The immunochemical structure and surface arrangement of Leishmania donovani lipophosphoglycan determined using monoclonal antibodies

Tolson, Douglas L.; Turco, Salvatore J.; Beecroft, Robert P.; Pearson, Terry W.

doi:10.1016/0166-6851(89)90113-8

Cited by 108 publications

(73 citation statements)

References 19 publications

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“…1) (55). Gal-modified PG repeats can be recognized by the monoclonal antibody WIC79.3 (56), whereas unmodified PG repeats can be recognized by monoclonal antibody CA7AE (39). We used these in immunoblotting to probe the effects of LPG5A and LPG5B mutations on PG synthesis; similar results were obtained by immunofluorescence microscopy (data not shown).…”

Section: Lpg5a or Lpg5b Is Sufficient For Pg Synthesis But The Lpg5asupporting

confidence: 56%

Two Functionally Divergent UDP-Gal Nucleotide Sugar Transporters Participate in Phosphoglycan Synthesis in Leishmania major

Capul

Barron

Dobson

et al. 2007

Journal of Biological Chemistry

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In the protozoan parasite Leishmania, abundant surface and secreted molecules, such as lipophosphoglycan (LPG) and proteophosphoglycans (PPGs), contain extensive galactose in the form of phosphoglycans (PGs) based on (Gal-Man-PO 4 ) repeating units. PGs are synthesized in the parasite Golgi apparatus and require transport of cytoplasmic nucleotide sugar precursors to the Golgi lumen by nucleotide sugar transporters (NSTs). GDP-Man transport is mediated by the LPG2 gene product, and here we focused on transporters for UDP-Gal. Data base mining revealed 12 candidate NST genes in the L. major genome, including LPG2 as well as a candidate endoplasmic reticulum UDP-glucose transporter (HUT1L) and several pseudogenes. Gene knock-out studies established that two genes (LPG5A and LPG5B) encoded UDP-Gal NSTs. Although the single lpg5A ؊ and lpg5B ؊ mutants produced PGs, an lpg5A ؊ /5B ؊ double mutant was completely deficient. PG synthesis was restored in the lpg5A ؊ /5B ؊ mutant by heterologous expression of the human UDP-Gal transporter, and heterologous expression of LPG5A and LPG5B rescued the glycosylation defects of the mammalian Lec8 mutant, which is deficient in UDP-Gal uptake. Interestingly, the LPG5A and LPG5B functions overlap but are not equivalent, since the lpg5A ؊ mutant showed a partial defect in LPG but not PPG phosphoglycosylation, whereas the lpg5B ؊ mutant showed a partial defect in PPG but not LPG phosphoglycosylation. Identification of these key NSTs in Leishmania will facilitate the dissection of glycoconjugate synthesis and its role(s) in the parasite life cycle and further our understanding of NSTs generally.

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Section: Lpg5a or Lpg5b Is Sufficient For Pg Synthesis But The Lpg5asupporting

confidence: 56%

Two Functionally Divergent UDP-Gal Nucleotide Sugar Transporters Participate in Phosphoglycan Synthesis in Leishmania major

Capul

Barron

Dobson

et al. 2007

Journal of Biological Chemistry

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“…The PG domain contains species and strain-specific substitutions (Tolson et al, 1989;McConville et al, 1990;Ilg et al, 1992) and developmentally regulated polymorphisms (Glaser et al, 1991;Sacks, 1992;Moody et al, 1993). Furthermore, it recently has become apparent that L. major strains differ in their PG structure Dobson, Scholtes et al, 2003).…”

Section: Discussionmentioning

confidence: 99%

Transcriptional Inhibition of Interleukin-12 Promoter Activity in Leishmania Spp.–Infected Macrophages

