Developmental dynamics of mitochondrial mRNA abundance and editing reveal roles for temperature and the differentiation-repressive kinase RDK1 in cytochrome oxidase subunit II mRNA editing

Developmental regulation of mitochondrial uridine insertion/deletion editing in Trypanosoma brucei is necessary to modulate parasite metabolism as it shifts from dependence on glycolysis for ATP production in the mammalian bloodstream form (BSF) to oxidative phosphorylation in the insect procyclic form (PCF). However, the timing and stimuli that regulate mRNA editing have been poorly characterized. Here, we utilized a pleomorphic T. brucei strain and quantitative RT-PCR and droplet digital PCR analyses to evaluate the changes in total mRNA abundance and editing as parasites progressively differentiate from slender BSF to PCF and investigate the effect of individual stimuli on mitochondrial gene expression. We observed little change during the slender-to-stumpy BSF transition. Rather, we found that mainly the mitochondrial cytochrome (COI, COII, COIII, and CYb) mRNAs are upregulated within 24 h after stumpy BSF is stimulated to differentiate to PCF in vitro and during in vivo tsetse fly infections. Temperature reduction from 37°C to 27°C is a critical factor for increasing the editing of COII and COIII mRNAs and COIV protein expression but not the editing of CYb mRNA or RISP protein expression. We further demonstrate that the depletion of the differentiationrepressive kinase RDK1 couples with temperature reduction to stimulate COII mRNA editing, and the accessory factor p22 is required for the cold-responsive upregulation of COII mRNA editing. Overall, we show that cytochrome mRNAs are regulated during development by distinct stimuli through a variety of methods to increase their abundance and/or editing.