Human infections caused by parasitic protozoans and helminths are among the world's
leading causes of death. More than a million people die each year from diseases like
malaria and neglected tropical diseases like leishmaniasis, trypanosomiasis, and schistosomiasis.
Patients also endure disabilities that cause lifelong suffering and that affect productivity
and development [1]. More insidiously, parasites generate important economic losses,
since they often also infect commercially valuable animals. Worldwide, exposure to
parasites is increasing due to growing international travel and migrations, as well
as climate changes, which affect the geographic distribution of the parasite vectors.
The parasitic threat is also aggravated by the rise of the immunocompromised population,
which is particularly sensitive to parasite infections (e.g., individuals with AIDS
and other immunodeficiencies).
A common feature of protozoan parasites and helminths is the synthesis of glycoconjugates
and glycan-binding proteins for protection and to interact and respond to changes
in their environment. To address the many challenges associated with the study of
the structure, the biosynthesis, and the biology of parasitic glycans, the authors
of this article have established GlycoPar, a European Marie Curie training program
steered by some of the world's academic leaders in the field of parasite glycobiology,
in close association with European industrial enterprises. The main scientific goal
of this network is the description of novel paradigms and models by which parasite
glycoconjugates play a role in the successful colonization of the different hosts.
By means of a training-through-research program, the aim of the network is to contribute
to the training of a generation of young scientists capable of tackling the challenges
posed by parasite glycobiology.
Parasites Are Covered by a Protective Glycocalyx
Due to the complexity of their life cycles, parasites need to sequentially exploit
various host species to complete the different stages involved in their survival and
development. The interactions with their different hosts are critical for the completion
of each life stage and are often based on carbohydrate recognition. In particular,
parasites have developed different strategies to escape the immune and defense systems
of the different infected organisms. Their surfaces are covered by glycoconjugates
of varied natures, often of types absent from mammals. This so-called glycocalyx is
protective against the host defense systems but may also be implicated in "hijacking"
proteins involved in host innate immunity (Fig 1) [2,3]. Thus, through a "glycan gimmickry"
designated process, helminths express host-like glycans that interact with host lectins
to modulate the immune response [4]. Furthermore, the walls that protect different
parasitic cysts from harsh environments are also rich in polysaccharides and polysaccharide-binding
lectins [5]. Thereby, glycans are crucial for parasite virulence and survival.
10.1371/journal.ppat.1005169.g001
Fig 1
The surfaces of parasites, such as Trypanosoma brucei brucei, are covered by glycoconjugates
forming a protective glycocalyx against the host defense systems.
False-color scanning electron microscopy (EM) of a T. b. brucei procyclic interacting
with cell microvilli in the tsetse fly proventriculus (bottom panel). Transmission
EM of ruthenium-red stained ultrathin sections showing the surface glycocalyx of T.
b. brucei procyclic cells (middle panel). Scheme summarizing the main surface glycosylphosphatidylinositol
(GPI)-anchored (EP- and GPEET-procyclins and trans-sialidases) and transmembrane (including
polytopic) glycoproteins and glycolipids expressed by T. b. brucei procyclics (top
panel) [2,24]. Open rectangles linked to GPI molecules represent side chains characteristic
of surface glycoconjugates from procyclic T. b. brucei. GIPLs: glycoinositolphospholipids,
or free GPIs. EM images obtained by C. Rose, A. Beckett, L. Tetley, I. Prior, and
A. Acosta-Serrano.
Since glycans are central to host–parasite interactions, their study constitutes a
fertile, but currently largely unexploited, area for therapeutic applications. Research
in parasite glycosylation provides new opportunities for the discovery of vaccine
candidates and for the development of novel chemotherapy approaches and diagnostic
tools. Thus, for instance, besides its effect modulating the host immune response
against the infection, glycans from Schistosoma mansoni are currently being explored
as targets for vaccination and/or serodiagnosis of human schistosomiasis [6]. Nevertheless,
there are many challenges associated with working with parasites, including problems
in obtaining sufficient amounts of biological material for analytical purposes, difficulties
of culturing the different life stages, and, on occasion, the lack of tools for functional
genomics and molecular biology approaches. Glycans add another level of difficulty
to these studies, due to their extensive diversity and exquisite complexity. In contrast
to nucleic acid and proteins, their biosynthesis is only indirectly template-driven
and generates an important amount of structural variability in biological systems.
