Spatial remodeling of sphingomyelin synthesis by Zika virus drives neuroinflammation and cortical developmental injury
Abstract
Neuroinflammation can disrupt brain development, yet the cellular mechanisms driving pathogenic inflammatory responses in the developing brain remain poorly understood. Here, we identify ectopic sphingomyelin synthesis at the endoplasmic reticulum–Golgi intermediate compartment (ERGIC) as a mechanism linking Zika virus (ZIKV) infection to neuroinflammation and cortical developmental injury. Using super-resolution imaging and lipidomics, we show that the viral protein NS4A redirects sphingomyelin synthase 1 (SGMS1) and ceramide transfer protein (CERT1) to the ERGIC, establishing ectopic sphingomyelin synthesis hub. The resulting sphingomyelin accumulation at the ERGIC promotes homo-oligomerization and activation of TMED10, an ERGIC-resident protein translocator that mediates unconventional protein secretion, thereby driving interleukin-1β (IL-1β) release and neuroinflammation. In mice, inhibition of sphingomyelin synthesis attenuated IL-1β release and neuroinflammation while preserving cortical development. Furthermore, functional mutational screening identified key TMED10 residues required for sphingomyelin sensing. Disruption of this lipid-sensing mechanism via point mutation suppressed ZIKV-induced inflammatory responses in vitro and ameliorated neuroinflammation and cortical developmental defects in vivo. These findings establish an ERGIC sphingomyelin–TMED10 axis linking spatial lipid remodeling to pathogenic inflammatory signaling and identify a potential therapeutic target for inflammation-driven neurodevelopmental injury.
References
1. Mogensen, T.H. Pathogen Recognition and Inflammatory Signaling in Innate Immune Defenses. Clin Microbiol Rev 22, 240–273 (2009).
2. Medzhitov, R. Origin and physiological roles of inflammation. Nature 454, 428–435 (2008).
3. Ostrem, L.M., et al. Fetal brain response to maternal inflammation requires microglia. Development (2024).
4. Otero, A.M. & Antonson, A.M. At the crux of maternal immune activation: Viruses, microglia, microbes, and IL-17A. Immunol Rev 311, 205–223 (2022).
5. Couch, A.C.M., Berger, T., Hanger, B., Matuleviciute, R. & Vernon, A.C. Maternal Immune Activation Primes Deficiencies in Adult Hippocampal Neurogenesis. Brain Behav Immun, 410–422 (2021).
6. Martín-Guerrero, S.M., et al. Maternal immune activation imprints translational dysregulation and differential MAP2 phosphorylation in descendant neural stem cells. Mol Psychiatr 30, 2994–3007 (2025).
7. Assir, M.Z.K., et al. Modeling maternal immune activation in 3D ex vivo human fetal brain cerebroids reveals IL-17A-driven disruption of cortical development. Nat Neurosci 29, 2456–2468 (2026).
8. Mlakar, J., et al. Zika Virus Associated with Microcephaly. New Engl J Med 374, 951–958 (2016).
9. Rasmussen, S.A., Jamieson, D.J., Honein, M.A. & Petersen, L.R. Zika Virus and Birth Defects - Reviewing the Evidence for Causality. New Engl J Med 374, 1981–1987 (2016).
10. Li, C., et al. Zika Virus Disrupts Neural Progenitor Development and Leads to Microcephaly in Mice Cell Stem Cell 19, 672–672 (2016).
11. Tang, H.L., et al. Zika Virus Infects Human Cortical Neural Progenitors and Attenuates Their Growth. Cell Stem Cell 18, 587–590 (2016).
12. Komarasamy, T.V., Adnan, N.A.A., James, W. & Balasubramaniam, V.R.M.T. Zika Virus Neuropathogenesis: The Different Brain Cells, Host Factors and Mechanisms Involved. Front Immunol 13, 773191 (2022).
13. Garcez, P.P., et al. Zika virus impairs growth in human neurospheres and brain organoids. Science 352, 816–818 (2016).
14. Wang, J., et al. Zika virus infected primary microglia impairs NPCs proliferation and differentiation. Biochem Bioph Res Co 497, 619–625 (2018).
15. Hassaan, N.A. & Xing, L. The mechanisms of Zika virus-induced neuropathogenesis. Braz J Microbiol 55, 3933–3943 (2024).
16. de Sousa, J.R., et al. Inflammasome activation results in severe damage to the central nervous system in fatal Zika virus microcephaly cases. Cytokine 111, 255–264 (2018).
17. Lima, M.C., et al. The Transcriptional and Protein Profile From Human Infected Neuroprogenitor Cells Is Strongly Correlated to Zika Virus Microcephaly Cytokines Phenotype Evidencing a Persistent Inflammation in the CNS. Front Immunol 10(2019).
18. Nascimento-Carvalho, G.C., et al. Zika-exposed microcephalic neonates exhibit higher degree of inflammatory imbalance in cerebrospinal fluid. Sci Rep-Uk 11, 8474 (2021).
