title: RNA-binding protein DUS16 plays an essential role in primary miRNA processing in the unicellular alga Chlamydomonas reinhardtii.
abstract: Canonical microRNAs (miRNAs) are embedded in duplexed stem-loops in long precursor transcripts and are excised by sequential cleavage by DICER nuclease(s). In this miRNA biogenesis pathway, dsRNA-binding proteins play important roles in animals and plants by assisting DICER. However, these RNA-binding proteins are poorly characterized in unicellular organisms. Here we report that a unique RNA-binding protein, Dull slicer-16 (DUS16), plays an essential role in processing of primary-miRNA (pri-miRNA) transcripts in the unicellular green alga Chlamydomonas reinhardtii In animals and plants, dsRNA-binding proteins involved in miRNA biogenesis harbor two or three dsRNA-binding domains (dsRBDs), whereas DUS16 contains one dsRBD and also an ssRNA-binding domain (RRM). The null mutant of DUS16 showed a drastic reduction in most miRNA species. Production of these miRNAs was complemented by expression of full-length DUS16, but the expression of RRM- or dsRBD-truncated DUS16 did not restore miRNA production. Furthermore, DUS16 is predominantly localized to the nucleus and associated with nascent (unspliced form) pri-miRNAs and the DICER-LIKE 3 protein. These results suggest that DUS16 recognizes pri-miRNA transcripts cotranscriptionally and promotes their processing into mature miRNAs as a component of a microprocessor complex. We propose that DUS16 is an essential factor for miRNA production in Chlamydomonas and, because DUS16 is functionally similar to the dsRNA-binding proteins involved in miRNA biogenesis in animals and land plants, our report provides insight into this mechanism in unicellular eukaryotes.
authors: Department of Environmental Systems Engineering, Kochi University of Technology, Kami, Kochi 782-8502, Japan; tomohito@nibb.ac.jp.
authors: Tomohito
authors: 0000-0001-9157-0209
authors: T
authors: Yamasaki
authors: Department of Genetics, Stanford University School of Medicine, Stanford, CA 94305;
authors: Masayuki
authors: M
authors: Onishi
authors: School of Biological Sciences, University of Nebraska-Lincoln, Lincoln, NE 68588; Center for Plant Science Innovation, University of Nebraska-Lincoln, Lincoln, NE 68588.
authors: Eun-Jeong
authors: EJ
authors: Kim
authors: School of Biological Sciences, University of Nebraska-Lincoln, Lincoln, NE 68588; Center for Plant Science Innovation, University of Nebraska-Lincoln, Lincoln, NE 68588.
authors: Heriberto
authors: H
authors: Cerutti
authors: Department of Environmental Systems Engineering, Kochi University of Technology, Kami, Kochi 782-8502, Japan;
authors: Takeshi
authors: T
authors: Ohama
mesh_terms: D020418:Algal Proteins / Q000235:genetics*; D029681:Arabidopsis Proteins / Q000235:genetics; D016825:Chlamydomonas reinhardtii / Q000235:genetics*; D004277:DNA, Single-Stranded / Q000235:genetics; D035683:MicroRNAs / Q000096:biosynthesis / Q000235:genetics*; D012323:RNA Processing, Post-Transcriptional / Q000235:genetics; D016601:RNA-Binding Proteins / Q000235:genetics*; D043244:Ribonuclease III / Q000235:genetics
chemical_list: D020418:Algal Proteins; D029681:Arabidopsis Proteins; D004277:DNA, Single-Stranded; D035683:MicroRNAs; D016601:RNA-Binding Proteins; C519654:DCL3 protein, Arabidopsis; D043244:Ribonuclease III
keywords: Chlamydomonas; dsRNA-binding protein; microRNA biogenesis; mutagenesis; small RNA-seq