MDV-encoded protein kinase US3 phosphorylates WTAP to inhibit transcriptomic m6A modification and cellular protein translation

Marek's disease virus (MDV)-encoded U_S3 is a highly conserved serine/threonine protein kinase in alpha-herpesviruses. In other alpha-herpesviruses, such as pseudorabies virus (PRV), U_S3 phosphorylates the N6-methyladenosine (m⁶A) methyltransferase Wilms tumor 1-associated protein (WTAP), inhibiting m⁶A modification. However, the role and mechanism of U_S3-mediated WTAP phosphorylation during MDV infection remain undefined. Our study revealed that MDV infection in vitro does not alter WTAP expression, while significant changes in WTAP expression occur during the MDV life cycle in vivo. We demonstrated that MDV-encoded U_S3 interacts with and co-localizes with WTAP in the nucleus. Further analysis showed that U_S3 binds to WTAP's C-terminal domain and phosphorylates WTAP at S273, S305, S314, and S375. Notably, the interaction between U_S3 and WTAP does not affect WTAP stability but inhibits transcriptomic m⁶A modification and cellular protein translation. Therefore, these findings enhance our understanding of the molecular mechanisms underlying MDV infection.