The disruption of GDP-fucose de novo biosynthesis suggests the presence of a novel fucose-containing glycoconjugate in Plasmodium asexual blood stages

Abstract

Glycosylation is an important posttranslational protein modification in all eukaryotes. Besides glycosylphosphatidylinositol (GPI) anchors and N-glycosylation, O-fucosylation has been recently reported in key sporozoite proteins of the malaria parasite. Previous analyses showed the presence of GDP-fucose (GDP-Fuc), the precursor for all fucosylation reactions, in the blood stages of Plasmodium falciparum. The GDP-Fuc de novo pathway, which requires the action of GDP-mannose 4,6-dehydratase (GMD) and GDP-L-fucose synthase (FS), is conserved in the parasite genome, but the importance of fucose metabolism for the parasite is unknown. To functionally characterize the pathway we generated a PfGMD mutant and analyzed its phenotype. Although the labelling by the fucose-binding Ulex europaeus agglutinin I (UEA-I) was completely abrogated, GDP-Fuc was still detected in the mutant. This unexpected result suggests the presence of an alternative mechanism for maintaining GDP-Fuc in the parasite. Furthermore, PfGMD null mutant exhibited normal growth and invasion rates, revealing that the GDP-Fuc de novo metabolic pathway is not essential for the development in culture of the malaria parasite during the asexual blood stages. Nonetheless, the function of this metabolic route and the GDP-Fuc pool that is generated during this stage may be important for gametocytogenesis and sporogonic development in the mosquito.

Protein O-fucosylation in Plasmodium falciparum ensures efficient infection of mosquito and vertebrate hosts

“…The punctate distribution pattern within the ER suggests that POFUT2-HA localizes within sub-domains of the ER, the presence of which has been described previously 34 . Detection of POFUT2 expression in asexual parasites is consistent with reports that GDP-fucose is biosynthesized in blood stage P. falciparum parasites 35 , though it does not appear to be essential 36 . It is unclear what, if any, protein(s) might be O-fucosylated by POFUT2 in the blood stage (Fig.…”

“…Both mutant clones developed within erythrocytes at the same rate as NF54 parasites, indicating that POFUT2 is not essential for asexual blood stage growth (Supplementary Fig. 8C ), in agreement with GDP-fucose being dispensable 36 and the absence of predicted substrates in this stage (Fig. 1d ).…”

“…The recent discovery of O-glycosylation on CSP and TRAP in P. falciparum sporozoites represents a very important advance in our understanding of Plasmodium glycobiology 7 and complements recent work on the biosynthesis of sugar nucleotides in P. falciparum 35 , 36 . We have significantly built on these findings by identifying the ER-resident enzyme responsible for O-glycosylation in P. falciparum and confirming that POFUT2 is a fucosyltransferase that modifies parasite TSR domains specifically with L-fucose at the conserved serine/threonine residue within the CXX( S/T )C sequon.…”

Genetic metabolic complementation establishes a requirement for GDP-fucose in Leishmania

“…In T. gondii , the de novo GDP-fucose pathway appears to be essential, possibly due to a role in nuclear pore complex modification ( 50 ). Protein O -fucosylation of the surface CSP and TRAP sporozoite proteins of the malaria parasite Plasmodium falciparum has been described ( 57 ), although the de novo pathway for GDP-fucose synthesis does not appear to be essential ( 58 , 59 ).…”

Implications of Plasmodium Glycosylation on Vaccine Efficacy and Design

“…Even so, homologs of several genes required for protein glycosylation are present and conserved in the genomes of Plasmodium spp. and metabolomic analyses of parasite material has demonstrated the presence of the nucleotide sugars required for protein glycosylation, which alluded to the existence of as yet undiscovered parasite glycans.With the aid of modern protein mass spectrometry methods, several research groups have recently tackled the issues of glycan lability and relatively small sample sizes to begin characterizing the true diversity of Plasmodium protein glycosylation [6][7][8][9][10][11][12]. Evidence for the synthesis of short N-linked glycans has been reported in asexual blood stages of Plasmodium falciparum [6], and there is every reason to expect that these modifications will be found in other life cycle stages as well.…”