Toward Predicting Pandemic Potential: A Comparative Analysis of Virus‐Host Interactions Between Diverse Influenza A Viruses and the Human Innate Immune System - 2026
Toward Predicting Pandemic Potential: A Comparative Analysis of Virus‐Host Interactions Between Diverse Influenza A Viruses and the Human Innate Immune System
Gerodez, Antoine; Dos Santos, Melanie; Attia, Mikaëlet al.
[en] ABSTRACT
Zoonotic infections caused by influenza A viruses (IAVs) of animal origin are a major public health concern, as they may represent the initial step toward the emergence of a pandemic strain. The host range of IAVs is shaped by species‐specific virus‐host interactions, and several host barriers have been identified that restrict the adaptation of avian viruses to humans. We used a comparative interaction screen by mammalian cell‐based Nanoluciferase two‐hybrid (mN2H) assay to investigate whether differences in the interaction networks between viral proteins from human, avian, and zoonotic IAVs and a curated panel of human innate immune factors could serve as indicators of the zoonotic potential of avian IAVs. The results revealed both conserved and strain‐specific interactions, while underscoring limitations in comparing interactions across independent screens. Complementary functional analyses by siRNA knockdowns identified cellular factors and pathways potentially involved in host restriction, as well as pro‐viral factors that could represent potential antiviral targets. The comparative interactomic mN2H approach holds promise for evaluating avian IAV zoonotic potential, but our findings highlighted the need for further studies using broader cellular libraries to fully validate its predictive value.
Gerodez, Antoine ; Sciensano, National Influenza Centre Brussels Belgium ; Sciensano, Avian Virology and Immunology Brussels Belgium
Dos Santos, Melanie ; Institut Pasteur, Interactomics RNA and Immunity Laboratory Paris France
Attia, Mikaël ; Institut Pasteur, Interactomics RNA and Immunity Laboratory Paris France
De Geest, Killian; Sciensano, Avian Virology and Immunology Brussels Belgium
Hamid, Ibrahim; Sciensano, National Influenza Centre Brussels Belgium
Tesse, Rodrigue; Sciensano, National Influenza Centre Brussels Belgium ; UMONS - Université de Mons > Faculté des Sciences > Service de biologie cellulaire
Fdillate, Ilham; Sciensano, National Influenza Centre Brussels Belgium
Boschmans, Marc; Sciensano, Avian Virology and Immunology Brussels Belgium
Van Borm, Steven ; Sciensano, Avian Virology and Immunology Brussels Belgium
Steensels, Mieke ; Sciensano, Avian Virology and Immunology Brussels Belgium
Lambrecht, Benedicte ; Sciensano, Avian Virology and Immunology Brussels Belgium
Tafforeau, Lionel ; Université de Mons - UMONS > Faculté des Sciences > Service de Biologie cellulaire
Demeret, Caroline ; Institut Pasteur, Interactomics RNA and Immunity Laboratory Paris France
Barbezange, Cyril ; Sciensano, National Influenza Centre Brussels Belgium
Toward Predicting Pandemic Potential: A Comparative Analysis of Virus‐Host Interactions Between Diverse Influenza A Viruses and the Human Innate Immune System
A. D. Iuliano, K. M. Roguski, H. H. Chang, et al., “Estimates of Global Seasonal Influenza-Associated Respiratory Mortality: A Modelling Study,” Lancet 391 (2018): 1285–1300, https://doi.org/10.1016/S0140-6736(17)33293-2.
N. M. Bouvier and P. Palese, “The Biology of Influenza Viruses,” Vaccine 26, no. 4 (2008): D49–D53, https://doi.org/10.1016/j.vaccine.2008.07.039.
W. Hao, L. Wang, and S. Li, “Roles of the Non-Structural Proteins of Influenza A Virus,” Pathogens 9 (2020): 812, https://doi.org/10.3390/pathogens9100812.
S. Fereidouni, E. Starick, K. Karamendin, et al., “Genetic Characterization of a New Candidate Hemagglutinin Subtype of influenza A Viruses,” Emerging Microbes & Infections 12 (2023): 2225645, https://doi.org/10.1080/22221751.2023.2225645.
I. Wendel, M. Matrosovich, and H. D. Klenk, “SnapShot: Evolution of Human Influenza A Viruses,” Cell Host & Microbe 17 (2015): 416, https://doi.org/10.1016/j.chom.2015.02.001.
