Expression cloning of sialyltransferase involved in malignant progression of human RCC

本研究利用表达克隆和PCR方法，对与人肾细胞癌（RCC）恶性进展相关的神经节苷脂第一种物质--单唾液酸半乳糖苷脂（MSGG）合成有关的α 2，3-唾液酸转移酶进行了鉴定。迄今为止，我们还不能通过表达克隆法获得α 2，3-唾液酸转移酶，但通过该方法鉴定了rRNA的5'前体、胎盘转化生长因子β、未知cDNA等的浓缩cDNA。本实验通过PCR方法获得了hST 3Gal Ⅰ和hST 3Gal Ⅳ，并对它们与MSGG表达的关系进行了研究。北方分析表明，hST 3 Gal I与肾细胞癌的恶性进展无明显相关性，稳定转染hST 3 Gal I的细胞无MSGG表达，稳定转染hST 3 Gal IV的细胞MSGG表达中度增加，但在几乎所有正常肾组织中，hST 3 Gal IV转录本明显增加，而在大多数癌组织中，hST 3 Gal IV转录本减少。提示hST 3Gal IV与肾分化相关。根据这些结果，我们推测可能存在另一种α 2，3-唾液酸转移酶，它决定着MSGG的合成，目前对这种唾液酸转移酶的研究还在进行中。在本研究中，我们鉴定了N-乙酰氨基半乳糖二唾液酸型链副糖苷（GalNAc DSPG-1）作为一种新的神经节苷脂，并发现DSGG分布在中胚层起源的细胞中。我们还发现，MSGG的表达与GM 3在人RCC中的表达呈负相关。

In all attempt to identify α 2, 3-sialyltransferase involved in synthesis of monosialosyl galactosyl globoside (MSGG), the first substance of globo-series ganglioside, associated with the malignant progression of human renal cell carcinoma (RCC), we utilized both expression cloning procedure and PCR method. Thus far, we could not obtain α 2, 3-sialyltransferase by expression cloning procedure, but 5' precursor of rRNA, placental transforming growth factor β, unknown cDNA, etc. were identified as concentrated cDNA by this procedure. We are now investigating the relationship between these cDNA and expression of MSGG.We obtained hST3Gal I and hST3Gal IV by PCR method. By Northern analysis, there was no clear correlation between hST3Gal I and the malignant progression of human RCC.None of the stable transfectants with hST3Gal I examined showed expression of MSGG.Stable transfectant with hST3Gal IV showed moderate increase of MSGG.However, hST3Gal IV transcript increased prominently in almost all normal renal tissues and decreased in most cases of cancer tissues. It was suggested that hST3Gal IV is associated with renal differentiation. From these results, it was anticipated that there is another a 2, 3-sialyltransferase determining the sysnthesis of MSGG.Investigation of this putative sialyltransferase is still underway. During this research, we identified N-acetyl-galactosaminyl disialosyl typel chain paragloboside (GalNAc DSPG-1) as a novel ganglioside, and found that DSGG is distributed in cells of mesoderm origin. We also found that expression of MSGG is inversely related to that of GM3 in human RCC.