Coronavirus PEDV nucleocapsid protein interacts with p53 to induce cell cycle arrest in S-phase and promotes viral replication.

Su, Mingjun; Shi, Da; Xing, Xiaoxu; Qi, Shanshan; Yang, Dan; Zhang, Jiyu; Han, Yuru; Zhu, Qinghe; Sun, Haibo; Wang, Xiaoran; Wu, Haoyang; Wang, Meijiao; Wei, Shan; Li, Chunqiu; Guo, Donghua; Feng, Li; Sun, Dongbo

Subversion of the host cell cycle to facilitate viral replication is a common feature of coronavirus infections. Coronavirus nucleocapsid (N) protein could modulate host cell cycle, but the mechanistic details remain largely unknown. Here, we investigated manipulation of porcine epidemic diarrhea virus (PEDV) N protein on cell cycle and its influence on viral replication. Results indicated that PEDV N-induced Vero E6 cell cycle arrest at S-phase, which promoted viral replication (P < 0.05). S-phase arrest was dependent on N protein nuclear localization signal S71NWHFYYLGTGPHADLRYRT90 and interaction between N protein and p53. In the nucleus, the binding of N protein to p53 maintained consistently high-level expression of p53, which activated p53-DREAM pathway. The key domain of the N protein interacting with p53 was revealed to be S171RGNSQNRGNNQGRGASQNRGGNN194 (NS171-N194), in which G183RG185 are core sites. NS171-N194 and G183RG185 were essential for N-induced S-phase arrest. Moreover, small molecular drugs targeting the NS171-N194 domain of PEDV N protein were screened through molecular docking. Hyperoside could antagonize N protein-induced S-phase arrest by interfering with interaction between N protein and p53 and inhibit viral replication (P < 0.05). The above experiments were also validated in porcine intestinal cells, and resulting data were in line with that of Vero E6 cells. Therefore, these results revealed that PEDV N protein interacted with p53 to activate p53-DREAM pathway, and subsequently induced S-phase arrest to create a favorable environment for virus replication. These findings provided new insight into the PEDV-host interaction and the design of novel antiviral strategies against PEDV.