European Food Safety Authority, European Centre for Disease Prevention and Control, Influenza, European Union Reference Laboratory for Avian Influenza. L. Alexakis, H. Buczkowski, M. Ducatez, et al., “Avian Influenza Overview December 2024–March 2025,” EFSA Journal 23 (2025): 9352, https://doi.org/10.2903/j.efsa.2025.9352.
World Health Organization. Avian Influenza Weekly Update Number 983, 2025, https://cdn.who.int/media/docs/default-source/wpro---documents/emergency/surveillance/avian-influenza/ai_20250131.pdf?sfvrsn=5f006f99_149.
Y.-H. Wang, J.-J. Chen, J. Ma, et al., “Early-Warning Signals and the Role of H9N2 in the Spillover of Avian Influenza Viruses,” Med 6 (2025): 100639, https://doi.org/10.1016/j.medj.2025.100639.
J. S. Long, B. Mistry, S. M. Haslam, and W. S. Barclay, “Host and Viral Determinants of Influenza A Virus Species Specificity,” Nature Reviews Microbiology 17 (2019): 67–81, https://doi.org/10.1038/s41579-018-0115-z.
G. Malik and Y. Zhou, “Innate Immune Sensing of Influenza A Virus,” Viruses 12 (2020): 755, https://doi.org/10.3390/v12070755.
W. An, S. Lakhina, J. Leong, K. Rawat, and M. Husain, “Host Innate Antiviral Response to influenza A Virus Infection: From Viral Sensing to Antagonism and Escape,” Pathogens 13 (2024): 561, https://doi.org/10.3390/pathogens13070561.
Z. Ji, X. Wang, and X. Liu, “NS1: A Key Protein in the “Game” Between Influenza A Virus and Host in Innate Immunity,” Frontiers in Cellular and Infection Microbiology 11 (2021): 670177, https://doi.org/10.3389/fcimb.2021.670177.
R. M. Pinto, S. Bakshi, S. Lytras, et al., “BTN3A3 Evasion Promotes the Zoonotic Potential of Influenza A Viruses,” Nature 619 (2023): 338–347, https://doi.org/10.1038/s41586-023-06261-8.
C. M. Deeg, E. Hassan, P. Mutz, et al., “In Vivo Evasion of MxA by Avian Influenza Viruses Requires Human Signature in the Viral Nucleoprotein,” Journal of Experimental Medicine 214 (2017): 1239–1248, https://doi.org/10.1084/jem.20161033.
M. Weber, H. Sediri, U. Felgenhauer, et al., “Influenza Virus Adaptation PB2-627K Modulates Nucleocapsid Inhibition by the Pathogen Sensor RIG-I,” Cell Host & Microbe 17 (2015): 309–319, https://doi.org/10.1016/j.chom.2015.01.005.
P. Cassonnet, C. Rolloy, G. Neveu, et al., “Benchmarking a Luciferase Complementation Assay for Detecting Protein Complexes,” Nature Methods 8 (2011): 990–992, https://doi.org/10.1038/nmeth.1773.
S. G. Choi, J. Olivet, P. Cassonnet, et al., “Maximazing Binary Interactome Mapping With a Minimal Number of Assays,” Nature Communications 10 (2019): 3907, https://doi.org/10.1038/s41467-019-11809-2.
P. Braun, M. Tasan, M. Dreze, et al., “An Experimentally Derived Confidence Score for Binary Protein-Protein Interactions,” Nature Methods 6 (2009): 91–97, https://doi.org/10.1038/nmeth.1281.
E. Biquand, J. Poirson, M. Karim, et al., “Comparative Profiling of Ubiquitin Proteasome System Interplay With Influenza A Virus PB2 Polymerase Protein Recapitulating Virus Evolution in Humans,” mSphere 2 (2017): e00330–e00417, https://doi.org/10.1128/msphere.00330-17.
M. Muller, Y. Jacob, L. Jones, et al., “Large Scale Genotype Comparison of Human Papillomavirus E2-Host Interaction Networks Provides New Insights for E2 Molecular Functions,” PLOS Pathogens 8 (2012): 1002761, https://doi.org/10.1371/journal.ppat.1002761.
M. Matrosovich, T. Matrosovich, J. Carr, N. A. Roberts, and H.-D. Klenk, “Overexpression of the α-2,6-Sialyltransferase in MDCK Cells Increases Influenza Virus Sensitivity to Neuraminidase Inhibitors,” Journal of Virology 77 (2003): 8418–8425, https://doi.org/10.1128/jvi.77.15.8418-8425.2003.
