PAD1 quantification data.

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.