Visualizing a correlation between siRNA localization, cellular uptake, and RNAi in living cells.
RNA interference (RNAi) is the process by which short-interfering RNA (siRNA) target a specific mRNA for degradation through interactions with an RNA-induced silencing complex (RISC). Here, a clear correlation between siRNA localization, cellular uptake, and RNAi activity was discovered by delivering siRNA into cells using siRNA-TAT(47-57) peptide, siRNA-TAT(47-57)-derived oligocarbamate conjugates, or nanoparticles. For successful RNAi, the localization of siRNA was distinctly perinuclear, suggesting that siRNA is targeted to these regions for interactions with RISC to induce RNAi. siRNA sequence variation and the presence of the target mRNA apparently did not change the subcellular localization pattern of siRNA. Intriguingly, siRNA conjugated to TAT(47-57) peptide or TAT(47-57)-derived oligocarbamate resulted in efficient RNAi activity and perinuclear localization of siRNA that was distinctly different from nonconjugated free TAT peptide nucleolar localization. These results suggest that interactions with RISC dictate siRNA localization even when siRNA is conjugated to TAT(47-57) peptide.
Chiu
Ya-Lin
YL
Department of Biochemistry and Molecular Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01605, USA.
Ali
Akbar
A
Chu
Chia-Ying
CY
Cao
Hong
H
Rana
Tariq M
TM
D001483:Base Sequence; D001704:Biopolymers / Q000737:chemistry / Q000378:metabolism; D002219:Carbamates / Q000737:chemistry; D002470:Cell Survival; D042863:Cyclin-Dependent Kinase 9 / Q000235:genetics / Q000378:metabolism; D003593:Cytoplasm / Q000378:metabolism; D015696:Gene Products, tat / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D049329:Nanostructures; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D034443:RNA Transport; D034741:RNA, Small Interfering / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
D001704:Biopolymers; D002219:Carbamates; D015696:Gene Products, tat; D010446:Peptide Fragments; D034741:RNA, Small Interfering; C467969:CDK9 protein, human; D042863:Cyclin-Dependent Kinase 9