Cell Adhesion Regulation Mechanism by Sialyl-Le^X Structure.

Sialyl-Le^X 结构的细胞粘附调节机制。

• 批准号: 09670161
• 负责人: NAKAMURA Mitsuru
• 金额: $ 1.98万
• 依托单位: JICHI MEDICAL SCHOOL
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1997
• 资助国家: 日本
• 起止时间: 1997 至 1998
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-09670161/
• 关键词: E-Selectin; Sialyl-LeX; Counter Ligand; Cell Adhesion; E-セレクチン; シリアルルイス-X; B細胞分化; コア2GlcNAcの転移酵素; シアリルルイスX; GATA

We have demonstrated through remodeling of the glycosyltrasferase machineries in human pre-B leukemia cells that cell surface sialyl-LeX (sLeX) antigen expression level is controlled by core 2 N-acetylglucosaminyltransferase (C2GnT) gene. In addition, we have revealed the followings.1. sLeX antigen is not expressed on every protein but expressed on a specific core glycoprotein with molecular size of l5OkDa (gp 150). sLeX antigen determinants are expressed at the termini of 0-linked oligosaccharide chains of gplSO.2. Analyzing transcription regulation mechanism of C2GnT expression by luciferase assay, EMSA, and RNA blotting, we have demonstrated the followings.(1) Among three TATA boxes in the C2GnT transcription regulation area, a deletion mutant containing the most proximate -182 TATA to the transcription initiation site showed the highest luciferase activity.(2) C2GnT gene expressing cells had 10-fold transient transcription activity of the non-expressing cells. Transient transcription activity was down-regulated to 1/10 by inhibiting C2GnT gene expression among cell differentiation. These are in a good agreement with changes of C2GnT message, C2GnT enzyme activity, and cell surface sLeX antigen expression levels.(3) Involved transcription factor binding sites are GATA, NF-IL6, and Spi. The expression of GATA3, NF-1L6, C/EBPgamma, Sp3, and Sp4 genes correlated with C2GnT gene expression level.

我们通过重组人前B白血病细胞中糖基转移酶机制，证明细胞表面唾液酸LeX（sLeX）抗原表达水平受核心2 N-乙酰氨基葡萄糖转移酶（C2 GnT）基因控制。此外，我们还揭示了以下内容。1. sLeX抗原不是在所有蛋白质上表达，而是在分子大小为150 kDa的特异性核心糖蛋白（gp 150）上表达。sLeX抗原决定簇在gp15.2的0-连接寡糖链的末端表达。通过荧光素酶分析、EMSA和RNA印迹分析C2 GnT表达的转录调控机制，我们证明了以下几点。(1)在C2 GnT转录调控区的三个TATA盒中，含有最接近转录起始位点的-182 TATA的缺失突变体显示出最高的荧光素酶活性。(2)C2 GnT基因表达细胞具有非表达细胞的10倍瞬时转录活性。通过抑制C2 GnT基因在细胞分化过程中的表达，瞬时转录活性下调至1/10。这些与C2 GnT信息、C2 GnT酶活性和细胞表面sLeX抗原表达水平的变化非常一致。(3)涉及的转录因子结合位点是加塔、NF-IL 6和Spi。GATA 3、NF-1 L 6、C/EBP γ、Sp3和Sp 4基因表达水平与C2 GnT基因表达水平相关。

项目成果

Nakamura M,Kudo T,Narimatsu H,Furukawa Y,Kikuchi J,Asakura S,Yang W,Iwase S,Hatake K,and Miura Y: "Single Glycosyltrans-ferase, Core 2 beta1*6N-acetylglucosaminyltrans-ferase, Regulates Cell Surface Sialyl-Le^X Ex-pression Level in Human Pre-B Lymphocytic

Nakamura M、Kudo T、Narimatsu H、Furukawa Y、Kikuchi J、Asakura S、Yang W、Iwase S、Hatake K 和 Miura Y：“单一糖基转移酶，核心 2 beta1*6N-乙酰氨基葡萄糖转移酶，调节细胞表面

Nakamura,M.: "Single Glycosyltransferase,Core2β1→6N-Acetylglucosaminyltransferase,Regulates Cell Surface Sialyl-LeX Expression Level in Human Pre-B lymphocytic Leutemia Cell Line KM3" I.Biol.Chem.273. 26779-26789 (1998)

Nakamura, M.：“单一糖基转移酶，Core2β1→6N-乙酰氨基葡萄糖转移酶，调节人前 B 淋巴细胞白血病细胞系 KM3 中的细胞表面 Sialyl-LeX 表达水平”I.Biol.Chem.273 (1998)。

Kudo T,Ikehara Y,Togayachi A,Morozumi K,Watanabe M,Nakamura M,Nishihara S,and Narimatsu H.: "Up-regulation of a Set of Glycosyl-transferase Genes in Human Colorectal Cancer." Lab.Invest.78. 797-811 (1998)

Kudo T、Ikehara Y、Togayachi A、Morozumi K、Watanabe M、Nakamura M、Nishihara S 和 Narimatsu H.：“人类结直肠癌中一组糖基转移酶基因的上调”。

Ishii,A.: "Expression Cloning and Functional Characterization of Human cDNA for Ganglioside GM3 Synthase" J.Biol.Chem.273. 31652-31655 (1998)

Ishii,A.：“神经节苷脂 GM3 合酶的人 cDNA 的表达克隆和功能表征”J.Biol.Chem.273。

Kudo,T.: "Up-regulation of a set of glycosyltransferase genes in human colorectal cancer" Lab.Invest. 78. 797-811 (1998)

Kudo,T.：“人类结直肠癌中一组糖基转移酶基因的上调”Lab.Invest。