, deep mutational scanning can offer insight into the folding state of a protein in living cells, and we apply this method to probe concentration-dependent changes in the folding of α-synuclein in a popular yeast model of pathology. We discover that at a wide range of cellular concentrations, α-synuclein is highly biased toward formation of a membrane-bound amphiphilic helix that imparts toxicity. Population of this toxic state can be disrupted by mutations that reduce membrane affinity, which shift the folding equilibrium away from the membrane-bound state. Reduced-affinity variants exhibit distinct sensitivity to concentration relative to variants with WT-like affinity, likely because these variants are expressed at concentrations closer to their dissociation constant for membrane binding. These results show how mutational scanning can provide high-resolution insights into the folding landscape of proteins in living cells, which is likely to be of special utility for studying proteins that misfold and/or aggregate.
    Keywords:  concentration dependence; membrane binding; mutational scanning; protein misfolding; yeast; α‐synuclein
    DOI:  https://doi.org/10.1002/pro.70456
  38. Nat Chem Biol. 2026 Jan 19.
    Unconventional monooxygenation by the O2-dependent tRNA wobble uridine hydroxylase TrhO.
    Kiroo Shin, Da Bean Han, Hyun Woo Kim, Jungwook Kim.
      Modifications at the wobble position of transfer RNA (tRNA) are critical for accurate codon recognition and efficient translation. 5-Hydroxyuridine serves as a key intermediate for more complex wobble uridine derivatives commonly found in bacterial tRNAs and is synthesized by either prephenate-dependent TrhP or dioxygen-dependent TrhO. Despite its biological importance, structural and mechanistic insights into these enzymes have remained elusive. Here, we report the cryo-electron microscopy structure of Bacillus subtilis TrhO-tRNAAla complex. Combined with biochemical analyses, our results reveal that TrhO functions without any metal or organic cofactor, unlike most other oxygenases. We propose that the conserved C179 reacts with dioxygen to form a thiohydroperoxy intermediate, which is cleaved to produce 5-hydroxyuridine and a sulfenic acid at C179. The oxidized cysteine subsequently forms a disulfide bond with the adjacent C185, protecting the catalytic cysteine from irreversible overoxidation. These findings broaden our understanding of cofactor-independent dioxygen use in aromatic ring hydroxylation.
    DOI:  https://doi.org/10.1038/s41589-025-02129-2
  39. bioRxiv. 2025 Dec 02. pii: 2025.11.30.691457. [Epub ahead of print]
    Arsenic Trioxide Underpins Delayed Neuroinflammation and Impaired Synaptic Integrity involving Integrative Stress Response Signaling.
    Jasim Khan, Sazzad Khan, Himanshi Singh, Jianfeng Xiao, Daniel Johnson, Mohammad Athar, Mohammad Moshahid Khan.
      Arsenic trioxide (ATO), an industrial and environmental chemical with potential for accidental or intentional exposure, exerts profound neurotoxic effects. Its single acute exposure has been linked to delayed neurological and neurodegenerative outcomes. However, the molecular and cellular pathways driving these long-term manifestations remain poorly defined. In this study, combining human iPSC-derived neuronal and in vivo mouse model, we uncovered that the acute ATO exposure activates cellular stress pathway, which drives long-term neurotoxic and associated functional outcomes. We found that ATO induces integrated stress response (ISR) signaling in human iPSC-derived neurons. To model the impact of accidental ATO exposure in vivo , we administered a single high dose of ATO to C57BL/6J wild type mice. Four weeks post-treatment, ATO-exposed mice displayed neuropsychological symptoms and cognitive deficits. Brain analyses of ATO-challenged mice revealed elevated ISR activity marked by the increased phosphorylation of PERK and eIF2α and upregulation of the transcription factors, CHOP and ATF4. Transcriptomic profiling using bulk RNAseq revealed activation of pathways associated with stress and neuroinflammatory responses. Consistently, increased DNA damage and dysregulation of the STING-mediated innate immune response were also found in the brain of ATO-challenged mice. Pharmacological inhibition of the ISR with a small-molecule inhibitor, ISRIB mitigated ISR activation and preserved synaptic integrity in mouse hippocampal cells. In conclusion, our data identify ISR activation and DNA damage-driven immune dysregulation as key pathogenic drivers of ATO-induced delayed neurotoxicity and cognitive deficits and highlight ISR inhibitors as promising therapeutics to mitigate these effects.
    DOI:  https://doi.org/10.64898/2025.11.30.691457
  40. J Virol. 2026 Jan 21. e0145925
    Limited effect of short- to mid-term storage conditions on an Australian farmland soil RNA virome.
    Sabrina Sadiq, PeiPei Xue, Yijia Tang, Mingming Du, Kate Van Brussel, Alex B McBratney, Edward C Holmes, Budiman Minasny.
