Nucleotide sugar transporters: Structure and function, and physiological regulation

核苷酸糖转运蛋白：结构和功能以及生理调节

• 批准号: 11480172
• 负责人: KAWAKITA Masao
• 金额: $ 9.98万
• 依托单位: Kogakuin University (2001)Tokyo Metropolitan Organization for Medical Research (1999-2000)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-11480172/
• 关键词: Nucleotide sugar transporter; UDP-galactose; Glycobiology; Colon cancer; Glycoconjugates; UDP-N-acetylglucosamine; CMP-sialic acid; UGTre17輸送体; UST74C輸送体; 多重特異的糖ヌクレオチド輸送体; UDP-ガラクトース輸送体; UDP-ガラクトース輸送体遺伝子; 可溶化再構成; ヒスチジンータグ; CMP-シアル酸輸送体

Several nucleotide sugar transporters including human UDP-GlcNAc transporter, human CMP -sialic acid transporter, human UDP-GlcA/UDP-GalNAc/UDP-GlcNAc multi-specific transporter (hUGTrel7), Drosophila multi-specific UDP-sugar transporter (UST74C) were cloned and characterized. Substrate specificity of these transporters were convincingly defined by their functional expression in Saccharomyces cerevisiae Golgi membranes. HUGTrel7 is localized to the ER membranes and may be involved in glucuronidation of xenobiotics and the biosynthesis of proteoglycans. UST74C is the product of Drosophila frc gene whose defect results in phenotypes resembling wingless and Notch. Analysis of the phenotypes of frc mutants suggested that change s in nucleotide-sugar levels could differently affect these signaling pathways.Molecular chimeras between UDP-Gal transporter and CMP-sialic acid transporter were constructed and analyzed in order to identify submolecular regions responsible for the determination of … More substrate specificity. We found that chimeras that contain transmembrane helices 2,3 and 7 derived from CMP-sialic acid transporter in the UDP-Gal transporter background could transport both UDP-Gal and CMP-sialic acid. This indicates that the regions which are critical for determining the substrate specificity of these two transporters resided in different submolecular sites in the two transporters, and that these different determinants could be present within one protein without interfering each other's function.The amount of mRNA for UDP-Gal transporter was significantly increased in colon cancer tissues compared with nonmalignant mucosa tissues. The increase was more prominent in patients with advanced colorectal cancer of Duke's stages C and D. Transfection of UDP-Gal transporter cDNA to cultured colon cancer cells led to increased expression of Thomsen-Freidenreich antigen and of sialyl Lewis A and sialyl Lewis X determinants in transfected cells, which resulted in markedly enhanced cell adhesion to vascular E-selectin. These findings suggest that nucleotide sugar transporters can act as one of the regulatory elements of cellular behavior through their effects on the structure of cell surface glycoconjugates. Less

克隆并鉴定了几种核苷酸糖转运蛋白，包括人UDP-GlcNAc转运蛋白、人cMP-唾液酸转运蛋白、人UDP-Glca/UDP-GalNAc/UDP-GlcNAc多特异性转运蛋白(HUGTrel7)、果蝇多特异性UDP-糖转运蛋白(UST74C)。这些转运蛋白在酿酒酵母高尔基膜中的功能表达令人信服地确定了它们的底物特异性。HUGTrel7定位于内质网细胞膜，可能参与外源物质的葡萄糖醛酸化和蛋白多糖的生物合成。UST74C是果蝇FRC基因的产物，其缺陷导致了类似于无翼和Notch的表型。对frc突变体的表型分析表明，核苷酸-糖水平的变化可能会对这些信号通路产生不同的影响。构建了UDP-Gal转运体和Cp-唾液酸转运体之间的分子嵌合体，并对其进行了分析，以确定与…相关的亚分子区域底物专一性更强。我们发现，在UDP-Gal转运体背景下，含有来自Cp-唾液酸转运体的跨膜螺旋2，3和7的嵌合体可以同时转运UDP-Gal和Cp-唾液酸。这表明，决定这两个转运蛋白底物特异性的关键区域位于两个转运蛋白的不同亚分子位置，这些不同的决定因素可以存在于同一蛋白中，而不会相互干扰功能。结肠癌组织中UDP-Gal转运蛋白的mRNA含量显著高于非恶性粘膜组织。UDP-Gal转运蛋白的表达在Duke‘s分期为C、D期的晚期结直肠癌患者中更为明显。UDP-Gal转运体基因导入培养的结肠癌细胞后，可导致Thomsen-Fredenreich抗原、唾液酸化Lewis A和Sialyl Lewis X决定簇的表达增加，从而显著增强细胞与血管E-选择素的粘附性。这些发现表明，核苷酸糖转运体可以通过影响细胞表面糖结合物的结构而作为细胞行为的调节元件之一。较少

项目成果

Kainuma, M. et al.: "Coexpression of α1, 2 galactosyltransferase and UDP-galactose transporter efficiently galactosylates N- and O-glycans in Saccharomyces cerevisiae"Glycobiology. 9. 133-141 (1999)

Kainuma, M. 等人：“α1, 2 半乳糖基转移酶和 UDP-半乳糖转运蛋白的共表达可有效地在酿酒酵母中半乳糖基化 N-和 O-聚糖”，《糖生物学》9. 133-141 (1999)。

Segawa, H.: "Human and Drosophila UDP-galactose transporters transport UDP-N-acetylgalactosamine in addition to UDP-galactose"Eur.J.Biochem.. 269. 128-138 (2002)

Sekawa, H.：“人和果蝇 UDP-半乳糖转运蛋白除了 UDP-半乳糖外还转运 UDP-N-乙酰半乳糖胺”Eur.J.Biochem.. 269. 128-138 (2002)

Aoki, K.他2名: "Substrate Recognition by UDP-galactose and CMP-sialic acid transporters : different sets of transmembrance helices are utilized for the specific recognition of UDP-galactose and CMP-sialic acid"J.Biol.Chem.. 276. 21555-21561 (2001)

Aoki, K. 和另外 2 人：“UDP-半乳糖和 CMP-唾液酸转运蛋白的底物识别：不同组的跨膜螺旋用于 UDP-半乳糖和 CMP-唾液酸的特异性识别”J.Biol.Chem.. 276.21555-21561（2001）

Aoki, K. et al.: "Expression and activity of chimeric molecules between human UDP-galactose transporter and CMP-sialic acid transporter"J. Biochem.. 126. 940-950 (1999)

Aoki, K. 等人：“人 UDP-半乳糖转运蛋白和 CMP-唾液酸转运蛋白之间的嵌合分子的表达和活性”J.

Kumamoto, K.他6名: "Increased expression of UDP-galactose transporter messenger RNA in human colon cancer tissues and its implication in synthesis of Thomsen-Friedenreich antigen and sialyl Lewis A/X determinants"Cancer Res.. 61. 4620-4627 (2001)

Kumamoto, K. 和其他 6 人：“人结肠癌组织中 UDP-半乳糖转运蛋白信使 RNA 的表达增加及其对 Thomsen-Friedenreich 抗原和唾液酸 Lewis A/X 决定簇合成的影响”Cancer Res.. 61. 4620-4627 (2001)