19. Dang, J., et al. Zika Virus Depletes Neural Progenitors in Human Cerebral Organoids through Activation of the Innate Immune Receptor TLR3. Cell Stem Cell 19, 258–265 (2016).
20. Lum, F.M., et al. Zika Virus Infects Human Fetal Brain Microglia and Induces Inflammation. Clin Infect Dis 64, 914–920 (2017).
21. Xu, Y.-P., et al. Zika virus infection induces RNAi-mediated antiviral immunity in human neural progenitors and brain organoids. Cell Research 29, 265–273 (2019).
22. Jeong, G.U., et al. Zika Virus Infection Induces Interleukin-1β-Mediated Inflammatory Responses by Macrophages in the Brain of an Adult Mouse Model. J Virol 97, e0055623 (2023).
23. Wang, W.B., et al. Zika virus infection induces host inflammatory responses by facilitating NLRP3 inflammasome assembly and interleukin-1β secretion. Nature Communications 9, 106 (2018).
24. Miner, J.J., et al. Zika Virus Infection during Pregnancy in Mice Causes Placental Damage and Fetal Demise. Cell 165, 1080–1090 (2016).
25. Yockey, L.J., et al. Vaginal Exposure to Zika Virus during Pregnancy Leads to Fetal Brain Infection. Cell 166, 1247–1256.e1244 (2016).
26. Cugola, F.R., et al. The Brazilian Zika virus strain causes birth defects in experimental models. Nature 534, 267–271 (2016).
27. Liu, X.Y. & Quan, N. Microglia and CNS Interleukin-1: Beyond Immunological Concepts. Front Neurol 9(2018).
28. Muzio, L., Viotti, A. & Martino, G. Microglia in Neuroinflammation and Neurodegeneration: From Understanding to Therapy. Front Neurosci-Switz 15(2021).
29. Monif, M., et al. Interleukin-1β has trophic effects in microglia and its release is mediated by P2X7R pore. J Neuroinflamm 13, 173 (2016).
30. He, W.T., et al. Gasdermin D is an executor of pyroptosis and required for interleukin-1β secretion. Cell Research 25, 1285–1298 (2015).
31. Kayagaki, N., et al. Caspase-11 cleaves gasdermin D for non-canonical inflammasome signalling. Nature 526, 666–671 (2015).
32. Shi, J.J., et al. Cleavage of GSDMD by inflammatory caspases determines pyroptotic cell death. Nature 526, 660–665 (2015).
33. Lieberman, J., Wu, H. & Kagan, J.C. Gasdermin D activity in inflammation and host defense. Sci Immunol 4, eaav1447 (2019).
34. Lei, J., et al. IL-1 receptor antagonist therapy mitigates placental dysfunction and perinatal injury following Zika virus infection. Jci Insight 4, e122678 (2019).
35. Zhang, M., et al. A translocation pathway for vesicle-mediated unconventional protein secretion. Cell 181, 637–652 (2020).
36. Huitema, K., van den Dikkenberg, J., Brouwers, J.F.H.M. & Holthuis, J.C.M. Identification of a family of animal sphingomyelin synthases. Embo J 23, 33–44 (2004).
37. Hanada, K., et al. Molecular machinery for non-vesicular trafficking of ceramide. Nature 426, 803–809 (2003).
38. Hannun, Y.A. & Obeid, L.M. Sphingolipids and their metabolism in physiology and disease. Nat Rev Mol Cell Bio 19, 175–191 (2018).
39. Othman, M.A., et al. Malabaricone C as Natural Sphingomyelin Synthase Inhibitor against Diet-Induced Obesity and Its Lipid Metabolism in Mice. Acs Med Chem Lett 10, 1154–1158 (2019).
40. Yasuda, S., et al. A novel inhibitor of ceramide trafficking from endoplasmic reticulum to the site of sphingomyelin synthesis J Biol Chem 288, 24162–24162 (2013).
41. Abdelbasset, M., et al. Differential contributions of fetal mononuclear phagocytes to Zika virus neuroinvasion versus neuroprotection during congenital infection. Cell 187, 7511–7532 (2024).
42. Yeang, C., Ding, T.B., Chirico, W.J. & Jiang, X.C. Subcellular Targeting Domains of Sphingomyelin Synthase 1 and 2. Nutr Metab 8, 89 (2011).
43. Subathra, M., Qureshi, A. & Luberto, C. Sphingomyelin Synthases Regulate Protein Trafficking and Secretion. Plos One 6, e23644 (2011).
44. Kumagai, K. & Hanada, K. Structure, functions and regulation of CERT, a lipid-transfer protein for the delivery of ceramide at the ER-Golgi membrane contact sites. Febs Lett 593, 2366–2377 (2019).
45. Arita, M., et al. Phosphatidylinositol 4-Kinase III Beta Is a Target of Enviroxime-Like Compounds for Antipoliovirus Activity. J Virol 85, 2364–2372 (2011).
