Figure 5

A vital sugar code for ricin toxicity. (A) Proposed sugar code for ricin toxicity. α1,3-fucose residues of Lewis X structures (SSEA-1⁽⁺⁾) impair α2,3-sialylation of terminal galactoses (i.e., sialyl Lewis X formation, SSEA-1⁽⁻⁾), leading to enhanced exposure of terminal galactoses and ricin binding. Absence of fucosylation allows more efficient sialylation of terminal galactoses and thus is assumed to inhibit ricin binding. GlcNAc (N-acetylglucosamine), Gal (Galactose), NeuNAc (N-Acetylneuraminic acid, one type of sialic acid). (B) Slc35c1 mutant and WT MEFs were stained for α2,3-sialic acid using MALII and counterstained with DAPI to image nuclei. Scale bar, 50 μm. (C) HL-60 cells were treated with the fucosylation inhibitor 2F-peracetyl fucose (FI-1, 100 μM) or the sialylation inhibitor 3F_ax-peracetyl Neu5Ac (SI, 250 μM) for 3 days. The amount of the fucose containing, un-sialylated epitope SSEA-1 (CD15) was determined via flow cytometry. SSEA-1 expression of vehicle-treated cells (control) as well as an isotype-matched control (isotype control) is shown. (D) HL-60 cells were pretreated with inhibitors of fucosylation (FI-1, 100 μM) or sialylation (SI, 250 μM) and exposed to different amounts of ricin thereafter. The survival of the cells was determined using Alamar Blue. Data are representative of three independent experiments. (E, F) Slc35c1 wild type (WT) and mutant (KO) mESCs (E) and MEFs (F) were treated with SI (250 μM) and their sensitivity to ricin was assessed using Alamar Blue. Data in D-F are shown as mean ± SD of triplicate cultures. Experiments were repeated three times with similar results. ^*P < 0.05, ^**P < 0.01, ^***P < 0.001; NS, not significant (Student's t-test).