Abstract
Despite many efforts and diverse approaches, developing an effective herpesvirus vaccine remains a great challenge. Traditional inactivated and live-attenuated vaccines always raise efficacy or safety concerns. This study used Pseudorabies virus (PRV), a swine herpes virus, as a model. We attempted to develop a live but replication-incompetent PRV by genetic code expansion (GCE) technology. Premature termination codon (PTC) harboring PRV was successfully rescued in the presence of orthogonal system MbpylRS/tRNAPyl pair and unnatural amino acids (UAA). However, UAA incorporating efficacy seemed extremely low in our engineered PRV PTC virus. Furthermore, we failed to establish a stable transgenic cell line containing orthogonal translation machinery for PTC virus replication, and we demonstrated that orthogonal tRNAPyl is a key limiting factor. This study is the first to demonstrate that orthogonal translation system-mediated amber codon suppression strategy could precisely control PRV-PTC engineered virus replication. To our knowledge, this is the first reported PTC herpesvirus generated by GCE technology. Our work provides a proof-of-concept for generating UAAs-controlled PRV-PTC virus, which can be used as a safe and effective vaccine.
Highlights
Evaluation of unnatural amino acids (UAA) Site-Specific Incorporation for Potential Pseudorabies virus (PRV) gB Premature termination codon (PTC) Sites gB was recognized as an essential gene for PRV replication
The results demonstrated that MbpylRS/tRNAPyl pair could be successfully delivered by
Harboring PRV, and the results suggested that PRV-PTC virus was successfully rescued in the presence of orthogonal system MbpylRS/tRNAPyl pair and UAA
Summary
State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Shanghai Veterinary Research Institute, The Chinese Academy of Agricultural Sciences, Henan Provincial Engineering and Technology Center of Animal Disease Diagnosis and Integrated Control, These authors contributed to this work. This study used Pseudorabies virus (PRV), a swine herpes virus, as a model. UAA incorporating efficacy seemed extremely low in our engineered PRV PTC virus. This study is the first to demonstrate that orthogonal translation system-mediated amber codon suppression strategy could precisely control PRV-PTC engineered virus replication. To our knowledge, this is the first reported PTC herpesvirus generated by GCE technology. Our work provides a proof-of-concept for generating UAAs-controlled PRV-PTC virus, which can be used as a safe and effective vaccine. Developing a safe and effective PRV vaccine is one of the best choices for PRV control in related animals and humans. The inactivated PRV vaccine mainly induces a humoral immune response, lacks effective
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