CoChat
Sign Up
Cited by 7Enzyme Structure and FunctionRNA Research and SplicingRNA and protein synthesis mechanisms

The DOI matches its Crossref record; 2 of the 2 references it cites were found in a registry or an index.

Molecular basis for the calcium-dependent activation of the ribonuclease EndoU

Open access
Malard, Florian, Dias, Kristen, Baudy, Margaux, Thore, Stéphane, Vialet, Brune, Barthélémy, Philippe, Fribourg, Sébastien, Karginov, Fedor V., Campagne, Sébastien·2025·Nature Communications
DOI 10.1038/s41467-025-58462-6PMC PMC11961692

Published version of a preprint

Open this paper in CoChat — free account

Abstract

Abstract Ribonucleases (RNases) are ubiquitous enzymes that process or degrade RNA, essential for cellular functions and immune responses. The EndoU-like superfamily includes endoribonucleases conserved across bacteria, eukaryotes, and certain viruses, with an ancient evolutionary link to the ribonuclease A-like superfamily. Both bacterial EndoU and animal RNase A share a similar fold and function independently of cofactors. In contrast, the eukaryotic EndoU catalytic domain requires divalent metal ions for catalysis, possibly due to an N-terminal extension near the catalytic core. In this study, we used biophysical and computational techniques along with in vitro assays to investigate the calcium-dependent activation of human EndoU. We determined the crystal structure of EndoU bound to calcium and found that calcium binding remote from the catalytic triad triggers water-mediated intramolecular signaling and structural changes, activating the enzyme through allostery. Calcium-binding involves residues from both the catalytic core and the N-terminal extension, indicating that the N-terminal extension interacts with the catalytic core to modulate activity in response to calcium. Our findings suggest that similar mechanisms may be present across all eukaryotic EndoUs, highlighting a unique evolutionary adaptation that connects endoribonuclease activity to cellular signaling in eukaryotes.

Overview

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Integrity signals

Registry

Crossref · 10.1038/s41467-025-58462-6

Cites

2 of 2 references verified0 citation errors · 0 not checkable

Statistics

Verification

Claims the paper makes

The claims and statistics of this paper haven’t been checked yet.

Run the check — free, no account needed. It reads the open-access text and checks the sources it cites.

A few free checks a day · papers already verified open instantly · results are public

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

The self-check needs the paper’s full text.

