Osanai T, Watanabe Y, Sanai Y
Department of Biochemical Cell Research, Tokyo Metropolitan Institute of Medical Science (RINSHOKEN), Japan.
Biochem Biophys Res Commun. 1997 Dec 18;241(2):327-33. doi: 10.1006/bbrc.1997.7817.
We have studied ganglioside alterations and their enzymatic basis during the course of neural differentiation of mouse embryonic carcinoma cell line P19. This cell line can differentiate into neurons and astrocytes on cell aggregation after treatment with retinoic acid (RA) or into muscle cells on dimethyl sulfoxide (DMSO) treatment. GD3, detected on immunostaining after thin-layer chromatography (TLC) with monoclonal antibody (MAb) R24, was markedly present in aggregates treated with RA. GM3 synthase (alpha 2,3-sialyltransferase, SAT-I) in neurons was found to exhibit the highest activity. GD3 synthase (alpha 2,8-sialyltransferase, SAT-II) and GD3 synthase mRNA, as analyzed by Northern blotting, were also markedly present in aggregates and neurons induced by RA. However, on treatment with DMSO, which induces muscle cells, there was no change in the level of GD3 synthase activity, and its transcript was hardly detected during the course of muscle differentiation. GT1b synthase (alpha 2,3-sialyltransferase, SAT-IV) was present at similar levels in undifferentiated cells and aggregates treated with RA, but a higher level was observed in neurons. On the other hand, the level of GQ1b synthase (alpha 2,8-sialyltransferase, SAT-V) in RA-induced aggregates was significantly higher than that in neurons. These results show that RA but not DMSO induces the expression of GM3, GD3, GT1b and GQ1b synthases, and particularly GD3 synthase mRNA, in the ganglioside biosynthetic pathway during the neural differentiation of embryonic carcinoma P19 cells.
我们研究了小鼠胚胎癌细胞系P19神经分化过程中神经节苷脂的变化及其酶学基础。该细胞系在用视黄酸（RA）处理后细胞聚集时可分化为神经元和星形胶质细胞，在用二甲基亚砜（DMSO）处理时可分化为肌肉细胞。用单克隆抗体（MAb）R24进行薄层色谱（TLC）免疫染色后检测到的GD3，在经RA处理的聚集体中显著存在。发现神经元中的GM3合酶（α2,3-唾液酸转移酶，SAT-I）具有最高活性。通过Northern印迹分析，GD3合酶（α2,8-唾液酸转移酶，SAT-II）和GD3合酶mRNA在经RA诱导的聚集体和神经元中也显著存在。然而，在用诱导肌肉细胞的DMSO处理时，GD3合酶活性水平没有变化，并且在肌肉分化过程中几乎检测不到其转录本。GT1b合酶（α2,3-唾液酸转移酶，SAT-IV）在未分化细胞和经RA处理的聚集体中的水平相似，但在神经元中观察到更高的水平。另一方面，RA诱导的聚集体中GQ1b合酶（α2,8-唾液酸转移酶，SAT-V）的水平显著高于神经元中的水平。这些结果表明，在胚胎癌P19细胞的神经分化过程中，RA而非DMSO诱导神经节苷脂生物合成途径中GM3、GD3、GT1b和GQ1b合酶的表达，特别是GD3合酶mRNA的表达。