      Soils represent one of the largest and most diverse reservoirs of microbial life on Earth, yet their associated RNA viruses remain underexplored compared to animal and aquatic systems. Viral discovery in soils has been further limited by technical hurdles, particularly difficulties in obtaining sufficient yields of high-quality RNA for sequencing. To address this, we evaluated a range of storage and preservation strategies, including the use of commercial preservative solutions and ultra-cold snap-freezing, followed by standardized RNA extraction, sequencing, and virus discovery pipelines. This work aimed to establish minimum sample storage requirements that maintain RNA integrity, generate sufficient RNA sequencing data, and subsequently enable reliable soil virome characterization. While no preservative solution proved effective, "neat" soil samples were stable at 2°C-8°C and -30°C for at least 2 weeks, and at -80°C for at least 3 months, with no measurable reduction in RNA quality, sequencing data, or viral abundance and diversity. From 32 resulting libraries, we identified 1,475 putative novel RNA viruses, with the majority belonging to the microbe-associated phylum Lenarviricota. Several novel viruses formed divergent clusters with other environmentally derived sequences distantly related to traditionally animal-associated families, such as the Astroviridae and Picornaviridae. Furthermore, unique clusters within the Picobirnaviridae, Alsuvirucetes, Ghabrivirales, and Amabiliviricetes comprised exclusively Australian viruses, suggesting instances of region-specific evolution. Together, these findings highlight soils as rich reservoirs of RNA viral diversity and provide practical minimum standards for storage, expanding opportunities to investigate the ecological and evolutionary roles of RNA viruses in terrestrial systems.IMPORTANCERNA viruses are the most abundant and diverse biological entities on Earth and are likely present in all other organisms and ecosystems, including soil-dwelling invertebrates, microbes, and plants. Despite this, their diversity and role in soil systems remain largely unknown. Methodological challenges in preserving and extracting sufficient quantities of RNA from soils have hindered the study of these communities. Here, we identified 1,475 previously undescribed RNA viruses in Australian soils while systematically testing different preservation strategies. The significance of our research lies in the demonstration that snap-freezing soil is a viable and robust storage strategy for at least 3 months, while also highlighting the extraordinary scale of viral diversity present in terrestrial environments. This work establishes a foundation for reliable exploration of terrestrial RNA viruses, improving the accessibility of more remote environmental viromes and enabling future efforts to integrate them into broader models of microbial ecology and ecosystem function.
    Keywords:  Australia; RNA virus; ecology; environmental; meta-transcriptomics; preservation; soil
    DOI:  https://doi.org/10.1128/jvi.01459-25
  41. J Biochem Mol Toxicol. 2026 Feb;40(2): e70702
    CircDYRK1A Suppresses Gastric Cancer Progression via Sponging miR-331-5p to Regulate GADD45A.
    Shan Chen, JingFu Liu, Zhen Li, Xianren Ye.
      Numerous studies have demonstrated that circRNAs exhibit abnormal expression patterns in cancer and contribute to the regulation of malignant tumor progression. This study aims to investigate the role of circDYRK1A in the progression of Gastric Cancer (GC). Bioinformatics and qRT-PCR analyses revealed significant downregulation of circDYRK1A in GC tissues and cell lines. The circular structure and stability of circDYRK1A were confirmed via Sanger sequencing, electrophoresis, and RNase R treatment. Subcellular localization experiments indicated predominant cytoplasmic distribution. The effects of circDYRK1A on the functions of GC cells were evaluated through CCK-8, EDU proliferation and Transwell assays. The interaction between circDYRK1A and miR-331-5p/GADD45A was validated by dual luciferase assay, RNA immunoprecipitation, western blotting, and rescue assay. The research results showed that circDYRK1A was significantly down-regulated in GC and inhibited the proliferation, migration and invasion of GC cells by the miR-331-5p/GADD45A axis. This study lays a theoretical foundation for the molecular mechanism by which circDYRK1A inhibits the progression of GC and opens up new potential therapeutic approaches for targeted treatment of GC.
    Keywords:  circRNA; gastric cancer; miR‐331‐5p
    DOI:  https://doi.org/10.1002/jbt.70702
  42. G3 (Bethesda). 2026 Jan 22. pii: jkag016. [Epub ahead of print]
    The RNA-binding protein NOVA-1 regulates circRNA expression, alternative splicing, and aging in C. elegans.
    Emmanuel Adeyemi, Hussam Z Alshareef, Jaffar M Bhat, Pedro Miura, Alexander M van der Linden.
      Circular RNA (circRNA) biogenesis is regulated by RNA-binding proteins (RBPs) that alter back-splicing of exons in protein coding genes. However, few in vivo roles for RBPs in the regulation of circRNA biogenesis have been characterized. We previously showed that many circRNAs increase with age in C. elegans, and that loss of circ-crh-1, an abundant age-accumulated circRNA, extends mean lifespan. Given the established role of the mammalian RBP NOVA2 in promoting circRNA biogenesis, we investigated whether nova-1, the sole C. elegans homolog of NOVA1/2, similarly regulates circRNA expression and function in vivo. RNA-sequencing of nova-1 mutants compared to wild-type identified 686 circRNAs. Of these, 103 were differentially expressed in nova-1 mutants compared to wild-type, with 76 upregulated and 27 downregulated circRNAs, suggesting NOVA-1 acts as a negative regulator of a subset of circRNAs. nova-1 mutants also exhibited linear alternative splicing changes, primarily in alternative 3' splice site usage and exon skipping, and showed minimal overlap with circRNA loci. Notably, circ-crh-1 represented a shared regulatory target, suggesting NOVA-1 may coordinate splicing regulation with the production of crh-1 circRNAs. Motif analysis further revealed that over half of the NOVA-1-regulated splicing events contained YCAY motif sites, with crh-1 harboring a high density of sites, consistent with its alternative 3' splice site usage and circRNA production. Finally, nova-1 mutants exhibited an extended mean lifespan and enhanced heat stress recovery. Together, these findings identify NOVA-1 as a key regulator of circRNA expression and alternative splicing in C. elegans, with likely downstream consequences for organismal lifespan and stress resilience.
    Keywords:   C. elegans ; NOVA proteins; RNA-binding protein; WormBase; aging; alternative splicing; back-splicing; circ-crh-1; circRNA
    DOI:  https://doi.org/10.1093/g3journal/jkag016
  43. J Mol Neurosci. 2026 Jan 20. 76(1): 12
    Circadian Regulation of m6A RNA Methylation in Migraine: Mechanisms and Therapeutic Implications.
