tdrd5Pachytene piRNAs are the most abundant piRNAs in mammalian adult testes. They aregenerated from long precursor transcripts by the primary piRNA biogenesis pathway but thefactors involved in pachytene piRNA precursors processing are poorly understood. Here weshow that the Tudor domain-containing 5 (TDRD5) protein is essential for pachytene piRNAbiogenesis in mice. Conditional inactivation of TDRD5 in mouse postnatal germ cells revealsthat TDRD5 selectively regulates the production of pachytene piRNAs from abundant piRNA-producing precursors, with little effect on low-abundant piRNAs. Unexpectedly, TDRD5 is notrequired for the 5′end processing of the precursors, but is crucial for promoting production ofpiRNAs from the other regions of the transcript. Furthermore, we show that TDRD5 is anRNA-binding protein directly associating with piRNA precursors. These observationsestablish TDRD5 as a piRNA biogenesis factor and reveal two genetically separable steps atthe start of pachytene piRNA processing

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