G. Kayali, A. Kandeil, R. El-Shesheny, et al., “Avian Influenza A(H5N1) Virus in Egypt,” Emerging Infectious Diseases 22 (2016): 379–388, https://doi.org/10.3201/eid2203.150593.
R. El-Shesheny, A. Kandeil, A. Mostafa, M. A. Ali, and R. J. Webby, “H5 Influenza Viruses in Egypt,” Cold Spring Harbor Perspectives in Medicine 11 (2021): a038745, https://doi.org/10.1101/cshperspect.a038745.
A. Fusaro, B. Zecchin, E. Giussani, et al., “High Pathogenic Avian Influenza A(H5) Viruses of Clade 2.3.4.4b in Europe—Why Trends of Virus Evolution are More Difficult to Predict,” Virus Evolution 10 (2024): veae027, https://doi.org/10.1093/ve/veae027.
S. G. Choi, J. Olivet, P. Cassonnet, et al., “Maximizing Binary Interactome Mapping With a Minimal Number of Assays,” Nature Communications 10 (2019): 3907, https://doi.org/10.1038/s41467-019-11809-2.
V. Navratil, B. de Chassey, L. Meyniel, F. Pradezynski, C. Rabourdin-Combe, and V. Lotteau, “System-Level Comparison of Protein−Protein Interactions Between Viruses and the Human Type I Interferon System Network,” Journal of Proteome Research 9 (2010): 3527–3536, https://doi.org/10.1021/pr100326j.
M. Morita, K. Kuba, A. Ichikawa, et al., “The Lipid Mediator Protectin D1 Inhibits Influenza Virus Replication and Improves Severe Influenza,” Cell 153 (2013): 112–125, https://doi.org/10.1016/j.cell.2013.02.027.
A. J. Eisfeld, E. Kawakami, T. Watanabe, G. Neumann, and Y. Kawaoka, “RAB11A is Essential for Transport of the Influenza Virus Genome to the Plasma Membrane,” Journal of Virology 85 (2011): 6117–6126, https://doi.org/10.1128/jvi.00378-11.
Z. Chen, C. Wang, X. Feng, et al., “Interactomes of SARS-CoV-2 and Human Coronaviruses Reveal Host Factors Potentially Affecting Pathogenesis,” EMBO Journal 40 (2021): 107776, https://doi.org/10.15252/embj.2021107776.
L. Wang, B. Fu, W. Li, et al., “Comparative Influenza Protein Interactomes Identify the Role of Plakophilin 2 in Virus Restriction,” Nature Communications 8 (2017): 13876, https://doi.org/10.1038/ncomms13876.
L. Martin-Sancho, S. Tripathi, A. Rodriguez-Frandsen, et al., “Restriction Factor Compendium for Influenza A Virus Reveals a Mechanism for Evasion of Autophagy,” Nature Microbiology 6 (2021): 1319–1333, https://doi.org/10.1038/s41564-021-00964-2.
S. Fernbach, E. E. Spieler, I. Busnadiego, et al., “Restriction Factor Screening Identifies RABGAP1L-Mediated Disruption of Endocytosis as a Host Antiviral Defense,” Cell Reports 38 (2022): 110549, https://doi.org/10.1016/j.celrep.2022.110549.
M.-L. Goulet, D. Olagnier, Z. Xu, et al., “Systems Analysis of a RIG-I Agonist Inducing Broad Spectrum Inhibition of Virus Infectivity,” PLOS Pathogens 9 (2013): 1003298, https://doi.org/10.1371/journal.ppat.1003298.
M. Kandasamy, A. Suryawanshi, S. Tundup, et al., “RIG-I Signaling Is Critical for Efficient Polyfunctional T Cell Responses During Influenza Virus Infection,” PLOS Pathogens 12 (2016): 1005754, https://doi.org/10.1371/journal.ppat.1005754.
D. M. Hammerbeck, G. R. Burleson, C. J. Schuller, et al., “Administration of a Dual Toll-Like Receptor 7 and Toll-Like Receptor 8 Agonist Protects Against Influenza in Rats,” Antiviral Research 73 (2007): 1–11, https://doi.org/10.1016/j.antiviral.2006.07.011.
D. J. Lynn, C. Chan, M. Naseer, et al., “Curating the Innate Immunity Interactome,” BMC Systems Biology 4 (2010): 117, https://doi.org/10.1186/1752-0509-4-117.
H. Hong, H.-K. Choi, and T.-Y. Yoon, “Untangling the Complexity of Membrane Protein Folding,” Current Opinion in Structural Biology 72 (2022): 237–247, https://doi.org/10.1016/j.sbi.2021.11.013.
