A Family of Dual-Activity Glycosyltransferase-Phosphorylases Mediates Mannogen Turnover and Virulence in Leishmania Parasites

M Fleur Sernee,Julie E Ralton,Tracy L Nero,Lukasz F Sobala,Joachim Kloehn,Marcel A Vieira-Lara,Simon A Cobbold,Lauren Stanton,Douglas E.V Pires,Eric Hanssen,Alexandra Males,Tom Ward,Laurence M Bastidas,Phillip L Van Der Peet,Michael W Parker,David B Ascher,Spencer J Williams,Gideon J Davies,Malcolm J Mcconville

doi:10.1016/j.chom.2019.08.009

M Fleur Sernee, Julie E Ralton + Show 17 more

Open Access

https://doi.org/10.1016/j.chom.2019.08.009

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Abstract

Parasitic protists belonging to the genus Leishmania synthesize the non-canonical carbohydrate reserve, mannogen, which is composed of β-1,2-mannan oligosaccharides. Here, we identify a class of dual-activitymannosyltransferase/phosphorylases (MTPs) that catalyze both the sugar nucleotide-dependent biosynthesis and phosphorolytic turnover of mannogen. Structural and phylogenic analysis shows that while the MTPs are structurally related to bacterial mannan phosphorylases, they constitute a distinct family of glycosyltransferases (GT108) that have likely been acquired by horizontal gene transfer from gram-positive bacteria. The seven MTPs catalyze the constitutive synthesis and turnover of mannogen. This metabolic rheostat protects obligate intracellular parasite stages from nutrient excess, and is essential for thermotolerance and parasite infectivity in the mammalian host. Our results suggest that the acquisition and expansion of the MTP family in Leishmania increased the metabolic flexibility of these protists and contributed to their capacity to colonize new host niches.

Highlights

The Trypanosomatidae are parasitic protists that cause diseases such as human African trypanosomiasis (Trypanosoma brucei), Chagas disease (T. cruzi), and human leishmaniasis (Leishmania spp) that are estimated to chronically or acutely infect more than 100 million people worldwide (Stuart et al, 2008; Singh et al, 2014; Burza et al, 2018; Banuls et al, 2011)
We show that the synthesis and turnover of mannogen regulates Leishmania central carbon metabolism and cellular stress responses under both nutrient replete, as well as nutrient-limiting conditions, and is essential for parasite infectivity in the mammalian host
Detection of Mannogen Synthase and Phosphorylase Activities in Leishmania Extracts In support of earlier studies (Ralton et al, 2003; van der Peet et al, 2006, 2012), mannogen synthesis was shown to be mediated by guanosine diphosphate (GDP)-Man-dependent mannosyltransferase(s), as incubation of L. mexicana cell-free extracts with GDP-3H-Man and either native mannogen or the synthetic substrate, octyl-a-Dmannopyranoside (OM1), resulted in the synthesis of labeled oligosaccharides (Figure 1B) comprising linear chains of b-1,2-linked mannose (Figures S1A and S1B)

Summary

Introduction

The Trypanosomatidae are parasitic protists that cause diseases such as human African trypanosomiasis (Trypanosoma brucei), Chagas disease (T. cruzi), and human leishmaniasis (Leishmania spp) that are estimated to chronically or acutely infect more than 100 million people worldwide (Stuart et al, 2008; Singh et al, 2014; Burza et al, 2018; Banuls et al, 2011) These pathogens often have complex life cycles, occupying extracellular and intracellular niches in different insect, plant, and vertebrate hosts. Leishmania and closely related trypanosomatids (including the plant pathogen Phytomonas and insect parasites Leptomonas, Crithidia, and Herpetomonas) have subsequently acquired a new pathway of carbohydrate reserve biosynthesis (Ralton et al, 2003; Sernee et al, 2006) These parasites synthesize mannogen, a family of linear oligosaccharides made up of 2-60 b-1,2-linked mannose residues (Previato et al, 1984; Keegan and Blum, 1992; Ralton et al, 2003). How the acquisition of mannogen synthesis contributes to the parasitic lifestyle of Leishmania remains unclear

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