Safeguarding spermatogenesis from retrotransposon insertions by forming ecDNA.

Retrotransposon mobilization in germline cells enables the rewriting of genetic information to drive genome innovation, species evolution, and adaptation through the generation of de novo mutations. However, uncontrolled mobilization can cause DNA breaks and genome instability, often leading to sterility. How germ cells balance retrotransposon-induced genome innovation with the need for genomic integrity remains poorly understood. Here, we used Drosophila spermatogenesis as a model to investigate retrotransposon mobilization dynamics. Although many retrotransposon families are transcriptionally active, we found that the LTR-retrotransposon nomad completes the full mobilization cascade-including mRNA export, protein translation, and reverse transcription-to produce double-stranded DNA (dsDNA) the most efficiently. Strikingly, despite successfully generating dsDNA, nomad rarely achieves genomic reintegration. Instead, its newly synthesized DNA predominantly forms extrachromosomal circular DNA (ecDNA). These findings suggest that ecDNA formation acts as a protective mechanism to sequester retrotransposon-derived DNA and prevent widespread genomic integration during spermatogenesis, thereby preserving genome stability while allowing limited retrotransposon activity.