[Targeted inhibition of Rabies virus gene expression by a chimeric multidomain protein mediated shRNA delivery].
OBJECTIVE
In this study, a new chimeric protein SEG expressed in previous work was applied to evaluate its translocating efficiency of shRNA to rabies virus infected cells in mice, meanwhile, the capability of anti-rabies virus was investigated.


METHODS
Rabies virus strain CVS-24 was inoculated into the hind leg to establish a mouse model of rabies in a dose of 50 LD₅₀; 12 h thereafter the mice were injected intravenously with shRNA-producing plasmid mixed with SEG. To test shRNA delivery, single-cell suspensions from brain, spleen and liver were examined by flow cytometry. Rabies virus in brain tissue of mice was detected by qRT-PCR, RT-PCR, western blot and directed immunofluorescence assay. Mice were monitored for survival and serum samples were tested for IFN-α levels.


RESULTS
No green fluorescent protein (GFP) was seen in the spleen or liver, suggesting that SEG allows specific targeting of RV-infected cells. RT-PCR and western blot showed that mice treated with SEG-shRNA had lower rabies virus RNA and protein levels than the controls. Real-time PCR showed that rabies virus was reduced 4.88 fold compared to the mock cells. Survival of RV-infected mouse showed a significant protection from rabies virus infection by SEG-shRNA treatment. The survival was up to 50% whereas the control group all died. IFN was not induced in SEG-shRNA treated animals.


CONCLUSIONS
shRNA-producing plasmid was specifically delivered into rabies virus infected cells using the SEG protein, and effectively inhibited rabies virus geneexpression and replication in vivo. SEG-shRNA can be used for adjuvant treatment for rabies.
Ruimei
R
Yang
Hualei
H
Wang
Hu
H
Shan
Songtao
S
Yang
Xianzhu
X
Xia
D000818:Animals; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016503:Drug Delivery Systems; D015967:Gene Expression Regulation, Viral; D006801:Humans; D051379:Mice; D017434:Protein Structure, Tertiary; D034741:RNA, Small Interfering / Q000235:genetics* / Q000378:metabolism / Q000627:therapeutic use; D011818:Rabies / Q000628:therapy* / Q000821:virology*; D011820:Rabies virus / Q000235:genetics* / Q000502:physiology; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014157:Transcription Factors / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D004268:DNA-Binding Proteins; C052130:GAL4 protein, S cerevisiae; D034741:RNA, Small Interfering; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors