title: A PPR-DYW protein is required for splicing of a group II intron of cox1 pre-mRNA in Physcomitrella patens.
abstract: The pentatricopeptide repeat (PPR) protein family is involved in various steps of RNA metabolism in plastids and mitochondria. To investigate the function of a DYW sub-class PPR protein in the moss Physcomitrella patens, we constructed and characterized knockout mutants of the PpPPR_43 gene, which encodes a mitochondrial localized PPR protein with a C-terminal DYW domain. The disruptants showed poor growth of moss protonemata. To investigate whether mitochondrial transcripts were affected by disruption of PpPPR_43, we sequenced the cDNA to detect RNA editing events and performed RT-PCR analyses to measure steady-state mitochondrial transcript levels. Disruption of PpPPR_43 did not result in defective RNA editing, but a substantial reduction in the level of mature cox1 transcript was observed in the disruptants. RT-PCR analysis showed that the 3rd intron of cox1 pre-mRNA was not spliced out in the disruptants, but the 1st, 2nd and 4th introns were efficiently spliced out. This suggests that PpPPR_43 is an intron 3-specific splicing factor. The role of the C-terminal domains of PpPPR_43 in intron 3 splicing was analyzed by complementation experiments with truncated constructs lacking the DYW domain or both the E and DYW domains. Both truncated genes completely restored splicing in the PpPPR_43 knockout mutant. This indicates that the E and DYW domains of PpPPR_43 are not required for splicing, and can be deleted without loss of cox1 intron 3 splicing.
authors: Center for Gene Research, Nagoya University, Chikusa-Ku, Nagoya 468-8602, Japan.
authors: Mizuho
authors: M
authors: Ichinose
authors: Eiji
authors: E
authors: Tasaki
authors: Chieko
authors: C
authors: Sugita
authors: Mamoru
authors: M
authors: Sugita
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D015152:Blotting, Northern; D019068:Bryopsida / Q000235:genetics* / Q000378:metabolism; D003576:Electron Transport Complex IV / Q000235:genetics*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D007438:Introns / Q000235:genetics*; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D010940:Plant Proteins / Q000235:genetics* / Q000378:metabolism; D021122:Protein Subunits / Q000235:genetics; D012313:RNA / Q000235:genetics / Q000378:metabolism; D012322:RNA Precursors / Q000235:genetics* / Q000378:metabolism; D012326:RNA Splicing; D000077278:RNA, Mitochondrial; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D024101:Mitochondrial Proteins; D010940:Plant Proteins; D021122:Protein Subunits; D012322:RNA Precursors; D000077278:RNA, Mitochondrial; D049452:Green Fluorescent Proteins; D012313:RNA; D003576:Electron Transport Complex IV