Citations

Cites

Flagged first, then by citations

Research works printed but not linked: 67

  1. Kushner, S. R. mRNA Decay in Prokaryotes and Eukaryotes: Different Approaches to a Similar Problem. IUBMB Life 56, 585–594 (2004). · DOI
  2. Evguenieva‐Hackenberg, E. & Klug, G. RNA Degradation in Archaea and Gram‐Negative Bacteria Different from Escherichia coli. Prog. Mol. Biol. Transl. Sci. 85, 275–317 (2009). · DOI
  3. Schmid, M. & Jensen, T. H. The exosome: a multipurpose RNA-decay machine. Trends Biochem. Sci. 33, 501–510 (2008). · DOI
  4. Tomecki, R. & Dziembowski, A. Novel endoribonucleases as central players in various pathways of eukaryotic RNA metabolism. RNA 16, 1692–1724 (2010). · DOI
  5. Paysan-Lafosse, T. et al. InterPro in 2022. Nucleic Acids Res. 51, D418–D427 (2023). · DOI
  6. Mushegian, A., Sorokina, I., Eroshkin, A. & Dlakić, M. An ancient evolutionary connection between Ribonuclease A and EndoU families. RNA 26, 803–813 (2020). · DOI
  7. Raines, R. T. Ribonuclease A. Chem. Rev. 98, 1045–1066 (1998). · DOI
  8. Smyth, D. G., Stein, W. H. & Moore, S. The sequence of amino acid residues in bovine pancreatic ribonuclease: revisions and confirmations. J. Biol. Chem. 238, 227–34 (1963). · DOI
  9. Findlay, D., Herries, D., Mathias, A., Rabin, B. & Ross, C. The active site and mechanism of action of bovine pancreatic ribonuclease. 7. The catalytic mechanism. Biochemical J. 85, 152–153 (1962). · DOI
  10. AVEY, H. P. et al. Structure of Ribonuclease. Nature 213, 557–562 (1967). · DOI
  11. Michalska, K. et al. Functional plasticity of antibacterial EndoU toxins. Mol. Microbiol. 109, 509–527 (2018). · DOI
  12. Caffarelli, E., Maggi, L., Fatica, A., Jiricny, J. & Bozzoni, I. A Novel Mn++-Dependent Ribonuclease That Functions in U16 SnoRNA Processing inX. Laevis. Biochem. Biophys. Res. Commun. 233, 514–517 (1997). · DOI
  13. Gioia, U. et al. Functional Characterization of XendoU, the Endoribonuclease Involved in Small Nucleolar RNA Biosynthesis. J. Biol. Chem. 280, 18996–19002 (2005). · DOI
  14. Laneve, P. et al. Purification, Cloning, and Characterization of XendoU, a Novel Endoribonuclease Involved in Processing of Intron-encoded Small Nucleolar RNAs in Xenopus laevis. J. Biol. Chem. 278, 13026–13032 (2003). · DOI
  15. Mistry, J. et al. Pfam: The protein families database in 2021. Nucleic Acids Res. 49, D412–D419 (2021). · DOI · Paper page
  16. Bohn, H. & Winckler, W. Isolierung und Charakterisierung eines neuen Plazenta-Gewebsproteins (PP11). Arch. Gynecol. 229, 293–301 (1980). · DOI
  17. Laneve, P. et al. The Tumor Marker Human Placental Protein 11 Is an Endoribonuclease. J. Biol. Chem. 283, 34712–34719 (2008). · DOI
  18. Xu, C. et al. Integrated Analysis Reveals ENDOU as a Biomarker in Head and Neck Squamous Cell Carcinoma Progression. Front. Oncol. 10, 522332 (2021). · DOI
  19. Basic, V., Zhang, B., Domert, J., Pellas, U. & Tot, T. Integrative meta-analysis of gene expression profiles identifies FEN1 and ENDOU as potential diagnostic biomarkers for cervical squamous cell carcinoma. Oncol. Lett. 22, 840 (2021). · DOI
  20. Lou, Y., Ren, L. & Wang, L. Hsa_circ_0049396 inhibited oral squamous cell carcinoma progression by regulating the miR-663b/ENDOU axis. Genomics 115, 110553 (2023). · DOI
  21. Inaba, N. et al. Immunohistochemical detection of pregnancy-specific protein (SP1) and placenta-specific tissue proteins (PP5, PP10, PP11 and PP12) in ovarian adenocarcinomas. Oncodev. Biol. Med. 3, 379–89 (1982).
  22. Inaba, N., Renk, T., Wurster, K., Rapp, W. & Bohn, H. Ectopic synthesis of pregnancy specific beta 1-glycoprotein (SP1) and placental specific tissue proteins (PP5, PP10, PP11, PP12) in nontrophoblastic malignant tumours possible markers in oncology. Klin. Wochenschr. 58, 789–791 (1980). · DOI
  23. Schwarz, D. S. & Blower, M. D. The calcium-dependent ribonuclease XendoU promotes ER network formation through local RNA degradation. J. Cell Biol. 207, 41–57 (2014). · DOI
  24. Zhao, G. & Blackstone, C. ER Morphology: Sculpting with XendoU. Curr. Biol. 24, R1170–R1172 (2014). · DOI
  25. Poe, J. C. et al. EndoU is a novel regulator of AICD during peripheral B cell selection. J. Exp. Med. 211, 57–69 (2014). · DOI
  26. Laneve, P. et al. Drosophila CG3303 is an essential endoribonuclease linked to TDP-43-mediated neurodegeneration. Sci. Rep. 7, 41559 (2017). · DOI
  27. Ujisawa, T. et al. Endoribonuclease ENDU-2 regulates multiple traits including cold tolerance via cell autonomous and nonautonomous controls in Caenorhabditis elegans. Proc. Natl Acad. Sci. 115, 8823–8828 (2018). · DOI
  28. Zhang, D., de Souza, R. F., Anantharaman, V., Iyer, L. M. & Aravind, L. Polymorphic toxin systems: Comprehensive characterization of trafficking modes, processing, mechanisms of action, immunity and ecology using comparative genomics. Biol. Direct 7, 18 (2012). · DOI
  29. Nedialkova, D. D. et al. Biochemical Characterization of Arterivirus Nonstructural Protein 11 Reveals the Nidovirus-Wide Conservation of a Replicative Endoribonuclease. J. Virol. 83, 5671–5682 (2009). · DOI
  30. Bhardwaj, K., Guarino, L. & Kao, C. C. The Severe Acute Respiratory Syndrome Coronavirus Nsp15 Protein Is an Endoribonuclease That Prefers Manganese as a Cofactor. J. Virol. 78, 12218–12224 (2004). · DOI
  31. Renzi, F. et al. The structure of the endoribonuclease XendoU: From small nucleolar RNA processing to severe acute respiratory syndrome coronavirus replication. Proc. Natl Acad. Sci. 103, 12365–12370 (2006). · DOI
  32. Heng, T. S. P. et al. The Immunological Genome Project: networks of gene expression in immune cells. Nat. Immunol. 9, 1091–1094 (2008). · DOI · Paper page
  33. Uhlén, M. et al. Tissue-based map of the human proteome. Science 347, 1260419 (2015). · DOI
  34. Groves, T. et al. In vitro maturation of clonal CD4+CD8+ cell lines in response to TCR engagement. J. Immunol. 154, 5011–22 (1995). · DOI
