The piRNA pathway is an RNA-silencing pathway involving small non-coding RNAs and PIWI proteins.This pathway was discovered a decade ago as a defense mechanism against transposons. Since then, this field has experienced rapid progress. Recent studies have implicated piRNAs and PIWI proteins in a broad range of functions, including epigenetic and post-transcriptional gene regulation, genome rearrangements, germ cell development, and stem cell biology. The aim of this EMBO Workshop is to discuss the latest findings in all aspects of piRNA biology. We will address piRNA biogenesis; the role of piRNAs/PIWI proteins in genome defense; epigenetic regulation; gene regulation; genome rearrangements; germ cell development and fertility; stem cell biology; somatic functions; and cancers. We have invited the leading specialists in the field, who have expressed their enthusiasm for the workshop. We have organized the first and second EMBO Workshops, on "Multiple functions of piRNAs and PIWI proteins" and “piRNAs and PIWI proteins”, respectively, in Montpellier in 2016 and 2018. Both workshops were big success and numerous participants suggested to continue this workshop as a biannual series. Without any doubt, this EMBO workshop would have a strong impact on the piRNA field and beyond.