    Shikha Baghel Chauhan, Ayushi Bhandari, Chirag Jain, Indu Singh.
      Migraine is a complex neurological disorder showing distinct circadian rhythmicity in its onset and intensity. Attacks often follow daily patterns, suggesting that molecular clock mechanisms modulate neuronal excitability and pain signaling. Recent advances in neuroepigenetics identify RNA modifications particularly N6-methyladenosine (m6A) as rapid regulators of gene expression that respond to circadian cues. Exploring this circadian epitranscriptomic interaction may clarify time-of-day variations in migraine risk and drug response. This review integrates recent molecular and translational studies examining the interplay between m6A RNA methylation, circadian clock genes, and migraine pathophysiology. Evidence from transcriptomic, neurochemical, and pharmacological research was analyzed to understand how rhythmic RNA modifications affect calcitonin gene-related peptide (CGRP) signaling, neuroinflammation, and chronotherapeutic outcomes. Altered timing or function of m6A enzymes disrupts rhythmic transcription of core clock genes, enhancing CGRP release and inflammatory mediators such as interleukin-6, tumor necrosis factor-α, and nitric oxide. These changes heighten neuronal sensitivity and reduce the migraine threshold. Circadian variations in RNA methylation also influence drug-metabolizing enzymes and transporters, contributing to time-dependent differences in the efficacy of triptans, β-blockers, and CGRP antagonists.Integrating circadian and epitranscriptomic insights offers a pathway to precision migraine therapy. Profiling time-specific RNA modifications and tailoring drug administration to biological timing could improve efficacy, minimize side effects, and guide development of novel disease-modifying treatments targeting m6A-regulated pathways.
    Keywords:  Biomarkers; CGRP; Chronopharmacology; Chronotherapy; Circadian rhythms; Epitranscriptomics; M6A; Migraine
    DOI:  https://doi.org/10.1007/s12031-025-02468-8
  44. Med Oncol. 2026 Jan 20. 43(2): 122
    Mechanisms of astragalus polysaccharide enhancing STM2457 therapeutic efficacy in m6A-mediated OSCC treatment.
    Xi Wang.
      Oral squamous cell carcinoma (OSCC) is a prevalent malignancy with poor prognosis due to therapeutic resistance and tumor heterogeneity. N6-methyladenosine (m6A) modification, mediated by methyltransferase-like 3 (METTL3), drives OSCC progression via the HNRNPA2B1/FOXQ1 axis. STM2457, a selective METTL3 inhibitor, and Astragalus polysaccharide (APS), a traditional Chinese medicine with antitumor properties, hold promise for enhancing OSCC treatment, but their combined efficacy remains unexplored. In CAL27 OSCC cells, optimal concentrations of STM2457 (15 µM) and APS (100 µg/mL) were determined using EdU assays. Effects on m6A levels, METTL3, HNRNPA2B1, and FOXQ1 expression, and mRNA stability were assessed via RT-qPCR, Western blot, and RIP-qPCR. Cell proliferation, migration, invasion, and epithelial-mesenchymal transition (EMT) were evaluated using EdU, wound healing, and Transwell assays. In vivo efficacy was tested in nude mouse xenografts treated with STM2457 (50 mg/kg/day) and APS (80 mg/kg/day), with tumor growth and FOXQ1 expression analyzed. Combined STM2457 and APS treatment significantly reduced m6A levels, METTL3, HNRNPA2B1, and FOXQ1 expression, and mRNA stability compared to single-drug treatments, approaching or surpassing METTL3 silencing effects. The combination markedly suppressed cell proliferation, migration, invasion, and EMT, with increased E-cadherin and decreased N-cadherin levels. Regarding the role of APS in regulating m6A, we acknowledge that the current evidence is speculative and requires further mechanistic validation. In vivo, combination therapy significantly reduced tumor growth and FOXQ1 expression, outperforming single-drug treatments. STM2457 and APS enhance the inhibitory effects on OSCC progression by targeting the METTL3/HNRNPA2B1/FOXQ1 axis, offering a potential therapeutic strategy that integrates precision oncology with traditional herbal medicine. Further mechanistic and clinical studies are warranted to validate these findings.
    Keywords:  Astragalus polysaccharide; M6A; METTL3; Oral squamous cell carcinoma; STM2457
    DOI:  https://doi.org/10.1007/s12032-026-03254-1
  45. FEBS Open Bio. 2026 Jan 19.
    Nuclear pore links Fob1-dependent rDNA damage relocation to lifespan control.
    Yamato Okada, Mina Iwaki, Kyosuke Hagiri, Rei Izumi, Masahiko Harata, Chihiro Horigome.
      In budding yeast, the replication fork blocking protein Fob1 arrests replication forks at the ribosomal RNA gene (rDNA) locus, leading to DNA double-strand breaks that promote genomic instability and limit replicative lifespan. rDNA damage has been reported to drive exit from the nucleolus, and persistent double-strand breaks can relocate to the nuclear periphery, but how these spatial transitions are organized and how they influence genome stability and aging remain unclear. Here, we analyze the subnuclear localization of a site-specific rDNA break and its functional relationship with nuclear pores. Using quantitative fluorescence microscopy, we show that damaged rDNA accumulates at the nucleolar-nucleoplasmic interface adjacent to the nuclear envelope. This position represents the minimal movement required to leave the nucleolar interior while maintaining contact with the nuclear periphery, in a manner reminiscent of nucleolar caps of higher eukaryotes. Cells defective in nuclear pore association display pronounced rDNA instability that is largely, but not completely, suppressed by deletion of Fob1, with partial restoration of rDNA stability. Disruption of nuclear pore association also shortens replicative lifespan, and this defect is partially rescued by Fob1 deletion, indicating that nuclear pores affect longevity through both Fob1-dependent and Fob1-independent pathways. These findings refine current models of rDNA damage handling in budding yeast and support a role for nuclear pores in spatially organizing Fob1-induced rDNA damage to maintain rDNA stability and replicative lifespan.
