State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, The Chinese Academy of Agricultural Sciences, Harbin 150069, China.
Shanghai Veterinary Research Institute, The Chinese Academy of Agricultural Sciences, Shanghai 200241, China.
Viruses. 2022 Mar 10;14(3):572. doi: 10.3390/v14030572.
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/tRNA 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 tRNA 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.
尽管已经付出了许多努力并采用了多种方法，但开发有效的疱疹病毒疫苗仍然是一个巨大的挑战。传统的灭活疫苗和减毒活疫苗总是存在疗效或安全性方面的问题。本研究以猪疱疹病毒（PRV）为模型，尝试利用遗传密码扩展（GCE）技术开发一种活但复制缺陷的 PRV。在正交系统 MbpylRS/tRNA 对和非天然氨基酸（UAA）的存在下，成功拯救了携带提前终止密码子（PTC）的 PRV。然而，我们构建的 PRV PTC 病毒中 UAA 掺入效率似乎非常低。此外，我们未能建立一个稳定的含有正交翻译机制的转染细胞系来复制 PTC 病毒，并且我们证明了正交 tRNA 是一个关键的限制因素。本研究首次证明了正交翻译系统介导的琥珀密码子抑制策略可以精确控制 PRV-PTC 工程病毒的复制。据我们所知，这是首次利用 GCE 技术生成 PTC 疱疹病毒的报道。我们的工作为生成 UAA 控制的 PRV-PTC 病毒提供了概念验证，该病毒可作为一种安全有效的疫苗。
