Uridine insertion/deletion editing activities

The uridine nucleotide insertion and deletion editing of trypanosomatid mitochondrial mRNAs is catalyzed by a macromolecular complex, the editosome. Many investigations of RNA editing involve some assessment of editosome activity either in vitro or in vivo. Assays to detect insertion or deletion editing activity on RNAs in vitro have been particularly useful, and can include the initial endonucleolytic step (full-round) or bypass it (precleaved). Additional assays to examine individual catalytic steps have also proved useful to dissect particular steps in editing. Detection of RNA editing activity in vivo has been significantly advanced by the application of real-time PCR technology, which can simultaneously assay several edited and pre-edited targets. Here we describe these assays to assess editing both in vitro (full-round insertion and deletion; precleaved insertion and deletion; individual TUTase, ligase, or helicase activity) and in vivo (real-time PCR).