Jayakumar

Widenmaier

et al. 2008

Journal of Parasitology

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To establish and persist within a host, Leishmania spp. parasites delay the onset of cell-mediated immunity by suppressing interleukin-12 (IL-12) production from host macrophages. Although it is established that Leishmania spp.-infected macrophages have impaired IL-12 production, the mechanisms that account for this suppression remain to be completely elucidated. Using a luciferase reporter assay assessing IL-12 transcription, we report here that Leishmania major, Leishmania donovani, and Leishmania chagasi inhibit IL-12 transcription in response to interferon-gamma, lipopolysaccharide, and CD40 ligand and that Leishmania spp. lipophosphoglycan, phosphoglycans, and major surface protein are not necessary for inhibition. In addition, all the Leishmania spp. strains and life-cycle stages tested inhibited IL-12 promoter activity. Our data further reveal that autocrineacting host factors play no role in the inhibitory response and that phagocytosis signaling is necessary for inhibition of IL-12.The hallmark of an intracellular pathogen is its ability to survive within the intracellular niche. These organisms must be resistant to, or able to evade, the host cell's microbiocidal mechanisms. This dilemma is particularly relevant to Leishmania spp. because these organisms primarily reside within vertebrate macrophages (MP). These host cells become activated for microbial destruction and cytokine secretion by exposure to immune modulators, such as interferon-gamma (IFN-γ) and CD40 ligand (CD40L) found on activated T-cells. One strategy Leishmania spp. parasites use to avoid host cell activation is to interfere with the signaling pathways that induce MP to become microbicidal (Gregory and Olivier, 2005); another is inhibiting or delaying the production of activating cytokines (Belkaid et al., 2000).The most consistent dysfunction reported from Leishmania spp.-infected MP is the aberrant production of inflammatory cytokines, specifically an inhibition of interleukin-12 (IL-12) production (McDowell and Sacks, 1999). Being essential for T-helper 1 (Th1) cell differentiation, the inability of Leishmania spp.-infected MP to produce IL-12 allows Leishmania spp. to evade acquired resistance by postponing IL-12 production and the induction of IFN-γ, thereby allowing the establishment of the infection; both clinical and experimental studies indicate that the onset of Th1-mediated immunity and Leishmania spp. killing is indeed delayed (Melby, 1991). Inhibition of MP IL-12 production and resulting Th1 responses is not unique to Leishmania; viruses (Chehimi et al., 1994;Chougnet et al., 1996;Karp et 1996) and bacteria (Marth and Kelsall, 1997;Sutterwala et al., 1997;Matsunaga et al., 2003) also exploit this mechanism to avoid clearance.The general nature of impaired IL-12 production in Leishmania spp.-infected MP has extended to every IL-12 agonist that has been tested. Leishmania spp.-infected MP are unable to produce IL-12 even in response to strong inflammatory stimuli, including microbial stimuli, e.g., lipopolysacchar...

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“…For blotting with the anti-serum CA7AE which recognizes the phosphoglycan [Gal(␤1-4)Man(␣ 1-PO 4 ) → 6] repeating disaccharide units present on LPG, PPG and secretory acid phosphatase [22], 2-3 × 10 8 cells in 1 ml of 0.15N NaCl were suspended in 50 mM Tris-HCl, pH 8, 1 mM PMSF, and sonicated on ice with a probe sonicator for 20 s. 2× SDS sample buffer with 400 mM dithiothreitol was added and debris was pelleted at 15,000 × g for 5 min at 25 • C [23]. 5 × 10 6 cell equivalents were loaded per lane for 12% SDS-PAGE with a 5% stacker and run at 60 V. The gel was blotted to nitrocellulose membranes and blocked overnight with 5% milk in PBS.…”

Section: Western Blotting and Detection Of Secretory Acid Phosphatasementioning

confidence: 99%

An in vitro system for developmental and genetic studies of Leishmania donovani phosphoglycans

Goyard

Segawa

Gordon

et al. 2003

Molecular and Biochemical Parasitology

164

183

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The immunochemical structure and surface arrangement of Leishmania donovani lipophosphoglycan determined using monoclonal antibodies

Cited by 108 publications

References 19 publications

Two Functionally Divergent UDP-Gal Nucleotide Sugar Transporters Participate in Phosphoglycan Synthesis in Leishmania major

Two Functionally Divergent UDP-Gal Nucleotide Sugar Transporters Participate in Phosphoglycan Synthesis in Leishmania major

Transcriptional Inhibition of Interleukin-12 Promoter Activity in Leishmania Spp.–Infected Macrophages

An in vitro system for developmental and genetic studies of Leishmania donovani phosphoglycans

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