    Keywords:  Fob1; Saccharomyces cerevisiae; genome stability; lifespan; nuclear pore; ribosomal RNA gene (rDNA)
    DOI:  https://doi.org/10.1002/2211-5463.70193
  46. Noncoding RNA Res. 2026 Apr;17 128-149
    SNHG5 enhances colorectal cancer metastasis through RNA-protein interaction with GNB2 and activation of canonical Wnt signaling.
    Xinyi Chen, Mu Yang, Xiaoxiao Luo, Xianglin Yuan.
       Background and purpose: Colorectal cancer (CRC) is one of the most prevalent and lethal malignancies worldwide, with distant metastasis-particularly to the liver-representing the primary cause of poor prognosis. Long non-coding RNAs (lncRNAs) have emerged as critical regulators of CRC progression, yet the mechanisms by which they modulate G protein signaling during hepatic metastasis remain unclear. This study aimed to determine the role of the lncRNA SNHG5 in CRC liver metastasis and to explore whether G protein-related mechanisms are involved in this process.
    Methods: We established murine MC38 CRC sublines with distinct metastatic capacities (F0 and F3) and performed RNA sequencing to identify key lncRNAs. Biotin-labeled RNA pull-down coupled with mass spectrometry was used to identify SNHG5-interacting proteins. The SNHG5-GNB2 interaction was validated using RIP, RNA-FISH, and Western blot analyses. Functional rescue assays, in vivo liver metastasis models, and Wnt pathway activity measurements were conducted to delineate downstream effects. Public transcriptomic datasets from GEO and TCGA were used to assess the expression patterns and prognostic relevance of SNHG5 and GNB2 in CRC and metastatic lesions.
    Results: SNHG5 was significantly upregulated in the highly metastatic F3 subline and predominantly localized in the cytoplasm. Pull-down and proteomic analysis identified GNB2, a classical G protein β-subunit, as a direct binding partner of SNHG5. Functionally, SNHG5 promoted cell proliferation, migration, epithelial-mesenchymal transition (EMT), and suppressed apoptosis, while GNB2 overexpression partially rescued the tumor-suppressive phenotypes induced by SNHG5 silencing. Mechanistically, the SNHG5-GNB2 axis enhanced Wnt/β-catenin signaling via increased p-GSK3β and β-catenin levels, thereby driving EMT. Transcriptomic analyses further revealed that GNB2 is upregulated in CRC and liver metastases and is associated with poor prognosis. Multi-omics data suggested additional roles for this axis in immune evasion, metabolic reprogramming, and remodeling of the metastatic microenvironment.
    Conclusion: This study provides the first evidence that SNHG5 promotes CRC liver metastasis through direct interaction with GNB2 and subsequent activation of the Wnt/β-catenin pathway. The SNHG5-GNB2 axis orchestrates a multilayered regulatory network that integrates EMT induction, immune suppression, and metabolic adaptation, highlighting its potential as a mechanistic driver and therapeutic target in metastatic CRC.
    Keywords:  Bioinformatics analysis; Bioinformatics databases; Colorectal cancer; GNB2; Liver metastasis; Long non-coding RNA; RNA–protein interaction; SNHG5; Wnt/β-catenin signaling
    DOI:  https://doi.org/10.1016/j.ncrna.2025.12.002
  47. Chemistry. 2026 Jan 20. e03291
    Recent Advances in Chemical Probing Strategies for RNA Structure Determination in Vivo.
    Maryana Yarshova, Jieyu Zhao, Chun Kit Kwok.
      The intricate structures of RNA molecules facilitate their diverse cellular functions. These structures are shaped by the cellular environment, a context that in silico and in vitro methods typically cannot reconstitute, making it more difficult to study the structure of RNA in cells. In response to these challenges, RNA structure probing using cell-permeable chemicals has emerged as an effective method to capture the RNA structural landscape in its native environment. The integration of these probes with advanced adduct detection techniques, particularly second- and third-generation sequencing, has propelled the field forward, facilitating a deeper understanding of the RNA structurome within its precise functional context, including the examination of RNA structure at the single-molecule and single-cell levels, within specific subcellular compartments, and across various stages of RNA biogenesis and regulation. This Review summarizes the significant advances in the field of RNA structure probing, focusing on the development of novel structural probes, strategies for RNA structure reconstruction, innovative methodologies that offer extended applicability to address unique biological questions, and concludes with an outlook on future directions in the field.
    Keywords:  RNA structure; RNA‐RNA interactions; chemical probing; nucleic acids; structure determination
    DOI:  https://doi.org/10.1002/chem.202503291
  48. Protein Sci. 2026 Feb;35(2): e70445
    Recognition of small Tim chaperones by the mitochondrial Yme1 protease.
    Mariella Quispe-Carbajal, Lauren Todd, Steven E Glynn.
