Double-stranded RNA is internalized by scavenger receptor-mediated endocytosis in Drosophila S2 cells.
Double-stranded RNA (dsRNA) fragments are readily internalized and processed by Drosophila S2 cells, making these cells a widely used tool for the analysis of gene function by gene silencing through RNA interference (RNAi). The underlying mechanisms are insufficiently understood. To identify components of the RNAi pathway in S2 cells, we developed a screen based on rescue from RNAi-induced lethality. We identified Argonaute 2, a core component of the RNAi machinery, and three gene products previously unknown to be involved in RNAi in Drosophila: DEAD-box RNA helicase Belle, 26 S proteasome regulatory subunit 8 (Pros45), and clathrin heavy chain, a component of the endocytic machinery. Blocking endocytosis in S2 cells impaired RNAi, suggesting that dsRNA fragments are internalized by receptor-mediated endocytosis. Indeed, using a candidate gene approach, we identified two Drosophila scavenger receptors, SR-CI and Eater, which together accounted for more than 90% of the dsRNA uptake into S2 cells. When expressed in mammalian cells, SR-CI was sufficient to mediate internalization of dsRNA fragments. Our data provide insight into the mechanism of dsRNA internalization by Drosophila cells. These results have implications for dsRNA delivery into mammalian cells.
Department of Pediatrics and Biocenter Oulu, University of Oulu, FIN-90014, Oulu, Finland.
Johanna
J
Ulvila
Mataleena
M
Parikka
Anni
A
Kleino
Raija
R
Sormunen
R Alan
RA
Ezekowitz
Christine
C
Kocks
Mika
M
Rämet
D000818:Animals; D002460:Cell Line; D004331:Drosophila melanogaster; D004705:Endocytosis; D015723:Gene Library; D020868:Gene Silencing; D005821:Genetic Techniques; D008156:Luciferases / Q000378:metabolism; D034622:RNA Interference; D012330:RNA, Double-Stranded / Q000737:chemistry*; D051116:Receptors, Scavenger / Q000378:metabolism*
D012330:RNA, Double-Stranded; D051116:Receptors, Scavenger; D008156:Luciferases