T. P. Peacock, L. Moncla, G. Dudas, et al., “The Global H5N1 Influenza Panzootic in Mammals,” Nature 637 (2025): 304–313, https://doi.org/10.1038/s41586-024-08054-z.
M. Declercq, E. Biquand, M. Karim, et al., “Influenza A Virus co-opts ERI1 Exonuclease Bound to Histone mRNA to Promote Viral Transcription,” Nucleic Acids Research 48 (2020): 10428–10440, https://doi.org/10.1093/nar/gkaa771.
A. Gerodez, F. E. Dufrasne, O. Denis, et al., “The Cellular Activating Protein 1 cFos Regulates Influenza A Virus Replication,” Journal of General Virology 107 (2026): 002194, https://doi.org/10.1099/jgv.0.002194.
J. Xie, S. Zhang, Y. Hu, et al., “Regulatory Roles of c-jun in H5N1 Influenza Virus Replication and Host Inflammation,” Biochimica et Biophysica Acta (BBA)—Molecular Basis of Disease 1842 (2014): 2479–2488, https://doi.org/10.1016/j.bbadis.2014.04.017.
J. Wang, J. Sun, J. Hu, et al., “A77 1726, the Active Metabolite of the Anti-Rheumatoid Arthritis Drug Leflunomide, Inhibits influenza A Virus Replication In Vitro and In Vivo by Inhibiting the Activity of Janus Kinases,” FASEB Journal 34 (2020): 10132–10145, https://doi.org/10.1096/fj.201902793RR.
T. Watanabe, E. Kawakami, J. E. Shoemaker, et al., “Influenza Virus-Host Interactome Screen as a Platform for Antiviral Drug Development,” Cell Host & Microbe 16 (2014): 795–805, https://doi.org/10.1016/j.chom.2014.11.002.
J. Han, J. T. Perez, C. Chen, et al., “Genome-Wide CRISPR/Cas9 Screen Identifies Host Factors Essential for Influenza Virus Replication,” Cell Reports 23 (2018): 596–607, https://doi.org/10.1016/j.celrep.2018.03.045.
S. Zhang, C. Huo, J. Xiao, et al., “p-STAT1 Regulates the Influenza A Virus Replication and Inflammatory Response In Vitro and Vivo,” Virology 537 (2019): 110–120, https://doi.org/10.1016/j.virol.2019.08.023.
J. Zhang, Y. Wu, Y. Wang, et al., “Influenza A Virus Infection Activates STAT3 to Enhance SREBP2 Expression, Cholesterol Biosynthesis, and Virus Replication,” iScience 27 (2024): 110424, https://doi.org/10.1016/j.isci.2024.110424.
A. Mecate-Zambrano, S. Sukumar, and G. Seebohm, et al., “Discrete Spatio-Temporal Regulation of Tyrosine Phosphorylation Directs Influenza A Virus M1 Protein Towards Its Function in Virion Assembly,” PLOS Pathogens (2020): 1008775, https://doi.org/10.1371/journal.ppat.1008775.
S. Wang, Z. Zhao, Y. Bi, L. Sun, X. Liu, and W. Liu, “Tyrosine 132 Phosphorylation of Influenza A Virus M1 Protein is Crucial for Virus Replication by Controlling the Nuclear Import of M1,” Journal of Virology 87 (2013): 6182–6191, https://doi.org/10.1128/jvi.03024-12.
X. Hu, J. Li, M. Fu, X. Zhao, and W. Wang, “The JAK/STAT Signaling Pathway: From Bench to Clinic,” Signal Transduction and Targeted Therapy 6 (2021): 402, https://doi.org/10.1038/s41392-021-00791-1.
Y. Du, F. Yang, Q. Wang, et al., “Influenza A Virus Antagonizes Type I and Type II Interferon Responses via SOCS1-Dependent Ubiquitination and Degradation of JAK1,” Virology Journal 17 (2020): 74, https://doi.org/10.1186/s12985-020-01348-4.
H. Yang, Y. Dong, Y. Bian, et al., “The Influenza Virus PB2 Protein Evades Antiviral Innate Immunity by Inhibiting JAK1/STAT Signalling,” Nature Communications 13 (2022): 6288, https://doi.org/10.1038/s41467-022-33909-2.
S. Dam, M. Kracht, S. Pleschka, and M. L. Schmitz, “The influenza A Virus Genotype Determines the Antiviral Function of NF-κB,” Journal of Virology 90 (2016): 7980–7990, https://doi.org/10.1128/jvi.00946-16.
