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Evidence that intracellular stages of Leishmania major utilize amino sugars as a major carbon source.

ABSTRACT: Intracellular parasites, such as Leishmania spp, must acquire suitable carbon sources from the host cell in order to replicate. Here we present evidence that intracellular amastigote stages of Leishmania exploit amino sugars in the phagolysosome of mammalian macrophages as a source of carbon and energy. L. major parasites are capable of using N-acetylglucosamine and glucosamine as primarily carbon sources and contain key enzymes required for conversion of these sugars to fructose-6-phosphate. The last step in this pathway is catalyzed by glucosamine-6-phosphate deaminase (GND), which was targeted to glycosomes via a canonical C-terminal targeting signal when expressed as a GFP fusion protein. Mutant parasites lacking GND were unable to grow in medium containing amino sugars as sole carbohydrate source and rapidly lost viability, concomitant with the hyper-accumulation of hexosamine-phosphates. Expression of native GND, but not a cytosolic form of GND, in ?gnd parasites restored hexosamine-dependent growth, indicating that toxicity is due to depletion of glycosomal pools of ATP. Non-lethal increases in hexosamine phosphate levels in both ?gnd and wild type parasites was associated with a defect in promastigote metacyclogenesis, suggesting that hexosamine phosphate levels may influence parasite differentiation. Promastigote and amastigote stages of the ?gnd mutant were unable to replicate within macrophages and were either completely cleared or exhibited reduced lesion development in highly susceptible Balb/c mice. Our results suggest that hexosamines are a major class of sugars in the macrophage phagolysosome and that catabolism of scavenged amino sugars is required to sustain essential metabolic pathways and prevent hexosamine toxicity.

SUBMITTER: Naderer T

PROVIDER: S-EPMC3009595 | biostudies-literature | 2010

REPOSITORIES: biostudies-literature

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Evidence that intracellular stages of Leishmania major utilize amino sugars as a major carbon source.

Naderer Thomas T Heng Joanne J McConville Malcolm J MJ

PLoS pathogens 20101223 12

Intracellular parasites, such as Leishmania spp, must acquire suitable carbon sources from the host cell in order to replicate. Here we present evidence that intracellular amastigote stages of Leishmania exploit amino sugars in the phagolysosome of mammalian macrophages as a source of carbon and energy. L. major parasites are capable of using N-acetylglucosamine and glucosamine as primarily carbon sources and contain key enzymes required for conversion of these sugars to fructose-6-phosphate. Th ...[more]

PMID: 21203480

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Project description:The life cycle of the Leishmania parasite in the sand fly vector involves differentiation into several distinctive forms, each thought to represent an adaptation to specific microenvironments in the midgut of the fly. Based on transcriptome sequencing (RNA-Seq) results, we describe the first high-resolution analysis of the transcriptome dynamics of four distinct stages of Leishmania major as they develop in a natural vector, Phlebotomus duboscqi The early transformation from tissue amastigotes to procyclic promastigotes in the blood-fed midgut was accompanied by the greatest number of differentially expressed genes, including the downregulation of amastins, and upregulation of multiple cell surface proteins, sugar and amino acid transporters, and genes related to glucose metabolism and cell cycle progression. The global changes accompanying post-blood meal differentiation of procyclic promastigotes to the nectomonad and metacyclic stages were less extensive, though each displayed a unique signature. The transcriptome of nectomonads, which has not been studied previously, revealed changes consistent with cell cycle arrest and the upregulation of genes associated with starvation and stress, including autophagic pathways of protein recycling. Maturation to the infective, metacyclic stage was accompanied by changes suggesting preadaptation to the intracellular environment of the mammalian host, demonstrated by the amastigote-like profiles of surface proteins and metabolism-related genes. Finally, a direct comparison between sand fly-derived and culture-derived metacyclics revealed a reassuring similarity between the two forms, with the in vivo forms distinguished mainly by a stronger upregulation of transcripts associated with nutrient stress.IMPORTANCE The life cycle of Leishmania parasites in the sand fly vector includes their growth and development as morphologically distinct forms of extracellular promastigotes found within the different microenvironments of the gut. Based on RNA-Seq, we provide here the first high-resolution, transcriptomic analysis of Leishmania insect stages during their cyclical development in vivo, from tissue amastigotes ingested with the blood meal to infective, metacyclic promastigotes that initiate infection in the mammalian host. The most extensive genetic reprogramming occurred during the early transformation of amastigotes to rapidly dividing procyclic promastigotes in the blood-fed midgut, with major changes in the abundance of mRNAs for surface proteins and metabolism. The post-blood meal-adapted nectomonad stage was characterized by the downregulation of cell cycle-related genes and the upregulation of stress- and starvation-related genes. Finally, the transcriptome of metacyclic promastigotes shifted to a more amastigote-like profile, suggesting their preadaptation to the intracellular host environment.

Dataset Information

Evidence that intracellular stages of Leishmania major utilize amino sugars as a major carbon source.

Publications

Evidence that intracellular stages of Leishmania major utilize amino sugars as a major carbon source.

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