Characterization of lipophosphoglycan from a ricin-resistant mutant of Leishmania major
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1Walter and Eliza Hall Institute of Medical Research, Post Office Royal Melbourne Hospital, Victoria, Australia.
Abstract:
One of the virulence factors of the protozoan parasite Leishmania major is the surface glycoconjugate, lipophosphoglycan (LPG). A Ricin-resistant mutant of L.major was generated and characterised with respect to its virulence in mice and the structure and expression of LPG. The LPG from this mutant (1F6-B5) retained the tripartite structure of wild-type LPG, comprising a glycosylphosphatidylinositol (GPI) anchor linked to a phosphorylated disaccharide backbone terminating in a nonreducing neutral oligosaccharide cap. The structure of the GPI anchor and the major capping oligosaccharide were identical to wild-type LPG. However, there were variations in the number of phosphorylated repeats (PO4-6Gal(beta 1-4)Man(alpha 1-) comprising the backbone of LPG, although the degree of substitution with side branches (approx. 95%) was similar to that of wild-type LPG. Thus, the mutant LPG was shorter in length having, on average, 15 repeat units per molecule compared with 30 in the wild-type LPG. The mutant LPG contained both arabinose (Ara(beta 1-2)[Gal(beta 1-3)-]1,2) and galactose ([Gal(beta 1-3)-]1-8) capped side branches linked to the backbone. In contrast to wild-type LPG, the number of arabinose-capped side chains was significantly reduced, and a new population of galactose-capped (Gal(beta 1-3)]5-8) side branches was present. The level of LPG expression in mutant parasites was approximately one-tenth of the wild-type parasite. The mutant parasites were avirulent in mice. Over a period of 18 months, they did not cause lesions and organisms could not be isolated from the draining lymph nodes.
Insights
A Leishmania major mutant with altered lipophosphoglycan (LPG) structure and reduced expression was avirulent in mice. This study investigates the role of LPG in Leishmania major virulence.
Area of Science:
- Parasitology
- Molecular Biology
- Glycobiology
Background:
- Lipophosphoglycan (LPG) is a key virulence factor on the surface of the protozoan parasite Leishmania major.
- Understanding LPG structure and expression is crucial for developing anti-Leishmania strategies.
Purpose of the Study:
- To characterize a Ricin-resistant Leishmania major mutant regarding its virulence and lipophosphoglycan (LPG) structure and expression.
- To elucidate the relationship between LPG modifications and Leishmania major pathogenicity.
Main Methods:
- Generation and characterization of a Ricin-resistant Leishmania major mutant.
- Structural analysis of lipophosphoglycan (LPG) from the mutant and wild-type parasites.
- Assessment of Leishmania major mutant virulence in a murine model.
Main Results:
- The mutant's LPG retained its tripartite structure but had fewer phosphorylated repeat units, resulting in a shorter backbone compared to wild-type LPG.
- Significant alterations in LPG side-chain capping were observed, with reduced arabinose-capped chains and the emergence of galactose-capped chains.
- LPG expression was reduced approximately tenfold in the mutant, which proved to be avirulent in mice, causing no lesions and showing no recoverable organisms.
Conclusions:
- Lipophosphoglycan (LPG) structure, length, and expression level are critical determinants of Leishmania major virulence.
- Modifications in LPG, particularly reduced length and altered capping, render the parasite avirulent.
- This mutant provides a valuable tool for further investigation into LPG function in Leishmania pathogenesis.