A Slicer-Mediated Mechanism for Repeat-Associated siRNA 5' End Formation in Drosophila

• DOI: 10.1126/science.1140494
• Publication Date: 2007-02-23T01:57:12Z
• Authors: Lalith S. Gunawar...; Kuniaki Saito; Kazumichi M. Nishida; Keita Miyoshi; Yoshinori Kawamura; Tomoko Nagami; Haruhiko Siomi; Mikiko C. Siomi
• Keywords: Male; 0303 health sciences; Models, Genetic; Retroelements; Recombinant Fusion Proteins; Molecular Sequence Data; Ovary; Proteins; 03 medical and health sciences; Drosophila melanogaster; Peptide Initiation Factors; Argonaute Proteins; Testis; Animals; Drosophila Proteins; RNA-Induced Silencing Complex; Female; RNA Interference; Amino Acid Sequence; RNA, Small Interfering; Gene Library

In Drosophila , repeat-associated small interfering RNAs (rasiRNAs) are produced in the germ line by a Dicer-independent pathway and function through the PIWI subfamily of Argonautes to ensure silencing of retrotransposons. We sequenced small RNAs associated with the PIWI subfamily member AGO3. Although other members of PIWI, Aubergine (Aub) and Piwi, associated with rasiRNAs derived mainly from the antisense strand of retrotransposons, AGO3-associated rasiRNAs arose mainly from the sense strand. Aub- and Piwi-associated rasiRNAs showed a strong preference for uracil at their 5′ ends, and AGO3-associated rasiRNAs showed a strong preference for adenine at nucleotide 10. Comparisons between AGO3- and Aub-associated rasiRNAs revealed pairs of rasiRNAs showing complementarities in their first 10 nucleotides. Aub and AGO3 exhibited Slicer activity in vitro. These data support a model in which formation of a 5′ terminus within rasiRNA precursors is guided by rasiRNAs originating from transcripts of the other strand in concert with the Slicer activity of PIWI.