Optimizing encephalomyocarditis virus VP1 protein assembly on pseudorabies virus envelope via US9 protein anchoring

• Published in: Virulence 2025-12, Vol.16 (1), p.2445235
• Main Authors: Ren, Bao Ru, Qin, Hui, Zhang, Yan Fang, Du, Xuan Ying, Sun, Hao Chen, Luo, Yang, Gan, Yan Zi, Wang, Jian Long, Hu, Wei, Cao, Sheng Bo, Yang, Yang
• Format: Article
• Language: English
• Subjects: Animals; Antibodies, Neutralizing - immunology; Antibodies, Viral - immunology; Capsid Proteins - genetics; Capsid Proteins - immunology; Capsid Proteins - metabolism; Cardiovirus Infections - immunology; Cardiovirus Infections - prevention & control; Cardiovirus Infections - virology; Encephalomyocarditis virus; Encephalomyocarditis virus - genetics; Encephalomyocarditis virus - immunology; Encephalomyocarditis virus - metabolism; Female; Herpesvirus 1, Suid - genetics; Herpesvirus 1, Suid - immunology; Intracellular Signaling Peptides and Proteins; Lipoproteins; Mice; Mice, Inbred BALB C; Pseudorabies - immunology; Pseudorabies - prevention & control; Pseudorabies - virology; pseudorabies virus; surface display; Swine; Swine Diseases - immunology; Swine Diseases - prevention & control; Swine Diseases - virology; US9 C-terminal; vaccine vector; Viral Envelope Proteins - genetics; Viral Envelope Proteins - immunology; Viral Envelope Proteins - metabolism; Viral Proteins - genetics; Viral Proteins - immunology; Viral Proteins - metabolism; Viral Vaccines - genetics; Viral Vaccines - immunology
• ISSN: 2150-5594; 2150-5608; 2150-5608
• DOI: 10.1080/21505594.2024.2445235

Live herpesvirus-vectored vaccines are critical in veterinary medicine, but they can sometimes offer insufficient protection due to suboptimal antigen expression or localization. Encephalomyocarditis virus (EMCV) is a significant zoonotic threat, with VP1 protein as a key immunogen on its capsid. To enhance immunogenicity, we explored the use of recombinant pseudorabies virus (rPRV) as a vaccine vector against EMCV. In silico analysis indicated that fusing VP1 with US9 enhances the formation of a type II transmembrane heterodimer. We constructed six rPRV groups expressing different VP1 variants and found that VP1 fused with US9's C-terminal (US9-VP1) enhances VP1's membrane localization and its incorporation into the PRV envelope, unlike wild-type VP1. Immunogold electron microscopy illustrated that rPRV with deleted US8 and US9, supplemented with US8 regulatory sequence (rΔ89-U9VP1), improved VP1 incorporation into the viral envelope. Post-immunization, only rΔ89-U9VP1 provided 100% protection against EMCV in mice and induced high levels of virus-neutralizing antibodies in piglets. Additionally, rPRV expressing VP1 stimulated robust T-cell responses, as demonstrated by flow cytometry and ELISpot assays. This study introduces rPRV as a potential EMCV vaccine, demonstrating that the selection of the US9 C-terminal domain and US8 regulatory sequence significantly enhances the presentation of heterologous antigens, improving vaccine efficacy.