title: 3' end formation of PIWI-interacting RNAs in vitro.
abstract: PIWI-interacting RNAs (piRNAs) are 23-30 nucleotides small RNAs that act with PIWI proteins to silence transposon activity in animal gonads. In contrast to microRNAs and small interfering RNAs, the biogenesis of piRNAs, including how 3' ends are formed, remains largely unknown. Here, by using lysate from BmN4, a silkworm ovary-derived cell line, we have developed a cell-free system that recapitulates key steps of piRNA biogenesis: loading of long single-stranded precursor RNAs into PIWI proteins with 5'-nucleotide bias, followed by Mg(2+)-dependent 3' to 5' exonucleolytic trimming and 2'-O-methylation at 3' ends. Importantly, 3' end methylation is tightly coupled with trimming and yet is not a prerequisite for determining the mature piRNA length. Our system provides a biochemical framework for dissecting piRNA biogenesis.
authors.affiliation: Department of Agricultural and Environmental Biology, Graduate School of Agricultural and Life Sciences, The University of Tokyo, Yayoi 1-1-1, Bunkyo-ku, Tokyo 113-8657, Japan.
authors.forename: Shinpei
authors.initials: S
authors.lastname: Kawaoka
authors.forename: Natsuko
authors.initials: N
authors.lastname: Izumi
authors.forename: Susumu
authors.initials: S
authors.lastname: Katsuma
authors.forename: Yukihide
authors.initials: Y
authors.lastname: Tomari
mesh_terms: D000818:Animals; D012831:Bombyx / Q000235:genetics*; D002460:Cell Line; D019476:Insect Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D026721:RNA 3' End Processing; D034741:RNA, Small Interfering / Q000378:metabolism*
chemical_list: D019476:Insect Proteins; D034741:RNA, Small Interfering