The Host Protein CAD Regulates the Replication of FMDV through the Function of Pyrimidines' De Novo Synthesis

Foot-and-mouth disease virus (FMDV) is a single-stranded picornavirus that causes economically devastating disease in even-hooved animals. There has been little research on the function of host cells during FMDV infection. We aimed to shed light on key host factors associated with FMDV replication during acute infection. We found that HDAC1 overexpression in host cells induced upregulation of FMDV RNA and protein levels. Activation of the AKT-mammalian target of rapamycin (mTOR) signaling pathway using bpV(HOpic) or SC79 also promoted FMDV replication. Furthermore, short hairpin RNA (shRNA)-induced suppression of carbamoyl-phosphate synthetase 2, aspartate transcarbamylase, and dihydroorotase (CAD), a transcription factor downstream of the AKT-mTOR signaling pathway, resulted in downregulation of FMDV RNA and protein levels. Coimmunoprecipitation assays showed that the ACTase domain of CAD could interact with the FMDV 2C protein, suggesting that the ACTase domain of CAD may be critical in FMDV replication. CAD proteins participate in pyrimidine synthesis. Inhibition of FMDV replication by deletion of the ACTase domain of CAD in host cells could be reversed by supplementation with uracil. These results revealed that the contribution of the CAD ACTase domain to FMDV replication is dependent on pyrimidine synthesis. Our research shows that HDAC1 promotes FMDV replication by regulating pyrimidine synthesis from CAD via the AKT-mTOR signaling pathway. Foot-and-mouth disease virus is an animal virus of the family that seriously harms the development of animal husbandry and foreign trade of related products, and there is still a lack of effective means to control its harm. Replication complexes would generate during FMDV replication to ensure efficient replication cycles. 2C is a common viral protein in the replication complex of virus, which is thought to be an essential component of membrane rearrangement and viral replication complex formation. The host protein CAD is a key protein in the pyrimidines synthesis. In our research, the interaction of CAD and FMDV 2C was demonstrated in FMDV-infected BHK-21 cells, and it colocalized with 2C in the replication complex. The inhibition of the expression of FMDV 3D protein through interference with CAD and supplementation with exogenous pyrimidines reversed this inhibition, suggesting that FMDV might recruit CAD through the 2C protein to ensure pyrimidine supply during replication. In addition, we also found that FMDV