High throughput sequencing revolution reveals conserved fundamentals of U‐indel editing

Abstract

Among Euglenozoans, mitochondrial RNA editing occurs in the diplonemids and in the kinetoplastids that include parasitic trypanosomes. Yet U-indel editing, in which open reading frames (ORFs) on mRNAs are generated by insertion and deletion of uridylates in locations dictated by guide RNAs, appears confined to kinetoplastids. The nature of guide RNA and edited mRNA populations has been cursorily explored in a surprisingly extensive number of species over the years, although complete sets of fully edited mRNAs for most kinetoplast genomes are largely missing. Now, however, high throughput sequencing technologies have had an enormous impact on what we know and will learn about the mechanisms, benefits, and final edited products of U-indel editing. Tools including PARERS, TREAT, and T-Aligner function to organize and make sense of U-indel mRNA transcriptomes, which are comprised of mRNAs harboring uridylate indels both consistent and inconsistent with translatable products. From high throughput sequencing data come arguments that partially edited mRNAs containing "junction regions" of noncanonical editing are editing intermediates, and conversely, arguments that they are dead-end products. These data have also revealed that the percent of a given transcript population that is fully or partially edited varies dramatically between transcripts and organisms. Outstanding questions that are being addressed include the prevalence of sequences that apparently encode alternative ORFs, diversity of editing events in ORF termini and 5' and 3' untranslated regions, and the differences that exist in this byzantine process between species. High throughput sequencing technologies will also undoubtedly be harnessed to probe U-indel editing's evolutionary origins. This article is categorized under: RNA Processing > RNA Editing and Modification RNA Evolution and Genomics > Computational Analyses of RNA.

Intrinsic and regulated properties of minimally edited trypanosome mRNAs

“…In general, IPSs may be bounded on their 5′ ends by either pre-edited or non-canonically edited junction sequence. Junctions are variable edited sequences that match neither pre-edited nor fully edited sequence and are present in partially edited mRNAs between the 3′ fully edited and 5′ pre-edited regions (31). The abundant presence of junction sequences in numerous kinetoplastid species has been known for more than two decades and was recently confirmed by high-throughput methods (8,32–36).…”

MRB10130 is a RESC assembly factor that promotes kinetoplastid RNA editing initiation and progression

“…A novel and extreme type of RNA editing occurs in the mitochondria of kinetoplastid flagellates, entailing specific insertion and deletion of uridylate (U) residues to correct frame shifts and create translatable open reading frames (ORFs) (Hashimi et al 2013;Aphasizheva and Aphasizhev 2016;Read et al 2016;Cruz-Reyes et al 2018;Zimmer et al 2018). U insertion/deletion (U-indel) editing is essential for survival of these organisms, which include several deadly human pathogens (Stuart et al 2008), because fully edited mRNAs encode proteins involved in mitochondrial bioenergetics and translation.…”

“…Junctions are regions of variable lengths that are edited to a sequence other than the final edited sequence. At least some junctions likely represent regions of active editing that will undergo iterative corrections until the final edited sequence is achieved and editing can progress in a 5 ′ direction (Simpson et al 2017;Zimmer et al 2018). Indeed, two components of the editing machinery were recently shown to be essential for both junction formation and 3 ′ to 5 ′ editing progression, suggesting a link between the two (Simpson et al 2017).…”

Developmental regulation of edited CYb and COIII mitochondrial mRNAs is achieved by distinct mechanisms in Trypanosoma brucei

“…An editing stop site is the 5′-most ES of continuous and uninterrupted canonical editing; thus, every ES that is 3′ of the editing stop site is also canonically edited ( 42 ). The region 5′ of the editing stop site can match the pre-edited sequence, a junction sequence in the process of active editing, or an alternative edited sequence that does not match the published canonical edited sequence ( 31 ).…”

“…Several scientific advances of the past decade permit us to now return to this intriguing question with a new arsenal of approaches. These advances include the HTS revolution that provided comprehensive gRNA libraries and several bioinformatic approaches to analyze mitochondrial mRNA populations ( 31 , 42 , 61 , 62 , 79–81 ), thereby allowing us to gain single nucleotide level insight into changes in editing progression during the life cycle. A second critical advance is the ability to trigger metacyclogenesis in PCF in vitro by overexpression of a single RNA-binding protein ( 66 ), which allowed us to perform the first analysis of mRNA editing throughout the T. brucei life cycle, including EMF and MCF stages.…”