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PIWI-interacting RNAs (piRNAs) repress transposons to protect the germline genome from DNA damage caused by transposon transposition. In  Drosophila , the  Traffic jam  ( Tj ) mRNA is consumed to produce piRNA in its 3′-UTR. A  cis  element located within the 3′-UTR,  Tj-cis , is necessary for piRNA biogenesis. In this study, we analyzed the structure of the  Tj-cis  RNA, a 100-nt RNA corresponding to the  Tj-cis  element, by the SHAPE and NMR analyses and found that a stable hairpin structure formed in the 5′ half of the  Tj-cis  RNA. The tertiary structure of the 16-nt stable hairpin was analyzed by NMR, and a novel stem–loop structure, the T-hairpin, was found. In the T-hairpin, four uridine residues are exposed to the solvent, suggesting that this stem–loop is the target of Yb protein, a Tudor domain-containing piRNA biogenesis factor. The piRNA biogenesis assay showed that both the T-hairpin and the 3′ half are required for the function of the  Tj-cis  element, suggesting that both the T-hairpin and the 3′ half are recognized by Yb protein.

T-hairpin structure found in the RNA element involved in piRNA biogenesis