Suppression of Sialyl Lewis X Expression and E-selectin-mediated Cell Adhesion in Cultured Human Lymphoid Cells by Transfection of Antisense cDNA of an α1→3 Fucosyltransferase (Fuc-T VII)

• Published in: The Journal of biological chemistry Vol. 271; no. 49; p. 31556
• Main Authors: Nozomu Hiraiwa, Taeko Dohi, Naoko Kawakami-Kimura, Miki Yumen, Katsuyuki Ohmori, Michiyuki Maeda, Reiji Kannagi
• Format: Journal Article
• Language: English
• Published: American Society for Biochemistry and Molecular Biology 06.12.1996
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.271.49.31556

The antisense cDNA approach was used to identify the endogenous fucosyltransferase species responsible for synthesis of the sialyl Lewis X (NeuAcα2→3 Galβ1→4[Fucα1→3]GlcNAcβ1→R) determinant in human lymphoid cells. The cultured human adult T-cell leukemia cell line, ED40515-N, expressed the message of α1→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 (Galβ1→4 [Fucα1→3]GlcNAcβ1→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 α1→3 fucosyltransferase against sialylated type 2 chain substrate. This was accompanied by diminution of cell adhesive activity toward E-selectin on interleukin-1β-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 α1→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.