      Yme1 is a conserved ATP-dependent protease that maintains mitochondrial function by degrading proteins in the intermembrane space. However, how Yme1 selects substrates within the crowded mitochondrial environment is poorly understood. An established substrate of Yme1 in yeast is the Tim10 subunit of the small Tim9-Tim10 protein chaperone complex, which is degraded following disruption of the subunit's internal disulfide bonds. Here, we use biochemical and biophysical approaches to examine initial substrate binding and degradation of small Tim proteins by Yme1 and shed light on the molecular mechanism of substrate selection. We show that Yme1 preferentially binds Tim10 over other small Tim proteins by forming a strong interaction with the subunit irrespective of the presence of its disulfide bonds. This interaction is primarily mediated by Tim10's flexible N-terminal "tentacle," though substrate unfolding exposes additional contact sites that enhance engagement. Notably, the human ortholog TIMM13 is also recognized by yeast Yme1, suggesting conservation of recognition strategy across species. Yme1 also binds to the assembled Tim9-Tim10 chaperone but independently of the Tim10 N-terminal tentacle. These findings suggest that Yme1 interacts with both the functional chaperone complex and the disassembled Tim10 monomers but only commits to degradation after disruption of its disulfide bonds.
    Keywords:  AAA+ proteases; intermembrane space; i‐AAA; mitochondrial proteostasis
    DOI:  https://doi.org/10.1002/pro.70445
  49. J Physiol Pharmacol. 2026 Jan;76(6): 681-693
    RNA-binding protein Epiplakin 1 promotes ovarian cancer immune escape and distal metastasis by enhancing the RNA stability of yes-associated protein.
    X Li, Z G Chen, F Xu, Y Q Liu.
      This study investigated the mechanism of the RNA-binding protein Epiplakin 1 (EPPK1) in ovarian cancer (OC). EPPK1 expression in OC tissues was analyzed using gene expression profiling interactive analysis (GEPIA). The association between EPPK1 expression and patient overall survival (OS) and progression-free survival (PFS) was evaluated through Kaplan-Meier analysis. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot were performed to quantify EPPK1 levels in 25 clinical OC specimens. An EPPK1-knockdown xenograft model was established to examine tumor growth and distant metastasis in vivo. The roles of EPPK1 and Yes-associated protein (YAP) in OC cell proliferation, migration, and immune escape were assessed in vitro. The EPPK1-YAP interaction was characterized. Analysis of the GEPIA database revealed elevated EPPK1 expression in ovarian serous cystadenocarcinoma, with Kaplan-Meier survival curves demonstrating significantly shorter OS and PFS in patients with high EPPK1 expression over a 5-year period. RT-qPCR and Western blot analyses confirmed EPPK1 upregulation in clinical OC specimens. Functional studies demonstrated that altered EPPK1 and YAP expression modulated OC cell proliferation, migration, invasion, and immune escape capabilities both in vitro and in vivo. Mechanistically, EPPK1 binds to and enhances YAP mRNA stability. Notably, YAP overexpression rescued the inhibitory effects of EPPK1 knockdown on OC cell proliferation, migration, invasion, and immune evasion in vitro. The results indicate that EPPK1 promotes immune evasion and distant metastasis in OC by stabilizing YAP RNA.
    Keywords:  Epiplakin 1; RNA stability; RNA-binding protein; immune escape; metastasis; oncogeny; ovarian cancer; programmed death-ligand 1; yes-associated protein
    DOI:  https://doi.org/10.26402/jpp.2025.6.06
  50. Front Plant Sci. 2025 ;16 1712714
    Mobile RNAs as systemic signaling beyond boundaries for plant stress resistance.
    Shujin Lin, Haining Li, Shiyan Bai, Xiao Han.
      Plant mobile RNAs-including small RNAs (miRNAs, siRNAs), mRNAs, lncRNAs, and tRNA fragments-function as systemic signaling molecules that traverse cellular, tissue, and species boundaries to coordinate plant adaptation to environmental stresses. Here, we summarize the critical roles of mobile RNAs in mediating systemic adaptation to abiotic challenges and biotic interactions. Crucially, we highlight the diverse transport mechanisms enabling their movement and discuss the emerging functional versatility of mobile RNAs, which extends beyond transcriptional regulation to encompass epigenetic modifications, resource allocation, and cross-species communication. These fundamental insights are driving transformative applications: Mobile RNAs provide the foundation for developing systemic RNAi-based biopesticides and are being integrated with CRISPR-Cas technologies to overcome delivery barriers and enable heritable, transgene-free genome editing in crops. Understanding and harnessing mobile RNA networks offers unprecedented potential for engineering resilient crops and implementing precise, sustainable crop protection strategies to address global food security challenges.
    Keywords:  RNA trafficking; RNAi applications; mobile RNAs; stress resistance; systemic signaling
    DOI:  https://doi.org/10.3389/fpls.2025.1712714
  51. Arch Biochem Biophys. 2026 Jan 20. pii: S0003-9861(26)00017-2. [Epub ahead of print] 110746
    RNA binding motif protein 15 promotes intracerebral hemorrhage progression by regulating mitophagy through E2F transcription factor 1.
    Jiwei Sun, Ning Li, Yesen Zhang, Jian Song, Yi Han.
      RNA binding motif protein 15 (RBM15) has been implicated in the progression of various diseases. However, its role in intracerebral hemorrhage (ICH) remains inadequately understood. To investigate RBM15's involvement in ICH, an ICH model was established using HT-22 cells and C57BL/6 mice. Elevated RBM15 expression levels were observed, suggesting its critical role in ICH pathogenesis. Our study further revealed that RBM15 depletion significantly mitigated ICH-induced brain injury and enhanced neurological function. Furthermore, RBM15 was found to mediate the N6-methyladenosine (m6A) methylation of E2F transcription factor 1 (E2F1) mRNA, thereby enhancing its stability. Further analysis confirmed that RBM15 regulates mitophagy via E2F1, with E2F1 overexpression effectively counteracting mitophagy in the absence of RBM15. Collectively, these findings indicate that targeting RBM15 may offer a promising therapeutic strategy for ICH.
