UDP-GlcNAc2-epimerase regulates cell surface sialylation and ceramide-induced cell death in human malignant lymphoma

Abstract

Abstract. Stress signals induce ceramide (cer) through sphingomyelinase activation, and metabolites of cer such as sphingosine (Sph) and sphingosine-1-phoshate (S-1-P) play a significant role in many biological processes. This study aimed to elucidate the association between the alteration in cell surface sialylation and ceramide-induced cell death in the human Burkitt's lymphoma cell line, HBL-8. The highly sialylated 3G3 clone was less sensitive to C6-ceramideinduced cell death. On the other hand, the hyposialylated 3D2 clone was more sensitive to C6-ceramide-induced cell death. Neuraminidase treatment or knockdown by siRNA of uridine diphosphate-N-acetylglucosamine 2-epimerase (UDP-GlcNAc2-epimerase), which is a key enzyme of sialic acid biosynthesis, enhanced the amount of cell death induced by C6-ceramide in the highly sialylated 3G3 clone. Sialic acid metabolic complementation assays using several precursors of sialic acid showed that cell surface resialylation by N-acetyl-D-mannosamine (ManNAc) inhibited C6-ceramideinduced cell death. The amount of cell death by C6-ceramide was enhanced after pretreatment with phosphatidylinositol 3-kinase (PI3K) inhibitor, LY294002 in both clones. In addition, clone 3G3 was less sensitive to Sph than the 3D2 clone. In conclusion, in human malignant lymphoma, ceramide and its metabolite-induced cell death is regulated by the amount of sialic acid on the cell surface which in turn is regulated by mRNA expression of UDP-GlcNAc2-epimerase.

UDP-GlcNAc 2-Epimerase/ManNAc Kinase (GNE): A Master Regulator of Sialic Acid Synthesis

“…In addition, Morris hepatoma cell lines were reported with decreased GNE expression [60], as did a chicken hepatoma cell line [132], although no direct links to GNE promoter methylation status were reported for these lines. Human Burkitt's lymphoma cell lines demonstrated sensitivity to ceramide-induced cell death regulated by GNE expression levels [133]. Hyposialylation due to GNE down-regulation was demonstrated in at least two non-cancerous cell lines.…”

Glycosylation in lymphoma: Biology and glycotherapy

“…. Stress within the vessels may induce the apoptosis of lymphoma cells, whereas sialylation is known to protect lymphoma cells from stress signals and may rescue lymphoma cells from apoptosis. We speculate that sialylation of cell surface glycoproteins may regulate cell adhesion in lymphoma, and desialylation by neuraminidase or knockdown of ST6Gal1 may enhance cell adhesion to galectin or extracellular matrix, thereby inhibiting the invasiveness or metastasis of lymphoma cells and providing a potential strategy for lymphoma glycotherapy by desialylation.…”

“…. Knockdown of GNE in HBL‐8 3G3 cells appears to inhibit cell growth and enhance cell death caused by ceramide, an inducer of apoptosis or cell death . The survival of tumor cells from stress such as hypoxia, oxidative stress, or apoptosis may be related to increased metastatic tumors, and thus the high survival rate of 3G3 clone cells following sialylation may be associated with high metastasis in a SCID animal model.…”

“…Resialylation by ManNAc inhibits ceramide‐induced cell death and inhibits necrosis and death due to lactate dehydrogenase (LDH) release from the cytoplasm caused by membrane permeability, suggesting that resialylation may be closely associated with enhanced HBL‐8 cell growth or the inhibition of cell death. RT‐PCR and flow cytometry using small interfering RNA (siRNA) for GNE showed that the knockdown of GNE in the 3G3 clone of HBL‐8 reduced GNE mRNA in real time and decreased cell surface sialylation of the sialic acid‐specific lectin from Maachia amurensis . In addition, reduced cell surface sialylation on the 3G3 clone of HBL‐8 by knockdown of GNE reduced the cell growth rate and increased sensitivity to ceramide‐induced cell death.…”

Recent Progress and New Perspectives in Lymphoma Glycobiology

“…In addition, clone 3G3 was less sensitive to Sph than the 3D2 clone. In conclusion, in human malignant lymphoma, Cer and its metabolitesinduced cell death is regulated by the amount of sialic acid on the cell surface, which in turn is regulated by mRNA expression of UDP -GlcNAc2 -epimerase 67) . Goni et al recently reported that Cer and sphingolipids modified the physical properties of the cell membrane 68) , and Montes et al reported that Cer induced an increased efflux or permeability in cell membranes 69) .…”