Authors

G Amegatcher¹; A Ivens¹; K R Matthews¹; A Schnaufer¹;  ¹ Institute of Immunology & Infection Research, University of Edinburgh, UK

Discussion

In organisms such as yeast, mammals and plants, ‘mitochondrial retrograde signalling’ pathways, including the ‘unfolded protein response (UPRmt), convey information on the functional status of this organelle to the nucleus and modulate expression of nuclear genes accordingly1,2,3. Although successful completion of the life cycle of Trypanosoma brucei depends on stringent regulation of mitochondrial activity, it is not known if similar signalling pathways exist in these parasites. As T. brucei differentiates from the slender bloodstream form via the transmission competent stumpy form to the procyclic insect form it undergoes dramatic remodelling of its morphology and metabolism, including mitochondrial activity. As a first step towards exploring potential retrograde signalling pathways in Trypanosoma brucei we have compared the transcriptome of strain EATRO 1125 (AnTat1.1 90:134) with a genetically engineered derivate devoid of mitochondrial DNA (akinetoplastic, or AK cells), before and after differentiation from slender to stumpy forms. Our main findings are: (i) In control (‘WT’) and AK parasites, 242 and 433 genes, respectively, showed significant upregulation in stumpy forms (we considered a difference of  2-fold with a p-value  0.05 as significant). 102 upregulated genes were shared, including genes previously reported to be upregulated in stumpy cells such as PIP39 (Tb927.9.6090), dihydrolipoyl dehydrogenase (Tb927.11.16730) and EP1 procyclin (Tb927.10.10260). The upregulation of other procyclins was much less pronounced in AK stumpy cells compared to WT stumpy cells. On the other hand, 10 hypothetical proteins showed more than 4-fold upregulation exclusively in stumpy AK cells. (ii) Genes involved in the glycolytic pathway were generally downregulated in stumpy cells. We also observed robust downregulation in stumpy cells of numerous histones and of two genes involved in kDNA maintenance, mitochondrial DNA ligase LIG k alpha (Tb927.7.610) and cysteine peptidase PNT1 (Tb927.11.6550). (iii) When we compared slender AK vs. WT cells we observed only 27 robust changes (either up or down), which mostly concerned genes annotated as ‘pseudogene’, or ‘hypothetical unlikely’, including a putative UDP-Gal or UDP-GlcNAc-dependent glycosyltransferase pseudogene that was ~5-fold increased in AK slender cells. The same mRNA was ~3-fold increased in AK stumpy cells compared to WT stumpy cells. Other changes in AK stumpy cells concerned a hypothetical protein (~3-fold upregulated) and a putative adenylosuccinate lyase (~3-fold downregulated), but overall we observed only a limited number of robust changes (13 up, 9 down), including the decreased levels of procyclin mRNAs mentioned above. In summary, our transcriptomic studies suggest that absence of the mitochondrial genome has a surprisingly limited effect on levels of nuclearly encoded mRNAs in bloodstream stage T. brucei