Role of Leishmania (Leishmania) amazonensis amastigote glycosphingolipids in macrophage infectivity

Abstract

The role of glycosphingolipids (GSLs) present in amastigote forms of Leishmania (Leishmania) amazonensis during infection of macrophages was analyzed, with particular emphasis on GSLs presenting the terminal Galpß1-3Galpα disaccharide. Macrophage invasion by L. (L.) amazonensis amastigotes was reduced by 37% when the disaccharide Galpß1-3Galp (1 mM) was added to the culture medium. The putative macrophage receptor/lectin for ß-Gal-globotriaosylceramide (Galpß1-3Galpα1-4Galpß1-4Glcpß1-1Cer) and other structurally related GSLs from L. (L.) amazonensis amastigotes were analyzed by micelles and parasite binding assay to peritoneal macrophage proteins fractionated by SDS-PAGE under nonreducing conditions. Micelles containing purified amastigote GSLs or a suspention of L. (L.) amazonensis amastigotes fixed with 2% formaldehyde were incubated with nitrocellulose membrane containing the macrophage proteins transferred by Western blotting. Binding of micelles containing purified GSLs from amastigote forms or fixed L. (L.) amazonensis amastigotes to nitrocellulose membrane was probed using monoclonal antibody ST-3, which recognizes the glycoepitope Galpß1-3Galpα1-R present either in the micelle preparation or on the amastigote surface. Macrophage protein with molecular mass ~30 kDa bound the amastigote GSL and appeared to be a doublet on electrophoresis. The specificity of this interaction was confirmed using fixed L. (L.) chagasi amastigotes, which do not express GSLs such as ß-Galpglobotriaosylceramides, and which do not bind to 30-kDa protein. Correspondence

Current relevance of fungal and trypanosomatid glycolipids and sphingolipids: studies defining structures conspicuously absent in mammals

“…These mAbs were able to inhibit up to 80% of macrophage invasion by amastigotes, suggesting that GSLs are essential for L. (L.) amazonensis binding to macrophages (Straus et al 1993. Moreover, Gal pβ1-3Gal p disaccharide, which is part of one of the L. (L.) amazonensis amastigote GSLs, Gal pβ1-3Gal pα1-3Gal pβ1-4Glc pβ1-1Cer (referred herein to as β-Gal-globotriaosylceramide), recognized by mAb ST-3, was also able to inhibit ∼40% of macrophage invasion by these parasites, corroborating therefore the importance of these GSLs in L. (L.) amazonensis amastigote infection (Straus et al 1993, Tanaka et al 2007). Recently, a possible receptor for L. (L.) amazonensis amastigote GSLs was identified as being a 30 kDa protein from macrophage (Tanaka et al 2007).…”

“…Moreover, Gal pβ1-3Gal p disaccharide, which is part of one of the L. (L.) amazonensis amastigote GSLs, Gal pβ1-3Gal pα1-3Gal pβ1-4Glc pβ1-1Cer (referred herein to as β-Gal-globotriaosylceramide), recognized by mAb ST-3, was also able to inhibit ∼40% of macrophage invasion by these parasites, corroborating therefore the importance of these GSLs in L. (L.) amazonensis amastigote infection (Straus et al 1993, Tanaka et al 2007). Recently, a possible receptor for L. (L.) amazonensis amastigote GSLs was identified as being a 30 kDa protein from macrophage (Tanaka et al 2007). Interestingly, this protein was not able to bind to Leishmania (Leishmania) chagasi amastigotes, which cause a visceral type of leishmaniasis and lack β-Gal-globotriaosylceramide, suggesting that GSLs of L. (L.) amazonensis amastigotes confer a particular specificity towards the host (Tanaka et al 2007).…”

“…Recently, a possible receptor for L. (L.) amazonensis amastigote GSLs was identified as being a 30 kDa protein from macrophage (Tanaka et al 2007). Interestingly, this protein was not able to bind to Leishmania (Leishmania) chagasi amastigotes, which cause a visceral type of leishmaniasis and lack β-Gal-globotriaosylceramide, suggesting that GSLs of L. (L.) amazonensis amastigotes confer a particular specificity towards the host (Tanaka et al 2007). tributed in membrane rafts (Fig.…”

“…Recently, we demonstrated that Aureobasidin-A (AbA), an IPC synthase inhibitor, was able to decrease growth rate of promastigotes of L. (L.) (Tanaka et al 2007. In addition, AbA-treated promastigotes were viable, suggesting that IPC is important for cell division events.…”

Structures of Glycolipids Found in Trypanosomatids: Contribution to Parasite Functions

“…The role of this glycolipid in macrophage infectivity was confirmed using Mabs directed to this molecule. A putative receptor/lectin of macrophages with a molecular mass of 30kDa for L.amazonensis GSL was suggested [21].…”