  35. Waterhouse, A. et al. SWISS-MODEL: homology modelling of protein structures and complexes. Nucleic Acids Res. 46, W296–W303 (2018). · DOI
  36. Svergun, D., Barberato, C. & Koch, M. H. J. CRYSOL – a Program to Evaluate X-ray Solution Scattering of Biological Macromolecules from Atomic Coordinates. J. Appl. Crystallogr. 28, 768–773 (1995). · DOI
  37. Malard, F., Karginov, F. V. & Campagne, S. 1H, 13C and 15N backbone resonance assignment of the calcium-activated EndoU endoribonuclease. Biomol. NMR Assign. 18, 263–267 (2024). · DOI
  38. Abramson, J. et al. Addendum: Accurate structure prediction of biomolecular interactions with AlphaFold 3. Nature 636, E4–E4 (2024). · DOI
  39. Yang, W., Lee, J. Y. & Nowotny, M. Making and Breaking Nucleic Acids: Two-Mg2+-Ion Catalysis and Substrate Specificity. Mol. Cell 22, 5–13 (2006). · DOI · Paper page
  40. Cuchillo, C. M. et al. The role of 2’, 3’‐cyclic phosphodiesters in the bovine pancreatic ribonuclease A catalysed cleavage of RNA: intermediates or products? FEBS Lett. 333, 207–210 (1993). · DOI
  41. Qi, W. et al. The secreted endoribonuclease ENDU-2 from the soma protects germline immortality in C. elegans. Nat. Commun. 12, 1262 (2021). · DOI
  42. Keen, C. L., Uriu-Adams, J. Y., Ensunsa, J. L. & Gershwin, M. E. Trace Elements/Minerals and Immunity. In: Handbook of Nutrition and Immunity 117–140 (Humana Press, 2004) https://doi.org/10.1007/978-1-59259-790-1_6. · DOI
  43. Jenne, D. & Stanley, K. K. Nucleotide sequence and organization of the human S-protein gene: repeating peptide motifs in the ‘pexin’ family and a model for their evolution. Biochemistry 26, 6735–6742 (1987). · DOI
  44. Alquraini, A. et al. The interaction of lubricin/proteoglycan 4 (PRG4) with toll-like receptors 2 and 4: an anti-inflammatory role of PRG4 in synovial fluid. Arthritis Res. Ther. 17, 353 (2015). · DOI
  45. Goding, J. W., Grobben, B. & Slegers, H. Physiological and pathophysiological functions of the ecto-nucleotide pyrophosphatase/phosphodiesterase family. Biochim. Biophys. Acta Mol. Basis Dis. 1638, 1–19 (2003). · DOI
  46. Droujinine, I. A. et al. Proteomics of protein trafficking by in vivo tissue-specific labeling. Nat. Commun. 12, 2382 (2021). · DOI
  47. Bootman, M. D. & Bultynck, G. Fundamentals of Cellular Calcium Signaling: A Primer. Cold Spring Harb. Perspect. Biol. 12, a038802 (2020). · DOI
  48. Pillon, M. C. et al. Cryo-EM structures of the SARS-CoV-2 endoribonuclease Nsp15 reveal insight into nuclease specificity and dynamics. Nat. Commun. 12, 636 (2021). · DOI
  49. Huang, T. et al. Kinetic analysis of RNA cleavage by coronavirus Nsp15 endonuclease: Evidence for acid–base catalysis and substrate-dependent metal ion activation. J. Biol. Chem. 299, 104787 (2023). · DOI
  50. Wang, X. & Zhu, B. SARS-CoV-2 nsp15 preferentially degrades AU-rich dsRNA via its dsRNA nickase activity. Nucleic Acids Res. 52, 5257–5272 (2024). · DOI
  51. Bhardwaj, K., Sun, J., Holzenburg, A., Guarino, L. A. & Kao, C. C. RNA Recognition and Cleavage by the SARS Coronavirus Endoribonuclease. J. Mol. Biol. 361, 243–256 (2006). · DOI
  52. Frazier, M. N. et al. Characterization of SARS2 Nsp15 nuclease activity reveals it’s mad about U. Nucleic Acids Res. 49, 10136–10149 (2021). · DOI
  53. Kim, Y. et al. Tipiracil binds to uridine site and inhibits Nsp15 endoribonuclease NendoU from SARS-CoV-2. Commun. Biol. 4, 193 (2021). · DOI
  54. Ivanov, K. A. et al. Major genetic marker of nidoviruses encodes a replicative endoribonuclease. Proc. Natl Acad. Sci. 101, 12694–12699 (2004). · DOI
  55. Jamet, A. et al. A New Family of Secreted Toxins in Pathogenic Neisseria Species. PLoS Pathog. 11, e1004592 (2015). · DOI
  56. Sternburg, E. L., Dias, K. C. & Karginov, F. V. Selection-dependent and Independent Generation of CRISPR/Cas9-mediated Gene Knockouts in Mammalian Cells. J. Vis. Exper. https://doi.org/10.3791/55903 (2017). · DOI
  57. Liu, H. & Naismith, J. H. An efficient one-step site-directed deletion, insertion, single and multiple-site plasmid mutagenesis protocol. BMC Biotechnol. 8, 91 (2008). · DOI
  58. Kabsch, W. XDS. Acta Crystallogr D. Biol. Crystallogr 66, 125–132 (2010). · DOI · Paper page
  59. Liebschner, D. et al. Macromolecular structure determination using X-rays, neutrons and electrons: recent developments in Phenix. Acta Crystallogr D. Struct. Biol. 75, 861–877 (2019). · DOI · Paper page
  60. Müller, P., Köpke, S. & Sheldrick, G. M. Is the bond-valence method able to identify metal atoms in protein structures? Acta Crystallogr D. Biol. Crystallogr 59, 32–37 (2003). · DOI
  61. Zheng, H. et al. CheckMyMetal: a macromolecular metal-binding validation tool. Acta Crystallogr D. Struct. Biol. 73, 223–233 (2017). · DOI
  62. Van Der Spoel, D. et al. GROMACS: Fast, flexible, and free. J. Comput Chem. 26, 1701–1718 (2005). · DOI
  63. Lee, J. et al. CHARMM-GUI Input Generator for NAMD, GROMACS, AMBER, OpenMM, and CHARMM/OpenMM Simulations Using the CHARMM36 Additive Force Field. J. Chem. Theory Comput 12, 405–413 (2016). · DOI
  64. Thureau, A., Roblin, P. & Pérez, J. BioSAXS on the SWING beamline at Synchrotron SOLEIL. J. Appl. Crystallogr. 54, 1698–1710 (2021). · DOI
  65. David, G. & Pérez, J. Combined sampler robot and high-performance liquid chromatography: a fully automated system for biological small-angle X-ray scattering experiments at the Synchrotron SOLEIL SWING beamline. J. Appl. Crystallogr. 42, 892–900 (2009). · DOI
  66. Hopkins, J. B., Gillilan, R. E. & Skou, S. BioXTAS RAW: improvements to a free open-source program for small-angle X-ray scattering data reduction and analysis. J. Appl. Crystallogr. 50, 1545–1553 (2017). · DOI
  67. Franke, D. et al. ATSAS 2.8: a comprehensive data analysis suite for small-angle scattering from macromolecular solutions. J. Appl. Crystallogr. 50, 1212–1225 (2017). · DOI

