Gene silencing in Echinococcus multilocularis protoscoleces using RNA interference.
We investigated the potential of gene silencing in Echinococcus multilocularis protoscoleces using RNA interference (RNAi). For the introduction of siRNA, soaking and electroporation were first examined for their effects on the viability of protoscoleces and their efficacy for siRNA introduction. Consequently, electroporation using 100 V and 800 μF showed the optimal results. This electroporation procedure was then evaluated for its ability to induce RNAi in protoscoleces using siRNAs targeting the 14-3-3 and elp genes. It was found that the levels of 14-3-3 and elp mRNA in 14-3-3 siRNA- and elp siRNA-treated protoscoleces were reduced to 21.8 ± 2.6 and 35.5 ± 0.4% of those of the untreated control by day 3, respectively. Moreover, the target proteins significantly decreased in the siRNA-treated samples by day 15. In the analysis of viability, the untreated control, electroporation control, 14-3-3 siRNA-treated, and elp siRNA-treated samples displayed 98.4 ± 1.4, 83.0 ± 2.5, 58.0 ± 23.0, and 55.1 ± 14.6% viability, respectively, on day 15. In conclusion, we successfully demonstrated that RNAi mediated the knock-down of target gene expression in E. multilocularis protoscoleces at both the transcriptional and translational levels.
Mizukami
Chiaki
C
Laboratory of Parasitology, Department of Disease Control, Graduate School of Veterinary Medicine, Hokkaido University, Sapporo 060-0818, Japan.
Spiliotis
Markus
M
Gottstein
Bruno
B
Yagi
Kinpei
K
Katakura
Ken
K
Oku
Yuzaburo
Y
D048948:14-3-3 Proteins / Q000235:genetics / Q000378:metabolism; D000818:Animals; D000947:Antigens, Helminth / Q000235:genetics / Q000378:metabolism; D000954:Antigens, Surface / Q000235:genetics / Q000378:metabolism; D048210:Echinococcus multilocularis / Q000235:genetics / Q000254:growth & development / Q000378:metabolism* / Q000648:ultrastructure; D018274:Electroporation; D015870:Gene Expression / Q000187:drug effects; D020868:Gene Silencing; D015801:Helminth Proteins / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism*
D048948:14-3-3 Proteins; D000947:Antigens, Helminth; D000954:Antigens, Surface; C072149:Elp protein, Echinococcus multilocularis; D015801:Helminth Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering