[Objective]Normally,mature small RNAs(sRNAs)mainly reside in cytoplasmic loci.We supposed that shuttling these small RNAs intentionally from cytoplasm to nucleus may shift their roles from post-transcriptional gene scilencing(PTGS)to transcriptional gene scilencing(TGS)and this may present a promising new RNAi strategy against viral infection.However,delivery of DNA or siRNA with nuclear targeting has often been limited by the nuclear transport mechanisms.Here,we described an Argonaute1 protein-based gene delivery system to facilitate nuclear translocation of siRNAs with an extremely high efficiency.[Methods]First,we interrogated the distributions and movements of chemical synthetic sRNAs labeled with Cy3 or FAM fluorochrome in Hela cells after transfection.Second,we untilized the innate sRNAs biogenesis pathway to form the effector complexes through incorporation of siRNAs and the transformed Argonaute1 protein(AGO1)in vivo.Western blot(WB)and indirect immuno-fluorescence(IIF)assays were performed to confirm the silencing effect and colocalizations among core components of the protein complexes,respectively.[Results]We found that exogenous sRNAs appeared in lysosomes in early phase,afterwards,localized in processing bodies(p-bodies).Those sRNAs were confined in local regions and displayed distinct motion modes with respect to their locations.It was obvious that sRNAs were solely distributed in the cytoplasm rather than in the nucleus.However,after we pre-transfected a designed AGO1construct,most of the sRNAs were strikingly redistributed to the nucleus.As we expected,the gene silencing of PTGS was alleviated,which was quite well confirmed by WB and IIF assays.[Conclusion]Compared with conventional siRNA-based gene therapy targeting messager RNAs(mRNAs),delivery of transcription start element-associated siRNAs by way of TGS may minimize off-target effects during gene silencing.However,it remains unclear whether siRNAs delivered into the nucleus maintained in an active state by routine methods.The approach we reported here efficiently overcame the barrier of siRNA delivery to the nucleus.Most important of all,siRNAs were loaded into effector complexes successfully.Further work will be necessary to evaluate the TGS effect based on viral infection models such as AIDS and influenza.