Targeted gene silencing in mouse germ cells by insertion of a homologous DNA into a piRNA generating locus

• Published in: Genome research 2013-02, Vol.23 (2), p.292-299
• Main Authors: Yamamoto, Yasuhiro, Watanabe, Toshiaki, Hoki, Yuko, Shirane, Kenjiro, Li, Yufeng, Ichiiyanagi, Kenji, Kuramochi-Miyagawa, Satomi, Toyoda, Atsushi, Fujiyama, Asao, Oginuma, Masayuki, Suzuki, Hitomi, Sado, Takashi, Nakano, Toru, Sasaki, Hiroyuki
• Format: Article
• Language: English
• Subjects: Animals; DNA - genetics; Gene Expression Regulation; Gene Silencing; Gene Targeting; Genes, Reporter; Genetic Loci; Germ Cells - metabolism; Male; Mice; Mice, Transgenic; RNA Processing, Post-Transcriptional; RNA, Small Interfering - genetics
• ISSN: 1088-9051; 1549-5469
• DOI: 10.1101/gr.137224.112

In germ cells, early embryos, and stem cells of animals, PIWI-interacting RNAs (piRNAs) have an important role in silencing retrotransposons, which are vicious genomic parasites, through transcriptional and post-transcriptional mechanisms. To examine whether the piRNA pathway can be used to silence genes of interest in germ cells, we have generated knock-in mice in which a foreign DNA fragment was inserted into a region generating pachytene piRNAs. The knock-in sequence was transcribed, and the resulting RNA was processed to yield piRNAs in postnatal testes. When reporter genes possessing a sequence complementary to portions of the knock-in sequence were introduced, they were greatly repressed after the time of pachytene piRNA generation. This repression mainly occurred at the post-transcriptional level, as degradation of the reporter RNAs was accelerated. Our results show that the piRNA pathway can be used as a tool for sequence-specific gene silencing in germ cells and support the idea that the piRNA generating regions serve as traps for retrotransposons, enabling the host cell to generate piRNAs against active retrotransposons.