Reduced Susceptibility to Transmissible Gastroenteritis Virus in Porcine ANPEP N783A-Edited Cell Clone

Transmissible gastroenteritis virus (TGEV) uses porcine aminopeptidase N (pAPN), encoded by the alanyl aminopeptidase gene (ANPEP), as a cellular receptor. Complete ANPEP disruption can restrict TGEV infection but may compromise APN-associated functions. Here, we investigated whether limited modification of pAPN at residue N783 could reduce cellular susceptibility to TGEV without eliminating APN protein expression or the aminopeptidase activity examined in this study. Prime editing was then used to generate an ANPEP N783A-edited clone in PE7-expressing PK15 cells. The edited clone retained APN protein expression, cell-normalized aminopeptidase activity, and membrane-associated APN staining. Following TGEV exposure at multiplicities of infection of 0.01, 0.1, and 1, intracellular RNA containing the TGEV nucleocapsid (N) gene sequence at 24 h post-infection was reduced by approximately 83%, 73%, and 74%, respectively. The edited cells also maintained higher relative viability under most infected conditions, with the greatest difference observed at 36 hpi and an MOI of 1 (83% vs. 42%). Relative LDH release was reduced by approximately 28% and 22% at MOIs of 0.1 and 1, respectively. The edited cells also displayed weaker dead-cell fluorescence and milder morphological damage. These findings provide preliminary, clone-specific evidence that an ANPEP N783A-edited clone can display reduced susceptibility to TGEV while retaining the APN-related phenotypes examined.