This complexity is critical in molecular recognition events including cell–cell, cell–matrix,
and cell–molecule interactions during essential steps of pathogenesis. Thus, the thorough
characterization of parasite glycobiology requires systematic approaches that focus
on the description of the glycosylation precursors, the glycan-processing enzymes,
and the structure and functional significance of parasitic glycans. In addition, most
of the medically and veterinarially important parasites are phylogenetically ancient
organisms and represent good models for studying evolutionary aspects of eukaryotic
glycosylation. Thus, the study of parasite glycans may unravel novel mechanisms also
present in higher eukaryotes. Excellent examples are the description of the structure
of glycosylphosphatidylinositol (GPI) membrane anchors in African trypanosomes [7]
or the discovery of the glycoprotein quality control cycle, thanks to seminal studies
on the N-glycosylation pathway of trypanosomatid parasites [8]. Interestingly enough,
different parasitic protists present variable lengths in their N-glycan precursors
that directly affect this N-glycan-dependent quality control system [9].
The Metabolic Precursors of Parasite Glycosylation
Glycan synthesis requires activated monosaccharides, mainly in the form of nucleotide
sugars that will be used by glycosyltransferase enzymes as glycosyl donor substrates
in glycosylation reactions. Therefore, the presence of activated sugars is a prerequisite
for glycan biosynthesis, and their availability influences the glycan structures that
may be synthesized by a parasite (the glycome). Thus, valuable information about the
glycome can be gained from the identification and quantification of the sugar nucleotide
pools maintained during the life stages of different parasites. For example, the capping
of Leishmania major surface lipophosphoglycan with arabinose side chains, which is
required for detachment of the infectious parasites from the sand fly midgut, correlates
with a strong increase of the GDP-α-D-arabinopyranose pool [10].
Sugar nucleotides are formed by de novo pathways requiring the bioconversion of an
existing sugar or sugar nucleotide or by salvage pathways involving the activation
of the sugar using a kinase and a pyrophosphorylase. The conservation of specific
biosynthetic pathways in the parasite genomes are strong hints of the presence of
nucleotide sugar pools [11,12]. Monosaccharide activation usually takes place in the
cytoplasm, although in Trypanosoma brucei brucei and possibly other kinetoplastid
parasites, these biosynthetic reactions occur in a specific organelle called glycosome
[13]. Since sugar nucleotides are mostly used by glycosyltransferases in the endoplasmic
reticulum and/or the Golgi apparatus, they must be translocated to these cellular
compartments by specific transporters (Fig 2). This metabolic compartmentalization
and the study of the transporters involved also offer new opportunities for the selective
inhibition of crucial glycosylation reactions in parasites.
10.1371/journal.ppat.1005169.g002
Fig 2
Glycosylation processes involve different cellular compartments.
Glycan biosynthesis and cellular compartments involved in the glycosylation process.
Sugars are carried across the plasma membrane into cells or are salvaged from degraded
glycoconjugates at lysosomes. Through biosynthetic and interconversion reactions,
monosaccharides are activated into different nucleotide sugars. Sugar activation generally
takes place in the cytoplasm, although several enzymes involved in sugar nucleotide
biosynthesis in T. b. brucei are localized in the glycosome. After being activated,
sugar nucleotides are transported into the endoplasmic reticulum/Golgi apparatus and
used by different glycosyltransferases (GT). Glycosyltransferases and other glycan-processing
enzymes define the assembly and final structure of glycans that are secreted or located
in the cell surface, forming a protective glycocalyx. Sugar nucleotide transporters
are marked with an asterisk (*).
Parasitic Glycan-Processing Enzymes and Glycan-Binding Proteins
Glycosyltransferases transfer sugar moieties from activated donors to specific acceptor
molecules, generating glycosidic linkages between carbohydrates or between a carbohydrate
and a noncarbohydrate moiety. Therefore, they define the assembly and final structure
of glycan chains, which can be linear or branched and of various lengths. Glycoside
hydrolases, the enzymes that hydrolyze glycosidic bonds, form another main group of
carbohydrate-active enzymes that also play important roles in determining the final
structure of mature glycans. The combined action of several of these enzymes in the
secretory pathway leads to a vast and diverse array of glycan structures. Additionally,
parasitic glycan-binding proteins interact with specific parasite and host glycan
structures present in the surface of cells.
