Authors

M Hammond⁵; J Pyrih⁶; L Cadena⁴; M Svobodova⁴; A Alves⁷; S Dean⁸; JD Sunter¹; R Wheeler⁹; K Gull³; C Benz⁵; V Raskova⁴; I Durante⁴; J Lukeš²;  ¹ University of Oxford, UK;  ² Institute of Parasitology, Biology Centre, ASCR, Czech Republic;  ³ University of Oxford, Sir William Dunn School of Pathology, UK;  ⁴ Institute of Parasitology, BioCenter, České Budějovice, Czech Republic;  ⁵ Institute of Parasitology, Czech Academy of Sciences, České Budějovice, Czech Republic;  ⁶ Department of Biochemistry, University of Cambridge, UK;  ⁷ Oxford Brookes University, UK;  ⁸ Division of Biomedical Sciences, Warwick Medical School, UK;  ⁹ Peter Medawar Building for Pathogen Research, Nuffield Department of Medicine, University of Oxford, UK

Discussion

The TrypTag project represents a methodological approach to tag every protein encoded in the nuclear genome of Trypanosoma brucei. A specialised mitochondrial investigation of this study (MitoTag), revealed for the first time a fluorescent tag association with the concentrated mitochondrial DNA structure known as the kinetoplast, documented in several hundred mitochondrial proteins. Combined with transmembrane domain prediction, MitoTag enabled the sub-localisation of 1,053 mitochondrial proteins to this organelle's four compartments, circumventing the need for electron microscopy or other intensive sub-localisation methods. Furthermore, we demonstrate a method to distinguish genuine kinetoplast proteins from artificial tagging-induced associations among mitochondrial proteins, and accordingly demonstrate over a dozen novel kinetoplast components. From this, we expand the functions of the kinetoplast to metabolic pathways of dUMP synthesis and One Carbon metabolism. This constitutes the largest single expansion of the kinetoplast repertoire to date and represents an exciting development for a complex long considered an attractive drug target due to its clade-specific presence.