Short hairpin RNA-mediated inhibition of HSV-1 gene expression and function during HSV-1 infection in Vero cells.
OBJECTIVE
To evaluate the efficiency of 3 short hairpin RNA (shRNA) interfering with the herpes simplex virus type 1 (HSV-1) gene coding glycoprotein D (gD) for inhibiting the gD expression and virus replication in vitro.


METHODS
Vero cells were selected for an in vitro model of infection. Three shRNA sequences (shRNAgD1, -gD2, and -gD3) targeting specifically the gD gene of HSV-1 were selected for evaluating the antiviral effects. The antiviral effects of shRNA in the cells infected with HSV-1 were evaluated by cytopathic effect (CPE) observations and plaque assays. The transcription level of viral RNA and the gD expression were studied by RT-PCR, Western blotting, and flow cytometry.


RESULTS
With the 3 shRNA at a final concentration of 120 nmol/L, a significant inhibition of CPE in the HSV-1-infected cells was observed. The ED50 of shRNA-gD1, gD2, and gD3 were 48.74+/-2.57, 57.13+/-3.24, and 114.64+/-5.12 nmol/L, respectively. The gD gene decreased significantly after viral infection in the Vero cells pretreated with shRNA compared to the virus group. The expressions of the gD protein, determined by Western blotting and flow cytometry, were also drastically decreased in shRNA-transfected cells.


CONCLUSIONS
Exogenous shRNA molecules can suppress the HSV-1 gD expression. They are inhibitors of HSV replication during infection in Vero cells.
Liu
Yuan-yuan
YY
State Key Laboratory of Virology, Institute of Medical Virology, School of Medicine, Wuhan University, Wuhan 430071, China.
Deng
Hai-ying
HY
Yang
Guang
G
Jiang
Wen-lin
WL
Grossin
Laurent
L
Yang
Zhan-qiu
ZQ
D000818:Animals; D002522:Chlorocebus aethiops; D003588:Cytopathogenic Effect, Viral / Q000187:drug effects; D015967:Gene Expression Regulation, Viral / Q000235:genetics / Q000502:physiology*; D018259:Herpesvirus 1, Human / Q000235:genetics* / Q000502:physiology*; D055029:Inverted Repeat Sequences / Q000235:genetics; D012367:RNA, Viral / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000096:biosynthesis / Q000235:genetics; D014779:Virus Replication / Q000187:drug effects
D012367:RNA, Viral; D014759:Viral Envelope Proteins; C039447:glycoprotein D, Human herpesvirus 1