Sulfatide is expressed in both erythrocytes and platelets of bovine origin.
ABSTRACT A novel sulfated glycosphingolipid containing a sulfated galactosyl residue was isolated from bovine erythrocyte ghosts, and purified to homogeneity by column chromatography on DEAE-Sephadex and silica beads. Structural characterization included compositional analyses, permethylation studies, proton nuclear magnetic resonance (NMR) spectroscopy, negative secondary ion mass spectrometry (SIMS), solvolysis and immunostaining on thin-layer chromatogram. As a result, the structure of this glycolipid is proposed as HSO3-Gal beta 1-1 Cer. The ceramide portion contained d18:1, d18:0 and t18:0, and the predominant fatty acid consisted of palmitate and palmitate with a hydroxy group, as deduced by both compositional analysis and negative SIMS mass spectrometry. The component of this glycosphingolipid probably originates from erythrocytes and platelets as indicated by the results of flow cytometry analysis using Sulph I monoclonal antibody. The yield of galactosyl sulfatide was about 0.37 mg/kg wet bovine erythrocyte membranes, about three times that of human kidney. Our results strongly suggest that galactosylceramide sulfate on erythroid cells may play an important biological role in cell to cell interaction and recognition.
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ABSTRACT: Platelets contact each other at the site of vascular injury to stop bleeding. One negative regulator of platelet aggregation is Disabled-2 (Dab2), which is released to the extracellular surface upon platelet activation. Dab2 inhibits platelet aggregation through its phosphotyrosine-binding (PTB) domain by competing with fibrinogen for alphaIIbbeta3 integrin receptor binding by an unknown mechanism. Using protein-lipid overlay and liposome-binding assays, we identified that the N-terminal region of Dab2, including its PTB domain (N-PTB), specifically interacts with sulfatides. Moreover, we determined that such interaction is mediated by two conserved basic motifs with a dissociation constant (K(d)) of 0.6 microM as estimated by surface plasmon resonance (SPR) analysis. In addition, liposome-binding assays combined with mass spectroscopy studies revealed that thrombin, a strong platelet agonist, cleaved N-PTB at a site located between the basic motifs, a region that becomes protected from thrombin cleavage when bound to sulfatides. Sulfatides on the platelet surface interact with coagulation proteins, playing a major role in haemostasis. Our results show that sulfatides recruit N-PTB to the platelet surface, sequestering it from integrin receptor binding during platelet activation. This is a transient recruitment that follows N-PTB internalization by an actin-dependent process. Our experimental data support a model where two pools of Dab2 co-exist at the platelet surface, in both sulfatide- and integrin receptor-bound states, and their balance controls the extent of the clotting response.PLoS ONE 01/2009; 4(11):e8007. · 4.09 Impact Factor