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An antigenically distinct lipophosphoglycan on amastigotes of Leishmania major
T A Glaser1, S F Moody, E Handman
1Walter and Eliza Hall Institute of Medical Research, Melbourne, Victoria, Australia.
Abstract:
We show that lipophosphoglycan (LPG) on the surface of amastigotes of Leishmania major is antigenically and biochemically distinct from promastigote LPG. A rabbit antiserum raised against the amastigote integral membrane fraction detected LPG spanning the region of Mr 55,000-100,000 on Western blots of the amastigote integral membrane fraction, but did not recognize the promastigote integral membrane fraction. WIC 79.3, a monoclonal antibody which recognizes L. major metacyclic promastigote LPG, did not recognize the amastigote integral membrane fraction on Western blots. The antigen recognized by this rabbit antiserum was shown to be LPG by its migration pattern on SDS-PAGE, the presence of terminal galactose residues, recognition by a monoclonal antibody to LPG, WIC 108.3, the biosynthetic incorporation of label from [3H]glucose and [32P]phosphate, a hydrophobic chromatography elution profile similar to promastigote LPG, and the presence of a lipid anchor sensitive to phosphatidylinositol-specific phospholipase C. The temporal regulation of LPG expression during parasite differentiation was studied in vitro. During amastigote-to-promastigote transformation, the amastigote-specific form of LPG disappeared after subculture at 48 h. The WIC 79.3 epitope was not detected by Western blotting on transforming parasites until 48 h in culture. During promastigote-to-amastigote transformation, the amastigote-specific form of LPG was detected 12 h after infection. WIC 79.3 epitopes gradually diminished over 48 h. The results demonstrate the developmentally regulated expression of an antigenically distinct LPG on amastigotes of L. major.
Insights
Lipophosphoglycan (LPG) on Leishmania major amastigotes is distinct from promastigote LPG. This amastigote-specific LPG is developmentally regulated during parasite differentiation, indicating distinct roles in infection.
Area of Science:
- Parasitology
- Molecular Biology
- Immunology
Background:
- Lipophosphoglycan (LPG) is a major surface glycoconjugate of Leishmania parasites.
- LPG plays crucial roles in parasite-host interactions and immune evasion.
- Distinct forms of LPG may be expressed during different life cycle stages.
Purpose of the Study:
- To investigate the antigenic and biochemical differences of lipophosphoglycan (LPG) between amastigote and promastigote forms of Leishmania major.
- To characterize the temporal expression of amastigote-specific LPG during in vitro differentiation.
- To understand the developmental regulation of LPG in Leishmania major.
Main Methods:
- Western blotting using specific antisera and monoclonal antibodies.
- Analysis of LPG by SDS-PAGE and hydrophobic chromatography.
- Biosynthetic labeling with [3H]glucose and [32P]phosphate.
- Enzymatic digestion with phosphatidylinositol-specific phospholipase C.
- In vitro differentiation assays (amastigote-to-promastigote and promastigote-to-amastigote).
Main Results:
- Leishmania major amastigote LPG is antigenically and biochemically distinct from promastigote LPG.
- A rabbit antiserum detected amastigote-specific LPG (Mr 55,000-100,000) not recognized by antibodies against promastigote LPG.
- Amastigote-specific LPG expression was temporally regulated during in vitro differentiation, appearing after 12 hours of promastigote-to-amastigote transformation and disappearing after 48 hours of amastigote-to-promastigote transformation.
- The amastigote-specific LPG possesses a lipid anchor sensitive to phosphatidylinositol-specific phospholipase C.
Conclusions:
- Leishmania major expresses an antigenically distinct form of LPG on amastigotes compared to promastigotes.
- LPG expression is developmentally regulated, with stage-specific forms appearing during parasite differentiation.
- These findings highlight the dynamic nature of the Leishmania parasite surface and suggest stage-specific functions for LPG.