Exploring the activity of the putative Δ6-desaturase and its role in bloodstream form life-cycle transitions in Trypanosoma brucei

TL;DR

This study explores how a desaturase enzyme in the parasite Trypanosoma brucei affects fatty acid production and life-cycle transitions between hosts.

Contribution

The paper identifies the specific substrates and products of the Δ6-desaturase and links its activity to lipid remodeling and morphological changes during life-cycle transitions.

Findings

Δ6-desaturase produces 22:6 and 22:5 from 20:4 and 22:4 in T. brucei.

Overexpression of Δ6-desaturase increases inositol-phosphoryl ceramide in bloodstream forms.

Desaturase activity is linked to differentiation proteins and lipid changes during life-cycle transitions.

Abstract

Trypanosomatids 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 Δ6-desaturase (Tb11.v5.0580) in the native organism T. brucei, whose expression level varies dramatically between life cycle stages. Utilising FA analysis via GC-MS, we were able to elucidate i) via genetic manipulation of the level of expression of Δ6-desaturases in both procyclic (PCF) and bloodstream (BSF) forms of T. brucei and ii) via supplementation of the media with various levels of FA sources, that docosahexaenoic acid (22:6) and/or docosapentaenoic acid (22:5) are the products, while arachidonic acid (20:4) and/or docosatetraenoic acid (22:4) are the substrates of this Δ6-desaturase. Surprisingly, 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 Δ6-desaturase in low-fat media in BSF. The formation of IPC is normally only observed in the stumpy and procyclic forms of T. brucei. Therefore, the expression levels of Δ6-desaturases, which increases between BSF, stumpy and PCF, might be involved in the cascade(s) of metabolic events that contributes to these remodelling of the lipid pools and ultimately morphological changes, which are key to the transition between these life-cycle stages. We were in fact able to show that the overexpression of Δ6-desaturase is indeed linked to the expression of protein associated with differentiation (PAD1) in stumpy, and of the upregulation of some proteins and metabolites which are normally upregulated in stumpy and PCF.

Trypanosoma brucei is a unicellular parasite that causes human and animal African trypanosomiasis, which is transmitted by the bite of the Tsetse fly. These parasites have the special ability to make their own pool of fatty molecules by assembling and modifying the fatty acid building blocks that they take up from their mammalian and insect hosts. 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 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 environment, the cells respond by making i) more or less LC-PUFAs and ii) by forming different type of lipids and sphingolipids for their cellular membranes. We highlighted that by tuning the level of activity of the desaturase enzyme 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 they are exposed to when changing between hosts. Hence, these important shifts between different life-cycle stages get the parasites ready and adapted for survival as they go from a mammalian host to an insect vector and back again.

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