Lipophosphoglycan expression and virulence in ricin-resistant variants of Leishmania major.
Lipophosphoglycan (LPG) of Leishmania is a polymorphic molecule comprising an alkylglycerol anchor, a conserved oligosaccharide core and a species-specific polymer of oligosaccharide repeats jointed by phosphodiester bonds. This molecule, together with the membrane polypeptide gp63, has been implicated as a parasite receptor for host macrophages. To examine the role of LPG in parasite infectivity glycosylation variants of Leishmania major were generated by chemical mutagenesis of a virulent cloned line V121 and variants with modified LPG selected using the galactose-specific lectin Ricinus communis II (RCA II). Twenty RCA II-resistant primary clones were generated. Analysis of LPG profile by immunoblotting using LPG-specific monoclonal and polyclonal antibodies revealed that some of the clones were LPG-deficient. Three clones that did not bind any LPG-specific antibodies but expressed normal levels of the Mr 63,000 glycoprotein (gp63), a second parasite receptor for host, were chosen for detailed studies. All three clones expressed, at least to some extent, a surface molecule which could be labeled by mild periodate oxidation and sodium borotritide and behaved like LPG by hydrophobic interaction chromatography. All clones also bound a well-characterized monoclonal antibody L157 directed to the core oligosaccharide of LPG, but did not bind another monoclonal antibody, CA7AE, to an epitope on a repeating unit shared by Leishmania donovani and L. major LPG. A third monoclonal antibody, 5E6, recognizing LPG on the surface of wild-type V121 promastigotes bound only to RCA II-resistant clone 3A2-C3 and was restricted to an internal structure. The LPG molecule that this clone expressed was a form of LPG by its chromatographic behavior and by its monosaccharide and alkylglycerol composition. Clone 3A2-C3 was the only one to infect mice in vivo and survive in macrophages in vitro, albeit at a much reduced rate compared to wild-type V121 promastigotes. The data suggest that some form of LPG may be necessary to ensure parasite infectivity.
Elhay
M
M
Walter and Eliza Hall Institute of Medical Research, Parkville, Victoria, Australia.
Kelleher
M
M
Bacic
A
A
McConville
M J
MJ
Tolson
D L
DL
Pearson
T W
TW
Handman
E
E
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D004351:Drug Resistance; D014644:Genetic Variation; D006028:Glycosphingolipids / Q000096:biosynthesis / Q000235:genetics* / Q000276:immunology; D007895:Leishmania tropica / Q000187:drug effects / Q000235:genetics* / Q000472:pathogenicity; D007896:Leishmaniasis / Q000235:genetics; D051379:Mice; D008807:Mice, Inbred BALB C; D012276:Ricin / Q000494:pharmacology*; D014774:Virulence / Q000235:genetics*
D000911:Antibodies, Monoclonal; D006028:Glycosphingolipids; C008290:lipophosphonoglycan; D012276:Ricin