Structural insights into Crimean-Congo haemorrhagic fever virus RNA polymerase reveal a dual-site non-nucleoside inhibition mechanism
Abstract
Crimean-Congo hemorrhagic fever virus (CCHFV) is a priority pathogen defined by its wide geographic distribution and high case-fatality rates, yet no approved vaccines or therapeutics exist. Its massive ~450 kDa RNA-dependent RNA polymerase (L protein)—the multi-domain and multi-functional machinery for viral transcription and replication—has remained structurally enigmatic due to its exceptional size and complexity. Here we present cryo-EM structures of the CCHFV L protein in apo, promoter-bound, and inhibitor-bound states. Structural analysis reveals that the L protein adopts the canonical architecture shared across the order Bunyavirales, with high structural conservation extending to individual subdomains and the 'hook-like' recognition of the vRNA promoter. Unexpectedly, we found that the non-nucleotide inhibitor suramin inhibits the polymerase through a unique dual-site mechanism. It not only competitively occludes the 5' vRNA-binding pocket but also functions as a “molecular glue” at the distal Linker–Fingers interface, likely allosterically restricting essential conformational dynamics. These findings provide a detailed structural framework for CCHFV replication and offer a novel paradigm for designing antivirals against Bunyavirales.
References
McFadden, E. et al. Engineering and structures of Crimean-Congo hemorrhagic fever virus glycoprotein complexes. Cell 188, 303-315 e313(2025). https://doi.org/10.1016/j.cell.2024.11.008
Ritter, M. et al. The low-density lipoprotein receptor and apolipoprotein E associated with CCHFV particles mediate CCHFV entry into cells. Nat Commun 15,4542(2024). https://doi.org/10.1038/s41467-024-48989-5
Monteil, V. M. et al. Crimean-Congo haemorrhagic fever virus uses LDLR to bind and enter host cells. Nat Microbiol9, 1499-1512(2024). https://doi.org/10.1038/s41564-024-01672-3
Golden, J. W. et al. GP38-targeting monoclonal antibodies protect adult mice against lethal Crimean-Congo hemorrhagic fever virus infection. Sci Adv 5, eaaw9535(2019). https://doi.org/10.1126/sciadv.aaw9535
Hawman, D. W. & Feldmann, H. Crimean-Congo haemorrhagic fever virus. Nat Rev Microbiol 21,463-477(2023). https://doi.org/10.1038/s41579-023-00871-9
Frank, M. G. et al. Crimean-Congo Hemorrhagic Fever Virus for Clinicians-Diagnosis, Clinical Management, and Therapeutics. Emerg Infect Dis 30, 864-873(2024). https://doi.org/10.3201/eid3005.231648
Bente, D. A. et al. Crimean-Congo hemorrhagic fever: history, epidemiology, pathogenesis, clinical syndrome and genetic diversity. Antiviral Res 100,159-189(2013). https://doi.org/10.1016/j.antiviral.2013.07.006
WHO. Crimean-Congo haemorrhagic fever. World Health Organization, Geneva, Switzerland, 2018.
Ma, J. et al. Identification of a new orthonairovirus associated with human febrile illness in China. Nat Med 27,434-439(2021). https://doi.org/10.1038/s41591-020-01228-y
Zhang, X. A. et al. A New Orthonairovirus Associated with Human Febrile Illness. N Engl J Med 391,821-831(2024). https://doi.org/10.1056/NEJMoa2313722
Zhang, M. Z. et al. Human Infection with a Novel Tickborne Orthonairovirus Species in China. N Engl J Med 392,200-202(2025). https://doi.org/10.1056/NEJMc2410853
Iglesias-Rivas, P., Gonzalez-Vazquez, L. D. & Arenas, M. Molecular Evolution and Phylogeography of the Crimean-Congo Hemorrhagic Fever Virus. Viruses 17(2025). https://doi.org/10.3390/v17081054
Zivcec, M., Scholte, F. E., Spiropoulou, C. F., Spengler, J. R. & Bergeron, E. Molecular Insights into Crimean-Congo Hemorrhagic Fever Virus. Viruses 8, 106(2016). https://doi.org/10.3390/v8040106
Kuang, W. et al. Structure and function of the nairovirus cap-snatching endonuclease. Nucleic Acids Res 54(2026). https://doi.org/10.1093/nar/gkaf1515
Leventhal, S. S. et al. Antibodies targeting the Crimean-Congo Hemorrhagic Fever Virus nucleoprotein protect via TRIM21. Nat Commun 15, 9236(2024). https://doi.org/10.1038/s41467-024-53362-7
