title: DNA interference: DNA-induced gene silencing in the appendicularian Oikopleura dioica.
abstract: RNA interference is widely employed as a gene-silencing system in eukaryotes for host defence against invading nucleic acids. In response to invading double-stranded RNA (dsRNA), mRNA is degraded in sequence-specific manner. So far, however, DNA interference (DNAi) has been reported only in plants, ciliates and archaea, and has not been explored in Metazoa. Here, we demonstrate that linear double-stranded DNA promotes both sequence-specific transcription blocking and mRNA degradation in developing embryos of the appendicularian Oikopleura dioica. Introduced polymerase chain reaction (PCR) products or linearized plasmids encoding Brachyury induced tail malformation and mRNA degradation. This malformation was also promoted by DNA fragments of the putative 5'-flanking region and intron without the coding region. PCR products encoding Zic-like1 and acetylcholine esterase also induced loss of sensory organ and muscle acetylcholinesterase activity, respectively. Co-injection of mRNA encoding EGFP and mCherry, and PCR products encoding these fluorescent proteins, induced sequence-specific decrease in the green or red fluorescence, respectively. These results suggest that O. dioica possesses a defence system against exogenous DNA and RNA, and that DNA fragment-induced gene silencing would be mediated through transcription blocking as well as mRNA degradation. This is the first report of DNAi in Metazoa.
authors.affiliation: Department of Biological Sciences, Graduate School of Science, Osaka University, 1-1 Machikaneyama-cho, Toyonaka, Osaka 560-0043, Japan otamazako@bio.sci.osaka-u.ac.jp.
authors.forename: Tatsuya
authors.initials: T
authors.lastname: Omotezako
authors.affiliation: Department of Biological Sciences, Graduate School of Science, Osaka University, 1-1 Machikaneyama-cho, Toyonaka, Osaka 560-0043, Japan.
authors.forename: Takeshi A
authors.initials: TA
authors.lastname: Onuma
authors.affiliation: Department of Biological Sciences, Graduate School of Science, Osaka University, 1-1 Machikaneyama-cho, Toyonaka, Osaka 560-0043, Japan.
authors.forename: Hiroki
authors.initials: H
authors.lastname: Nishida
mesh_terms: D000110:Acetylcholinesterase / Q000235:genetics / Q000378:metabolism; D000818:Animals; D004247:DNA / Q000235:genetics*; D005326:Fetal Proteins / Q000235:genetics* / Q000378:metabolism; D020868:Gene Silencing; D007438:Introns; D008969:Molecular Sequence Data; D009132:Muscles / Q000201:enzymology; D012333:RNA, Messenger / Q000378:metabolism; D012679:Sense Organs / Q000201:enzymology; D020825:T-Box Domain Proteins / Q000235:genetics* / Q000378:metabolism; D013623:Tail / Q000002:abnormalities; D014561:Urochordata / Q000196:embryology / Q000235:genetics*
chemical_list: D005326:Fetal Proteins; D012333:RNA, Messenger; D020825:T-Box Domain Proteins; D004247:DNA; D000110:Acetylcholinesterase
keywords: Brachyury; DNA interference; Oikopleura dioica; chordate; post-transcriptional gene silencing; transcriptional gene silencing