Sequence-based families of glycosyltransferases, glycoside hydrolases, and carbohydrate-binding
proteins group together according to their function, indicating that the acquisition
of the specificities of these enzymes evolved from common progenitors. Therefore,
despite the huge diversity of glycans, the activities and molecular mechanism of the
enzymes involved in their biosynthesis can often be inferred from their sequences
[14]. Nevertheless, because of the substantial evolutionary distance between protozoan
parasites and higher eukaryotes, it can be challenging to define the precise function
of specific parasitic glycosyltransferases from sequence similarity [15,16] or by
inference from the final structures determined by a particular glycosylation pathway
[17,18]. Glycosyltransferases and other glycan-processing enzymes involved in the
biosynthesis of glycans essential for the survival and infectivity of parasites might
be exploited as drug targets. Therefore, increasing our knowledge of the different
parasitic glycosylation pathways and their biological relevance will contribute to
uncovering the therapeutic potential therein.
Parasite Glycomics and the Biological Function of Glycoconjugates
The characterization and quantification of the complete set of glycans and glycoconjugates
made by a cell or organism at a given time is defined as glycomics. Since glycosylation
is the most structurally diverse, and one of the most abundant, protein and lipid
modifications, the description of the spectrum of all glycan structures—the glycome—of
even just a single cell type is a huge challenge. Nevertheless, to shed light on the
structure–function relationship of parasite glycans at the molecular level, a detailed
knowledge of their structures is an important prerequisite that can only be achieved
through the use of different analytical methodologies and glycoproteomics and glycolipidomics
strategies. Currently, mass spectrometry is a key tool in glycomics and has revealed
highly unusual glycans from a number of unicellular and metazoan parasites [19].
The assessment of the functional significance of the different glycosylation states
will only be achieved by employing adequate screening and/or genetic tools that, in
the case of particular parasites, are still in the development stage [20]. Host receptor
molecules can specifically recognize glycans, and these glycan–receptor interactions
are related to migration, invasion, adhesion, toxin production, and other essential
processes during the course of parasitic infections. By a thorough exploration of
the glycomic capacity of parasites and its influence on the interactions with their
hosts, the code defined by the different glycan structures can be gradually characterized.
In addition, glycomic approaches can be illuminating in the discovery of novel antigenic
glycans for the development of diagnostic tools or glycovaccines. An important step
in this respect would be the development of glycan microarrays reflecting parasite
glycomes in order to identify binding partners in the human proteome, such as components
of the innate immune system. Similarly, identifying host glycan structures recognized
by parasite proteins with lectin-like properties will be fundamental for describing
host–parasite interactions in parasitic diseases.
Future Perspectives: The Translation of Parasitic Glycobiology
Glycobiology has become a well-established area of study in recent decades and is
currently providing drug targets against several pathogens and diseases. Ethambutol,
Caspofungin, Zanamivir, and Oseltamivir are well-known examples of commercial drugs
in use—as therapies against tuberculosis, candidiasis, aspergillosis, and influenza—that
target glycosylation and carbohydrate processing. In this regard, echinocandins, antifungal
drugs that target β-1,3-glucan synthesis, also inhibit oocyst wall biosynthesis in
Eimeria [21]. Similarly, bacterial polysaccharide–protein conjugate vaccines have
recently revolutionized vaccination strategies. This approach may be applied to prevent
or treat parasitic diseases, using parasite-derived xeno-glycans absent in the human
glycome [6,22]. Furthermore, the identification of parasitic glycan antigen structures
and monoclonal antibodies to these epitopes holds unprecedented promise for the development
of novel diagnostic procedures for various parasitic infections [23]. Thus, through
profound and systematic approaches to this important but frequently neglected area
of pathogenic parasite research, knowledge about the biology of these organisms will
be extended, and novel methods to tackle them will likely be uncovered.