Effect of Inhibitors of GLycoprotein Processing on Cytokine Secretion and Production in Anti CD3-Stimulated T Cells

Abstract

We have investigated the effect of inhibitors of glycoprotein processing on cytokine secretion and production in anti CD3-stimulated T cells to elucidate the role of carbohydrate in the triggering of T cell function. The inhibitors of glycoprotein processing, especially mannnosidase inhibitors, enhanced the anti CD3-induced production of interleukin-2 (IL-2), which is a cytokine without the linkage sequence of N-linked oligosaccharides. On the other hand, N-methyl-1-deoxynojirimycin (NMdNM, an inhibitor of processing glucosidase I), 1-deoxynojirimycin (dNM, an inhibitor of processing glucosidase I and II) and bromoconduritol (BCD, an inhibitor of processing glucosidase II) inhibited the secretion of interleukin-4 (IL-4), interferon-γ (IFN-γ), or interleukin-5 (IL-5) into culture supernatants of anti CD3-stimulated T cells, which had N-linked oligosaccharides. Mannosidase inhibitors, 1-deoxymannojirimycin (dMAN, and inhibitor of processing mannosidase I) and swainsonine (SWN, an inhibitor of processing mannosidase II) did not inhibit the secretion or production of IL-4, IFN-γ and IL-5. To confirm the inhibition of N-linked oligosaccharide processing in the cytokines by the above inhibitors, the binding of IFN-γ to lectins with various sugar-binding specificities was investigated. All inhibitors reduced the binding of IFN-γ to PHA E4, which had a high affinity to bi- or tri-antennary complex type N-linked oligosaccharides with bisecting N-acetylglucosamine. Similarly, all inhibitors reduced the binding of IFN-γ to PHA L4, which had high affinity to tri- or tetra-antennary complex type N-linked oligosaccharides with β1-6-linked branching. SWN and dMAN increased the binding of IFN-γ to concanavalin A (ConA), which had a high affinity to bi-antennary complex type, hybrid type and high-mannose type N-linked oligosaccharides. These results suggest that the processing inhibitors used here inhibit the N-linked oligosaccharide processing of cytokines, and the inhibition of processing enzyme glucosidases I and II induces a decreased secretion of cytokines with N-linked oligosaccharides.