PIWIL3 forms a complex with TDRKH in mammalian oocytes

PIWIs are crucial guardians of genome integrity, particularly in germ cells. While mammalian PIWIs have been primarily studied in mouse and rat, a homologue for the human PIWIL3 geneis absent in the Muridae family, and hence the unique function of PIWIL3 in germ cells cannot be effectively modeled by mouse knockouts. Herein, we investigated the expression,distribution and interaction of PIWIL3 in bovine oocytes. We localized PIWIL3 to mitochondria, and demonstrated that PIWIL3 expression is stringently controlled both spatially and temporally before and after fertilization. Moreover, we identified PIWIL3 in a mitochondrial-recruited three-membered complex with TDRKH and PNLDC1, and demonstrated by mutagenesis that PIWIL3 N-terminal arginine modifications are required for complex assembly. Finally we sequenced the piRNAs bound to PIWIL3-TDRKH-PNLDC1 and report here that about 50% of these piRNAs map to transposable elements, recapitulating the important role of PIWIL3 in maintaining genome integrity in mammalian oocytes. Small RNA sequencing from GV stage bovine oocytes (Tdrkh co-Immunoprecipitation vs. Input)