    Keywords:  E2F transcription factor 1; RNA binding motif protein 15; intracerebral hemorrhage; mitophagy
    DOI:  https://doi.org/10.1016/j.abb.2026.110746
  52. Nucleic Acids Res. 2026 Jan 14. pii: gkag029. [Epub ahead of print]54(2):
    A fold switch regulates conformation of an alphavirus RNA-dependent RNA polymerase.
    Jamie J Arnold, Sean M Braet, Luiz C Vieira, Ibrahim M Moustafa, David W Gohara, Julia A Fecko, Yuan-Wei Norman Su, Abha Jain, David Aponte-Diaz, Claus O Wilke, Ganesh S Anand, Neela H Yennawar, Craig E Cameron.
      Alphaviruses are mosquito-vectored, positive-strand RNA viruses causing rheumatic and neurological diseases. Like all RNA viruses, they encode an RNA-dependent RNA polymerase (RdRp, nsP4). Purification of an nsP4 derivative capable of processive RNA synthesis from a heteropolymeric template has been unsuccessful. Prior studies indicated O'nyong-nyong virus (ONNV) nsP4 is soluble and requires additional nonstructural proteins for activity. We performed biochemical and biophysical characterization of ONNV nsP4, including analytical ultracentrifugation and small-angle X-ray scattering (SAXS), revealing an extended conformation inconsistent with AlphaFold predictions of a compact structure. Fold switching was required for the extended conformation. Hydrogen-deuterium exchange mass spectrometry confirmed the fold-switched, extended state. Phylogenetic analysis showed conservation of residues contributing to both extended and compact states, implying functional roles for each. The extended form exhibited weak RNA binding and no polymerase activity on primed templates. The SAXS envelope of a precursor containing 50 amino acids from the nsP3 C-terminus (CT50-P34) matched the compact state. We propose precursor forms adopt the compact conformation. At the replication site, proteolytic cleavage would convert the precursor to an active polymerase. Polymerase dissociation upon completion of synthesis would induce fold switching to the inactive, extended state, precluding cytoplasmic activity that would activate intracellular immune responses.
    DOI:  https://doi.org/10.1093/nar/gkag029
  53. Cancer Causes Control. 2026 Jan 21. 37(2): 35
    Contributions of selenoproteins to breast cancer etiology and racial disparity.
    Soumen Bera, Li Liu, Weiwei Ma, Ziqiao Xu, Maria Sverdlov, Ryan Deaton, Virgilia Macias, Klara Valyi-Nagy, Andre Kajdacsy-Balla, Kent Hoskins, Elizabeth L Wiley, Irida Kastrati, Alan M Diamond.
       PURPOSE: Breast cancer etiology is multifactorial with African American women experiencing a significant health disparity in clinical presentation and outcomes. The selenium-containing protein SELENOF has been implicated in breast carcinogenesis by cell culture and animal studies. SELENOF translation is highly regulated in part by the RNA helicase eIF4a3, which binds to the key regulatory regions in the SELENOF mRNA and suppress its translation. In addition, SELENOP, the primary selenium transporter, plays a critical role in selenium delivery to tissues and may influence selenoprotein synthesis. This study aimed to examine the levels of SELENOF and eIF4a3, along with SELENOF and SELENOP genotypes, in breast cancer tissues from African American and Caucasian women METHODS: To study their roles in breast cancer outcome and racial disparity, human tissues were assessed by multiplex immunofluorescence staining with antibodies directed against SELENOF and eIF4a3 and DNA from these tissues were genotyped for previously studied variations in SELENOF and the selenium transporter protein SELENOP RESULTS: Elevated levels of both SELENOF and eIF4a3 were observed in breast cancer tissues. SELENOF expression and genotype varied by HER2 status, while SELENOP genotypes were associated with breast cancer and showed age-related differences. SELENOF and eIF4a3 were also higher in tissues derived from African American women, who also exhibited higher frequency of a SELENOP polymorphism in the non-coding region of the gene CONCLUSION: These findings suggest that SELENOF, eIF4a3, and SELENOP may contribute to breast cancer progression and racial disparities in outcomes. Their differential expression and genetic variation highlight potential molecular mechanisms underlying these disparities and may inform future therapeutic or diagnostic strategies.
    Keywords:  Breast cancer; Polymorphisms; Racial disparity; Selenoproteins; Tissue microarray
    DOI:  https://doi.org/10.1007/s10552-025-02123-y
  54. Res Vet Sci. 2026 Jan 14. pii: S0034-5288(26)00015-9. [Epub ahead of print]202 106063
    Structure of the nucleoli in the spermatocytes of three canid species.
    Katarzyna Andraszek, Marta Kuchta-Gładysz, Piotr Niedbała.
      The nucleolus is a structure of the cell nucleus associated mainly with ribosome biogenesis and indirectly responsible for protein biosynthesis. The nucleolus also plays a role in the regulation of cell growth and cell cycle progression, the detection and repair of DNA damage, and the export or degradation of mRNA. It is also involved in the most critical processes for the organism, such as ageing at the cellular level and broadly defined carcinogenesis. Analysis of the structure of nucleoli, which are the products of nucleolar organizer regions, can be an alternative source of information on the activity of rRNA-encoding genes. The material for the study was testes collected post mortem from sexually mature males of three canid species: silver fox (Vulpes vulpes), blue fox (Alopex lagopus) and Chinese raccoon dog (Nyctereutes procyonoides procyonoides). The morphology and morphometry of nucleoli identified in early primary spermatocytes were analysed. The values for all morphometric parameters of the spermatocyte nucleus were lowest for the silver fox and highest for the raccoon dog. In the case of nucleolus parameters, the lowest values were found in the raccoon dog and the lowest in the blue fox. The size of the nucleoli and spermatocyte nucleus proved to be a species-specific trait associated with the karyotype and number of nucleolar organizer regions in the species. The nucleolus is believed to play an important role in controlling meiosis. It is involved in preventing chromosome segregation during crossing over, and proteins associated with the nucleoli identify damaged synaptonemal complexes. Epigenetic regulations at the level of the nucleolus and rRNA genes remain a crucial subject of genetic research.
