Physical and functional coupling of RNA-dependent RNA polymerase and Dicer in the biogenesis of endogenous siRNAs.
Many classes of small RNA (sRNA) involved in RNA silencing are generated by double-stranded RNA (dsRNA) processing. Although principles of sRNA biogenesis have emerged, newly identified classes of sRNAs have features that suggest additional biogenesis mechanisms. Tetrahymena thermophila expresses one such class, comprising sRNAs of 23 and 24 nucleotides (nt) with an absolute strand bias in accumulation. Here we demonstrate sRNA production by the T. thermophila Dicer Dcr2 and the RNA-dependent RNA polymerase Rdr1, which purifies as a multisubunit RNA-dependent RNA polymerase complex (RDRC). Dcr2 and RDRC interact, stimulating Dcr2 activity. Moreover, Dcr2 specificity is influenced by RDRC beyond this physical interaction, as Dcr2 generates discrete 23- and 24-nt sRNAs only from dsRNA with a 5'-triphosphate. These findings suggest that sRNA strand bias arises from Dcr2 processing polarity, conferred by physical and functional coupling of RDRC and Dicer enzymes.
Lee
Suzanne R
SR
Department of Molecular and Cell Biology, University of California - Berkeley, 142 Life Sciences Addition 3200, Berkeley, California 94720-3200, USA.
Collins
Kathleen
K
D000595:Amino Acid Sequence; D000818:Animals; D017125:Genes, Protozoan; D008969:Molecular Sequence Data; D015800:Protozoan Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D016053:RNA, Protozoan / Q000096:biosynthesis*; D034741:RNA, Small Interfering / Q000096:biosynthesis* / Q000737:chemistry; D012324:RNA-Dependent RNA Polymerase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D043244:Ribonuclease III / Q000737:chemistry* / Q000378:metabolism; D013379:Substrate Specificity; D016808:Tetrahymena thermophila / Q000201:enzymology*
D015800:Protozoan Proteins; D016053:RNA, Protozoan; D034741:RNA, Small Interfering; D012324:RNA-Dependent RNA Polymerase; D043244:Ribonuclease III