46. Zheng, J.F., et al. TMEDs mediate versatile cargo transport in vesicle-dependent unconventional secretion. J Cell Biol 225, e202503075 (2025).
47. Schwinn, M.K., et al. CRISPR-Mediated Tagging of Endogenous Proteins with a Luminescent Peptide. ACS chemical biology, 467 (2017).
48. Passaro, S., et al. Boltz-2: Towards Accurate and Efficient Binding Affinity Prediction. bioRxiv, 2025.2006.2014.659707 (2025).
49. Leier, H.C., et al. A global lipid map defines a network essential for Zika virus replication. Nature Communications 11, 3652 (2020).
50. Diop, F., et al. Zika virus infection modulates the metabolomic profile of microglial cells. Plos One 13, e0206093 (2018).
51. Hatton, C.F. & Duncan, C.J.A. Microglia Are Essential to Protective Antiviral Immunity: Lessons From Mouse Models of Viral Encephalitis. Front Immunol 10(2019).
52. Sun, Y.X., et al. A dual role of ERGIC-localized Rabs in TMED10-mediated unconventional protein secretion. Nat Cell Biol 26, 1623–1623 (2024).
53. L.J. REED , H.M. A SIMPLE METHOD OF ESTIMATING FIFTY PER CENT ENDPOINTS. American Journal of Epidemiology 27, 493–497 (1938).
54. Zeng, J.X., et al. The Zika Virus Capsid Disrupts Corticogenesis by Suppressing Dicer Activity and miRNA Biogenesis. Cell Stem Cell 27, 618–632.e619 (2020).
55. Kim, J.Y., Grunke, S.D., Levites, Y., Golde, T.E. & Jankowsky, J.L. Intracerebroventricular Viral Injection of the Neonatal Mouse Brain for Persistent and Widespread Neuronal Transduction. Jove-J Vis Exp, 51863 (2014).
56. Jang, A. & Lehtinen, M.K. In utero intracerebroventricular delivery of adeno-associated viral vectors to target mouse choroid plexus and cerebrospinal fluid. Star Protoc 4, 101975 (2023).
57. Rathore, A.P.S., Saron, W.A.A., Lim, T., Johan, N. & St John, A.L. Maternal immunity and antibodies to dengue virus promote infection and Zika virus-induced microcephaly in fetuses. Sci Adv 5, eaav3208 (2019).
58. Yuan, L., et al. A single mutation in the prM protein of Zika virus contributes to fetal microcephaly. Science 358, 933–936 (2017).
59. Chen, T., et al. ImageGP 2 for enhanced data visualization and reproducible analysis in biomedical research. Imeta 3, e239 (2024).
60. Cho, K.F., et al. Proximity labeling in mammalian cells with TurboID and split-TurboID. Nat Protoc 15, 3971–3999 (2020).
61. Ge, L., Melville, D., Zhang, M. & Schekman, R. The ER-Golgi intermediate compartment is a key membrane source for the LC3 lipidation step of autophagosome biogenesis. Elife 2, e00947 (2013).
62. Ulmer, C.Z., Jones, C.M., Yost, R.A., Garrett, T.J. & Bowden, J.A. Optimization of Folch, Bligh-Dyer, and Matyash sample-to-extraction solvent ratios for human plasma-based lipidomics studies. Anal Chim Acta 1037, 351–357 (2018).
63. Reichl, B., et al. Evaluation and optimization of common lipid extraction methods in cerebrospinal fluid samples. J Chromatogr B 1153, 122271 (2020).
64. Maekawa, M. & Fairn, G.D. Molecular probes to visualize the location, organization and dynamics of lipids. J Cell Sci 127, 4801–4812 (2014).
65. Deng, Y., Rivera-Molina, F.E., Toomre, D.K. & Burd, C.G. Sphingomyelin is sorted at the trans Golgi network into a distinct class of secretory vesicle. Mol Biol Cell 27, 6677–6682 (2016).
66. Hao, Y.H., et al. Integrated analysis of multimodal single-cell data. Cell 184, 3573–3587 (2021).
67. Stuart, T., et al. Comprehensive Integration of Single-Cell Data. Cell 177, 1888–1902 (2019).
68. Stokes, C., et al. The human neural cell atlas of Zika virus infection in developing brain tissue. Cell Rep Med 6, 102189 (2025).
69. Subramanian, A., et al. Gene set enrichment analysis: A knowledge-based approach for interpreting genome-wide expression profiles. P Natl Acad Sci USA 102, 15545–15550 (2005).
70. Liberzon, A., et al. Molecular signatures database (MSigDB) 3.0. Bioinformatics 27, 1739–1740 (2011).
71. Arthur Liberzon, C.B., Helga Thorvaldsdóttir, Mahmoud Ghandi, Jill P Mesirov, Pablo Tamayo. The Molecular Signatures Database (MSigDB) hallmark gene set collection. Cell systems 1, 417–425 (2025).
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