Establishment Of A Stable IFITM-Expressing HeLa Cell Line And The Inhibitory Effects Of IFITM On CA16Virus Replication

Background: Hand, foot, and mouth disease（HFMD）is a common viral illness amonginfants and young children, with clinical characterization of prodromal fever followed bypharyngitis, mouth ulcers, and a rash on the hands and feet. Acute infection may also causesevere neurological disease such as encephalitis and meningitis that leads to significantmortality. Clinically, so far, no prophylactic or therapeutic method is available to treat HFMD.IFITM is an interferon stimulated protein that restricted many virus replication. These urgentissues and the potential for an HFMD pandemic in the future prompte us to exploit a moreeffective approach to combat CA16virus.Methods: At first, plasmid pTet-on was transfected and sifted by G418to generateHeLa-tet cell line. Then, each pTRE2-IFITM plasmid companied with pTK-Hyg plasmid wascotransfected into positive clones followed by hygromycin B selection to generateHeLa/Tet/pTRE2-IFITM1/2/3cells. The positive cell line was verified by Western Blottinganalysis to detect the expression of IFITM1/2/3proteins after doxycycline treatment.Results: Controllable IFITM-expressing HeLa cell line was infected with differentmultiplicity of infection (MOI) of CA16virus, then the level of virus RNA was detected byreal-time quantitative PCR (RT-qPCR) to determine the inhibitory effect of each IFITMprotein. The results demonstrated that IFITM proteins displayed strong restriction effect onCA16virus replication especially IFITM3.Conclusions: This study established the IFITM1/2/3controllable expressing HeLa cellline and provided suitable cell model for further study in the function and antiviral mechanismof IFITM proteins.