The insect-phase gRNA transcriptome in Trypanosoma brucei

One of the most striking examples of small RNA regulation of gene expression is the process of RNA editing in the mitochondria of trypanosomes. In these parasites, RNA editing involves extensive uridylate insertions and deletions within most of the mitochondrial messenger RNAs (mRNAs). Over 1200 small guide RNAs (gRNAs) are predicted to be responsible for directing the sequence changes that create start and stop codons, correct frameshifts and for many of the mRNAs generate most of the open reading frame. In addition, alternative editing creates the opportunity for unprecedented protein diversity. In Trypanosoma brucei, the vast majority of gRNAs are transcribed from minicircles, which are approximately one kilobase in size, and encode between three and four gRNAs. The large number (5000-10,000) and their concatenated structure make them difficult to sequence. To identify the complete set of gRNAs necessary for mRNA editing in T. brucei, we used Illumina deep sequencing of purified gRNAs from the procyclic stage. We report a near complete set of gRNAs needed to direct the editing of the mRNAs.