Akutsu, M., Ye, Y., Virdee, S., Chin, J. W. & Komander, D. Molecular basis for ubiquitin and ISG15 cross-reactivity in viral ovarian tumor domains. Proc Natl Acad Sci U S A 108,2228-2233(2011). https://doi.org/10.1073/pnas.1015287108
Spengler, J. R. et al. RIG-I Mediates an Antiviral Response to Crimean-Congo Hemorrhagic Fever Virus. J Virol 89,10219-10229(2015). https://doi.org/10.1128/JVI.01643-15
Scholte, F. E. M. et al. Crimean-Congo Hemorrhagic Fever Virus Suppresses Innate Immune Responses via a Ubiquitin and ISG15 Specific Protease. Cell Rep 20, 2396-2407(2017). https://doi.org/10.1016/j.celrep.2017.08.040
Peng, R. et al. Structural insight into arenavirus replication machinery. Nature 579,615-619(2020). https://doi.org/10.1038/s41586-020-2114-2
Xu, X. et al. Cryo-EM structures of Lassa and Machupo virus polymerases complexed with cognate regulatory Z proteins identify targets for antivirals. Nat Microbiol 6,921-931(2021). https://doi.org/10.1038/s41564-021-00916-w
Ma, J., Zhang, S. & Zhang, X. Structure of Machupo virus polymerase in complex with matrix protein Z. Nat Commun 12,6163(2021). https://doi.org/10.1038/s41467-021-26432-3
Kouba, T. et al. Conformational changes in Lassa virus L protein associated with promoter binding and RNA synthesis activity. Nat Commun 12, 7018(2021). https://doi.org/10.1038/s41467-021-27305-5
Durieux Trouilleton, Q., Barata-Garcia, S., Arragain, B., Reguera, J. & Malet, H. Structures of active Hantaan virus polymerase uncover the mechanisms of Hantaviridae genome replication. Nat Commun 14, 2954(2023). https://doi.org/10.1038/s41467-023-38555-w
Durieux Trouilleton, Q., Housset, D., Tarillon, P., Arragain, B. & Malet, H. Structural characterization of the oligomerization of full-length Hantaan virus polymerase into symmetric dimers and hexamers. Nat Commun 15, 2256(2024). https://doi.org/10.1038/s41467-024-46601-4
Keown, J. R., Carrique, L., Nilsson-Payant, B. E., Fodor, E. & Grimes, J. M. Structural characterization of the full-length Hantaan virus polymerase. PLoS Pathog 20, e1012781(2024). https://doi.org/10.1371/journal.ppat.1012781
Gerlach, P., Malet, H., Cusack, S. & Reguera, J. Structural Insights into Bunyavirus Replication and Its Regulation by the vRNA Promoter. Cell 161, 1267-1279(2015). https://doi.org/10.1016/j.cell.2015.05.006
Arragain, B. et al. Pre-initiation and elongation structures of full-length La Crosse virus polymerase reveal functionally important conformational changes. Nat Commun 11,3590(2020). https://doi.org/10.1038/s41467-020-17349-4
Arragain, B. et al. Structural snapshots of La Crosse virus polymerase reveal the mechanisms underlying Peribunyaviridae replication and transcription. Nat Commun 13,902(2022). https://doi.org/10.1038/s41467-022-28428-z
Wang, P. et al. Structure of severe fever with thrombocytopenia syndrome virus L protein elucidates the mechanisms of viral transcription initiation. Nat Microbiol 5,864-871(2020). https://doi.org/10.1038/s41564-020-0712-2
Vogel, D. et al. Structural and functional characterization of the severe fever with thrombocytopenia syndrome virus L protein. Nucleic Acids Res 48, 5749-5765(2020). https://doi.org/10.1093/nar/gkaa253
Williams, H. M. et al. Structural insights into viral genome replication by the severe fever with thrombocytopenia syndrome virus L protein. Nucleic Acids Res 51, 1424-1442(2023). https://doi.org/10.1093/nar/gkac1249
Wang, X. et al. Structure of Rift Valley Fever Virus RNA-Dependent RNA Polymerase. J Virol 96,e0171321(2022). https://doi.org/10.1128/JVI.01713-21
Li, J. et al. Structural basis for the activation of plant bunyavirus replication machinery and its dual-targeted inhibition by ribavirin. Nat Plants 11, 518-530(2025). https://doi.org/10.1038/s41477-025-01940-y
Oestereich, L. et al. Evaluation of antiviral efficacy of ribavirin, arbidol, and T-705(favipiravir) in a mouse model for Crimean-Congo hemorrhagic fever. PLoS Negl Trop Dis 8,e2804(2014). https://doi.org/10.1371/journal.pntd.0002804
