Supplementary file 1_The role and mechanism of p53 F229V mutation in inhibiting pseudorabies virus replication.docx

The p53 protein is a key transcription factor that regulates cellular responses to stress and is widely recognized as a host restriction factor against various viral infections. However, its specific role in pseudorabies virus (PRV) replication, pathogenesis, and host response remains unclear. This study identified a swine p53 (sp53) mutation in the PK15 cell line under prolonged passage stress, characterized by an amino acid substitution at position 229, replacing phenylalanine with valine (sp53 F229V). This mutation eliminates the transcriptional activity of wild-type p53 and confers resistance to PRV infection. Notably, it reverses p53’s original pro-viral role, converting it into an inhibitor of PRV proliferation. Further analysis revealed that the PRV early protein EP0 promotes the degradation of sp53 F229V through the proteasome pathway. These findings indicate that a defined p53 alteration can decouple transcriptional and antiviral functions in a mutation-specific, context-dependent manner. The EP0–p53 interface emerges as a candidate target to modulate PRV replication, pending validation. These findings were obtained in vitro and require in vivo validation in pigs to determine their relevance to PRV pathogenesis.

p53蛋白是一类关键转录因子，可调控细胞对应激的应答反应，且被广泛认定为对抗多种病毒感染的宿主限制因子。然而，其在伪狂犬病病毒（pseudorabies virus, PRV）的复制、致病机制以及宿主应答过程中的具体作用仍有待阐明。本研究在长期传代应激的PK15细胞系中发现了一处猪p53（swine p53, sp53）突变，该突变位于第229位氨基酸位点，表现为苯丙氨酸被缬氨酸取代（sp53 F229V）。此突变可消除野生型p53的转录活性，并使细胞获得PRV感染抗性。值得关注的是，该突变逆转了p53原本的促病毒功能，将其转变为PRV增殖的抑制剂。进一步分析显示，PRV早期蛋白EP0可通过蛋白酶体通路促进sp53 F229V的降解。上述研究结果表明，特定的p53改变能够以突变特异性、情境依赖的方式，解偶联其转录功能与抗病毒功能。EP0与p53的互作界面有望成为调控PRV复制的候选靶点，尚待后续验证。本研究所有发现均基于体外实验，需在猪体内开展验证以明确其与PRV致病机制的相关性。