The role of phosphoglycans inLeishmania–sand fly interactions

Abstract

Leishmania promastigotes synthesize an abundance of phosphoglycans, either attached to the cell surface through phosphatidylinositol anchors (lipophosphoglycan, LPG) or secreted as proteincontaining glycoconjugates. These phosphoglycans are thought to promote the survival of the parasite within both its vertebrate and invertebrate hosts. The relative contributions of different phosphoglycan-containing molecules in Leishmania-sand fly interactions were tested by using mutants specifically deficient in either total phosphoglycans or LPG alone. Leishmania donovani promastigotes deficient in both LPG and protein-linked phosphoglycans because of loss of LPG2 (encoding the Golgi GDP-Man transporter) failed to survive the hydrolytic environment within the early blood-fed midgut. In contrast, L. donovani and Leishmania major mutants deficient solely in LPG expression because of loss of LPG1 (involved in biosynthesis of the core oligosaccharide LPG domain) had only a slight reduction in the survival and growth of promastigotes within the early blood-fed midgut. The ability of the LPG1-deficient promastigotes to persist in the midgut after blood meal excretion was completely lost, and this defect was correlated with their inability to bind to midgut epithelial cells in vitro. For both mutants, when phosphoglycan expression was restored to wild-type levels by reintroduction of LPG1 or LPG2 (as appropriate), then the wild-type phenotype was also restored. We conclude, first, that LPG is not essential for survival in the early blood-fed midgut but, along with other secreted phosphoglycan-containing glycoconjugates, can protect promastigotes from the digestive enzymes in the gut and, second, that LPG is required to mediate midgut attachment and to maintain infection in the fly during excretion of the digested blood meal.

Regulated Degradation of an Endoplasmic Reticulum Membrane Protein in a Tubular Lysosome inLeishmania mexicana

“…These GPI-anchored macromolecules and free GPIs are most highly expressed in the promastigote (sandfly) stage and are thought to form a protective surface glycocalyx. They also mediate specific hostparasite interactions in the midgut of the sandfly vector and are required for promastigote invasion of macrophages in the mammalian host (Ilg, 2000;Sacks et al, 2000;Spath et al, 2000). Recent studies with the use of L. mexicana promastigotes suggest that the protein anchor and LPG anchor precursors and free GPIs are assembled on distinct phosphatidylinositol molecular species in a subcompartment of the ER (Ralton and McCon-ville, 1998;Ilgoutz et al, 1999b).…”

Manipulación genética y el estudio del parásito protozoario Leishmania.

“…Las aproximaciones genéticas también se han realizado por medio de complementación génica, en la que se empieza a partir de un fenotipo mutante o variante para identificar el gen involucrado (49,72,(76)(77)(78)(79)(80)83,88). Los genes responsables de los defectos son identificados por una transfección en masa de los mutantes con una librería cósmida de ADN de la especie de Leishmania silvestre y se seleccionan por recobrar la función a través de la complementación.…”

“…Los genes identificados por complementación fueron lpg1 y lpg2 (transportador de la GDP-manosa a la luz del aparato de Golgi). Los mutantes carentes de ambos genes no sobrevivieron en el ambiente hidrolítico dentro del intestino medio del insecto; además, la unión del parásito al intestino medio del insecto y el mantenimiento de la infección luego de la excreción de la sangre del infectado se vieron comprometidas (83,88). Los estudios subsecuentes han extendido esta aproximación para la identificación de más de 10 genes involucrados en la síntesis de LPG y glicoconjugados relacionados (72,(76)(77)(78)(79)(80)(81)90).…”

Probing Elongating and Branching β-d-Galactosyltransferase Activities in Leishmania Parasites by Making Use of Synthetic Phosphoglycans

“…There was only one product of digestion, which co-migrated with standard galactose (Figure 6, lanes 3 and 7). Direct treatment of Bacic, 22,23 our data would imply that the addition of the side-chain (β1-3)Gal residues was compatible with a mechanism independent of the synthesis of the repeating backbone units.In the case of L. major LPG, branching structures containing two or three (β1-3)Gal units in the side chain are common, but larger structures with up to 11 (β1-3)Gal units in the side chain have been isolated from L. major amastigote LPG (and are present in LPGs at very low levels) 15 and from a L. major mutant cell line LPG. 37 It was postulated that the rate of addition of the first Gal onto exogenous linear acceptor (initiating side-chain GalT) is much higher than the rate of addition of the second and the third galactose residues (elongating side-chain GalTs).…”