title: Organelle trafficking of chimeric ribozymes and genetic manipulation of mitochondria.
abstract: With the expansion of the RNA world, antisense strategies have become widespread to manipulate nuclear gene expression but organelle genetic systems have remained aside. The present work opens the field to mitochondria. We demonstrate that customized RNAs expressed from a nuclear transgene and driven by a transfer RNA-like (tRNA-like) moiety are taken up by mitochondria in plant cells. The process appears to follow the natural tRNA import specificity, suggesting that translocation indeed occurs through the regular tRNA uptake pathway. Upon validation of the strategy with a reporter sequence, we developed a chimeric catalytic RNA composed of a specially designed trans-cleaving hammerhead ribozyme and a tRNA mimic. Organelle import of the chimeric ribozyme and specific target cleavage within mitochondria were demonstrated in transgenic tobacco cell cultures and Arabidopsis thaliana plants, providing the first directed knockdown of a mitochondrial RNA in a multicellular eukaryote. Further observations point to mitochondrial messenger RNA control mechanisms related to the plant developmental stage and culture conditions. Transformation of mitochondria is only accessible in yeast and in the unicellular alga Chlamydomonas. Based on the widespread tRNA import pathway, our data thus make a breakthrough for direct investigation and manipulation of mitochondrial genetics.
authors.affiliation: Institut de Biologie Moléculaire des Plantes, CNRS and Université de Strasbourg, 12 rue du Général Zimmer, 67084 Strasbourg, France.
authors.forename: Romain
authors.initials: R
authors.lastname: Val
authors.forename: Eliza
authors.initials: E
authors.lastname: Wyszko
authors.forename: Clarisse
authors.initials: C
authors.lastname: Valentin
authors.forename: Maciej
authors.initials: M
authors.lastname: Szymanski
authors.forename: Anne
authors.initials: A
authors.lastname: Cosset
authors.forename: Malek
authors.initials: M
authors.lastname: Alioua
authors.forename: Theo W
authors.initials: TW
authors.lastname: Dreher
authors.forename: Jan
authors.initials: J
authors.lastname: Barciszewski
authors.forename: André
authors.initials: A
authors.lastname: Dietrich
mesh_terms: D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D005818:Genetic Engineering; D008928:Mitochondria / Q000378:metabolism*; D008969:Molecular Sequence Data; D012313:RNA / Q000378:metabolism; D034443:RNA Transport; D016337:RNA, Catalytic / Q000737:chemistry / Q000378:metabolism*; D000077278:RNA, Mitochondrial; D012343:RNA, Transfer / Q000737:chemistry / Q000378:metabolism; D014026:Nicotiana / Q000235:genetics / Q000378:metabolism
chemical_list: D016337:RNA, Catalytic; D000077278:RNA, Mitochondrial; C427204:hammerhead ribozyme; D012313:RNA; D012343:RNA, Transfer