Introduction
The giant grouper (Epinephelus lanceolatus) is a cornerstone of Taiwan's aquaculture industry; however, infection by nervous necrosis virus (NNV) during the larval and juvenile stages results in mortality rates as high as 90%. Consequently, elucidating the host immune mechanisms following viral infection is critical for developing effective preventive strategies. Myxovirus resistance (Mx) proteins are key downstream effectors of the type I interferon (IFN) system. Although our previous research demonstrated that ggMx1,2, 3 mRNA levels are regulated by NNV, it remains unclear whether these three Mx isoforms possess distinct temporal roles in inhibiting NNV replication, and the precise regulatory mechanisms of their respective proteins have yet to be fully clarified. In this study, we characterized the distribution and dynamic shifts of the three ggMx protein under viral stress.
Materials and Methods
GF-1 cells were utilized as the experimental model. The overexpression of ggMx1, 2, and 3 protein were characterized under NNV challenge. Subcellular distribution and dynamic protein shifts were visualized using high-resolution confocal immunofluorescence imaging. To investigate physical interactions between the host immune factors and the pathogen, colocalization analysis was performed between Mx proteins and the NNV capsid protein.
Results
Confocal immunofluorescence imaging revealed the distribution and dynamic shifts of Mx proteins, demonstrating that following the overexpression of Mx1, 2, and 3, the peak suppression of NNV occurred sequentially, starting with Mx1 protein, followed by Mx2 protein, and finally Mx3 protein. Colocalization analysis confirmed physical interactions between Mx proteins and the NNV capsid protein, with peak colocalization observed at 30 hours for Mx1 and Mx2, and 36 hours for Mx3.
Discussion
This study provides novel insights into the temporal and spatial regulation of the innate immune response in giant grouper. The sequential peak suppression of NNV by different Mx isoforms suggests a coordinated, stage-specific antiviral defense strategy. The physical interaction and peak colocalization times indicate that Mx1 and Mx2 act earlier in the infection cycle compared to Mx3. Collectively, these findings elucidate the dual response mechanisms of ggMx to biological and viral stress, providing profound insights into the interaction of NNV and Mx and supporting the development in aquaculture.
Acknowledgment
This research was supported by the National Science and Technology Council (NSTC 112-2313-B-006-006-MY3 and NSTC 114-2312-B-006-002).