Metadata

Authors
Malard, F., Dias, K., Baudy, M., Thore, S., Vialet, B., Barthélémy, P., Fribourg, S., Karginov, F. V., & Campagne, S.
Affiliations
  • Centre National de la Recherche Scientifique (FR)
  • University of California, Riverside (US)
Corresponding author
  • Fribourg, Sébastien
  • Karginov, Fedor V.
  • Campagne, Sébastien
ORCID
Venue
Nature Communications
Publisher
Springer Science and Business Media LLC
Published
2025-04-01
Volume
16
Issue
1
Pages
3110-3110
ISSN
2041-1723 (ISSN-L)
DOI
10.1038/s41467-025-58462-6
PMC
PMC11961692
OpenAlex
W4409050148
Cited by
7
References
2
Citations by year
2026: 5 · 2025: 1
FWCI
1.27
Type
journal article
Language
en
Open access
Licence
Locations
  • Nature Communications (publishedVersion, cc-by-nc-nd)
  • Research Square (acceptedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by-nc-nd)
  • PubMed Central (publishedVersion, cc-by-nc-nd)
  • PubMed (publishedVersion)
  • eScholarship (California Digital Library) (submittedVersion, cc-by-nc-nd)
  • PubMed Central (submittedVersion, cc-by)
  • HAL (submittedVersion)
  • RePEc: Research Papers in Economics (submittedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
Funders
  • Institut National de la Santé et de la Recherche Médicale
  • Centre National de la Recherche Scientifique
  • National Institutes of Health (1r01gm152548-01a1)
  • National Institute of General Medical Sciences (R01 GM152548)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Structural Biology › Enzyme Structure and Function
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Research and Splicing
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
Keywords
endoribonuclease, calcium-dependent activation, allosteric activation, N-terminal extension, crystal structure, RNA cleavage, metal ion catalysis

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

What counts as a claim ›How we measure confidence ›How we check this ›

Are you an author, or do you see a problem with this page? Request a review