EXPRESSION OF LIPOPHOSPHOGLYCAN, HIGH-MOLECULAR-WEIGHT PHOSPHOGLYCAN AND GLYCOPROTEIN-63 IN PROMASTIGOTES AND AMASTIGOTES OF LEISHMANIA-MEXICANA

The abundant surface glycoconjugate of Leishmania promastigotes, lipophosphoglycan (LPG), forms a blue-colored complex (lambda(max) = 649 nm) with the cationic dye Stains-all, which can be quantitated densitometrically on polyacrylamide gels of cell lysates. Promastigotes of Leishmania mexicana, Leishmania major and Leishmania donovani yield values of 1-3 x 10(6) LPG molecules cell-1. In amastigotes the LPG content is down-regulated below the detection limit ( < 10(3) molecules cell-1) in L. mexicana and L. donovani, but remains significant in L. major (2 x (10)3 molecules cell-1). In the case of L. mexicana, these results are supported by immunological studies. Using several monoclonal and polyclonal antibodies, LPG is undetectable by immunoblotting in lysates of either amastigotes or infected macrophages and the amastigote surface is devoid of LPG as judged by immunofluorescence and immunoelectron microscopy. Immunoblotting experiments demonstrate that amastigotes synthesize hydrophilic high-molecular weight compounds which stain blue with Stains-all and cross-react with the monoclonal and polyvalent antibodies suggesting the presence of similar phosphoglycan structures as in LPG. The high-molecular weight phosphoglycan appears to be located in the lumen of the flagellar pocket of mouse lesion amastigotes and may be secreted from there into the lumen of the parasitophorous vacuole of parasitized macrophages. In L. mexicana promastigotes the surface protease gp63 is amphiphilic and comprises about 1% of the cellular proteins. In contrast, in amastigotes gp63-related proteins are predominantly hydrophilic; they amount to only about 0.1% of the cellular proteins and are mainly located in the lumen of the extended lysosomes (megasomes) characteristic for this species.