Project description:Trypanosoma brucei gambiense is the causative agent of the fatal human disease African sleeping sickness. Here we have compared the transcriptome of two different life cycle stages, the potentially human-infective bloodstream form and the non-human-infective procyclic stage, using digital gene expression (DGE) analysis.
Project description:Mitochondrial metabolic remodeling is a hallmark of the Trypanosoma brucei digenetic life cycle since the insect stage utilizes the cost-effective oxidative phosphorylation to generate ATP, while bloodstream cells switch to less energetically efficient aerobic glycolysis. Due to difficulties in acquiring enough parasites from the tsetse fly vector for biochemical analysis, the dynamics of the parasite´s mitochondrial metabolic rewiring in the vector have remained obscure. Here, we took advantage of in vitro-induced differentiation to follow changes at the RNA levels.
Project description:Trypanosoma brucei gambiense is the causative agent of the fatal human disease African sleeping sickness. Using Digital Gene Expression we have compared the transcriptome of a group 1 T.b.gambiense (Eliane) and a T.b.brucei (STIB 247).
Project description:Some organisms like the human and animal parasite Trypanosoma brucei add a leader sequence to their mRNAs through a reaction called trans-splicing. Until now the splice sites for most mRNAs were unknown in T. brucei. Using high throughput sequencing we have developed a method to identify the splice sites and at the same time measure the abundance of the corresponding mRNAs. Analyzing three different life cycle stages of the parasite we identified the vast majority of splice sites in the organism and, to our great surprise, uncovered more than 2500 alternative splicing events, many of which appeared to be specific for one of the life cycle stages. Alternative splicing is a result of the addition of the leader sequence to different positions on the mRNA, leading to mixed mRNA populations that can encode for proteins with varying properties. One of the most obvious changes caused by alternative splicing is the gain or loss of targeting signals, leading to differential localization of the corresponding proteins. Based on our findings we hypothesize that alternative splicing is a major mechanism to regulate gene expression in T. brucei and could contribute to protein diversity in the parasite.
Project description:The host range of African trypanosomes is influenced by innate protective molecules in the blood of primates. A subfraction of human high-density lipoprotein (HDL) containing apolipoprotein A-I, apolipoprotein L-I, and haptoglobin-related protein is toxic to Trypanosoma brucei brucei but not the human sleeping sickness parasite Trypanosoma brucei rhodesiense. It is thought that T. b. rhodesiense evolved from a T. b. brucei-like ancestor and expresses a defense protein that ablates the antitrypanosomal activity of human HDL. To directly investigate this possibility, we developed an in vitro selection to generate human HDL-resistant T. b. brucei. Here we show that conversion of T. b. brucei from human HDL sensitive to resistant correlates with changes in the expression of the variant surface glycoprotein (VSG) and abolished uptake of the cytotoxic human HDLs. Complete transcriptome analysis of the HDL-susceptible and -resistant trypanosomes confirmed that VSG switching had occurred but failed to reveal the expression of other genes specifically associated with human HDL resistance, including the serum resistance-associated gene (SRA) of T. b. rhodesiense. In addition, we found that while the original active expression site was still utilized, expression of three expression site-associated genes (ESAG) was altered in the HDL-resistant trypanosomes. These findings demonstrate that resistance to human HDLs can be acquired by T. b. brucei. Keywords: Trypanosoma, VSG, antigenic switching, HDL-resistance
