Gene Loss and Error-Prone RNA Editing in the Mitochondrion of Perkinsela , an Endosymbiotic Kinetoplastid

Abstract

Perkinsela is an enigmatic early-branching kinetoplastid protist that lives as an obligate endosymbiont inside Paramoeba (Amoebozoa). We have sequenced the highly reduced mitochondrial genome of Perkinsela, which possesses only six protein-coding genes (cox1, cox2, cox3, cob, atp6, and rps12), despite the fact that the organelle itself contains more DNA than is present in either the host or endosymbiont nuclear genomes. An in silico analysis of two Perkinsela strains showed that mitochondrial RNA editing and processing machineries typical of kinetoplastid flagellates are generally conserved, and all mitochondrial transcripts undergo U-insertion/deletion editing. Canonical kinetoplastid mitochondrial ribosomes are also present. We have developed software tools for accurate and exhaustive mapping of transcriptome sequencing (RNA-seq) reads with extensive U-insertions/deletions, which allows detailed investigation of RNA editing via deep sequencing. With these methods, we show that up to 50% of reads for a given edited region contain errors of the editing system or, less likely, correspond to alternatively edited transcripts.

Massive mitochondrial DNA content in diplonemid and kinetoplastid protists

“…These post‐transcriptional events are specified by hundreds of minicircle‐encoded guide (g) RNAs, which along with numerous large protein complexes produce a mere dozen mitochondrial proteins . Recent genomic and transcriptomic evidence confirms earlier predictions that uridine‐indel RNA editing is present in all Kinetoplastea , with the notable exception of petite mutants of T. brucei , who lack the mitochondrial genome as such .…”

“…Electron microscopic studies from almost 50 years ago revealed that some bodonid flagellates contain huge amounts of kinetoplast DNA. However, their size and coding capacity has only recently started to become clear . Only direct comparison with sequenced nuclear genomes, as carried out here, allows estimation of their size.…”

“…The last examined kinetoplastid was the early branching Perkinsela sp. DAPI staining and electron microscopy revealed that this aflagellar endosymbiont contains significantly more DNA in its inflated mitochondrion than in its small nucleus . The nuclear genome of Perkinsela was recently sequenced and found to be ~9.5 Mbp in size (diploid); with only 5,252 genes, it is the most reduced genome among this group of protists .…”

“…Its structure also varies, ranging from free circles and small catenanes to long linear or circular molecules. Invariably, however, the bodonids carry much more DNA in their mitochondria than do the trypanosomatids .…”

Unlocking the biological potential ofEuglena gracilis: evolution, cell biology and significance to parasitism

“…We find no evidence for RNA-editing in E. gracilis, nor for the TAC, both of which are consistent with the presence of a mitochondrial genome composed of only short linear DNA molecules (encoding only seven different proteins) and a conventional mitochondrial mRNA transcription system . Specifically, only 14 of 51 proteins involved in RNA-editing in T. brucei (David, et al, 2015) had reciprocal best BLAST ! 13…”