alpha-Neup5Ac-(2--3)-beta-D-Galp-(1--4)-[alpha-L-Fucp-(1--3)]-D-GlcpNAc and Leukemia--Lymphoid

The antisense cDNA approach was used to identify the endogenous fucosyltransferase species responsible for synthesis of the sialyl Lewis X (NeuAcalpha2-->3 Galbeta1-->4[Fucalpha1-->3]GlcNAcbeta1-->R) determinant in human lymphoid cells. The cultured human adult T-cell leukemia cell line, ED40515-N, expressed the message of alpha1-->3 fucosyltransferase (Fuc-T) IV and VII, with a low level of the Fuc-T III and VI message, and manifested the sialyl Lewis X as well as Lewis X (Galbeta1-->4 [Fucalpha1-->3]GlcNAcbeta1-->R) determinant at the cell surface. Transfection of this cell line with the pRc/CMV vector containing an antisense human Fuc-T VII construct (pRc/CMV/5'FT7AS) resulted in a significant decrease of endogenous Fuc-T VII message and a marked reduction in the cell surface expression of sialyl Lewis X determinant as well as a reduction in the enzymatic activity of alpha1-->3 fucosyltransferase against sialylated type 2 chain substrate. This was accompanied by diminution of cell adhesive activity toward E-selectin on interleukin-1beta-treated endothelial cells. These results indicated that the synthesis of the sialyl Lewis X determinants that were functionally active as E-selectin ligands was mainly mediated by Fuc-T VII in these lymphoid cells. On the other hand, the message of Fuc-T IV showed no significant change in the transfectant clones, and the surface expression of the Lewis X antigen as well as the enzymatic activity of alpha1-->3 fucosyltransferase against non-sialylated type 2 chain substrate was well preserved. The clear contrast between the diminished expression of sialyl Lewis X and the conserved manifestation of Lewis X in the transfectant clones suggested that the synthesis of sialyl Lewis X and that of Lewis X are independently regulated by different fucosyltransferases in human lymphoid cells. Fuc-T VII must be involved in the synthesis of sialyl Lewis X, while the synthesis of Lewis X is mediated by an enzyme other than Fuc-T VII, most probably Fuc-T IV.

Topics: Adult; Blotting, Northern; Cell Adhesion; Cells, Cultured; DNA, Complementary; E-Selectin; Flow Cytometry; Fucosyltransferases; Humans; Leukemia, Lymphoid; Lewis Blood Group Antigens; Oligonucleotides, Antisense; Oligosaccharides; RNA, Messenger; Sialyl Lewis X Antigen; Transfection; Tumor Cells, Cultured