title: Electroporation of DNA into Physarum polycephalum Mitochondria: Effects on Transcription and RNA Editing in Isolated Organelles.
abstract: Mitochondrial RNAs in the acellular slime moldPhysarum polycephalumcontain nucleotides that are not encoded in the mitochondrial genes from which they are transcribed. These site-specific changes are quite extensive, comprising ~4% of the residues within mRNAs and ~2% of rRNAs and tRNAs. These "extra" nucleotides are added co-transcriptionally, but the means by which this is accomplished have not been elucidated. Thecox1mRNA also contains four sites of C to U changes, which occur post-transcriptionally, most likely via targeted deamination. The currently available in vitro systems for studyingP. polycephalumediting are limited in that the template is the entire ~63,000 bp mitochondrial genome. This presents a significant challenge when trying to define the signals that specify editing sites. In an attempt to overcome this issue, a method for introducing DNA into isolatedP. polycephalummitochondria via electroporation has been developed. Exogenous DNA is expressed, but the transcripts synthesized from these templates are not edited under the conditions tested. However, transcripts derived from the mitochondrial genome are accurately edited after electroporation, indicating that the editing machinery is still functional. These findings suggest that this method may ultimately provide a feasible approach to elucidating editing signals.
authors.affiliation: Center for RNA Molecular Biology, Case Western Reserve University, Cleveland, OH 44120, USA. jmg13@case.edu.
authors.forename: Jonatha M
authors.initials: JM
authors.lastname: Gott
authors.affiliation: Center for RNA Molecular Biology, Case Western Reserve University, Cleveland, OH 44120, USA. gmnaegele@gmail.com.
authors.forename: Gregory M
authors.initials: GM
authors.lastname: Naegele
authors.affiliation: Visual Sciences Research Center, Case Western Reserve University, Cleveland, OH 44120, USA. sjh36@case.edu.
authors.forename: Scott J
authors.initials: SJ
authors.lastname: Howell
keywords: Physarum polycephalum; RNA editing; electroporation; mitochondria