Exploring the activity and the essentiality of the putative Delta6-desaturase in the procyclic and bloodstream forms of Trypanosoma brucei

Abstract

AbstractTrypanosomatids have been shown to possess an exclusive and finely regulated biosynthetic pathway for de novo synthesis of fatty acids (FAs) and particularly of polyunsaturated fatty acids (PUFAs). The key enzymes for the process of unsaturation are known as desaturases. In this work, we explored the biocatalytic activity of the putative {Delta}6-desaturase (Tb11.v5.0580) in the native organism T. brucei. Utilising fatty acid analysis via GC-MS, we were able to elucidate via genetic manipulation of the level of expression of {Delta}6-desaturases in both procyclic (PCF) and bloodstream (BSF) forms of T. brucei and via supplementation of the media with various levels of FA sources, that docosahexaenoic acid (22:6) and/or docosapentaenoic acid (22:5), and arachidonic acid (20:4) and/or docosatetraenoic acid (22:4) are the products and the substrates respectively of this {Delta}6-desaturases. Interestingly, we were able to observe, via lipidomic analysis with ESI-MS/MS, an increase in inositol-phosphoryl ceramide (IPC) in response to the overexpression of {Delta}6-desaturases in low-fat media, both in PCF and rather surprisingly in BSF. The formation of IPC is normally only observed in the stumpy and procyclic forms of T. brucei. Therefore, the expression levels of {Delta}6-desaturases, which varies between BSF and PCF, might be involved in the cascade(s) of metabolic events that cause these remodelling of the lipid pools and ultimately morphological changes, which are key to the transition between these life-cycle stages. Author summaryTrypanosoma brucei is a unicellular parasite that causes human and animal African trypanosomiasis. These parasites have the special ability to make their own pool of fat molecules by assembling and modifying the fatty acid building blocks that they take up from the human and animal hosts and from the insect vector. In this study, we investigated the unknown activity of a desaturase enzyme. By modulating its activity, we showed that it can make different levels of high-value long chain polyunsaturated fatty acids (LC-PUFAs) often known as omega-6 and omega-3. If we increase or reduce the fat sources available from the outer environment, the cells respond by making more or less LC-PUFAs and by forming different type of lipids and sphingolipids for their cellular membranes. We highlighted that by tuning the level of activity of the desaturase and varying the type and amounts of fat sources available to the cells, T. brucei can alter their morphology. This is key for the parasites to adapt to the various environments and the nutrients therein that are often constantly changing within the host, allowing the shift between different life-stages during the complex life cycle from the insect vector to the host and back.