| The fucosyl residue in cancer metastasis-related Lewis sugar antigens was selectedto study the function of a single sugar residue in glycans. Lewis antigens were divided intotwo major classes as sialyl and non-sialyl ones. The fucosylation step in sialyl Lewisantigen X (SLex) synthesis was catalyzed by α1,3fucosyltransferase-VII (α1,3FucT-VII), aterminal glycosyltransferase. This enzyme leads to the termination of glycan processingand the structure of the glycan can be no longer processed. Therefore, α1,3FucT-VII, whichadds only one fucosyl residue at glycan terminal may seemed as a good tool in the study ofthe function of a single sugar residue. By using the method of plasmid transfection, the cDNA of α1,3FucT-VII wastransfected into human H7721 hepatocarcinoma cell line, which expresses low level ofα1,3FucT-VII, and a series of biological effects after α1,3FucT-VII transfection was widelyobserved. These effects may reflect the function of the fucosyl residue in sialyl Lewis X insome degree. This thesis was divided into four parts; Part 1. Effect of α1,3FucT-VII Cdna transfection on the expression of SLex antigen on some cell surface receptors At first, the stable cell lines with different expressions of α1,3FucT-VII wereestablished. After characterization of the cell lines with the detection of α1,3FucT-VIImRNA and cell surface SLex (the product of α1,3FucT-VII), it was found that the cellsmock-transfected with the vector (Mock/H7721) expressed α1,3FucT-VII lowly, and theother two cell lines transfected with α1,3FucT-VII cDNA expressed moderate and highlevel of α1,3FucT-VII, they were named as FucTVII-M and FucTVII-H respectively. The receptor of insulin (InR) and epidermal growth factor (EGFR) were chosen as thetwo representatives of surface receptors to study the effects of α1,3FucT-VII transfectionon the SLex expression and the function of the receptors. InRs consists of α and β subunitsas a hetrodimer, but EGFR is a monomer. In addition, the integrin α5β1, receptor offibronection (Fn), which is a non-growth hormone receptor related to cell adhesion and 6migration was selected as another target of our research. The results were as following: 1. The expressions of surface and intracellular InR and EGFR were not altered afterthe transfection of α1,3FucT-VII, the immuno-precipitated InR and EGFR were alsounchaged. 2. The SLex expression on the α-subunit of InR was increased after the transfectionof α1,3FucT-VII, but that of EGFR was not changed. 3. The tyrosine auto-phophorylation of InR β-subunit was enhanced, while that ofEGFR was not altered after the transfection of α1,3FucT-VII. 4. The expression of insulin substrate-1 (IRS-1) was down regulated by thetransfection of α1,3FucT-VII, whereas, the tyrosine phophorylation per IRS-1protein wassignificantly up regulated. 5. The transfection of α1,3FucT-VII did not affect the relative SLex content onintegrin α5 and β1 subunits (SLex content /α5 or β1 protein) and the expression of β1subunit, but up regulated the expression of α5 subunit. 6. The cell adhesion and spreading on Fn were promoted after α1,3FucT-VIItransfection The above-mentioned SLex expression on InR α-subunit, tyrosine phosphorylation ofInR β subunit and IRS-1, expression of integrin α5 subunit as well as the cell adhesion/spreading on Fn showed the same order of FucTVII-H > FucTVII-M > Mock cells, whichwas in accordance with the order of the expression of α1,3FucT-VII mRNA. Hence, all theabove changes were probably mediated by the increase of SLex content on InR resultedfrom the transfection of α1,3FucT-VII. The only exception was the expression of IRS-1protein with the order of FucTVII-H < FucTVII-M < Mock cells. Part-2 Effect of α1,3FucT-VII cDNA transfection on some important signaling pathways...
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