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

Author:

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

Abstract

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.

Publisher

Cold Spring Harbor Laboratory

Subject

Genetics(clinical),Genetics

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