Tasdelen Fisgin, N., Ergonul, O., Doganci, L. & Tulek, N. The role of ribavirin in the therapy of Crimean-Congo hemorrhagic fever: early use is promising. Eur J Clin Microbiol Infect Dis 28,929-933(2009). https://doi.org/10.1007/s10096-009-0728-2
Welch, S. R. et al. Identification of 2'-deoxy-2'-fluorocytidine as a potent inhibitor of Crimean-Congo hemorrhagic fever virus replication using a recombinant fluorescent reporter virus. Antiviral Res 147,91-99(2017). https://doi.org/10.1016/j.antiviral.2017.10.008
Pitts, J. et al. Oral dosing of the nucleoside analog obeldesivir is efficacious against RSV infection in African green monkeys. Nat Commun 16, 6437(2025). https://doi.org/10.1038/s41467-025-61595-3
Cross, R. W. et al. Oral administration of obeldesivir protects nonhuman primates against Sudan ebolavirus. Science 383,eadk6176(2024). https://doi.org/10.1126/science.adk6176
Yuan, B. et al. Structure of the Ebola virus polymerase complex. Nature 610, 394-401(2022). https://doi.org/10.1038/s41586-022-05271-2
Yin, W. et al. Structural basis for inhibition of the SARS-CoV-2 RNA polymerase by suramin. Nat Struct Mol Biol 28, 319-325(2021). https://doi.org/10.1038/s41594-021-00570-0
Mastrangelo, E. et al. Structure-based inhibition of Norovirus RNA-dependent RNA polymerases. J Mol Biol 419,198-210(2012). https://doi.org/10.1016/j.jmb.2012.03.008
Jiao, L. et al. Structure of severe fever with thrombocytopenia syndrome virus nucleocapsid protein in complex with suramin reveals therapeutic potential. J Virol 87,6829-6839(2013). https://doi.org/10.1128/JVI.00672-13
Ellenbecker, M., Lanchy, J. M. & Lodmell, J. S. Inhibition of Rift Valley fever virus replication and perturbation of nucleocapsid-RNA interactions by suramin. Antimicrob Agents Chemother 58,7405-7415(2014). https://doi.org/10.1128/AAC.03595-14
Cross, R. W. et al. Oral 4'-fluorouridine rescues nonhuman primates from advanced Lassa fever. Nature(2026). https://doi.org/10.1038/s41586-025-09906-y
Johnson, S. et al. Ribavirin for treating Crimean Congo haemorrhagic fever. Cochrane Database Syst Rev 6,CD012713(2018). https://doi.org/10.1002/14651858.CD012713.pub2
Ascioglu, S., Leblebicioglu, H., Vahaboglu, H. & Chan, K. A. Ribavirin for patients with Crimean-Congo haemorrhagic fever: a systematic review and meta-analysis. J Antimicrob Chemother 66,1215-1222(2011). https://doi.org/10.1093/jac/dkr136
Hawman, D. W. et al. Efficacy of favipiravir(T-705) against Crimean-Congo hemorrhagic fever virus infection in cynomolgus macaques. Antiviral Res 181,104858(2020). https://doi.org/10.1016/j.antiviral.2020.104858
Tipih, T. et al. Favipiravir and Ribavirin protect immunocompetent mice from lethal CCHFV infection. Antiviral Res 218,105703(2023). https://doi.org/10.1016/j.antiviral.2023.105703
Hawman, D. W. et al. Favipiravir(T-705) but not ribavirin is effective against two distinct strains of Crimean-Congo hemorrhagic fever virus in mice. Antiviral Res 157,18-26(2018). https://doi.org/10.1016/j.antiviral.2018.06.013
Brun, R., Blum, J., Chappuis, F. & Burri, C. Human African trypanosomiasis. Lancet 375,148-159(2010). https://doi.org/10.1016/S0140-6736(09)60829-1
Punjani, A., Rubinstein, J. L., Fleet, D. J. & Brubaker, M. A. cryoSPARC: algorithms for rapid unsupervised cryo-EM structure determination. Nat Methods 14, 290-296(2017). https://doi.org/10.1038/nmeth.4169
Pettersen, E. F. et al. UCSF ChimeraX: Structure visualization for researchers, educators, and developers. Protein Sci 30,70-82(2021). https://doi.org/10.1002/pro.3943
Casanal, A., Lohkamp, B. & Emsley, P. Current developments in Coot for macromolecular model building of Electron Cryo-microscopy and Crystallographic Data. Protein Sci 29,1069-1078(2020). https://doi.org/10.1002/pro.3791
Afonine, P. V. et al. Real-space refinement in PHENIX for cryo-EM and crystallography. Acta Crystallogr D Struct Biol 74, 531-544(2018). https://doi.org/10.1107/S2059798318006551
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