    Keywords:  Blue fox; Chinese raccoon dog; Nucleolus; Silver fox; Spermatogenesis
    DOI:  https://doi.org/10.1016/j.rvsc.2026.106063
  55. Nucleic Acids Res. 2026 Jan 22. pii: gkaf1524. [Epub ahead of print]54(3):
    Structural plasticity enables broad cAn binding and dual activation of CRISPR-associated ribonuclease Cdn1.
    Wenxuan Zhang, Jianping Kong, Yuqin Zeng, Yunning Su, Sijun Zhang, Yutao Li, Chunyi Hu, Qihua Chen, Yibei Xiao, Meiling Lu.
      Prokaryotes have naturally evolved diverse RNA-guided defense systems against viral infections, with the type III CRISPR-Cas systems representing the most intricate. These systems feature accessory proteins activated by cyclic oligoadenylates (cOAs) produced upon target RNA recognition, synergizing with the CRISPR-Cas machinery to defend against exogenous invaders. Typically, each accessory protein is activated by only one specific cOA type. Here, we characterize Cdn1, a type III-B CRISPR accessory protein from Psychrobacter lutiphocae, which binds to cA3, cA4, and cA6, but activated by cA4 and cA6 with different efficacies to catalyze ssRNA cleavage. Combined structural and biochemical analyses reveal that cOA binding triggers dramatic conformational reorganization, including the formation of a dimerization interface of nuclease domains, the emergence of substrate binding cleft, and the reconstruction of a metal-dependent catalytic center essential for RNA cleavage. This dual activation mechanism illustrates evolutionary innovation within CRISPR-associated Rossman-fold nucleases. We propose that such structural plasticity evolved to maximize defensive resilience during microbial competition and horizontal gene transfer, while preserving broad-spectrum antiviral ability. These findings not only elucidate the activation mechanisms of Cdn1 within the type III systems but also underscore the functional complexity and adaptability of CRISPR-Cas ancillary proteins.
    DOI:  https://doi.org/10.1093/nar/gkaf1524
  56. Diabetes. 2026 Jan 21. pii: db250424. [Epub ahead of print]
    Optimizing Single-Cell Long-Read Sequencing for Enhanced Isoform Detection in Pancreatic Islets.
    Maria S Hansen, Christopher J Hill, Lori Sussel, Kristen L Wells.
      Alternative splicing is an essential mechanism for generating protein diversity by producing distinct isoforms from a single gene. Dysregulation of splicing that affects pancreatic function and immune tolerance has been linked to both types 1 and 2 diabetes. Next-generation sequencing technologies, with their short read lengths, are limited in their ability to accurately detect splice variants. Long-read sequencing technologies offer the potential to overcome these limitations by providing full-length transcript information; however, their application in single-cell RNA sequencing has been hindered by technical challenges, including insufficient read lengths and higher error rates. Furthermore, cell types that produce high levels of a single transcript, such as islet endocrine cells, can obscure identification of lower-abundance transcripts. In this study, we optimized a protocol for single-cell long-read sequencing in pancreatic islets to improve read length and transcript detection. Our findings demonstrate that 5' library preparation protocols outperform 3' protocols, resulting in better transcript identification. Furthermore, we show that targeted depletion of insulin transcripts enhances the detection of informative reads, highlighting the utility of transcript-depletion strategies. This optimized protocol enables isoform-specific gene expression analysis and reveals differential transcript usage across the various cell types in pancreatic islets. By leveraging this approach, we gain deeper insights into the transcriptomic complexity and cellular heterogeneity within pancreatic islets.
    ARTICLE HIGHLIGHTS: This study addresses the limitations of current single-cell long-read RNA sequencing technologies in detecting full-length transcripts and isoform diversity, particularly in pancreatic islets. We demonstrate that optimizing single-cell library preparation protocols reproducibly enhances read length and transcript identification in pancreatic islets. Combined with targeted insulin depletion and extended reverse transcription, 5' capture methods significantly improved read length and isoform detection compared with standard protocols, while maximizing the number of informative reads. These improvements yield longer reads in single-cell experiments, substantially enhancing transcript identification and enabling more accurate analysis of isoform diversity.
    DOI:  https://doi.org/10.2337/db25-0424
  57. bioRxiv. 2025 Dec 06. pii: 2025.12.05.692659. [Epub ahead of print]
    PIWIL4 regulates gene expression and piRNA levels in RSV-infected airway epithelial cells.
    Tiziana Corsello, Tianshuang Liu, Andrzej S Kudlicki, Yuanyi Zhang, Nicholas Dillman, Sehar Fazal, Roberto P Garofalo, Antonella Casola.
