Correlation of the cyclin A expression level with porcine circovirus type 2 propagation efficiency
Tóm tắt
Porcine circovirus type 2 (PCV2) is an important pathogen in swine, and it is assumed that PCV2 replication is cell cycle dependent (especially during S phase). However, the cellular molecules that regulate PCV2 replication have not been fully identified. Here, we cloned the porcine cyclin A (CycA) and CDK2 genes, the major regulators of the S phase, and established CycA or CDK2 overexpression and lower-expression cell lines. The propagation efficiency of strains PCV2a/CL or PCV2b/YJ in these cell lines was investigated using a capture enzyme-linked immunosorbent assay (ELISA) or an immunoperoxidase monolayer assay (IPMA), and the cell cycle was analyzed by flow cytometry. The results showed that CycA overexpression suppressed PCV2 replication. In contrast, CycA down-regulation by shRNA induced increases during the S and G2/M phases and resulted in increased PCV2 propagation. In contrast, overexpression or lower expression of CDK2 exhibited no significant influence on PCV2 replication. Furthermore, the subcellular localization of the PCV2 replicase protein (Rep) and capsid protein (Cap), CycA, and CDK2 in PK-15 cells was analyzed by confocal microscopy. The results showed that overexpression of CycA, rather than CDK2, altered normal nuclear localization of PCV2-Rep, which was transferred to the cytoplasm. In conclusion, PCV2 replication is both S- and G2/M-phase dependent and CycA, is an important regulator of the PCV2 life cycle.
Tài liệu tham khảo
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