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Modification of the second translation initiation site restricts the replication of foot-and-mouth disease virus in PK-15 cells
- Source :
- Applied Microbiology and Biotechnology
- Publication Year :
- 2020
- Publisher :
- Springer Science and Business Media LLC, 2020.
-
Abstract
- Abstract The translation initiation of foot-and-mouth disease virus (FMDV) occurs at two alternative initiation sites (Lab AUG and Lb AUG). Usually, the Lb AUG is more favorably used to initiate protein synthesis than the Lab AUG. To explore the effect of Lb AUG on FMDV replication and obtain FMDV with restricted replication, this initiation codon was mutated to a variety of non-AUG codons (UGG, AUC, CUG, and AAA). Fortunately, the modifications did not prevent viral viability but influenced replication characteristics of some FMDV mutants in a cell-specific manner, as was shown by the similar replication in BHK-21 cells and delayed growth kinetics in PK-15 cells. This attenuated phenotype of FMDV mutants in PK-15 cells was found to be correlated with reduced abilities to cleave eIF4GI and suppress interference (IFN) expression. As leader (L) protein was reported to be responsible for eIF4GI cleavage and inhibition of IFN expression, the in vivo L protein synthesis was examined during the infection of FMDV mutants. Our results showed that not only the total yield of L proteins was severely influenced but also the individual yield of L protein was seen to be affected, which implied that both the relative usage of the two initiation sites and overall translation efficiency were changed by Lb AUG modifications. In addition, the in vitro translation activity was also negatively regulated by Lb AUG mutations. Collectively, these findings suggested that the restricted replications of Lb AUG-modified FMDVs were related to the delayed eIF4GI cleavage and decreased ability to block IFN expression but were mainly determined by the inefficient translation initiation. FMDVs precisely with modifications of Lb AUG initiation codon may represent safer seed viruses for vaccine production. Key points • The polyprotein translation of FMDV initiates at two alternative initiation sites (Lab AUG and Lb AUG). In order to explore the effect of Lb AUG on FMDV replication and obtain FMDV with restricted replication, the Lb initiation AUG was mutated to a variety of non-AUG codons (UGG, AUC, CUG, and AAA), and four FMDV mutants with Lb AUG modification were generated. • We found that partial FMDV mutants grew almost as well as WT virus in BHK-21 cells, a typical cell line used for FMD vaccine production, but displayed impaired replication in IFN-competent PK-15 cells. • The attenuation of mutant FMDVs in PK-15 cells was found to be correlated with delayed eIF4GI cleavage and decreased ability to block IFN expression. • We proved that the attenuated phenotype of Lb AUG-modified FMDVs was mainly determined by the inefficient translation initiation, as demonstrated by the decrease of total yield of L proteins and individual production of L protein. • We successfully generated genetically engineered FMDV with attenuated phenotype. The approach of precise engineering of FMDV with the modification of initiation codon provides a safe platform to produce inactivated antigen vaccines.
- Subjects :
- Virus replication
viruses
Mutant
Codon, Initiator
Applied Microbiology and Biotechnology
Virus
Cell Line
FMDV
03 medical and health sciences
Eukaryotic translation
Start codon
Translation initiation
Protein biosynthesis
Animals
030304 developmental biology
0303 health sciences
biology
030306 microbiology
General Medicine
biology.organism_classification
Virology
Applied Microbial and Cell Physiology
Viral replication
Foot-and-Mouth Disease Virus
Cell culture
Foot-and-Mouth Disease
Leader protein
Foot-and-mouth disease virus
Protein Processing, Post-Translational
Vaccine
Biotechnology
Subjects
Details
- ISSN :
- 14320614 and 01757598
- Volume :
- 104
- Database :
- OpenAIRE
- Journal :
- Applied Microbiology and Biotechnology
- Accession number :
- edsair.doi.dedup.....8fec781fd42347f107b9ccb8e6e0e915
- Full Text :
- https://doi.org/10.1007/s00253-020-10810-w