      Respiratory syncytial virus (RSV) is a leading cause of acute lower respiratory tract infections with significant morbidity and mortality in young children, the elderly and immunocompromised hosts. Despite its clinical burden, no effective RSV vaccine or therapy exists for infants, only prophylactic treatment. Small non-coding RNAs have emerged as important regulators of host-pathogen interactions. PIWI-interacting RNAs (piRNAs) are a distinct class of small non-coding RNAs known for maintaining the genome complexity and integrity in gonadal cells. However, there is growing evidence of their role in controlling gene expression in somatic cells. The biogenesis and function of piRNAs is associated with P-element Induced Wimpy testis (Piwi) proteins, whose function in the respiratory epithelium in response to infections remains largely unexplored. Here, we characterize the expression and function of the Piwi-like protein PIWIL4 in the context of RSV infection. We found that PIWIL4 is expressed in both primary and immortalized small airway epithelial cells and is significantly induced at the mRNA and protein levels following RSV infection or poly I:C stimulation, a proxy of viral infection. Immunofluorescence microscopy revealed that PIWIL4 was primarily nuclear in uninfected cells but translocated to the cytoplasm upon RSV exposure. While siRNA-mediated knockdown of PIWIL4 did not significantly affect RSV replication, it led to decreased secretion of several cytokines, chemokines and growth factors, indicating a role in modulating host innate immune responses. Transcriptomic analysis of PIWIL4-silenced iSAE cells showed significant changes in gene expression both in basal conditions and upon RSV infection. Ingenuity Pathway analysis of differentially expressed genes underscored the role of PIWIL4 in modulation of interferon signaling, cytokine production, stress and metabolic responses, as well as airway remodeling pathways. Silencing of PIWIL4 also resulted in global alteration of piRNA expression both in uninfected and infected cells. However, the predicted targets of the differentially expressed piRNAs had limited overlap with the differentially expressed genes identified by transcriptomics, suggesting a function of PIWIL4 in regulating airway epithelial cell responses at least in part independent of piRNAs. Taken together, our study uncovers an important role for PIWIL4 in somatic cells and position it as a key regulator of airway epithelial innate immunity. A better understanding of the mechanisms by which PIWIL4 affects host cells responses following a pathogen exposure may identify novel therapeutic strategies for RSV, as well as other viral respiratory infections.
    DOI:  https://doi.org/10.64898/2025.12.05.692659
  58. Phys Rev E. 2025 Dec;112(6-1): 064403
    Tuning nuclear rheology through transient chromatin cross-links.
    Yin-Dong Zhang, Chao-Hao Wu, Han-Xuan Shi, Holger Merlitz, Kerry S Bloom, M Gregory Forest, Chen-Xu Wu, Xue-Zheng Cao.
      In eukaryotic cells, the nucleolus is a pivotal subnuclear organelle, instrumental in ribosomal RNA synthesis and nuclear organization. Although the unique viscoelastic properties of the nucleolus are associated with transient interactions between chromatin and regulatory proteins, the specific mechanistic details driving nucleolar phase separation and mechanical responses have remained largely undefined. In this study, we employ a computational approach to elucidate chromatin-protein interactions within the nucleolus of budding yeast, using a sophisticated bead-spring polymer model. This model integrates DNA and nucleolar architectures with dynamic simulations of interactions involving chromosomal structural maintenance proteins and rDNA transcriptional regulators through systematically varied cross-linking kinetics. Our findings reveal that modulations in protein-DNA interactions critically dictate the phase behavior, relaxation dynamics, and viscoelastic properties of the nucleolus, underscoring a complex but precise regulatory mechanism at play. Notably, protein-mediated bridging emerges as a critical factor enhancing nucleolar condensation and modulating stress relaxation, highlighting the transformative role of transient cross-linking in nuclear mechanics regulation. These insights not only deepen our understanding of nucleolar function but also open avenues for interventions in genetic engineering and disease therapeutics.
    DOI:  https://doi.org/10.1103/tn2w-kzb8
  59. J Inorg Biochem. 2026 Jan 03. pii: S0162-0134(26)00002-4. [Epub ahead of print]277 113213
    Persulfidation of the zinc finger protein ZRANB2 modulates its RNA binding and alternative splicing function.
    Matthew S Hursey, Abigail D Reitz, Samuel E Fidler, Sarah L J Michel.
      Zinc finger Ran-binding domain-containing protein 2 (ZRANB2) is an RNA-binding protein that plays a key role in alternative splicing. It contains two N-terminal RanBP2-type ZF domains in which four cysteine residues coordinate Zn(II) in a tetrahedral geometry to afford proper folding and function. Persulfidation, a post-translational modification in which cysteine thiols (-SH) are converted to persulfides (-SSH) by hydrogen sulfide (H2S), has emerged as a means for regulating ZF activity. ZRANB2 is frequently identified as persulfidated in chemoselective proteomics screens, and here, we evaluate the direct modification of ZRANB2 by H2S. Using a recombinantly expressed two-domain construct (ZRANB2-2D), we report that Zn(II)-bound ZRANB2-2D undergoes persulfidation when exposed to H2S and oxygen with superoxide generated as an intermediate. This modification induces a loss of Zn(II)-dependent structure and abrogates binding to an RNA oligonucleotide from exon 3 of the transformer-2 protein homolog beta (TRA2B) RNA, a splicing target of ZRANB2, as well as to an optimized RNA oligonucleotide. Consistent with impaired RNA binding, cellular treatment with H2S leads to decreased formation of a TRA2B splice product, suggesting a connection to persulfidation of ZRANB2 in cells. Notably, addition of a reductant restores ZRANB2-2D RNA-binding activity in vitro. These results position persulfidation as a rheostat for modulating ZF protein function, exemplified here by its role in regulating ZRANB2 RNA binding and splicing.
    Keywords:  Alternative splicing; Hydrogen Sulfide; Persulfidation; RNA binding; ZRANB2; Zinc finger
    DOI:  https://doi.org/10.1016/j.jinorgbio.2026.113213
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