Abstract
The baculovirus Autographa californica multiple nucleopolyhedrovirus (AcMNPV) possesses a gene, ac-pcna or ac49, which encodes a protein with similarity to proliferating cell nuclear antigen (PCNA). Homologs of this gene code for DNA polymerase processivity factors and are essential in the DNA replication systems. But the function of ac-pcna still remains unclear. To define the function of Ac-pcna in AcMNPV and Sf-pcna in host Sf9 cells, Bac-to-Bac baculovirus expression system was used to generate two recombinant baculoviruses: AcMNPV-Ac-pcna-EGFP and AcMNPV-Sf-pcna-EGFP. Results indicated that AcMNPV-mediated overexpression of Ac-PCNA and Sf-PCNA could stimulate replication of AcMNPV genome in the host Sf9 cells. Meanwhile, either AcMNPV-Ac-pcna-EGFP or AcMNPV-Sf-pcna-EGFP had a significant stimulating effect on Sf9 genome replication during infection. We also found that Ac-PCNA and Sf-PCNA could promote the production of budded virus. Ac-PCNA could improve the transcription level of ie2 gene dramatically and further improved the transcription of late gene, for example 38 K and vp39, at 12 h p.i.. Moreover, insecticidal potency test showed that the larvae of Beet armyworm in the AcMNPV-Ac-pcna-EGFP and AcMNPV-Sf-pcna-EGFP groups had a higher mortality rate (83.33 and 91.67%), a lower pupation rate (16.67 and 8.33%), and a lower emergence rate (6.67 and 3.33%), compared with those in AcMNPV-EGFP group. The function of Ac-PCNA and Sf-PCNA was confirmed in this study, which provided the theoretical foundation for using and modifying AcMNPV.
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Acknowledgements
This project was supported by grants from ‘National Natural Science Foundation of China (No.31272100 and 31372199)’, ‘Natural Science Foundation of Shanxi Province (No.2014011038-1)’.
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Supplementary Table 1
Sequence of primers used to amplify Ac-pcna, Sf-pcna, hr3, hsp90 (DOC 26 kb)
Supplementary Table 2
Sequence of primers used to amplify β-actin, 38K, vp39, ie2 in qPCR (DOC 27 kb)
Supplementary Table 3
Average weight of Beet armyworm larvae (g) (DOC 35 kb)
Supplementary Table 4
Mortality rate analysis of Beet armyworm larvae (DOC 45 kb)
Supplementary Table 5
Pupation rate analysis of Beet armyworm larvae (%) (DOC 33 kb)
Supplementary Table 6
Emergence rate analysis of Beet armyworm larvae (%) (DOC 33 kb)
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Fu, Y., Wang, R. & Liang, A. Function analysis of Ac-PCNA and Sf-PCNA during the Autographa californica multiple nucleopolyhedrovirus infection process. Mol Cell Biochem 443, 57–68 (2018). https://doi.org/10.1007/s11010-017-3210-y
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DOI: https://doi.org/10.1007/s11010-017-3210-y