Cyprinid herpesvirus 2 (CyHV‐2), the etiological agent of herpesvirus haematopoietic necrosis (HVHN) in carp and goldfish, has caused significant economic losses in the aquaculture industry. During viral infection, the host initiates a series of active or passive defences to regulate the process of virus infection. Apoptosis is a key component of active cellular defence, and members of the Bcl‐2 family have been shown to play a critical role in the apoptotic process. However, the mechanism of action of the Bcl‐2 family in inducing apoptosis during CyHV‐2 infection remains unclear. In this study, we revealed the molecular mechanism of miRNA‐mediated silver crucian carp BAX (ccBax) in CyHV‐2‐induced apoptosis for the first time and demonstrated that the overexpression of miR‐124 suppressed ccBax expression and significantly down‐regulated apoptosis in caudal fin cells of Carassius auratus gibelio (GiCF), while miR‐124 inhibitors were the opposite. These studies indicated that miR‐124 inhibits CyHV‐2‐induced apoptosis by reducing the expression of ccBax. Furthermore, the fact that transfection of miR‐124 mimics promoted CyHV‐2 replication, whereas miR‐124 inhibitors inhibited CyHV‐2 replication, indicated that miR‐124 inhibited CyHV‐2‐induced apoptosis and contributed to viral replication. All these results suggested that miR‐124 suppresses virus‐induced apoptosis and promotes viral replication by targeting and regulating ccBax expression.

IntroductionRibonucleotide reductase (RR) is essential for the replication of the double-stranded DNA virus CyHV-2 due to its ability to catalyze the conversion of ribonucleotides to deoxyribonucleotides, and is a potential target for the development of antiviral drugs to control CyHV-2 infection.MethodsBioinformatic analysis was conducted to identify potential homologues of RR in CyHV-2. The transcription and translation levels of ORF23 and ORF141, which showed high homology to RR, were measured during CyHV-2 replication in GICF. Co-localization experiments and immunoprecipitation were performed to investigate the interaction between ORF23 and ORF141. siRNA interference experiments were conducted to evaluate the effect of silencing ORF23 and ORF141 on CyHV-2 replication. The inhibitory effect of hydroxyurea, a nucleotide reductase inhibitor, on CyHV-2 replication in GICF cells and RR enzymatic activity in vitro was also evaluated.ResultsORF23 and ORF141 were identified as potential viral ribonucleotide reductase homologues in CyHV-2, and their transcription and translation levels increased with CyHV-2 replication. Co-localization experiments and immunoprecipitation suggested an interaction between the two proteins. Simultaneous silencing of ORF23 and ORF141 effectively inhibited the replication of CyHV-2. Additionally, hydroxyurea inhibited the replication of CyHV-2 in GICF cells and the in vitro enzymatic activity of RR.ConclusionThese results suggest that the CyHV-2 proteins ORF23 and ORF141 function as viral ribonucleotide reductase and their function makes an effect to CyHV-2 replication. Targeting ribonucleotide reductase could be a crucial strategy for developing new antiviral drugs against CyHV-2 and other herpesviruses.